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Sample records for aeruginosa biofilms mediates

  1. Direct evaluation of Pseudomonas aeruginosa biofilm mediators in a chronic infection model.

    Science.gov (United States)

    Byrd, Matthew S; Pang, Bing; Hong, Wenzhou; Waligora, Elizabeth A; Juneau, Richard A; Armbruster, Chelsie E; Weimer, Kristen E D; Murrah, Kyle; Mann, Ethan E; Lu, Haiping; Sprinkle, April; Parsek, Matthew R; Kock, Nancy D; Wozniak, Daniel J; Swords, W Edward

    2011-08-01

    Biofilms contribute to Pseudomonas aeruginosa persistence in a variety of diseases, including cystic fibrosis, burn wounds, and chronic suppurative otitis media. However, few studies have directly addressed P. aeruginosa biofilms in vivo. We used a chinchilla model of otitis media, which has previously been used to study persistent Streptococcus pneumoniae and Haemophilus influenzae infections, to show that structures formed in vivo are biofilms of bacterial and host origin within a matrix that includes Psl, a P. aeruginosa biofilm polysaccharide. We evaluated three biofilm and/or virulence mediators of P. aeruginosa known to affect biofilm formation in vitro and pathogenesis in vivo--bis-(3',5')-cyclic dimeric GMP (c-di-GMP), flagella, and quorum sensing--in a chinchilla model. We show that c-di-GMP overproduction has a positive impact on bacterial persistence, while quorum sensing increases virulence. We found no difference in persistence attributed to flagella. We conclude from these studies that a chinchilla otitis media model provides a means to evaluate pathogenic mediators of P. aeruginosa and that in vitro phenotypes should be examined in multiple infection systems to fully understand their role in disease.

  2. Pseudomonas aeruginosa biofilms in cystic fibrosis

    DEFF Research Database (Denmark)

    Høiby, Niels; Ciofu, Oana; Bjarnsholt, Thomas

    2010-01-01

    The persistence of chronic Pseudomonas aeruginosa lung infections in cystic fibrosis (CF) patients is due to biofilm-growing mucoid (alginate-producing) strains. A biofilm is a structured consortium of bacteria, embedded in a self-produced polymer matrix consisting of polysaccharide, protein...... and DNA. In CF lungs, the polysaccharide alginate is the major part of the P. aeruginosa biofilm matrix. Bacterial biofilms cause chronic infections because they show increased tolerance to antibiotics and resist phagocytosis, as well as other components of the innate and the adaptive immune system....... As a consequence, a pronounced antibody response develops, leading to immune complex-mediated chronic inflammation, dominated by polymorphonuclear leukocytes. The chronic inflammation is the major cause of the lung tissue damage in CF. Biofilm growth in CF lungs is associated with an increased frequency...

  3. Pyoverdine and PQS Mediated Subpopulation Interactions Involved in Pseudomonas aeruginosa Biofilm Formation

    DEFF Research Database (Denmark)

    Yang, Liang; Nilsson, Martin; Gjermansen, Morten

    2009-01-01

    Using flow chamber-grown Pseudomonas aeruginosa biofilms as model system, we show in the present study that formation of heterogeneous biofilms may occur through mechanisms that involve complex subpopulation interactions. One example of this phenomenon is expression of the iron...

  4. Silver against Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Kirketerp-Møller, K.; Kristiansen, S.

    2007-01-01

    bacteria in both the planktonic and biofilm modes of growth. The action of silver on mature in vitro biofilms of Pseudomonas aeruginosa, a primary pathogen of chronic infected wounds, was investigated. The results show that silver is very effective against mature biofilms of P. aeruginosa......, but that the silver concentration is important. A concentration of 5-10 ig/mL silver sulfadiazine eradicated the biofilm whereas a lower concentration (1 ig/mL) had no effect. The bactericidal concentration of silver required to eradicate the bacterial biofilm was 10-100 times higher than that used to eradicate...... planktonic bacteria. These observations strongly indicate that the concentration of silver in currently available wound dressings is much too low for treatment of chronic biofilm wounds. It is suggested that clinicians and manufacturers of the said wound dressings consider whether they are treating wounds...

  5. A Biofilm Matrix-Associated Protease Inhibitor Protects Pseudomonas aeruginosa from Proteolytic Attack.

    Science.gov (United States)

    Tseng, Boo Shan; Reichhardt, Courtney; Merrihew, Gennifer E; Araujo-Hernandez, Sophia A; Harrison, Joe J; MacCoss, Michael J; Parsek, Matthew R

    2018-04-10

    Pseudomonas aeruginosa produces an extracellular biofilm matrix that consists of nucleic acids, exopolysaccharides, lipid vesicles, and proteins. In general, the protein component of the biofilm matrix is poorly defined and understudied relative to the other major matrix constituents. While matrix proteins have been suggested to provide many functions to the biofilm, only proteins that play a structural role have been characterized thus far. Here we identify proteins enriched in the matrix of P. aeruginosa biofilms. We then focused on a candidate matrix protein, the serine protease inhibitor ecotin (PA2755). This protein is able to inhibit neutrophil elastase, a bactericidal enzyme produced by the host immune system during P. aeruginosa biofilm infections. We show that ecotin binds to the key biofilm matrix exopolysaccharide Psl and that it can inhibit neutrophil elastase when associated with Psl. Finally, we show that ecotin protects both planktonic and biofilm P. aeruginosa cells from neutrophil elastase-mediated killing. This may represent a novel mechanism of protection for biofilms to increase their tolerance against the innate immune response. IMPORTANCE Proteins associated with the extracellular matrix of bacterial aggregates called biofilms have long been suggested to provide many important functions to the community. To date, however, only proteins that provide structural roles have been described, and few matrix-associated proteins have been identified. We developed a method to identify matrix proteins and characterized one. We show that this protein, when associated with the biofilm matrix, can inhibit a bactericidal enzyme produced by the immune system during infection and protect biofilm cells from death induced by the enzyme. This may represent a novel mechanism of protection for biofilms, further increasing their tolerance against the immune response. Together, our results are the first to show a nonstructural function for a confirmed matrix

  6. Effects of ginseng on Pseudomonas aeruginosa motility and biofilm formation

    DEFF Research Database (Denmark)

    Wu, Hong; Lee, Baoleri; Yang, Liang

    2011-01-01

    protected animal models from developing chronic lung infection by P. aeruginosa. In the present study, the effects of ginseng on the formation of P. aeruginosa biofilms were further investigated in vitro and in vivo. Ginseng aqueous extract at concentrations of 0.5-2.0% did not inhibit the growth of P......Biofilm-associated chronic Pseudomonas aeruginosa lung infections in patients with cystic fibrosis are virtually impossible to eradicate with antibiotics because biofilm-growing bacteria are highly tolerant to antibiotics and host defense mechanisms. Previously, we found that ginseng treatments....... aeruginosa, but significantly prevented P. aeruginosa from forming biofilm. Exposure to 0.5% ginseng aqueous extract for 24 h destroyed most 7-day-old mature biofilms formed by both mucoid and nonmucoid P. aeruginosa strains. Ginseng treatment enhanced swimming and twitching motility, but reduced swarming...

  7. Pseudomonas aeruginosa Biofilm, a Programmed Bacterial Life for Fitness.

    Science.gov (United States)

    Lee, Keehoon; Yoon, Sang Sun

    2017-06-28

    A biofilm is a community of microbes that typically inhabit on surfaces and are encased in an extracellular matrix. Biofilms display very dissimilar characteristics to their planktonic counterparts. Biofilms are ubiquitous in the environment and influence our lives tremendously in both positive and negative ways. Pseudomonas aeruginosa is a bacterium known to produce robust biofilms. P. aeruginosa biofilms cause severe problems in immunocompromised patients, including those with cystic fibrosis or wound infection. Moreover, the unique biofilm properties further complicate the eradication of the biofilm infection, leading to the development of chronic infections. In this review, we discuss the history of biofilm research and general characteristics of bacterial biofilms. Then, distinct features pertaining to each stage of P. aeruginosa biofilm development are highlighted. Furthermore, infections caused by biofilms on their own or in association with other bacterial species ( i.e. , multispecies biofilms) are discussed in detail.

  8. Complement activation by Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Jensen, E T; Kharazmi, A; Garred, P

    1993-01-01

    In chronic infections, such as the bronchopulmonary Pseudomonas aeruginosa infection in cystic fibrosis (CF) patients, bacteria persist despite an intact host immune defense and frequent antibiotic treatment. An important reason for the persistence of the bacteria is their capacity for the biofilm...... mode of growth. In this study we investigated the role of biofilms in activation of complement, a major contributor to the inflammatory process. Complement activation by P. aeruginosa was examined in a complement consumption assay, production of C3 and factor B conversion products assessed by crossed...... immuno-electrophoresis, C5a generation tested by a PMN chemotactic assay, and terminal complement complex formation measured by ELISA. Two of the four assays showed that P. aeruginosa grown in biofilm activated complement less than planktonic bacteria, and all assays showed that activation by intact...

  9. Cell death in Pseudomonas aeruginosa biofilm development

    DEFF Research Database (Denmark)

    Webb, J.S.; Thompson, L.S.; James, S.

    2003-01-01

    Bacteria growing in biofilms often develop multicellular, three-dimensional structures known as microcolonies. Complex differentiation within biofilms of Pseudomonas aeruginosa occurs, leading to the creation of voids inside microcolonies and to the dispersal of cells from within these voids....... However, key developmental processes regulating these events are poorly understood. A normal component of multicellular development is cell death. Here we report that a repeatable pattern of cell death and lysis occurs in biofilms of P. aeruginosa during the normal course of development. Cell death...... occurred with temporal and spatial organization within biofilms, inside microcolonies, when the biofilms were allowed to develop in continuous-culture flow cells. A subpopulation of viable cells was always observed in these regions. During the onset of biofilm killing and during biofilm development...

  10. Effects of Iron on DNA Release and Biofilm Development by Pseudomonas Aeruginosa

    DEFF Research Database (Denmark)

    Yang, Liang; Barken, Kim Bundvig; Skindersø, Mette Elena

    2007-01-01

    Extracellular DNA is one of the major matrix components in Pseudomonas aeruginosa biofilms. It functions as an intercellular connector and plays a role in stabilization of the biofilms. Evidence that DNA release in P. aeruginosa PAO1 biofilms is controlled by the las-rhl and pqs quorum-sensing sy......Extracellular DNA is one of the major matrix components in Pseudomonas aeruginosa biofilms. It functions as an intercellular connector and plays a role in stabilization of the biofilms. Evidence that DNA release in P. aeruginosa PAO1 biofilms is controlled by the las-rhl and pqs quorum......-sensing systems has been previously presented. This paper provides evidence that DNA release in P. aeruginosa PAO1 biofilms is also under iron regulation. Experiments involving cultivation of P. aeruginosa in microtitre trays suggested that pqs expression, DNA release and biofilm formation were favoured in media...

  11. Pseudomonas aeruginosa biofilm infections

    DEFF Research Database (Denmark)

    Tolker-Nielsen, Tim

    2014-01-01

    Bacteria in natural, industrial and clinical settings predominantly live in biofilms, i.e., sessile structured microbial communities encased in self-produced extracellular matrix material. One of the most important characteristics of microbial biofilms is that the resident bacteria display...... a remarkable increased tolerance toward antimicrobial attack. Biofilms formed by opportunistic pathogenic bacteria are involved in devastating persistent medical device-associated infections, and chronic infections in individuals who are immune-compromised or otherwise impaired in the host defense. Because...... the use of conventional antimicrobial compounds in many cases cannot eradicate biofilms, there is an urgent need to develop alternative measures to combat biofilm infections. The present review is focussed on the important opportunistic pathogen and biofilm model organism Pseudomonas aeruginosa. Initially...

  12. Biomolecular Mechanisms of Pseudomonas aeruginosa and Escherichia coli Biofilm Formation

    Science.gov (United States)

    Laverty, Garry; Gorman, Sean P.; Gilmore, Brendan F.

    2014-01-01

    Pseudomonas aeruginosa and Escherichia coli are the most prevalent Gram-negative biofilm forming medical device associated pathogens, particularly with respect to catheter associated urinary tract infections. In a similar manner to Gram-positive bacteria, Gram-negative biofilm formation is fundamentally determined by a series of steps outlined more fully in this review, namely adhesion, cellular aggregation, and the production of an extracellular polymeric matrix. More specifically this review will explore the biosynthesis and role of pili and flagella in Gram-negative adhesion and accumulation on surfaces in Pseudomonas aeruginosa and Escherichia coli. The process of biofilm maturation is compared and contrasted in both species, namely the production of the exopolysaccharides via the polysaccharide synthesis locus (Psl), pellicle Formation (Pel) and alginic acid synthesis in Pseudomonas aeruginosa, and UDP-4-amino-4-deoxy-l-arabinose and colonic acid synthesis in Escherichia coli. An emphasis is placed on the importance of the LuxR homologue sdiA; the luxS/autoinducer-II; an autoinducer-III/epinephrine/norepinephrine and indole mediated Quorum sensing systems in enabling Gram-negative bacteria to adapt to their environments. The majority of Gram-negative biofilms consist of polysaccharides of a simple sugar structure (either homo- or heteropolysaccharides) that provide an optimum environment for the survival and maturation of bacteria, allowing them to display increased resistance to antibiotics and predation. PMID:25438014

  13. Biomolecular Mechanisms of Pseudomonas aeruginosa and Escherichia coli Biofilm Formation

    Directory of Open Access Journals (Sweden)

    Garry Laverty

    2014-07-01

    Full Text Available Pseudomonas aeruginosa and Escherichia coli are the most prevalent Gram-negative biofilm forming medical device associated pathogens, particularly with respect to catheter associated urinary tract infections. In a similar manner to Gram-positive bacteria, Gram-negative biofilm formation is fundamentally determined by a series of steps outlined more fully in this review, namely adhesion, cellular aggregation, and the production of an extracellular polymeric matrix. More specifically this review will explore the biosynthesis and role of pili and flagella in Gram-negative adhesion and accumulation on surfaces in Pseudomonas aeruginosa and Escherichia coli. The process of biofilm maturation is compared and contrasted in both species, namely the production of the exopolysaccharides via the polysaccharide synthesis locus (Psl, pellicle Formation (Pel and alginic acid synthesis in Pseudomonas aeruginosa, and UDP-4-amino-4-deoxy-l-arabinose and colonic acid synthesis in Escherichia coli. An emphasis is placed on the importance of the LuxR homologue sdiA; the luxS/autoinducer-II; an autoinducer-III/epinephrine/norepinephrine and indole mediated Quorum sensing systems in enabling Gram-negative bacteria to adapt to their environments. The majority of Gram-negative biofilms consist of polysaccharides of a simple sugar structure (either homo- or heteropolysaccharides that provide an optimum environment for the survival and maturation of bacteria, allowing them to display increased resistance to antibiotics and predation.

  14. 2-Furaldehyde diethyl acetal from tender coconut water (Cocos nucifera) attenuates biofilm formation and quorum sensing-mediated virulence of Chromobacterium violaceum and Pseudomonas aeruginosa.

    Science.gov (United States)

    Sethupathy, Sivasamy; Nithya, Chari; Pandian, Shunmugiah Karutha

    2015-01-01

    The aim of this study was to evaluate the anti-biofilm and quorum sensing inhibitory (QSI) potential of tender coconut water (TCW) against Chromobacterium violaceum and Pseudomonas aeruginosa. TCW significantly inhibited the QS regulated violacein, virulence factors and biofilm production without affecting their growth. qRT-PCR analysis revealed the down-regulation of autoinducer synthase, transcriptional regulator and virulence genes. Mass-spectrometric analysis of a petroleum ether extract of the TCW hydrolyte revealed that 2-furaldehyde diethyl acetal (2FDA) and palmitic acid (PA) are the major compounds. In vitro bioassays confirmed the ability of 2FDA to inhibit the biofilm formation and virulence factors. In addition, the combination of PA with 2FDA resulted in potent inhibition of biofilm formation and virulence factors. The results obtained strongly suggest that TCW can be exploited as a base for designing a novel antipathogenic drug formulation to treat biofilm mediated infections caused by P. aeruginosa.

  15. Extracellular DNA Shields against Aminoglycosides in Pseudomonas aeruginosa Biofilms

    DEFF Research Database (Denmark)

    Chiang, Wen-Chi; Nilsson, Martin; Jensen, Peter Østrup

    2013-01-01

    Within recent years, it has been established that extracellular DNA is a key constituent of the matrix of microbial biofilms. In addition, it has recently been demonstrated that DNA binds positively charged antimicrobials such as aminoglycosides and antimicrobial peptides. In the present study, we...... provide evidence that extracellular DNA shields against aminoglycosides in Pseudomonas aeruginosa biofilms. We show that exogenously supplemented DNA integrates into P. aeruginosa biofilms and increases their tolerance toward aminoglycosides. We provide evidence that biofilms formed by a DNA release......-deficient P. aeruginosa quorum-sensing mutant are more susceptible to aminoglycoside treatment than wild-type biofilms but become rescued from the detrimental action of aminoglycosides upon supplementation with exogenous DNA. Furthermore, we demonstrate that exposure to lysed polymorphonuclear leukocytes...

  16. Multiple roles of biosurfactants in structural biofilm development by Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Pamp, Sünje Johanna; Tolker-Nielsen, Tim

    2007-01-01

    Recent studies have indicated that biosurfactants produced by Pseudomonas aeruginosa play a role both in maintaining channels between multicellular structures in biofilms and in dispersal of cells from biofilms. Through the use of flow cell technology and enhanced confocal laser scanning microscopy......, we have obtained results which suggest that the biosurfactants produced by P. aeruginosa play additional roles in structural biofilm development. We present genetic evidence that during biofilm development by P. aeruginosa, biosurfactants promote microcolony formation in the initial phase...... and facilitate migration-dependent structural development in the later phase. P. aeruginosa rhl4 mutants, deficient in synthesis of biosurfactants, were not capable of forming microcolonies in the initial phase of biofilm formation. Experiments involving two-color-coded mixed-strain biofilms showed that P...

  17. Pseudomonas aeruginosa forms Biofilms in Acute InfectionIndependent of Cell-to-Cell Signaling

    Energy Technology Data Exchange (ETDEWEB)

    Schaber, J. Andy; Triffo, W.J.; Suh, Sang J.; Oliver, Jeffrey W.; Hastert, Mary C.; Griswold, John A.; Auer, Manfred; Hamood, Abdul N.; Rumbaugh, Kendra P.

    2006-09-20

    Biofilms are bacterial communities residing within a polysaccharide matrix that are associated with persistence and antibiotic resistance in chronic infections. We show that the opportunistic pathogen Pseudomonas aeruginosa forms biofilms within 8 hours of infection in thermally-injured mice, demonstrating that biofilms contribute to bacterial colonization in acute infections. P. aeruginosa biofilms were visualized within burned tissue surrounding blood vessels and adipose cells. Although quorum sensing (QS), a bacterial signaling mechanism, coordinates differentiation of biofilms in vitro, wild type and QS-deficient P. aeruginosa formed similar biofilms in vivo. Our findings demonstrate that P. aeruginosa forms biofilms on specific host tissues independent of QS.

  18. Interference of Pseudomonas aeruginosa signalling and biofilm formation for infection control

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Tolker-Nielsen, Tim; Høiby, Niels

    2010-01-01

    Pseudomonas aeruginosa is the best described bacterium with regards to quorum sensing (QS), in vitro biofilm formation and the development of antibiotic tolerance. Biofilms composed of P. aeruginosa are thought to be the underlying cause of many chronic infections, including those in wounds...... and in the lungs of patients with cystic fibrosis. In this review, we provide an overview of the molecular mechanisms involved in QS, QS-enabled virulence, biofilm formation and biofilm-enabled antibiotic tolerance. We now have substantial knowledge of the multicellular behaviour of P. aeruginosa in vitro. A major...

  19. Comparison of UVB and UVC irradiation disinfection efficacies on Pseudomonas Aeruginosa (P. aeruginosa) biofilm

    Science.gov (United States)

    Argyraki, A.; Markvart, M.; Nielsen, Anne; Bjarnsholt, T.; Bjørndal, L.; Petersen, P. M.

    2016-04-01

    Disinfection routines are important in all clinical applications. The uprising problem of antibiotic resistance has driven major research efforts towards alternative disinfection approaches, involving light-based solutions. Pseudomonas aeruginosa (P. aeruginosa) is a common bacterium that can cause skin, soft tissue, lungs, kidney and urinary tract infections. Moreover, it can be found on and in medical equipment causing often cross infections in hospitals. The objective of this study was to test the efficiency, of two different light-based disinfection treatments, namely UVB and UVC irradiation, on P. aeruginosa biofilms at different growth stages. In our experiments a new type of UV light emitting diodes (LEDs) were used to deliver UV irradiation on the biofilms, in the UVB (296nm) and UVC (266nm) region. The killing rate was studied as a function of dose for 24h grown biofilms. The dose was ramped from 72J/m2 to 10000J/m2. It was shown that UVB irradiation was more effective than UVC irradiation in inactivating P. aeruginosa biofilms. No colony forming units (CFU) were observed for the UVB treated biofilms when the dose was 10000 J/m2 (CFU in control sample: 7.5 x 104). UVB irradiation at a dose of 20000J/m2 on mature biofilms (72h grown) resulted in a 3.9 log killing efficacy. The fact that the wavelength of 296nm exists in daylight and has such disinfection ability on biofilms gives new perspectives for applications within disinfection at hospitals.

  20. Enzyme-mediated quenching of the Pseudomonas quinolone signal (PQS promotes biofilm formation of Pseudomonas aeruginosa by increasing iron availability

    Directory of Open Access Journals (Sweden)

    Beatrix Tettmann

    2016-12-01

    Full Text Available The 2-alkyl-3-hydroxy-4(1H-quinolone 2,4-dioxygenase HodC was previously described to cleave the Pseudomonas quinolone signal, PQS, which is exclusively used in the complex quorum sensing (QS system of Pseudomonas aeruginosa, an opportunistic pathogen employing QS to regulate virulence and biofilm development. Degradation of PQS by exogenous addition of HodC to planktonic cells of P. aeruginosa attenuated production of virulence factors, and reduced virulence in planta. However, proteolytic cleavage reduced the efficacy of HodC. Here, we identified the secreted protease LasB of P. aeruginosa to be responsible for HodC degradation. In static biofilms of the P. aeruginosa PA14 lasB::Tn mutant, the catalytic activity of HodC led to an increase in viable biomass in newly formed but also in established biofilms, and reduced the expression of genes involved in iron metabolism and siderophore production, such as pvdS, pvdL, pvdA and pvdQ. This is likely due to an increase in the levels of bioavailable iron by degradation of PQS, which is able to sequester iron from the surrounding environment. Thus, HodC, despite its ability to quench the production of virulence factors, is contraindicated for combating P. aeruginosa biofilms.

  1. Quorum-Quenching Human Designer Cells for Closed-Loop Control of Pseudomonas aeruginosa Biofilms.

    Science.gov (United States)

    Sedlmayer, Ferdinand; Jaeger, Tina; Jenal, Urs; Fussenegger, Martin

    2017-08-09

    Current antibiotics gradually lose their efficacy against chronic Pseudomonas aeruginosa infections due to development of increased resistance mediated by biofilm formation, as well as the large arsenal of microbial virulence factors that are coordinated by the cell density-dependent phenomenon of quorum sensing. Here, we address this issue by using synthetic biology principles to rationally engineer quorum-quencher cells with closed-loop control to autonomously dampen virulence and interfere with biofilm integrity. Pathogen-derived signals dynamically activate a synthetic mammalian autoinducer sensor driving downstream expression of next-generation anti-infectives. Engineered cells were able to sensitively score autoinducer levels from P. aeruginosa clinical isolates and mount a 2-fold defense consisting of an autoinducer-inactivating enzyme to silence bacterial quorum sensing and a bipartite antibiofilm effector to dissolve the biofilm matrix. The self-guided cellular device fully cleared autoinducers, potentiated bacterial antibiotic susceptibility, substantially reduced biofilms, and alleviated cytotoxicity to lung epithelial cells. We believe this strategy of dividing otherwise coordinated pathogens and breaking up their shielded stronghold represents a blueprint for cellular anti-infectives in the postantibiotic era.

  2. A characterization of DNA release in Pseudomonas aeruginosa cultures and biofilms

    DEFF Research Database (Denmark)

    Allesen-Holm, Marie; Barken, Kim Bundvig; Yang, Liang

    2006-01-01

    -type P. aeruginosa biofilms stained with different DNA stains suggested that the extracellular DNA is located primarily in the stalks of mushroom-shaped multicellular structures, with a high concentration especially in the outer part of the stalks forming a border between the stalk-forming bacteria...... to whole-genome DNA. Evidence that the extracellular DNA in P. aeruginosa biofilms and cultures is generated via lysis of a subpopulation of the bacteria was obtained through experiments where extracellular beta-galactosidase released from lacZ-containing P. aeruginosa strains was assessed. Experiments...... and the cap-forming bacteria. Biofilms formed by lasIrhlI, pqsA and fliMpilA mutants contained less extracellular DNA than biofilms formed by the wild type, and the mutant biofilms were more susceptible to treatment with sodium dodecyl sulphate than the wild-type biofilm....

  3. Stratified growth in Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Werner, E.; Roe, F.; Bugnicourt, A.

    2004-01-01

    In this study, stratified patterns of protein synthesis and growth were demonstrated in Pseudomonas aeruginosa biofilms. Spatial patterns of protein synthetic activity inside biofilms were characterized by the use of two green fluorescent protein (GFP) reporter gene constructs. One construct...... synthesis was restricted to a narrow band in the part of the biofilm adjacent to the source of oxygen. The zone of active GFP expression was approximately 60 Am wide in colony biofilms and 30 Am wide in flow cell biofilms. The region of the biofilm in which cells were capable of elongation was mapped...... by treating colony biofilms with carbenicillin, which blocks cell division, and then measuring individual cell lengths by transmission electron microscopy. Cell elongation was localized at the air interface of the biofilm. The heterogeneous anabolic patterns measured inside these biofilms were likely a result...

  4. [The effect of biyuanshu oral liquid on the formation of Pseudomonas aeruginosa biofilms in vitro].

    Science.gov (United States)

    Liu, Xiang; Chen, Haihong; Wang, Shengqing

    2012-07-01

    To observe the effect of biyuanshu oral liquid on the formation of pseudomonas aeruginosa biofilms in vitro. Pseudomonas aeruginosa biofilm was established by plate culture and detected by Scanning electron microscopy and AgNO3 staining. After treated with different dosages of biyuanshu oral liquid and erythromycin, the pseudomonas aeruginosa biofilms were observed by AgNO3 staining and the number of viable bacteria were measured by serial dilution. The pseudomonas aeruginosa biofilms could be detected by SEM at the seventh culture day and it was consistent with the detection of AgNO3 staining. The biyuanshu oral liquid and erythromycin have the effect on inhibiting the formation of pseudomonas aeruginosa biofilms. But with the already formed pseudomonas aeruginosa biofilms the inhibition was not significant. The serial dilution method showed that the viable counts of bacteria of biyuanshu oral liquid and erythromycin treated groups were significantly lower than those untreated groups (P formation of pseudomonas aeruginosa biofilms in vitro.

  5. N-acylhomoserine-lactone-mediated communication between Pseudomonas aeruginosa and Burkholderia cepacia in mixed biofilms

    DEFF Research Database (Denmark)

    Riedel, K.; Hentzer, Morten; Geisenberger, O.

    2001-01-01

    Pseudomonas aeruginosa and Burkholderia cepacia are capable of forming mixed biofilms in the lungs of cystic fibrosis patients. Both bacteria employ quorum-sensing systems, which rely on N-acylhomoserine lactone (AHL) signal molecules, to co- ordinate expression of virulence factors with the form...

  6. Colistin-Tobramycin Combinations Are Superior to Monotherapy Concerning the Killing of Biofilm Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Herrmann, G.; Yang, Liang; Wu, H.

    2010-01-01

    Background. Antibiotic combination therapy might be more efficient than single antibiotics to combat Pseudomonas aeruginosa biofilms in the airways of patients with cystic fibrosis. We tested the ability of colistin sulphatetobramycin combinations and single antibiotics to kill P. aeruginosa...... biofilms. Methods. P. aeruginosa biofilms were generated in vitro and in rat lungs. In a pilot study, 5 patients with cystic fibrosis inhaled colistin and then tobramycin for 4 weeks. The changes in P. aeruginosa counts and lung function were assessed before and after therapy. Results. Antibiotic...... combination therapy significantly reduced the number of P. aeruginosa cells in P. aeruginosa biofilm models in vitro. When rats were challenged with 1 x 10(7) cfu of P. aeruginosa, which was embedded in alginate beads, mortality rates, lung pathologic findings, and bacterial colony-forming unit counts were...

  7. Pseudomonas aeruginosa PAO1 exopolysaccharides are important for mixed species biofilm community development and stress tolerance

    Directory of Open Access Journals (Sweden)

    Saravanan ePeriasamy

    2015-08-01

    Full Text Available Pseudomonas aeruginosa PAO1 produces three polysaccharides, alginate, Psl and Pel that play distinct roles in attachment and biofilm formation for monospecies biofilms. Considerably less is known about their role in the development of mixed species biofilm communities. This study has investigated the roles of alginate, Psl and Pel during biofilm formation of P. aeruginosa in a defined and experimentally informative mixed species biofilm community, consisting of P. aeruginosa, Pseudomonas protegens and Klebsiella pneumoniae. Loss of the Psl polysaccharide had the biggest impact on the integration of P. aeruginosa in the mixed species biofilms, where the percent composition of the psl mutant was significantly lower (0.06% than its wild-type parent (2.44%. In contrast, loss of the Pel polysaccharide had no impact on mixed species biofilm development. Loss of alginate or its overproduction resulted in P. aeruginosa representing 8.4% and 18.11%, respectively, of the mixed species biofilm. Dual species biofilms of P. aeruginosa and K. pneumoniae were not affected by loss of alginate, Pel or Psl, while the mucoid P. aeruginosa strain achieved a greater biomass than its parent strain. When P. aeruginosa was grown with P. protegens, loss of the Pel or alginate polysaccharides resulted in biofilms that were not significantly different from biofilms formed by the wild-type PAO1. In contrast, overproduction of alginate resulted in biofilms that were comprised of 35-40% of P. aeruginosa, which was significantly higher than the wild-type (5-20%. Loss of the Psl polysaccharide significantly reduced the percentage composition of P. aeruginosa in dual species biofilms with P. protegens (<1%. Loss of the Psl polysaccharide significantly disrupted the communal stress resistance of the three species biofilms. Thus, the polysaccharide composition of an individual species significantly impacts mixed species biofilm development and the emergent properties of such

  8. An update on Pseudomonas aeruginosa biofilm formation, tolerance, and dispersal

    DEFF Research Database (Denmark)

    Harmsen, Morten; Yang, Liang; Pamp, Sünje Johanna

    2010-01-01

    We review the recent advances in the understanding of the Pseudomonas aeruginosa biofilm lifestyle from studies using in vitro laboratory setups such as flow chambers and microtiter trays. Recent work sheds light on the role of nutrients, motility, and quorum sensing in structure formation in P....... aeruginosa biofilms. The second messenger, c-di-GMP, is established as an important regulator of the synthesis of polysaccharide and protein components of the biofilm matrix. Extracellular DNA is shown to be an essential component of the biofilm matrix. It has become apparent that biofilm formation involves...... interactions between different subpopulations. The molecular mechanisms underlying the tolerance of biofilm bacteria to antimicrobial agents are beginning to be unraveled, and new knowledge has been obtained regarding the environmental cues and regulatory mechanisms involved in biofilm dispersal....

  9. Spatial transcriptomes within the Pseudomonas aeruginosa biofilm architecture.

    Science.gov (United States)

    Heacock-Kang, Yun; Sun, Zhenxin; Zarzycki-Siek, Jan; McMillan, Ian A; Norris, Michael H; Bluhm, Andrew P; Cabanas, Darlene; Fogen, Dawson; Vo, Hung; Donachie, Stuart P; Borlee, Bradley R; Sibley, Christopher D; Lewenza, Shawn; Schurr, Michael J; Schweizer, Herbert P; Hoang, Tung T

    2017-12-01

    Bacterial cooperative associations and dynamics in biofilm microenvironments are of special interest in recent years. Knowledge of localized gene-expression and corresponding bacterial behaviors within the biofilm architecture at a global scale has been limited, due to a lack of robust technology to study limited number of cells in stratified layers of biofilms. With our recent pioneering developments in single bacterial cell transcriptomic analysis technology, we generated herein an unprecedented spatial transcriptome map of the mature in vitro Pseudomonas aeruginosa biofilm model, revealing contemporaneous yet altered bacterial behaviors at different layers within the biofilm architecture (i.e., surface, middle and interior of the biofilm). Many genes encoding unknown functions were highly expressed at the biofilm-solid interphase, exposing a critical gap in the knowledge of their activities that may be unique to this interior niche. Several genes of unknown functions are critical for biofilm formation. The in vivo importance of these unknown proteins was validated in invertebrate (fruit fly) and vertebrate (mouse) models. We envisage the future value of this report to the community, in aiding the further pathophysiological understanding of P. aeruginosa biofilms. Our approach will open doors to the study of bacterial functional genomics of different species in numerous settings. © 2017 The Authors. Molecular Microbiology Published by John Wiley & Sons Ltd.

  10. Inhibition of quorum sensing in Pseudomonas aeruginosa biofilm bacteria by a halogenated furanone compound

    DEFF Research Database (Denmark)

    Hentzer, Morten; Riedel, K.; Rasmussen, Thomas Bovbjerg

    2002-01-01

    Novel molecular tools have been constructed which allow for in situ detection of N-acyl homoserine lactone (AHL)-mediated quorum sensing in Pseudomonas aeruginosa biofilms. The reporter responds to AHL activation of LasR lay expression of an unstable version of the green-fluorescent protein (Gfp...

  11. Targeting quorum sensing in Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Jakobsen, Tim Holm; Bjarnsholt, Thomas; Jensen, Peter Østrup

    2013-01-01

    Bacterial resistance to conventional antibiotics combined with an increasing acknowledgement of the role of biofilms in chronic infections has led to a growing interest in new antimicrobial strategies that target the biofilm mode of growth. In the aggregated biofilm mode, cell-to-cell communication...... alternative antibacterial strategies. Here, we review state of the art research of quorum sensing inhibitors against the opportunistic human pathogen Pseudomonas aeruginosa, which is found in a number of biofilm-associated infections and identified as the predominant organism infecting the lungs of cystic...

  12. Novel Targets for Treatment of Pseudomonas aeruginosa Biofilms

    DEFF Research Database (Denmark)

    Alhede, Morten; Alhede, Maria; Bjarnsholt, Thomas

    2014-01-01

    Pseudomonas aeruginosa causes infection in all parts of the human body. The bacterium is naturally resistant to a wide range of antibiotics. In addition to resistance mechanisms such as efflux pumps, the ability to form aggregates, known as biofilm, further reduces Pseudomonas aeruginosa...

  13. Tolerance of Pseudomonas aeruginosa in in-vitro biofilms to high-level peracetic acid disinfection.

    Science.gov (United States)

    Akinbobola, A B; Sherry, L; Mckay, W G; Ramage, G; Williams, C

    2017-10-01

    Biofilm has been suggested as a cause of disinfection failures in flexible endoscopes where no lapses in the decontamination procedure can be identified. To test this theory, the activity of peracetic acid, one of the widely used disinfectants in the reprocessing of flexible endoscopes, was evaluated against both planktonic and sessile communities of Pseudomonas aeruginosa. To investigate the ability of P. aeruginosa biofilm to survive high-level peracetic acid disinfection. The susceptibility of planktonic cells of P. aeruginosa and biofilms aged 24, 48, 96, and 192 h to peracetic acid was evaluated by estimating their viability using resazurin viability and plate count methods. The biomass of the P. aeruginosa biofilms was also quantified using Crystal Violet assay. Planktonic cells of P. aeruginosa were treated with 5-30 ppm concentration of peracetic acid in the presence of 3.0 g/L of bovine serum albumin (BSA) for 5 min. Biofilms of P. aeruginosa were also treated with various peracetic acid concentrations (100-3000 ppm) for 5 min. Planktonic cells of P. aeruginosa were eradicated by 20 ppm of peracetic acid, whereas biofilms showed an age-dependent tolerance to peracetic acid, and 96 h biofilm was only eradicated at peracetic acid concentration of 2500 ppm. Ninety-six-hour P. aeruginosa biofilm survives 5 min treatment with 2000 ppm of peracetic acid, which is the working concentration used in some endoscope washer-disinfectors. This implies that disinfection failure of flexible endoscopes might occur when biofilms build up in the lumens of endoscopes. Copyright © 2017. Published by Elsevier Ltd.

  14. Biofilm Filtrates of Pseudomonas aeruginosa Strains Isolated from Cystic Fibrosis Patients Inhibit Preformed Aspergillus fumigatus Biofilms via Apoptosis.

    Science.gov (United States)

    Shirazi, Fazal; Ferreira, Jose A G; Stevens, David A; Clemons, Karl V; Kontoyiannis, Dimitrios P

    2016-01-01

    Pseudomonas aeruginosa (Pa) and Aspergillus fumigatus (Af) colonize cystic fibrosis (CF) patient airways. Pa culture filtrates inhibit Af biofilms, and Pa non-CF, mucoid (Muc-CF) and nonmucoid CF (NMuc-CF) isolates form an ascending inhibitory hierarchy. We hypothesized this activity is mediated through apoptosis induction. One Af and three Pa (non-CF, Muc-CF, NMuc-CF) reference isolates were studied. Af biofilm was formed in 96 well plates for 16 h ± Pa biofilm filtrates. After 24 h, apoptosis was characterized by viability dye DiBAc, reactive oxygen species (ROS) generation, mitochondrial membrane depolarization, DNA fragmentation and metacaspase activity. Muc-CF and NMuc-CF filtrates inhibited and damaged Af biofilm (pbiofilms (3.7- fold) compared to treatment with filtrates from Muc-CF- (2.5- fold) or non-CF Pa (1.7- fold). Depolarization of mitochondrial potential was greater upon exposure to NMuc-CF (2.4-fold) compared to Muc-CF (1.8-fold) or non-CF (1.25-fold) (pbiofilm, compared to control, mediated by metacaspase activation. In conclusion, filtrates from CF-Pa isolates were more inhibitory against Af biofilms than from non-CF. The apoptotic effect involves mitochondrial membrane damage associated with metacaspase activation.

  15. [Effect of Pseudomonas aeruginosa exometabolites on planktonic and biofilm cultures of Escherichia coli].

    Science.gov (United States)

    Kuznetsova, M V; Karpunina, T I; Maslennikova, I L; Nesterova, L Iu; Demakov, V A

    2012-01-01

    Study the effect of P. aeruginosa exometabolites on planktonic and biofilm cultures of bioluminescent E. coli strain. E. coli K12 TG1 (pF1 lux+ Ap(r)) recombinant bioluminescent strain, P. aeruginosa ATCC 27853 reference strain and 2 nosocomial isolates were used. Pyocyanin and pyoverdin content in supernatant of P. aeruginosa over-night cultures was evaluated according to E. Deziel et al. (2001). Planktonic and biofilm cultures of E. coli were obtained in 96-well plates (LB, statically, 37 degrees C), optical density of plankton, film biomass (OD600, OD580) and bioluminescence in plankton and biofilm were evaluated in microplate reader Infiniti M200 (Tecan, Austria). P. aeruginosa exometabolites increased the duration of lag-phase in E. coli, and short term exposition inhibited luminescence of planktonic cells. These effects are determined by bactericidal action ofpyocyanin and pyoverdin. Supernatants ofover-night cultures of P. aeruginosa inhibit formation of biofilm and disrupt the formed biofilm of E. coli. Effect of pyocyanin and pyoverdin on these processes is not established, other factors may have higher significance. Bioluminescence of E. coli K12 TGI that reflects the energetic status of the cell allows to evaluate and prognose the character of coexistence of P. aeruginosa in combined with E. coli planktonic and biofilm culture.

  16. The MerR-like regulator BrlR confers biofilm tolerance by activating multidrug efflux pumps in Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Liao, Julie; Schurr, Michael J; Sauer, Karin

    2013-08-01

    A defining characteristic of biofilms is antibiotic tolerance that can be up to 1,000-fold greater than that of planktonic cells. In Pseudomonas aeruginosa, biofilm tolerance to antimicrobial agents requires the biofilm-specific MerR-type transcriptional regulator BrlR. However, the mechanism by which BrlR mediates biofilm tolerance has not been elucidated. Genome-wide transcriptional profiling indicated that brlR was required for maximal expression of genes associated with antibiotic resistance, in particular those encoding the multidrug efflux pumps MexAB-OprM and MexEF-OprN. Chromatin immunoprecipitation (ChIP) analysis revealed a direct regulation of these genes by BrlR, with DNA binding assays confirming BrlR binding to the promoter regions of the mexAB-oprM and mexEF-oprN operons. Quantitative reverse transcriptase PCR (qRT-PCR) analysis further indicated BrlR to be an activator of mexAB-oprM and mexEF-oprN gene expression. Moreover, immunoblot analysis confirmed increased MexA abundance in cells overexpressing brlR. Inactivation of both efflux pumps rendered biofilms significantly more susceptible to five different classes of antibiotics by affecting MIC but not the recalcitrance of biofilms to killing by bactericidal agents. Overexpression of either efflux pump in a ΔbrlR strain partly restored tolerance of ΔbrlR biofilms to antibiotics. Expression of brlR in mutant biofilms lacking both efflux pumps partly restored antimicrobial tolerance of biofilms to wild-type levels. Our results indicate that BrlR acts as an activator of multidrug efflux pumps to confer tolerance to P. aeruginosa biofilms and to resist the action of antimicrobial agents.

  17. C-di-GMP regulates Pseudomonas aeruginosa stress response to tellurite during both planktonic and biofilm modes of growth

    DEFF Research Database (Denmark)

    Chua, Song Lin; Sivakumar, Krishnakumar; Rybtke, Morten Levin

    2015-01-01

    tellurite (TeO3(2-)) exposure induced the intracellular content of the secondary messenger cyclic di-GMP (c-di-GMP) of Pseudomonas aeruginosa. Two diguanylate cyclases (DGCs), SadC and SiaD, were responsible for the increased intracellular content of c-di-GMP. Enhanced c-di-GMP levels by TeO3(2-) further...... increased P. aeruginosa biofilm formation and resistance to TeO3(2-). P. aeruginosa ΔsadCΔsiaD and PAO1/p(lac)-yhjH mutants with low intracellular c-di-GMP content were more sensitive to TeO3(2-) exposure and had low relative fitness compared to the wild-type PAO1 planktonic and biofilm cultures exposed...... to TeO3(2-). Our study provided evidence that c-di-GMP level can play an important role in mediating stress response in microbial communities during both planktonic and biofilm modes of growth....

  18. Assembly and development of the Pseudomonas aeruginosa biofilm matrix.

    Directory of Open Access Journals (Sweden)

    Luyan Ma

    2009-03-01

    Full Text Available Virtually all cells living in multicellular structures such as tissues and organs are encased in an extracellular matrix. One of the most important features of a biofilm is the extracellular polymeric substance that functions as a matrix, holding bacterial cells together. Yet very little is known about how the matrix forms or how matrix components encase bacteria during biofilm development. Pseudomonas aeruginosa forms environmentally and clinically relevant biofilms and is a paradigm organism for the study of biofilms. The extracellular polymeric substance of P. aeruginosa biofilms is an ill-defined mix of polysaccharides, nucleic acids, and proteins. Here, we directly visualize the product of the polysaccharide synthesis locus (Psl exopolysaccharide at different stages of biofilm development. During attachment, Psl is anchored on the cell surface in a helical pattern. This promotes cell-cell interactions and assembly of a matrix, which holds bacteria in the biofilm and on the surface. Chemical dissociation of Psl from the bacterial surface disrupted the Psl matrix as well as the biofilm structure. During biofilm maturation, Psl accumulates on the periphery of 3-D-structured microcolonies, resulting in a Psl matrix-free cavity in the microcolony center. At the dispersion stage, swimming cells appear in this matrix cavity. Dead cells and extracellular DNA (eDNA are also concentrated in the Psl matrix-free area. Deletion of genes that control cell death and autolysis affects the formation of the matrix cavity and microcolony dispersion. These data provide a mechanism for how P. aeruginosa builds a matrix and subsequently a cavity to free a portion of cells for seeding dispersal. Direct visualization reveals that Psl is a key scaffolding matrix component and opens up avenues for therapeutics of biofilm-related complications.

  19. Investigating the link between imipenem resistance and biofilm formation by Pseudomonas aeruginosa.

    Science.gov (United States)

    Musafer, Hadeel K; Kuchma, Sherry L; Naimie, Amanda A; Schwartzman, Joseph D; Al-Mathkhury, Harith J Fahad; O'Toole, George A

    2014-07-01

    Pseudomonas aeruginosa, a ubiquitous environmental organism, is a difficult-to-treat opportunistic pathogen due to its broad-spectrum antibiotic resistance and its ability to form biofilms. In this study, we investigate the link between resistance to a clinically important antibiotic, imipenem, and biofilm formation. First, we observed that the laboratory strain P. aeruginosa PAO1 carrying a mutation in the oprD gene, which confers resistance to imipenem, showed a modest reduction in biofilm formation. We also observed an inverse relationship between imipenem resistance and biofilm formation for imipenem-resistant strains selected in vitro, as well as for clinical isolates. We identified two clinical isolates of P. aeruginosa from the sputum of cystic fibrosis patients that formed robust biofilms, but were sensitive to imipenem (MIC ≤ 2 μg/ml). To test the hypothesis that there is a general link between imipenem resistance and biofilm formation, we performed transposon mutagenesis of these two clinical strains to identify mutants defective in biofilm formation, and then tested these mutants for imipenem resistance. Analysis of the transposon mutants revealed a role for previously described biofilm factors in these clinical isolates of P. aeruginosa, including mutations in the pilY1, pilX, pilW, algC, and pslI genes, but none of the biofilm-deficient mutants became imipenem resistant (MIC ≥ 8 μg/ml), arguing against a general link between biofilm formation and resistance to imipenem. Thus, assessing biofilm formation capabilities of environmental isolates is unlikely to serve as a good predictor of imipenem resistance. We also discuss our findings in light of the limited literature addressing planktonic antibiotic resistance factors that impact biofilm formation.

  20. Evolution and adaptation in Pseudomonas aeruginosa biofilms driven by mismatch repair system-deficient mutators.

    Directory of Open Access Journals (Sweden)

    Adela M Luján

    Full Text Available Pseudomonas aeruginosa is an important opportunistic pathogen causing chronic airway infections, especially in cystic fibrosis (CF patients. The majority of the CF patients acquire P. aeruginosa during early childhood, and most of them develop chronic infections resulting in severe lung disease, which are rarely eradicated despite intensive antibiotic therapy. Current knowledge indicates that three major adaptive strategies, biofilm development, phenotypic diversification, and mutator phenotypes [driven by a defective mismatch repair system (MRS], play important roles in P. aeruginosa chronic infections, but the relationship between these strategies is still poorly understood. We have used the flow-cell biofilm model system to investigate the impact of the mutS associated mutator phenotype on development, dynamics, diversification and adaptation of P. aeruginosa biofilms. Through competition experiments we demonstrate for the first time that P. aeruginosa MRS-deficient mutators had enhanced adaptability over wild-type strains when grown in structured biofilms but not as planktonic cells. This advantage was associated with enhanced micro-colony development and increased rates of phenotypic diversification, evidenced by biofilm architecture features and by a wider range and proportion of morphotypic colony variants, respectively. Additionally, morphotypic variants generated in mutator biofilms showed increased competitiveness, providing further evidence for mutator-driven adaptive evolution in the biofilm mode of growth. This work helps to understand the basis for the specific high proportion and role of mutators in chronic infections, where P. aeruginosa develops in biofilm communities.

  1. Cooperative pathogenicity in cystic fibrosis: Stenotrophomonas maltophilia modulates Pseudomonas aeruginosa virulence in mixed biofilm

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    Arianna ePompilio

    2015-09-01

    Full Text Available The present study was undertaken in order to understand more about the interaction occurring between S. maltophilia and P. aeruginosa, which are frequently co-isolated from CF airways. For this purpose, S. maltophilia RR7 and P. aeruginosa RR8 strains, co-isolated from the lung of a chronically infected CF patient during a pulmonary exacerbation episode, were evaluated for reciprocal effect during planktonic growth, adhesion and biofilm formation onto both polystyrene and CF bronchial cell monolayer, motility, as well as for gene expression in mixed biofilms. P. aeruginosa significantly affected S. maltophilia growth in both planktonic and biofilm cultures, due to an inhibitory activity probably requiring direct contact. Conversely, no effect was observed on P. aeruginosa by S. maltophilia. Compared with monocultures, the adhesiveness of P. aeruginosa on CFBE41o- cells was significantly reduced by S. maltophilia, which probably acts by reducing P. aeruginosa's swimming motility. An opposite trend was observed for biofilm formation, confirming the findings obtained using polystyrene. When grown in mixed biofilm with S. maltophilia, P. aeruginosa significantly over-expressed aprA, and algD - codifying for protease and alginate, respectively - while the quorum sensing related rhlR and lasI genes were down-regulated. The induced alginate expression by P. aeruginosa might be responsible for the protection of S. maltophilia against tobramycin activity we observed in mixed biofilms. Taken together, our results suggest that the existence of reciprocal interference of S. maltophilia and P. aeruginosa in CF lung is plausible. In particular, S. maltophilia might confer some selective fitness advantage to P. aeruginosa under the specific conditions of chronic infection or, alternatively, increase the virulence of P. aeruginosa thus leading to pulmonary exacerbation.

  2. Activity of Bacteriophages in Removing Biofilms of Pseudomonas aeruginosa Isolates from Chronic Rhinosinusitis Patients

    Directory of Open Access Journals (Sweden)

    Stephanie A. Fong

    2017-09-01

    Full Text Available Introduction:Pseudomonas aeruginosa infections are prevalent amongst chronic rhinosinusitis (CRS sufferers. Many P. aeruginosa strains form biofilms, leading to treatment failure. Lytic bacteriophages (phages are viruses that infect, replicate within, and lyse bacteria, causing bacterial death.Aim: To assess the activity of a phage cocktail in eradicating biofilms of ex vivo P.aeruginosa isolates from CRS patients.Methods: P. aeruginosa isolates from CRS patients with and without cystic fibrosis (CF across three continents were multi-locus sequence typed and tested for antibiotic resistance. Biofilms grown in vitro were treated with a cocktail of four phages (CT-PA. Biofilm biomass was measured after 24 and 48 h, using a crystal violet assay. Phage titrations were performed to confirm replication of the phages. A linear mixed effects model was applied to assess the effects of treatment, time, CF status, and multidrug resistance on the biomass of the biofilm.Results: The isolates included 44 strain types. CT-PA treatment significantly reduced biofilm biomass at both 24 and 48 h post-treatment (p < 0.0001, regardless of CF status or antibiotic resistance. Biomass was decreased by a median of 76% at 48 h. Decrease in biofilm was accompanied by a rise in phage titres for all except one strain.Conclusion: A single dose of phages is able to significantly reduce biofilms formed in vitro by a range of P.aeruginosa isolates from CRS patients. This represents an exciting potential and novel targeted treatment for P. aeruginosa biofilm infections and multidrug resistant bacteria.

  3. Respiratory syncytial virus infection enhances Pseudomonas aeruginosa biofilm growth through dysregulation of nutritional immunity.

    Science.gov (United States)

    Hendricks, Matthew R; Lashua, Lauren P; Fischer, Douglas K; Flitter, Becca A; Eichinger, Katherine M; Durbin, Joan E; Sarkar, Saumendra N; Coyne, Carolyn B; Empey, Kerry M; Bomberger, Jennifer M

    2016-02-09

    Clinical observations link respiratory virus infection and Pseudomonas aeruginosa colonization in chronic lung disease, including cystic fibrosis (CF) and chronic obstructive pulmonary disease. The development of P. aeruginosa into highly antibiotic-resistant biofilm communities promotes airway colonization and accounts for disease progression in patients. Although clinical studies show a strong correlation between CF patients' acquisition of chronic P. aeruginosa infections and respiratory virus infection, little is known about the mechanism by which chronic P. aeruginosa infections are initiated in the host. Using a coculture model to study the formation of bacterial biofilm formation associated with the airway epithelium, we show that respiratory viral infections and the induction of antiviral interferons promote robust secondary P. aeruginosa biofilm formation. We report that the induction of antiviral IFN signaling in response to respiratory syncytial virus (RSV) infection induces bacterial biofilm formation through a mechanism of dysregulated iron homeostasis of the airway epithelium. Moreover, increased apical release of the host iron-binding protein transferrin during RSV infection promotes P. aeruginosa biofilm development in vitro and in vivo. Thus, nutritional immunity pathways that are disrupted during respiratory viral infection create an environment that favors secondary bacterial infection and may provide previously unidentified targets to combat bacterial biofilm formation.

  4. Induction of beta-lactamase production in Pseudomonas aeruginosa biofilm

    DEFF Research Database (Denmark)

    Giwercman, B; Jensen, E T; Høiby, N

    1991-01-01

    Imipenem induced high levels of beta-lactamase production in Pseudomonas aeruginosa biofilms. Piperacillin also induced beta-lactamase production in these biofilms but to a lesser degree. The combination of beta-lactamase production with other protective properties of the biofilm mode of growth c...... could be a major reason for the persistence of this sessile bacterium in chronic infections....

  5. Biofilm production by clinical isolates of Pseudomonas aeruginosa and structural changes in LasR protein of isolates non biofilm-producing

    Directory of Open Access Journals (Sweden)

    Jailton Lobo da Costa Lima

    2018-03-01

    Full Text Available Introduction: Biofilm production is an important mechanism for the survival of Pseudomonas aeruginosa and its relationship with antimicrobial resistance represents a challenge for patient therapeutics. P. aeruginosa is an opportunistic pathogen frequently associated to nosocomial infections, especially in imunocompromised hosts. Objectives: Analyze the phenotypic biofilm production in P. aeruginosa isolates, describe clonal profiles, and analyze quorum sensing (QS genes and the occurrence of mutations in the LasR protein of non-biofilm producing isolates. Methods: Isolates were tested for biofilm production by measuring cells adherence to the microtiter plates. Clonal profile analysis was carried out through ERIC-PCR, QS genes were by specific PCR. Results: The results showed that 77.5% of the isolates were considered biofilm producers. The results of genotyping showed 38 distinct genetic profiles. As for the occurrence of the genes, 100% of the isolates presented the lasR, rhlI and rhlR genes, and 97.5%, presented the lasI gene. In this study nine isolates were not biofilm producers. However, all presented the QS genes. Amplicons related to genes were sequenced in three of the nine non-biofilm-producing isolates (all presenting different genetic similarity profile and aligned to the sequences of those genes in P. aeruginosa strain PAO1 (standard biofilm-producing strain. Alignment analysis showed an insertion of three nucleotides (T, C and G causing the addition of an amino acid valine in the sequence of the LasR protein, in position 53. Conclusion: The modeling of the resulting LasR protein showed a conformational change in its structure, suggesting that this might be the reason why these isolates are unable to produce biofilm. Keywords: Pseudomonas aeruginosa, Biofilm, Multiresistance, Quorum sensing (QS

  6. Inhibition and dispersal of Agrobacterium tumefaciens biofilms by a small diffusible Pseudomonas aeruginosa exoproduct(s).

    Science.gov (United States)

    Hibbing, Michael E; Fuqua, Clay

    2012-06-01

    Environmental biofilms often contain mixed populations of different species. In these dense communities, competition between biofilm residents for limited nutrients such as iron can be fierce, leading to the evolution of competitive factors that affect the ability of competitors to grow or form biofilms. We have discovered a compound(s) present in the conditioned culture fluids of Pseudomonas aeruginosa that disperses and inhibits the formation of biofilms produced by the facultative plant pathogen Agrobacterium tumefaciens. The inhibitory activity is strongly induced when P. aeruginosa is cultivated in iron-limited conditions, but it does not function through iron sequestration. In addition, the production of the biofilm inhibitory activity is not regulated by the global iron regulatory protein Fur, the iron-responsive extracytoplasmic function σ factor PvdS, or three of the recognized P. aeruginosa quorum-sensing systems. In addition, the compound(s) responsible for the inhibition and dispersal of A. tumefaciens biofilm formation is likely distinct from the recently identified P. aeruginosa dispersal factor, cis-2-decenoic acid (CDA), as dialysis of the culture fluids showed that the inhibitory compound was larger than CDA and culture fluids that dispersed and inhibited biofilm formation by A. tumefaciens had no effect on biofilm formation by P. aeruginosa.

  7. Molecular Determinants of the Thickened Matrix in a Dual-Species Pseudomonas aeruginosa and Enterococcus faecalis Biofilm.

    Science.gov (United States)

    Lee, Keehoon; Lee, Kang-Mu; Kim, Donggeun; Yoon, Sang Sun

    2017-11-01

    Biofilms are microbial communities that inhabit various surfaces and are surrounded by extracellular matrices (ECMs). Clinical microbiologists have shown that the majority of chronic infections are caused by biofilms, following the introduction of the first biofilm infection model by J. W. Costerton and colleagues (J. Lam, R. Chan, K. Lam, and J. W. Costerton, Infect Immun 28:546-556, 1980). However, treatments for chronic biofilm infections are still limited to surgical removal of the infected sites. Pseudomonas aeruginosa and Enterococcus faecalis are two frequently identified bacterial species in biofilm infections; nevertheless, the interactions between these two species, especially during biofilm growth, are not clearly understood. In this study, we observed phenotypic changes in a dual-species biofilm of P. aeruginosa and E. faecalis , including a dramatic increase in biofilm matrix thickness. For clear elucidation of the spatial distribution of the dual-species biofilm, P. aeruginosa and E. faecalis were labeled with red and green fluorescence, respectively. E. faecalis was located at the lower part of the dual-species biofilm, while P. aeruginosa developed a structured biofilm on the upper part. Mutants with altered exopolysaccharide (EPS) productions were constructed in order to determine the molecular basis for the synergistic effect of the dual-species biofilm. Increased biofilm matrix thickness was associated with EPSs, not extracellular DNA. In particular, Pel and Psl contributed to interspecies and intraspecies interactions, respectively, in the dual-species P. aeruginosa and E. faecalis biofilm. Accordingly, targeting Pel and Psl might be an effective part of eradicating P. aeruginosa polymicrobial biofilms. IMPORTANCE Chronic infection is a serious problem in the medical field. Scientists have observed that chronic infections are closely associated with biofilms, and the vast majority of infection-causing biofilms are polymicrobial. Many studies

  8. High-Throughput Genetic Screen Reveals that Early Attachment and Biofilm Formation Are Necessary for Full Pyoverdine Production by Pseudomonas aeruginosa

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    Donghoon Kang

    2017-09-01

    Full Text Available Pseudomonas aeruginosa is a re-emerging, multidrug-resistant, opportunistic pathogen that threatens the lives of immunocompromised patients, patients with cystic fibrosis, and those in critical care units. One of the most important virulence factors in this pathogen is the siderophore pyoverdine. Pyoverdine serves several critical roles during infection. Due to its extremely high affinity for ferric iron, pyoverdine gives the pathogen a significant advantage over the host in their competition for iron. In addition, pyoverdine can regulate the production of multiple bacterial virulence factors and perturb host mitochondrial homeostasis. Inhibition of pyoverdine biosynthesis decreases P. aeruginosa pathogenicity in multiple host models. To better understand the regulation of pyoverdine production, we developed a high-throughput genetic screen that uses the innate fluorescence of pyoverdine to identify genes necessary for its biosynthesis. A substantial number of hits showing severe impairment of pyoverdine production were in genes responsible for early attachment and biofilm formation. In addition to genetic disruption of biofilm, both physical and chemical perturbations also attenuated pyoverdine production. This regulatory relationship between pyoverdine and biofilm is particularly significant in the context of P. aeruginosa multidrug resistance, where the formation of biofilm is a key mechanism preventing access to antimicrobials and the immune system. Furthermore, we demonstrate that the biofilm inhibitor 2-amino-5,6-dimethylbenzimidazole effectively attenuates pyoverdine production and rescues Caenorhabditis elegans from P. aeruginosa-mediated pathogenesis. Our findings suggest that targeting biofilm formation in P. aeruginosa infections may have multiple therapeutic benefits and that employing an unbiased, systems biology-based approach may be useful for understanding the regulation of specific virulence factors and identifying novel anti

  9. Inhibitory effect of zinc oxide nanoparticles on pseudomonas aeruginosa biofilm formation

    Directory of Open Access Journals (Sweden)

    Mohammad Hassani Sangani

    2015-04-01

    Full Text Available Objective(s: Bacterial biofilm formation causes many persistent and chronic infections. The matrix protects biofilm bacteria from exposure to innate immune defenses and antibiotic treatments. The purpose of this study was to evaluate the biofilm formation of clinical isolates of Pseudomonas aeruginosa and the activity of zinc oxide nanoparticles (ZnO NPs on biofilm. Materials and Methods: After collecting bacteria from clinical samples of hospitalized patients, the ability of organisms were evaluated to create biofilm by tissue culture plate (TCP assay. ZnO NPs were synthesized by sol gel method and the efficacy of different concentrations (50- 350 µg/ml of ZnO NPs was assessed on biofilm formation and also elimination of pre-formed biofilm by using TCP method. Results:The average diameter of synthesized ZnO NPs was 20 nm. The minimum inhibitory concentration of nanoparticles was 150- 158 μg/ml and the minimum bactericidal concentration was higher (325 µg/ml. All 15 clinical isolates of P. aeruginosa were able to produce biofilm. Treating the organisms with nanoparticles at concentrations of 350 μg/ml resulted in more than 94% inhibition in OD reduction%. Molecular analysis showed that the presence of mRNA of pslA gene after treating bacteria with ZnO NPs for 30 minutes. Conclusion: The results showed that ZnO NPs can inhibit the establishment of P. aeruginosa biofilms and have less effective in removing pre-formed biofilm. However the tested nanoparticles exhibited anti-biofilm effect, but mRNA of pslA gene could be still detected in the medium by RT-PCR technique after 30 minutes treatment with ZnO.

  10. Drosophila melanogaster as an animal model for the study of Pseudomonas aeruginosa biofilm infections in vivo.

    Directory of Open Access Journals (Sweden)

    Heidi Mulcahy

    2011-10-01

    Full Text Available Pseudomonas aeruginosa is an opportunistic pathogen capable of causing both acute and chronic infections in susceptible hosts. Chronic P. aeruginosa infections are thought to be caused by bacterial biofilms. Biofilms are highly structured, multicellular, microbial communities encased in an extracellular matrix that enable long-term survival in the host. The aim of this research was to develop an animal model that would allow an in vivo study of P. aeruginosa biofilm infections in a Drosophila melanogaster host. At 24 h post oral infection of Drosophila, P. aeruginosa biofilms localized to and were visualized in dissected Drosophila crops. These biofilms had a characteristic aggregate structure and an extracellular matrix composed of DNA and exopolysaccharide. P. aeruginosa cells recovered from in vivo grown biofilms had increased antibiotic resistance relative to planktonically grown cells. In vivo, biofilm formation was dependent on expression of the pel exopolysaccharide genes, as a pelB::lux mutant failed to form biofilms. The pelB::lux mutant was significantly more virulent than PAO1, while a hyperbiofilm strain (PAZHI3 demonstrated significantly less virulence than PAO1, as indicated by survival of infected flies at day 14 postinfection. Biofilm formation, by strains PAO1 and PAZHI3, in the crop was associated with induction of diptericin, cecropin A1 and drosomycin antimicrobial peptide gene expression 24 h postinfection. In contrast, infection with the non-biofilm forming strain pelB::lux resulted in decreased AMP gene expression in the fly. In summary, these results provide novel insights into host-pathogen interactions during P. aeruginosa oral infection of Drosophila and highlight the use of Drosophila as an infection model that permits the study of P. aeruginosa biofilms in vivo.

  11. The implication of Pseudomonas aeruginosa biofilms in infections

    DEFF Research Database (Denmark)

    Rybtke, Morten T; Jensen, Peter Østrup; Høiby, Niels

    2011-01-01

    Biofilm formation by bacteria is recognized as a major problem in chronic infections due to their recalcitrance against the immune defense and available antibiotic treatment schemes. The opportunistic pathogen Pseudomonas aeruginosa has drawn special attention in this regard due to its severity o...... treatment strategies where the underlying targets are less prone for resistance development as bacteria, in retrospect, have a unique ability to evade the actions of classic antibiotics.......Biofilm formation by bacteria is recognized as a major problem in chronic infections due to their recalcitrance against the immune defense and available antibiotic treatment schemes. The opportunistic pathogen Pseudomonas aeruginosa has drawn special attention in this regard due to its severity......-up of the extracellular matrix encasing the biofilm-associated bacteria as well as the elaborate signaling mechanisms employed by the bacterium enables it to withstand the continuous stresses imposed by the immune defense and administered antibiotics resulting in a state of chronic inflammation that damages the host...

  12. Spatiotemporal pharmacodynamics of meropenem- and tobramycin-treated Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Haagensen, Janus Anders Juul; Verotta, Davide; Huang, Liusheng

    2017-01-01

    The selection and dose of antibiotic therapy for biofilm-related infections are based on traditional pharmacokinetic studies using planktonic bacteria. The objective of this study was to characterize the time course and spatial activity of human exposure levels of meropenem and tobramycin against...... Pseudomonas aeruginosa biofilms grown in an in vitro flow-chamber model. Pharmacokinetic profiles of meropenem and tobramycin used in human therapy were administered to GFP-labelled P. aeruginosa PAO1 grown in flow chambers for 24 or 72 h. Images were acquired using confocal laser scanning microscopy...... throughout antibiotic treatment. Bacterial biomass was measured using COMSTAT and pharmacokinetic/pharmacodynamic models were fitted using NONMEM7. Meropenem treatment resulted in more rapid and sustained killing of both the 24 and 72 h PAO1 biofilm compared with tobramycin. Biofilm regrowth after antibiotic...

  13. Development of Topical Treatment for Pseudomonas aeruginosa Wound Infections by Quorum-Sensing Inhibitors Mediated by Poly(amidoamine) (PAMAM) Dendrimers

    Science.gov (United States)

    2013-01-01

    dendrimers would provide added benefits as a delivery vehicle of QSI compounds to inhibit PA biofilms, by both increasing the transport of QSI as drug...Pseudomonas aeruginosa Wound Infections by Quorum-Sensing Inhibitors Mediated by Poly(amidoamine) (PAMAM) Dendrimers PRINCIPAL INVESTIGATOR...Development of Topical Treatment for Pseudomonas aeruginosa Wound Infections by Quorum-Sensing Inhibitors Mediated by Poly(amidoamine) (PAMAM) Dendrimers

  14. High β-Lactamase Levels Change the Pharmacodynamics of β-Lactam Antibiotics in Pseudomonas aeruginosa Biofilms

    Science.gov (United States)

    Ciofu, Oana; Yang, Liang; Wu, Hong; Song, Zhijun; Oliver, Antonio; Høiby, Niels

    2013-01-01

    Resistance to β-lactam antibiotics is a frequent problem in Pseudomonas aeruginosa lung infection of cystic fibrosis (CF) patients. This resistance is mainly due to the hyperproduction of chromosomally encoded β-lactamase and biofilm formation. The purpose of this study was to investigate the role of β-lactamase in the pharmacokinetics (PK) and pharmacodynamics (PD) of ceftazidime and imipenem on P. aeruginosa biofilms. P. aeruginosa PAO1 and its corresponding β-lactamase-overproducing mutant, PAΔDDh2Dh3, were used in this study. Biofilms of these two strains in flow chambers, microtiter plates, and on alginate beads were treated with different concentrations of ceftazidime and imipenem. The kinetics of antibiotics on the biofilms was investigated in vitro by time-kill methods. Time-dependent killing of ceftazidime was observed in PAO1 biofilms, but concentration-dependent killing activity of ceftazidime was observed for β-lactamase-overproducing biofilms of P. aeruginosa in all three models. Ceftazidime showed time-dependent killing on planktonic PAO1 and PAΔDDh2Dh3. This difference is probably due to the special distribution and accumulation in the biofilm matrix of β-lactamase, which can hydrolyze the β-lactam antibiotics. The PK/PD indices of the AUC/MBIC and Cmax/MBIC (AUC is the area under concentration-time curve, MBIC is the minimal biofilm-inhibitory concentration, and Cmax is the maximum concentration of drug in serum) are probably the best parameters to describe the effect of ceftazidime in β-lactamase-overproducing P. aeruginosa biofilms. Meanwhile, imipenem showed time-dependent killing on both PAO1 and PAΔDDh2Dh3 biofilms. An inoculum effect of β-lactams was found for both planktonic and biofilm P. aeruginosa cells. The inoculum effect of ceftazidime for the β-lactamase-overproducing mutant PAΔDDh2Dh3 biofilms was more obvious than for PAO1 biofilms, with a requirement of higher antibiotic concentration and a longer period of treatment

  15. The Pseudomonas aeruginosa Type III Translocon Is Required for Biofilm Formation at the Epithelial Barrier

    DEFF Research Database (Denmark)

    Tran, Cindy S; Rangel, Stephanie M; Almblad, Henrik

    2014-01-01

    Clinical infections by Pseudomonas aeruginosa, a deadly Gram-negative, opportunistic pathogen of immunocompromised hosts, often involve the formation of antibiotic-resistant biofilms. Although biofilm formation has been extensively studied in vitro on glass or plastic surfaces, much less is known...... about biofilm formation at the epithelial barrier. We have previously shown that when added to the apical surface of polarized epithelial cells, P. aeruginosa rapidly forms cell-associated aggregates within 60 minutes of infection. By confocal microscopy we now show that cell-associated aggregates...... a previously unappreciated function for the type III translocon in the formation of P. aeruginosa biofilms at the epithelial barrier and demonstrate that biofilms may form at early time points of infection....

  16. Cross-regulation by CrcZ RNA controls anoxic biofilm formation in Pseudomonas aeruginosa

    Science.gov (United States)

    Pusic, Petra; Tata, Muralidhar; Wolfinger, Michael T.; Sonnleitner, Elisabeth; Häussler, Susanne; Bläsi, Udo

    2016-12-01

    Pseudomonas aeruginosa (PA) can thrive in anaerobic biofilms in the lungs of cystic fibrosis (CF) patients. Here, we show that CrcZ is the most abundant PA14 RNA bound to the global regulator Hfq in anoxic biofilms grown in cystic fibrosis sputum medium. Hfq was crucial for anoxic biofilm formation. This observation complied with an RNAseq based transcriptome analysis and follow up studies that implicated Hfq in regulation of a central step preceding denitrification. CrcZ is known to act as a decoy that sequesters Hfq during relief of carbon catabolite repression, which in turn alleviates Hfq-mediated translational repression of catabolic genes. We therefore inferred that CrcZ indirectly impacts on biofilm formation by competing for Hfq. This hypothesis was supported by the findings that over-production of CrcZ mirrored the biofilm phenotype of the hfq deletion mutant, and that deletion of the crcZ gene augmented biofilm formation. To our knowledge, this is the first example where competition for Hfq by CrcZ cross-regulates an Hfq-dependent physiological process unrelated to carbon metabolism.

  17. Mannitol enhances antibiotic sensitivity of persister bacteria in Pseudomonas aeruginosa biofilms.

    Directory of Open Access Journals (Sweden)

    Nicolas Barraud

    Full Text Available The failure of antibiotic therapies to clear Pseudomonas aeruginosa lung infection, the key mortality factor for cystic fibrosis (CF patients, is partly attributed to the high tolerance of P. aeruginosa biofilms. Mannitol has previously been found to restore aminoglycoside sensitivity in Escherichia coli by generating a proton-motive force (PMF, suggesting a potential new strategy to improve antibiotic therapy and reduce disease progression in CF. Here, we used the commonly prescribed aminoglycoside tobramycin to select for P. aeruginosa persister cells during biofilm growth. Incubation with mannitol (10-40 mM increased tobramycin sensitivity of persister cells up to 1,000-fold. Addition of mannitol to pre-grown biofilms was able to revert the persister phenotype and improve the efficacy of tobramycin. This effect was blocked by the addition of a PMF inhibitor or in a P. aeruginosa mutant strain unable to metabolise mannitol. Addition of glucose and NaCl at high osmolarity also improved the efficacy of tobramycin although to a lesser extent compared to mannitol. Therefore, the primary effect of mannitol in reverting biofilm associated persister cells appears to be an active, physiological response, associated with a minor contribution of osmotic stress. Mannitol was tested against clinically relevant strains, showing that biofilms containing a subpopulation of persister cells are better killed in the presence of mannitol, but a clinical strain with a high resistance to tobramycin was not affected by mannitol. Overall, these results suggest that in addition to improvements in lung function by facilitating mucus clearance in CF, mannitol also affects antibiotic sensitivity in biofilms and does so through an active, physiological response.

  18. Biofilm formation and disinfectant resistance of Salmonella sp. in mono- and dual-species with Pseudomonas aeruginosa.

    Science.gov (United States)

    Pang, X Y; Yang, Y S; Yuk, H G

    2017-09-01

    This study aimed to evaluate the biofilm formation and disinfectant resistance of Salmonella cells in mono- and dual-species biofilms with Pseudomonas aeruginosa, and to investigate the role of extracellular polymeric substances (EPS) in the protection of biofilms against disinfection treatment. The populations of Salmonella in mono- or dual-species biofilms with P. aeruginosa on stainless steel (SS) coupons were determined before and after exposure to commercial disinfectant, 50 μg ml -1 chlorine or 200 μg ml -1 Ecolab ® Whisper™ V (a blend of four effective quaternary ammonium compounds (QAC)). In addition, EPS amount from biofilms was quantified and biofilm structures were observed using scanning electron microscopy (SEM). Antagonistic interactions between Salmonella and P. aeruginosa resulted in lower planktonic population level of Salmonella, and lower density in dual-species biofilms compared to mono-species biofilms. The presence of P. aeruginosa significantly enhanced disinfectant resistance of S. Typhimurium and S. Enteritidis biofilm cells for 2 days, and led to an average of 50% increase in polysaccharides amount in dual-species biofilms than mono-species biofilms of Salmonella. Microscopy observation showed the presence of large microcolonies covered by EPS in dual-species biofilms but not in mono-species ones. The presence of P. aeruginosa in dual-species culture inhibited the growth of Salmonella cells in planktonic phase and in biofilms, but protected Salmonella cells in biofilms from disinfection treatment, by providing more production of EPS in dual-species biofilms than mono-species ones. This study provides insights into inter-species interaction, with regard to biofilm population dynamics and disinfectant resistance. Thus, a sanitation protocol should be designed considering the protective role of secondary species to pathogens in biofilms on SS surface which has been widely used at food surfaces and manufacturers. © 2017 The Society

  19. Anti-Biofilm and Antivirulence Activities of Metabolites from Plectosphaerella cucumerina against Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Jinwei Zhou

    2017-05-01

    Full Text Available This study reported the efficacy of the metabolites of Plectosphaerella cucumerina, one phyllosphere fungus from Orychophragmus violaceus, against Pseudomonas aeruginosa quorum sensing (QS and QS-regulated biofilms. The minimum inhibitory concentration (MIC of the ethyl acetate (EtOAc extract from P. cucumerina against P. aeruginosa PAO1 was 1.25 mg mL−1. At sub-MIC concentrations, P. cucumerina extract (0.25–1 mg mL−1 not only inhibited biofilm formation but also disrupted preformed biofilms of P. aeruginosa PAO1 without affecting its growth. Fluorescence and scanning electron microscope (SEM showed architectural disruption of the biofilms when treated with P. cucumerina metabolites. Further investigation demonstrated that metabolites in P. cucumerina attenuated the QS-dependent virulence factors. LC-MS/MS spectra coupled with experimentally standard samples suggested that patulin and emodin might act as the principal components possessing anti-biofilm and antivirulence activities. This is the first report of (1 the isolation of P. cucumerina from the phyllosphere of O. violaceus and (2 anti-biofilm, antivirulence, and biofilm disruption activities of this fungus. Thus, this study provides fascinating new pathways for screening antipathogenic agents.

  20. Anaerobic Corrosion of 304 Stainless Steel Caused by the Pseudomonas aeruginosa Biofilm

    Directory of Open Access Journals (Sweden)

    Ru Jia

    2017-11-01

    Full Text Available Pseudomonas aeruginosa is a ubiquitous bacterium capable of forming problematic biofilms in many environments. They cause biocorrosion of medical implants and industrial equipment and infrastructure. Aerobic corrosion of P. aeruginosa against stainless steels has been reported by some researchers while there is a lack of reports on anaerobic P. aeruginosa corrosion in the literature. In this work, the corrosion by a wild-type P. aeruginosa (strain PAO1 biofilm against 304 stainless steel (304 SS was investigated under strictly anaerobic condition for up to 14 days. The anaerobic corrosion of 304 SS by P. aeruginosa was reported for the first time. Results showed that the average sessile cell counts on 304 SS coupons after 7- and 14-day incubations were 4.8 × 107 and 6.2 × 107 cells/cm2, respectively. Scanning electron microscopy and confocal laser scanning microscopy corroborated the sessile cell counts. The X-ray diffraction analysis identified the corrosion product as iron nitride, confirming that the corrosion was caused by the nitrate reducing biofilm. The largest pit depths on 304 SS surfaces after the 7- and 14-day incubations with P. aeruginosa were 3.9 and 7.4 μm, respectively. Electrochemical tests corroborated the pitting data.

  1. Identification of biofilm-associated cluster (bac in Pseudomonas aeruginosa involved in biofilm formation and virulence.

    Directory of Open Access Journals (Sweden)

    Camille Macé

    Full Text Available Biofilms are prevalent in diseases caused by Pseudomonas aeruginosa, an opportunistic and nosocomial pathogen. By a proteomic approach, we previously identified a hypothetical protein of P. aeruginosa (coded by the gene pA3731 that was accumulated by biofilm cells. We report here that a Delta pA3731 mutant is highly biofilm-defective as compared with the wild-type strain. Using a mouse model of lung infection, we show that the mutation also induces a defect in bacterial growth during the acute phase of infection and an attenuation of the virulence. The pA3731 gene is found to control positively the ability to swarm and to produce extracellular rhamnolipids, and belongs to a cluster of 4 genes (pA3729-pA3732 not previously described in P. aeruginosa. Though the protein PA3731 has a predicted secondary structure similar to that of the Phage Shock Protein, some obvious differences are observed compared to already described psp systems, e.g., this unknown cluster is monocistronic and no homology is found between the other proteins constituting this locus and psp proteins. As E. coli PspA, the amount of the protein PA3731 is enlarged by an osmotic shock, however, not affected by a heat shock. We consequently named this locus bac for biofilm-associated cluster.

  2. HD-GYP domain proteins regulate biofilm formation and virulence in Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Ryan, Robert P.; Lucey, Jean; O'Donovan, Karen

    2009-01-01

    residues (YN-GYP). Here we have investigated the role of these proteins in biofilm formation, virulence factor synthesis and virulence of P. aeruginosa. Mutation of PA4108 and PA4781 led to an increase in the level of cyclic-di-GMP in P. aeruginosa, consistent with the predicted activity of the encoded......2572 had a negative influence on swarming that was cryptic and was revealed only after removal of an uncharacterized C-terminal domain. Mutation of PA4108, PA4781 and PA2572 had distinct effects on biofilm formation and architecture of P. aeruginosa. All three proteins contributed to virulence of P...

  3. Two genetic loci produce distinct carbohydrate-rich structural components of the Pseudomonas aeruginosa biofilm matrix.

    Science.gov (United States)

    Friedman, Lisa; Kolter, Roberto

    2004-07-01

    Pseudomonas aeruginosa forms biofilms, which are cellular aggregates encased in an extracellular matrix. Molecular genetics studies of three common autoaggregative phenotypes, namely wrinkled colonies, pellicles, and solid-surface-associated biofilms, led to the identification of two loci, pel and psl, that are involved in the production of carbohydrate-rich components of the biofilm matrix. The pel gene cluster is involved in the production of a glucose-rich matrix material in P. aeruginosa strain PA14 (L. Friedman and R. Kolter, Mol. Microbiol. 51:675-690, 2004). Here we investigate the role of the pel gene cluster in P. aeruginosa strain ZK2870 and identify a second genetic locus, termed psl, involved in the production of a mannose-rich matrix material. The 11 predicted protein products of the psl genes are homologous to proteins involved in carbohydrate processing. P. aeruginosa is thus able to produce two distinct carbohydrate-rich matrix materials. Either carbohydrate-rich matrix component appears to be sufficient for mature biofilm formation, and at least one of them is required for mature biofilm formation in P. aeruginosa strains PA14 and ZK2870. Copyright 2004 American Society for Microbiology

  4. Influence of glyphosate in planktonic and biofilm growth of Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Ilana Schneider Lima

    2014-09-01

    Full Text Available This study evaluated the impact of different concentrations of glyphosate (Rondup® on planktonic and biofilm growth of P. aeruginosa. Aerobic and anaerobic cultures of P. aeruginosa ATCC®15442 inoculated in MHB + glyphosate (0.845 ppm, 1.690 ppm, 8.45 ppm, 16.90 ppm, 84.50 ppm, 169 ppm, 845 ppm, and 1690 ppm and cultured in normoxia and anoxia, following their OD560nm every hour for 24 h. Biofilms of adapted cells were formed in the presence of glyphosate (0.845 to 1690 ppm in normoxia and anoxia for 36 h. Glyphosate at concentrations higher than 84.5 ppm reduces the cell density of planktonic aerobic cultures (p 0.05, and more pronounced over 169 ppm. Anaerobic biofilms have their growth more readily favored (p < 0.05, regardless of concentration. In a concentration-dependent manner, glyphosate interferes with the growth ability of P. aeruginosa ATCC®15442.

  5. Effect of biosurfactants on Pseudomonas aeruginosa and Staphylococcus aureus biofilms in a BioFlux channel.

    Science.gov (United States)

    Diaz De Rienzo, M A; Stevenson, P S; Marchant, R; Banat, I M

    2016-07-01

    Recent studies have indicated that biosurfactants play a role both in maintaining channels between multicellular structures in biofilms and in dispersal of cells from biofilms. A combination of caprylic acid (0.01 % v/v) together with rhamnolipids (0.04 % v/v) was applied to biofilms of Pseudomonas aeruginosa ATCC 15442, Staphylococcus aureus ATCC 9144 and a mixed culture under BioFlux flowthrough conditions and caused disruption of the biofilms. The biofilms were also treated with a combination of rhamnolipids (0.04 % v/v) and sophorolipids (0.01 %). Control treatments with PBS 1× had no apparent effect on biofilm disruption. The Gram-positive bacterium (S. aureus ATCC 9144) was more sensitive than P. aeruginosa ATCC 15442 in terms of disruption and viability as shown by Live/Dead staining. Disruption of biofilms of P. aeruginosa ATCC 15442 was minimal. Oxygen consumption by biofilms, after different treatments with biosurfactants, confirms that sophorolipid on its own is unable to kill/inhibit cells of P. aeruginosa ATCC 15442, and even when used in combination with rhamnolipids, under static conditions, no decrease in the cell viability was observed. Cells in biofilms exposed to mono-rhamnolipids (0.04 % v/v) showed behaviour typical of exposure to bacteriostatic compounds, but when exposed to di-rhamnolipids (0.04 % v/v), they displayed a pattern characteristic of bactericidal compounds.

  6. Nanoindentation of Pseudomonas aeruginosa bacterial biofilm using atomic force microscopy

    International Nuclear Information System (INIS)

    Baniasadi, Mahmoud; Xu, Zhe; Du, Yingjie; Lu, Hongbing; Minary-Jolandan, Majid; Gandee, Leah; Zimmern, Philippe

    2014-01-01

    Bacterial biofilms are a source of many chronic infections. Biofilms and their inherent resistance to antibiotics are attributable to a range of health issues including affecting prosthetic implants, hospital-acquired infections, and wound infection. Mechanical properties of biofilm, in particular, at micro- and nano-scales, are governed by microstructures and porosity of the biofilm, which in turn may contribute to their inherent antibiotic resistance. We utilize atomic force microscopy (AFM)-based nanoindentation and finite element simulation to investigate the nanoscale mechanical properties of Pseudomonas aeruginosa bacterial biofilm. This biofilm was derived from human samples and represents a medically relevant model. (paper)

  7. Comparison of UVB and UVC irradiation disinfection efficacies on Pseudomonas Aeruginosa biofilm

    DEFF Research Database (Denmark)

    Argyraki, Aikaterini; Markvart, M.; Nielsen, Anne

    2016-01-01

    skin, soft tissue, lungs, kidney and urinary tract infections. Moreover, it can be found on and in medical equipment causing often cross infections in hospitals. The objective of this study was to test the efficiency, of two different light-based disinfection treatments, namely UVB and UVC irradiation......, on P. aeruginosa biofilms at different growth stages. In our experiments a new type of UV light emitting diodes (LEDs) were used to deliver UV irradiation on the biofilms, in the UVB (296nm) and UVC (266nm) region. The killing rate was studied as a function of dose for 24h grown biofilms. The dose......Disinfection routines are important in all clinical applications. The uprising problem of antibiotic resistance has driven major research efforts towards alternative disinfection approaches, involving light-based solutions. Pseudomonas aeruginosa (P. aeruginosa) is a common bacterium that can cause...

  8. Biofilm Formation Mechanisms of Pseudomonas aeruginosa Predicted via Genome-Scale Kinetic Models of Bacterial Metabolism

    Science.gov (United States)

    2016-03-15

    RESEARCH ARTICLE Biofilm Formation Mechanisms of Pseudomonas aeruginosa Predicted via Genome-Scale Kinetic Models of Bacterial Metabolism Francisco G...jaques.reifman.civ@mail.mil Abstract A hallmark of Pseudomonas aeruginosa is its ability to establish biofilm -based infections that are difficult to...eradicate. Biofilms are less susceptible to host inflammatory and immune responses and have higher antibiotic tolerance than free-living planktonic

  9. High beta-Lactamase Levels Change the Pharmacodynamics of beta-Lactam Antibiotics in Pseudomonas aeruginosa Biofilms

    DEFF Research Database (Denmark)

    Wang, Hengzhuang; Ciofu, Oana; Yang, Liang

    2013-01-01

    the role of beta-lactamase in the pharmacokinetics (PK) and pharmacodynamics (PD) of ceftazidime and imipenem on P. aeruginosa biofilms. P. aeruginosa PAO1 and its corresponding beta-lactamase-overproducing mutant, PA Delta DDh2Dh3, were used in this study. Biofilms of these two strains in flow chambers......, microtiter plates, and on alginate beads were treated with different concentrations of ceftazidime and imipenem. The kinetics of antibiotics on the biofilms was investigated in vitro by time-kill methods. Time-dependent killing of ceftazidime was observed in PAO1 biofilms, but concentration-dependent killing...... activity of ceftazidime was observed for beta-lactamase-overproducing biofilms of P. aeruginosa in all three models. Ceftazidime showed time-dependent killing on planktonic PAO1 and PA Delta DDh2Dh3. This difference is probably due to the special distribution and accumulation in the biofilm matrix of beta...

  10. Polysaccharides serve as scaffold of biofilms formed by mucoid Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Yang, Liang; Hengzhuang, Wang; Wu, Hong

    2012-01-01

    Chronic lung infection by mucoid Pseudomonas aeruginosa is one of the major pathologic features in patients with cystic fibrosis. Mucoid P. aeruginosa is notorious for its biofilm forming capability and resistance to immune attacks. In this study, the roles of extracellular polymeric substances f...

  11. Combined treatment of Pseudomonas aeruginosa biofilms with bacteriophages and chlorine.

    Science.gov (United States)

    Zhang, Yanyan; Hu, Zhiqiang

    2013-01-01

    Bacterial biofilms are a growing concern in a broad range of areas. In this study, a mixture of RNA bacteriophages isolated from municipal wastewater was used to control and remove biofilms. At the concentrations of 400 and 4 × 10(7) PFU/mL, the phages inhibited Pseudomonas aeruginosa biofilm formation by 45 ± 15% and 73 ± 8%, respectively. At the concentrations of 6,000 and 6 × 10(7) PFU/mL, the phages removed 45 ± 9% and 75 ± 5% of pre-existing P. aeruginosa biofilms, respectively. Chlorine reduced biofilm growth by 86 ± 3% at the concentration of 210 mg/L, but it did not remove pre-existing biofilms. However, a combination of phages (3 × 10(7) PFU/mL) and chlorine at this concentration reduced biofilm growth by 94 ± 2% and removed 88 ± 6% of existing biofilms. In a continuous flow system with continued biofilm growth, a combination of phages (a one-time treatment at the concentration of 1.9 × 10(8) PFU/mL for 1 h first) with chlorine removed 97 ± 1% of biofilms after Day 5 while phage and chlorine treatment alone removed 89 ± 1% and 40 ± 5%, respectively. For existing biofilms, a combined use of a lower phage concentration (3.8 × 10(5) PFU/mL) and chlorination with a shorter time duration (12 h) followed by continuous water flushing removed 96 ± 1% of biofilms in less than 2 days. Laser scanning confocal microscopy supplemented with electron microscopy indicated that the combination treatment resulted in biofilms with lowest cell density and viability. These results suggest that the combination treatment of phages and chlorine is a promising method to control and remove bacterial biofilms from various surfaces. Copyright © 2012 Wiley Periodicals, Inc.

  12. Hyperbaric oxygen sensitizes anoxic Pseudomonas aeruginosa biofilm to ciprofloxacin

    DEFF Research Database (Denmark)

    Kolpen, Mette; Lerche, Christian J; Kragh, Kasper Nørskov

    2017-01-01

    Chronic Pseudomonas aeruginosa lung infection is characterized by the presence of endobronchial antibiotic-tolerant biofilm subject to strong oxygen (O2) depletion due to the activity of surrounding polymorphonuclear leukocytes. The exact mechanisms affecting the antibiotic susceptibility...... metabolism activity and the endogenous formation of reactive O2 radicals (ROS). In this study we aimed to apply hyperbaric oxygen treatment (HBOT) in order to sensitize anoxic P. aeruginosa agarose-biofilms established to mimic situations with intense O2 consumption by the host response in the cystic...... fibrosis (CF) lung. Application of HBOT resulted in enhanced bactericidal activity of ciprofloxacin at clinically relevant durations and was accompanied by indications of restored aerobic respiration, involvement of endogenous lethal oxidative stress and increased bacterial growth. The findings highlight...

  13. Interactions between polymorphonuclear leukocytes and Pseudomonas aeruginosa biofilms on silicone implants in vivo

    DEFF Research Database (Denmark)

    van Gennip, Maria; Hultqvist, Louise Dahl; Alhede, Morten

    2012-01-01

    (PMNs). In contrast, the number of cells of a P. aeruginosa rhlA mutant that cannot produce rhamnolipids was significantly reduced on the implants by day 1, and the bacteria were actively phagocytosed by infiltrating PMNs. In addition, we identified extracellular wire-like structures around the bacteria......Chronic infections with Pseudomonas aeruginosa persist because the bacterium forms biofilms that are tolerant to antibiotic treatment and the host immune response. Scanning electron microscopy and confocal laser scanning microscopy were used to visualize biofilm development in vivo following...... intraperitoneal inoculation of mice with bacteria growing on hollow silicone tubes, as well as to examine the interaction between these bacteria and the host innate immune response. Wild-type P. aeruginosa developed biofilms within 1 day that trapped and caused visible cavities in polymorphonuclear leukocytes...

  14. Hyperbaric oxygen sensitizes anoxic Pseudomonas aeruginosa biofilm to ciprofloxacin

    DEFF Research Database (Denmark)

    Kolpen, Mette; Lerche, Christian J; Kragh, Kasper Nørskov

    2017-01-01

    fibrosis (CF) lung. Application of HBOT resulted in enhanced bactericidal activity of ciprofloxacin at clinically relevant durations and was accompanied by indications of restored aerobic respiration, involvement of endogenous lethal oxidative stress and increased bacterial growth. The findings highlight...... that oxygenation by HBOT improves the bactericidal activity of ciprofloxacin on P. aeruginosa biofilm and suggest that bacterial biofilms is sensitized to antibiotics by supplying hyperbaric O2....

  15. Non-Tuberculous Mycobacteria multispecies biofilms in cystic fibrosis: development of an in vitro Mycobacterium abscessus and Pseudomonas aeruginosa dual species biofilm model.

    Science.gov (United States)

    Rodríguez-Sevilla, Graciela; García-Coca, Marta; Romera-García, David; Aguilera-Correa, John Jairo; Mahíllo-Fernández, Ignacio; Esteban, Jaime; Pérez-Jorge, Concepción

    2018-04-01

    Lung disease in cystic fibrosis (CF) is characterized by the progressive colonization of the respiratory tract by different bacteria, which develop polymicrobial biofilms. In the past decades, there has been an increase in the number of CF patients infected with Non-Tuberculous Mycobacteria (NTM). Although Mycobacterium abscessus is the main NTM isolated globally, little is known about M. abscessus multispecies biofilm formation. In the present study we developed an in vitro model to study the phenotypic characteristics of biofilms formed by M. abscessus and Pseudomonas aeruginosa, a major pathogen in CF. For that purpose, dual species biofilms were grown on polycarbonate membranes with a fixed concentration of P. aeruginosa and different inoculums of M. abscessus. The biofilms were sampled at 24, 48, and 72 h and bacteria were quantified in specific media. The results revealed that the increasing initial concentration of M. abscessus in dual species biofilms had an effect on its population only at 24 and 48 h, whereas P. aeruginosa was not affected by the different concentrations used of M. abscessus. Time elapsed increased biofilm formation of both species, specially between 24 and 48 h. According to the results, the conditions to produce a mature dual species biofilm in which the relative species distribution remained stable were 72 h growth of the mixed microbial culture at a 1:1 ratio. A significant decrease in mycobacterial population in dual compared to single species biofilms was found, suggesting that P. aeruginosa has a negative influence on M. abscessus. Finally, in a proof of concept experiment, young and mature dual species biofilms were exposed to clarithromycin. Copyright © 2018 Elsevier GmbH. All rights reserved.

  16. Determination of the spatiotemporal dependence of Pseudomonas aeruginosa biofilm viability after treatment with NLC-colistin

    Directory of Open Access Journals (Sweden)

    Sans-Serramitjana E

    2017-06-01

    Full Text Available Eulalia Sans-Serramitjana,1 Marta Jorba,1 José Luis Pedraz,2 Teresa Vinuesa,1 Miguel Viñas1 1Laboratory of Molecular Microbiology and Antimicrobials, Department of Pathology and Experimental Therapeutics, University of Barcelona, Barcelona, 2Laboratory of Pharmaceutics, University of the Basque Country (UPV/EHU, Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina, Vitoria, Spain Abstract: The emergence of colistin-resistant Pseudomonas aeruginosa in cystic fibrosis (CF patients, particularly after long-term inhalation treatments, has been recently reported. Nanoencapsulation may enable preparations to overcome the limitations of conventional pharmaceutical forms. We have determined the time-dependent viability of P. aeruginosa biofilms treated with both free and nanoencapsulated colistin. We also examined the relationship between the optimal anti-biofilm activity of nanostructured lipid carrier (NLC-colistin and the structural organization of the biofilm itself. The results showed the more rapid killing of P. aeruginosa bacterial biofilms by NLC-colistin than by free colistin. However, the two formulations did not differ in terms of the final percentages of living and dead cells, which were higher in the inner than in the outer layers of the treated biofilms. The effective anti-biofilm activity of NLC-colistin and its faster killing effect recommend further studies of its use over free colistin in the treatment of P. aeruginosa infections in CF patients. Keywords: cystic fibrosis, colistin sulfate, lipid nanoparticles, P. aeruginosa, confocal laser scanning microscopy, anti-biofilm activity

  17. Evolution and Adaptation in Pseudomonas aeruginosa Biofilms Driven by Mismatch Repair System-Deficient Mutators

    DEFF Research Database (Denmark)

    Luján, Adela M.; Maciá, María D.; Yang, Liang

    2011-01-01

    , which are rarely eradicated despite intensive antibiotic therapy. Current knowledge indicates that three major adaptive strategies, biofilm development, phenotypic diversification, and mutator phenotypes [driven by a defective mismatch repair system (MRS)], play important roles in P. aeruginosa chronic...... infections, but the relationship between these strategies is still poorly understood. We have used the flow-cell biofilm model system to investigate the impact of the mutS associated mutator phenotype on development, dynamics, diversification and adaptation of P. aeruginosa biofilms. Through competition...... diversification, evidenced by biofilm architecture features and by a wider range and proportion of morphotypic colony variants, respectively. Additionally, morphotypic variants generated in mutator biofilms showed increased competitiveness, providing further evidence for mutator-driven adaptive evolution...

  18. Formation of hydroxyl radicals contributes to the bactericidal activity of ciprofloxacin against Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Jensen, Peter Ø; Briales, Alejandra; Brochmann, Rikke P; Wang, Hengzhuang; Kragh, Kasper N; Kolpen, Mette; Hempel, Casper; Bjarnsholt, Thomas; Høiby, Niels; Ciofu, Oana

    2014-04-01

    Antibiotic-tolerant, biofilm-forming Pseudomonas aeruginosa has long been recognized as a major cause of chronic lung infections of cystic fibrosis patients. The mechanisms involved in the activity of antibiotics on biofilm are not completely clear. We have investigated whether the proposed induction of cytotoxic hydroxyl radicals (OH˙) during antibiotic treatment of planktonically grown cells may contribute to action of the commonly used antibiotic ciprofloxacin on P. aeruginosa biofilms. For this purpose, WT PAO1, a catalase deficient ΔkatA and a ciprofloxacin resistant mutant of PAO1 (gyrA), were grown as biofilms in microtiter plates and treated with ciprofloxacin. Formation of OH˙ and total amount of reactive oxygen species (ROS) was measured and viability was estimated. Formation of OH˙ and total ROS in PAO1 biofilms treated with ciprofloxacin was shown but higher levels were measured in ΔkatA biofilms, and no ROS production was seen in the gyrA biofilms. Treatment with ciprofloxacin decreased the viability of PAO1 and ΔkatA biofilms but not of gyrA biofilms. Addition of thiourea, a OH˙ scavenger, decreased the OH˙ levels and killing of PAO1 biofilm. Our study shows that OH˙ is produced by P. aeruginosa biofilms treated with ciprofloxacin, which may contribute to the killing of biofilm subpopulations. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  19. Different Dose-Dependent Modes of Action of C-Type Natriuretic Peptide on Pseudomonas aeruginosa Biofilm Formation

    Directory of Open Access Journals (Sweden)

    Florie Desriac

    2018-04-01

    Full Text Available We have previously shown that the C-type Natriuretic Peptide (CNP, a peptide produced by lungs, is able to impact Pseudomonas aeruginosa physiology. In the present work, the effect of CNP at different concentrations on P. aeruginosa biofilm formation was studied and the mechanisms of action of this human hormone on P. aeruginosa were deciphered. CNP was shown to inhibit dynamic biofilm formation in a dose-dependent manner without affecting the bacterial growth at any tested concentrations. The most effective concentrations were 1 and 0.1 µM. At 0.1 µM, the biofilm formation inhibition was fully dependent on the CNP sensor protein AmiC, whereas it was only partially AmiC-dependent at 1 µM, revealing the existence of a second AmiC-independent mode of action of CNP on P. aeruginosa. At 1 µM, CNP reduced both P. aeruginosa adhesion on glass and di-rhamnolipid production and also increased the bacterial membrane fluidity. The various effects of CNP at 1 µM and 0.1 µM on P. aeruginosa shown here should have major consequences to design drugs for biofilm treatment or prevention.

  20. Improved Biofilm Antimicrobial Activity of Polyethylene Glycol Conjugated Tobramycin Compared to Tobramycin in Pseudomonas aeruginosa Biofilms.

    Science.gov (United States)

    Du, Ju; Bandara, H M H N; Du, Ping; Huang, Hui; Hoang, Khang; Nguyen, Dang; Mogarala, Sri Vasudha; Smyth, Hugh D C

    2015-05-04

    The objective of this study was to develop a functionally enhanced antibiotic that would improve the therapeutic activity against bacterial biofilms. Tobramycin was chemically conjugated with polyethylene glycol (PEG) via site-specific conjugation to form PEGylated-tobramycin (Tob-PEG). The antibacterial efficacy of Tob-PEG, as compared to tobramycin, was assessed on the planktonic phase and biofilms phase of Pseudomonas aeruginosa. The minimum inhibitory concentration (MIC80) of Tob-PEG was higher (13.9 μmol/L) than that of tobramycin (1.4 μmol/L) in the planktonic phases. In contrast, the Tob-PEG was approximately 3.2-fold more effective in eliminating bacterial biofilms than tobramycin. Specifically, Tob-PEG had a MIC80 lower than those exhibited by tobramycin (27.8 μmol/L vs 89.8 μmol/L). Both confocal laser scanning microscopy and scanning electron microscopy further confirmed these data. Thus, modification of antimicrobials by PEGylation appears to be a promising approach for overcoming the bacterial resistance in the established biofilms of Pseudomonas aeruginosa.

  1. Antibiotic resistance in Pseudomonas aeruginosa biofilms: towards the development of novel anti-biofilm therapies.

    Science.gov (United States)

    Taylor, Patrick K; Yeung, Amy T Y; Hancock, Robert E W

    2014-12-10

    The growth of bacteria as structured aggregates termed biofilms leads to their protection from harsh environmental conditions such as physical and chemical stresses, shearing forces, and limited nutrient availability. Because of this highly adapted ability to survive adverse environmental conditions, bacterial biofilms are recalcitrant to antibiotic therapies and immune clearance. This is particularly problematic in hospital settings where biofilms are a frequent cause of chronic and device-related infections and constitute a significant burden on the health-care system. The major therapeutic strategy against infections is the use of antibiotics, which, due to adaptive resistance, are often insufficient to clear biofilm infections. Thus, novel biofilm-specific therapies are required. Specific features of biofilm development, such as surface adherence, extracellular matrix formation, quorum sensing, and highly regulated biofilm maturation and dispersal are currently being studied as targets to be exploited in the development of novel biofilm-specific treatments. Using Pseudomonas aeruginosa for illustrative purposes, this review highlights the antibiotic resistance mechanisms of biofilms, and discusses current research into novel biofilm-specific therapies. Copyright © 2014 Elsevier B.V. All rights reserved.

  2. The efficacy of sewage influent-isolated bacteriophages on Pseudomonas aeruginosa in a mixed-species biofilm

    KAUST Repository

    Yap, Scott

    2016-12-01

    The growth of environmentally persistent biofilms in cooling towers causes several associated problems, including microbiologically-induced corrosion (MIC) and biofouling. Current chemical control methods are not only ineffective against biofilms and costly to procure, they also have downstream environmental impacts when released untreated, or incur additional treatment costs. Bacteriophages are alternative biofilm control agents that have the potential to be more effective, cheaper to produce and yet have a more benign effect on the environment. In this study, biofilms grown under conditions simulating seawater fed cooling towers were characterized and the differences in growth and community make-up across time and different substrates were assessed. An MIC associated bacterium common in cooling tower water, P. aeruginosa, was chosen. Seven bacteriophage strains found to be effective against the chosen bacterium were isolated from wastewater influent. The relative effectiveness of these strains was measured against P. aeruginosa across different salinities. Separate biofilms fed with P. aeruginosa enriched seawater were characterized and the effectiveness of the isolated strains, singly and in cocktails, against the enriched biofilms was measured.

  3. The Pseudomonas aeruginosa transcriptome in planktonic cultures and static biofilms using RNA sequencing.

    Directory of Open Access Journals (Sweden)

    Andreas Dötsch

    Full Text Available In this study, we evaluated how gene expression differs in mature Pseudomonas aeruginosa biofilms as opposed to planktonic cells by the use of RNA sequencing technology that gives rise to both quantitative and qualitative information on the transcriptome. Although a large proportion of genes were consistently regulated in both the stationary phase and biofilm cultures as opposed to the late exponential growth phase cultures, the global biofilm gene expression pattern was clearly distinct indicating that biofilms are not just surface attached cells in stationary phase. A large amount of the genes found to be biofilm specific were involved in adaptation to microaerophilic growth conditions, repression of type three secretion and production of extracellular matrix components. Additionally, we found many small RNAs to be differentially regulated most of them similarly in stationary phase cultures and biofilms. A qualitative analysis of the RNA-seq data revealed more than 3000 putative transcriptional start sites (TSS. By the use of rapid amplification of cDNA ends (5'-RACE we confirmed the presence of three different TSS associated with the pqsABCDE operon, two in the promoter of pqsA and one upstream of the second gene, pqsB. Taken together, this study reports the first transcriptome study on P. aeruginosa that employs RNA sequencing technology and provides insights into the quantitative and qualitative transcriptome including the expression of small RNAs in P. aeruginosa biofilms.

  4. Selective labelling and eradication of antibiotic-tolerant bacterial populations in Pseudomonas aeruginosa biofilms

    Science.gov (United States)

    Chua, Song Lin; Yam, Joey Kuok Hoong; Hao, Piliang; Adav, Sunil S.; Salido, May Margarette; Liu, Yang; Givskov, Michael; Sze, Siu Kwan; Tolker-Nielsen, Tim; Yang, Liang

    2016-01-01

    Drug resistance and tolerance greatly diminish the therapeutic potential of antibiotics against pathogens. Antibiotic tolerance by bacterial biofilms often leads to persistent infections, but its mechanisms are unclear. Here we use a proteomics approach, pulsed stable isotope labelling with amino acids (pulsed-SILAC), to quantify newly expressed proteins in colistin-tolerant subpopulations of Pseudomonas aeruginosa biofilms (colistin is a ‘last-resort' antibiotic against multidrug-resistant Gram-negative pathogens). Migration is essential for the formation of colistin-tolerant biofilm subpopulations, with colistin-tolerant cells using type IV pili to migrate onto the top of the colistin-killed biofilm. The colistin-tolerant cells employ quorum sensing (QS) to initiate the formation of new colistin-tolerant subpopulations, highlighting multicellular behaviour in antibiotic tolerance development. The macrolide erythromycin, which has been previously shown to inhibit the motility and QS of P. aeruginosa, boosts biofilm eradication by colistin. Our work provides insights on the mechanisms underlying the formation of antibiotic-tolerant populations in bacterial biofilms and indicates research avenues for designing more efficient treatments against biofilm-associated infections. PMID:26892159

  5. Selective labelling and eradication of antibiotic-tolerant bacterial populations in Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Chua, Song Lin; Yam, Joey Kuok Hoong; Hao, Piliang; Adav, Sunil S; Salido, May Margarette; Liu, Yang; Givskov, Michael; Sze, Siu Kwan; Tolker-Nielsen, Tim; Yang, Liang

    2016-02-19

    Drug resistance and tolerance greatly diminish the therapeutic potential of antibiotics against pathogens. Antibiotic tolerance by bacterial biofilms often leads to persistent infections, but its mechanisms are unclear. Here we use a proteomics approach, pulsed stable isotope labelling with amino acids (pulsed-SILAC), to quantify newly expressed proteins in colistin-tolerant subpopulations of Pseudomonas aeruginosa biofilms (colistin is a 'last-resort' antibiotic against multidrug-resistant Gram-negative pathogens). Migration is essential for the formation of colistin-tolerant biofilm subpopulations, with colistin-tolerant cells using type IV pili to migrate onto the top of the colistin-killed biofilm. The colistin-tolerant cells employ quorum sensing (QS) to initiate the formation of new colistin-tolerant subpopulations, highlighting multicellular behaviour in antibiotic tolerance development. The macrolide erythromycin, which has been previously shown to inhibit the motility and QS of P. aeruginosa, boosts biofilm eradication by colistin. Our work provides insights on the mechanisms underlying the formation of antibiotic-tolerant populations in bacterial biofilms and indicates research avenues for designing more efficient treatments against biofilm-associated infections.

  6. Development and (evidence for) destruction of biofilm with Pseudomonas aeruginosa as architect

    Science.gov (United States)

    Uzcategui, Valerie N.; Donadeo, John J.; Lombardi, Daniel R.; Costello, Michael J.; Sauer, Richard L.

    1991-01-01

    Disinfection and maintenance of an acceptable level of asepsis in spacecraft potable water delivery systems is a formidable task. The major area of research for this project has been to monitor the formation and growth of biofilm, and biofilm attached microorganisms, on stainless steel surfaces (specifically coupons), and the use of ozone for the elimination of these species in a closed loop system. A number of different techniques have been utilized during the course of a typical run. Scraping and sonication of coupon surfaces with subsequent plating as well as epifluorescence microscopy have been utilized to enumerate biofilm protected Pseudomonas aeruginosa. In addition, scanning electron microscopy is the method of choice to examine the integrity of the biofilm. For ozone determinations, the indigo decolorization spectrophotometric method seems most reliable. Both high- and low-nutrient cultured P. aeruginosa organisms were the target species for the ozone disinfection experiments.

  7. In Vivo Pharmacokinetics/Pharmacodynamics of Colistin and Imipenem in Pseudomonas aeruginosa Biofilm Infection

    Science.gov (United States)

    Wu, Hong; Ciofu, Oana; Song, Zhijun; Høiby, Niels

    2012-01-01

    Many Pseudomonas aeruginosa isolates from the airways of patients with cystic fibrosis (CF) are sensitive to antibiotics in susceptibility testing, but eradication of the infection is difficult. The main reason is the biofilm formation in the airways of patients with CF. The pharmacokinetics (PKs) and pharmacodynamics (PDs) of antimicrobials can reliably be used to predict whether antimicrobial regimens will achieve the maximum bactericidal effect against infections. Unfortunately, however, most PK/PD studies of antimicrobials have been done on planktonic cells and very few PK/PD studies have been done on biofilms, partly due to the lack of suitable models in vivo. In the present study, a biofilm lung infection model was developed to provide an objective and quantitative evaluation of the PK/PD profile of antimicrobials. Killing curves were set up to detect the antimicrobial kinetics on planktonic and biofilm P. aeruginosa cells in vivo. Colistin showed concentration-dependent killing, while imipenem showed time-dependent killing on both planktonic and biofilm P. aeruginosa cells in vivo. The parameter best correlated to the elimination of bacteria in lung by colistin was the area under the curve (AUC) versus MIC (AUC/MIC) for planktonic cells or the AUC versus minimal biofilm inhibitory concentration (MBIC; AUC/MBIC) for biofilm cells. The best-correlated parameter for imipenem was the time that the drug concentration was above the MIC for planktonic cells (TMIC) or time that the drug concentration was above the MBIC (TMBIC) for biofilm cells. However, the AUC/MIC of imipenem showed a better correlation with the efficacy of imipenem for biofilm infections (R2 = 0.89) than planktonic cell infections (R2 = 0.38). The postantibiotic effect (PAE) of colistin and imipenem was shorter in biofilm infections than planktonic cell infections in this model. PMID:22354300

  8. Factors Affecting Catalase Expression in Pseudomonas aeruginosa Biofilms and Planktonic Cells

    OpenAIRE

    Frederick, Jesse R.; Elkins, James G.; Bollinger, Nikki; Hassett, Daniel J.; McDermott, Timothy R.

    2001-01-01

    Previous work with Pseudomonas aeruginosa showed that catalase activity in biofilms was significantly reduced relative to that in planktonic cells. To better understand biofilm physiology, we examined possible explanations for the differential expression of catalase in cells cultured in these two different conditions. For maximal catalase activity, biofilm cells required significantly more iron (25 μM as FeCl3) in the medium, whereas planktonic cultures required no addition of iron. However, ...

  9. Inhibition of biofilm formation, quorum sensing and infection in Pseudomonas aeruginosa by natural products-inspired organosulfur compounds.

    Directory of Open Access Journals (Sweden)

    Nathaniel C Cady

    Full Text Available Using a microplate-based screening assay, the effects on Pseudomonas aeruginosa PAO1 biofilm formation of several S-substituted cysteine sulfoxides and their corresponding disulfide derivatives were evaluated. From our library of compounds, S-phenyl-L-cysteine sulfoxide and its breakdown product, diphenyl disulfide, significantly reduced the amount of biofilm formation by P. aeruginosa at levels equivalent to the active concentration of 4-nitropyridine-N-oxide (NPO (1 mM. Unlike NPO, which is an established inhibitor of bacterial biofilms, our active compounds did not reduce planktonic cell growth and only affected biofilm formation. When used in a Drosophila-based infection model, both S-phenyl-L-cysteine sulfoxide and diphenyl disulfide significantly reduced the P. aeruginosa recovered 18 h post infection (relative to the control, and were non-lethal to the fly hosts. The possibility that the observed biofilm inhibitory effects were related to quorum sensing inhibition (QSI was investigated using Escherichia coli-based reporters expressing P. aeruginosa lasR or rhIR response proteins, as well as an endogenous P. aeruginosa reporter from the lasI/lasR QS system. Inhibition of quorum sensing by S-phenyl-L-cysteine sulfoxide was observed in all of the reporter systems tested, whereas diphenyl disulfide did not exhibit QSI in either of the E. coli reporters, and showed very limited inhibition in the P. aeruginosa reporter. Since both compounds inhibit biofilm formation but do not show similar QSI activity, it is concluded that they may be functioning by different pathways. The hypothesis that biofilm inhibition by the two active compounds discovered in this work occurs through QSI is discussed.

  10. Effect of bacteriocin and exopolysaccharides isolated from probiotic on P. aeruginosa PAO1 biofilm.

    Science.gov (United States)

    Sharma, Vivek; Harjai, Kusum; Shukla, Geeta

    2018-03-01

    Microorganisms develop biofilms on indwelling medical devices and are associated with biofilm-related infections, resulting in substantial morbidity and mortality. Therefore, to prevent and control biofilm-associated infections, the present study was designed to assess the anti-biofilm potential of postbiotics derived from probiotic organisms against most prevalent biofilm-forming Pseudomonas aeruginosa PAO1. Eighty lactic acid bacteria isolated from eight neonatal fecal samples possessed antibacterial activity against P. aeruginosa PAO1. Among these, only four lactic acid bacteria produced both bacteriocin and exopolysaccharides but only one isolate was found to maximally attenuate the P. aeruginosa PAO1 biofilm. More specifically, the phenotypic and probiotic characterization showed that the isolated lactic acid bacteria were gram positive, non-motile, and catalase and oxidase negative; tolerated acidic and alkaline pH; has bile salt concentration; showed 53% hydrophobicity; and was found to be non-hemolytic. Phylogenetically, the organism was found to be probiotic Lactobacillus fermentum with accession no. KT998657. Interestingly, pre-coating of a microtiter plate either with bacteriocin or with exopolysaccharides as well as their combination significantly (p < 0.05) reduced the number of viable cells forming biofilms to 41.7% compared with simultaneous coating of postbiotics that had 72.4% biofilm-forming viable cells as observed by flow cytometry and confocal laser scanning microscopy. Therefore, it can be anticipated that postbiotics as the natural biointerventions can be employed as the prophylactic agents for medical devices used to treat gastrointestinal and urinary tract infections.

  11. Baicalin inhibits biofilm formation, attenuates the quorum sensing-controlled virulence and enhances Pseudomonas aeruginosa clearance in a mouse peritoneal implant infection model.

    Directory of Open Access Journals (Sweden)

    Jing Luo

    Full Text Available The quorum sensing (QS circuit plays a role in the precise regulation of genes controlling virulence factors and biofilm formation in Pseudomonas aeruginosa. QS-controlled biofilm formation by Pseudomonas aeruginosa in clinical settings has remained controversial due to emerging drug resistance; therefore, screening diverse compounds for anti-biofilm or anti-QS activities is important. This study demonstrates the ability of sub-minimum inhibitory concentrations (sub-MICs of baicalin, an active natural compound extracted from the traditional Chinese medicinal Scutellaria baicalensis, to inhibit the formation of Pseudomonas aeruginosa biofilms and enhance the bactericidal effects of various conventional antibiotics in vitro. In addition, baicalin exerted dose-dependent inhibitory effects on virulence phenotypes (LasA protease, LasB elastase, pyocyanin, rhamnolipid, motilities and exotoxin A regulated by QS in Pseudomonas aeruginosa. Moreover, the expression levels of QS-regulatory genes, including lasI, lasR, rhlI, rhlR, pqsR and pqsA, were repressed after sub-MIC baicalin treatment, resulting in significant decreases in the QS signaling molecules 3-oxo-C12-HSL and C4-HSL, confirming the ability of baicalin-mediated QS inhibition to alter gene and protein expression. In vivo experiments indicated that baicalin treatment reduces Pseudomonas aeruginosa pathogenicity in Caenorhabditis elegans. Greater worm survival in the baicalin-treated group manifested as an increase in the LT50 from 24 to 96 h. In a mouse peritoneal implant infection model, baicalin treatment enhanced the clearance of Pseudomonas aeruginosa from the implants of mice infected with Pseudomonas aeruginosa compared with the control group. Moreover, the combination of baicalin and antibiotics significantly reduced the numbers of colony-forming units in the implants to a significantly greater degree than antibiotic treatment alone. Pathological and histological analyses revealed

  12. Spaceflight promotes biofilm formation by Pseudomonas aeruginosa.

    Directory of Open Access Journals (Sweden)

    Wooseong Kim

    Full Text Available Understanding the effects of spaceflight on microbial communities is crucial for the success of long-term, manned space missions. Surface-associated bacterial communities, known as biofilms, were abundant on the Mir space station and continue to be a challenge on the International Space Station. The health and safety hazards linked to the development of biofilms are of particular concern due to the suppression of immune function observed during spaceflight. While planktonic cultures of microbes have indicated that spaceflight can lead to increases in growth and virulence, the effects of spaceflight on biofilm development and physiology remain unclear. To address this issue, Pseudomonas aeruginosa was cultured during two Space Shuttle Atlantis missions: STS-132 and STS-135, and the biofilms formed during spaceflight were characterized. Spaceflight was observed to increase the number of viable cells, biofilm biomass, and thickness relative to normal gravity controls. Moreover, the biofilms formed during spaceflight exhibited a column-and-canopy structure that has not been observed on Earth. The increase in the amount of biofilms and the formation of the novel architecture during spaceflight were observed to be independent of carbon source and phosphate concentrations in the media. However, flagella-driven motility was shown to be essential for the formation of this biofilm architecture during spaceflight. These findings represent the first evidence that spaceflight affects community-level behaviors of bacteria and highlight the importance of understanding how both harmful and beneficial human-microbe interactions may be altered during spaceflight.

  13. Involvement of bacterial migration in the development of complex multicellular structures in Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Klausen, Mikkel; Aaes-Jorgensen, A.; Molin, Søren

    2003-01-01

    development, we have performed an investigation with time-lapse confocal laser scanning microscopy of biofilms formed by various combinations of colour-coded P. aeruginosa wild type and motility mutants. We show that mushroom-shaped multicellular structures in P. aeruginosa biofilms can form in a sequential...... process involving a non-motile bacterial subpopulation and a migrating bacterial subpopulation. The non-motile bacteria form the mushroom stalks by growth in certain foci of the biofilm. The migrating bacteria form the mushroom caps by climbing the stalks and aggregating on the tops in a process which...

  14. Contribution of Cell Elongation to the Biofilm Formation of Pseudomonas aeruginosa during Anaerobic Respiration

    Science.gov (United States)

    Park, Yongjin; Yoon, Sang Sun

    2011-01-01

    Pseudomonas aeruginosa, a gram-negative bacterium of clinical importance, forms more robust biofilm during anaerobic respiration, a mode of growth presumed to occur in abnormally thickened mucus layer lining the cystic fibrosis (CF) patient airway. However, molecular basis behind this anaerobiosis-triggered robust biofilm formation is not clearly defined yet. Here, we identified a morphological change naturally accompanied by anaerobic respiration in P. aeruginosa and investigated its effect on the biofilm formation in vitro. A standard laboratory strain, PAO1 was highly elongated during anaerobic respiration compared with bacteria grown aerobically. Microscopic analysis demonstrated that cell elongation likely occurred as a consequence of defective cell division. Cell elongation was dependent on the presence of nitrite reductase (NIR) that reduces nitrite (NO2 −) to nitric oxide (NO) and was repressed in PAO1 in the presence of carboxy-PTIO, a NO antagonist, demonstrating that cell elongation involves a process to respond to NO, a spontaneous byproduct of the anaerobic respiration. Importantly, the non-elongated NIR-deficient mutant failed to form biofilm, while a mutant of nitrate reductase (NAR) and wild type PAO1, both of which were highly elongated, formed robust biofilm. Taken together, our data reveal a role of previously undescribed cell biological event in P. aeruginosa biofilm formation and suggest NIR as a key player involved in such process. PMID:21267455

  15. Evaluation of combinations of putative anti-biofilm agents and antibiotics to eradicate biofilms of Staphylococcus aureus and Pseudomonas aeruginosa.

    Science.gov (United States)

    Belfield, Katherine; Bayston, Roger; Hajduk, Nadzieja; Levell, Georgia; Birchall, John P; Daniel, Matija

    2017-09-01

    To evaluate potential anti-biofilm agents for their ability to enhance the activity of antibiotics for local treatment of localized biofilm infections. Staphylococcus aureus and Pseudomonas aeruginosa in vitro biofilm models were developed. The putative antibiotic enhancers N-acetylcysteine, acetylsalicylic acid, sodium salicylate, recombinant human deoxyribonuclease I, dispersin B, hydrogen peroxide and Johnson's Baby Shampoo (JBS) were tested for their anti-biofilm activity alone and their ability to enhance the activity of antibiotics for 7 or 14 days, against 5 day old biofilms. The antibiotic enhancers were paired with rifampicin and clindamycin against S. aureus and gentamicin and ciprofloxacin against P. aeruginosa. Isolates from biofilms that were not eradicated were tested for antibiotic resistance. Antibiotic levels 10× MIC and 100× MIC significantly reduced biofilm, but did not consistently eradicate it. Antibiotics at 100× MIC with 10% JBS for 14 days was the only treatment to eradicate both staphylococcal and pseudomonal biofilms. Recombinant human deoxyribonuclease I significantly reduced staphylococcal biofilm. Emergence of resistance of surviving isolates was minimal and was often associated with the small colony variant phenotype. JBS enhanced the activity of antibiotics and several other promising anti-biofilm agents were identified. Antibiotics with 10% JBS eradicated biofilms produced by both organisms. Such combinations might be useful in local treatment of localized biofilm infections. © The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  16. Activity of Bacteriophages in Removing Biofilms of Pseudomonas aeruginosa Isolates from Chronic Rhinosinusitis Patients

    NARCIS (Netherlands)

    Fong, Stephanie A.; Drilling, Amanda; Morales, Sandra; Cornet, Marjolein E.; Woodworth, Bradford A.; Fokkens, Wytske J.; Psaltis, Alkis J.; Vreugde, Sarah; Wormald, Peter-John

    2017-01-01

    Introduction:Pseudomonas aeruginosa infections are prevalent amongst chronic rhinosinusitis (CRS) sufferers. Many P. aeruginosa strains form biofilms, leading to treatment failure. Lytic bacteriophages (phages) are viruses that infect, replicate within, and lyse bacteria, causing bacterial death.

  17. Decrease of Pseudomonas aeruginosa biofilm formation by food waste materials

    Czech Academy of Sciences Publication Activity Database

    Maděrová, Z.; Horská, K.; Kim, S.-R.; Lee, Ch.-H.; Pospíšková, K.; Šafaříková, Miroslava; Šafařík, Ivo

    2016-01-01

    Roč. 73, č. 9 (2016), s. 2143-2149 ISSN 0273-1223 Institutional support: RVO:60077344 Keywords : biofilm * food waste materials * magnetic spent grain * Pseudomonas aeruginosa Subject RIV: EI - Biotechnology ; Bionics Impact factor: 1.197, year: 2016

  18. Glutathione-Disrupted Biofilms of Clinical Pseudomonas aeruginosa Strains Exhibit an Enhanced Antibiotic Effect and a Novel Biofilm Transcriptome

    Science.gov (United States)

    Das, Theerthankar; Ibugo, Amaye; Buckle, Edwina; Manefield, Mike; Manos, Jim

    2016-01-01

    Pseudomonas aeruginosa infections result in high morbidity and mortality rates for individuals with cystic fibrosis (CF), with premature death often occurring. These infections are complicated by the formation of biofilms in the sputum. Antibiotic therapy is stymied by antibiotic resistance of the biofilm matrix, making novel antibiofilm strategies highly desirable. Within P. aeruginosa biofilms, the redox factor pyocyanin enhances biofilm integrity by intercalating with extracellular DNA. The antioxidant glutathione (GSH) reacts with pyocyanin, disrupting intercalation. This study investigated GSH disruption by assaying the physiological effects of GSH and DNase I on biofilms of clinical CF isolates grown in CF artificial sputum medium (ASMDM+). Confocal scanning laser microscopy showed that 2 mM GSH, alone or combined with DNase I, significantly disrupted immature (24-h) biofilms of Australian epidemic strain (AES) isogens AES-1R and AES-1M. GSH alone greatly disrupted mature (72-h) AES-1R biofilms, resulting in significant differential expression of 587 genes, as indicated by RNA-sequencing (RNA-seq) analysis. Upregulated systems included cyclic diguanylate and pyoverdine biosynthesis, the type VI secretion system, nitrate metabolism, and translational machinery. Biofilm disruption with GSH revealed a cellular physiology distinct from those of mature and dispersed biofilms. RNA-seq results were validated by biochemical and quantitative PCR assays. Biofilms of a range of CF isolates disrupted with GSH and DNase I were significantly more susceptible to ciprofloxacin, and increased antibiotic effectiveness was achieved by increasing the GSH concentration. This study demonstrated that GSH, alone or with DNase I, represents an effective antibiofilm treatment when combined with appropriate antibiotics, pending in vivo studies. PMID:27161630

  19. The catabolite repression control protein Crc plays a role in the development of antimicrobial-tolerant subpopulations in Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Zhang, Lianbo; Chiang, Wen-Chi; Gao, Qingguo

    2012-01-01

    Bacteria form complex surface-attached biofilm communities in nature. Biofilm cells differentiate into subpopulations which display tolerance towards antimicrobial agents. However, the signal transduction pathways regulating subpopulation differentiation in biofilms are largely unelucidated. In t....... In the present study, we show that the catabolite repression control protein Crc regulates the metabolic state of Pseudomonas aeruginosa cells in biofilms, and plays an important role in the development of antimicrobial-tolerant subpopulations in P. aeruginosa biofilms....

  20. Distinct roles of extracellular polymeric substances in Pseudomonas aeruginosa biofilm development

    DEFF Research Database (Denmark)

    Yang, Liang; Hu, Yifan; Liu, Yang

    2011-01-01

    Bacteria form surface attached biofilm communities as one of the most important survival strategies in nature. Biofilms consist of water, bacterial cells and a wide range of self‐generated extracellular polymeric substances (EPS). Biofilm formation is a dynamic self‐assembly process and several d...... polysaccharide is more important than Pel polysaccharide in P. aeruginosa PAO1 biofilm formation and antibiotic resistance. Our study thus suggests that different EPS materials play distinct roles during bacterial biofilm formation.......Bacteria form surface attached biofilm communities as one of the most important survival strategies in nature. Biofilms consist of water, bacterial cells and a wide range of self‐generated extracellular polymeric substances (EPS). Biofilm formation is a dynamic self‐assembly process and several...... distinguishable stages are observed during bacterial biofilm development. Biofilm formation is shown to be coordinated by EPS production, cell migration, subpopulation differentiation and interactions. However, the ways these different factors affect each other and contribute to community structural...

  1. Anti-biofilm activities from marine cold adapted bacteria against staphylococci and Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Rosanna ePapa

    2015-12-01

    Full Text Available Microbial biofilms have great negative impacts on the world’s economy and pose serious problems to industry, public health and medicine. The interest in the development of new approaches for the prevention and treatment of bacterial adhesion and biofilm formation has increased. Since, bacterial pathogens living in biofilm induce persistent chronic infections due to the resistance to antibiotics and host immune system. A viable approach should target adhesive properties without affecting bacterial vitality in order to avoid the appearance of resistant mutants. Many bacteria secrete anti-biofilm molecules that function in regulating biofilm architecture or mediating the release of cells from it during the dispersal stage of biofilm life cycle. Cold-adapted marine bacteria represent an untapped reservoir of biodiversity able to synthesize a broad range of bioactive compounds, including anti-biofilm molecules.The anti-biofilm activity of cell-free supernatants derived from sessile and planktonic cultures of cold-adapted bacteria belonging to Pseudoalteromonas, Psychrobacter and Psychromonas species were tested against Staphylococcus aureus, Staphylococcus epidermidis and Pseudomonas aeruginosa strains. Reported results demonstrate that we have selected supernatants, from cold-adapted marine bacteria, containing non-biocidal agents able to destabilize biofilm matrix of all tested pathogens without killing cells. A preliminary physico-chemical characterization of supernatants was also performed, and these analyses highlighted the presence of molecules of different nature that act by inhibiting biofilm formation. Some of them are also able to impair the initial attachment of the bacterial cells to the surface, thus likely containing molecules acting as anti-biofilm surfactant molecules.The described ability of cold-adapted bacteria to produce effective anti-biofilm molecules paves the way to further characterization of the most promising molecules

  2. Exposure of Bacterial Biofilms to Electrical Current Leads to Cell Death Mediated in Part by Reactive Oxygen Species.

    Science.gov (United States)

    Brinkman, Cassandra L; Schmidt-Malan, Suzannah M; Karau, Melissa J; Greenwood-Quaintance, Kerryl; Hassett, Daniel J; Mandrekar, Jayawant N; Patel, Robin

    2016-01-01

    Bacterial biofilms may form on indwelling medical devices such as prosthetic joints, heart valves and catheters, causing challenging-to-treat infections. We have previously described the 'electricidal effect', in which bacterial biofilms are decreased following exposure to direct electrical current. Herein, we sought to determine if the decreased bacterial quantities are due to detachment of biofilms or cell death and to investigate the role that reactive oxygen species (ROS) play in the observed effect. Using confocal and electron microscopy and flow cytometry, we found that direct current (DC) leads to cell death and changes in the architecture of biofilms formed by Gram-positive and Gram-negative bacteria. Reactive oxygen species (ROS) appear to play a role in DC-associated cell death, as there was an increase in ROS-production by Staphylococcus aureus and Staphylococcus epidermidis biofilms following exposure to DC. An increase in the production of ROS response enzymes catalase and superoxide dismutase (SOD) was observed for S. aureus, S. epidermidis and Pseudomonas aeruginosa biofilms following exposure to DC. Additionally, biofilms were protected from cell death when supplemented with antioxidants and oxidant scavengers, including catalase, mannitol and Tempol. Knocking out SOD (sodAB) in P. aeruginosa led to an enhanced DC effect. Microarray analysis of P. aeruginosa PAO1 showed transcriptional changes in genes related to the stress response and cell death. In conclusion, the electricidal effect results in death of bacteria in biofilms, mediated, at least in part, by production of ROS.

  3. Metabolomics-Based Screening of Biofilm-Inhibitory Compounds against Pseudomonas aeruginosa from Burdock Leaf

    Directory of Open Access Journals (Sweden)

    Zaixiang Lou

    2015-09-01

    Full Text Available Screening of anti-biofilm compounds from the burdock leaf based on metabolomics is reported here. The crystal violet assay indicated 34% ethanol elution fraction of burdock leaf could completely inhibit biofilm formation of Pseudomonas aeruginosa at 1 mg·mL−1. Then, the chemical composition of burdock leaf fraction was analyzed by ultra-performance liquid chromatography-mass spectrometry (UPLC-MS and 11 active compounds (chlorogenic acid, caffeic acid, p-coumaric acid, quercetin, ursolic acid, rutin, cynarin, luteolin, crocin, benzoic acid, and Tenacissoside I were identified. Lastly, UPLC-MS analysis was employed to obtain the metabolic fingerprints of burdock leaf fractions before and after inhibiting the biofilm of Pseudomonas aeruginosa. The metabolic fingerprints were transformed to data, analyzed with PLS-DA (partial least squares discriminant analysis and the peaks whose area was significantly changed were found out. Thus, 81 compounds were screened as potential anti-biofilm ingredients. Among them, rutin, ursolic acid, caffeic acid, p-coumaric acid and quercetin were identified and confirmed as the main anti-biofilm compounds in burdock leaf. The study provided basic anti-biofilm profile data for the compounds in burdock leaf, as well as provided a convenient method for fast screening of anti-biofilm compounds from natural plants.

  4. Carbapenem-resistant Pseudomonas aeruginosa: association with virulence genes and biofilm formation

    Directory of Open Access Journals (Sweden)

    Iara Rossi Gonçalves

    Full Text Available Abstract Pseudomonas aeruginosa is an opportunistic pathogen that causes frequently nosocomial infections, currently becoming more difficult to treat due to the various resistance mechanisms and different virulence factors. The purpose of this study was to determine the risk factors independently associated with the development of bacteremia by carbapenem-resistant P. aeruginosa, the frequency of virulence genes in metallo-β-lactamases producers and to evaluate their ability to produce biofilm. We conducted a case–control study in the Uberlândia Federal University – Hospital Clinic, Brazil. Polymerase Chain Reaction was performed for metallo-β-lactamases and virulence genes. Adhesion and biofilm assays were done by quantitative tests. Among the 157 strains analyzed, 73.9% were multidrug-resistant, 43.9% were resistant to carbapenems, 16.1% were phenotypically positive for metallo-β-lactamases, and of these, 10.7% were positive for blaSPM gene and 5.3% positive for blaVIM. The multivariable analysis showed that mechanical ventilation, enteral/nasogastric tubes, primary bacteremia with unknown focus, and inappropriate therapy were independent risk factors associated with bacteremia. All tested strains were characterized as strongly biofilm producers. A higher mortality was found among patients with bacteremia by carbapenem-resistant P. aeruginosa strains, associated independently with extrinsic risk factors, however it was not evident the association with the presence of virulence and metallo-β-lactamases genes.

  5. Chemical Analysis of Cellular and Extracellular Carbohydrates of a Biofilm-Forming Strain Pseudomonas aeruginosa PA14

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    Coulon, Charlène; Vinogradov, Evgeny; Filloux, Alain; Sadovskaya, Irina

    2010-01-01

    Background Pseudomonas aeruginosa is a Gram-negative bacterium and an opportunistic pathogen, which causes persisting life-threatening infections in cystic fibrosis (CF) patients. Biofilm mode of growth facilitates its survival in a variety of environments. Most P. aeruginosa isolates, including the non-mucoid laboratory strain PA14, are able to form a thick pellicle, which results in a surface-associated biofilm at the air-liquid (A–L) interface in standing liquid cultures. Exopolysaccharides (EPS) are considered as key components in the formation of this biofilm pellicle. In the non-mucoid P. aeruginosa strain PA14, the “scaffolding” polysaccharides of the biofilm matrix, and the molecules responsible for the structural integrity of rigid A–L biofilm have not been identified. Moreover, the role of LPS in this process is unclear, and the chemical structure of the LPS O-antigen of PA14 has not yet been elucidated. Principal Findings In the present work we carried out a systematic analysis of cellular and extracellular (EC) carbohydrates of P. aeruginosa PA14. We also elucidated the chemical structure of the LPS O-antigen by chemical methods and 2-D NMR spectroscopy. Our results showed that it is composed of linear trisaccharide repeating units, identical to those described for P. aeruginosa Lanýi type O:2a,c (Lanýi-Bergman O-serogroup 10a, 10c; IATS serotype 19) and having the following structure: -4)-α-L-GalNAcA-(1–3)-α-D-QuiNAc-(1–3)- α-L-Rha-(1-. Furthermore, an EC O-antigen polysaccharide (EC O-PS) and the glycerol-phosphorylated cyclic β-(1,3)-glucans were identified in the culture supernatant of PA14, grown statically in minimal medium. Finally, the extracellular matrix of the thick biofilm formed at the A-L interface contained, in addition to eDNA, important quantities (at least ∼20% of dry weight) of LPS-like material. Conclusions We characterized the chemical structure of the LPS O-antigen and showed that the O-antigen polysaccharide is

  6. Electron transfer mediators accelerated the microbiologically influence corrosion against carbon steel by nitrate reducing Pseudomonas aeruginosa biofilm.

    Science.gov (United States)

    Jia, Ru; Yang, Dongqing; Xu, Dake; Gu, Tingyue

    2017-12-01

    Electron transfer is a rate-limiting step in microbiologically influenced corrosion (MIC) caused by microbes that utilize extracellular electrons. Cross-cell wall electron transfer is necessary to transport the electrons released from extracellular iron oxidation into the cytoplasm of cells. Electron transfer mediators were found to accelerate the MIC caused by sulfate reducing bacteria. However, there is no publication in the literature showing the effect of electron transfer mediators on MIC caused by nitrate reducing bacteria (NRB). This work demonstrated that the corrosion of anaerobic Pseudomonas aeruginosa (PAO1) grown as a nitrate reducing bacterium biofilm on C1018 carbon steel was enhanced by two electron transfer mediators, riboflavin and flavin adenine dinucleotide (FAD) separately during a 7-day incubation period. The addition of either 10ppm (w/w) (26.6μM) riboflavin or 10ppm (12.7μM) FAD did not increase planktonic cell counts, but they increased the maximum pit depth on carbon steel coupons considerably from 17.5μm to 24.4μm and 25.0μm, respectively. Riboflavin and FAD also increased the specific weight loss of carbon steel from 2.06mg/cm 2 to 2.34mg/cm 2 and 2.61mg/cm 2 , respectively. Linear polarization resistance, electrochemical impedance spectroscopy and potentiodynamic polarization curves all corroborated the pitting and weight loss data. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Inhibitory activity of Iranian plant extracts on growth and biofilm formation by Pseudomonas aeruginosa

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    Mansouri, S.

    2013-01-01

    Full Text Available Aims: Pseudomonas aeruginosa is a drug resistance opportunistic bacterium. Biofilm formation is key factor for survivalof P. aeruginosa in various environments. Polysaccharides may be involved in biofilm formation. The purpose of thisstudy was to evaluate antimicrobial and anti-biofilm activities of seven plant extracts with known alpha-glucosidaseinhibitory activities on different strains of P. aeruginosa.Methodology and results: Plants were extracted with methanol by the maceration method. Antimicrobial activities weredetermined by agar dilution and by growth yield as measured by OD560nm of the Luria Bertani broth (LB culture with orwithout extracts. In agar dilution method, extracts of Quercus infectoria inhibited the growth of all, while Myrtuscommunis extract inhibited the growth of 3 out of 8 bacterial strains with minimum inhibitory concentration (MIC of 1000μg/mL. All extracts significantly (p≤0.003 reduced growth rate of the bacteria in comparison with the control withoutextracts in LB broth at sub-MIC concentrations (500 μg/mL. All plant extracts significantly (p≤0.003 reduced biofilmformation compared to the controls. Glycyrrhiza glabra and Q. infectoria had the highest anti-biofilm activities. Nocorrelation between the alpha-glucosidase inhibitory activity with growth or the intensity of biofilm formation was found.Conclusion, significance and impact of study: Extracts of Q. infectoria and M. communis had the most antimicrobial,while Q. infectoria and G. glabra had the highest anti-biofilm activities. All plant extracts had anti-biofilm activities withmarginal effect on growth, suggesting that the mechanisms of these activities are unrelated to static or cidal effects.Further work to understand the relation between antimicrobial and biofilm formation is needed for development of newmeans to fight the infectious caused by this bacterium in future.

  8. Magnesium limitation is an environmental trigger of the Pseudomonas aeruginosa biofilm lifestyle.

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    Heidi Mulcahy

    Full Text Available Biofilm formation is a conserved strategy for long-term bacterial survival in nature and during infections. Biofilms are multicellular aggregates of cells enmeshed in an extracellular matrix. The RetS, GacS and LadS sensors control the switch from a planktonic to a biofilm mode of growth in Pseudomonas aeruginosa. Here we detail our approach to identify environmental triggers of biofilm formation by investigating environmental conditions that repress expression of the biofilm repressor RetS. Mg(2+ limitation repressed the expression of retS leading to increased aggregation, exopolysaccharide (EPS production and biofilm formation. Repression of retS expression under Mg(2+ limitation corresponded with induced expression of the GacA-controlled small regulatory RNAs rsmZ and rsmY and the EPS biosynthesis operons pel and psl. We recently demonstrated that extracellular DNA sequesters Mg(2+ cations and activates the cation-sensing PhoPQ two-component system, which leads to increased antimicrobial peptide resistance in biofilms. Here we show that exogenous DNA and EDTA, through their ability to chelate Mg(2+, promoted biofilm formation. The repression of retS in low Mg(2+ was directly controlled by PhoPQ. PhoP also directly controlled expression of rsmZ but not rsmY suggesting that PhoPQ controls the equilibrium of the small regulatory RNAs and thus fine-tunes the expression of genes in the RetS pathway. In summary, Mg(2+ limitation is a biologically relevant environmental condition and the first bonafide environmental signal identified that results in transcriptional repression of retS and promotes P. aeruginosa biofilm formation.

  9. A Phytoanticipin Derivative, Sodium Houttuyfonate, Induces in Vitro Synergistic Effects with Levofloxacin against Biofilm Formation by Pseudomonas aeruginosa

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    Jing Shao

    2012-09-01

    Full Text Available Antibiotic resistance has become the main deadly factor in infections, as bacteria can protect themselves by hiding in a self-constructed biofilm. Consequently, more attention is being paid to the search for “non-antibiotic drugs” to solve this problem. Phytoanticipins, the natural antibiotics from plants, could be a suitable alternative, but few works on this aspect have been reported. In this study, a preliminary study on the synergy between sodium houttuyfonate (SH and levofloxacin (LFX against the biofilm formation of Pseudomonas aeruginosa was performed. The minimal inhibitory concentrations (MIC of LFX and SH, anti-biofilm formation and synergistic effect on Pseudomonas aeruginosa, and quantification of alginate were determined by the microdilution method, crystal violet (CV assay, checkerboard method, and hydroxybiphenyl colorimetry. The biofilm morphology of Pseudomonas aeruginosa was observed by fluorescence microscope and scanning electric microscope (SEM. The results showed that: (i LFX and SH had an obvious synergistic effect against Pseudomonas aeruginosa with MIC values of 0.25 μg/mL and 128 μg/mL, respectively; (ii ½ × MIC SH combined with 2 × MIC LFX could suppress the biofilm formation of Pseudomonas aeruginosa effectively, with up to 73% inhibition; (iii the concentration of alginate decreased dramatically by a maximum of 92% after treatment with the combination of antibiotics; and (iv more dead cells by fluorescence microscope and more removal of extracellular polymeric structure (EPS by SEM were observed after the combined treatment of LFX and SH. Our experiments demonstrate the promising future of this potent antimicrobial agent against biofilm-associated infections.

  10. Pathogenic factors of Pseudomonas aeruginosa – the role of biofilm in pathogenicity and as a target for phage therapy

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    Fairoz Al-Wrafy

    2017-02-01

    Full Text Available Pseudomonas aeruginosa is an opportunistic pathogen that can cause several acute and chronic infections in humans, and it has become an important cause of nosocomial infections and antibiotic resistance. Biofilm represents an important virulence factor for these bacteria, plays a role in P. aeruginosa infections and avoidance of immune defence mechanisms, and has the ability to protect the bacteria from antibiotics. Alginate, Psl and Pel, three exopolysaccharides, are the main components in biofilm matrix, with many biological functions attributed to them, especially with respect to the protection of the bacterial cell from antibiotics and the immune system. Pseudomonas infections, biofilm formation and development of resistance to antibiotics all require better understanding to achieve the best results using alternative treatment with phage therapy. This review describes the P. aeruginosa pathogenicity and virulence factors with a special focus on the biofilm and its role in infection and resistance to antibiotics and summarizes phage therapy as an alternative approach in treatment of P. aeruginosa infections.

  11. Antimicrobial properties of Kalanchoe blossfeldiana: a focus on drug resistance with particular reference to quorum sensing-mediated bacterial biofilm formation.

    Science.gov (United States)

    Sarkar, Ratul; Mondal, Chaitali; Bera, Rammohan; Chakraborty, Sumon; Barik, Rajib; Roy, Paramita; Kumar, Alekh; Yadav, Kirendra K; Choudhury, Jayanta; Chaudhary, Sushil K; Samanta, Samir K; Karmakar, Sanmoy; Das, Satadal; Mukherjee, Pulok K; Mukherjee, Joydeep; Sen, Tuhinadri

    2015-07-01

    This study attempts to investigate the antimicrobial properties of Kalanchoe blossfeldiana with a particular reference to quorum sensing (QS)-mediated biofilm formation. The methanol extract of K. blossfeldiana leaves (MEKB) was evaluated for antimicrobial properties including QS-controlled production of biofilm (including virulence factor, motility and lactone formation) in Pseudomonas aeruginosa. Methanol extract of K. blossfeldiana was also evaluated for anti-cytokine (tumour necrosis factor-alpha, interleukin-6 and interleukin-1 beta) properties in peripheral blood mononuclear cells (PBMC). Methanol extract of K. blossfeldiana exhibited antimicrobial effect on clinical isolates, as well as standard reference strains. Pseudomonas aeruginosa exposed to MEKB (subminimum inhibitory concentration (MIC)) displayed reduced biofilm formation, whereas supra-MIC produced destruction of preformed biofilms. Methanol extract of K. blossfeldiana reduced the secretion of virulence factors (protease and pyoverdin) along with generation of acyl homoserine lactone (AHL). Confocal laser scanning microscopy images indicate reduction of biofilm thickness. The extract also reduced cytokine formation in lipopolysaccharide-stimulated PBMC. Kalanchoe blossfeldiana was found to interfere with AHL production, which in turn may be responsible for downregulating QS-mediated production of biofilm and virulence. This first report on the antibiofilm and anticytokine properties of this plant may open up new vistas for future exploration of this plant for combating biofilm-related resistant infections. © 2015 Royal Pharmaceutical Society.

  12. Stigmatellin Y - An anti-biofilm compound from Bacillus subtilis BR4 possibly interferes in PQS-PqsR mediated quorum sensing system in Pseudomonas aeruginosa.

    Science.gov (United States)

    Boopathi, Seenivasan; Vashisth, Rajesh; Manoharan, Prabu; Kandasamy, Ruckmani; Sivakumar, Natesan

    2017-05-15

    Hitherto this is the first report pertaining to production of biofilm inhibitory compound(s) (BIC) from Bacillus subtilis BR4 against Pseudomonas aeruginosa (ATCC 27853) coupled with production optimization. In order to achieve this, combinations of media components were formulated by employing statistical tools such as Plackett-Burman analysis and central composite rotatable design (CCRD). It was evident that at 35mlL -1 glycerol and 3.8gL -1 casamino acid, anti-biofilm activity and production of extracellular protein significantly increased by 1.5-fold and 1.2-fold, respectively. These results corroborate that the combination of glycerol and casamino acid plays a key role in the production of BIC. Further, metabolic profiling of BIC was carried out using liquid chromatography/tandem mass spectrometry (LC-MS/MS) based on m/z value. The presence of Stigmatellin Y was predicted with monoisotopic neutral mass of 484.2825Da. In support of optimization study, higher production of BIC was confirmed in the optimized-media-grown BR4 (OPT-BR4) than in the ideal-media-grown BR4 (ID-BR4) by LC-MS/MS analysis. PqsR in P. aeruginosa is a potential target for anti-virulent therapy. Molecular docking study has revealed that Stigmatellin Y interacts with PqsR in the similar orientation like a cognate signal (PQS) and synthetic inhibitor. In addition, Stigmatellin Y was found to exhibit interaction with four more amino acid residues of PqsR to establish strong affinity. Stigmatellin Y thus might play a role of competitor for PQS to distract PQS-PqsR mediated communication in P. aeruginosa. The present investigation thus paves new avenues to develop anti-Pseudomonas virulent therapy. Copyright © 2017 Elsevier Ltd. All rights reserved.

  13. Differentiation and distribution of colistin- and sodium dodecyl sulfate-tolerant cells in Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Haagensen, Janus Anders Juul; Klausen, M; Ernst, RK

    2007-01-01

    During Pseudomonas aeruginosa flow cell biofilm development, the cell population differentiates into a nonmotile subpopulation which forms microcolonies and a migrating subpopulation which eventually colonizes the top of the microcolonies, resulting in the development of mushroom-shaped multicell......During Pseudomonas aeruginosa flow cell biofilm development, the cell population differentiates into a nonmotile subpopulation which forms microcolonies and a migrating subpopulation which eventually colonizes the top of the microcolonies, resulting in the development of mushroom......-targeting antibacterial agents. All biofilm-associated cells were sensitive to the antibacterial agents when tested in standard plate assays. A mutation eliminating the production of type IV pili, and hence surface-associated motility, prevented the formation of regular mushroom-shaped structures in the flow cell...... that only the cap-forming subpopulation in biofilms treated with colistin expresses the pmr operon. These results suggest that increased antibiotic tolerance in biofilms may be a consequence of differentiation into distinct subpopulations with different phenotypic properties....

  14. Clustered Regularly Interspaced Short Palindromic Repeat-Dependent, Biofilm-Specific Death of Pseudomonas aeruginosa Mediated by Increased Expression of Phage-Related Genes.

    Science.gov (United States)

    Heussler, Gary E; Cady, Kyle C; Koeppen, Katja; Bhuju, Sabin; Stanton, Bruce A; O'Toole, George A

    2015-05-12

    The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated (CRISPR/Cas) system is an adaptive immune system present in many archaea and bacteria. CRISPR/Cas systems are incredibly diverse, and there is increasing evidence of CRISPR/Cas systems playing a role in cellular functions distinct from phage immunity. Previously, our laboratory reported one such alternate function in which the type 1-F CRISPR/Cas system of the opportunistic pathogen Pseudomonas aeruginosa strain UCBPP-PA14 (abbreviated as P. aeruginosa PA14) inhibits both biofilm formation and swarming motility when the bacterium is lysogenized by the bacteriophage DMS3. In this study, we demonstrated that the presence of just the DMS3 protospacer and the protospacer-adjacent motif (PAM) on the P. aeruginosa genome is necessary and sufficient for this CRISPR-dependent loss of these group behaviors, with no requirement of additional DMS3 sequences. We also demonstrated that the interaction of the CRISPR system with the DMS3 protospacer induces expression of SOS-regulated phage-related genes, including the well-characterized pyocin operon, through the activity of the nuclease Cas3 and subsequent RecA activation. Furthermore, our data suggest that expression of the phage-related genes results in bacterial cell death on a surface due to the inability of the CRISPR-engaged strain to downregulate phage-related gene expression, while these phage-related genes have minimal impact on growth and viability under planktonic conditions. Deletion of the phage-related genes restores biofilm formation and swarming motility while still maintaining a functional CRISPR/Cas system, demonstrating that the loss of these group behaviors is an indirect effect of CRISPR self-targeting. The various CRISPR/Cas systems found in both archaea and bacteria are incredibly diverse, and advances in understanding the complex mechanisms of these varied systems has not only increased our knowledge of host

  15. l-Methionine anti-biofilm activity against Pseudomonas aeruginosa is enhanced by the cystic fibrosis transmembrane conductance regulator potentiator, ivacaftor.

    Science.gov (United States)

    Cho, Do-Yeon; Lim, Dong-Jin; Mackey, Calvin; Weeks, Christopher G; Peña Garcia, Jaime A; Skinner, Daniel; Grayson, Jessica W; Hill, Harrison S; Alexander, David K; Zhang, Shaoyan; Woodworth, Bradford A

    2018-05-01

    Biofilms may contribute to refractory chronic rhinosinusitis (CRS), as they lead to antibiotic resistance and failure of effective clinical treatment. l-Methionine is an amino acid with reported biofilm-inhibiting properties. Ivacaftor is a cystic fibrosis transmembrane conductance regulator (CFTR) potentiator with mild antimicrobial activity via inhibition of bacterial DNA gyrase and topoisomerase IV. The objective of this study was to evaluate whether co-treatment with ivacaftor and l-methionine can reduce the formation of Pseudomonas aeruginosa biofilms. P aeruginosa (PAO-1 strain) biofilms were studied in the presence of l-methionine and/or ivacaftor. For static biofilm assays, PAO-1 was cultured in a 48-well plate for 72 hours with stepwise combinations of these agents. Relative biofilm inhibitions were measured according to optical density of crystal violet stain at 590 nm. Live/dead assays (BacTiter-Glo™ assay, Promega) were imaged with laser scanning confocal microscopy. An agar diffusion test was used to confirm antibacterial effects of the drugs. l-Methionine (0.5 μM) significantly reduced PAO-1 biofilm mass (32.4 ± 18.0%; n = 4; p l-methionine (two-way analysis of variane, p = 0.0415) compared with corresponding concentrations of l-methionine alone. Ivacaftor enhanced the anti-biofilm activity of l-methionine against the PAO-1 strain of P aeruginosa. Further studies evaluating the efficacy of ivacaftor/l-methionine combinations for P aeruginosa sinusitis are planned. © 2018 ARS-AAOA, LLC.

  16. Pel is a cationic exopolysaccharide that cross-links extracellular DNA in the Pseudomonas aeruginosa biofilm matrix.

    Science.gov (United States)

    Jennings, Laura K; Storek, Kelly M; Ledvina, Hannah E; Coulon, Charlène; Marmont, Lindsey S; Sadovskaya, Irina; Secor, Patrick R; Tseng, Boo Shan; Scian, Michele; Filloux, Alain; Wozniak, Daniel J; Howell, P Lynne; Parsek, Matthew R

    2015-09-08

    Biofilm formation is a complex, ordered process. In the opportunistic pathogen Pseudomonas aeruginosa, Psl and Pel exopolysaccharides and extracellular DNA (eDNA) serve as structural components of the biofilm matrix. Despite intensive study, Pel's chemical structure and spatial localization within mature biofilms remain unknown. Using specialized carbohydrate chemical analyses, we unexpectedly found that Pel is a positively charged exopolysaccharide composed of partially acetylated 1→4 glycosidic linkages of N-acetylgalactosamine and N-acetylglucosamine. Guided by the knowledge of Pel's sugar composition, we developed a tool for the direct visualization of Pel in biofilms by combining Pel-specific Wisteria floribunda lectin staining with confocal microscopy. The results indicate that Pel cross-links eDNA in the biofilm stalk via ionic interactions. Our data demonstrate that the cationic charge of Pel is distinct from that of other known P. aeruginosa exopolysaccharides and is instrumental in its ability to interact with other key biofilm matrix components.

  17. Prevalence of Pseudomonas aeruginosa and Acinetobacter spp. in subgingival biofilm and saliva of subjects with chronic periodontal infection

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    Renata Souto

    2014-06-01

    Full Text Available P. aeruginosa and Acinetobacter spp. are important pathogens associated with late nosocomial pneumonia in hospitalized and institutionalized individuals. The oral cavity may be a major source of these respiratory pathogens, particularly in the presence of poor oral hygiene and periodontal infection. This study investigated the prevalence of P. aeruginosa and Acinetobacter spp. in subgingival biofilm and saliva of subjects with periodontal disease or health. Samples were obtained from 55 periodontally healthy (PH and 169 chronic periodontitis (CP patients. DNA was obtained from the samples and detection of P. aeruginosa and Acinetobacter spp. was carried out by multiplex and nested PCR. P. aeruginosa and Acinetobacter spp. were detected in 40% and 45% of all samples, respectively. No significant differences in the distribution of these microorganisms between men and women, subgingival biofilm and saliva samples, patients 35 years of age, and smokers and non-smokers were observed regardless periodontal status (p > 0.05. In contrast, the frequencies of P. aeruginosa and Acinetobacter spp. in saliva and biofilm samples were significantly greater in CP than PH patients (p < 0.01. Smokers presenting P. aeruginosa and high frequencies of supragingival plaque were more likely to present CP than PH. P. aeruginosa and Acinetobacter spp. are frequently detected in the oral microbiota of CP. Poor oral hygiene, smoking and the presence of P. aeruginosa are strongly associated with periodontitis.

  18. Selective labelling and eradication of antibiotic-tolerant bacterial populations in Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Chua, Song Lin; Yam, Joey Kuok Hoong; Hao, Piliang

    2016-01-01

    Drug resistance and tolerance greatly diminish the therapeutic potential of antibiotics against pathogens. Antibiotic tolerance by bacterial biofilms often leads to persistent infections, but its mechanisms are unclear. Here we use a proteomics approach, pulsed stable isotope labelling with amino...... acids (pulsed-SILAC), to quantify newly expressed proteins in colistin-tolerant subpopulations of Pseudomonas aeruginosa biofilms (colistin is a 'last-resort' antibiotic against multidrug-resistant Gram-negative pathogens). Migration is essential for the formation of colistin-tolerant biofilm...

  19. Increased bactericidal activity of colistin on Pseudomonas aeruginosa biofilms in anaerobic conditions

    DEFF Research Database (Denmark)

    Kolpen, Mette; Appeldorff, Cecilie F.; Brandt, Sarah

    2016-01-01

    that production of OH˙may not contribute significantly to the bactericidal activity of colistin on P. aeruginosa biofilm. Thus, we investigated the effect of colistin treatment on biofilm of wild-type PAO1, a catalase-deficient mutant (katA) and a colistin-resistant CF isolate cultured in microtiter plates...

  20. Motility of Pseudomonas aeruginosa contributes to SOS-inducible biofilm formation.

    Science.gov (United States)

    Chellappa, Shakinah T; Maredia, Reshma; Phipps, Kara; Haskins, William E; Weitao, Tao

    2013-12-01

    DNA-damaging antibiotics such as ciprofloxacin induce biofilm formation and the SOS response through autocleavage of SOS-repressor LexA in Pseudomonas aeruginosa. However, the biofilm-SOS connection remains poorly understood. It was investigated with 96-well and lipid biofilm assays. The effects of ciprofloxacin were examined on biofilm stimulation of the SOS mutant and wild-type strains. The stimulation observed in the wild-type in which SOS was induced was reduced in the mutant in which LexA was made non-cleavable (LexAN) and thus SOS non-inducible. Therefore, the stimulation appeared to involve SOS. The possible mechanisms of inducible biofilm formation were explored by subproteomic analysis of outer membrane fractions extracted from biofilms. The data predicted an inhibitory role of LexA in flagellum function. This premise was tested first by functional and morphological analyses of flagellum-based motility. The flagellum swimming motility decreased in the LexAN strain treated with ciprofloxacin. Second, the motility-biofilm assay was performed, which tested cell migration and biofilm formation. The results showed that wild-type biofilm increased significantly over the LexAN. These results suggest that LexA repression of motility, which is the initial event in biofilm development, contributes to repression of SOS-inducible biofilm formation. Copyright © 2013 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  1. A three-phase in-vitro system for studying Pseudomonas aeruginosa adhesion and biofilm formation upon hydrogel contact lenses

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    Kohlmann Thomas

    2010-11-01

    Full Text Available Abstract Background Pseudomonas aeruginosa is commonly associated with contact lens (CL -related eye infections, for which bacterial adhesion and biofilm formation upon hydrogel CLs is a specific risk factor. Whilst P. aeruginosa has been widely used as a model organism for initial biofilm formation on CLs, in-vitro models that closely reproduce in-vivo conditions have rarely been presented. Results In the current investigation, a novel in-vitro biofilm model for studying the adherence of P. aeruginosa to hydrogel CLs was established. Nutritional and interfacial conditions similar to those in the eye of a CL wearer were created through the involvement of a solid:liquid and a solid:air interface, shear forces and a complex artificial tear fluid. Bioburdens varied depending on the CL material and biofilm maturation occurred after 72 h incubation. Whilst a range of biofilm morphologies were visualised including dispersed and adherent bacterial cells, aggregates and colonies embedded in extracellular polymer substances (EPS, EPS fibres, mushroom-like formations, and crystalline structures, a compact and heterogeneous biofilm morphology predominated on all CL materials. Conclusions In order to better understand the process of biofilm formation on CLs and to test the efficacy of CL care solutions, representative in-vitro biofilm models are required. Here, we present a three-phase biofilm model that simulates the environment in the eye of a CL wearer and thus generates biofilms which resemble those commonly observed in-situ.

  2. The Oxidative Stress Agent Hypochlorite Stimulates c-di-GMP Synthesis and Biofilm Formation in Pseudomonas aeruginosa

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    Nikola Strempel

    2017-11-01

    Full Text Available The opportunistic human pathogen Pseudomonas aeruginosa is able to survive under a variety of often harmful environmental conditions due to a multitude of intrinsic and adaptive resistance mechanisms, including biofilm formation as one important survival strategy. Here, we investigated the adaptation of P. aeruginosa PAO1 to hypochlorite (HClO, a phagocyte-derived host defense compound and frequently used disinfectant. In static biofilm assays, we observed a significant enhancement in initial cell attachment in the presence of sublethal HClO concentrations. Subsequent LC-MS analyses revealed a strong increase in cyclic-di-GMP (c-di-GMP levels suggesting a key role of this second messenger in HClO-induced biofilm development. Using DNA microarrays, we identified a 26-fold upregulation of ORF PA3177 coding for a putative diguanylate cyclase (DGC, which catalyzes the synthesis of the second messenger c-di-GMP – an important regulator of bacterial motility, sessility and persistence. This DGC PA3177 was further characterized in more detail demonstrating its impact on P. aeruginosa motility and biofilm formation. In addition, cell culture assays attested a role for PA3177 in the response of P. aeruginosa to human phagocytes. Using a subset of different mutants, we were able to show that both Pel and Psl exopolysaccharides are effectors in the PA3177-dependent c-di-GMP network.

  3. Biofilm and metallo beta-lactamase production among the strains of Pseudomonas aeruginosa and Acinetobacter spp. at a Tertiary Care Hospital in Kathmandu, Nepal

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    Bandana Baniya

    2017-11-01

    Full Text Available Abstract Introduction Pseudomonas aeruginosa and Acinetobacter spp. are found to be associated with biofilm and metallo-β-lactamase production and are the common causes of serious infections mainly in hospitalized patients. So, the main aims of this study were to determine the rates of biofilm production and metallo beta-lactamase production (MBL among the strains of Pseudomonas aeruginosa and Acinetobacter spp. isolated from hospitalized patients. Methods A total of 85 P. aeruginosa isolates and 50 Acinetobacter spp. isolates isolated from different clinical specimens from patients admitted to Shree Birendra Hospital, Kathmandu, Nepal from July 2013 to May 2014 were included in this study. The bacterial isolates were identified with the help of biochemical tests. Modified Kirby-Bauer disc diffusion technique was used for antimicrobial susceptibility testing. Combined disc diffusion technique was used for the detection of MBL production, while Congo red agar method and tube adherence method were used for detection of biofilm production. Results Around 16.4% of P. aeruginosa isolates and 22% of the strains of Acinetobacter spp. were metallo β-lactamase producers. Out of 85 P. aeruginosa isolates, 23 (27.05% were biofilm producers according to tube adherence test while, only 13 (15.29% were biofilm producers as per Congo red agar method. Similarly, out of 50 Acinetobacter spp. 7 (14% isolates were biofilm producers on the basis of tube adherence test, while only 5 (10% were positive for biofilm production by Congo red agar method. Highest rates of susceptibility of P. aeruginosa as well as Acinetobacter spp. were seen toward colistin. Conclusion In our study, biofilm production and metallo beta-lactamase production were observed among Pseudomonas aeruginosa and Acinetobacter spp. However, no statistically significant association could be established between biofilm production and metallo beta-lactamase production.

  4. One-step purification and characterization of alginate lyase from a clinical Pseudomonas aeruginosa with destructive activity on bacterial biofilm

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    Parinaz Ghadam

    2017-05-01

    Full Text Available Objective(s: Pseudomonas aeruginosais a Gram-negative and aerobic rod bacterium that displays mucoid and non-mucoid phenotype. Mucoid strains secrete alginate, which is the main agent of biofilms in chronic P. aeruginosa infections, show high resistance to antibiotics; consequently, the biological disruption of mucoid P. aeruginosa biofilms is an attractive area of study for researchers. Alginate lyase gene (algl is a member of alginate producing operon which by glycosidase activity produces primer for other enzymes in this cluster. Also this activity can destroy the extracellular alginate; therefore this enzyme participates in alginate production and destruction pathway. Alginate lyase causes detachment of a biofilm by reducing its adhesion to the surfaces, and increases phagocytosis and antibiotic susceptibility. In this study, alginate lyase was purified in just one step and its properties were investigated. Materials and Methods: The purification was done by affinity chromatography, analysed by SDS-PAGE, and its effect on P. aeruginosa biofilms was surveyed by micro titer plate assay and SEM. The substrate specificity of the enzyme was determined by PCR. Results: Alginate lyase from isolate 48 was purified in one step. It is more thermally resistant than alginate lyase from Pseudomonas aeruginosa PAO1 and poly M, poly G and poly MG alginate were the substrate of this enzyme. Moreover, it has an eradication effect on biofilms from P. aeruginosa 48 and PAO1. Conclusion: In this study an alginate lyase with many characteristics suitable in medicine such as thermal stability, effective on poly M alginate, and bacterial biofilm destructive was introduced and purified.

  5. Searching for new strategies against biofilm infections: Colistin-AMP combinations against Pseudomonas aeruginosa and Staphylococcus aureus single- and double-species biofilms.

    Directory of Open Access Journals (Sweden)

    Paula Jorge

    Full Text Available Antimicrobial research is being pressured to look for more effective therapeutics for the ever-growing antibiotic-resistant infections, and antimicrobial peptides (AMP and antimicrobial combinations are promising solutions. This work evaluates colistin-AMP combinations against two major pathogens, Pseudomonas aeruginosa and Staphylococcus aureus, encompassing non- and resistant strains. Colistin (CST combined with the AMP temporin A (TEMP-A, citropin 1.1 (CIT-1.1 and tachyplesin I linear analogue (TP-I-L was tested against planktonic, single- and double-species biofilm cultures. Overall synergy for planktonic P. aeruginosa and synergy/additiveness for planktonic S. aureus were observed. Biofilm growth prevention was achieved with synergy and additiveness. Pre-established 24 h-old biofilms were harder to eradicate, especially for S. aureus and double-species biofilms; still, some synergy and addictiveness was observed for higher concentrations, including for the biofilms of resistant strains. Different treatment times and growth media did not greatly influence AMP activity. CST revealed low toxicity compared with the other AMP but its combinations were toxic for high concentrations. Overall, combinations reduced effective AMP concentrations, mainly in prevention scenarios. Improvement of effectiveness and toxicity of therapeutic strategies will be further investigated.

  6. Biofilm formation by Staphylococcus epidermidis on peritoneal dialysis catheters and the effects of extracellular products from Pseudomonas aeruginosa.

    Science.gov (United States)

    Pihl, Maria; Arvidsson, Anna; Skepö, Marie; Nilsson, Martin; Givskov, Michael; Tolker-Nielsen, Tim; Svensäter, Gunnel; Davies, Julia R

    2013-04-01

    Biofilm formation by Staphylococcus epidermidis is a cause of infections related to peritoneal dialysis (PD). We have used a PD catheter flow-cell model in combination with confocal scanning laser microscopy and atomic force microscopy to study biofilm formation by S. epidermidis. Adherence to serum-coated catheters was four times greater than to uncoated ones, suggesting that S. epidermidis binds to serum proteins on the catheter surface. Pseudomonas aeruginosa biofilm supernatant interfered with the formation of a serum protein coat thereby reducing the capacity for biofilm formation in S. epidermidis. Supernatants from ΔpelA, ΔpslBCD and ΔrhlAB strains of P. aeruginosa showed no differences from the wild-type supernatant indicating that the effect on serum coat formation was not due to rhamnolipids or the PelA and PslBCD polysaccharides. Supernatant from P. aeruginosa also dispersed established S. epidermidis biofilms. Supernatants lacking PelA or PslBCD showed no differences from the wild type but that from a ΔrhlAB strain, showed reduced, but not abolished, capacity for dispersal. This suggests that rhamnolipids are involved but not wholly responsible for the effect. Thus, supernatants from P. aeruginosa contain promising substances for the prevention and treatment of biofilm infections, although further work is required to identity more active components. © 2013 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  7. Anti-Biofilm and Immunomodulatory Activities of Peptides That Inhibit Biofilms Formed by Pathogens Isolated from Cystic Fibrosis Patients

    Directory of Open Access Journals (Sweden)

    César de la Fuente-Núñez

    2014-10-01

    Full Text Available Cystic fibrosis (CF patients often acquire chronic respiratory tract infections due to Pseudomonas aeruginosa and Burkholderia cepacia complex (Bcc species. In the CF lung, these bacteria grow as multicellular aggregates termed biofilms. Biofilms demonstrate increased (adaptive resistance to conventional antibiotics, and there are currently no available biofilm-specific therapies. Using plastic adherent, hydroxyapatite and flow cell biofilm models coupled with confocal and scanning electron microscopy, it was demonstrated that an anti-biofilm peptide 1018 prevented biofilm formation, eradicated mature biofilms and killed biofilms formed by a wide range of P. aeruginosa and B. cenocepacia clinical isolates. New peptide derivatives were designed that, compared to their parent peptide 1018, showed similar or decreased anti-biofilm activity against P. aeruginosa biofilms, but increased activity against biofilms formed by the Gram-positive bacterium methicillin resistant Staphylococcus aureus. In addition, some of these new peptide derivatives retained the immunomodulatory activity of 1018 since they induced the production of the chemokine monocyte chemotactic protein-1 (MCP-1 and suppressed lipopolysaccharide-mediated tumor necrosis factor-α (TNF-α production by human peripheral blood mononuclear cells (PBMC and were non-toxic towards these cells. Peptide 1018 and its derivatives provide promising leads for the treatment of chronic biofilm infections and hyperinflammatory lung disease in CF patients.

  8. Novel multiscale modeling tool applied to Pseudomonas aeruginosa biofilm formation.

    Directory of Open Access Journals (Sweden)

    Matthew B Biggs

    Full Text Available Multiscale modeling is used to represent biological systems with increasing frequency and success. Multiscale models are often hybrids of different modeling frameworks and programming languages. We present the MATLAB-NetLogo extension (MatNet as a novel tool for multiscale modeling. We demonstrate the utility of the tool with a multiscale model of Pseudomonas aeruginosa biofilm formation that incorporates both an agent-based model (ABM and constraint-based metabolic modeling. The hybrid model correctly recapitulates oxygen-limited biofilm metabolic activity and predicts increased growth rate via anaerobic respiration with the addition of nitrate to the growth media. In addition, a genome-wide survey of metabolic mutants and biofilm formation exemplifies the powerful analyses that are enabled by this computational modeling tool.

  9. Novel multiscale modeling tool applied to Pseudomonas aeruginosa biofilm formation.

    Science.gov (United States)

    Biggs, Matthew B; Papin, Jason A

    2013-01-01

    Multiscale modeling is used to represent biological systems with increasing frequency and success. Multiscale models are often hybrids of different modeling frameworks and programming languages. We present the MATLAB-NetLogo extension (MatNet) as a novel tool for multiscale modeling. We demonstrate the utility of the tool with a multiscale model of Pseudomonas aeruginosa biofilm formation that incorporates both an agent-based model (ABM) and constraint-based metabolic modeling. The hybrid model correctly recapitulates oxygen-limited biofilm metabolic activity and predicts increased growth rate via anaerobic respiration with the addition of nitrate to the growth media. In addition, a genome-wide survey of metabolic mutants and biofilm formation exemplifies the powerful analyses that are enabled by this computational modeling tool.

  10. The ABC of Biofilm Drug Tolerance: the MerR-Like Regulator BrlR Is an Activator of ABC Transport Systems, with PA1874-77 Contributing to the Tolerance of Pseudomonas aeruginosa Biofilms to Tobramycin.

    Science.gov (United States)

    Poudyal, Bandita; Sauer, Karin

    2018-02-01

    A hallmark of biofilms is their tolerance to killing by antimicrobial agents. In Pseudomonas aeruginosa , biofilm drug tolerance requires the c-di-GMP-responsive MerR transcriptional regulator BrlR. However, the mechanism by which BrlR mediates biofilm drug tolerance has not been elucidated. Here, we demonstrate that BrlR activates the expression of at least 7 ABC transport systems, including the PA1874-PA1875-PA1876-PA1877 (PA1874-77) operon, with chromatin immunoprecipitation and DNA binding assays confirming BrlR binding to the promoter region of PA1874-77. Insertional inactivation of the 7 ABC transport systems rendered P. aeruginosa PAO1 biofilms susceptible to tobramycin or norfloxacin. Susceptibility was linked to drug accumulation, with BrlR contributing to norfloxacin accumulation in a manner dependent on multidrug efflux pumps and the PA1874-77 ABC transport system. Inactivation of the respective ABC transport system, furthermore, eliminated the recalcitrance of biofilms to killing by tobramycin but not norfloxacin, indicating that drug accumulation is not linked to biofilm drug tolerance. Our findings indicate for the first time that BrlR, a MerR-type transcriptional activator, activates genes encoding several ABC transport systems, in addition to multiple multidrug efflux pump genes. Moreover, our data confirm a BrlR target contributing to drug tolerance, likely countering the prevailing dogma that biofilm tolerance arises from a multiplicity of factors. Copyright © 2018 American Society for Microbiology.

  11. Phenotypes of Non-Attached Pseudomonas aeruginosa Aggregates Resemble Surface Attached Biofilm

    DEFF Research Database (Denmark)

    Alhede, Morten; Kragh, Kasper Nørskov; Qvortrup, Klaus

    2011-01-01

    For a chronic infection to be established, bacteria must be able to cope with hostile conditions such as low iron levels, oxidative stress, and clearance by the host defense, as well as antibiotic treatment. It is generally accepted that biofilm formation facilitates tolerance to these adverse......, RT-PCR as well as traditional culturing techniques to study the properties of Pseudomonas aeruginosa aggregates. We found that non-attached aggregates from stationary-phase cultures have comparable growth rates to surface attached biofilms. The growth rate estimations indicated that, independently...... were also found to be strikingly similar to flow-cell biofilms. Our data indicate that the tolerance of both biofilms and non-attached aggregates towards antibiotics is reversible by physical disruption. We provide evidence that the antibiotic tolerance is likely to be dependent on both...

  12. Study on Hydro-Alcoholic Extract Effect of Pomegranate Peel on Pseudomonas aeruginosa Biofilm Formation

    Directory of Open Access Journals (Sweden)

    R. Habibipour

    2015-10-01

    Full Text Available Introduction & Objective: Microorganisms form biomass as biofilm in response to many factors, in order to adapt to hostile extracellular environments and biocides. Using different herbal compounds are of those strategies to deal with biofilm. It has been proved that plants extracts such as pomegranate, raspberry and chamomile essential oils have anti-biofilm effects. This study aimed to evaluate the effect of different concentrations of black peel pomegranate ex-tract on Pseudomonas aeruginosa biofilm formation. Materials & Methods: In this experimental research the anti-biofilm effect, reducing the amount of biofilm formation and growth kinetics of Pseudomonas aeruginosa in different treatments was measured by microtiter and plate colorimetric crystal violet method. Biofilm formation was also examined using a microscope. Statistical analysis of data obtained from the reading of the ELISA was performed using SPSS software, P value 0.05. Results: Findings of this study showed that bacteria cannot form any biofilm in first 6 hours of incubation, in all treatments. The amount of biofilm formation after 12 hours in 0.01 and 0.05 g/ mL treatments were medium. Among treatments, after 18 and 24 hours of incubation 0.001 g/ mL concentration of pomegranate peel extract had medium and strong inhibitory effect on biofilm formation, respectively. Conclusion: Results of this study showed that biofilm formation and biofilm reduction percent-age is directly related to the duration of exposure of bacteria that could be due to the different phases of growth. Growth kinetics study also revealed that in the majority of treatments the growth was incremental up to about 15 hours and decrement afterwards due to the effective-ness of different treatments. After 18 hours, treatments have greatest influence on biofilm formation. The foregoing has been fully confirmed by the results of microscopic slides. (Sci J Hamadan Univ Med Sci 2015; 22 (3: 195-202

  13. Pseudomonas aeruginosa extracellular products inhibit staphylococcal growth, and disrupt established biofilms produced by Staphylococcus epidermidis

    DEFF Research Database (Denmark)

    Qin, Zhiqiang; Yang, Liang; Qu, Di

    2009-01-01

    Multiple bacterial species often coexist as communities, and compete for environmental resources. Here, we describe how an opportunistic pathogen, Pseudomonas aeruginosa, uses extracellular products to interact with the nosocomial pathogen Staphylococcus epidermidis. S. epidermidis biofilms and p...... of a novel strategy for controlling S. epidermidis biofilms....

  14. Quorum-sensing regulation of the biofilm matrix genes (pel) of Pseudomonas aeruginosa.

    Science.gov (United States)

    Sakuragi, Yumiko; Kolter, Roberto

    2007-07-01

    Quorum sensing (QS) has been previously shown to play an important role in the development of Pseudomonas aeruginosa biofilms (D. G. Davies et al., Science 280:295-298, 1998). Although QS regulation of swarming and DNA release has been shown to play important roles in biofilm development, regulation of genes directly involved in biosynthesis of biofilm matrix has not been described. Here, transcription of the pel operon, essential for the production of a glucose-rich matrix exopolysaccharide, is shown to be greatly reduced in lasI and rhlI mutants. Chemical complementation of the lasI mutant with 3-oxo-dodecanoyl homoserine lactone restores pel transcription to the wild-type level and biofilm formation ability. These findings thus connect QS signaling and transcription of genes responsible for biofilm matrix biosynthesis.

  15. Ginger Extract Inhibits Biofilm Formation by Pseudomonas aeruginosa PA14

    Science.gov (United States)

    Kim, Han-Shin; Park, Hee-Deung

    2013-01-01

    Bacterial biofilm formation can cause serious problems in clinical and industrial settings, which drives the development or screening of biofilm inhibitors. Some biofilm inhibitors have been screened from natural products or modified from natural compounds. Ginger has been used as a medicinal herb to treat infectious diseases for thousands of years, which leads to the hypothesis that it may contain chemicals inhibiting biofilm formation. To test this hypothesis, we evaluated ginger’s ability to inhibit Pseudomonas aeruginosa PA14 biofilm formation. A static biofilm assay demonstrated that biofilm development was reduced by 39–56% when ginger extract was added to the culture. In addition, various phenotypes were altered after ginger addition of PA14. Ginger extract decreased production of extracellular polymeric substances. This finding was confirmed by chemical analysis and confocal laser scanning microscopy. Furthermore, ginger extract formed noticeably less rugose colonies on agar plates containing Congo red and facilitated swarming motility on soft agar plates. The inhibition of biofilm formation and the altered phenotypes appear to be linked to a reduced level of a second messenger, bis-(3′-5′)-cyclic dimeric guanosine monophosphate. Importantly, ginger extract inhibited biofilm formation in both Gram-positive and Gram-negative bacteria. Also, surface biofilm cells formed with ginger extract detached more easily with surfactant than did those without ginger extract. Taken together, these findings provide a foundation for the possible discovery of a broad spectrum biofilm inhibitor. PMID:24086697

  16. Selective proteomic analysis of antibiotic-tolerant cellular subpopulations in pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Babin, Brett M.; Atangcho, Lydia; van Eldijk, Mark B.

    2017-01-01

    involved in central carbon metabolism. We differentiated the immediate proteomic response, characterized by an increase in flagellar motility, from the long-term adaptive strategy, which included the upregulation of purine synthesis. This targeted, selective analysis of a bacterial subpopulation...... amino acid tagging (BONCAT) method to enable selective proteomic analysis of a Pseudomonas aeruginosa biofilm subpopulation. Through controlled expression of a mutant methionyl-tRNA synthetase, we targeted BONCAT labeling to cells in the regions of biofilm microcolonies that showed increased tolerance...... demonstrates how the study of proteome dynamics can enhance our understanding of biofilm heterogeneity and antibiotic tolerance. IMPORTANCE Bacterial growth is frequently characterized by behavioral heterogeneity at the single-cell level. Heterogeneity is especially evident in the physiology of biofilms...

  17. Morphological bactericidal fast-acting effects of peracetic acid, a high-level disinfectant, against Staphylococcus aureus and Pseudomonas aeruginosa biofilms in tubing.

    Science.gov (United States)

    Chino, T; Nukui, Y; Morishita, Y; Moriya, K

    2017-01-01

    The bactericidal effect of disinfectants against biofilms is essential to reduce potential endoscopy-related infections caused by contamination. Here, we investigated the bactericidal effect of a high-level disinfectant, peracetic acid (PAA), against Staphylococcus aureus and Pseudomonas aeruginosa biofilm models in vitro. S. aureus and P. aeruginosa biofilms were cultured at 35 °C for 7 days with catheter tubes. The following high-level disinfectants (HLDs) were tested: 0.3% PAA, 0.55% ortho-phthalaldehyde (OPA), and 2.0% alkaline-buffered glutaraldehyde (GA). Biofilms were exposed to these agents for 1-60 min and observed after 5 min and 30 min by transmission and scanning electron microscopy. A Student's t test was performed to compare the exposure time required for bactericidal effectiveness of the disinfectants. PAA and GA were active within 1 min and 5 min, respectively, against S. aureus and P. aeruginosa biofilms. OPA took longer than 10 min and 30 min to act against S. aureus and P. aeruginosa biofilms, respectively ( p  < 0.01). Treatment with PAA elicited changes in cell shape after 5 min and structural damage after 30 min. Amongst the HLDs investigated, PAA elicited the most rapid bactericidal effects against both biofilms. Additionally, treatment with PAA induced morphological alterations in the in vitro biofilm models, suggesting that PAA exerts fast-acting bactericidal effects against biofilms associated with endoscopy-related infections. These findings indicate that the exposure time for bactericidal effectiveness of HLDs for endoscope reprocessing in healthcare settings should be reconsidered.

  18. Corneal Biofilms: From Planktonic to Microcolony Formation in an Experimental Keratitis Infection with Pseudomonas Aeruginosa.

    Science.gov (United States)

    Saraswathi, Padmanabhan; Beuerman, Roger W

    2015-10-01

    Microbial biofilms commonly comprise part of the infectious scenario, complicating the therapeutic approach. The purpose of this study was to determine in a mouse model of corneal infection if mature biofilms formed and to visualize the stages of biofilm formation. A bacterial keratitis model was established using Pseudomonas aeruginosa ATCC 9027 (1 × 10(8) CFU/ml) to infect the cornea of C57BL/6 black mouse. Eyes were examined post-infection (PI) on days 1, 2, 3, 5, and 7, and imaged by slit lamp microscopy, and light, confocal, and electron microscopy to identify the stages of biofilm formation and the time of appearance. On PI day 1, Gram staining showed rod-shaped bacteria adherent on the corneal surface. On PI days 2 and 3, bacteria were seen within webs of extracellular polymeric substance (EPS) and glycocalyx secretion, imaged by confocal microscopy. Scanning electron microscopy demonstrated microcolonies of active infectious cells bound with thick fibrous material. Transmission electron microscopy substantiated the formation of classical biofilm architecture with P. aeruginosa densely packed within the extracellular polymeric substances on PI days 5 and 7. Direct visual evidence showed that biofilms routinely developed on the biotic surface of the mouse cornea. The mouse model can be used to develop new approaches to deal therapeutically with biofilms in corneal infections. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  19. Pseudomonas aeruginosa biofilm aggravates skin inflammatory response in BALB/c mice in a novel chronic wound model

    DEFF Research Database (Denmark)

    Trøstrup, Hannah; Thomsen, Kim; Christophersen, Lars J

    2013-01-01

    model in C3H/HeN and BALB/c mice. The chronic wound was established by an injection of seaweed alginate-embedded P. aeruginosa PAO1 beneath a third-degree thermal lesion providing full thickness skin necrosis, as in human chronic wounds. Cultures revealed growth of PA, and both alginate with or without......Chronic wounds are presumed to persist in the inflammatory state, preventing healing. Emerging evidence indicates a clinical impact of bacterial biofilms in soft tissues, including Pseudomonas aeruginosa (PA) biofilms. To further investigate this, we developed a chronic PA biofilm wound infection...... bacteria organized in clusters, resembling biofilms, and inflammation located adjacent to the PA. The chronic wound infection showed a higher number of PAO1 in the BALB/c mice at day 4 after infection as compared to C3H/HeN mice (p

  20. Human Tear Fluid Reduces Culturability of Contact Lens Associated Pseudomonas aeruginosa Biofilms but Induces Expression of the Virulence Associated Type III Secretion System

    Science.gov (United States)

    Wu, Yvonne T.; Tam, Connie; Zhu, Lucia S.; Evans, David J.; Fleiszig, Suzanne M. J.

    2017-01-01

    Purpose The type III secretion system (T3SS) is a significant virulence determinant for Pseudomonas aeruginosa. Using a rodent model, we found that contact lens (CL)-related corneal infections were associated with lens surface biofilms. Here, we studied the impact of human tear fluid on CL-associated biofilm growth and T3SS expression. Methods P. aeruginosa biofilms were formed on contact lenses for up to 7 days with or without human tear fluid, then exposed to tear fluid for 5 or 24 h. Biofilms were imaged using confocal microscopy. Bacterial culturability was quantified by viable counts, and T3SS gene expression measured by RT-qPCR. Controls included trypticase soy broth, PBS and planktonic bacteria. Results With or without tear fluid, biofilms grew to ~108 cfu viable bacteria by 24 h. Exposing biofilms to tear fluid after they had formed without it on lenses reduced bacterial culturability ~180-fold (pbacteria [5.46 ± 0.24-fold for T3SS transcriptional activitor exsA (p=.02), and 3.76 ± 0.36-fold for T3SS effector toxin exoS (p=.01)]. Tear fluid further enhanced exsA and exoS expression in CL-grown biofilms, but not planktonic bacteria, by 2.09 ± 0.38-fold (p = 0.04) and 1.89 ± 0.26-fold (p<.001), respectively. Conclusions Considering the pivitol role of the T3SS in P. aeruginosa infections, its induction in CL-grown P. aeruginosa biofilms by tear fluid might contribute to the pathogenesis of CL-related P. aeruginosa keratitis. PMID:27670247

  1. Pseudomonas aeruginosa uses T3SS to inhibit diabetic wound healing.

    Science.gov (United States)

    Goldufsky, Josef; Wood, Stephen J; Jayaraman, Vijayakumar; Majdobeh, Omar; Chen, Lin; Qin, Shanshan; Zhang, Chunxiang; DiPietro, Luisa A; Shafikhani, Sasha H

    2015-01-01

    Diabetic foot ulcers are responsible for more hospitalizations than any other complication of diabetes. Bacterial infection is recognized as an important factor associated with impaired healing in diabetic ulcers. Pseudomonas aeruginosa is the most frequently detected Gram-negative pathogen in diabetic ulcers. P. aeruginosa infection has been shown to impair healing in diabetic wounds in a manner that correlates with its ability to form biofilm. While the majority of infections in diabetic ulcers are biofilm associated, 33% of infections are nonbiofilm in nature. P. aeruginosa is the most prevalent Gram-negative pathogen in all diabetic wound types, which suggests that the deleterious impact of P. aeruginosa on healing in diabetic wounds goes beyond its ability to form biofilm and likely involves other factors. The Type III Secretion System (T3SS) virulence structure is required for the pathogenesis of all P. aeruginosa clinical isolates, suggesting that it may also play a role in the inhibition of wound repair in diabetic skin ulcers. We evaluated the role of T3SS in mediating P. aeruginosa-induced tissue damage in the wounds of diabetic mice. Our data demonstrate that P. aeruginosa establishes a robust and persistent infection in diabetic wounds independent of its ability to form biofilm and causes severe wound damage in a manner that primarily depends on its T3SS. © 2015 by the Wound Healing Society.

  2. Psl Produced by Mucoid Pseudomonas aeruginosa Contributes to the Establishment of Biofilms and Immune Evasion.

    Science.gov (United States)

    Jones, Christopher J; Wozniak, Daniel J

    2017-06-20

    Despite years of research and clinical advances, chronic pulmonary infections with mucoid Pseudomonas aeruginosa remain the primary concern for cystic fibrosis patients. Much of the research on these strains has focused on the contributions of the polysaccharide alginate; however, it is becoming evident that the neutral polysaccharide Psl also contributes to biofilm formation and the maintenance of chronic infections. Here, we demonstrate that Psl produced by mucoid strains has significant roles in biofilm structure and evasion of immune effectors. Though mucoid strains produce less Psl than nonmucoid strains, the Psl that is produced is functional, since it mediates adhesion to human airway cells and epithelial cell death. Additionally, Psl protects mucoid bacteria from opsonization and killing by complement components in human serum. Psl production by mucoid strains stimulates a proinflammatory response in the murine lung, leading to reduced colonization. To determine the relevance of these data to clinical infections, we tested Psl production and biofilm formation of a panel of mucoid clinical isolates. We demonstrated three classes of mucoid isolates, those that produce Psl and form robust biofilms, those that did not produce Psl and have a poor biofilm phenotype, and exopolysaccharide (EPS) redundant strains. Collectively, these experimental results demonstrate that Psl contributes to the biofilm formation and immune evasion of many mucoid strains. This is a novel role for Psl in the establishment and maintenance of chronic pulmonary infections by mucoid strains. IMPORTANCE Cystic fibrosis patients are engaged in an ongoing battle against chronic lung infections by the bacterium Pseudomonas aeruginosa One key factor contributing to the maintenance of chronic infections is the conversion to a mucoid phenotype, where the bacteria produce copious amounts of the polysaccharide alginate. Once the bacteria become mucoid, existing treatments are poorly effective. We

  3. Antibacterial, anti-swarming and anti-biofilm formation activities of Chamaemelum nobile against Pseudomonas aeruginosa.

    Science.gov (United States)

    Kazemian, Hossein; Ghafourian, Sobhan; Heidari, Hamid; Amiri, Pouya; Yamchi, Jalil Kardan; Shavalipour, Aref; Houri, Hamidreza; Maleki, Abbas; Sadeghifard, Nourkhoda

    2015-01-01

    Chamomile (Chamaemelum nobile) is widely used throughout the world, and has anti-inflammatory, deodorant, bacteriostatic, antimicrobial, carminative, sedative, antiseptic, anti-catarrhal, and spasmolytic properties. Because of the increasing incidence of drug-resistant bacteria, the development of natural antibacterial sources such as medical herbs for the treatment of infectious diseases is necessary. Extracts from different plant parts such as the leaves, flowers, fruit, and bark of Combretum albiflorum, Laurus nobilis , and Sonchus oleraceus were found to possess anti-quorum sensing (QS) activities. In this study, we evaluated the effect of C. nobile against Pseudomonas aeruginosa biofilm formation. The P. aeruginosa samples were isolated from patients with different types of infection, including wound infection, septicemia, and urinary tract infection. The flowers of C. nobile were dried and the extract was removed using a rotary device and then dissolved in dimethyl sulfoxide at pH 7.4. The microdilution method was used to evaluate the minimum inhibitory concentration (MIC) of this extract on P. aeruginosa , and biofilm inhibition was assayed. Eighty percent of the isolated samples (16/20) could form a biofilm, and most of these were isolated from wound infections. The biofilm inhibitory concentration of the C. nobile extract was 6.25-25mg/ml, whereas the MIC was 12.5-50mg/ml. The anti-QS property of C. nobile may play an important role in its antibacterial activity, thus offering an additional strategy in the fight against bacterial infections. However, molecular investigation is required to explore the exact mechanisms of the antibacterial action and functions of this phytocompound.

  4. Positively charged biopolymeric nanoparticles for the inhibition of Pseudomonas aeruginosa biofilms

    International Nuclear Information System (INIS)

    Chronopoulou, Laura; Di Domenico, Enea Gino; Ascenzioni, Fiorentina; Palocci, Cleofe

    2016-01-01

    Currently, many microbial infections have the potential to become lethal owing to the development of antimicrobial resistance by means of different mechanisms and mainly on the basis of the fact that many drugs are unable to reach therapeutic levels in the target sites. This requires the use of high doses and frequent administrations, causing adverse side effects or in some cases toxicity. The use of nanoparticle systems could help overcome such problems and increase drug efficacy. In the present study, we developed a new drug delivery system based on the use of biopolymeric nanovectors loaded with tobramycin (Tb), which is the standard antibiotic for the treatment of Cystic Fibrosis-associated P. aeruginosa lung infections. Tb-loaded biopolymeric nanoparticles composed by dextran sulfate (DS) and chitosan (CS) were prepared by ionotropic gelation. We optimized drug entrapment in DS/CS nanoparticles, obtaining particles of 170 nm and with a drug loading of 400 µg Tb/mg of nanoparticles. In accord with in vitro release experiments, such preparations were able to release approximately 25 % of their cargo in 60 h. In vitro, the antimicrobial efficacy of the drug delivery system on P. aeruginosa biofilm was tested and compared to the effects of free drug revealing that this formulation can reduce the viability of P. aeruginosa biofilms for 48 h with a single-dose administration.

  5. Positively charged biopolymeric nanoparticles for the inhibition of Pseudomonas aeruginosa biofilms

    Energy Technology Data Exchange (ETDEWEB)

    Chronopoulou, Laura [University of Rome La Sapienza, Department of Chemistry (Italy); Di Domenico, Enea Gino [IRCCS, Department of Clinical Pathology and Microbiology, San Gallicano Institute (Italy); Ascenzioni, Fiorentina [University of Rome La Sapienza, Department of Biology and Biotechnology C. Darwin (Italy); Palocci, Cleofe, E-mail: cleofe.palocci@uniroma1.it [University of Rome La Sapienza, Department of Chemistry (Italy)

    2016-10-15

    Currently, many microbial infections have the potential to become lethal owing to the development of antimicrobial resistance by means of different mechanisms and mainly on the basis of the fact that many drugs are unable to reach therapeutic levels in the target sites. This requires the use of high doses and frequent administrations, causing adverse side effects or in some cases toxicity. The use of nanoparticle systems could help overcome such problems and increase drug efficacy. In the present study, we developed a new drug delivery system based on the use of biopolymeric nanovectors loaded with tobramycin (Tb), which is the standard antibiotic for the treatment of Cystic Fibrosis-associated P. aeruginosa lung infections. Tb-loaded biopolymeric nanoparticles composed by dextran sulfate (DS) and chitosan (CS) were prepared by ionotropic gelation. We optimized drug entrapment in DS/CS nanoparticles, obtaining particles of 170 nm and with a drug loading of 400 µg Tb/mg of nanoparticles. In accord with in vitro release experiments, such preparations were able to release approximately 25 % of their cargo in 60 h. In vitro, the antimicrobial efficacy of the drug delivery system on P. aeruginosa biofilm was tested and compared to the effects of free drug revealing that this formulation can reduce the viability of P. aeruginosa biofilms for 48 h with a single-dose administration.

  6. Positively charged biopolymeric nanoparticles for the inhibition of Pseudomonas aeruginosa biofilms

    Science.gov (United States)

    Chronopoulou, Laura; Di Domenico, Enea Gino; Ascenzioni, Fiorentina; Palocci, Cleofe

    2016-10-01

    Currently, many microbial infections have the potential to become lethal owing to the development of antimicrobial resistance by means of different mechanisms and mainly on the basis of the fact that many drugs are unable to reach therapeutic levels in the target sites. This requires the use of high doses and frequent administrations, causing adverse side effects or in some cases toxicity. The use of nanoparticle systems could help overcome such problems and increase drug efficacy. In the present study, we developed a new drug delivery system based on the use of biopolymeric nanovectors loaded with tobramycin (Tb), which is the standard antibiotic for the treatment of Cystic Fibrosis-associated P. aeruginosa lung infections. Tb-loaded biopolymeric nanoparticles composed by dextran sulfate (DS) and chitosan (CS) were prepared by ionotropic gelation. We optimized drug entrapment in DS/CS nanoparticles, obtaining particles of 170 nm and with a drug loading of 400 µg Tb/mg of nanoparticles. In accord with in vitro release experiments, such preparations were able to release approximately 25 % of their cargo in 60 h. In vitro, the antimicrobial efficacy of the drug delivery system on P. aeruginosa biofilm was tested and compared to the effects of free drug revealing that this formulation can reduce the viability of P. aeruginosa biofilms for 48 h with a single-dose administration.

  7. Morphological bactericidal fast-acting effects of peracetic acid, a high-level disinfectant, against Staphylococcus aureus and Pseudomonas aeruginosa biofilms in tubing

    Directory of Open Access Journals (Sweden)

    T. Chino

    2017-12-01

    Full Text Available Abstract Background The bactericidal effect of disinfectants against biofilms is essential to reduce potential endoscopy-related infections caused by contamination. Here, we investigated the bactericidal effect of a high-level disinfectant, peracetic acid (PAA, against Staphylococcus aureus and Pseudomonas aeruginosa biofilm models in vitro. Methods S. aureus and P. aeruginosa biofilms were cultured at 35 °C for 7 days with catheter tubes. The following high-level disinfectants (HLDs were tested: 0.3% PAA, 0.55% ortho-phthalaldehyde (OPA, and 2.0% alkaline-buffered glutaraldehyde (GA. Biofilms were exposed to these agents for 1–60 min and observed after 5 min and 30 min by transmission and scanning electron microscopy. A Student’s t test was performed to compare the exposure time required for bactericidal effectiveness of the disinfectants. Results PAA and GA were active within 1 min and 5 min, respectively, against S. aureus and P. aeruginosa biofilms. OPA took longer than 10 min and 30 min to act against S. aureus and P. aeruginosa biofilms, respectively (p < 0.01. Treatment with PAA elicited changes in cell shape after 5 min and structural damage after 30 min. Conclusions Amongst the HLDs investigated, PAA elicited the most rapid bactericidal effects against both biofilms. Additionally, treatment with PAA induced morphological alterations in the in vitro biofilm models, suggesting that PAA exerts fast-acting bactericidal effects against biofilms associated with endoscopy-related infections. These findings indicate that the exposure time for bactericidal effectiveness of HLDs for endoscope reprocessing in healthcare settings should be reconsidered.

  8. Evolution of antibiotic resistance in biofilm and planktonic P. aeruginosa populations exposed to sub-inhibitory levels of ciprofloxacin

    DEFF Research Database (Denmark)

    Ahmed, Marwa N.; Porse, Andreas; Sommer, Morten Otto Alexander

    2018-01-01

    in planktonic cultures and are less studied in biofilms. We experimentally evolved P. aeruginosa PAO1 colony-biofilms and stationary-phase planktonic cultures for seven passages in the presence of sub-inhibitory levels (0.1 mg/L) of ciprofloxacin (CIP) and performed a genotypic (whole bacterial population......The opportunistic Gram-negative pathogen Pseudomonas aeruginosa, known for its intrinsic and acquired antibiotic resistance, has a notorious ability to form biofilms, which often facilitate chronic infections. The evolutionary paths to antibiotic resistance have mainly been investigated......-dependent adaptations. A general trend towards a reduction in type IV-pili dependent motility (twitching) in CIP-evolved populations, and towards loss of virulence associated traits in the populations evolved in the absence of antibiotic, was observed. In conclusion, our data indicate that biofilms facilitate...

  9. Pseudomonas aeruginosa biofilm hampers murine central wound healing by suppression of vascular epithelial growth factor

    DEFF Research Database (Denmark)

    Trøstrup, Hannah; Lerche, Christian J; Christophersen, Lars J

    2018-01-01

    -wounded, infection-susceptible BALB/c mice, which display delayed wound closure compared to C3H/HeN mice. Wounds were evaluated histopathologically 4, 7 or 10 days post-infection. Photoplanimetry evaluated necrotic areas. P. aeruginosa biofilm suppressed vascular endothelial growth factor levels centrally in BALB....../c wounds but increased peripheral levels 4-7 days post-infection. Central zones of the burn wound displayed lower levels of central vascular endothelial growth factor as observed 4 and 7 days post-infection in BALB/c mice compared to their C3H/HeN counterparts. Biofilm suppressed early, centrally located S......100A8/A9 in BALB/c and centrally and peripherally later on in C3H/HeN wounds as compared to uninfected mice. Peripheral polymorphonuclear-dominated inflammation and larger necrosis were observed in BALB/c wounds. In conclusion, P. aeruginosa biofilm modulates wounds by suppressing central...

  10. Regulatory and Metabolic Networks for the Adaptation of Pseudomonas aeruginosa Biofilms to Urinary Tract-Like Conditions

    Science.gov (United States)

    Dohnt, Katrin; Haddad, Isam; Jänsch, Lothar; Klein, Johannes; Narten, Maike; Pommerenke, Claudia; Scheer, Maurice; Schobert, Max; Schomburg, Dietmar; Thielen, Bernhard; Jahn, Dieter

    2013-01-01

    Biofilms of the Gram-negative bacterium Pseudomonas aeruginosa are one of the major causes of complicated urinary tract infections with detrimental outcome. To develop novel therapeutic strategies the molecular adaption strategies of P. aeruginosa biofilms to the conditions of the urinary tract were investigated thoroughly at the systems level using transcriptome, proteome, metabolome and enzyme activity analyses. For this purpose biofilms were grown anaerobically in artificial urine medium (AUM). Obtained data were integrated bioinformatically into gene regulatory and metabolic networks. The dominating response at the transcriptome and proteome level was the adaptation to iron limitation via the broad Fur regulon including 19 sigma factors and up to 80 regulated target genes or operons. In agreement, reduction of the iron cofactor-dependent nitrate respiratory metabolism was detected. An adaptation of the central metabolism to lactate, citrate and amino acid as carbon sources with the induction of the glyoxylate bypass was observed, while other components of AUM like urea and creatinine were not used. Amino acid utilization pathways were found induced, while fatty acid biosynthesis was reduced. The high amounts of phosphate found in AUM explain the reduction of phosphate assimilation systems. Increased quorum sensing activity with the parallel reduction of chemotaxis and flagellum assembly underscored the importance of the biofilm life style. However, reduced formation of the extracellular polysaccharide alginate, typical for P. aeruginosa biofilms in lungs, indicated a different biofilm type for urinary tract infections. Furthermore, the obtained quorum sensing response results in an increased production of virulence factors like the extracellular lipase LipA and protease LasB and AprA explaining the harmful cause of these infections. PMID:23967252

  11. Regulatory and metabolic networks for the adaptation of Pseudomonas aeruginosa biofilms to urinary tract-like conditions.

    Directory of Open Access Journals (Sweden)

    Petra Tielen

    Full Text Available Biofilms of the Gram-negative bacterium Pseudomonas aeruginosa are one of the major causes of complicated urinary tract infections with detrimental outcome. To develop novel therapeutic strategies the molecular adaption strategies of P. aeruginosa biofilms to the conditions of the urinary tract were investigated thoroughly at the systems level using transcriptome, proteome, metabolome and enzyme activity analyses. For this purpose biofilms were grown anaerobically in artificial urine medium (AUM. Obtained data were integrated bioinformatically into gene regulatory and metabolic networks. The dominating response at the transcriptome and proteome level was the adaptation to iron limitation via the broad Fur regulon including 19 sigma factors and up to 80 regulated target genes or operons. In agreement, reduction of the iron cofactor-dependent nitrate respiratory metabolism was detected. An adaptation of the central metabolism to lactate, citrate and amino acid as carbon sources with the induction of the glyoxylate bypass was observed, while other components of AUM like urea and creatinine were not used. Amino acid utilization pathways were found induced, while fatty acid biosynthesis was reduced. The high amounts of phosphate found in AUM explain the reduction of phosphate assimilation systems. Increased quorum sensing activity with the parallel reduction of chemotaxis and flagellum assembly underscored the importance of the biofilm life style. However, reduced formation of the extracellular polysaccharide alginate, typical for P. aeruginosa biofilms in lungs, indicated a different biofilm type for urinary tract infections. Furthermore, the obtained quorum sensing response results in an increased production of virulence factors like the extracellular lipase LipA and protease LasB and AprA explaining the harmful cause of these infections.

  12. Regulatory and metabolic networks for the adaptation of Pseudomonas aeruginosa biofilms to urinary tract-like conditions.

    Science.gov (United States)

    Tielen, Petra; Rosin, Nathalie; Meyer, Ann-Kathrin; Dohnt, Katrin; Haddad, Isam; Jänsch, Lothar; Klein, Johannes; Narten, Maike; Pommerenke, Claudia; Scheer, Maurice; Schobert, Max; Schomburg, Dietmar; Thielen, Bernhard; Jahn, Dieter

    2013-01-01

    Biofilms of the Gram-negative bacterium Pseudomonas aeruginosa are one of the major causes of complicated urinary tract infections with detrimental outcome. To develop novel therapeutic strategies the molecular adaption strategies of P. aeruginosa biofilms to the conditions of the urinary tract were investigated thoroughly at the systems level using transcriptome, proteome, metabolome and enzyme activity analyses. For this purpose biofilms were grown anaerobically in artificial urine medium (AUM). Obtained data were integrated bioinformatically into gene regulatory and metabolic networks. The dominating response at the transcriptome and proteome level was the adaptation to iron limitation via the broad Fur regulon including 19 sigma factors and up to 80 regulated target genes or operons. In agreement, reduction of the iron cofactor-dependent nitrate respiratory metabolism was detected. An adaptation of the central metabolism to lactate, citrate and amino acid as carbon sources with the induction of the glyoxylate bypass was observed, while other components of AUM like urea and creatinine were not used. Amino acid utilization pathways were found induced, while fatty acid biosynthesis was reduced. The high amounts of phosphate found in AUM explain the reduction of phosphate assimilation systems. Increased quorum sensing activity with the parallel reduction of chemotaxis and flagellum assembly underscored the importance of the biofilm life style. However, reduced formation of the extracellular polysaccharide alginate, typical for P. aeruginosa biofilms in lungs, indicated a different biofilm type for urinary tract infections. Furthermore, the obtained quorum sensing response results in an increased production of virulence factors like the extracellular lipase LipA and protease LasB and AprA explaining the harmful cause of these infections.

  13. Passive control of quorum sensing: prevention of Pseudomonas aeruginosa biofilm formation by imprinted polymers.

    Science.gov (United States)

    Piletska, Elena V; Stavroulakis, Georgios; Larcombe, Lee D; Whitcombe, Michael J; Sharma, Anant; Primrose, Sandy; Robinson, Gary K; Piletsky, Sergey A

    2011-04-11

    Here we present the first molecular imprinted polymer (MIP) that is able to attenuate the biofilm formation of the opportunistic human pathogen Pseudomonas aeruginosa through specific sequestration of its signal molecule N-(3-oxododecanoyl)-L-homoserine lactone (3-oxo-C(12)-AHL). The MIP was rationally designed using computational modeling, and its capacity and specificity and that of a corresponding blank polymer toward signal molecule of P. aeruginosa (3-oxo-C(12)-AHL) and its analogue were tested. The biofilm formation in the presence of polymers and without polymers was studied using scanning confocal laser microscopy. Staining with crystal violet dye was used for the quantification of the biofilm formation. A significant reduction of the biofilm growth was observed in the presence of MIP (>80%), which was superior to that of the resin prepared without template, which showed a reduction of 40% in comparison with biofilm, which was grown without polymer addition. It was shown that 3-oxo-C(12)-AHL-specific MIP prevented the development of quorum-sensing-controlled phenotypes (in this case, biofilm formation) from being up-regulated. The developed MIP could be considered as a new tool for the elimination of life-threatening infections in a multitude of practical applications; it could, for example, be grafted on the surface of medical devices such as catheters and lenses, be a component of paints, or be used as a wound adsorbent.

  14. Effects of Photoactivated Titanium Dioxide Nanopowders and Coating on Planktonic and Biofilm Growth of Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Polo, Andrea; Diamanti, Maria Vittoria; Bjarnsholt, Thomas

    2011-01-01

    eradication of P. aeruginosa planktonic cells (initial concentration 10(8) cells/ml) in 24 h compared to a 3-log reduction caused by UV-A light alone. In contrast, neither the photocatalytic treatment with TiO(2) film nor that with TiO(2) nanopowder had any effect on P. aeruginosa biofilms at all...

  15. Combination of hypothiocyanite and lactoferrin (ALX-109) enhances the ability of tobramycin and aztreonam to eliminate Pseudomonas aeruginosa biofilms growing on cystic fibrosis airway epithelial cells.

    Science.gov (United States)

    Moreau-Marquis, Sophie; Coutermarsh, Bonita; Stanton, Bruce A

    2015-01-01

    Chelating iron may be a promising new therapy to eliminate Pseudomonas aeruginosa biofilms in the lungs of cystic fibrosis (CF) patients. Here, we investigate whether ALX-109 [a defined combination of an investigational drug containing lactoferrin (an iron-binding glycoprotein) and hypothiocyanite (a bactericidal agent)], alone and in combination with tobramycin or aztreonam, reduces P. aeruginosa biofilms grown on human CF airway epithelial cells. P. aeruginosa (PAO1 and six clinical isolates of Pseudomonas) biofilms grown at the apical surface of confluent monolayers of CF airway epithelial cells were treated with ALX-109, either alone or in combination with tobramycin or aztreonam. Bacterial cfu remaining after treatment were determined by plate counting. ALX-109 alone reduced PAO1 biofilm formation, but had no effect on established biofilms. ALX-109 enhanced the ability of tobramycin and aztreonam to inhibit PAO1 biofilm formation and to reduce established PAO1 biofilms. ALX-109 and tobramycin were additive in disrupting established biofilms formed by six clinical isolates of P. aeruginosa obtained from the sputum of CF patients. Mucoid P. aeruginosa isolates were most susceptible to the combination of ALX-109 and tobramycin. In addition, ALX-109 also enhanced the ability of aztreonam to reduce established PAO1 biofilms. Inhalation therapy combining hypothiocyanite and lactoferrin with TOBI(®) (tobramycin) or Cayston(®) (aztreonam) may be beneficial to CF patients by decreasing the airway bacterial burden of P. aeruginosa. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  16. Bactericidal Effect of Calcium Oxide (Scallop-Shell Powder) Against Pseudomonas aeruginosa Biofilm on Quail Egg Shell, Stainless Steel, Plastic, and Rubber.

    Science.gov (United States)

    Jung, Soo-Jin; Park, Shin Young; Kim, Seh Eun; Kang, Ike; Park, Jiyong; Lee, Jungwon; Kim, Chang-Min; Chung, Myung-Sub; Ha, Sang-Do

    2017-07-01

    The aim of this study was to evaluate the bactericidal effect of calcium oxide (CaO) against Pseudomonas aeruginosa biofilms on quail eggshells and major egg contacting surfaces (stainless steel, plastic, and rubber). The samples were subjected to CaO treatments (0%, 0.01%, 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, and 0.30%) for 1 min. All the CaO treatments significantly reduced P. aeruginosa biofilms on all tested surfaces as compared to controls. In comparison of biofilm stability, the strongest and most resistant biofilm was formed on eggshell against the CaO treatment, followed by rubber, stainless steel, and plastic. In evaluation of bactericidal effect, the largest reduction (3.16 log CFU) was observed in plastic even at the lowest concentration of CaO (0.01%), whereas the least reduction was found in eggshells, regardless of CaO concentration. In addition, stainless steel showed a significant reduction in biofilm formation at all concentrations except 0.10% to 0.15% CaO. At 0.30% CaO, the reduction of P. aeruginosa in biofilms on stainless steel, plastic, rubber, and eggshell were 5.48, 6.37, 4.87, and 3.14 log CFU/cm 2 (CFU/egg), respectively. Biofilm reduction after CaO treatment was also observed by field emission scanning electron microscopy (FE-SEM). Based on the FE-SEM images, we observed that P. aeruginosa biofilms formed compact aggregations on eggshell surfaces with CaO treatments up to 0.30%. More specifically, a 0.20% CaO treatment resulted in the reductions of 3 to 6 log CFU in all materials. © 2017 Institute of Food Technologists®.

  17. Anti-biofilm activity of biogenic selenium nanoparticles and selenium dioxide against clinical isolates of Staphylococcus aureus, Pseudomonas aeruginosa, and Proteus mirabilis.

    Science.gov (United States)

    Shakibaie, Mojtaba; Forootanfar, Hamid; Golkari, Yaser; Mohammadi-Khorsand, Tayebe; Shakibaie, Mohammad Reza

    2015-01-01

    The aim of the present study was to investigate the anti-biofilm activity of biologically synthesized selenium nanoparticles (Se NPs) against the biofilm produced by clinically isolated bacterial strains compared to that of selenium dioxide. Thirty strains of Staphylococcus aureus, Pseudomonas aeruginosa, and Proteus mirabilis were isolated from various specimens of the patients hospitalized in different hospitals (Kerman, Iran). Quantification of the biofilm using microtiter plate assay method introduced 30% of S. aureus, 13% of P. aeruginosa and 17% of P. mirabilis isolates as severely adherent strains. Transmission electron micrograph (TEM) of the purified Se NPs (produced by Bacillus sp. MSh-1) showed individual and spherical nano-structure in the size range of 80-220nm. Obtained results of the biofilm formation revealed that selenium nanoparticles inhibited the biofilm of S. aureus, P. aeruginosa, and P. mirabilis by 42%, 34.3%, and 53.4%, respectively, compared to that of the non-treated samples. Effect of temperature and pH on the biofilm formation in the presence of Se NPs and SeO2 was also evaluated. Copyright © 2014 Elsevier GmbH. All rights reserved.

  18. Antibacterial, anti-swarming and anti-biofilm formation activities of Chamaemelum nobile against Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Hossein Kazemian

    2015-08-01

    Full Text Available AbstractINTRODUCTION:Chamomile ( Chamaemelum nobile is widely used throughout the world, and has anti-inflammatory, deodorant, bacteriostatic, antimicrobial, carminative, sedative, antiseptic, anti-catarrhal, and spasmolytic properties. Because of the increasing incidence of drug-resistant bacteria, the development of natural antibacterial sources such as medical herbs for the treatment of infectious diseases is necessary. Extracts from different plant parts such as the leaves, flowers, fruit, and bark of Combretum albiflorum, Laurus nobilis , and Sonchus oleraceus were found to possess anti-quorum sensing (QS activities. In this study, we evaluated the effect of C. nobile against Pseudomonas aeruginosa biofilm formationMETHODS:The P. aeruginosa samples were isolated from patients with different types of infection, including wound infection, septicemia, and urinary tract infection. The flowers of C. nobile were dried and the extract was removed using a rotary device and then dissolved in dimethyl sulfoxide at pH 7.4. The microdilution method was used to evaluate the minimum inhibitory concentration (MIC of this extract on P. aeruginosa , and biofilm inhibition was assayed.RESULTS:Eighty percent of the isolated samples (16/20 could form a biofilm, and most of these were isolated from wound infections. The biofilm inhibitory concentration of the C. nobile extract was 6.25-25mg/ml, whereas the MIC was 12.5-50mg/ml.CONCLUSIONS:The anti-QS property of C. nobile may play an important role in its antibacterial activity, thus offering an additional strategy in the fight against bacterial infections. However, molecular investigation is required to explore the exact mechanisms of the antibacterial action and functions of this phytocompound.

  19. THE FORMATION OF BIOFILMS BY PSEUDOMONAS AERUGINOSA STRAINS, AND METHODS OF ITS CONTROL (REVIEW

    Directory of Open Access Journals (Sweden)

    Sarkis-Ivanova VV

    2017-03-01

    Full Text Available The majority of natural bacterial populations exist in the form of biofilms, highly-structured multicellular communities incorporated in extracellular polymer matrix of own production. For a considerable part of clinically significant species this form of existence provides optimal conditions for reaching of pathogenic and colonizational potential, and also encourages saving of metabolically inactive part of population which is characterized by a low level of sensitivity to antibiotics impact. The latter plays a significant role in formation of chronic persistent infections resistant to antibiotics treatment. Members of microbial group are united basing on the principle which excludes antagonism, determines their nutritional, energetic and another connections between them and environment. Such a connection of microorganisms community behavior received a special definition of “quorum sensing”. Quorum sensing is an ability of some bacteria (probably, also another microorganisms to communicate and coordinate their behavior through secretion of substances which are the signals for coordination of certain behavior or interaction between bacteria of the same type or subtype depending on their growth tightness. When the concentration of excreted signaling agents reaches a borderline value the group of bacteria starts working as a single organism. Furthermore, signaling agents for Gram-negative and Gram-positive microorganisms differ. For instance, for Pseudomonas aeruginosa microorganism a signaling molecule of quorum sensing is acyl homoserine lactone, and biofilm created by this pathogen produces pyocyanin which functions as an electronic transporter in current generation. A large-scale investigation of P.aeruginosa biofilm morphology, mechanisms of its production and degradation, peculiarities of formation under different conditions and impact of various factors are at the infancy stage. According to undivided opinion of scientists who

  20. Nanoscale investigation on Pseudomonas aeruginosa biofilm formed on porous silicon using atomic force microscopy.

    Science.gov (United States)

    Kannan, Ashwin; Karumanchi, Subbalakshmi Latha; Krishna, Vinatha; Thiruvengadam, Kothai; Ramalingam, Subramaniam; Gautam, Pennathur

    2014-01-01

    Colonization of surfaces by bacterial cells results in the formation of biofilms. There is a need to study the factors that are important for formation of biofilms since biofilms have been implicated in the failure of semiconductor devices and implants. In the present study, the adhesion force of biofilms (formed by Pseudomonas aeruginosa) on porous silicon substrates of varying surface roughness was quantified using atomic force microscopy (AFM). The experiments were carried out to quantify the effect of surface roughness on the adhesion force of biofilm. The results show that the adhesion force increased from 1.5 ± 0.5 to 13.2 ± 0.9 nN with increase in the surface roughness of silicon substrate. The results suggest that the adhesion force of biofilm is affected by surface roughness of substrate. © 2014 Wiley Periodicals, Inc.

  1. Silver nanoparticles impede the biofilm formation by Pseudomonas aeruginosa and Staphylococcus epidermidis.

    Science.gov (United States)

    Kalishwaralal, Kalimuthu; BarathManiKanth, Selvaraj; Pandian, Sureshbabu Ram Kumar; Deepak, Venkataraman; Gurunathan, Sangiliyandi

    2010-09-01

    Biofilms are ensued due to bacteria that attach to surfaces and aggregate in a hydrated polymeric matrix. Formation of these sessile communities and their inherent resistance to anti-microbial agents are the source of many relentless and chronic bacterial infections. Such biofilms are responsible play a major role in development of ocular related infectious diseases in human namely microbial keratitis. Different approaches have been used for preventing biofilm related infections in health care settings. Many of these methods have their own demerits that include chemical based complications; emergent antibiotic resistant strains, etc. silver nanoparticles are renowned for their influential anti-microbial activity. Hence the present study over the biologically synthesized silver nanoparticles, exhibited a potential anti-biofilm activity that was tested in vitro on biofilms formed by Pseudomonas aeruginosa and Staphylococcus epidermidis during 24-h treatment. Treating these organisms with silver nanoparticles resulted in more than 95% inhibition in biofilm formation. The inhibition was known to be invariable of the species tested. As a result this study demonstrates the futuristic application of silver nanoparticles in treating microbial keratitis based on its potential anti-biofilm activity. Copyright 2010 Elsevier B.V. All rights reserved.

  2. Effect of nitrofurans and NO generators on biofilm formation by Pseudomonas aeruginosa PAO1 and Burkholderia cenocepacia 370.

    Science.gov (United States)

    Zaitseva, Julia; Granik, Vladimir; Belik, Alexandr; Koksharova, Olga; Khmel, Inessa

    2009-06-01

    Antibacterial drugs in the nitrofuran series, such as nitrofurazone, furazidin, nitrofurantoin and nifuroxazide, as well as the nitric oxide generators sodium nitroprusside and isosorbide mononitrate in concentrations that do not suppress bacterial growth, were shown to increase the capacity of pathogenic bacteria Pseudomonas aeruginosa PAO1 and Burkholderia cenocepacia 370 to form biofilms. At 25-100microg/ml, nitrofurans 2-2.5-fold enhanced biofilm formation of P. aeruginosa PAO1, and NO donors 3-6-fold. For B. cenocepacia 370, the enhancement was 2-5-fold (nitrofurans) and 4.5-fold (sodium nitroprusside), respectively.

  3. Antibiotic resistance and ndvB gene expression among biofilm ...

    African Journals Online (AJOL)

    A novel antibiotic resistant mechanism among biofilms is glucan-mediated sequestration in which ndvB gene encodes a glucosyltransferase involved in the formation of this glucans. We studied the biofilm formation and antibiotic susceptibility pattern of P. aeruginosa isolated from clinical samples, and measured the ...

  4. Pharmacokinetics/Pharmacodynamics of Colistin and Imipenem on Mucoid and Nonmucoid Pseudomonas aeruginosa Biofilms

    DEFF Research Database (Denmark)

    Hengzhuang, Wang; Wu, Hong; Ciofu, Oana

    2011-01-01

    The time course activity of Colistin and Imipenem on mucoid and non-mucoid biofilm-growing P. aeruginosa showed that compared with planktonic bacteria, the kinetics of Colistin and Imipenem retained the concentration- and time-dependent killings, respectively but higher doses of antibiotics and f...

  5. [Formation of the Pseudomonas aeruginosa PAO1 biofilms in the presence of hydrogen peroxide; the effect of the AiiA gene].

    Science.gov (United States)

    Pliuta, V A; Andreenko, Iu V; Kuznetsov, A E; Khmel', I A

    2013-01-01

    In the natural ecosystems, most bacteria exist as specifically organized biofilms attached to various surfaces; the biofilms have a complex architecture and are surrounded by an exopolymeric matrix. The bacteria in the biofilms are extremely resistant to antibacterial agents. The ability of the pathogenic bacteria to produce biofilms causes serious problems in medicine. Therefore, the study of the action of different compounds with antibacterial activity is of great interest. In this work, we studied the effect of the hydrogen peroxide (H2O2) on the formation of biofilms by Pseudomonas aeruginosa PAO1. It was shown that H2O2 in concentrations that do not suppress bacterial growth (or suppress it only weakly) stimulates the formation of the biofilms. At higher concentrations, H2O2 inhibits the formation of the biofilms. In order to determine if the stimulation of the biofilm formation depends on Quorum Sensing (QS) regulation, the plasmid pME6863 containing the heterologous gene aiiA encoding the N-acyl-homoserine lactonase AiiA was introduced into P. aeruginosa PAO1. The synthesis by cells of this enzyme degrading N-acyl-homoserine lactones (AHL), signaling molecules of the QS systems, led to the absence of the stimulation of the biofilm formation by the action of H2O2. This fact indicates that the stimulation of the biofilm formation in the presence of H2O2 depends on the functioning of the QS systems of the gene expression regulation of P. aeruginosa PAO1.

  6. Novel Multiscale Modeling Tool Applied to Pseudomonas aeruginosa Biofilm Formation

    OpenAIRE

    Biggs, Matthew B.; Papin, Jason A.

    2013-01-01

    Multiscale modeling is used to represent biological systems with increasing frequency and success. Multiscale models are often hybrids of different modeling frameworks and programming languages. We present the MATLAB-NetLogo extension (MatNet) as a novel tool for multiscale modeling. We demonstrate the utility of the tool with a multiscale model of Pseudomonas aeruginosa biofilm formation that incorporates both an agent-based model (ABM) and constraint-based metabolic modeling. The hybrid mod...

  7. Experimental Pseudomonas aeruginosa mediated rhino sinusitis in mink

    DEFF Research Database (Denmark)

    Kirkeby, S.; Hammer, A. S.; Høiby, N.

    2017-01-01

    The nasal and sinus cavities in children may serve as reservoirs for microorganisms that cause recurrent and chronic lung infections. This study evaluates whether the mink can be used as an animal model for studying Pseudomonas aeruginosa mediated rhino-sinusitis since there is no suitable...... in the infected mink shows features of carbohydrate expression comparable to what has been described in the respiratory system after Pseudomonas aeruginosa infection in humans. It is suggested that the mink is suitable for studying Pseudomonas aeruginosa mediated rhino-sinusitis....

  8. Biofilm formation by Staphylococcus epidermidis on peritoneal dialysis catheters and the effects of extracellular products from Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Pihl, Maria; Arvidsson, Anna; Skepö, Marie

    2013-01-01

    Biofilm formation by Staphylococcus epidermidis is a cause of infections related to peritoneal dialysis (PD). We have used a PD catheter flow-cell model in combination with confocal scanning laser microscopy and atomic force microscopy to study biofilm formation by S. epidermidis. Adherence....... aeruginosa contain promising substances for the prevention and treatment of biofilm infections, although further work is required to identity more active components....

  9. An Antipersister Strategy for Treatment of Chronic Pseudomonas aeruginosa Infections.

    Science.gov (United States)

    Koeva, Martina; Gutu, Alina D; Hebert, Wesley; Wager, Jeffrey D; Yonker, Lael M; O'Toole, George A; Ausubel, Frederick M; Moskowitz, Samuel M; Joseph-McCarthy, Diane

    2017-12-01

    Bacterial persisters are a quasidormant subpopulation of cells that are tolerant to antibiotic treatment. The combination of the aminoglycoside tobramycin with fumarate as an antibacterial potentiator utilizes an antipersister strategy that is aimed at reducing recurrent Pseudomonas aeruginosa infections by enhancing the killing of P. aeruginosa persisters. Stationary-phase cultures of P. aeruginosa were used to generate persister cells. A range of tobramycin concentrations was tested with a range of metabolite concentrations to determine the potentiation effect of the metabolite under a variety of conditions, including a range of pH values and in the presence of azithromycin or cystic fibrosis (CF) patient sputum. In addition, 96-well dish biofilm and colony biofilm assays were performed, and the cytotoxicity of the tobramycin-fumarate combination was determined utilizing a lactate dehydrogenase (LDH) assay. Enhanced killing of up to 6 orders of magnitude of P. aeruginosa persisters over a range of CF isolates, including mucoid and nonmucoid strains, was observed for the tobramycin-fumarate combination compared to killing with tobramycin alone. Furthermore, significant fumarate-mediated potentiation was seen in the presence of azithromycin or CF patient sputum. Fumarate also reduced the cytotoxicity of tobramycin-treated P. aeruginosa to human epithelial airway cells. Finally, in mucoid and nonmucoid CF isolates, complete eradication of P. aeruginosa biofilm was observed in the colony biofilm assay due to fumarate potentiation. These data suggest that a combination of tobramycin with fumarate as an antibacterial potentiator may be an attractive therapeutic for eliminating recurrent P. aeruginosa infections in CF patients through the eradication of bacterial persisters. Copyright © 2017 American Society for Microbiology.

  10. The biofilm-specific antibiotic resistance gene ndvB is important for expression of ethanol oxidation genes in Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Beaudoin, Trevor; Zhang, Li; Hinz, Aaron J; Parr, Christopher J; Mah, Thien-Fah

    2012-06-01

    Bacteria growing in biofilms are responsible for a large number of persistent infections and are often more resistant to antibiotics than are free-floating bacteria. In a previous study, we identified a Pseudomonas aeruginosa gene, ndvB, which is important for the formation of periplasmic glucans. We established that these glucans function in biofilm-specific antibiotic resistance by sequestering antibiotic molecules away from their cellular targets. In this study, we investigate another function of ndvB in biofilm-specific antibiotic resistance. DNA microarray analysis identified 24 genes that were responsive to the presence of ndvB. A subset of 20 genes, including 8 ethanol oxidation genes (ercS', erbR, exaA, exaB, eraR, pqqB, pqqC, and pqqE), was highly expressed in wild-type biofilm cells but not in ΔndvB biofilms, while 4 genes displayed the reciprocal expression pattern. Using quantitative real-time PCR, we confirmed the ndvB-dependent expression of the ethanol oxidation genes and additionally demonstrated that these genes were more highly expressed in biofilms than in planktonic cultures. Expression of erbR in ΔndvB biofilms was restored after the treatment of the biofilm with periplasmic extracts derived from wild-type biofilm cells. Inactivation of ethanol oxidation genes increased the sensitivity of biofilms to tobramycin. Together, these results reveal that ndvB affects the expression of multiple genes in biofilms and that ethanol oxidation genes are linked to biofilm-specific antibiotic resistance.

  11. Resistance of Pseudomonas aeruginosa to liquid disinfectants on contaminated surfaces before formation of biofilms.

    Science.gov (United States)

    Sagripanti, J L; Bonifacino, A

    2000-01-01

    A comparison was made of the effectiveness of popular disinfectants (Cavicide, Cidexplus, Clorox, Exspor, Lysol, Renalin, and Wavicide) under conditions prescribed for disinfection in the respective product labels on Pseudomonas aeruginosa either in suspension or deposited onto surfaces of metallic or polymeric plastic devices. The testing also included 7 nonformulated germicidal agents (glutaraldehyde, formaldehyde, peracetic acid, hydrogen peroxide, sodium hypochlorite, phenol, and cupric ascorbate) commonly used in disinfection and decontamination. Results showed that P. aeruginosa is on average 300-fold more resistant when present on contaminated surfaces than in suspension. This increase in resistance agrees with results reported in studies of biofilms, but unexpectedly, it precedes biofilm formation. The surface to which bacteria are attached can influence the effectiveness of disinfectants. Viable bacteria attached to devices may require dislodging through more than a one-step method for detection. The data, obtained with a sensitive and quantitative test, suggest that disinfectants are less effective on contaminated surfaces than generally acknowledged.

  12. Synergistic effect of 14-alpha-lipoyl andrographolide and various antibiotics on the formation of biofilms and production of exopolysaccharide and pyocyanin by Pseudomonas aeruginosa.

    Science.gov (United States)

    Zeng, Xiangping; Liu, Xiangyang; Bian, Jiang; Pei, Gang; Dai, Huanqin; Polyak, Steven W; Song, Fuhang; Ma, Li; Wang, Yuqiang; Zhang, Lixin

    2011-06-01

    Pseudomonas aeruginosa produces a biofilm that provides the bacteria with an effective barrier against antibiotics. Here, we investigated the synergy of various antibiotics with 14-alpha-lipoyl andrographolide (AL-1), focusing upon synthesis of the biofilm. AL-1 also inhibited the production of the exopolysaccharide and pyocyanin components. We propose that AL-1 may potentially serve as a cotherapy to combat P. aeruginosa.

  13. Antimicrobial activity of synthetic cationic peptides and lipopeptides derived from human lactoferricin against Pseudomonas aeruginosa planktonic cultures and biofilms.

    Science.gov (United States)

    Sánchez-Gómez, Susana; Ferrer-Espada, Raquel; Stewart, Philip S; Pitts, Betsey; Lohner, Karl; Martínez de Tejada, Guillermo

    2015-07-07

    Infections by Pseudomonas aeruginosa constitute a serious health threat because this pathogen -particularly when it forms biofilms - can acquire resistance to the majority of conventional antibiotics. This study evaluated the antimicrobial activity of synthetic peptides based on LF11, an 11-mer peptide derived from human lactoferricin against P. aeruginosa planktonic and biofilm-forming cells. We included in this analysis selected N-acylated derivatives of the peptides to analyze the effect of acylation in antimicrobial activity. To assess the efficacy of compounds against planktonic bacteria, microdilution assays to determine the minimal inhibitory concentration (MIC), minimum bactericidal concentration (MBC) and time-kill studies were conducted. The anti-biofilm activity of the agents was assessed on biofilms grown under static (on microplates) and dynamic (in a CDC-reactor) flow regimes. The antimicrobial activity of lipopeptides differed from that of non-acylated peptides in their killing mechanisms on planktonic and biofilm-forming cells. Thus, acylation enhanced the bactericidal activity of the parental peptides and resulted in lipopeptides that were uniformly bactericidal at their MIC. In contrast, acylation of the most potent anti-biofilm peptides resulted in compounds with lower anti-biofilm activity. Both peptides and lipopeptides displayed very rapid killing kinetics and all of them required less than 21 min to reduce 1,000 times the viability of planktonic cells when tested at 2 times their MBC. The peptides, LF11-215 (FWRIRIRR) and LF11-227 (FWRRFWRR), displayed the most potent anti-biofilm activity causing a 10,000 fold reduction in cell viability after 1 h of treatment at 10 times their MIC. At that concentration, these two compounds exhibited low citotoxicity on human cells. In addition to its bactericidal activity, LF11-227 removed more that 50 % of the biofilm mass in independent assays. Peptide LF11-215 and two of the shortest and least

  14. Heterogeneity of biofilms formed by nonmucoid Pseudomonas aeruginosa isolates from patients with cystic fibrosis

    DEFF Research Database (Denmark)

    Lee, Baoleri; Haagensen, Janus Anders Juul; Ciofu, O.

    2005-01-01

    lactone quorum-sensing molecules). Compared with wild-type strain PAO 1, most P. aeruginosa isolates demonstrated different degrees of reduction of adherence on polystyrene surfaces. The in vitro biofilm formation of isolates was investigated in a hydrodynamic flow system. Confocal laser scanning...... to phenotypic changes and in vitro biofilm formation. The physiological alterations were associated with a loss of motility (35% were nonmotile) and with decreased production of virulence factors (pyocyanin, proteases) and quorum-sensing molecules (45% of the isolates were unable to produce 3-O-C(12)-homoserine...

  15. Attenuation of quorum-sensing-dependent virulence factors and biofilm formation by medicinal plants against antibiotic resistant Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    P. Sankar Ganesh

    2018-01-01

    Full Text Available Pseudomonas aeruginosa use small signaling molecules such as acyl homoserine lactones (AHLs, which play an important role in release virulence factors and toxin for further establishment of host infection. Thus, involving with the QS system would provide alternative ways of preventing the pathogenicity. In the present study, totally six medicinal plants (Terminalia bellerica, Celastrus paniculatus, Kingiodendron pinnatum, Schleichera oleosa, Melastoma malabathricum, Garcinia gummi-gutta were screened for anti-QS activity using biomonitor strain of Chromobacterium violaceum CV12472. The primary screening of antimicrobial activity of all the plant extracts have inhibited the growth of tested bacterial species. Of these at the sub-minimum inhibitory concentration the methanol extract of T. bellerica (0.0625–0.5 mg/ml has significantly inhibited violacein production (20.07–66.22% in C. violaceum (CV12472. Consequently, the extract of T. bellerica has reduced the production of pyocyanin, exopolysaccharide and biofilm formation in P. aeruginosa strains. Fluorescence and scanning electron microscopy analysis confirmed the reduction of biofilm formation in P. aeruginosa strains when treated with T. bellerica. GC–MS analysis showed the active compounds inhibited the production of virulence factors of P. aeruginosa. The results suggest the possible use of this T. bellerica as an anti-QS and anti-biofilm agent to control Pseudomonas infection. Interference of QS provides an important means for the inhibition of bacterial virulence and thus aids in treatment strategies.

  16. Pseudomonas aeruginosa Biofilm Infections

    DEFF Research Database (Denmark)

    Rybtke, Morten; Hultqvist, Louise Dahl; Givskov, Michael

    2015-01-01

    Studies of biopsies from infectious sites, explanted tissue and medical devises have provided evidence that biofilms are the underlying cause of a variety of tissue-associated and implant-associated recalcitrant human infections. With a need for novel anti-biofilm treatment strategies, research...... in biofilm infection microbiology, biofilm formation mechanisms and biofilm-associated antimicrobial tolerance has become an important area in microbiology. Substantial knowledge about biofilm formation mechanisms, biofilm-associated antimicrobial tolerance and immune evasion mechanisms has been obtained...... through work with biofilms grown in in vitro experimental setups, and the relevance of this information in the context of chronic infections is being investigated by the use of animal models of infection. Because our current in vitro experimental setups and animal models have limitations, new advanced...

  17. Reinforcement of the bactericidal effect of ciprofloxacin on Pseudomonas aeruginosa biofilm by hyperbaric oxygen treatment

    DEFF Research Database (Denmark)

    Kolpen, Mette; Mousavi, Nabi; Sams, Thomas

    2016-01-01

    Chronic Pseudomonas aeruginosa lung infection is the most severe complication in cystic fibrosis patients. It is characterised by antibiotic-tolerant biofilms in the endobronchial mucus with zones of oxygen (O2) depletion mainly due to polymorphonuclear leucocyte activity. Whilst the exact mechan...

  18. Requirements for Pseudomonas aeruginosa Type I-F CRISPR-Cas Adaptation Determined Using a Biofilm Enrichment Assay.

    Science.gov (United States)

    Heussler, Gary E; Miller, Jon L; Price, Courtney E; Collins, Alan J; O'Toole, George A

    2016-11-15

    CRISPR (clustered regularly interspaced short palindromic repeat)-Cas (CRISPR-associated protein) systems are diverse and found in many archaea and bacteria. These systems have mainly been characterized as adaptive immune systems able to protect against invading mobile genetic elements, including viruses. The first step in this protection is acquisition of spacer sequences from the invader DNA and incorporation of those sequences into the CRISPR array, termed CRISPR adaptation. Progress in understanding the mechanisms and requirements of CRISPR adaptation has largely been accomplished using overexpression of cas genes or plasmid loss assays; little work has focused on endogenous CRISPR-acquired immunity from viral predation. Here, we developed a new biofilm-based assay system to enrich for Pseudomonas aeruginosa strains with new spacer acquisition. We used this assay to demonstrate that P. aeruginosa rapidly acquires spacers protective against DMS3vir, an engineered lytic variant of the Mu-like bacteriophage DMS3, through primed CRISPR adaptation from spacers present in the native CRISPR2 array. We found that for the P. aeruginosa type I-F system, the cas1 gene is required for CRISPR adaptation, recG contributes to (but is not required for) primed CRISPR adaptation, recD is dispensable for primed CRISPR adaptation, and finally, the ability of a putative priming spacer to prime can vary considerably depending on the specific sequences of the spacer. Our understanding of CRISPR adaptation has expanded largely through experiments in type I CRISPR systems using plasmid loss assays, mutants of Escherichia coli, or cas1-cas2 overexpression systems, but there has been little focus on studying the adaptation of endogenous systems protecting against a lytic bacteriophage. Here we describe a biofilm system that allows P. aeruginosa to rapidly gain spacers protective against a lytic bacteriophage. This approach has allowed us to probe the requirements for CRISPR adaptation in

  19. Macrolides decrease the minimal inhibitory concentration of anti-pseudomonal agents against Pseudomonas aeruginosa from cystic fibrosis patients in biofilm

    Directory of Open Access Journals (Sweden)

    Lutz Larissa

    2012-09-01

    Full Text Available Abstract Background Biofilm production is an important mechanism for bacterial survival and its association with antimicrobial resistance represents a challenge for the patient treatment. In this study we evaluated the in vitro action of macrolides in combination with anti-pseudomonal agents on biofilm-grown Pseudomonas aeruginosa recovered from cystic fibrosis (CF patients. Results A total of 64 isolates were analysed. The biofilm inhibitory concentration (BIC results were consistently higher than those obtained by the conventional method, minimal inhibitory concentration, (MIC for most anti-pseudomonal agents tested (ceftazidime: P = 0.001, tobramycin: P = 0.001, imipenem: P P = 0.005. When macrolides were associated with the anti-pseudomonal agents, the BIC values were reduced significantly for ceftazidime (P  0.001 and tobramycin (P  0.001, regardless the concentration of macrolides. Strong inhibitory quotient was observed when azithromycin at 8 mg/L was associated with all anti-pseudomonal agents tested in biofilm conditions. Conclusions P. aeruginosa from CF patients within biofilms are highly resistant to antibiotics but macrolides proved to augment the in vitro activity of anti-pseudomonal agents.

  20. Natural isothiocyanates express antimicrobial activity against developing and mature biofilms of Pseudomonas aeruginosa.

    Science.gov (United States)

    Kaiser, Stefan J; Mutters, Nico T; Blessing, Brigitte; Günther, Frank

    2017-06-01

    The antimicrobial properties of natural isothiocyanates (ITCs) found in plants such as nasturtium (Tropaeolum majus) and horseradish (Armoracia rusticana), and the need of new chemotherapeutic options for treatment of infections caused by multidrug-resistant and biofilm-forming Gram-negative bacteria such as Pseudomonas aeruginosa (Pa), led us to evaluate the effects of three major ITCs, allylisothiocyanate (AITC), benzylisothiocyanate (BITC), and phenylethyl-isothiocyanate (PEITC), and a mixture (ITCM) adapted to the ITC composition after release of active components out of natural sources. Out of 105Pa isolates 27 isolates with increased biofilm formation were selected for testing. The effects of ITCs on Pa were evaluated regarding (1) planktonic bacterial proliferation, (2) biofilm formation, (3) metabolic activity in mature biofilms, and (4) synergism of ITCs and antibiotics. (1) Each ITC had anti-Pa activity. Mean minimum inhibitory concentrations (MICs) were (μg/ml, mean±standard deviation): AITC 103±6.9; BITC, 2145±249; PEITC 29,423±1652; and ITCM, 140±5. (2) Treating bacteria with PEITC and ITCM in concentrations below the MIC significantly inhibited biofilm formation. Particularly, ITCM reduced biofilm mass and bacterial proliferation. (3) ITCs significantly inhibited metabolic activity in mature biofilms. (4) Combining ITCs with meropenem synergistically increased antimicrobial efficacy on Pa biofilms. ITCs represent a promising group of natural anti-infective compounds with activity against Pa biofilms. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  1. Disinfection of Pseudomonas aeruginosa biofilm contaminated tube lumens with ultraviolet C light emitting diodes

    DEFF Research Database (Denmark)

    Bak, Jimmy; Ladefoged, Søren D; Tvede, Michael

    2010-01-01

    with biofilm was investigated. It was shown that UVC light propagation was possible through both Teflon and catheter tubes (silicone). The disinfection efficiency of the diodes was demonstrated on tubes contaminated artificially with a Pseudomonas aeruginosa biofilm. The tubes were connected to a flow system...... and biofilms were produced during a 3 day period. Tubes in lengths of 10 (Teflon, silicone) and 20 cm (Teflon) were contaminated. Tubes for control and for UVC treatment were contaminated in parallel. Biofilms were sampled from the total inner surface of the tubes. Colony counts on the control samples were...... in the range of 5 x 10(5)-1.3 x 10(9) CFU ml(-1), with disinfection rates in the range 96-100%. The applied UVC doses corresponded to treatment times between 15 and 300 min. Disinfection (100%) was obtained in 10 cm Teflon tubes exposed for 30 min (detection limit...

  2. Disinfection of Pseudomonas aeruginosa biofilm contaminated tube lumens with ultraviolet C light emitting diodes

    DEFF Research Database (Denmark)

    Bak, Jimmy; Ladefoged, S.D.; Tvede, M.

    2010-01-01

    with biofilm was investigated. It was shown that UVC light propagation was possible through both Teflon and catheter tubes ( silicone). The disinfection efficiency of the diodes was demonstrated on tubes contaminated artificially with a Pseudomonas aeruginosa biofilm. The tubes were connected to a flow system...... and biofilms were produced during a 3 day period. Tubes in lengths of 10 ( Teflon, silicone) and 20 cm ( Teflon) were contaminated. Tubes for control and for UVC treatment were contaminated in parallel. Biofilms were sampled from the total inner surface of the tubes. Colony counts on the control samples were...... in the range of 5 x 10(5)-1.3 x 10(9) CFU ml(-1), with disinfection rates in the range 96-100%. The applied UVC doses corresponded to treatment times between 15 and 300 min. Disinfection (100%) was obtained in 10 cm Teflon tubes exposed for 30 min (detection limit...

  3. Pseudomonas aeruginosa Exhibits Deficient Biofilm Formation in the Absence of Class II and III Ribonucleotide Reductases Due to Hindered Anaerobic Growth.

    Science.gov (United States)

    Crespo, Anna; Pedraz, Lucas; Astola, Josep; Torrents, Eduard

    2016-01-01

    Chronic lung infections by the ubiquitous and extremely adaptable opportunistic pathogen Pseudomonas aeruginosa correlate with the formation of a biofilm, where bacteria grow in association with an extracellular matrix and display a wide range of changes in gene expression and metabolism. This leads to increased resistance to physical stress and antibiotic therapies, while enhancing cell-to-cell communication. Oxygen diffusion through the complex biofilm structure generates an oxygen concentration gradient, leading to the appearance of anaerobic microenvironments. Ribonucleotide reductases (RNRs) are a family of highly sophisticated enzymes responsible for the synthesis of the deoxyribonucleotides, and they constitute the only de novo pathway for the formation of the building blocks needed for DNA synthesis and repair. P. aeruginosa is one of the few bacteria encoding all three known RNR classes (Ia, II, and III). Class Ia RNRs are oxygen dependent, class II are oxygen independent, and class III are oxygen sensitive. A tight control of RNR activity is essential for anaerobic growth and therefore for biofilm development. In this work we explored the role of the different RNR classes in biofilm formation under aerobic and anaerobic initial conditions and using static and continuous-flow biofilm models. We demonstrated the importance of class II and III RNR for proper cell division in biofilm development and maturation. We also determined that these classes are transcriptionally induced during biofilm formation and under anaerobic conditions. The molecular mechanism of their anaerobic regulation was also studied, finding that the Anr/Dnr system is responsible for class II RNR induction. These data can be integrated with previous knowledge about biofilms in a model where these structures are understood as a set of layers determined by oxygen concentration and contain cells with different RNR expression profiles, bringing us a step closer to the understanding of this

  4. The LapG protein plays a role in Pseudomonas aeruginosa biofilm formation by controlling the presence of the CdrA adhesin on the cell surface

    DEFF Research Database (Denmark)

    Rybtke, Morten; Berthelsen, Jens; Yang, Liang

    2015-01-01

    Pseudomonas aeruginosa is a clinically relevant species involved in biofilm-based chronic infections. We provide evidence that the P. aeruginosa LapG protein functions as a periplasmic protease that can cleave the protein adhesin CdrA off the cell surface, and thereby plays a role in biofilm...... formation and biofilm dispersal. The P. aeruginosa LapG protein is shown to be a functional homolog of the Pseudomonas putida LapG protein which has previously been shown to function as a periplasmic protease that targets the surface adhesin LapA. Transposon mutagenesis and characterization of defined...... and whole-cell protein fractions showed that CdrA was retained in the whole-cell protein fraction when LapG was absent, whereas it was found in the culture supernatant when LapG was present. The finding that CdrA is a target of LapG in P. aeruginosa is surprising because CdrA has no homology to LapA....

  5. A 96-well-plate-based optical method for the quantitative and qualitative evaluation of Pseudomonas aeruginosa biofilm formation and its application to susceptibility testing.

    Science.gov (United States)

    Müsken, Mathias; Di Fiore, Stefano; Römling, Ute; Häussler, Susanne

    2010-08-01

    A major reason for bacterial persistence during chronic infections is the survival of bacteria within biofilm structures, which protect cells from environmental stresses, host immune responses and antimicrobial therapy. Thus, there is concern that laboratory methods developed to measure the antibiotic susceptibility of planktonic bacteria may not be relevant to chronic biofilm infections, and it has been suggested that alternative methods should test antibiotic susceptibility within a biofilm. In this paper, we describe a fast and reliable protocol for using 96-well microtiter plates for the formation of Pseudomonas aeruginosa biofilms; the method is easily adaptable for antimicrobial susceptibility testing. This method is based on bacterial viability staining in combination with automated confocal laser scanning microscopy. The procedure simplifies qualitative and quantitative evaluation of biofilms and has proven to be effective for standardized determination of antibiotic efficiency on P. aeruginosa biofilms. The protocol can be performed within approximately 60 h.

  6. Inoculation density and nutrient level determine the formation of mushroom-shaped structures in Pseudomonas aeruginosa biofilms

    Science.gov (United States)

    Ghanbari, Azadeh; Dehghany, Jaber; Schwebs, Timo; Müsken, Mathias; Häussler, Susanne; Meyer-Hermann, Michael

    2016-09-01

    Pseudomonas aeruginosa often colonises immunocompromised patients and the lungs of cystic fibrosis patients. It exhibits resistance to many antibiotics by forming biofilms, which makes it hard to eliminate. P. aeruginosa biofilms form mushroom-shaped structures under certain circumstances. Bacterial motility and the environment affect the eventual mushroom morphology. This study provides an agent-based model for the bacterial dynamics and interactions influencing bacterial biofilm shape. Cell motility in the model relies on recently published experimental data. Our simulations show colony formation by immotile cells. Motile cells escape from a single colony by nutrient chemotaxis and hence no mushroom shape develops. A high number density of non-motile colonies leads to migration of motile cells onto the top of the colonies and formation of mushroom-shaped structures. This model proposes that the formation of mushroom-shaped structures can be predicted by parameters at the time of bacteria inoculation. Depending on nutrient levels and the initial number density of stalks, mushroom-shaped structures only form in a restricted regime. This opens the possibility of early manipulation of spatial pattern formation in bacterial colonies, using environmental factors.

  7. Inoculation density and nutrient level determine the formation of mushroom-shaped structures in Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Ghanbari, Azadeh; Dehghany, Jaber; Schwebs, Timo; Müsken, Mathias; Häussler, Susanne; Meyer-Hermann, Michael

    2016-09-09

    Pseudomonas aeruginosa often colonises immunocompromised patients and the lungs of cystic fibrosis patients. It exhibits resistance to many antibiotics by forming biofilms, which makes it hard to eliminate. P. aeruginosa biofilms form mushroom-shaped structures under certain circumstances. Bacterial motility and the environment affect the eventual mushroom morphology. This study provides an agent-based model for the bacterial dynamics and interactions influencing bacterial biofilm shape. Cell motility in the model relies on recently published experimental data. Our simulations show colony formation by immotile cells. Motile cells escape from a single colony by nutrient chemotaxis and hence no mushroom shape develops. A high number density of non-motile colonies leads to migration of motile cells onto the top of the colonies and formation of mushroom-shaped structures. This model proposes that the formation of mushroom-shaped structures can be predicted by parameters at the time of bacteria inoculation. Depending on nutrient levels and the initial number density of stalks, mushroom-shaped structures only form in a restricted regime. This opens the possibility of early manipulation of spatial pattern formation in bacterial colonies, using environmental factors.

  8. The Pseudomonas aeruginosa chemotaxis methyltransferase CheR1 impacts on bacterial surface sampling.

    Directory of Open Access Journals (Sweden)

    Juliane Schmidt

    Full Text Available The characterization of factors contributing to the formation and development of surface-associated bacterial communities known as biofilms has become an area of intense interest since biofilms have a major impact on human health, the environment and industry. Various studies have demonstrated that motility, including swimming, swarming and twitching, seems to play an important role in the surface colonization and establishment of structured biofilms. Thereby, the impact of chemotaxis on biofilm formation has been less intensively studied. Pseudomonas aeruginosa has a very complex chemosensory system with two Che systems implicated in flagella-mediated motility. In this study, we demonstrate that the chemotaxis protein CheR1 is a methyltransferase that binds S-adenosylmethionine and transfers a methyl group from this methyl donor to the chemoreceptor PctA, an activity which can be stimulated by the attractant serine but not by glutamine. We furthermore demonstrate that CheR1 does not only play a role in flagella-mediated chemotaxis but that its activity is essential for the formation and maintenance of bacterial biofilm structures. We propose a model in which motility and chemotaxis impact on initial attachment processes, dispersion and reattachment and increase the efficiency and frequency of surface sampling in P. aeruginosa.

  9. Rapid development in vitro and in vivo of resistance to ceftazidime in biofilm-growing Pseudomonas aeruginosa due to chromosomal beta-lactamase

    DEFF Research Database (Denmark)

    Bagge, N; Ciofu, O; Skovgaard, L T

    2000-01-01

    The aim of this study was to examine the development of resistance of biofilm-growing P. aeruginosa during treatment with ceftazidime. Biofilms were established in vitro using a modified Robbins device (MRD) and in vivo in the rat model of chronic lung infection. Three P. aeruginosa strains...... of ceftazidime to biofilms established in MDR, a statistically significant development of resistance to ceftazidime in PAO 579 or 19676A bacterial populations occurred. When ceftazidime was administered 4 h/day (200 mg/l) for 2 weeks, the frequency of resistant 19676A having MIC>25 mg/l was 4.4 10(-1) compared...... to 6.0-10(-5) in the control biofilm. The same trend was observed after continuous administration of ceftazidime. MICceftazidime of the more resistant variants was increased 500-fold for PAO 579 and 8-fold for 19676A, and the specific basal beta-lactamase activities from 19 to 1,400 units for PAO 579...

  10. Pseudomonas Aeruginosa: interactions with organisms in the environment and cells of the immune defence

    DEFF Research Database (Denmark)

    Skindersø, Mette Elena

    2008-01-01

    , which emphasises the urgent need for development of novel strategies that will help us to defeat this pathogen. P. aeruginosa biofilm cells display a multicellular-like coordinated behaviour and control expression of virulence factors, elements involved in biofilm development and immunomodulating...... factors by means of signal molecule mediated communication, known as quorum sensing. This thesis explores a strategy which aims to counteract P. aeruginosa virulence and pathogenicity by impeding its cell-to-cell communication. A treatment regime, which focuses on targeting bacterial communication instead......; two quorum sensing signal molecules; the Pseudomonas Quinolone Signal and N-3-oxododecanoyl L-homoserine lactone exhibit the ability to modulate activities of the immune defence in addition to functioning as quorum sensing mediators. The two signal molecules impair activities of immune cells crucial...

  11. A Genotypic Analysis of Five P. aeruginosa Strains after Biofilm Infection by Phages Targeting Different Cell Surface Receptors

    Directory of Open Access Journals (Sweden)

    Diana P. Pires

    2017-06-01

    Full Text Available Antibiotic resistance constitutes one of the most serious threats to the global public health and urgently requires new and effective solutions. Bacteriophages are bacterial viruses increasingly recognized as being good alternatives to traditional antibiotic therapies. In this study, the efficacy of phages, targeting different cell receptors, against Pseudomonas aeruginosa PAO1 biofilm and planktonic cell cultures was evaluated over the course of 48 h. Although significant reductions in the number of viable cells were achieved for both cases, the high level of adaptability of the bacteria in response to the selective pressure caused by phage treatment resulted in the emergence of phage-resistant variants. To further investigate the genetic makeup of phage-resistant variants isolated from biofilm infection experiments, some of these bacteria were selected for phenotypic and genotypic characterization. Whole genome sequencing was performed on five phage-resistant variants and all of them carried mutations affecting the galU gene as well as one of pil genes. The sequencing analysis further revealed that three of the P. aeruginosa PAO1 variants carry large deletions (>200 kbp in their genomes. Complementation of the galU mutants with wild-type galU in trans restored LPS expression on the bacterial cell surface of these bacterial strains and rendered the complemented strains to be sensitive to phages. This provides unequivocal evidence that inactivation of galU function was associated with resistance to the phages that uses LPS as primary receptors. Overall, this work demonstrates that P. aeruginosa biofilms can survive phage attack and develop phage-resistant variants exhibiting defective LPS production and loss of type IV pili that are well adapted to the biofilm mode of growth.

  12. Species-independent attraction to biofilms through electrical signaling

    Science.gov (United States)

    Humphries, Jacqueline; Xiong, Liyang; Liu, Jintao; Prindle, Arthur; Yuan, Fang; Arjes, Heidi A.; Tsimring, Lev; Süel, Gürol M.

    2017-01-01

    Summary Bacteria residing within biofilm communities can coordinate their behavior through cell-to-cell signaling. However, it remains unclear if these signals can also influence the behavior of distant cells that are not part of the community. Using a microfluidic approach, we find that potassium ion channel-mediated electrical signaling generated by a Bacillus subtilis biofilm can attract distant cells. Integration of experiments and mathematical modeling indicates that extracellular potassium emitted from the biofilm alters the membrane potential of distant cells, thereby directing their motility. This electrically-mediated attraction appears to be a generic mechanism that enables cross-species interactions, as Pseudomonas aeruginosa cells also become attracted to the electrical signal released by the B. subtilis biofilm. Cells within a biofilm community can thus not only coordinate their own behavior, but also influence the behavior of diverse bacteria at a distance through long-range electrical signaling. PMID:28086091

  13. Dynamics of development and dispersal in sessile microbial communities: examples from Pseudomonas aeruginosa and Pseudomonas putida model biofilms

    DEFF Research Database (Denmark)

    Klausen, M.; Gjermansen, Morten; Kreft, J.-U.

    2006-01-01

    Surface-associated microbial communities in many cases display dynamic developmental patterns. Model biofilms formed by Pseudomonas aeruginosa and Pseudomonas putida in laboratory flow-chamber setups represent examples of such behaviour. Dependent on the experimental conditions the bacteria...

  14. Candida albicans ethanol stimulates Pseudomonas aeruginosa WspR-controlled biofilm formation as part of a cyclic relationship involving phenazines.

    Directory of Open Access Journals (Sweden)

    Annie I Chen

    2014-10-01

    Full Text Available In chronic infections, pathogens are often in the presence of other microbial species. For example, Pseudomonas aeruginosa is a common and detrimental lung pathogen in individuals with cystic fibrosis (CF and co-infections with Candida albicans are common. Here, we show that P. aeruginosa biofilm formation and phenazine production were strongly influenced by ethanol produced by the fungus C. albicans. Ethanol stimulated phenotypes that are indicative of increased levels of cyclic-di-GMP (c-di-GMP, and levels of c-di-GMP were 2-fold higher in the presence of ethanol. Through a genetic screen, we found that the diguanylate cyclase WspR was required for ethanol stimulation of c-di-GMP. Multiple lines of evidence indicate that ethanol stimulates WspR signaling through its cognate sensor WspA, and promotes WspR-dependent activation of Pel exopolysaccharide production, which contributes to biofilm maturation. We also found that ethanol stimulation of WspR promoted P. aeruginosa colonization of CF airway epithelial cells. P. aeruginosa production of phenazines occurs both in the CF lung and in culture, and phenazines enhance ethanol production by C. albicans. Using a C. albicans adh1/adh1 mutant with decreased ethanol production, we found that fungal ethanol strongly altered the spectrum of P. aeruginosa phenazines in favor of those that are most effective against fungi. Thus, a feedback cycle comprised of ethanol and phenazines drives this polymicrobial interaction, and these relationships may provide insight into why co-infection with both P. aeruginosa and C. albicans has been associated with worse outcomes in cystic fibrosis.

  15. Phenotypes of non-attached Pseudomonas aeruginosa aggregates resemble surface attached biofilm.

    Directory of Open Access Journals (Sweden)

    Morten Alhede

    Full Text Available For a chronic infection to be established, bacteria must be able to cope with hostile conditions such as low iron levels, oxidative stress, and clearance by the host defense, as well as antibiotic treatment. It is generally accepted that biofilm formation facilitates tolerance to these adverse conditions. However, microscopic investigations of samples isolated from sites of chronic infections seem to suggest that some bacteria do not need to be attached to surfaces in order to establish chronic infections. In this study we employed scanning electron microscopy, confocal laser scanning microscopy, RT-PCR as well as traditional culturing techniques to study the properties of Pseudomonas aeruginosa aggregates. We found that non-attached aggregates from stationary-phase cultures have comparable growth rates to surface attached biofilms. The growth rate estimations indicated that, independently of age, both aggregates and flow-cell biofilm had the same slow growth rate as a stationary phase shaking cultures. Internal structures of the aggregates matrix components and their capacity to survive otherwise lethal treatments with antibiotics (referred to as tolerance and resistance to phagocytes were also found to be strikingly similar to flow-cell biofilms. Our data indicate that the tolerance of both biofilms and non-attached aggregates towards antibiotics is reversible by physical disruption. We provide evidence that the antibiotic tolerance is likely to be dependent on both the physiological states of the aggregates and particular matrix components. Bacterial surface-attachment and subsequent biofilm formation are considered hallmarks of the capacity of microbes to cause persistent infections. We have observed non-attached aggregates in the lungs of cystic fibrosis patients; otitis media; soft tissue fillers and non-healing wounds, and we propose that aggregated cells exhibit enhanced survival in the hostile host environment, compared with non

  16. Flow environment and matrix structure interact to determine spatial competition in Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Nadell, Carey D; Ricaurte, Deirdre; Yan, Jing; Drescher, Knut; Bassler, Bonnie L

    2017-01-13

    Bacteria often live in biofilms, which are microbial communities surrounded by a secreted extracellular matrix. Here, we demonstrate that hydrodynamic flow and matrix organization interact to shape competitive dynamics in Pseudomonas aeruginosa biofilms. Irrespective of initial frequency, in competition with matrix mutants, wild-type cells always increase in relative abundance in planar microfluidic devices under simple flow regimes. By contrast, in microenvironments with complex, irregular flow profiles - which are common in natural environments - wild-type matrix-producing and isogenic non-producing strains can coexist. This result stems from local obstruction of flow by wild-type matrix producers, which generates regions of near-zero shear that allow matrix mutants to locally accumulate. Our findings connect the evolutionary stability of matrix production with the hydrodynamics and spatial structure of the surrounding environment, providing a potential explanation for the variation in biofilm matrix secretion observed among bacteria in natural environments.

  17. Strain- and Substrate-Dependent Redox Mediator and Electricity Production by Pseudomonas aeruginosa.

    Science.gov (United States)

    Bosire, Erick M; Blank, Lars M; Rosenbaum, Miriam A

    2016-08-15

    Pseudomonas aeruginosa is an important, thriving member of microbial communities of microbial bioelectrochemical systems (BES) through the production of versatile phenazine redox mediators. Pure culture experiments with a model strain revealed synergistic interactions of P. aeruginosa with fermenting microorganisms whereby the synergism was mediated through the shared fermentation product 2,3-butanediol. Our work here shows that the behavior and efficiency of P. aeruginosa in mediated current production is strongly dependent on the strain of P. aeruginosa We compared levels of phenazine production by the previously investigated model strain P. aeruginosa PA14, the alternative model strain P. aeruginosa PAO1, and the BES isolate Pseudomonas sp. strain KRP1 with glucose and the fermentation products 2,3-butanediol and ethanol as carbon substrates. We found significant differences in substrate-dependent phenazine production and resulting anodic current generation for the three strains, with the BES isolate KRP1 being overall the best current producer and showing the highest electrochemical activity with glucose as a substrate (19 μA cm(-2) with ∼150 μg ml(-1) phenazine carboxylic acid as a redox mediator). Surprisingly, P. aeruginosa PAO1 showed very low phenazine production and electrochemical activity under all tested conditions. Microbial fuel cells and other microbial bioelectrochemical systems hold great promise for environmental technologies such as wastewater treatment and bioremediation. While there is much emphasis on the development of materials and devices to realize such systems, the investigation and a deeper understanding of the underlying microbiology and ecology are lagging behind. Physiological investigations focus on microorganisms exhibiting direct electron transfer in pure culture systems. Meanwhile, mediated electron transfer with natural redox compounds produced by, for example, Pseudomonas aeruginosa might enable an entire microbial

  18. The Pseudomonas aeruginosa PSL Polysaccharide Is a Social but Noncheatable Trait in Biofilms.

    Science.gov (United States)

    Irie, Yasuhiko; Roberts, Aled E L; Kragh, Kasper N; Gordon, Vernita D; Hutchison, Jaime; Allen, Rosalind J; Melaugh, Gavin; Bjarnsholt, Thomas; West, Stuart A; Diggle, Stephen P

    2017-06-20

    Extracellular polysaccharides are compounds secreted by microorganisms into the surrounding environment, and they are important for surface attachment and maintaining structural integrity within biofilms. The social nature of many extracellular polysaccharides remains unclear, and it has been suggested that they could function as either cooperative public goods or as traits that provide a competitive advantage. Here, we empirically tested the cooperative nature of the PSL polysaccharide, which is crucial for the formation of biofilms in Pseudomonas aeruginosa We show that (i) PSL is not metabolically costly to produce; (ii) PSL provides population-level benefits in biofilms, for both growth and antibiotic tolerance; (iii) the benefits of PSL production are social and are shared with other cells; (iv) the benefits of PSL production appear to be preferentially directed toward cells which produce PSL; (v) cells which do not produce PSL are unable to successfully exploit cells which produce PSL. Taken together, this suggests that PSL is a social but relatively nonexploitable trait and that growth within biofilms selects for PSL-producing strains, even when multiple strains are on a patch (low relatedness at the patch level). IMPORTANCE Many studies have shown that bacterial traits, such as siderophores and quorum sensing, are social in nature. This has led to an impression that secreted traits act as public goods, which are costly to produce but benefit both the producing cell and its surrounding neighbors. Theories and subsequent experiments have shown that such traits are exploitable by asocial cheats, but we show here that this does not always hold true. We demonstrate that the Pseudomonas aeruginosa exopolysaccharide PSL provides social benefits to populations but that it is nonexploitable, because most of the fitness benefits accrue to PSL-producing cells. Our work builds on an increasing body of work showing that secreted traits can have both private and public

  19. Anti-Pseudomonas aeruginosa IgY Antibodies Induce Specific Bacterial Aggregation and Internalization in Human Polymorphonuclear Neutrophils

    DEFF Research Database (Denmark)

    Thomsen, K.; Christophersen, L.; Bjarnsholt, T.

    2015-01-01

    with P. aeruginosa by augmenting the phagocytic competence of PMNs may postpone the deteriorating chronic biofilm infection. Anti-P. aeruginosa IgY antibodies significantly increase the PMN-mediated respiratory burst and subsequent bacterial killing of P. aeruginosa in vitro. The mode of action...... is attributed to IgY-facilitated formation of immobilized bacteria in aggregates, as visualized by fluorescence microscopy and the induction of increased bacterial hydrophobicity. Thus, the present study demonstrates that avian egg yolk immunoglobulins (IgY) targeting P. aeruginosa modify bacterial fitness...

  20. The anti-cancerous drug doxorubicin decreases the c-di-GMP content in Pseudomonas aeruginosa but promotes biofilm formation

    DEFF Research Database (Denmark)

    Groizeleau, Julie; Rybtke, Morten; Andersen, Jens Bo

    2016-01-01

    Current antibiotic treatments are insufficient in eradicating bacterial biofilms, which represent the primary cause of chronic bacterial infections. Thus, there is an urgent need for new strategies to eradicate biofilm infections. The second messenger c-di-GMP is a positive regulator of biofilm...... formation in many clinically relevant bacteria. It is hypothesized that drugs lowering the intracellular level of c-di-GMP will force biofilm bacteria into a more treatable planktonic lifestyle. To identify compounds capable of lowering c-di-GMP levels in Pseudomonas aeruginosa, we screened 5000 compounds...... for their potential c-di-GMP-lowering effect using a recently developed c-di-GMP biosensor strain. Our screen identified the anti-cancerous drug doxorubicin as a potent c-di-GMP inhibitor. In addition, the drug decreased the transcription of many biofilm-related genes. However, despite its effect on the c-di-GMP...

  1. Formation of hydroxyl radicals contributes to the bactericidal activity of ciprofloxacin against Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Jensen, Peter Østrup; Briales, Alejandra; Brochmann, Rikke Prejh

    2014-01-01

    induction of cytotoxic hydroxyl radicals (OH˙) during antibiotic treatment of planktonically grown cells may contribute to action of the commonly used antibiotic ciprofloxacin on P. aeruginosa biofilms. For this purpose, WT PAO1, a catalase deficient ΔkatA and a ciprofloxacin resistant mutant of PAO1 (gyr...

  2. Clearance of Pseudomonas aeruginosa Foreign-Body Biofilm Infections through Reduction of the Cyclic Di-GMP Level in the Bacteria

    DEFF Research Database (Denmark)

    Christensen, Louise D.; van Gennip, Maria; Rybtke, Morten Theil

    2013-01-01

    Opportunistic pathogenic bacteria can engage in biofilm-based infections that evade immune responses and develop into chronic conditions. Because conventional antimicrobials cannot efficiently eradicate biofilms, there is an urgent need to develop alternative measures to combat biofilm infections....... It has recently been established that the secondary messenger cyclic diguanosine monophosphate (c-di-GMP) functions as a positive regulator of biofilm formation in several different bacteria. In the present study we investigated whether manipulation of the c-di-GMP level in bacteria potentially can...... be used for biofilm control in vivo. We constructed a Pseudomonas aeruginosa strain in which a reduction in the c-di-GMP level can be achieved via induction of the Escherichia coli YhjH c-di-GMP phosphodiesterase. Initial experiments showed that induction of yhjH expression led to dispersal...

  3. Biofilms

    OpenAIRE

    López, Daniel; Vlamakis, Hera; Kolter, Roberto

    2010-01-01

    The ability to form biofilms is a universal attribute of bacteria. Biofilms are multicellular communities held together by a self-produced extracellular matrix. The mechanisms that different bacteria employ to form biofilms vary, frequently depending on environmental conditions and specific strain attributes. In this review, we emphasize four well-studied model systems to give an overview of how several organisms form biofilms: Escherichia coli, Pseudomonas aeruginosa, Bacillus subtilis, and ...

  4. Effect of Negative Pressure on Proliferation, Virulence Factor Secretion, Biofilm Formation, and Virulence-Regulated Gene Expression of Pseudomonas aeruginosa In Vitro

    Directory of Open Access Journals (Sweden)

    Guo-Qi Wang

    2016-01-01

    Full Text Available Objective. To investigate the effect of negative pressure conditions induced by NPWT on P. aeruginosa. Methods. P. aeruginosa was cultured in a Luria–Bertani medium at negative pressure of −125 mmHg for 24 h in the experimental group and at atmospheric pressure in the control group. The diameters of the colonies of P. aeruginosa were measured after 24 h. ELISA kit, orcinol method, and elastin-Congo red assay were used to quantify the virulence factors. Biofilm formation was observed by staining with Alexa Fluor® 647 conjugate of concanavalin A (Con A. Virulence-regulated genes were determined by quantitative RT-PCR. Results. As compared with the control group, growth of P. aeruginosa was inhibited by negative pressure. The colony size under negative pressure was significantly smaller in the experimental group than that in the controls (p<0.01. Besides, reductions in the total amount of virulence factors were observed in the negative pressure group, including exotoxin A, rhamnolipid, and elastase. RT-PCR results revealed a significant inhibition in the expression level of virulence-regulated genes. Conclusion. Negative pressure could significantly inhibit the growth of P. aeruginosa. It led to a decrease in the virulence factor secretion, biofilm formation, and a reduction in the expression level of virulence-regulated genes.

  5. Biofilms.

    Science.gov (United States)

    López, Daniel; Vlamakis, Hera; Kolter, Roberto

    2010-07-01

    The ability to form biofilms is a universal attribute of bacteria. Biofilms are multicellular communities held together by a self-produced extracellular matrix. The mechanisms that different bacteria employ to form biofilms vary, frequently depending on environmental conditions and specific strain attributes. In this review, we emphasize four well-studied model systems to give an overview of how several organisms form biofilms: Escherichia coli, Pseudomonas aeruginosa, Bacillus subtilis, and Staphylococcus aureus. Using these bacteria as examples, we discuss the key features of biofilms as well as mechanisms by which extracellular signals trigger biofilm formation.

  6. Melittin and its potential in the destruction and inhibition of the biofilm formation by Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa isolated from bovine milk.

    Science.gov (United States)

    Picoli, Tony; Peter, Cristina Mendes; Zani, João Luíz; Waller, Stefanie Bressan; Lopes, Matheus Gomes; Boesche, Kamilla Neutzling; Vargas, Gilberto D Ávila; Hübner, Silvia de Oliveira; Fischer, Geferson

    2017-11-01

    Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa stand out in veterinary and human medicine for their role in opportunistic infections and their pathogenic mechanisms, including the biofilms formation. It was investigated the antibacterial activity of melittin and antibiofilm of such bacteria. Twelve strains of these microorganisms isolated from bovine milk were used, as well as the strains S. aureus ATCC 12600, E. coli ATCC 8739 and Pseudomonas aeruginosa ATCC 15442. The minimum inhibitory concentrations (MIC) and minimum bactericidal concentration (MBC) were determined by broth microdilution technique. The biofilms were formed in 96-well plates and melittin on these colonies was added at different concentrations and times. Bacteria previously exposed to melittin were evaluated for inhibition of biofilm production. The MIC and MBC were respectively in μg/mL: S. aureus (6-7 and 32-64), E. coli (40-42.5 and 64-128) and P. aeruginosa (65-70 and 64-128). S. aureus biofilms were more sensitive to the action of melittin, since upon exposure to a concentration 10 times lower than the MIC for 4 h, was completely destroyed. In Gram negative bacteria, the pre-formed biofilm was destroyed only when exposed for 4 h under the MIC. With respect to inhibition of biofilm production, S. aureus was the most sensitive again because produced only 37.2% of the biofilm formed by the control (without previous exposure to melittin), when exposed to the MIC, and at a concentration hundred times smaller than MIC, this microorganism produced 75.2% of the biofilm. E. coli was the most resistant bacteria and produced 56.3% of the biofilm, even if previously exposed to melittin MIC. Melittin presents desirable effects in combating microorganisms studied both at your disposal, biofilm destruction and inhibition of the formation, and maybe used in future studies of new strategies to combat infections caused by these pathogens. Copyright © 2017 Elsevier Ltd. All

  7. Pseudomonas aeruginosa Biofilms

    DEFF Research Database (Denmark)

    Alhede, Maria; Bjarnsholt, Thomas; Givskov, Michael

    2014-01-01

    biofilms, which protect the aggregated, biopolymer-embedded bacteria from the detrimental actions of antibiotic treatments and host immunity. A key component in the protection against innate immunity is rhamnolipid, which is a quorum sensing (QS)-regulated virulence factor. QS is a cell-to-cell signaling...... mechanism used to coordinate expression of virulence and protection of aggregated biofilm cells. Rhamnolipids are known for their ability to cause hemolysis and have been shown to cause lysis of several cellular components of the human immune system, for example, macrophages and polymorphonuclear leukocytes...

  8. Development and maturation of Escherichia coli K-12 biofilms

    DEFF Research Database (Denmark)

    Reisner, A.; Haagensen, J.A.J.; Schembri, Mark

    2003-01-01

    The development and maturation of E. coli biofilms in flow-chambers was investigated. We found that the presence of transfer constitutive IncF plasmids induced biofilm development forming structures resembling those reported for Pseudomonas aeruginosa . The development occurred in a step...... occurred in conjugation pilus proficient plasmid-carrying strains. The final shapes of the expanding structures in the mature biofilm seem to be determined by the pilus configuration, as various mutants affected in the processing and activity of the transfer pili displayed differently structured biofilms....... We further provide evidence that flagella, type 1 fimbriae, curli and Ag43 are all dispensable for the observed biofilm maturation. In addition, our results indicate that cell-to-cell signalling mediated by autoinducer 2 (AI-2) is not required for differentiation of E. coli within a biofilm community...

  9. In vitro production of biofilm in a flow cell system in a strain of Pseudomonas aeruginosa and Staphylococcus aureus and determination of efficiency of ciprofloxacin against them

    Directory of Open Access Journals (Sweden)

    Soham Gupta

    2011-01-01

    Full Text Available Background: Microorganisms develop biofilm on various medical devices. The process is particularly relevant in public health since biofilm associated organisms are much more resistant to antibiotics and have a potential to cause infections in patients with indwelling medical devices. Materials and Methods: To determine the efficiency of an antibiotic against the biofilm it is inappropriate to use traditional technique of determining Minimum Inhibitory Concentration (MIC on the free floating laboratory phenotype. Thus we have induced formation of biofilm in two strains (Pseudomonas aeruginosa and Staphylococcus aureus, which showed heavy growth of biofilm in screening by Tube method in a flow cell system and determined their antibiotic susceptibility against ciprofloxacin by agar dilution method in the range (0.25 mg/ml to 8 mg/ml. The MIC value of ciprofloxacin for the biofilm produced organism was compared with its free form and a standard strain as control on the same plates. Observations: Both the biofilm produced strains showed a higher resistance (MIC > 8 mg/ml than its free form, which were 2 μg/ml for Pseudomonas aeruginosa and 4 mg/ml for Staphylococcus aureus. Thus biofilm can pose a threat in the patient treatment.

  10. Pseudomonas aeruginosa Alters Staphylococcus aureus Sensitivity to Vancomycin in a Biofilm Model of Cystic Fibrosis Infection

    Directory of Open Access Journals (Sweden)

    Giulia Orazi

    2017-07-01

    Full Text Available The airways of cystic fibrosis (CF patients have thick mucus, which fosters chronic, polymicrobial infections. Pseudomonas aeruginosa and Staphylococcus aureus are two of the most prevalent respiratory pathogens in CF patients. In this study, we tested whether P. aeruginosa influences the susceptibility of S. aureus to frontline antibiotics used to treat CF lung infections. Using our in vitro coculture model, we observed that addition of P. aeruginosa supernatants to S. aureus biofilms grown either on epithelial cells or on plastic significantly decreased the susceptibility of S. aureus to vancomycin. Mutant analyses showed that 2-n-heptyl-4-hydroxyquinoline N-oxide (HQNO, a component of the P. aeruginosa Pseudomonas quinolone signal (PQS system, protects S. aureus from the antimicrobial activity of vancomycin. Similarly, the siderophores pyoverdine and pyochelin also contribute to the ability of P. aeruginosa to protect S. aureus from vancomycin, as did growth under anoxia. Under our experimental conditions, HQNO, P. aeruginosa supernatant, and growth under anoxia decreased S. aureus growth, likely explaining why this cell wall-targeting antibiotic is less effective. P. aeruginosa supernatant did not confer additional protection to slow-growing S. aureus small colony variants. Importantly, P. aeruginosa supernatant protects S. aureus from other inhibitors of cell wall synthesis as well as protein synthesis-targeting antibiotics in an HQNO- and siderophore-dependent manner. We propose a model whereby P. aeruginosa causes S. aureus to shift to fermentative growth when these organisms are grown in coculture, leading to reduction in S. aureus growth and decreased susceptibility to antibiotics targeting cell wall and protein synthesis.

  11. Enterobactin-mediated iron transport in Pseudomonas aeruginosa.

    Science.gov (United States)

    Poole, K; Young, L; Neshat, S

    1990-01-01

    A pyoverdine-deficient strain of Pseudomonas aeruginosa was unable to grow in an iron-deficient minimal medium in the presence of the nonmetabolizable iron chelator ethylene diamine-di(omega-hydroxyphenol acetic acid) (EDDHA), although addition of enterobactin to EDDHA-containing minimal media did restore growth of the pyoverdine-deficient P. aeruginosa. Consistent with the apparent ability of enterobactin to provide iron to P. aeruginosa, enterobactin-dependent 55Fe3+ uptake was observed in cells of P. aeruginosa previously grown in an iron-deficient medium containing enterobactin (or enterobactin-containing Escherichia coli culture supernatant). This uptake was energy dependent, was observable at low concentrations (60 nM) of FeCl3, and was absent in cells cultured without enterobactin. A novel protein with a molecular weight of approximately 80,000 was identified in the outer membranes of cells grown in iron-deficient minimal medium containing enterobactin, concomitant with the induction of enterobactin-dependent iron uptake. A Tn501 insertion mutant lacking this protein was isolated and shown to be deficient in enterobactin-mediated iron transport at 60 nM FeCl3, although it still exhibited enterobactin-dependent growth in iron-deficient medium containing EDDHA. It was subsequently observed that the mutant was, however, capable of enterobactin-mediated iron transport at much higher concentrations (600 nM) of FeCl3. Indeed, enterobactin-dependent iron uptake at this concentration of iron was observed in both the mutant and parent strains irrespective of whether they had been cultured in the presence of enterobactin.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:2174865

  12. Effects of ambroxol on biofilm adhesion and viability of Pseudomonas aeruginosa quorum sensing defective strain

    Directory of Open Access Journals (Sweden)

    Qi LU

    2013-07-01

    Full Text Available Objective To investigate the effects of ambroxol on the biofilm viability and pristine adhesion of Pseudomonas aeruginosa wild (PAO1 and quorum sensing defective strain (QS, gene deletion of ∆lasI and ∆rhlI. Methods The biofilm was treated by different concentrations (0, 1.875, 3.75mg/ml of ambroxol. The number of colony was measured with agar plate, multifunction fluorometer was used to measure the fluorescence intensity of PAO1 and QS strains at the bottom of 96-well plate. The adhesion ratio (% was calculated to determine the effects of ambroxol on bacterial biofilm adhesion. Results Ambroxol treatment reduced the survival rate of the mutant strains compared to that of wild strain, even though the QS strain had increased the adhesion in the presence of ambroxol compared to that of wild strain (P<0.05. Conclusion Ambroxol has a property of significantly antagonizing quorum-sensing system, suggesting that it might be of importance in treatment against chronic Pseudomonasaeruginosainfections.

  13. Mitigation of a nitrate reducing Pseudomonas aeruginosa biofilm and anaerobic biocorrosion using ciprofloxacin enhanced by D-tyrosine.

    Science.gov (United States)

    Jia, Ru; Yang, Dongqing; Xu, Dake; Gu, Tingyue

    2017-07-31

    Pseudomonas aeruginosa (PA) is a ubiquitous microbe. It can form recalcitrant biofilms in clinical and industrial settings. PA biofilms cause infections in patients. They also cause biocorrosion of medical implants. In this work, D-tyrosine (D-tyr) was investigated as an antimicrobial enhancer for ciprofloxacin (CIP) against a wild-type PA biofilm (strain PAO1) on C1018 carbon steel in a strictly anaerobic condition. Seven-day biofilm prevention test results demonstrated that 2 ppm (w/w) D-tyr enhanced 30 ppm CIP by achieving extra 2-log sessile cell reduction compared with the 30 ppm CIP alone treatment. The cocktail of 30 ppm CIP + 2 ppm D-tyr achieved similar efficacy as the 80 ppm CIP alone treatment in the biofilm prevention test. Results also indicated that the enhanced antimicrobial treatment reduced weight loss and pitting corrosion. In the 3-hour biofilm removal test, the cocktail of 80 ppm CIP + 5 ppm D-tyr achieved extra 1.5-log reduction in sessile cell count compared with the 80 ppm CIP alone treatment. The cocktail of 80 ppm CIP + 5 ppm D-tyr achieved better efficacy than the 150 ppm CIP alone treatment in the biofilm removal test.

  14. Impact of a Novel, Anti-microbial Dressing on In Vivo, Pseudomonas aeruginosa Wound Biofilm: Quantitative Comparative Analysis using a Rabbit Ear Model

    Science.gov (United States)

    2014-12-01

    therapies such as debridement , lavage, and antimicrobials, but with little evidence that they improve chronic wound healing in a quantitative and... TYPE N/A 3. DATES COVERED - 4. TITLE AND SUBTITLE Impact of a novel, anti-microbial dressing on in vivo, Pseudomonas aeruginosa wound biofilm...study. Bacterial strains and culture Wild- type strains of P. aeruginosa (obtained from the labora- tory of Dr. Barbara H. Iglewski, University of

  15. The Small RNA ErsA of Pseudomonas aeruginosa Contributes to Biofilm Development and Motility through Post-transcriptional Modulation of AmrZ

    DEFF Research Database (Denmark)

    Falcone, Marilena; Ferrara, Silvia; Rossi, Elio

    2018-01-01

    . In this study, we show that a knock-out ersA mutant strain forms a flat and uniform biofilm, not characterized by mushroom-multicellular structures typical of a mature biofilm. Conversely, the knock-out mutant strain showed enhanced swarming and twitching motilities. To assess the influence of ErsA on the P....... aeruginosa transcriptome, we performed RNA-seq experiments comparing the knock-out mutant with the wild-type. More than 160 genes were found differentially expressed in the knock-out mutant. Parts of these genes, important for biofilm formation and motility regulation, are known to belong also to the Amr...

  16. Real-Time Monitoring of nfxB Mutant Occurrence and Dynamics in Pseudomonas aeruginosa Biofilm Exposed to Subinhibitory Concentrations of Ciprofloxacin

    DEFF Research Database (Denmark)

    Zaborskytė, Greta; Andersen, Jens Bo; Kragh, Kasper Nørskov

    2017-01-01

    Biofilm infections caused by Pseudomonas aeruginosa are frequently treated with ciprofloxacin (CIP); however, resistance rapidly develops. One of the primary resistance mechanisms is the overexpression of the MexCD-OprJ pump due to a mutation in nfxB, encoding the transcriptional repressor...

  17. In vitro anti-pseudomonal potential of Juglans regia and Otostegia limbata leaves extract against planktonic and biofilm form of Pseudomonas aeruginosa

    International Nuclear Information System (INIS)

    Khan, I.; Khan, K.; Nawaz, M.A.; Khan, U.

    2018-01-01

    In the present study the anti-pseudomonal potential of crude methanolic extracts, hexane, ethyl acetate and water fractions of the leaves of Juglans regia L. and Otostegia limbata (Benth.) Boiss.against planktonic and biofilm form of clinical strains (P1, P2 and P3 strains) of Pseudomonas aeruginosa (P. aeruginosa) were evaluated. Agar well diffusion and minimum inhibitory concentration (MIC) assays were used against planktonic, whereas pellicle inhibition and static biofilm inhibition assays were performed against biofilm form of P. aeruginosa. In well diffusion assay, the crude methanolic extract of J. regia showed good bacterial inhibition than O. limbata. The J. regiacrude methanol extract had significant (+; complete breakage of pellicle layer), good (++; partial breakage of pellicle layer) and moderate (+++; uniformthin layer of pellicle formation) pellicle inhibition activity, while O. limbata had moderate (+++; uniformthin layer of pellicle formation) to weak (++++; loose thick layer pellicle formation) pellicle inhibition effect. In MIC assays, hexane and water fractions of J. regia had high (86 vs. 77%) antibacterial activity, while crude methanolic extract of O. limbata showed 51% inhibition against the most resistant P3 strain at 1000 mu g/ml concentration. In static antibiofilm assay, hexane fraction of J. regia had high (63%) inhibition compared to crude methanolic extract of O. limbata(31%) against P3 strain. The present study highlights that J. regia extracts possesses high anti-pseudomonal properties as compared to O. limbata. (author)

  18. The in vitro effect of xylitol on chronic rhinosinusitis biofilms.

    Science.gov (United States)

    Jain, R; Lee, T; Hardcastle, T; Biswas, K; Radcliff, F; Douglas, R

    2016-12-01

    Biofilms have been implicated in chronic rhinosinusitis (CRS) and may explain the limited efficacy of antibiotics. There is a need to find more effective, non-antibiotic based therapies for CRS. This study examines the effects of xylitol on CRS biofilms and planktonic bacteria. Crystal violet assay and spectrophotometry were used to quantify the effects of xylitol (5% and 10% solutions) against Staphylococcus epidermidis, Pseudomonas aeruginosa, and Staphylococcus aureus. The disruption of established biofilms, inhibition of biofilm formation and effects on planktonic bacteria growth were investigated and compared to saline and no treatment. Xylitol 5% and 10% significantly reduced biofilm biomass (S. epidermidis), inhibited biofilm formation (S. aureus and P. aeruginosa) and reduced growth of planktonic bacteria (S. epidermidis, S. aureus, and P. aeruginosa). Xylitol 5% inhibited formation of S. epidermidis biofilms more effectively than xylitol 10%. Xylitol 10% reduced S. epidermidis planktonic bacteria more effectively than xylitol 5%. Saline, xylitol 5% and 10% disrupted established biofilms of S. aureus when compared with no treatment. No solution was effective against established P. aeruginosa biofilm. Xylitol has variable activity against biofilms and planktonic bacteria in vitro and may have therapeutic efficacy in the management of CRS.

  19. A Novel Insight into Dehydroleucodine Mediated Attenuation of Pseudomonas aeruginosa Virulence Mechanism

    Directory of Open Access Journals (Sweden)

    S. Mustafi

    2015-01-01

    Full Text Available Increasing resistance of Pseudomonas aeruginosa (P. aeruginosa to conventional treatments demands the search for novel therapeutic strategies. In this study, the antimicrobial activity of dehydroleucodine (DhL, a sesquiterpene lactone obtained from Artemisia (A. douglasiana, was screened against several pathogenic virulence effectors of P. aeruginosa. In vitro, minimum inhibitory concentration of DhL was determined against P. aeruginosa strains PAO1, PA103, PA14, and multidrug resistant clinical strain, CDN118. Results showed that DhL was active against each strain where PAO1 and PA103 showed higher susceptibility (MIC 0.48 mg/mL as compared to PA14 (MIC 0.96 mg/mL and CDN118 (MIC 0.98 mg/mL. Also, when PAO1 strain was grown in the presence of DhL (MIC50, 0.12 mg/mL, a delay in the generation time was noticed along with significant inhibition of secretory protease and elastase activities, interruption in biofilm attachment phase in a stationary culture, and a significant decline in Type III effector ExoS. At MIC50, DhL treatment increased the sensitivity of P. aeruginosa towards potent antibiotics. Furthermore, treatment of P. aeruginosa with DhL prevented toxin-induced apoptosis in macrophages. These observations suggest that DhL activity was at the bacterial transcriptional level. Hence, antimicrobial activity of DhL may serve as leads in the development of new anti-Pseudomonas pharmaceuticals.

  20. Mitophagy confers resistance to siderophore-mediated killing by Pseudomonas aeruginosa.

    Science.gov (United States)

    Kirienko, Natalia V; Ausubel, Frederick M; Ruvkun, Gary

    2015-02-10

    In the arms race of bacterial pathogenesis, bacteria produce an array of toxins and virulence factors that disrupt core host processes. Hosts mitigate the ensuing damage by responding with immune countermeasures. The iron-binding siderophore pyoverdin is a key virulence mediator of the human pathogen Pseudomonas aeruginosa, but its pathogenic mechanism has not been established. Here we demonstrate that pyoverdin enters Caenorhabditis elegans and that it is sufficient to mediate host killing. Moreover, we show that iron chelation disrupts mitochondrial homeostasis and triggers mitophagy both in C. elegans and mammalian cells. Finally, we show that mitophagy provides protection both against the extracellular pathogen P. aeruginosa and to treatment with a xenobiotic chelator, phenanthroline, in C. elegans. Although autophagic machinery has been shown to target intracellular bacteria for degradation (a process known as xenophagy), our report establishes a role for authentic mitochondrial autophagy in the innate immune defense against P. aeruginosa.

  1. A peptide from human β thymosin as a platform for the development of new anti-biofilm agents for Staphylococcus spp. and Pseudomonas aeruginosa.

    Science.gov (United States)

    Schillaci, Domenico; Spinello, Angelo; Cusimano, Maria Grazia; Cascioferro, Stella; Barone, Giampaolo; Vitale, Maria; Arizza, Vincenzo

    2016-08-01

    Conventional antibiotics might fail in the treatment of biofilm-associated infections causing infection recurrence and chronicity. The search for antimicrobial peptides has been performed with the aim to discover novel anti-infective agents active on pathogens in both planktonic and biofilm associated forms. The fragment 9-19 of human thymosin β4 was studied through 1 μs MD simulation. Two main conformations of the peptide were detected, both constituted by a central hydrophobic core and by the presence of peripheral charged residues suggesting a possible mechanism of interaction with two models of biological membranes, related to eukaryotic or bacterial membrane respectively. In addition, the peptide was chemically synthesized and its antimicrobial activity was tested in vitro against planktonic and biofilm form of a group of reference strains of Staphylococcus spp. and one P. aeruginosa strain. The human thymosin β4 fragment EIEKFDKSKLK showed antibacterial activity against staphylococcal strains and Pseudomonas aeruginosa ATCC 15442 at concentrations from 12.5 to 6.2 mg/ml and inhibited biofilm formation at sub-inhibitory concentrations (3.1-0.75 mg/ml). The activity of the fragment in inhibiting biofilm formation, could be due to the conformations highlighted by the MD simulations, suggesting its interaction with the bacterial membrane. Human thymosin β4 fragment can be considered a promising lead compound to develop novel synthetic or recombinant derivatives with improved pharmaceutical potential.

  2. Immune mediators of sea-cucumber Holothuria tubulosa (Echinodermata) as source of novel antimicrobial and anti-staphylococcal biofilm agents.

    Science.gov (United States)

    Schillaci, Domenico; Cusimano, Maria Grazia; Cunsolo, Vincenzo; Saletti, Rosaria; Russo, Debora; Vazzana, Mirella; Vitale, Maria; Arizza, Vincenzo

    2013-06-24

    The present study aims to investigate coelomocytes, immune mediators cells in the echinoderm Holothuria tubulosa, as an unusual source of antimicrobial and antibiofilm agents. The activity of the 5kDa peptide fraction of the cytosol from H. tubulosa coelomocytes (5-HCC) was tested against a reference group of Gram-negative and Gram-positive human pathogens. Minimal inhibitory concentrations (MICs) ranging from 125 to 500 mg/ml were determined against tested strains. The observed biological activity of 5-HCC could be due to two novel peptides, identified by capillary RP-HPLC/nESI-MS/MS, which present the common chemical-physical characteristics of antimicrobial peptides. Such peptides were chemically synthesized and their antimicrobial activity was tested. The synthetic peptides showed broad-spectrum activity at 12.5 mg/ml against the majority of the tested Gram-positive and Gram-negative strains, and they were also able to inhibit biofilm formation in a significant percentage at a concentration of 3.1 mg/ml against staphylococcal and Pseudomonas aeruginosa strains.The immune mediators in H. tubulosa are a source of novel antimicrobial peptides for the development of new agents against biofilm bacterial communities that are often intrinsically resistant to conventional antibiotics.

  3. Pattern formation in Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Parsek, Matthew R.; Tolker-Nielsen, Tim

    2008-01-01

    Bacteria are capable of forming elaborate multicellular communities called biofilms. Pattern formation in biofilms depends on cell proliferation and cellular migration in response to the available nutrients and other external cues, as well as on self-generated intercellular signal molecules...... and the production of an extracellular matrix that serves as a structural 'scaffolding' for the biofilm cells. Pattern formation in biofilms allows cells to position themselves favorably within nutrient gradients and enables buildup and maintenance of physiologically distinct subpopulations, which facilitates...... survival of one or more subpopulations upon environmental insult, and therefore plays an important role in the innate tolerance displayed by biofilms toward adverse conditions....

  4. Bacterial biofilm formation in different surfaces of food industries

    Directory of Open Access Journals (Sweden)

    Karine Angélica Dalla Costa

    2017-06-01

    Full Text Available The term biofilm describes the sessile microbial life form, characterized by microorganism adhesion to any surface and with the production of extracellular polymeric substances. In food industries, the formation of biofilms results in serious problems, since it can be a contamination source of the food product, compromising the final product quality and consumer health. The aim of this study was to verify the adhesion of biofilms (sessile cells of pathogenic and/or deteriorating bacteria against surfaces of the food industry. The bacterial species tested were Pseudomonas aeruginosa ATCC 27853, Staphylococcus aureus ATCC 29213, Listeria monocytogenes ATCC 19117 and Salmonella Typhimurium ATCC 14028. It was used stainless steel and polypropylene coupons as contact surfaces. The results demonstrated that P. aeruginosa and S. Typhimurium showed higher biofilm formation capacity. Statistically, there was no difference in count of P. aeruginosa and S. Typhimurium (p > 0.05 cells. The same occurred between L. monocytogenes and S. aureus. However, the counts of P. aeruginosa and S. Typhimurium cells were statistically higher than S. aureus and L. monocytogenes (p < 0.05. By means of scanning electron microscopy it was also found increased adhesion of P. aeruginosa. The results revealed that P. aeruginosa was the bacterial species with higher biofilm formation capacity among the others.

  5. Scaffold of Selenium Nanovectors and Honey Phytochemicals for Inhibition of Pseudomonas aeruginosa Quorum Sensing and Biofilm Formation

    Directory of Open Access Journals (Sweden)

    Prateeksha

    2017-03-01

    Full Text Available Honey is an excellent source of polyphenolic compounds that are effective in attenuating quorum sensing (QS, a chemical process of cell-to-cell communication system used by the opportunistic pathogen Pseudomonas aeruginosa to regulate virulence and biofilm formation. However, lower water solubility and inadequate bioavailability remains major concerns of these therapeutic polyphenols. Its therapeutic index can be improved by using nano-carrier systems to target QS signaling potently. In the present study, we fabricated a unique drug delivery system comprising selenium nanoparticles (SeNPs; non-viral vectors and polyphenols of honey (HP for enhancement of anti-QS activity of HP against P. aeruginosa PAO1. The developed selenium nano-scaffold showed superior anti-QS activity, anti-biofilm efficacy, and anti-virulence potential in both in-vitro and in-vivo over its individual components, SeNPs and HP. LasR is inhibited by selenium nano-scaffold in-vitro. Using computational molecular docking studies, we have also demonstrated that the anti-virulence activity of selenium nano-scaffold is reliant on molecular binding that occurs between HP and the QS receptor LasR through hydrogen bonding and hydrophobic interactions. Our preliminary investigations with selenium-based nano-carriers hold significant promise to improve anti-virulence effectiveness of phytochemicals by enhancing effective intracellular delivery.

  6. Pseudomonas aeruginosa in Swimming Pool Water: Evidences and Perspectives for a New Control Strategy

    Directory of Open Access Journals (Sweden)

    Marco Guida

    2016-09-01

    Full Text Available Pseudomonas aeruginosa is frequently isolated in swimming pool settings. Nine recreational and rehabilitative swimming pools were monitored according to the local legislation. The presence of P. aeruginosa was correlated to chlorine concentration. The ability of the isolates to form a biofilm on plastic materials was also investigated. In 59.5% of the samples, microbial contamination exceeded the threshold values. P. aeruginosa was isolated in 50.8% of these samples. The presence of P. aeruginosa was not correlated with free or total chlorine amount (R2 < 0.1. All the isolates were moderate- to strong-forming biofilm (Optical Density O.D.570 range 0.7–1.2. To control biofilm formation and P. aeruginosa colonization, Quantum FreeBioEnergy© (QFBE, FreeBioEnergy, Brisighella, Italy, has been applied with encouraging preliminary results. It is a new, promising control strategy based on the change of an electromagnetic field which is responsible for the proliferation of some microorganisms involved in biofilm formation, such as P. aeruginosa.

  7. Formação de biofilme por Pseudomonas aeruginosa sobre aço inoxidável em contato com leite e seu controle por óleos essenciais

    Directory of Open Access Journals (Sweden)

    Nádia Nara BATISTA

    2014-01-01

    Full Text Available Objetivo: Avaliar a ação bacteriostática e bactericida de diferentes óleos essenciais sobre células planctônicas de Pseudomonas aeruginosa ATCC 27853, bem como verificar a ação sanitizante, dos óleos essenciais que apresentarem a menor Concentração Mínima Inibitória (CMI, sobre o biofilme formado por esta espécie, Material e Métodos: A ação bacteriostática foi realizada por meio da determinação das CMIs dos óleos de Zingiber officinale, Eugenia caryophyllus, Elettaria cardamomum, Citrus limon e Citrus reticulata v, tangerine, O tempo de morte bacteriana foi determinado utilizando-se as CMIs de cada óleo essencial submetidos a diferentes tempos de contato, O biofilme de P, aeruginosa foi desenvolvido em cupons de aço inoxidável AISI 304 dispostos em placa de Petri contendo leite tratado por Ultra Alta Temperatura (UAT, sendo incubado sob agitação de 70 rpm, a 37 °C/96 horas, Células aderidas foram removidas através de swabs e enumeradas por contagem em placas após submissão a diferentes tratamentos, Resultados: Todos os óleos essenciais apresentaram efeito bacteriostático, se destacando Z, officinale, E, caryophyllus e E, cardamomum, por apresentarem menor CMI, O tempo de morte de P, aeruginosa foi de 10 minutos quando utilizadas soluções a base de E, cardamomum e E, caryophyllus, No entanto, quando testados em biofilme, apenas E, caryophyllus eliminou as células bacterianas viáveis de P, aeruginosa, Conclusão: E, caryophyllus é uma nova alternativa para o controle do biofilme de P, aeruginosa na indústria de alimentos, pois, além de sua alta atividade antimicrobiana, é um composto natural, o que atende as exigências do mercado consumidor.

  8. Studies on the biofilm produced by Pseudomonas aeruginosa grown in different metal fatty acid salt media and its application in biodegradation of fatty acids and bioremediation of heavy metal ions.

    Science.gov (United States)

    Abinaya Sindu, P; Gautam, Pennathur

    2017-01-01

    Metal fatty acid salts (MFAS) in untreated industrial effluents cause environmental pollution. The use of biocompatible agents for remediation may help in reducing the harm caused to the ambient aquatic organism. Pseudomonas aeruginosa is a ubiquitous organism that thrives under harsh conditions and is resistant to toxic metal ions. The present study shows a proof-of-concept of using this organism in the biodegradation of MFAS. MFAS were prepared and we studied their effect on the growth of the planktonic form and the formation of biofilm by P. aeruginosa. We observed biofilm formation in the presence of all the MFAS when used as the sole carbon source, albeit the quantity of biofilm formed in the presence of cadmium and copper was less. There was no effect on the planktonic form of the organism but the formation of biofilm increased in the presence of magnesium palmitate. This study shows that metal ions play a pivotal role in the formation of biofilm. HPLC (high-performance liquid chromatography) analysis of the biofilm polysaccharide showed that hexose sugar was a major component when compared with pentose sugar. The structure of biofilm polysaccharide and the coordination of the metal ion with the biofilm polysaccharide were confirmed by FTIR (Fourier transform infrared spectroscopy) and Raman spectroscopy.

  9. A novel technique using potassium permanganate and reflectance confocal microscopy to image biofilm extracellular polymeric matrix reveals non-eDNA networks in Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Swearingen, Matthew C; Mehta, Ajeet; Mehta, Amar; Nistico, Laura; Hill, Preston J; Falzarano, Anthony R; Wozniak, Daniel J; Hall-Stoodley, Luanne; Stoodley, Paul

    2016-02-01

    Biofilms are etiologically important in the development of chronic medical and dental infections. The biofilm extracellular polymeric substance (EPS) determines biofilm structure and allows bacteria in biofilms to adapt to changes in mechanical loads such as fluid shear. However, EPS components are difficult to visualize microscopically because of their low density and molecular complexity. Here, we tested potassium permanganate, KMnO4, for use as a non-specific EPS contrast-enhancing stain using confocal laser scanning microscopy in reflectance mode. We demonstrate that KMnO4 reacted with EPS components of various strains of Pseudomonas, Staphylococcus and Streptococcus, yielding brown MnO2 precipitate deposition on the EPS, which was quantifiable using data from the laser reflection detector. Furthermore, the MnO2 signal could be quantified in combination with fluorescent nucleic acid staining. COMSTAT image analysis indicated that KMnO4 staining increased the estimated biovolume over that determined by nucleic acid staining alone for all strains tested, and revealed non-eDNA EPS networks in Pseudomonas aeruginosa biofilm. In vitro and in vivo testing indicated that KMnO4 reacted with poly-N-acetylglucosamine and Pseudomonas Pel polysaccharide, but did not react strongly with DNA or alginate. KMnO4 staining may have application as a research tool and for diagnostic potential for biofilms in clinical samples. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  10. Deconvoluting the effects of surface chemistry and nanoscale topography: Pseudomonas aeruginosa biofilm nucleation on Si-based substrates.

    Science.gov (United States)

    Zhang, Jing; Huang, Jinglin; Say, Carmen; Dorit, Robert L; Queeney, K T

    2018-06-01

    The nucleation of biofilms is known to be affected by both the chemistry and topography of the underlying substrate, particularly when topography includes nanoscale (topography vs. chemistry is complicated by concomitant variation in both as a result of typical surface modification techniques. Analyzing the behavior of biofilm-forming bacteria exposed to surfaces with systematic, independent variation of both topography and surface chemistry should allow differentiation of the two effects. Silicon surfaces with reproducible nanotopography were created by anisotropic etching in deoxygenated water. Surface chemistry was varied independently to create hydrophilic (OH-terminated) and hydrophobic (alkyl-terminated) surfaces. The attachment and proliferation of Psuedomonas aeruginosa to these surfaces was characterized over a period of 12 h using fluorescence and confocal microscopy. The number of attached bacteria as well as the structural characteristics of the nucleating biofilm were influenced by both surface nanotopography and surface chemistry. In general terms, the presence of both nanoscale features and hydrophobic surface chemistry enhance bacterial attachment and colonization. However, the structural details of the resulting biofilms suggest that surface chemistry and topography interact differently on each of the four surface types we studied. Copyright © 2018 Elsevier Inc. All rights reserved.

  11. C-di-GMP regulates antimicrobial peptide resistance in Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Chua, Song Lin; Tan, Sean Yang-Yi; Rybtke, Morten Theil

    2013-01-01

    Bis-(3'-5')-cyclic dimeric GMP (c-di-GMP) is an intracellular second messenger which controls the life styles of many bacteria. A high intracellular level of c-di-GMP induces a biofilm lifestyle, whereas a low intracellular level of c-di-GMP stimulates dispersal of biofilms and promotes...... a planktonic lifestyle. Here, we used expression of different reporters to show that planktonic cells (PCells), biofilm cells (BCells) and cells dispersed from biofilms (DCells) had distinct intracellular c-di-GMP levels. Proteomics analysis showed that the low intracellular c-di-GMP level of DCells induced...... the expression of proteins required for the virulence and development of antimicrobial peptide resistance in P. aeruginosa. In accordance, P. aeruginosa cells with low c-di-GMP levels were found to be more resistant to colistin than P. aeruginosa cells with high c-di-GMP levels. This contradicts the current...

  12. Immune Modulating Topical S100A8/A9 Inhibits Growth of Pseudomonas aeruginosa and Mitigates Biofilm Infection in Chronic Wounds

    DEFF Research Database (Denmark)

    Trøstrup, Hannah; Lerche, Christian Johann; Christophersen, Lars

    2017-01-01

    in a murine model and P. aeruginosa growth in vitro. Seventy-six mice, inflicted with a full-thickness burn wound were challenged subcutaneously (s.c.) by 10⁶ colony-forming units (CFUs) of P. aeruginosa biofilm. Mice were subsequently randomized into two treatment groups, one group receiving recombinant...... murine S100A8/A9 and a group of vehicle controls (phosphate-buffered saline, PBS) all treated with s.c. injections daily for up to five days. Wounds were analyzed for quantitative bacteriology and contents of key inflammatory markers. Count of blood polymorphonuclear leukocytes was included. S100A8/A9...

  13. Garlic blocks quorum sensing and promotes rapid clearing of pulmonary Pseudomonas aeruginosa infections

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Jensen, P. Ø.; Rasmussen, Thomas Bovbjerg

    2005-01-01

    The opportunistic human pathogen Pseudomonas aeruginosa is the predominant micro-organism of chronic lung infections in cystic fibrosis patients. P. aeruginosa colonizes the lungs by forming biofilm microcolonies throughout the lung. Quorum sensing (QS) renders the biofilm bacteria highly tolerant......-treated biofilm. Garlic extract was administered as treatment for a mouse pulmonary infection model. Mice were treated with garlic extract or placebo for 7 days, with the initial 2 days being prophylactic before P. aeruginosa was instilled in the left lung of the mice. Bacteriology, mortality, histopathology...... and phagocytosis by PMNs, as well as leading to an improved outcome of pulmonary infections....

  14. Swimming Motility Mediates the Formation of Neutrophil Extracellular Traps Induced by Flagellated Pseudomonas aeruginosa.

    Directory of Open Access Journals (Sweden)

    Madison Floyd

    2016-11-01

    Full Text Available Pseudomonas aeruginosa is an opportunistic pathogen causing severe infections often characterized by robust neutrophilic infiltration. Neutrophils provide the first line of defense against P. aeruginosa. Aside from their defense conferred by phagocytic activity, neutrophils also release neutrophil extracellular traps (NETs to immobilize bacteria. Although NET formation is an important antimicrobial process, the details of its mechanism are largely unknown. The identity of the main components of P. aeruginosa responsible for triggering NET formation is unclear. In this study, our focus was to identify the main bacterial factors mediating NET formation and to gain insight into the underlying mechanism. We found that P. aeruginosa in its exponential growth phase promoted strong NET formation in human neutrophils while its NET-inducing ability dramatically decreased at later stages of bacterial growth. We identified the flagellum as the primary component of P. aeruginosa responsible for inducing NET extrusion as flagellum-deficient bacteria remained seriously impaired in triggering NET formation. Purified P. aeruginosa flagellin, the monomeric component of the flagellum, does not stimulate NET formation in human neutrophils. P. aeruginosa-induced NET formation is independent of the flagellum-sensing receptors TLR5 and NLRC4 in both human and mouse neutrophils. Interestingly, we found that flagellar motility, not flagellum binding to neutrophils per se, mediates NET release induced by flagellated bacteria. Immotile, flagellar motor-deficient bacterial strains producing paralyzed flagella did not induce NET formation. Forced contact between immotile P. aeruginosa and neutrophils restored their NET-inducing ability. Both the motAB and motCD genetic loci encoding flagellar motor genes contribute to maximal NET release; however the motCD genes play a more important role. Phagocytosis of P. aeruginosa and superoxide production by neutrophils were also

  15. Anaerobic Pseudomonas aeruginosa and other obligately anaerobic bacterial biofilms growing in the thick airway mucus of chronically infected cystic fibrosis patients: an emerging paradigm or "Old Hat"?

    Science.gov (United States)

    Su, Shengchang; Hassett, Daniel J

    2012-09-01

    The cystic fibrosis (CF) airway mucus is an ideal niche in which many bacteria can develop antibiotic- and phagocyte-resistance in unique structures known as "mode II biofilms" where bacteria are embedded within the mucus, yet unattached to airway epithelial cells. Pseudomonas aeruginosa is the dominant CF pathogen, yet herein the authors provide burgeoning evidence that obligate anaerobic bacteria (e.g., Prevotella) actually thrive within the CF mucus, a paradigmatic shift that chronic CF is an "aerobic" disease. Interestingly, CF organisms repress virulence factor production (e.g., P. aeruginosa) while others (e.g., S. aureus) increase them under anaerobic conditions. The authors shed additional light on (i) the anoxic nature of the CF airway mucus, (ii) the relative commonality of anaerobic bacteria isolated from CF sputum, (iii) virulence factor production and cross-talk between obligate anaerobes and P. aeruginosa relative to disease progression/remission, (iv) the role of mucoidy in CF, and (v) the role of nitrosative stress in activation of bacteriophage and pyocins within biofilms. The authors conclude with insight as to how we might treat some CF bacteria during mode II biofilm infections that utilizes a metabolite of bacterial anaerobic respiration and an aerobic oxidation product of airway-generated NO, acidified NO(2)(-).

  16. Chronic Pseudomonas aeruginosa biofilm infection impairs murine S100A8/A9 and neutrophil effector cytokines—implications for delayed wound closure?

    DEFF Research Database (Denmark)

    Trøstrup, Hannah; Lerche, Christian Johann; Christophersen, Lars Jackie

    2017-01-01

    The impact of Pseudomonas aeruginosa biofilm infections in chronic wounds and clinical implication for healing is receiving increased attention. However, the pathophysiology of host/pathogen interplay is not fully understood. By further revealing the mechanisms, necessary new treatment strategies...

  17. Acquisition and Role of Molybdate in Pseudomonas aeruginosa

    Science.gov (United States)

    Pederick, Victoria G.; Eijkelkamp, Bart A.; Ween, Miranda P.; Begg, Stephanie L.; Paton, James C.

    2014-01-01

    In microaerophilic or anaerobic environments, Pseudomonas aeruginosa utilizes nitrate reduction for energy production, a process dependent on the availability of the oxyanionic form of molybdenum, molybdate (MoO42−). Here, we show that molybdate acquisition in P. aeruginosa occurs via a high-affinity ATP-binding cassette permease (ModABC). ModA is a cluster D-III solute binding protein capable of interacting with molybdate or tungstate oxyanions. Deletion of the modA gene reduces cellular molybdate concentrations and results in inhibition of anaerobic growth and nitrate reduction. Further, we show that conditions that permit nitrate reduction also cause inhibition of biofilm formation and an alteration in fatty acid composition of P. aeruginosa. Collectively, these data highlight the importance of molybdate for anaerobic growth of P. aeruginosa and reveal novel consequences of nitrate reduction on biofilm formation and cell membrane composition. PMID:25172858

  18. Pseudomonas aeruginosa Psl Exopolysaccharide Interacts with the Antimicrobial Peptide LG21

    Directory of Open Access Journals (Sweden)

    Joyce Seow Fong Chin

    2017-09-01

    Full Text Available Biofilm formation by opportunistic pathogens serves as one of the major causes of chronic and persistent infections. Bacterial cells in the biofilms are embedded in their self-generated protective extracellular polymeric substances (EPS, which include exopolysaccharides, large adhesin proteins and extracellular DNA. In this study, we identified an antimicrobial peptide (AMP LG21 that is able to interact specifically with the Psl exopolysaccharide of Pseudomonas aeruginosa, thus it can be used as a diagnostic tool for P. aeruginosa biofilms. Molecular dynamics simulation analysis showed that residues numbered from 15 to 21 (WKRKRFG in LG21 are involved in interacting with Psl. Our study indicates that host immune systems might detect and interact with microbial biofilms through AMPs. Engineering biofilm EPS-targeting AMPs might provide novel strategies for biofilm detection and treatment.

  19. Microsensor and transcriptomic signatures of oxygen depletion in biofilms associated with chronic wounds: Biofilms and oxygen

    Energy Technology Data Exchange (ETDEWEB)

    James, Garth A. [Center for Biofilm Engineering, Montana State University, Bozeman Montana; Ge Zhao, Alice [Division of Dermatology, Department of Medicine, University of Washington, Seattle Washington; Usui, Marcia [Division of Dermatology, Department of Medicine, University of Washington, Seattle Washington; Underwood, Robert A. [Division of Dermatology, Department of Medicine, University of Washington, Seattle Washington; Nguyen, Hung [The Gene and Linda Voiland School of Chemical Engineering and Bioengineering, Washington State University, Pullman Washington; Beyenal, Haluk [The Gene and Linda Voiland School of Chemical Engineering and Bioengineering, Washington State University, Pullman Washington; deLancey Pulcini, Elinor [Center for Biofilm Engineering, Montana State University, Bozeman Montana; Agostinho Hunt, Alessandra [Department of Microbiology and Molecular Genetics, 5180 Biomedical and Physical Sciences, Michigan State University, East Lansing Michigan; Bernstein, Hans C. [Pacific Northwest National Laboratory, Chemical and Biological Signature Science, Richland Washington; Fleckman, Philip [Division of Dermatology, Department of Medicine, University of Washington, Seattle Washington; Olerud, John [Division of Dermatology, Department of Medicine, University of Washington, Seattle Washington; Williamson, Kerry S. [Center for Biofilm Engineering, Montana State University, Bozeman Montana; Franklin, Michael J. [Center for Biofilm Engineering, Montana State University, Bozeman Montana; Stewart, Philip S. [Center for Biofilm Engineering, Montana State University, Bozeman Montana

    2016-02-16

    Polymicrobial biofilms have been implicated in delayed wound healing, although the mechanisms by which biofilms impair wound healing are poorly understood. Many species of bacteria produce exotoxins and exoenzymes that may inhibit healing. In addition, oxygen consumption by biofilms may impede wound healing. In this study, we used oxygen microsensors to measure oxygen transects through in vitro-cultured biofilms, biofilms formed in vivo in a diabetic (db/db) mouse model, and ex vivo human chronic wound specimens. The results show that oxygen levels within both euthanized and live mouse wounds had steep gradients that reached minima ranging from 19 to 61% oxygen partial pressure, compared to atmospheric oxygen levels. The oxygen gradients in the mouse wounds were similar to those observed for clinical isolates cultured in vitro and for human ex vivo scabs. No oxygen gradients were observed for heat-killed scabs, suggesting that active metabolism by the viable bacteria contributed to the reduced oxygen partial pressure of the wounds. To characterize the metabolic activities of the bacteria in the mouse wounds, we performed transcriptomics analyses of Pseudomonas aeruginosa biofilms associated with the db/db mice wounds using Affymetrix microarrays. The results demonstrated that the bacteria expressed genes for metabolic activities associated with cell growth. Interestingly, the transcriptome results indicated that the bacteria within the wounds also experienced oxygen-limitation stress. Among the bacterial genes that were expressed in vivo were genes associated with the Anr-mediated hypoxia-stress response. Other bacterial stress response genes highly expressed in vivo were genes associated with stationary-phase growth, osmotic stress, and RpoH-mediated heat shock stress. Overall, the results support the hypothesis that the metabolic activities of bacteria in biofilms act as oxygen sinks in chronic wounds and that the depletion of oxygen contributes to the

  20. Recent advances in the treatment of Pseudomonas aeruginosa infections in cystic fibrosis

    DEFF Research Database (Denmark)

    Høiby, Niels

    2011-01-01

    Chronic Pseudomonas aeruginosa lung infection in cystic fibrosis (CF) patients is caused by biofilm-growing mucoid strains. Biofilms can be prevented by early aggressive antibiotic prophylaxis or therapy, and they can be treated by chronic suppressive therapy. New results from one small trial sug...... patients without P. aeruginosa infection did not improve lung function. Here I review the recent advances in the treatment of P. aeruginosa lung infections with a focus on inhalation treatments targeted at prophylaxis and chronic suppressive therapy....

  1. Bacteriophage Infectivity Against Pseudomonas aeruginosa in Saline Conditions

    KAUST Repository

    Scarascia, Giantommaso

    2018-05-02

    Pseudomonas aeruginosa is a ubiquitous member of marine biofilm, and reduces thiosulfate to produce toxic hydrogen sulfide gas. In this study, lytic bacteriophages were isolated and applied to inhibit the growth of P. aeruginosa in planktonic mode at different temperature, pH, and salinity. Bacteriophages showed optimal infectivity at a multiplicity of infection of 10 in saline conditions, and demonstrated lytic abilities over all tested temperature (25, 30, 37, and 45°C) and pH 6–9. Planktonic P. aeruginosa exhibited significantly longer lag phase and lower specific growth rates upon exposure to bacteriophages. Bacteriophages were subsequently applied to P. aeruginosa-enriched biofilm and were determined to lower the relative abundance of Pseudomonas-related taxa from 0.17 to 5.58% in controls to 0.01–0.61% in treated microbial communities. The relative abundance of Alphaproteobacteria, Pseudoalteromonas, and Planococcaceae decreased, possibly due to the phage-induced disruption of the biofilm matrix. Lastly, when applied to mitigate biofouling of ultrafiltration membranes, bacteriophages were determined to reduce the transmembrane pressure increase by 18% when utilized alone, and by 49% when used in combination with citric acid. The combined treatment was more effective compared with the citric acid treatment alone, which reported ca. 30% transmembrane pressure reduction. Collectively, the findings demonstrated that bacteriophages can be used as a biocidal agent to mitigate undesirable P. aeruginosa-associated problems in seawater applications.

  2. Chemical sanitizers to control biofilms formed by two Pseudomonas species on stainless steel surface

    Directory of Open Access Journals (Sweden)

    Danila Soares Caixeta

    2012-03-01

    Full Text Available The biofilm formation of Pseudomonas aeruginosa and Pseudomonas fluorescens on AISI 304 stainless steel in the presence of reconstituted skim milk under different temperatures was conducted, and the potential of three chemical sanitizers in removing the mono-species biofilms formed was compared. Pseudomonas aeruginosa cultivated in skim milk at 28 °C presented better growth rate (10.4 log CFU.mL-1 when compared with 3.7 and 4.2 log CFU.mL-1 for P. aeruginosa and P. fluorescens cultivated at 7 °C, respectively. Pseudomonas aeruginosa formed biofilm when cultivated at 28 °C. However, only the adhesion of P. aeruginosa and P. fluorescens was observed when incubated at 7 °C. The sodium dichloroisocyanurate was the most efficient sanitizer in the reduction of the adhered P. aeruginosa cells at 7 and 28 °C and those on the biofilm, respectively. The hydrogen peroxide was more effective in the reduction of adhered cells of P. fluorescens at 7 °C.

  3. Decontamination Efficacy of Ultraviolet Radiation against Biofilms of Common Nosocomial Bacteria.

    Science.gov (United States)

    Tingpej, Pholawat; Tiengtip, Rattana; Kondo, Sumalee

    2015-06-01

    Ultraviolet radiation (UV) is commonly used to destroy microorganisms in the health-care environment. However, the efficacy of UV radiation against bacteria growing within biofilms has never been studied. To measure the sterilization effectiveness of UV radiation against common healthcare associated pathogens growing within biofilms. Staphylococcus aureus, Methicillin-resistant S. aureus (MRSA), Streptococcus epidermidis, Escherichia coli, ESBL-producing E. coli, Pseudomonas aeruginosa and Acinetobacter baumannii were cultivated in the Calgary Biofilm Device. Their biofilms were placed 50 cm from the UV lamp within the Biosafety Cabinet. Viability test, crystal violet assay and a scanning electron microscope were used to evaluate the germicidal efficacy. Within 5 minutes, UV radiation could kill S. aureus, MRSA, S. epidermidis, A. baumannii and ESBL-producing E. coli completely while it required 20 minutes and 30 minutes respectively to kill E. coli and P. aeruginosa. However, the amounts of biomass and the ultrastructure between UV-exposed biofilms and controls were not significantly different. UV radiation is effective in inactivating nosocomial pathogens grown within biofilms, but not removing biofilms and EPS. The biofilm of P. aeruginosa was the most durable.

  4. Plant flavones enhance antimicrobial activity of respiratory epithelial cell secretions against Pseudomonas aeruginosa.

    Directory of Open Access Journals (Sweden)

    Benjamin M Hariri

    Full Text Available Flavones are a class of natural plant secondary metabolites that have anti-inflammatory and anti-bacterial effects. Some flavones also activate the T2R14 bitter taste receptor, which is expressed in motile cilia of the sinonasal epithelium and activates innate immune nitric oxide (NO production. Flavones may thus be potential therapeutics for respiratory infections. Our objective was to examine the anti-microbial effects of flavones on the common sinonasal pathogens Candida albicans, Staphylococcus aureus, and Pseudomonas aeruginosa, evaluating both planktonic and biofilm growth. Flavones had only very low-level antibacterial activity alone. They did not reduce biofilm formation, but did reduce production of the important P. aeruginosa inflammatory mediator and ciliotoxin pyocyanin. However, flavones exhibited synergy against P. aeruginosa in the presence of antibiotics or recombinant human lysozyme. They also enhanced the efficacy of antimicrobials secreted by cultured and primary human airway cells grown at air-liquid interface. This suggests that flavones may have anti-gram-negative potential as topical therapeutics when combined with antibiotics or in the context of innate antimicrobials secreted by the respiratory or other epithelia. This may have an additive effect when combined with T2R14-activated NO production. Additional studies are necessary to understand which flavone compounds or mixtures are the most efficacious.

  5. In Vitro Evaluation of Biofilm Dispersal as a Therapeutic Strategy To Restore Antimicrobial Efficacy

    DEFF Research Database (Denmark)

    Roizman, Dan; Vidaillac, Celine; Givskov, Michael

    2017-01-01

    As a proof-of-concept study, the direct impact of biofilm dispersal on the in vitro efficacy of imipenem and tobramycin was evaluated against 3-day-old biofilms of Pseudomonas aeruginosa. Arabinose induction of biofilm dispersal via activation of the phosphodiesterase YhjH in the P. aeruginosa en...

  6. Regulation of Initial Attachment of P. aeruginosa

    Science.gov (United States)

    2010-12-08

    shown to play a role in promoting biofilm formation in diverse P. aeruginosa strains (Friedman and Kolter , 2004; Jackson et al., 2004; Ryder et al...34House of Biofilm Cells". J. Bacteriol. 189: 7945-7947. Friedman, L., and Kolter , R. (2004) Genes involved in matrix formation in Pseudomonas

  7. The Diguanylate Cyclase HsbD Intersects with the HptB Regulatory Cascade to Control Pseudomonas aeruginosa Biofilm and Motility.

    Directory of Open Access Journals (Sweden)

    Martina Valentini

    2016-10-01

    Full Text Available The molecular basis of second messenger signaling relies on an array of proteins that synthesize, degrade or bind the molecule to produce coherent functional outputs. Cyclic di-GMP (c-di-GMP has emerged as a eubacterial nucleotide second messenger regulating a plethora of key behaviors, like the transition from planktonic cells to biofilm communities. The striking multiplicity of c-di-GMP control modules and regulated cellular functions raised the question of signaling specificity. Are c-di-GMP signaling routes exclusively dependent on a central hub or can they be locally administrated? In this study, we show an example of how c-di-GMP signaling gains output specificity in Pseudomonas aeruginosa. We observed the occurrence in P. aeruginosa of a c-di-GMP synthase gene, hsbD, in the proximity of the hptB and flagellar genes cluster. We show that the HptB pathway controls biofilm formation and motility by involving both HsbD and the anti-anti-sigma factor HsbA. The rewiring of c-di-GMP signaling into the HptB cascade relies on the original interaction between HsbD and HsbA and on the control of HsbD dynamic localization at the cell poles.

  8. Statistical Analysis of Pseudomonas aeruginosa Biofilm Development: Impact of Mutations in Genes Involved in Twitching Motility, Cell-to-Cell Signaling, and Stationary-Phase Sigma Factor Expression

    DEFF Research Database (Denmark)

    Heydorn, Arne; Ersbøll, Bjarne Kjær; Kato, Junichi

    2002-01-01

    of variance model comprising the four P. aeruginosa strains, five time points (55, 98, 146, 242, and 314 h), and three independent rounds of biofllm experiments. The results showed that the wild type, the DeltapilHIJK mutant, and the rpoS mutant display conspicuously different types of temporal biofilm...... development, whereas the lasI mutant was indistinguishable from the wild type at all time points. The wild type and the lasI mutant formed uniform, densely packed biofilms. The rpoS mutant formed densely packed biofilms that were significantly thicker than those of the wild type, whereas the Deltapil...

  9. The immune system vs. Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Jensen, Peter Østrup; Givskov, Michael; Bjarnsholt, Thomas

    2010-01-01

    Ilya Metchnikoff and Paul Ehrlich were awarded the Nobel price in 1908. Since then, numerous studies have unraveled a multitude of mechanistically different immune responses to intruding microorganisms. However, in the vast majority of these studies, the underlying infectious agents have appeared...... in the planktonic state. Accordingly, much less is known about the immune responses to the presence of biofilm-based infections (which is probably also due to the relatively short period of time in which the immune response to biofilms has been studied). Nevertheless, more recent in vivo and in vitro studies have...... revealed both innate as well as adaptive immune responses to biofilms. On the other hand, measures launched by biofilm bacteria to achieve protection against the various immune responses have also been demonstrated. Whether particular immune responses to biofilm infections exist remains to be firmly...

  10. The effects of hyperosmosis or high pH on a dual-species biofilm of Enterococcus faecalis and Pseudomonas aeruginosa : an in vitro study

    NARCIS (Netherlands)

    van der Waal, S. V.; van der Sluis, L. W. M.; Ozok, A. R.; Exterkate, R. A. M.; van Marle, J.; Wesselink, P. R.; de Soet, J. J.

    2011-01-01

    van der Waal SV, van der Sluis LWM, Ozok AR, Exterkate RAM, van Marle J, Wesselink PR, de Soet JJ. The effects of hyperosmosis or high pH on a dual-species biofilm of Enterococcus faecalis and Pseudomonas aeruginosa: an in vitro study. International Endodontic Journal, 44, 11101117, 2011. Aim To

  11. Antimicrobial Tolerance of Pseudomonas aeruginosa Biofilms Is Activated during an Early Developmental Stage and Requires the Two-Component Hybrid SagS

    Science.gov (United States)

    Gupta, Kajal; Marques, Cláudia N. H.; Petrova, Olga E.

    2013-01-01

    A hallmark characteristic of biofilms is their extraordinary tolerance to antimicrobial agents. While multiple factors are thought to contribute to the high level of antimicrobial tolerance of biofilms, little is known about the timing of induction of biofilm tolerance. Here, we asked when over the course of their development do biofilms gain their tolerance to antimicrobial agents? We demonstrate that in Pseudomonas aeruginosa, biofilm tolerance is linked to biofilm development, with transition to the irreversible attachment stage regulated by the two-component hybrid SagS, marking the timing when biofilms switch to the high-level tolerance phenotype. Inactivation of sagS rendered biofilms but not planktonic cells more susceptible to tobramycin, norfloxacin, and hydrogen peroxide. Moreover, inactivation of sagS also eliminated the recalcitrance of biofilms to killing by bactericidal antimicrobial agents, a phenotype comparable to that observed upon inactivation of brlR, which encodes a MerR-like transcriptional regulator required for biofilm tolerance. Multicopy expression of brlR in a ΔsagS mutant restored biofilm resistance and recalcitrance to killing by bactericidal antibiotics to wild-type levels. In contrast, expression of sagS did not restore the susceptibility phenotype of ΔbrlR mutant biofilms to wild-type levels, indicating that BrlR functions downstream of SagS. Inactivation of sagS correlated with reduced BrlR levels in biofilms, with the produced BrlR being impaired in binding to the previously described BrlR-activated promoters of the two multidrug efflux pump operons mexAB-oprM and mexEF-oprN. Our findings demonstrate that biofilm tolerance is linked to early biofilm development and SagS, with SagS contributing indirectly to BrlR activation. PMID:23995639

  12. Bioguided Fractionation Shows Cassia alata Extract to Inhibit Staphylococcus epidermidis and Pseudomonas aeruginosa Growth and Biofilm Formation

    Directory of Open Access Journals (Sweden)

    Samuel Takashi Saito

    2012-01-01

    Full Text Available Plant extracts have a long history to be used in folk medicine. Cassia alata extracts are known to exert antibacterial activity but details on compounds and mechanism of action remain poorly explored. We purified and concentrated the aqueous leaf extract of C. alata by reverse phase-solid phase extraction and screened the resulting CaRP extract for antimicrobial activity. CaRP extract exhibited antimicrobial activity for Pseudomonas aeruginosa, Staphylococcus epidermidis, S. aureus, and Bacillus subtilis. CaRP also inhibited biofilm formation of S. epidermidis and P. aeruginosa. Several bacterial growth-inhibiting compounds were detected when CaRP extract was fractionated by TLC chromatography coupled to bioautography agar overlay technique. HPLC chromatography of CaRP extract yielded 20 subfractions that were tested by bioautography for antimicrobial activity against S. aureus and S. epidermidis. Five bioactive fractions were detected and chemically characterized, using high-resolution mass spectrometry (qTOF-MS/MS. Six compounds from four fractions could be characterized as kaempferol, kaempferol-O-diglucoside, kaempferol-O-glucoside, quercetin-O-glucoside, rhein, and danthron. In the Salmonella/microsome assay CaRP showed weak mutagenicity (MI<3 only in strain TA98, pointing to a frameshift mutation activity. These results indicate that C. alata leaf extract contains a minimum of 7 compounds with antimicrobial activity and that these together or as single substance are active in preventing formation of bacterial biofilm, indicating potential for therapeutic applications.

  13. Bioguided Fractionation Shows Cassia alata Extract to Inhibit Staphylococcus epidermidis and Pseudomonas aeruginosa Growth and Biofilm Formation

    Science.gov (United States)

    Saito, Samuel Takashi; Trentin, Danielle da Silva; Macedo, Alexandre José; Pungartnik, Cristina; Gosmann, Grace; Silveira, Jaqueline de Deos; Guecheva, Temenouga Nikolova; Henriques, João Antonio Pêgas; Brendel, Martin

    2012-01-01

    Plant extracts have a long history to be used in folk medicine. Cassia alata extracts are known to exert antibacterial activity but details on compounds and mechanism of action remain poorly explored. We purified and concentrated the aqueous leaf extract of C. alata by reverse phase-solid phase extraction and screened the resulting CaRP extract for antimicrobial activity. CaRP extract exhibited antimicrobial activity for Pseudomonas aeruginosa, Staphylococcus epidermidis, S. aureus, and Bacillus subtilis. CaRP also inhibited biofilm formation of S. epidermidis and P. aeruginosa. Several bacterial growth-inhibiting compounds were detected when CaRP extract was fractionated by TLC chromatography coupled to bioautography agar overlay technique. HPLC chromatography of CaRP extract yielded 20 subfractions that were tested by bioautography for antimicrobial activity against S. aureus and S. epidermidis. Five bioactive fractions were detected and chemically characterized, using high-resolution mass spectrometry (qTOF-MS/MS). Six compounds from four fractions could be characterized as kaempferol, kaempferol-O-diglucoside, kaempferol-O-glucoside, quercetin-O-glucoside, rhein, and danthron. In the Salmonella/microsome assay CaRP showed weak mutagenicity (MI < 3) only in strain TA98, pointing to a frameshift mutation activity. These results indicate that C. alata leaf extract contains a minimum of 7 compounds with antimicrobial activity and that these together or as single substance are active in preventing formation of bacterial biofilm, indicating potential for therapeutic applications. PMID:22548121

  14. DNA-mediated bacterial aggregation is dictated by acid-base interactions

    NARCIS (Netherlands)

    Das, Theerthankar; Krom, Bastiaan P.; van der Mei, Henny C.; Busscher, Henk J.; Sharma, Prashant K.

    2011-01-01

    Extracellular DNA (eDNA) plays a significant role in bacterial biofilm formation and aggregation. Here, for the first time, we present a physico-chemical analysis of the DNA-mediated aggregation for three bacterial strains (Streptococcus mutans LT11, Pseudomonas aeruginosa PAO1 and Staphylococcus

  15. Membrane-anchored MucR mediates nitrate-dependent regulation of alginate production in Pseudomonas aeruginosa

    KAUST Repository

    Wang, Yajie

    2015-04-29

    Alginates exhibit unique material properties suitable for medical and industrial applications. However, if produced by Pseudomonas aeruginosa, it is an important virulence factor in infection of cystic fibrosis patients. The alginate biosynthesis machinery is activated by c-di-GMP imparted by the inner membrane protein, MucR. Here, it was shown that MucR impairs alginate production in response to nitrate in P. aeruginosa. Subsequent site-specific mutagenesis of MucR revealed that the second MHYT sensor motif (MHYT II, amino acids 121–124) of MucR sensor domain was involved in nitrate sensing. We also showed that both c-di-GMP synthesizing and degrading active sites of MucR were important for alginate production. Although nitrate and deletion of MucR impaired alginate promoter activity and global c-di-GMP levels, alginate yields were not directly correlated with alginate promoter activity or c-di-GMP levels, suggesting that nitrate and MucR modulate alginate production at a post-translational level through a localized pool of c-di-GMP. Nitrate increased pel promoter activity in the mucR mutant while in the same mutant the psl promoter activity was independent of nitrate. Nitrate and deletion of mucR did not impact on swarming motility but impaired attachment to solid surfaces. Nitrate and deletion of mucR promoted the formation of biofilms with increased thickness, cell density, and survival. Overall, this study provided insight into the functional role of MucR with respect to nitrate-mediated regulation of alginate biosynthesis. © 2015 Springer-Verlag Berlin Heidelberg

  16. Inhalable Antimicrobials for Treatment of Bacterial Biofilm-Associated Sinusitis in Cystic Fibrosis Patients

    DEFF Research Database (Denmark)

    Klodzinska, Sylvia Natalie; Priemel, Petra Alexandra; Rades, Thomas

    2016-01-01

    Bacterial biofilm-associated chronic sinusitis in cystic fibrosis (CF) patients caused by Pseudomonas aeruginosa infections and the lack of available treatments for such infections constitute a critical aspect of CF disease management. Currently, inhalation therapies to combat P. aeruginosa infec...... and management of biofilm infections caused by P. aeruginosa and discusses critical issues related to novel antimicrobial drug formulation design approaches.......Bacterial biofilm-associated chronic sinusitis in cystic fibrosis (CF) patients caused by Pseudomonas aeruginosa infections and the lack of available treatments for such infections constitute a critical aspect of CF disease management. Currently, inhalation therapies to combat P. aeruginosa....... aeruginosa from the respiratory tract after a first infection has been shown to delay chronic pulmonary infection with the bacteria for up to two years. The challenges with providing a suitable treatment for bacterial sinusitis include: (i) identifying a suitable antimicrobial compound; (ii) selecting...

  17. Biofilm inhibition formation of clinical strains of Pseudomonas aeruginosa mutans, photocatalytic activity of azo dye and GC-MS analysis of leaves of Lagerstroemia speciosa.

    Science.gov (United States)

    Sai Saraswathi, V; Kamarudheen, Neethu; Bhaskara Rao, K V; Santhakumar, K

    2017-04-01

    The investigation was conducted to analyse the bioactive compounds from the leaf extracts of L. speciosa by GC-MS. The extracts were screened for antibacterial and antibiofilm activities against potential clinical strains. The bioactive compounds from the leaves of L. speciosa were extracted by soxhlet continuous extraction method and their chemical composition was analysed by Gas Chromatography-Mass Spectroscopy (GC-MS). The antibacterial activity was evaluated against clinical strain like Staphylococcus aureus, Escherichia coli, P. aeruginosa and Salmonella typhi by well diffusion technique. We also screened for antibacterial property against common food borne pathogens namely Listeria monocytogenes and Bacillus cereus at varied concentration 250μml -1 to 1000μml -1 . Thereafter antibiofilm assay was carried out at from 250 to 1000μg/ml against P. aeruginosa (high biofilm forming pathogen) clinical strain by cover slip technique and the morphology of the pathogen was observed using Scanning Electron Microscopy-(SEM). It was observed that diverse class of secondary metabolites were found by GC-MS analysis for all the extracts upon the continuous extraction. It was found that only minimum inhibition was seen in alcoholic extract for antibacterial activity, whereas all other extracts showed negligible activity. P. aeruginosa biofilm inhibited to 93.0±2% and 91±2% at higher concentration (1000μg/ml) for methanolic and ethanolic extract respectively. Absence of extracellular matrix structure and the surface cracking of biofilm were viewed by SEM, which confirmed the antibiofilm activity. Hence this study reveals that L. speciosa showed significant antibiofilm activity against P. aeruginosa due to the phytoconstituents present in the leaf extracts which was well documented in the alcoholic extracts by GC-MS analysis. The methanolic and ethanolic extract showed good photocatalytic activity of 77.44% and 96.66% against azo dye degradation respectively. Further

  18. Mechanisms of humoral immune response against Pseudomonas aeruginosa biofilm infection in cystic fibrosis

    DEFF Research Database (Denmark)

    Mauch, Renan Marrichi; Jensen, Peter Østrup; Moser, Claus

    2018-01-01

    P. aeruginosa chronic lung infection is the major cause of morbidity and mortality in patients with cystic fibrosis (CF), and is characterized by a biofilm mode of growth, increased levels of specific IgG antibodies and immune complex formation. However, despite being designed to combat...... this infection, such elevated humoral response is not associated with clinical improvement, pointing to a lack of anti-pseudomonas effectiveness. The mode of action of specific antibodies, as well as their structural features, and even the background involving B-cell production, stimulation and differentiation...... into antibody-producing cells in the CF airways are poorly understood. Thus, the aim of this review is to discuss studies that have addressed the intrinsic features of the humoral immune response and provide new insights regarding its insufficiency in the CF context....

  19. Effects of PslG on the surface movement of Pseudomonas aeruginosa.

    Science.gov (United States)

    Zhang, Jingchao; He, Jing; Zhai, Chunhui; Ma, Luyan Z; Gu, Lichuan; Zhao, Kun

    2018-05-04

    PslG attracted a lot of attention recently due to its great potential abilities in inhibiting biofilms of Pseudomonas aeruginosa However, how PslG affects biofilm development still remains largely unexplored. Here, we focused on the surface motility of bacterial cells, which is critical for biofilm development. We studied the effect of PslG on bacterial surface movement in early biofilm development at a single-cell resolution by using a high-throughput bacterial tracking technique. The results showed that compared with no exogenous PslG addition, when PslG was added to the medium, bacterial surface movement was significantly faster by 4 ∼ 5 times, and was in a more random way with no clear preferred direction. A further study revealed that the fraction of walking mode increased when PslG was added, which then resulted in an elevated average speed. The differences of motility due to PslG addition led to a clear distinction in patterns of bacterial surface movement, and retarded microcolony formation greatly. Our results provide insight into developing new PslG-based biofilm-control techniques. IMPORTANCE Biofilms of Pseudomonas aeruginosa are a major cause for hospital-acquired infections. They are notoriously difficult to eradicate and pose serious health hazard to our human society. So finding new ways to control biofilms are urgently needed. Recent work on PslG showed that PslG might be a good candidate for inhibiting/disassembling biofilms of Pseudomonas aeruginosa through Psl-based regulation. However, to fully explore PslG functions in biofilm-control, a better understanding of PslG-Psl interactions is needed. Toward this end, we examined the effect of PslG on the surface movement of Pseudomonas aeruginosa in this work. The significance of our work is in greatly enhancing our understanding of the inhibiting mechanism of PslG on biofilms by providing a detailed picture of bacterial surface movement at a single-cell level, which will allow a full understanding

  20. Label-free molecular imaging of bacterial communities of the opportunistic pathogen Pseudomonas aeruginosa

    Science.gov (United States)

    Baig, Nameera; Polisetti, Sneha; Morales-Soto, Nydia; Dunham, Sage J. B.; Sweedler, Jonathan V.; Shrout, Joshua D.; Bohn, Paul W.

    2016-09-01

    Biofilms, such as those formed by the opportunistic human pathogen Pseudomonas aeruginosa are complex, matrix enclosed, and surface-associated communities of cells. Bacteria that are part of a biofilm community are much more resistant to antibiotics and the host immune response than their free-floating counterparts. P. aeruginosa biofilms are associated with persistent and chronic infections in diseases such as cystic fibrosis and HIV-AIDS. P. aeruginosa synthesizes and secretes signaling molecules such as the Pseudomonas quinolone signal (PQS) which are implicated in quorum sensing (QS), where bacteria regulate gene expression based on population density. Processes such as biofilms formation and virulence are regulated by QS. This manuscript describes the powerful molecular imaging capabilities of confocal Raman microscopy (CRM) and surface enhanced Raman spectroscopy (SERS) in conjunction with multivariate statistical tools such as principal component analysis (PCA) for studying the spatiotemporal distribution of signaling molecules, secondary metabolites and virulence factors in biofilm communities of P. aeruginosa. Our observations reveal that the laboratory strain PAO1C synthesizes and secretes 2-alkyl-4-hydroxyquinoline N-oxides and 2-alkyl-4-hydroxyquinolones in high abundance, while the isogenic acyl homoserine lactone QS-deficient mutant (ΔlasIΔrhlI) strain produces predominantly 2-alkyl-quinolones during biofilm formation. This study underscores the use of CRM, along with traditional biological tools such as genetics, for studying the behavior of microbial communities at the molecular level.

  1. Quantification of biofilm structures by the novel computer program COMSTAT.

    Science.gov (United States)

    Heydorn, A; Nielsen, A T; Hentzer, M; Sternberg, C; Givskov, M; Ersbøll, B K; Molin, S

    2000-10-01

    The structural organization of four microbial communities was analysed by a novel computer program, COMSTAT, which comprises ten features for quantifying three-dimensional biofilm image stacks. Monospecies biofilms of each of the four bacteria, Pseudomonas: putida, P. aureofaciens, P. fluorescens and P. aeruginosa, tagged with the green fluorescent protein (GFP) were grown in flow chambers with a defined minimal medium as substrate. Analysis by the COMSTAT program of four variables describing biofilm structure - mean thickness, roughness, substratum coverage and surface to volume ratio - showed that the four Pseudomonas: strains represent different modes of biofilm growth. P. putida had a unique developmental pattern starting with single cells on the substratum growing into micro-colonies, which were eventually succeeded by long filaments and elongated cell clusters. P. aeruginosa colonized the entire substratum, and formed flat, uniform biofilms. P. aureofaciens resembled P. aeruginosa, but had a stronger tendency to form micro-colonies. Finally, the biofilm structures of P. fluorescens had a phenotype intermediate between those of P. putida and P. aureofaciens. Analysis of biofilms of P. aureofaciens growing on 0.03 mM, 0.1 mM or 0.5 mM citrate minimal media showed that mean biofilm thickness increased with increasing citrate concentration. Moreover, biofilm roughness increased with lower citrate concentrations, whereas surface to volume ratio increased with higher citrate concentrations.

  2. Chemical sanitizers to control biofilms formed by two Pseudomonas species on stainless steel surface Sanificantes químicos no controle de biofilmes formados por duas espécies de Pseudomonas em superfície de aço inoxidável

    Directory of Open Access Journals (Sweden)

    Danila Soares Caixeta

    2012-03-01

    Full Text Available The biofilm formation of Pseudomonas aeruginosa and Pseudomonas fluorescens on AISI 304 stainless steel in the presence of reconstituted skim milk under different temperatures was conducted, and the potential of three chemical sanitizers in removing the mono-species biofilms formed was compared. Pseudomonas aeruginosa cultivated in skim milk at 28 °C presented better growth rate (10.4 log CFU.mL-1 when compared with 3.7 and 4.2 log CFU.mL-1 for P. aeruginosa and P. fluorescens cultivated at 7 °C, respectively. Pseudomonas aeruginosa formed biofilm when cultivated at 28 °C. However, only the adhesion of P. aeruginosa and P. fluorescens was observed when incubated at 7 °C. The sodium dichloroisocyanurate was the most efficient sanitizer in the reduction of the adhered P. aeruginosa cells at 7 and 28 °C and those on the biofilm, respectively. The hydrogen peroxide was more effective in the reduction of adhered cells of P. fluorescens at 7 °C.A capacidade de adesão e formação de biofilme por Pseudomonas aeruginosa e Pseudomonas fluorescens em aço inoxidável AISI 304, na presença de leite desnatado resconstituído sobre diferentes temperaturas foi conduzido e o potencial de três sanificantes químicos na remoção de biofilmes monoespécies foi comparado. Pseudomonas aeruginosa cultivada em leite desnatado a 28 °C apresentou melhor crescimento (10,4 log UFC.mL-1 quando comparado com 3,7 and 4,2 log UFC.mL-1 para P. aeruginosa e P. fluorescens cultivadas a 7 °C, respectivamente. Pseudomonas aeruginosa formou biofilme quando cultivada a 28 °C. Contudo foi observado somente adesão de P. aeruginosa e P. fluorescens quando incubada a 7 °C. O dicloroisocianurato de sódio foi o sanificante mais eficiente na redução de células aderidas e em biofilme de P. aeruginosa a 7 e 28 °C, respectivamente. O peróxido de hidrogênio foi o mais eficiente na redução de células aderidas de P. fluorescens a 7 °C.

  3. Facultative control of matrix production optimizes competitive fitness in Pseudomonas aeruginosa PA14 biofilm models.

    Science.gov (United States)

    Madsen, Jonas S; Lin, Yu-Cheng; Squyres, Georgia R; Price-Whelan, Alexa; de Santiago Torio, Ana; Song, Angela; Cornell, William C; Sørensen, Søren J; Xavier, Joao B; Dietrich, Lars E P

    2015-12-01

    As biofilms grow, resident cells inevitably face the challenge of resource limitation. In the opportunistic pathogen Pseudomonas aeruginosa PA14, electron acceptor availability affects matrix production and, as a result, biofilm morphogenesis. The secreted matrix polysaccharide Pel is required for pellicle formation and for colony wrinkling, two activities that promote access to O2. We examined the exploitability and evolvability of Pel production at the air-liquid interface (during pellicle formation) and on solid surfaces (during colony formation). Although Pel contributes to the developmental response to electron acceptor limitation in both biofilm formation regimes, we found variation in the exploitability of its production and necessity for competitive fitness between the two systems. The wild type showed a competitive advantage against a non-Pel-producing mutant in pellicles but no advantage in colonies. Adaptation to the pellicle environment selected for mutants with a competitive advantage against the wild type in pellicles but also caused a severe disadvantage in colonies, even in wrinkled colony centers. Evolution in the colony center produced divergent phenotypes, while adaptation to the colony edge produced mutants with clear competitive advantages against the wild type in this O2-replete niche. In general, the structurally heterogeneous colony environment promoted more diversification than the more homogeneous pellicle. These results suggest that the role of Pel in community structure formation in response to electron acceptor limitation is unique to specific biofilm models and that the facultative control of Pel production is required for PA14 to maintain optimum benefit in different types of communities. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  4. Role of inducers in detection of blaPDC-mediated oxyimino-cephalosporin resistance in Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Birson Ingti

    2017-01-01

    Interpretation & conclusions: P. aeruginosa harbouring inducible (chromosomal and plasmid-mediated AmpC β-lactamase is a matter of concern as it may limit therapeutic option. Using cefoxitin-ceftazidime-based test is simple and may be used for detecting inducible AmpC β-lactamase amongst P. aeruginosa.

  5. Fremmedlegemeinfektioner--nyt om biofilm og quorum sensing

    DEFF Research Database (Denmark)

    Høiby, Niels; Johansen, Helle Krogh; Ciofu, Oana

    2007-01-01

    Biofilms are structured consortia of bacteria embedded in self-produced polymer matrix. Biofilms are resistant to antibiotics, disinfectives and phagocytosis. The persistence of foreign body infections is due to biofilms. Chronic P. aeruginosa lung infection in cystic fibrosis patients is a biofilm....... Bacteria in biofilms communicate by means of quorum sensing which activates genes for virulence factors. Biofilms can be prevented by antibiotic prophylaxis or early therapy or by quorum sensing inhibitors which make them susceptible to antibiotics and phagocytosis....

  6. Increased resistance of contact lens related bacterial biofilms to antimicrobial activity of soft contact lens care solutions

    Science.gov (United States)

    Szczotka-Flynn, Loretta B.; Imamura, Yoshifumi; Chandra, Jyotsna; Yu, Changping; Mukherjee, Pranab K.; Pearlman, Eric; Ghannoum, Mahmoud A.

    2014-01-01

    PURPOSE To determine if clinical and reference strains of Pseudomonas aeruginosa, Serratia marcescens, and Staphylococcus aureus form biofilms on silicone hydrogel contact lenses, and ascertain antimicrobial activities of contact lens care solutions. METHODS Clinical and American Type Culture Collection (ATCC) reference strains of Pseudomonas aeruginosa, Serratia marcescens, and Staphylococcus aureus were incubated with lotrafilcon A lenses under conditions that facilitate biofilm formation. Biofilms were quantified by quantitative culturing (colony forming units, CFUs), and gross morphology and architecture were evaluated using scanning electron microscopy (SEM) and confocal microscopy. Susceptibilities of the planktonic and biofilm growth phases of the bacteria to five common multipurpose contact lens care solutions and one hydrogen peroxide care solution were assessed. RESULTS P. aeruginosa, S. marcescens, and S. aureus reference and clinical strains formed biofilms on lotrafilcon A silicone hydrogel contact lenses, as dense networks of cells arranged in multiple layers with visible extracellular matrix. The biofilms were resistant to commonly used biguanide preserved multipurpose care solutions. P. aeruginosa and S. aureus biofilms were susceptible to a hydrogen peroxide and a polyquaternium preserved care solution, whereas S. marcescens biofilm was resistant to a polyquaternium preserved care solution but susceptible to hydrogen peroxide disinfection. In contrast, the planktonic forms were always susceptible. CONCLUSIONS P. aeruginosa, S. marcescens, and S. aureus form biofilms on lotrafilcon A contact lenses, which in contrast to planktonic cells, are resistant to the antimicrobial activity of several soft contact lens care products. PMID:19654521

  7. Non-Identity-Mediated CRISPR-Bacteriophage Interaction Mediated via the Csy and Cas3 Proteins ▿#

    Science.gov (United States)

    Cady, Kyle C.; O'Toole, George A.

    2011-01-01

    Studies of the Escherichia, Neisseria, Thermotoga, and Mycobacteria clustered regularly interspaced short palindromic repeat (CRISPR) subtypes have resulted in a model whereby CRISPRs function as a defense system against bacteriophage infection and conjugative plasmid transfer. In contrast, we previously showed that the Yersinia-subtype CRISPR region of Pseudomonas aeruginosa strain UCBPP-PA14 plays no detectable role in viral immunity but instead is required for bacteriophage DMS3-dependent inhibition of biofilm formation by P. aeruginosa. The goal of this study is to define the components of the Yersinia-subtype CRISPR region required to mediate this bacteriophage-host interaction. We show that the Yersinia-subtype-specific CRISPR-associated (Cas) proteins Csy4 and Csy2 are essential for small CRISPR RNA (crRNA) production in vivo, while the Csy1 and Csy3 proteins are not absolutely required for production of these small RNAs. Further, we present evidence that the core Cas protein Cas3 functions downstream of small crRNA production and that this protein requires functional HD (predicted phosphohydrolase) and DEXD/H (predicted helicase) domains to suppress biofilm formation in DMS3 lysogens. We also determined that only spacer 1, which is not identical to any region of the DMS3 genome, mediates the CRISPR-dependent loss of biofilm formation. Our evidence suggests that gene 42 of phage DMS3 (DMS3-42) is targeted by CRISPR2 spacer 1 and that this targeting tolerates multiple point mutations between the spacer and DMS3-42 target sequence. This work demonstrates how the interaction between P. aeruginosa strain UCBPP-PA14 and bacteriophage DMS3 can be used to further our understanding of the diverse roles of CRISPR system function in bacteria. PMID:21398535

  8. Antimicrobial and Antibiofilm Activity and Machine Learning Classification Analysis of Essential Oils from Different Mediterranean Plants against Pseudomonas aeruginosa.

    Science.gov (United States)

    Artini, Marco; Patsilinakos, Alexandros; Papa, Rosanna; Božović, Mijat; Sabatino, Manuela; Garzoli, Stefania; Vrenna, Gianluca; Tilotta, Marco; Pepi, Federico; Ragno, Rino; Selan, Laura

    2018-02-23

    Pseudomonas aeruginosa is a ubiquitous organism and opportunistic pathogen that can cause persistent infections due to its peculiar antibiotic resistance mechanisms and to its ability to adhere and form biofilm. The interest in the development of new approaches for the prevention and treatment of biofilm formation has recently increased. The aim of this study was to seek new non-biocidal agents able to inhibit biofilm formation, in order to counteract virulence rather than bacterial growth and avoid the selection of escape mutants. Herein, different essential oils extracted from Mediterranean plants were analyzed for their activity against P. aeruginosa . Results show that they were able to destabilize biofilm at very low concentration without impairing bacterial viability. Since the action is not related to a bacteriostatic/bactericidal activity on P. aeruginosa , the biofilm change of growth in presence of the essential oils was possibly due to a modulation of the phenotype. To this aim, application of machine learning algorithms led to the development of quantitative activity-composition relationships classification models that allowed to direct point out those essential oil chemical components more involved in the inhibition of biofilm production. The action of selected essential oils on sessile phenotype make them particularly interesting for possible applications such as prevention of bacterial contamination in the community and in healthcare environments in order to prevent human infections. We assayed 89 samples of different essential oils as P. aeruginosa anti-biofilm. Many samples inhibited P. aeruginosa biofilm at concentrations as low as 48.8 µg/mL. Classification of the models was developed through machine learning algorithms.

  9. The metabolically active subpopulation in Pseudomonas aeruginosa biofilms survives exposure to membrane-targeting antimicrobials via distinct molecular mechanisms

    DEFF Research Database (Denmark)

    Chiang, Wen-Chi; Pamp, Sünje Johanna; Nilsson, Martin

    2012-01-01

    encoding lipopolysaccharide modification enzymes, as well as on the mexAB-oprM, mexCD-oprJ, and muxABC-opmB genes encoding antimicrobial efflux pumps, but does not depend on the mexPQ-opmE efflux pump genes. Development of chlorhexidine-tolerant subpopulations was found to depend on the mexCD-oprJ genes......, but does not depend on the pmr, mexAB-oprM, mexPQ-opmE, or muxABC-opmB genes. Tolerance to SDS and EDTA in P. aeruginosa biofilms is linked to metabolically active cells, but does not depend on the pmr, mexAB, mexCD, mexPQ, or muxABC genes. Our data suggest that the active subpopulation in P. aeruginosa......-targeting compounds colistin, EDTA, SDS, and chlorhexidine resulted in the same spatial distribution of live and dead bacteria, we investigated whether tolerance to these compounds originated from the same molecular mechanisms. Development of colistin-tolerant subpopulations was found to depend on the pmr genes...

  10. Fremmedlegemeinfektioner--nyt om biofilm og quorum sensing

    DEFF Research Database (Denmark)

    Høiby, Niels; Johansen, Helle Krogh; Ciofu, Oana

    2007-01-01

    Biofilms are structured consortia of bacteria embedded in self-produced polymer matrix. Biofilms are resistant to antibiotics, disinfectives and phagocytosis. The persistence of foreign body infections is due to biofilms. Chronic P. aeruginosa lung infection in cystic fibrosis patients is a biofilm....... Bacteria in biofilms communicate by means of quorum sensing which activates genes for virulence factors. Biofilms can be prevented by antibiotic prophylaxis or early therapy or by quorum sensing inhibitors which make them susceptible to antibiotics and phagocytosis. Udgivelsesdato: 2007-Nov-26...

  11. [Formation of microbial biofilms in causative agents of acute and chronic pyelonephritis].

    Science.gov (United States)

    Lagun, L V; Atanasova, Iu V; Tapal'skiĭ, D V

    2013-01-01

    Study the intensity of formation of microbial biofilms by Pseudomonas aeruginosa, Escherichia coli, Klebsiella pneumoniae, Staphylococcus aureus strains isolated during various forms of pyelonephritis. 150 clinical isolates of microorganisms isolated from urine ofpatientswith acute and chronic pyelonephritiswere included into the study. Determination of intensity of film-formation was carried out by staining of the formed biofilms by crystal violet with consequent extraction of the dye and measurement of its concentration in washout solution. Among causative agents ofpyelonephritis P. aeruginosa isolates had the maximum film-forming ability. The intensity of biofilm formation of these isolates was 2-3 time higher than staphylococcus and enterobacteria strains. Strains isolated from patients with chronic pyelonephritis by ability to form biofilms significantly surpassed strains isolated from acute pyelonephritis patients. A higher ability to form microbial biofilms for microorganisms--causative agents of pyelonephritis progressing against the background ofurolithiasis was noted. The ability to form biofilms is determined by both causative agent species and character of the infectious process in which this microorganism participates. Intensive formation of biofilms by E. coli, P. aeruginosa, K. pneumoniae, S. aureus clinical isolates may be an important factor of chronization of urinary tract infections.

  12. Synergistic antibacterial efficacy of early combination treatment with tobramycin and quorum-sensing inhibitors against Pseudomonas aeruginosa in an intraperitoneal foreign-body infection mouse model

    DEFF Research Database (Denmark)

    Christensen, Louise; van Gennip, Maria; Jakobsen, Tim H

    2012-01-01

    Quorum sensing (QS)-deficient Pseudomonas aeruginosa biofilms formed in vitro are more susceptible to tobramycin than QS-proficient P. aeruginosa biofilms, and combination treatment with a QS inhibitor (QSI) and tobramycin shows synergistic effects on the killing of in vitro biofilms. We extended...

  13. Effect of Lactoferrin on Oral Biofilm Formation

    Science.gov (United States)

    2009-10-01

    effect of Lf on the early stages of single-species and multi- species oral biofilm development. Streptococcus gordonii (Sg), Streptococcus mutans ...and biofilm development by Pseudomonas aeruginosa and Streptococcus mutans have been demonstrated, limited studies have been conducted on its effect...the effect of Lf on the early stages of single- species and multi-species oral biofilm development. Streptococcus gordonii, Streptococcus mutans

  14. Antibacterial effect of the laser-generated Se nanocoatings on Staphylococcus aureus and Pseudomonas aeruginosa biofilms

    Science.gov (United States)

    Ionin, A. A.; Ivanova, A. K.; Khmel'nitskii, R. A.; Klevkov, Yu V.; Kudryashov, S. I.; Levchenko, A. O.; Nastulyavichus, A. A.; Rudenko, A. A.; Saraeva, I. N.; Smirnov, N. A.; Zayarny, D. A.; Gonchukov, S. A.; Tolordava, E. R.

    2018-01-01

    The antibacterial properties of selenium nanoparticles (Se NPs) were successfully demonstrated in vitro for Staphylococcus aureus and Pseudomonas aeruginosa biofilms. The possible mechanisms of antibacterial impact included the emergence of reactive oxygen species, induced by free radicals on the NP surface and accompanied by subsequent oxidative stress, as well as mechanical decomposition of the mitochondrial membrane. Se nanocoatings were deposited on bare and silver-coated silica glass substrates via inkjet printing with concentrated nanoinks, prepared by infrared laser-ablative processing of a solid Se target in a 50%-isopropyl solution. The resulted porous nanofilms with high-percentage surface coverage, consisting of spherical Se NPs and Se nanorods, were characterized by means of standard microscopy techniques (optical, scanning electron, transmission), UV-vis-IR and EDX spectroscopy.

  15. Alginate overproduction affects Pseudomonas aeruginosa biofilm structure and function

    DEFF Research Database (Denmark)

    Hentzer, Morten; Teitzel, G.M.; Balzer, G.J.

    2001-01-01

    -resistant communities of microorganisms organized in biofilms. Although biofilm formation and the conversion to mucoidy are both important aspects of CF pathogenesis, the relationship between them is at the present unclear. In this study, we report that the overproduction of alginate affects biofilm development...... on an abiotic surface. Biofilms formed by an alginate- overproducing strain exhibit a highly structured architecture and are significantly more resistant to the antibiotic tobramycin than a biofilm formed by an isogenic nonmucoid strain. These results suggest that an important consequence of the conversion...

  16. Antimicrobial and Antibiofilm Activity and Machine Learning Classification Analysis of Essential Oils from Different Mediterranean Plants against Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Marco Artini

    2018-02-01

    Full Text Available Pseudomonas aeruginosa is a ubiquitous organism and opportunistic pathogen that can cause persistent infections due to its peculiar antibiotic resistance mechanisms and to its ability to adhere and form biofilm. The interest in the development of new approaches for the prevention and treatment of biofilm formation has recently increased. The aim of this study was to seek new non-biocidal agents able to inhibit biofilm formation, in order to counteract virulence rather than bacterial growth and avoid the selection of escape mutants. Herein, different essential oils extracted from Mediterranean plants were analyzed for their activity against P. aeruginosa. Results show that they were able to destabilize biofilm at very low concentration without impairing bacterial viability. Since the action is not related to a bacteriostatic/bactericidal activity on P. aeruginosa, the biofilm change of growth in presence of the essential oils was possibly due to a modulation of the phenotype. To this aim, application of machine learning algorithms led to the development of quantitative activity–composition relationships classification models that allowed to direct point out those essential oil chemical components more involved in the inhibition of biofilm production. The action of selected essential oils on sessile phenotype make them particularly interesting for possible applications such as prevention of bacterial contamination in the community and in healthcare environments in order to prevent human infections. We assayed 89 samples of different essential oils as P. aeruginosa anti-biofilm. Many samples inhibited P. aeruginosa biofilm at concentrations as low as 48.8 µg/mL. Classification of the models was developed through machine learning algorithms.

  17. Diagnosis of biofilm infections in cystic fibrosis patients

    DEFF Research Database (Denmark)

    Høiby, Niels; Bjarnsholt, Thomas; Moser, Claus

    2017-01-01

    Chronic Pseudomonas aeruginosa biofilm lung infection in cystic fibrosis patients is the best described biofilm infection in medicine. The initial focus can be the paranasal sinuses and then follows repeated colonization and infection of the lungs by aspiration. The matrix of the biofilms is domi...... by other pathogens e.g., Stenotrophomonas, Burkholderia multivorans, Achromobacter xylosoxidans and Mycobacterium abscessus complex....

  18. Anti-infective properties of Lactobacillus fermentum against Staphylococcus aureus and Pseudomonas aeruginosa.

    Science.gov (United States)

    Varma, Parvathi; Nisha, N; Dinesh, Kavitha R; Kumar, Anil V; Biswas, Raja

    2011-01-01

    Surgical wounds and implant-associated Staphylococcus aureus and Pseudomonas aeruginosa infections are often difficult to treat because of limited susceptibility of several of these strains to conventional antibiotics. As a result, there is a constant need for new alternative drugs. The aim of this study was to investigate the antimicrobial properties of Lactobacillus fermentum, a probiotic bacterium, which we have isolated from colonic biopsies. The inhibition of S. aureus and P. aeruginosa growth was evaluated by coincubating with L. fermentum strains. Growth inhibition was tested for several of their clinical isolates using agar well diffusion assays. For biofilm assay S. aureus and P. aeruginosa were grown on the glass slides and in 96-well plates in presence of 2.5 μg/ml culture filtrate of L. fermentum. Biofilms were photographed using confocal microscope or stained with 0.1% crystal violet. Reduction in the cytotoxicity of S. aureus and P. aeruginosa was observed in presence of 2.5 μg/ml L. fermentum-spent media. Using in vitroexperiments, we showed that L. fermentum-secreted compound(s) inhibits the growth, cytotoxicity and biofilm formation of several S. aureus and P. aeruginosa strains. Compound(s) present in the culture supernatant of L. fermentum may have promising applications in treating hospital-acquired infections. Copyright © 2011 S. Karger AG, Basel.

  19. A study on the long term effect of biofilm produced by biosurfactant producing microbe on medical implant

    International Nuclear Information System (INIS)

    Prabhawathi, Veluchamy; Thirunavukarasu, Kathirvel; Doble, Mukesh

    2014-01-01

    Low density polyethylene (LDPE) is used as a long term medical implant. Biofilm forming ability of two pathogenic microorganisms, namely, Bacillus subtilis (B. subtilis) and Pseudomonas aeruginosa (P. aeruginosa) on this polymer and the differences in the properties of these matrices are studied for a year. There are very few long term studies on biofilms formed on medical implants. After three months, colonies of B. subtilis were two times higher when compared to those of P. aeruginosa. And at the end of one year, they were two orders of magnitude higher than the later. The exopolysaccharide (EPS) and biosurfactant recovered from the polymer surface after three months were 21 and 10.4 μg/cm 2 for B. subtilis and 13 and 8.6 μg/cm 2 for P. aeruginosa. After one year, these were higher in B. subtilis (50 and 37.1 μg/cm 2 , respectively) than in P. aeruginosa (34.1 and 31.8 μg/cm 2 , respectively). B. subtilis consisted of protein controlling the community and sporulation development, while P. aeruginosa had either housekeeping or metabolic proteins. The EPS in the respective biofilm consisted of biosurfactants produced by B. subtilis (surfactins, m/z = 1029 to 1134) and P. aeruginosa (rhamnolipids, m/z = 568 to 705). Thermogravimetric analysis indicated that LDPE incubated with these organisms underwent a weight loss of 4 and 3% after three months and 11.1 and 9.2% after one year, respectively at 435 °C. Laccase and manganese peroxidase were detected in the biofilm which could be involved in the degradation. The biosurfactant of these microorganisms altered the hydrophobicity of the surface, favoring their attachment and proliferation. - Highlights: • Early P.aeru biofilm had genes needed for motility but later for housekeeping. • Early B. sub biofilm had genes needed for its formation but later for maturity. • Cells and matrix components in B. sub biofilm are higher than in P.aeru. • Compositions of these two biofilms are different. • So they need

  20. A study on the long term effect of biofilm produced by biosurfactant producing microbe on medical implant

    Energy Technology Data Exchange (ETDEWEB)

    Prabhawathi, Veluchamy; Thirunavukarasu, Kathirvel; Doble, Mukesh, E-mail: mukeshd@iitm.ac.in

    2014-07-01

    Low density polyethylene (LDPE) is used as a long term medical implant. Biofilm forming ability of two pathogenic microorganisms, namely, Bacillus subtilis (B. subtilis) and Pseudomonas aeruginosa (P. aeruginosa) on this polymer and the differences in the properties of these matrices are studied for a year. There are very few long term studies on biofilms formed on medical implants. After three months, colonies of B. subtilis were two times higher when compared to those of P. aeruginosa. And at the end of one year, they were two orders of magnitude higher than the later. The exopolysaccharide (EPS) and biosurfactant recovered from the polymer surface after three months were 21 and 10.4 μg/cm{sup 2} for B. subtilis and 13 and 8.6 μg/cm{sup 2} for P. aeruginosa. After one year, these were higher in B. subtilis (50 and 37.1 μg/cm{sup 2}, respectively) than in P. aeruginosa (34.1 and 31.8 μg/cm{sup 2}, respectively). B. subtilis consisted of protein controlling the community and sporulation development, while P. aeruginosa had either housekeeping or metabolic proteins. The EPS in the respective biofilm consisted of biosurfactants produced by B. subtilis (surfactins, m/z = 1029 to 1134) and P. aeruginosa (rhamnolipids, m/z = 568 to 705). Thermogravimetric analysis indicated that LDPE incubated with these organisms underwent a weight loss of 4 and 3% after three months and 11.1 and 9.2% after one year, respectively at 435 °C. Laccase and manganese peroxidase were detected in the biofilm which could be involved in the degradation. The biosurfactant of these microorganisms altered the hydrophobicity of the surface, favoring their attachment and proliferation. - Highlights: • Early P.aeru biofilm had genes needed for motility but later for housekeeping. • Early B. sub biofilm had genes needed for its formation but later for maturity. • Cells and matrix components in B. sub biofilm are higher than in P.aeru. • Compositions of these two biofilms are different.

  1. Mimicking disinfection and drying of biofilms in contaminated endoscopes

    NARCIS (Netherlands)

    Kovaleva, J.; Degener, J. E.; van der Mei, H. C.

    2010-01-01

    The effects of peracetic acid-based (PAA) disinfectant with, and without, additional drying on Candida albicans, Candida parapsilosis, Pseudomonas aeruginosa and Stenotrophomonas maltophilia, isolated from contaminated flexible endoscopes, in single-and dual-species biofilms were studied. Biofilms

  2. Antibiotic tolerance and microbial biofilms

    DEFF Research Database (Denmark)

    Folkesson, Anders

    Increased tolerance to antimicrobial agents is thought to be an important feature of microbes growing in biofilms. We study the dynamics of antibiotic action within hydrodynamic flow chamber biofilms of Escherichia coli and Pseudomonas aeruginosa using isogenic mutants and fluorescent gene...... expression reporters and we address the question of how biofilm organization affects antibiotic susceptibility. The dynamics of microbial killing is monitored by viable count determination, and confocal laser microscopy. Our work shows that the apparent increased antibiotic tolerance is due to the formation...... of antibiotic tolerant subpopulations within the biofilm. The formation of these subpopulations is highly variable and dependent on the antibiotic used, the biofilm structural organization and the induction of specific tolerance mechanisms....

  3. Expression, purification, crystallization and preliminary crystallographic analysis of PA3885 (TpbA) from Pseudomonas aeruginosa PAO1

    International Nuclear Information System (INIS)

    Yang, Wen; Li, Kan; Bai, Yuwei; Zhou, Ruimin; Zhou, Weihong; Bartlam, Mark

    2010-01-01

    PA3885 (TpbA), a tyrosine phosphatase, may function as a balancing factor between biofilm formation and motility in the opportunistic pathogen P. aeruginosa. Here, the expression, purification, crystallization and preliminary crystallographic analysis of PA3885 from P. aeruginosa PAO1 are reported. Biofilms are important in cell communication and growth in most bacteria and are also responsible for most human clinical infections and diseases. Quorum-sensing systems have been identified to be crucial for biofilm formation and regulation. PA3885 (TpbA), a tyrosine phosphatase, is reported to convert extracellular quorum-sensing signals into internal gene-cascade reactions that result in reduced biofilm formation in the opportunistic pathogen Pseudomonas aeruginosa. Here, PA3885 from P. aeruginosa PAO1 was expressed, purified and crystallized. Single crystals were studied by X-ray crystallography and native diffraction data were collected to 2.8 Å resolution. These crystals were determined to belong to space group C2. It was not possible to conclusively determine the number of proteins in the asymmetric unit from the preliminary X-ray diffraction data analysis alone and attempts to determine the crystal structure of PA3885 are currently under way

  4. Spectroscopic identification and anti-biofilm properties of polar metabolites from the medicinal plant Helichrysum italicum against Pseudomonas aeruginosa.

    Science.gov (United States)

    D'Abrosca, Brigida; Buommino, Elisabetta; D'Angelo, Grazia; Coretti, Lorena; Scognamiglio, Monica; Severino, Valeria; Pacifico, Severina; Donnarumma, Giovanna; Fiorentino, Antonio

    2013-11-15

    Two new acylated styrylpyrones, one 5-methoxy-1(3H)-isobenzofuranone glucoside and a hydroxymethyl-orcinol derivative, along with sixteen known aromatic metabolites, including lignans, quinic acid derivatives low-molecular weight phenol glucosides, have been isolated from the methanol extract of Helichrysum italicum, a medicinal plant typical of the Mediterranean vegetation. The structures of these compounds have been elucidated on the basis of extensive 2D-NMR spectroscopic analyses, including COSY, TOCSY, HSQC, CIGAR-HMBC, H2BC and HSQC-TOCSY, along with Q-TOF HRMS(2) analysis. Selected compounds were evaluated for their anti-biofilm properties against Pseudomonas aeruginosa. Copyright © 2013. Published by Elsevier Ltd.

  5. Alginate production affects Pseudomonas aeruginosa biofilm development and architecture, but is not essential for biofilm formation

    DEFF Research Database (Denmark)

    Stapper, A.P.; Narasimhan, G.; Oman, D.E.

    2004-01-01

    of their biofilm formation using confocal laser scanning microscopy. Biofilm Image Processing (BIP) and Community Statistics (COMSTAT) software programs were used to provide quantitative measurements of the two-dimensional biofilm images. All three strains formed distinguishable biofilm architectures, indicating...

  6. Biofilm Formation As a Response to Ecological Competition.

    Directory of Open Access Journals (Sweden)

    Nuno M Oliveira

    2015-07-01

    Full Text Available Bacteria form dense surface-associated communities known as biofilms that are central to their persistence and how they affect us. Biofilm formation is commonly viewed as a cooperative enterprise, where strains and species work together for a common goal. Here we explore an alternative model: biofilm formation is a response to ecological competition. We co-cultured a diverse collection of natural isolates of the opportunistic pathogen Pseudomonas aeruginosa and studied the effect on biofilm formation. We show that strain mixing reliably increases biofilm formation compared to unmixed conditions. Importantly, strain mixing leads to strong competition: one strain dominates and largely excludes the other from the biofilm. Furthermore, we show that pyocins, narrow-spectrum antibiotics made by other P. aeruginosa strains, can stimulate biofilm formation by increasing the attachment of cells. Side-by-side comparisons using microfluidic assays suggest that the increase in biofilm occurs due to a general response to cellular damage: a comparable biofilm response occurs for pyocins that disrupt membranes as for commercial antibiotics that damage DNA, inhibit protein synthesis or transcription. Our data show that bacteria increase biofilm formation in response to ecological competition that is detected by antibiotic stress. This is inconsistent with the idea that sub-lethal concentrations of antibiotics are cooperative signals that coordinate microbial communities, as is often concluded. Instead, our work is consistent with competition sensing where low-levels of antibiotics are used to detect and respond to the competing genotypes that produce them.

  7. The cep quorum-sensing system of Burkholderia cepacia H111 controls biofilm formation and swarming motility

    DEFF Research Database (Denmark)

    Huber, B.; Riedel, K.; Hentzer, Morten

    2001-01-01

    Burkholderia cepacia and Pseudomonas aeruginosa often co-exist as mixed biofilms in the lungs of patients suffering from cystic fibrosis (CF). Here, the isolation of random mini-Tn5 insertion mutants of B. cepacia H111 defective in biofilm formation on an abiotic surface is reported. It is demons......Burkholderia cepacia and Pseudomonas aeruginosa often co-exist as mixed biofilms in the lungs of patients suffering from cystic fibrosis (CF). Here, the isolation of random mini-Tn5 insertion mutants of B. cepacia H111 defective in biofilm formation on an abiotic surface is reported...

  8. Fluorescence-Based Reporter for Gauging Cyclic Di-GMP Levels in Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Rybtke, Morten T.; Borlee, Bradley R.; Murakami, Keiji

    2012-01-01

    The increased tolerance toward the host immune system and antibiotics displayed by biofilm-forming Pseudomonas aeruginosa and other bacteria in chronic infections such as cystic fibrosis bronchopneumonia is of major concern. Targeting of biofilm formation is believed to be a key aspect in the dev...

  9. Isolate-specific effects of patulin, penicillic Acid and EDTA on biofilm formation and growth of dental unit water line biofilm isolates.

    Science.gov (United States)

    Liaqat, Iram; Bachmann, Robert Thomas; Sabri, Anjum Nasim; Edyvean, Robert G J

    2010-08-01

    Dental unit water line (DUWL) contamination by opportunistic pathogens has its significance in nosocomial infection of patients, health care workers, and life-threatening infections to immunocompromized persons. Recently, the quorum sensing (QS) system of DUWL isolates has been found to affect their biofilm-forming ability, making it an attractive target for antimicrobial therapy. In this study, the effect of two quorum-sensing inhibitory compounds (patulin; PAT, penicillic acid; PA) and EDTA on planktonic growth, AI-2 signalling and in vitro biofilm formation of Pseudomonas aeruginosa, Achromobacter xylosoxidans and Achromobacter sp. was monitored. Vibrio harveyi BB170 bioassay and crystal violet staining methods were used to detect the AI-2 monitoring and biofilm formation in DUWL isolates, respectively. The V. harveyi BB170 bioassay failed to induce bioluminescence in A. xylosoxidans and Achromobacter sp., while P. aeruginosa showed AI-2 like activity suggesting the need of some pretreatments prior to bioassay. All strains were found to form biofilms within 72 h of incubation. The QSIs/EDTA combination have isolate-specific effects on biofilm formation and in some cases it stimulated biofilm formation as often as it was inhibited. However, detailed information about the anti-biofilm effect of these compounds is still lacking.

  10. Disinfection of bacterial biofilms in pilot-scale cooling tower systems.

    Science.gov (United States)

    Liu, Yang; Zhang, Wei; Sileika, Tadas; Warta, Richard; Cianciotto, Nicholas P; Packman, Aaron I

    2011-04-01

    The impact of continuous chlorination and periodic glutaraldehyde treatment on planktonic and biofilm microbial communities was evaluated in pilot-scale cooling towers operated continuously for 3 months. The system was operated at a flow rate of 10,080 l day(-1). Experiments were performed with a well-defined microbial consortium containing three heterotrophic bacteria: Pseudomonas aeruginosa, Klebsiella pneumoniae and Flavobacterium sp. The persistence of each species was monitored in the recirculating cooling water loop and in biofilms on steel and PVC coupons in the cooling tower basin. The observed bacterial colonization in cooling towers did not follow trends in growth rates observed under batch conditions and, instead, reflected differences in the ability of each organism to remain attached and form biofilms under the high-through flow conditions in cooling towers. Flavobacterium was the dominant organism in the community, while P. aeruginosa and K. pneumoniae did not attach well to either PVC or steel coupons in cooling towers and were not able to persist in biofilms. As a result, the much greater ability of Flavobacterium to adhere to surfaces protected it from disinfection, whereas P. aeruginosa and K. pneumoniae were subject to rapid disinfection in the planktonic state.

  11. Zingiber officinale: Its antibacterial activity on Pseudomonas aeruginosa and mode of action evaluated by flow cytometry.

    Science.gov (United States)

    Chakotiya, Ankita Singh; Tanwar, Ankit; Narula, Alka; Sharma, Rakesh Kumar

    2017-06-01

    Biofilm formation, low membrane permeability and efflux activity developed by Pseudomonas aeruginosa, play an important role in the mechanism of infection and antimicrobial resistance. In the present study we evaluate the antibacterial effect of Zingiber officinale against multi-drug resistant strain of P. aeruginosa. The study explores antibacterial efficacy and time-kill study concomitantly the effect of herbal extract on bacterial cell physiology with the use of flow cytometry and inhibition of biofilm formation. Z. officinale was found to inhibit the growth of P. aeruginosa, significantly. A major decline in the Colony Forming Units was observed with 3 log 10  at 12 h of treatment. Also it is found to affect the membrane integrity of the pathogen, as 70.06 ± 2.009% cells were found to stain with Propidium iodide. In case of efflux activity 86.9 ± 2.08% cells were found in Ethidium bromide positive region. Biofilm formation inhibition ability was found in the range of 68.13 ± 4.11% to 84.86 ± 2.02%. Z.officinale is effective for killing Multi-Drug Resistant P. aeruginosa clinical isolate by affecting the cellular physiology and inhibiting the biofilm formation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. In Vitro Efficacy of Nonantibiotic Treatments on Biofilm Disruption of Gram-Negative Pathogens and an In Vivo Model of Infectious Endometritis Utilizing Isolates from the Equine Uterus.

    Science.gov (United States)

    Ferris, Ryan A; McCue, Patrick M; Borlee, Grace I; Loncar, Kristen D; Hennet, Margo L; Borlee, Bradley R

    2016-03-01

    In this study, we evaluated the ability of the equine clinical treatments N-acetylcysteine, EDTA, and hydrogen peroxide to disrupt in vitro biofilms and kill equine reproductive pathogens (Escherichia coli, Pseudomonas aeruginosa, or Klebsiella pneumoniae) isolated from clinical cases. N-acetylcysteine (3.3%) decreased biofilm biomass and killed bacteria within the biofilms of E. coli isolates. The CFU of recoverable P. aeruginosa and K. pneumoniae isolates were decreased, but the biofilm biomass was unchanged. Exposure to hydrogen peroxide (1%) decreased the biofilm biomass and reduced the CFU of E. coli isolates, K. pneumoniae isolates were observed to have a reduction in CFU, and minimal effects were observed for P. aeruginosa isolates. Chelating agents (EDTA formulations) reduced E. coli CFU but were ineffective at disrupting preformed biofilms or decreasing the CFU of P. aeruginosa and K. pneumoniae within a biofilm. No single nonantibiotic treatment commonly used in equine veterinary practice was able to reduce the CFU and biofilm biomass of all three Gram-negative species of bacteria evaluated. An in vivo equine model of infectious endometritis was also developed to monitor biofilm formation, utilizing bioluminescence imaging with equine P. aeruginosa isolates from this study. Following infection, the endometrial surface contained focal areas of bacterial growth encased in a strongly adherent "biofilm-like" matrix, suggesting that biofilms are present during clinical cases of infectious equine endometritis. Our results indicate that Gram-negative bacteria isolated from the equine uterus are capable of producing a biofilm in vitro, and P. aeruginosa is capable of producing biofilm-like material in vivo. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  13. The chemical digestion of Ti6Al7Nb scaffolds produced by Selective Laser Melting reduces significantly ability of Pseudomonas aeruginosa to form biofilm.

    Science.gov (United States)

    Junka, Adam F; Szymczyk, Patrycja; Secewicz, Anna; Pawlak, Andrzej; Smutnicka, Danuta; Ziółkowski, Grzegorz; Bartoszewicz, Marzenna; Chlebus, Edward

    2016-01-01

    In our previous work we reported the impact of hydrofluoric and nitric acid used for chemical polishing of Ti-6Al-7Nb scaffolds on decrease of the number of Staphylococcus aureus biofilm forming cells. Herein, we tested impact of the aforementioned substances on biofilm of Gram-negative microorganism, Pseudomonas aeruginosa, dangerous pathogen responsible for plethora of implant-related infections. The Ti-6Al-7Nb scaffolds were manufactured using Selective Laser Melting method. Scaffolds were subjected to chemical polishing using a mixture of nitric acid and fluoride or left intact (control group). Pseudomonal biofilm was allowed to form on scaffolds for 24 hours and was removed by mechanical vortex shaking. The number of pseudomonal cells was estimated by means of quantitative culture and Scanning Electron Microscopy. The presence of nitric acid and fluoride on scaffold surfaces was assessed by means of IR and rentgen spetorscopy. Quantitative data were analysed using the Mann-Whitney test (P ≤ 0.05). Our results indicate that application of chemical polishing correlates with significant drop of biofilm-forming pseudomonal cells on the manufactured Ti-6Al-7Nb scaffolds ( p = 0.0133, Mann-Whitney test) compared to the number of biofilm-forming cells on non-polished scaffolds. As X-ray photoelectron spectroscopy revealed the presence of fluoride and nitrogen on the surface of scaffold, we speculate that drop of biofilm forming cells may be caused by biofilm-supressing activity of these two elements.

  14. Bisphenol A removal by a Pseudomonas aeruginosa immobilized on granular activated carbon and operating in a fluidized bed reactor

    Energy Technology Data Exchange (ETDEWEB)

    Mita, Luigi [National Laboratory on Endocrine Disruptors, National Institute of Biostructures and Biosystems (INBB), Via P. Castellino, 111, 80131 Naples (Italy); Institute of Genetic and Biophysics “ABT”, Via P. Castellino, 111, 80131 Naples Italy (Italy); Grumiro, Laura [National Laboratory on Endocrine Disruptors, National Institute of Biostructures and Biosystems (INBB), Via P. Castellino, 111, 80131 Naples (Italy); Rossi, Sergio [Institute of Genetic and Biophysics “ABT”, Via P. Castellino, 111, 80131 Naples Italy (Italy); Bianco, Carmen; Defez, Roberto [Institute of Biosciences and BioResources, Via P. Castellino, 111, 80131 Naples (Italy); Gallo, Pasquale [Dipartimento di Chimica, Istituto Zooprofilattico Sperimentale del Mezzogiorno, Via della Salute 2, 80055 Portici, Naples (Italy); Mita, Damiano Gustavo, E-mail: mita@igb.cnr.it [National Laboratory on Endocrine Disruptors, National Institute of Biostructures and Biosystems (INBB), Via P. Castellino, 111, 80131 Naples (Italy); Institute of Genetic and Biophysics “ABT”, Via P. Castellino, 111, 80131 Naples Italy (Italy); Diano, Nadia [National Laboratory on Endocrine Disruptors, National Institute of Biostructures and Biosystems (INBB), Via P. Castellino, 111, 80131 Naples (Italy); Department of Experimental Medicine, Second University of Naples, Via S.M. di Costantinopoli, 16, 80138 Naples Italy (Italy)

    2015-06-30

    Highlights: • A fluidized bed reactor, filled with a Pseudomonas aeruginosa immobilized on GAC, has been used for BPA removal. • BPA removal resulted from a biological activated carbon (BAC) process. • Equations describing the results have been indicated. • BPA removal was analyzed as a function of time and biofilm reuse. - Abstract: Serratia rubidiae, Pseudomonas aeruginosa and Escherichia coli K12 have been studied for their ability of Bisphenol A removal from aqueous systems and biofilm formation on activated granule carbon. Mathematical equations for biodegradation process have been elaborated and discussed. P. aeruginosa was found the best strain to be employed in the process of Bisphenol A removal. The yield in BPA removal of a P. aeruginosa biofilm grown on GAC and operating in a fluidized bed reactor has been evaluated. The results confirm the usefulness in using biological activated carbon (BAC process) to remove phenol compounds from aqueous systems.

  15. Bisphenol A removal by a Pseudomonas aeruginosa immobilized on granular activated carbon and operating in a fluidized bed reactor

    International Nuclear Information System (INIS)

    Mita, Luigi; Grumiro, Laura; Rossi, Sergio; Bianco, Carmen; Defez, Roberto; Gallo, Pasquale; Mita, Damiano Gustavo; Diano, Nadia

    2015-01-01

    Highlights: • A fluidized bed reactor, filled with a Pseudomonas aeruginosa immobilized on GAC, has been used for BPA removal. • BPA removal resulted from a biological activated carbon (BAC) process. • Equations describing the results have been indicated. • BPA removal was analyzed as a function of time and biofilm reuse. - Abstract: Serratia rubidiae, Pseudomonas aeruginosa and Escherichia coli K12 have been studied for their ability of Bisphenol A removal from aqueous systems and biofilm formation on activated granule carbon. Mathematical equations for biodegradation process have been elaborated and discussed. P. aeruginosa was found the best strain to be employed in the process of Bisphenol A removal. The yield in BPA removal of a P. aeruginosa biofilm grown on GAC and operating in a fluidized bed reactor has been evaluated. The results confirm the usefulness in using biological activated carbon (BAC process) to remove phenol compounds from aqueous systems

  16. Microenvironmental characteristics and physiology of biofilms in chronic infections of CF patients are strongly affected by the host immune response

    DEFF Research Database (Denmark)

    Jensen, Peter Østrup; Kolpen, Mette; Kragh, Kasper Nørskov

    2017-01-01

    in chronic lung infections of cystic fibrosis (CF) patients is very different from what is observed in vitro, for example, in biofilms grown in flow chambers. Dense in vitro biofilms of P. aeruginosa exhibit rapid O2 depletion within ... investigations show that P. aeruginosa persists in the chronically infected CF lung as relatively small cell aggregates that are surrounded by numerous PMNs, where the activity of PMNs is the major cause of O2 depletion rendering the P. aeruginosa aggregates anoxic. High levels of nitrate and nitrite enable P....... aeruginosa to persist fueled by denitrification in the PMN-surrounded biofilm aggregates. This configuration creates a potentially long-term stable ecological niche for P. aeruginosa in the CF lung, which is largely governed by slow growth and anaerobic metabolism and enables persistence and resilience...

  17. The clinical impact of bacterial biofilms

    DEFF Research Database (Denmark)

    Høiby, Niels; Ciofu, Oana; Johansen, Helle Krogh

    2011-01-01

    Bacteria survive in nature by forming biofilms on surfaces and probably most, if not all, bacteria (and fungi) are capable of forming biofilms. A biofilm is a structured consortium of bacteria embedded in a self-produced polymer matrix consisting of polysaccharide, protein and extracellular DNA....... Bacterial biofilms are resistant to antibiotics, disinfectant chemicals and to phagocytosis and other components of the innate and adaptive inflammatory defense system of the body. It is known, for example, that persistence of staphylococcal infections related to foreign bodies is due to biofilm formation....... Likewise, chronic Pseudomonas aeruginosa lung infections in cystic fibrosis patients are caused by biofilm growing mucoid strains. Gradients of nutrients and oxygen exist from the top to the bottom of biofilms and the bacterial cells located in nutrient poor areas have decreased metabolic activity...

  18. Killing mediated spatial structure in V. Cholerae biofilms

    Science.gov (United States)

    Yanni, David

    Most bacteria live in biofilms, which are implicated in 60 - 80 % of microbial infections in the body. The spatial structure of a biofilm confers advantages to its member-cells, such as antibiotic resistance, and is strongly affected by competition between strains and taxa. However, A complete picture of how competition affects the self-organized structure of these complex, far-from-equilibrium systems, is yet to emerge. To that end, we investigate phase separation dynamics driven by T6SS-facilitated bacterial warfare in a system composed of two strains of mutually antagonistic V. cholerae. T6SS is a contact mediated killing mechanism present in 25 % of all gram negative bacteria, and has been shown by recent work to play a major role in the spatial assortment of biofilms. T6SS events induce lysis, causing variations in local mechanical pressure, and acting as thermalizing events. We study cells immobilized in biofilms at the air-solid interface, so our experimental system represents a different type active matter, wherein activity is due to cell death and reproduction, not mobility. Here, we show how that activity imposes a constraint of minimal curvature on strain-strain interfaces; an effective Laplace pressure is characterized which governs interfacial dynamics.

  19. Role of bacterial efflux pumps in biofilm formation.

    Science.gov (United States)

    Alav, Ilyas; Sutton, J Mark; Rahman, Khondaker Miraz

    2018-02-28

    Efflux pumps are widely implicated in antibiotic resistance because they can extrude the majority of clinically relevant antibiotics from within cells to the extracellular environment. However, there is increasing evidence from many studies to suggest that the pumps also play a role in biofilm formation. These studies have involved investigating the effects of efflux pump gene mutagenesis and efflux pump inhibitors on biofilm formation, and measuring the levels of efflux pump gene expression in biofilms. In particular, several key pathogenic species associated with increasing multidrug resistance, such as Acinetobacter baumannii, Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus, have been investigated, whilst other studies have focused on Salmonella enterica serovar Typhimurium as a model organism and problematic pathogen. Studies have shown that efflux pumps, including AcrAB-TolC of E. coli, MexAB-OprM of P. aeruginosa, AdeFGH of A. baumannii and AcrD of S. enterica, play important roles in biofilm formation. The substrates for such pumps, and whether changes in their efflux activity affect biofilm formation directly or indirectly, remain to be determined. By understanding the roles that efflux pumps play in biofilm formation, novel therapeutic strategies can be developed to inhibit their function, to help disrupt biofilms and improve the treatment of infections. This review will discuss and evaluate the evidence for the roles of efflux pumps in biofilm formation and the potential approaches to overcome the increasing problem of biofilm-based infections.

  20. A computer investigation of chemically mediated detachment in bacterial biofilms.

    Science.gov (United States)

    Hunt, Stephen M; Hamilton, Martin A; Sears, John T; Harkin, Gary; Reno, Jason

    2003-05-01

    A three-dimensional computer model was used to evaluate the effect of chemically mediated detachment on biofilm development in a negligible-shear environment. The model, BacLAB, combines conventional diffusion-reaction equations for chemicals with a cellular automata algorithm to simulate bacterial growth, movement and detachment. BacLAB simulates the life cycle of a bacterial biofilm from its initial colonization of a surface to the development of a mature biofilm with cell areal densities comparable to those in the laboratory. A base model founded on well established transport equations that are easily adaptable to investigate conjectures at the biological level has been created. In this study, the conjecture of a detachment mechanism involving a bacterially produced chemical detachment factor in which high local concentrations of this detachment factor cause the bacteria to detach from the biofilm was examined. The results show that the often observed 'mushroom'-shaped structure can occur if detachment events create voids so that the remaining attached cells look like mushrooms.

  1. Pseudomonas aeruginosa Trent and zinc homeostasis.

    Science.gov (United States)

    Davies, Corey B; Harrison, Mark D; Huygens, Flavia

    2017-09-01

    Pseudomonas aeruginosa is a Gram-negative pathogen and the major cause of mortality in patients with cystic fibrosis. The mechanisms that P. aeruginosa strains use to regulate intracellular zinc have an effect on infection, antibiotic resistance and the propensity to form biofilms. However, zinc homeostasis in P. aeruginosa strains of variable infectivity has not been compared. In this study, zinc homeostasis in P. aeruginosa Trent, a highly infectious clinical strain, was compared to that of a laboratory P. aeruginosa strain, ATCC27853. Trent was able to tolerate higher concentrations of additional zinc in rich media than ATCC27853. Further, pre-adaptation to additional zinc enhanced the growth of Trent at non-inhibitory concentrations but the impact of pre-adaption on the growth of ATCC27853 under the same conditions was minimal. The results establish clear differences in zinc-induced responses in Trent and ATCC27853, and how zinc homeostasis can be a promising target for the development of novel antimicrobial strategies for P. aeruginosa infection in cystic fibrosis patients. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  2. Antibiotic resistance of bacterial biofilms

    DEFF Research Database (Denmark)

    Hoiby, N.; Bjarnsholt, T.; Givskov, M.

    2010-01-01

    A biofilm is a structured consortium of bacteria embedded in a self-produced polymer matrix consisting of polysaccharide, protein and DNA. Bacterial biofilms cause chronic infections because they show increased tolerance to antibiotics and disinfectant chemicals as well as resisting phagocytosis...... and other components of the body's defence system. The persistence of, for example, staphylococcal infections related to foreign bodies is due to biofilm formation. Likewise, chronic Pseudomonas aeruginosa lung infection in cystic fibrosis patients is caused by biofilm-growing mucoid strains....... Characteristically, gradients of nutrients and oxygen exist from the top to the bottom of biofilms and these gradients are associated with decreased bacterial metabolic activity and increased doubling times of the bacterial cells; it is these more or less dormant cells that are responsible for some of the tolerance...

  3. MEDICINAL PLANTS FROM BRAZILIAN CAATINGA: ANTIBIOFILM AND ANTIBACTERIAL ACTIVITIES AGAINST Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    DANIELLE SILVA TRENTIN

    2014-01-01

    Full Text Available The Caatinga biome covers a vast area in northeastern Brazil and presents a high level of biodiversity. It is known that about 400 plant species are used by semi-arid local communities for medical purposes. Based on ethnopharmacological reports, this study aims to screen 24 species from Caatinga regarding the ability to prevent biofilm formation and to inhibit the growth of Pseudomonas aeruginosa - a major opportunistic human pathogen and an important causative agent of morbidity and mortality. The effects of aqueous extracts, at 0.4 and 4.0 mg mL-1, on biofilm formation and on growth of P. aeruginosa ATCC 27853 were studied using the crystal violet assay and the OD600 absorbance, respectively. The most active extracts were analyzed by thinlayer chromatography and high performance liquid chromatography. Our investigation pointed extracts of four species with potential application for the control of P. aeruginosa: Anadenanthera colubrina (Vell. Brenan, Commiphora leptophloeos (Mart. J.B. Gillett, Myracrodruoun urundeuva Allemão, whose antibiofilm effects (89%, 56% and 79% inhibition of biofilm, respectively were associated with complete inhibition of bacterial growth, and Pityrocarpa moniliformis (Benth. Luckow & R.W. Jobson, which were able avoid 68% of biofilm formation and inhibited 30% bacterial growth. The qualitative phytochemical analyses reveal the complexity of the samples as well as the presence of compounds with high molecular weight.

  4. An investigation of Pseudomonas aeruginosa biofilm growth on novel nanocellulose fibre dressings.

    Science.gov (United States)

    Powell, Lydia C; Khan, Saira; Chinga-Carrasco, Gary; Wright, Chris J; Hill, Katja E; Thomas, David W

    2016-02-10

    Nanocellulose from wood is a novel biomaterial, which is highly fibrillated at the nanoscale. This affords the material a number of advantages, including self-assembly, biodegradability and the ability to absorb and retain moisture, which highlights its potential usefulness in clinical wound-dressing applications. In these in vitro studies, the wound pathogen Pseudomonas aeruginosa PAO1 was used to assess the ability of two nanocellulose materials to impair bacterial growth (nanocelluloses had a relatively small fraction of residual fibres (nanocellulose films and increased cell death when compared to a commercial control wound dressing, Aquacel(®). Nanocellulose suspensions inhibited bacterial growth, whilst UV-vis spectrophotometry and laser profilometry also revealed the ability of nanocellulose to form smooth, translucent films. Atomic force microscopy studies of the surface properties of nanocellulose demonstrated that PAO1 exhibited markedly contrasting morphology when grown on the nanocellulose film surfaces compared to an Aquacel(®) control dressing (p<0.05). This study highlights the potential utility of these biodegradable materials, from a renewable source, for wound dressing applications in the prevention and treatment of biofilm development. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Predation Efficacy of Bdellovibrio bacteriovorus on Multidrug-Resistant Clinical Pathogens and Their Corresponding Biofilms.

    Science.gov (United States)

    Sun, Yao; Ye, Jianzhong; Hou, Yuanbo; Chen, Huale; Cao, Jianming; Zhou, Tieli

    2017-09-25

    The aim of the present study was to evaluate the predation efficacy of Bdellovibrio bacteriovorus on multidrug-resistant (MDR) or extensive drug resistant (XDR) gram-negative pathogens and their corresponding biofilms. In this study, we examined the ability of B. bacteriovorus to prey on MDR and XDR gram-negative clinical bacteria, including Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii. Results showed that B. bacteriovorus was able to prey on all planktonic cultures, among which the most efficient predation was observed for drug-resistant E. coli, with a 3.11 log10 reduction in viability. Furthermore, B. bacteriovorus demonstrated promising efficacy in preventing biofilm formation and dispersing the established biofilm. Reductions in biofilm formation of E. coli, K. pneumoniae, P. aeruginosa, and A. baumannii co-cultured with B. bacteriovorus were 65.2%, 37.1%, 44.7%, and 36.8%, respectively. Meanwhile, the established biofilms of E. coli, K. pneumoniae, P. aeruginosa, and A. baumannii were significantly reduced by 83.4%, 81.8%, 83.1%, and 79.9%, respectively. A visual analysis supported by scanning electron microscopy demonstrated the role of B. bacteriovorus in removing the established biofilms. This study highlights the potential use of B. bacteriovorus as a biological control agent with the capability to prey on MDR/XDR gram-negative pathogens and eradicate biofilms.

  6. Subinhibitory concentration of kanamycin induces the Pseudomonas aeruginosa type VI secretion system.

    Directory of Open Access Journals (Sweden)

    Cerith Jones

    Full Text Available Pseudomonas aeruginosa is a Gram-negative bacterium found in natural environments including plants, soils and warm moist surfaces. This organism is also in the top ten of nosocomial pathogens, and prevalent in cystic fibrosis (CF lung infections. The ability of P. aeruginosa to colonize a wide variety of environments in a lasting manner is associated with the formation of a resistant biofilm and the capacity to efficiently outcompete other microorganisms. Here we demonstrate that sub-inhibitory concentration of kanamycin not only induces biofilm formation but also induces expression of the type VI secretion genes in the H1-T6SS cluster. The H1-T6SS is known for its role in toxin production and bacterial competition. We show that the antibiotic induction of the H1-T6SS only occurs when a functional Gac/Rsm pathway is present. These observations may contribute to understand how P. aeruginosa responds to antibiotic producing competitors. It also suggests that improper antibiotic therapy may enhance P. aeruginosa colonization, including in the airways of CF patients.

  7. Pseudomonas aeruginosa outer membrane vesicles triggered by human mucosal fluid and lysozyme can prime host tissue surfaces for bacterial adhesion

    Directory of Open Access Journals (Sweden)

    Matteo Maria Emiliano Metruccio

    2016-06-01

    Full Text Available Pseudomonas aeruginosa is a leading cause of human morbidity and mortality that often targets epithelial surfaces. Host immunocompromise, or the presence of indwelling medical devices, including contact lenses, can predispose to infection. While medical devices are known to accumulate bacterial biofilms, it is not well understood why resistant epithelial surfaces become susceptible to P. aeruginosa. Many bacteria, including P. aeruginosa, release Outer Membrane Vesicles (OMVs in response to stress that can fuse with host cells to alter their function. Here, we tested the hypothesis that mucosal fluid can trigger OMV release to compromise an epithelial barrier. This was tested using tear fluid and corneal epithelial cells in vitro and in vivo. After 1 h both human tear fluid, and the tear component lysozyme, greatly enhanced OMV release from P. aeruginosa strain PAO1 compared to PBS controls (~100 fold. TEM and SDS-PAGE showed tear fluid and lysozyme-induced OMVs were similar in size and protein composition, but differed from biofilm-harvested OMVs, the latter smaller with fewer proteins. Lysozyme-induced OMVs were cytotoxic to human corneal epithelial cells in vitro and murine corneal epithelium in vivo. OMV exposure in vivo enhanced Ly6G/C expression at the corneal surface, suggesting myeloid cell recruitment, and primed the cornea for bacterial adhesion (~4-fold, P < 0.01. Sonication disrupted OMVs retained cytotoxic activity, but did not promote adhesion, suggesting the latter required OMV-mediated events beyond cell killing. These data suggest that mucosal fluid induced P. aeruginosa OMVs could contribute to loss of epithelial barrier function during medical device-related infections.

  8. Transfer of bacteria between stainless steel and chicken meat: A CLSM and DGGE study of biofilms

    Directory of Open Access Journals (Sweden)

    Christine C. Gaylarde

    2016-08-01

    Full Text Available This study aimed to assess the interaction between bacteria and food processing surfaces using novel methods. Microbial cross contamination between stainless steel, a common food processing material, and raw chicken was studied using microbiological culture, specialized microscope and molecular techniques. Confocal laser scanning microscopy (CLSM allowed the visualization of biofilms containing single or dual species of Escherichia coli O157:H7, Salmonella typhimurium, Bacillus cereus, Staphylococcus aureus and Pseudomonas aeruginosa, formed after 6 days’ incubation on stainless steel or 4h on raw chicken. The results provided information on intra-biofilm location and stratification of species within dual species biofilms. Top-to-bottom Z-stack images revealed that, on both materials, S. typhimurium and E. coli attached concurrently, the former in greater numbers. E. coli and B. cereus segregated on steel, E. coli more frequent near the metal surface, B. cereus almost the only species in outer layers. Few cells of S. aureus, found at all depths, were seen in the 2.9 µm thick biofilm on steel with E. coli. Greatest attachment was shown by P. aeruginosa, followed by S. typhimurium, E. coli and finally Gram positive species. Large amounts of EPS in P. aeruginosa biofilms made visualization difficult on both materials, but especially on chicken meat, a limitation of this technique. Nevertheless, CLSM was useful for determining time sequence of adhesion and species makeup of thin biofilms. The technique showed that five min contact between bacterially-contaminated chicken and sterile steel resulted in greatest transfer of P. aeruginosa, followed by S. typhimurium. This was confirmed using DGGE. Gram positive bacteria transferred poorly. A biofilm containing 2.3 × 105  cfu·cm−2 B. cereus on steel transferred an undetectable number of cells to chicken after 5 min contact. This species was unable to form biofilm on chicken when incubated for 4 h

  9. A simple and low-cost biofilm quantification method using LED and CMOS image sensor.

    Science.gov (United States)

    Kwak, Yeon Hwa; Lee, Junhee; Lee, Junghoon; Kwak, Soo Hwan; Oh, Sangwoo; Paek, Se-Hwan; Ha, Un-Hwan; Seo, Sungkyu

    2014-12-01

    A novel biofilm detection platform, which consists of a cost-effective red, green, and blue light-emitting diode (RGB LED) as a light source and a lens-free CMOS image sensor as a detector, is designed. This system can measure the diffraction patterns of cells from their shadow images, and gather light absorbance information according to the concentration of biofilms through a simple image processing procedure. Compared to a bulky and expensive commercial spectrophotometer, this platform can provide accurate and reproducible biofilm concentration detection and is simple, compact, and inexpensive. Biofilms originating from various bacterial strains, including Pseudomonas aeruginosa (P. aeruginosa), were tested to demonstrate the efficacy of this new biofilm detection approach. The results were compared with the results obtained from a commercial spectrophotometer. To utilize a cost-effective light source (i.e., an LED) for biofilm detection, the illumination conditions were optimized. For accurate and reproducible biofilm detection, a simple, custom-coded image processing algorithm was developed and applied to a five-megapixel CMOS image sensor, which is a cost-effective detector. The concentration of biofilms formed by P. aeruginosa was detected and quantified by varying the indole concentration, and the results were compared with the results obtained from a commercial spectrophotometer. The correlation value of the results from those two systems was 0.981 (N = 9, P CMOS image-sensor platform. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Anti-Biofilm Efficacy of Nitric Oxide-Releasing Silica Nanoparticles

    OpenAIRE

    Hetrick, Evan M.; Shin, Jae Ho; Paul, Heather S.; Schoenfisch, Mark H.

    2009-01-01

    The ability of nitric oxide (NO)-releasing silica nanoparticles to kill biofilm-based microbial cells is reported. Biofilms of Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, and Candida albicans were formed in vitro and exposed to NO-releasing silica nanoparticles. Replicative viability experiments revealed that ≥ 99% of cells from each type of biofilm were killed via NO release, with the greatest efficacy (≥ 99.999% killing) against gram-negative...

  11. Pseudomonas aeruginosa and Saccharomyces cerevisiae Biofilm in Flow Cells

    DEFF Research Database (Denmark)

    Weiss Nielsen, Martin; Sternberg, Claus; Molin, Søren

    2011-01-01

    well-defined conditions(2,3). The system consists of a flow cell that serves as growth chamber for the biofilm. The flow cell is supplied with nutrients and oxygen from a medium flask via a peristaltic pump and spent medium is collected in a waste container. This construction of the flow system allows......Many microbial cells have the ability to form sessile microbial communities defined as biofilms that have altered physiological and pathological properties compared to free living microorganisms. Biofilms in nature are often difficult to investigate and reside under poorly defined conditions(1...... a continuous supply of nutrients and administration of e.g. antibiotics with minimal disturbance of the cells grown in the flow chamber. Moreover, the flow conditions within the flow cell allow studies of biofilm exposed to shear stress. A bubble trapping device confines air bubbles from the tubing which...

  12. Pseudomonas aeruginosa and Saccharomyces cerevisiae Biofilm in Flow Cells

    DEFF Research Database (Denmark)

    Weiss Nielsen, Martin; Sternberg, Claus; Molin, Søren

    2011-01-01

    or proteins compatible with CLSM analysis. This enables online visualization and allows investigation of niches in the developing biofilm. Microbial interrelationship, investigation of antimicrobial agents or the expression of specific genes, are of the many experimental setups that can be investigated......). Using a transparent substratum it is possible to device a system where simple biofilms can be examined in a non-destructive way in real-time: here we demonstrate the assembly and operation of a flow cell model system, for in vitro 3D studies of microbial biofilms generating high reproducibility under...... well-defined conditions(2,3). The system consists of a flow cell that serves as growth chamber for the biofilm. The flow cell is supplied with nutrients and oxygen from a medium flask via a peristaltic pump and spent medium is collected in a waste container. This construction of the flow system allows...

  13. Inhibition of biofilm formation by D-tyrosine: Effect of bacterial type and D-tyrosine concentration.

    Science.gov (United States)

    Yu, Cong; Li, Xuening; Zhang, Nan; Wen, Donghui; Liu, Charles; Li, Qilin

    2016-04-01

    D-Tyrosine inhibits formation and triggers disassembly of bacterial biofilm and has been proposed for biofouling control applications. This study probes the impact of D-tyrosine in different biofilm formation stages in both G+ and G- bacteria, and reveals a non-monotonic correlation between D-tyrosine concentration and biofilm inhibition effect. In the attachment stage, cell adhesion was studied in a flow chamber, where D-tyrosine caused significant reduction in cell attachment. Biofilms formed by Pseudomonas aeruginosa and Bacillus subtilis were characterized by confocal laser scanning microscopy as well as quantitative analysis of cellular biomass and extracellular polymeric substances. D-Tyrosine exhibited strong inhibitive effects on both biofilms with an effective concentration as low as 5 nM; the biofilms responded to D-tyrosine concentration change in a non-monotonic, bi-modal pattern. In addition, D-tyrosine showed notable and different impact on EPS production by G+ and G- bacteria. Extracellular protein was decreased in P. aeruginosa biofilms, but increased in those of B. subtilis. Exopolysaccharides production by P. aeruginosa was increased at low concentrations and reduced at high concentrations while no impact was found in B. subtilis. These results suggest that distinct mechanisms are at play at different D-tyrosine concentrations and they may be species specific. Dosage of D-tyrosine must be carefully controlled for biofouling control applications. Copyright © 2016 Elsevier Ltd. All rights reserved.

  14. Sinus biofilms in patients with cystic fibrosis: is adjusted eradication therapy needed?

    DEFF Research Database (Denmark)

    Aanaes, Kasper; Eickhardt, Steffen; Johansen, Helle Krogh

    2015-01-01

    The paranasal sinuses can be a focus for colonisation of the cystic fibrosis (CF) lungs with pathogens. In the sinuses, bacteria can adapt to the lung environment and enhance their antibiotic resistance, with biofilm formation thought to be the most important adaptive mechanism, causing recalcitr......The paranasal sinuses can be a focus for colonisation of the cystic fibrosis (CF) lungs with pathogens. In the sinuses, bacteria can adapt to the lung environment and enhance their antibiotic resistance, with biofilm formation thought to be the most important adaptive mechanism, causing...... recalcitrant disease. The presence of biofilms in CF sinuses is sparsely described. In this descriptive cross-sectional study, the sinus mucosa from 16 CF patients were analysed by fluorescence in situ hybridization using specific peptide nucleic acid (PNA-FISH) probes for Pseudomonas aeruginosa...... and Staphylococcus aureus to demonstrate the presence of biofilms. Small clusters of biofilm were visualised lining the sinus mucosa of CF patients. Biofilms were found in 10 out of 18 cases; 7 with intermittent lung colonisation, 2 chronically infected, and one lung transplanted patient. Finding P. aeruginosa...

  15. Controlled assembly of silver nano-fluid in Heliotropium crispum extract: A potent anti-biofilm and bactericidal formulation

    Science.gov (United States)

    Khan, Faria; Hashmi, Muhammad Uzair; Khalid, Nauman; Hayat, Muhammad Qasim; Ikram, Aamer; Janjua, Hussnain A.

    2016-11-01

    The study describes the optimized method for silver nanoparticle (AgNPs) synthesis using Heliotropium crispum (HC) plant extract. Optimization of physicochemical parameters resulted in stable and rapidly assembled AgNPs. FTIR results suggest presence of plant phytochemicals that helped in the reduction, stabilization and capping of AgNPs. The assembled Ag nano-composites displayed the peak surface plasmon resonance (SPR) around 428 nm. The presence of uniquely assembled Ag-biomolecule composites, cap and stabilize nanoparticles in aqueous plant suspension. Spherical, uniform-shaped AgNPs with low poly-dispersion and average particle size of 42 nm and was determined through dynamic light scattering (DLS) and scanning election microscopy (SEM) which present robust interaction with microbes. The study also evaluates the antimicrobial and anti-biofilm properties of biologically synthesized AgNPs on clinical isolates of MRSA, Pseudomonas aeruginosa and Acinetobacter baumannii. Minimum inhibitory concentration (0.5 mg mL-1) of nanoparticles that presented bactericidal effect was made through inhibition assays on bacterial strains. The concentration which presented potent bactericidal response was then evaluated through growth inhibition in liquid medium for anti-biofilm studies at 2.0 mg mL-1. HC-Ag nanoparticles mediated anti-biofilm effects on Pseudomonas aeruginosa was revealed through SEM. Complete breakdown of biofilm's extracellular polymeric substances resulted after incubation with AgNPs. Peptidoglycan cell wall destruction was also revealed on planktonic bacterial images after 24 h of incubation.

  16. Efflux mediated adaptive and cross resistance to ciprofloxacin and benzalkonium chloride in Pseudomonas aeruginosa of dairy origin.

    Science.gov (United States)

    Pagedar, Ankita; Singh, Jitender; Batish, Virender K

    2011-06-01

    The present study was undertaken to investigate the role of efflux pump activity (EPA) in conferring adaptive and cross resistances against ciprofloxacin (CF) and benzalkonium chloride (BC) in dairy isolates of Pseudomonas aeruginosa. Biofilm formation potential was correlated with development of adaptive resistance in originally resistant strains. Irrespective of parent strains's susceptibility, isolates developed substantial adaptive resistance against CF and BC. Significant difference was observed in ability of non resistant isolates to develop adaptive resistance against CF and BC (P Reduction in adaptive resistances due to EPI was more evident in originally non resistant strains, which reaffirms EPA as probable mechanism of adaptive resistance. The present study perhaps first of its kind, suggests an active role of EPA in conferring adaptive and cross resistances in food related P. aeruginosa isolates and supports reverse hypothesis that antibiotic-resistant organisms eventually become tolerant to other antibacterial agents as well. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Long-Chain 4-Aminoquinolines as Quorum Sensing Inhibitors in Serratia marcescens and Pseudomonas aeruginosa.

    Science.gov (United States)

    Aleksić, Ivana; Šegan, Sandra; Andrić, Filip; Zlatović, Mario; Moric, Ivana; Opsenica, Dejan M; Senerovic, Lidija

    2017-05-19

    Antibiotic resistance has become a serious global threat to public health; therefore, improved strategies and structurally novel antimicrobials are urgently needed to combat infectious diseases. Here we report a new type of highly potent 4-aminoquinoline derivatives as quorum sensing inhibitors in Serratia marcescens and Pseudomonas aeruginosa, exhibiting weak bactericidal activities (minimum inhibitory concentration (MIC) > 400 μM). Through detailed structure-activity study, we have identified 7-Cl and 7-CF 3 substituted N-dodecylamino-4-aminoquinolines (5 and 10) as biofilm formation inhibitors with 50% biofilm inhibition at 69 μM and 63 μM in S. marcescens and P. aeruginosa, respectively. These two compounds, 5 and 10, are the first quinoline derivatives with anti-biofilm formation activity reported in S. marcescens. Quantitative structure-activity relationship (QSAR) analysis identified structural descriptors such as Wiener indices, hyper-distance-path index (HDPI), mean topological charge (MTC), topological charge index (TCI), and log D(o/w) exp as the most influential in biofilm inhibition in this bacterial species. Derivative 10 is one of the most potent quinoline type inhibitors of pyocyanin production described so far (IC 50 = 2.5 μM). While we have demonstrated that 5 and 10 act as Pseudomonas quinolone system (PQS) antagonists, the mechanism of inhibition of S. marcescens biofilm formation with these compounds remains open since signaling similar to P. aeruginosa PQS system has not yet been described in Serratia and activity of these compounds on acylhomoserine lactone (AHL) signaling has not been detected. Our data show that 7-Cl and 7-CF 3 substituted N-dodecylamino-4-aminoquinolines present the promising scaffolds for developing antivirulence and anti-biofilm formation agents against multidrug-resistant bacterial species.

  18. Pseudomonas aeruginosa in Swimming Pool Water: Evidences and Perspectives for a New Control Strategy

    OpenAIRE

    Guida, Marco; Di Onofrio, Valeria; Gall?, Francesca; Gesuele, Renato; Valeriani, Federica; Liguori, Renato; Romano Spica, Vincenzo; Liguori, Giorgio

    2016-01-01

    Pseudomonas aeruginosa is frequently isolated in swimming pool settings. Nine recreational and rehabilitative swimming pools were monitored according to the local legislation. The presence of P. aeruginosa was correlated to chlorine concentration. The ability of the isolates to form a biofilm on plastic materials was also investigated. In 59.5% of the samples, microbial contamination exceeded the threshold values. P. aeruginosa was isolated in 50.8% of these samples. The presence of P. aerugi...

  19. Use of artificial sputum medium to test antibiotic efficacy against Pseudomonas aeruginosa in conditions more relevant to the cystic fibrosis lung.

    Science.gov (United States)

    Kirchner, Sebastian; Fothergill, Joanne L; Wright, Elli A; James, Chloe E; Mowat, Eilidh; Winstanley, Craig

    2012-06-05

    There is growing concern about the relevance of in vitro antimicrobial susceptibility tests when applied to isolates of P. aeruginosa from cystic fibrosis (CF) patients. Existing methods rely on single or a few isolates grown aerobically and planktonically. Predetermined cut-offs are used to define whether the bacteria are sensitive or resistant to any given antibiotic. However, during chronic lung infections in CF, P. aeruginosa populations exist in biofilms and there is evidence that the environment is largely microaerophilic. The stark difference in conditions between bacteria in the lung and those during diagnostic testing has called into question the reliability and even relevance of these tests. Artificial sputum medium (ASM) is a culture medium containing the components of CF patient sputum, including amino acids, mucin and free DNA. P. aeruginosa growth in ASM mimics growth during CF infections, with the formation of self-aggregating biofilm structures and population divergence. The aim of this study was to develop a microtitre-plate assay to study antimicrobial susceptibility of P. aeruginosa based on growth in ASM, which is applicable to both microaerophilic and aerobic conditions. An ASM assay was developed in a microtitre plate format. P. aeruginosa biofilms were allowed to develop for 3 days prior to incubation with antimicrobial agents at different concentrations for 24 hours. After biofilm disruption, cell viability was measured by staining with resazurin. This assay was used to ascertain the sessile cell minimum inhibitory concentration (SMIC) of tobramycin for 15 different P. aeruginosa isolates under aerobic and microaerophilic conditions and SMIC values were compared to those obtained with standard broth growth. Whilst there was some evidence for increased MIC values for isolates grown in ASM when compared to their planktonic counterparts, the biggest differences were found with bacteria tested in microaerophilic conditions, which showed a much

  20. Extracellular DNA as matrix component in microbial biofilms

    DEFF Research Database (Denmark)

    Chiang, Wen-Chi; Tolker-Nielsen, Tim

    2010-01-01

    Bacteria in nature primarily live in surface-associated communities commonly known as biofilms. Because bacteria in biofilms, in many cases, display tolerance to host immune systems, antibiotics, and biocides, they are often difficult or impossible to eradicate. Biofilm formation, therefore, leads...... to various persistent infections in humans and animals, and to a variety of complications in industry, where solid–water interfaces occur. Knowledge about the molecular mechanisms involved in biofilm formation is necessary for creating strategies to control biofilms. Recent studies have shown...... that extracellular DNA is an important component of the extracellular matrix of microbial biofilms. The present chapter is focussed on extracellular DNA as matrix component in biofilms formed by Pseudomonas aeruginosa as an example from the Gram-negative bacteria, and Streptococcus and Staphylococcus as examples...

  1. Bisphenol A removal by a Pseudomonas aeruginosa immobilized on granular activated carbon and operating in a fluidized bed reactor.

    Science.gov (United States)

    Mita, Luigi; Grumiro, Laura; Rossi, Sergio; Bianco, Carmen; Defez, Roberto; Gallo, Pasquale; Mita, Damiano Gustavo; Diano, Nadia

    2015-06-30

    Serratia rubidiae, Pseudomonas aeruginosa and Escherichia coli K12 have been studied for their ability of Bisphenol A removal from aqueous systems and biofilm formation on activated granule carbon. Mathematical equations for biodegradation process have been elaborated and discussed. P. aeruginosa was found the best strain to be employed in the process of Bisphenol A removal. The yield in BPA removal of a P. aeruginosa biofilm grown on GAC and operating in a fluidized bed reactor has been evaluated. The results confirm the usefulness in using biological activated carbon (BAC process) to remove phenol compounds from aqueous systems. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Pseudomonas aeruginosa biofilms in the respiratory tract of cystic fibrosis patients

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Jensen, Peter Østrup; Fiandaca, Mark J

    2009-01-01

    therapy, explanted lungs from 3 intensively treated chronically P. aeruginosa infected CF patients and routine sputum from 77 chronically P. aeruginosa infected CF patients. All samples were investigated microscopically using hematoxylin-eosin (HE), Gram and alcian-blue stain, PNA FISH...

  3. Bacteriophage-antibiotic synergism to control planktonic and biofilm ...

    African Journals Online (AJOL)

    Bacteriophage-antibiotic synergism to control planktonic and biofilm producing clinical isolates of Pseudomonas aeruginosa. Amina Amal Mahmoud Nouraldin, Manal Mohammad Baddour, Reem Abdel Hameed Harfoush, Sara AbdelAziz Mohamed Essa ...

  4. Paracentrin 1, a synthetic antimicrobial peptide from the sea-urchin Paracentrotus lividus, interferes with staphylococcal and Pseudomonas aeruginosa biofilm formation.

    Science.gov (United States)

    Schillaci, Domenico; Cusimano, Maria Grazia; Spinello, Angelo; Barone, Giampaolo; Russo, Debora; Vitale, Maria; Parrinello, Daniela; Arizza, Vincenzo

    2014-01-01

    The rise of antibiotic-resistance as well as the reduction of investments by pharmaceutical companies in the development of new antibiotics have stimulated the investigation for alternative strategies to conventional antibiotics. Many antimicrobial peptides show a high specificity for prokaryotes and a low toxicity for eukaryotic cells and, due to their mode of action the development of resistance is considered unlikely. We recently characterized an antimicrobial peptide that was called Paracentrin 1 from the 5-kDa peptide fraction from the coelomocyte cytosol of the Paracentrotus lividus. In this study, the chemically synthesized Paracentrin 1, was tested for its antimicrobial and antibiofilm properties against reference strains of Gram positive and Gram negative. The Paracentrin 1 was active against planktonic form of staphylococcal strains (reference and isolates) and Pseudomonas aeruginosa ATCC 15442 at concentrations ranging from 12.5 to 6.2 mg/ml. The Paracentrin 1 was able to inhibit biofilm formation of staphylococcal and Pseudomonas aeruginosa strains at concentrations ranging from 3.1 to 0.75 mg/ml. We consider the tested peptide as a good starting molecule for novel synthetic derivatives with improved pharmaceutical potential.

  5. The pvc operon regulates the expression of the Pseudomonas aeruginosa fimbrial chaperone/usher pathway (cup genes.

    Directory of Open Access Journals (Sweden)

    Uzma Qaisar

    Full Text Available The Pseudomonas aeruginosa fimbrial structures encoded by the cup gene clusters (cupB and cupC contribute to its attachment to abiotic surfaces and biofilm formation. The P. aeruginosa pvcABCD gene cluster encodes enzymes that synthesize a novel isonitrile functionalized cumarin, paerucumarin. Paerucumarin has already been characterized chemically, but this is the first report elucidating its role in bacterial biology. We examined the relationship between the pvc operon and the cup gene clusters in the P. aeruginosa strain MPAO1. Mutations within the pvc genes compromised biofilm development and significantly reduced the expression of cupB1-6 and cupC1-3, as well as different genes of the cupB/cupC two-component regulatory systems, roc1/roc2. Adjacent to pvc is the transcriptional regulator ptxR. A ptxR mutation in MPAO1 significantly reduced the expression of the pvc genes, the cupB/cupC genes, and the roc1/roc2 genes. Overexpression of the intact chromosomally-encoded pvc operon by a ptxR plasmid significantly enhanced cupB2, cupC2, rocS1, and rocS2 expression and biofilm development. Exogenously added paerucumarin significantly increased the expression of cupB2, cupC2, rocS1 and rocS2 in the pvcA mutant. Our results suggest that pvc influences P. aeruginosa biofilm development through the cup gene clusters in a pathway that involves paerucumarin, PtxR, and different cup regulators.

  6. Antimicrobial blue light inactivation of biofilms formed by clinical isolates of multidrug-resistant microorganisms

    Science.gov (United States)

    Ferrer-Espada, Raquel; Fang, Yanyan; Dai, Tianhong

    2018-02-01

    Antibiotic resistance is one of the most serious threats to public health. It is estimated that at least 23,000 people die each year in the USA as a direct result of antibiotic-resistant infections. In addition, many antibiotic-resistant microorganisms develop biofilms, surface-associated microbial communities that are extremely resistant to antibiotics and the immune system. A light-based approach, antimicrobial blue light (aBL), has attracted increasing attention due to its intrinsic antimicrobial effect without the involvement of exogenous photosensitizers. In this study, we investigated the effectiveness of this non-antibiotic approach against biofilms formed by multidrug-resistant (MDR) microorganisms. MDR Acinetobacter baumannii, Escherichia coli, Candida albicans, and Pseudomonas aeruginosa biofilms were grown either in 96-well microtiter plates for 24 h or in a CDC biofilm reactor for 48 h, and then exposed to aBL at 405 nm emitted from a light-emitting diode (LED). We demonstrated that, for the biofilms grown in the CDC biofilm reactor, approximately 1.88 log10 CFU reduction was achieved in A. baumannii, 2.78 log10 CFU in E. coli and 3.18 log10 CFU in P. aeruginosa after 162 J/cm2 , 576 J/cm2 and 500 J/cm2 aBL were delivered, respectively. For the biofilms formed in the 96-well microtiter plates, 5.67 and 2.46 log10 CFU reduction was observed in P. aeruginosa and C. albicans polymicrobial biofilm after an exposure of 216 J/cm2 . In conclusion, aBL is potentially an alternative non-antibiotic approach against MDR biofilm-related infections. Future studies are warranted to investigate other important MDR microorganisms, the mechanism of action of aBL, and aBL efficacy in vivo.

  7. Garlic as an inhibitor of Pseudomonas aeruginosa quorum sensing in cystic fibrosis--a pilot randomized controlled trial

    DEFF Research Database (Denmark)

    Smyth, Alan R; Cifelli, Paramita M; Ortori, Catharine A

    2010-01-01

    Pseudomonas aeruginosa forms biofilms in the cystic fibrosis lung. Quorum sensing (QS) controls biofilm maturation, immune evasion, antibiotic tolerance and virulence factor production. Garlic shows QS inhibitory activity in vitro and in animal models. We report the first clinical trial in man of...

  8. Biofilm removal technique using sands as a research tool for accessing microbial attachment on surface

    Directory of Open Access Journals (Sweden)

    Nathanon Trachoo

    2004-01-01

    Full Text Available Biofilms have profound impacts on improved survival of the constituent microorganisms in nature. Biofilms were believed to protect constituent microorganisms from sanitizer treatment, provide a more suitable habitat for microorganisms, and become a site for genetic material exchanges between microorganisms. As we realize more about the significance of biofilm, methods used for biofilm study should be consistently developed and evaluated. To determine microbial attachment on surfaces, usually biofilms are grown on substratum surfaces and removed by vortexing with glass beads or scraping. However, scraping is not as effective as vortexing with glass beads. Another approach is direct-agar overlaying which cannot be used with high density biofilm. In this experiment, we compared effectiveness of glass beads (298±28 μm in diameter and sands (width: 221±55 μm and length: 329±118 μm in removing biofilm of Pseudomonas aeruginosa by vortexing method. The results suggested that acid-washed sands, which are significantly less inexpensive than glass beads, were as effective as (P>0.05 analytical grade glass beads in Pseudomonas aeruginosa biofilm removal without inhibiting growth of the organism.

  9. Bacteriophage Therapy for Staphylococcus aureus Biofilm-Infected Wounds: A New Approach to Chronic Wound Care

    Science.gov (United States)

    2013-02-01

    lidocaine and 1:100,000 epineph- rine at the planned wound sites. Six full-thickness dermal wounds, 6 mm in diameter, were created on the ventral ear...action were ineffective against S. aureus biofilm, as was seen with P. aeruginosa biofilm.22 Given the durability of biofilm in the face of a harsh

  10. Antimikrobiální peptidy z jedu divoce žijící včely odstraňují biofilm a působí synergicky s antibiotiky proti Pseudomonas aeruginosa

    Czech Academy of Sciences Publication Activity Database

    Nešuta, Ondřej; Monincová, Lenka; Voburka, Zdeněk; Bednárová, Lucie; Slaninová, Jiřina; Čeřovský, Václav

    2013-01-01

    Roč. 107, č. 5 (2013), s. 429-430 ISSN 0009-2770. [Mezioborové setkání mladých biologů, biochemiků a chemiků /13./. 14.5.2013-17.5.2013, Žďár nad Sázavou] Institutional support: RVO:61388963 Keywords : antimicrobial peptides * Pseudomonas aeruginosa * Staphylococcus aureus * synergism * biofilm Subject RIV: CE - Biochemistry

  11. Sputum Active Polymyxin Lipopeptides: Activity against Cystic Fibrosis Pseudomonas aeruginosa Isolates and Their Interactions with Sputum Biomolecules.

    Science.gov (United States)

    Schneider-Futschik, Elena K; Paulin, Olivia K A; Hoyer, Daniel; Roberts, Kade D; Ziogas, James; Baker, Mark A; Karas, John; Li, Jian; Velkov, Tony

    2018-05-11

    The mucoid biofilm mode of growth of Pseudomonas aeruginosa ( P. aeruginosa) in the lungs of cystic fibrosis patients makes eradication of infections with antibiotic therapy very difficult. The lipopeptide antibiotics polymyxin B and colistin are currently the last-resort therapies for infections caused by multidrug-resistant P. aeruginosa. In the present study, we investigated the antibacterial activity of a series of polymyxin lipopeptides (polymyxin B, colistin, FADDI-003, octapeptin A 3 , and polymyxin A 2 ) against a panel of polymyxin-susceptible and polymyxin-resistant P. aeruginosa cystic fibrosis isolates grown under planktonic or biofilm conditions in artificial sputum and their interactions with sputum component biomolecules. In sputum media under planktonic conditions, the lipopeptides FADDI-003 and octapeptin A 3 displayed very promising activity against the polymyxin-resistant isolate FADDI-PA066 (polymyxin B minimum inhibitory concentration (MIC) = 32 mg/L), while retaining their activity against the polymyxin-sensitive strains FADDI-PA021 (polymyxin B MIC = 1 mg/L) and FADDI-PA020 (polymyxin B MIC = 2 mg/L). Polymyxin A 2 was only effective against the polymyxin-sensitive isolates. However, under biofilm growth conditions, the hydrophobic lipopeptide FADDI-003 was inactive compared to the more hydrophilic lipopeptides, octapeptin A 3 , polymyxin A 2 , polymyxin B, and colistin. Transmission electron micrographs revealed octapeptin A 3 caused reduction in the cell numbers in biofilm as well as biofilm disruption/"antibiofilm" activity. We therefore assessed the interactions of the lipopeptides with the component sputum biomolecules, mucin, deoxyribonucleic acid (DNA), surfactant, F-actin, lipopolysaccharide, and phospholipids. We observed the general trend that sputum biomolecules reduce lipopeptide antibacterial activity. Collectively, our data suggests that, in the airways, lipopeptide binding to component sputum biomolecules may reduce

  12. Detection of Pathogenic Biofilms with Bacterial Amyloid Targeting Fluorescent Probe, CDy11

    DEFF Research Database (Denmark)

    Kim, Jun Young; Sahu, Srikanta; Yau, Yin Hoe

    2016-01-01

    Bacterial biofilms are responsible for a wide range of persistent infections. In the clinic, diagnosis of biofilm-associated infections relies heavily on culturing methods, which fail to detect nonculturable bacteria. Identification of novel fluorescent probes for biofilm imaging will greatly...... facilitate diagnosis of pathogenic bacterial infection. Herein, we report a novel fluorescent probe, CDy11 (compound of designation yellow 11), which targets amyloid in the Pseudomonas aeruginosa biofilm matrix through a diversity oriented fluorescent library approach (DOFLA). CDy11 was further demonstrated...

  13. Antibiofilm potential of flavonoids extracted from Moringa oleifera seed coat against Staphylococcus aureus, Pseudomonas aeruginosa and Candida albicans.

    Science.gov (United States)

    Onsare, J G; Arora, D S

    2015-02-01

    The increased microbial drug resistance due to biofilms and the side effects associated with the use of conventional drugs is still a major concern in the medical fraternity. This work evaluates the antibiofilm potential of flavonoids extracted from Moringa oleifera seed coat (SC) in search for green and effective alternatives for overcoming menace of biofilms. The study evaluated the minimum inhibitory concentration (MIC) of flavonoids against respective test organisms, inhibition of initial cell attachment as well as disruption of preformed biofilms and metabolic activity of treated biofilms. Mutagenicity and cytotoxicity as well as characterization of the active component were also carried out. Although Pseudomonas aeruginosa showed the lowest MIC of 0.05 mg ml(-1), the action of flavonoids and gentamicin on initial cell attachment revealed a comparable effect against bacterial biofilms, i.e. Staphylococcus aureus and Pseudomonas aeruginosa with approx. 80% inhibition compared to Candida albicans. Disruption of the preformed biofilms revealed that susceptibility of P. aeruginosa began as early as 4 h of exposure to flavonoids with 88% growth inhibition at the end of 24-h incubation. Encouragingly, t-test analysis on the effect of the extract and the standard antibiotic against each organism indicated no significant variance at P < 0.05. A drastic low metabolic activity exhibited by the treated biofilms as compared to the untreated ones was further supportive of the antibiofilm potential of seed coat flavonoids. The bioactive component from M. oleifera seed coat has exhibited antibiofilm potential against the test organisms belonging to Gram positive, Gram negative and yeast. Antibiofilm potential and biosafety of plant-based flavonoids from M. oleifera seed coat reveal a prospective active principle that could be of use in biofilm-associated menace. © 2014 The Society for Applied Microbiology.

  14. Blue light enhances the antimicrobial activity of honey against Pseudomonas aeruginosa

    Science.gov (United States)

    Orlandi, Viviana Teresa; Bolognese, Fabrizio; Barbieri, Paola

    2018-02-01

    Pseudomonas aeruginosa may be isolated from skin wounds of burn patients, bedsore and diabetic ulcers. The healing of wounds is often impaired by the intrinsic antibiotic resistance, the tolerance to many antimicrobials and the ability to form biofilm of this opportunistic pathogen. Finding new topical treatments to combine with antibiotics is thus essential. Among natural products, the antimicrobial properties of honeys have been known for millennia. In this study honey and visible light have been combined to control the growth of P. aeruginosa PAO1. The irradiation by a broad spectrum light source of bacteria inoculated onto 2 % w/v fir and forest honeydew (HD) honeys caused a killing effect that the honeys alone or the light alone did not show. This antimicrobial activity was light energy-dose and honey-concentration dependent. Among the tested honeys, the fir and forest HD honeys were the most efficient ones. In particular, the irradiation by blue LED (λmax = 466 nm) yielded good rates of killing, that were significantly higher in comparison to irradiation alone and honey alone. Interestingly, a similar effect was obtained by plating bacteria on blue LED pre-irradiated HD honeys. The combined use of honey and blue light was also successful in inhibiting the biofilm formation of P. aeruginosa. The blue LED irradiation of PAO1 administered with 10 % w/v forest HD honey significantly enhanced the inhibition of biofilm formation in comparison to dark incubated honey.

  15. Biofilm initiation and growth of Pseudomonas aeruginosa on 316L stainless steel in low gravity in orbital space flight

    Science.gov (United States)

    Todd, Paul; Pierson, Duane L.; Allen, Britt; Silverstein, JoAnn

    The formation of biofilms by water microorganisms such as Pseudomonas aeruginosa in spacecraft water systems has been a matter of concern for long-duration space flight. Crewed spacecraft plumbing includes internal surfaces made of 316L stainless steel. Experiments were therefore undertaken to compare the ability of P. aeruginosa to grow in suspension, attach to stainless steel and to grow on stainless steel in low gravity on the space shuttle. Four categories of cultures were studied during two space shuttle flights (STS-69 and STS-77). Cultures on the ground were held in static horizontal or vertical cylindrical containers or were tumbled on a clinostat and activated under conditions identical to those for the flown cultures. The containers used on the ground and in flight were BioServe Space Technologies’ Fluid Processing Apparatus (FPA), an open-ended test tube with rubber septa that allows robotic addition of bacteria to culture media to initiate experiments and the addition of fixative to conclude experiments. Planktonic growth was monitored by spectrophotometry, and biofilms were characterized quantitatively by epifluorescence and scanning electron microscopy. In these experiments it was found that: (1) Planktonic growth in flown cultures was more extensive than in static cultures, as seen repeatedly in the history of space microbiology, and closely resembled the growth of tumbled cultures. (2) Conversely, the attachment of cells in flown cultures was as much as 8 times that in tumbled cultures but not significantly different from that in static horizontal and vertical cultures, consistent with the notion that flowing fluid reduces microbial attachment. (3) The final surface coverage in 8 days was the same for flown and static cultures but less by a factor of 15 in tumbled cultures, where coverage declined during the preceding 4 days. It is concluded that cell attachment to 316L stainless steel in the low gravity of orbital space flight is similar to that

  16. Wild Mushroom Extracts as Inhibitors of Bacterial Biofilm Formation

    Directory of Open Access Journals (Sweden)

    Maria José Alves

    2014-08-01

    Full Text Available Microorganisms can colonize a wide variety of medical devices, putting patients in risk for local and systemic infectious complications, including local-site infections, catheter-related bloodstream infections, and endocarditis. These microorganisms are able to grow adhered to almost every surface, forming architecturally complex communities termed biofilms. The use of natural products has been extremely successful in the discovery of new medicine, and mushrooms could be a source of natural antimicrobials. The present study reports the capacity of wild mushroom extracts to inhibit in vitro biofilm formation by multi-resistant bacteria. Four Gram-negative bacteria biofilm producers (Escherichia coli, Proteus mirabilis, Pseudomonas aeruginosa, and Acinetobacter baumannii isolated from urine were used to verify the activity of Russula delica, Fistulina hepatica, Mycena rosea, Leucopaxilus giganteus, and Lepista nuda extracts. The results obtained showed that all tested mushroom extracts presented some extent of inhibition of biofilm production. Pseudomonas aeruginosa was the microorganism with the highest capacity of biofilm production, being also the most susceptible to the extracts inhibition capacity (equal or higher than 50%. Among the five tested extracts against E. coli, Leucopaxillus giganteus (47.8% and Mycenas rosea (44.8% presented the highest inhibition of biofilm formation. The extracts exhibiting the highest inhibitory effect upon P. mirabilis biofilm formation were Sarcodon imbricatus (45.4% and Russula delica (53.1%. Acinetobacter baumannii was the microorganism with the lowest susceptibility to mushroom extracts inhibitory effect on biofilm production (highest inhibition—almost 29%, by Russula delica extract. This is a pioneer study since, as far as we know, there are no reports on the inhibition of biofilm production by the studied mushroom extracts and in particular against multi-resistant clinical isolates; nevertheless, other

  17. Ellagic acid derivatives from Terminalia chebula Retz. downregulate the expression of quorum sensing genes to attenuate Pseudomonas aeruginosa PAO1 virulence.

    Directory of Open Access Journals (Sweden)

    Sajal Sarabhai

    Full Text Available BACKGROUND: Burgeoning antibiotic resistance in Pseudomonas aeruginosa has necessitated the development of anti pathogenic agents that can quench acylhomoserine lactone (AHL mediated QS with least risk of resistance. This study explores the anti quorum sensing potential of T. chebula Retz. and identification of probable compounds(s showing anti QS activity and the mechanism of attenuation of P. aeruginosa PAO1 virulence factors. METHODS AND RESULTS: Methanol extract of T. chebula Retz. fruit showed anti QS activity using Agrobacterium tumefaciens A136. Bioactive fraction (F7, obtained by fractionation of methanol extract using Sephadex LH20, showed significant reduction (p<0.001 in QS regulated production of extracellular virulence factors in P. aeruginosa PAO1. Biofilm formation and alginate were significantly (p<0.05 reduced with enhanced (20% susceptibility to tobramycin. Real Time PCR of F7 treated P. aeruginosa showed down regulation of autoinducer synthase (lasI and rhlI and their cognate receptor (lasR and rhlR genes by 89, 90, 90 and 93%, respectively. Electrospray Ionization Mass Spectrometry also showed 90 and 64% reduction in the production of 3-oxo-C(12HSL and C(4HSL after treatment. Decrease in AHLs as one of the mechanisms of quorum quenching by F7 was supported by the reversal of inhibited swarming motility in F7-treated P. aeruginosa PAO1 on addition of C(4HSL. F7 also showed antagonistic activity against 3-oxo-C(12HSL-dependent QS in E. coli bioreporter. C. elegans fed on F7-treated P. aeruginosa showed enhanced survival with LT50 increasing from 24 to 72 h. LC-ESI-MS of F7 revealed the presence of ellagic acid derivatives responsible for anti QS activity in T. chebula extract. CONCLUSIONS: This is the first report on anti QS activity of T. chebula fruit linked to EADs which down regulate the expression of lasIR and rhlIR genes with concomitant decrease in AHLs in P. aeruginosa PAO1 causing attenuation of its virulence factors

  18. Pseudomonas aeruginosa disrupts Caenorhabditis elegans iron homeostasis, causing a hypoxic response and death.

    Science.gov (United States)

    Kirienko, Natalia V; Kirienko, Daniel R; Larkins-Ford, Jonah; Wählby, Carolina; Ruvkun, Gary; Ausubel, Frederick M

    2013-04-17

    The opportunistic pathogen Pseudomonas aeruginosa causes serious human infections, but effective treatments and the mechanisms mediating pathogenesis remain elusive. Caenorhabditis elegans shares innate immune pathways with humans, making it invaluable to investigate infection. To determine how P. aeruginosa disrupts host biology, we studied how P. aeruginosa kills C. elegans in a liquid-based pathogenesis model. We found that P. aeruginosa-mediated killing does not require quorum-sensing pathways or host colonization. A chemical genetic screen revealed that iron chelators alleviate P. aeruginosa-mediated killing. Consistent with a role for iron in P. aeruginosa pathogenesis, the bacterial siderophore pyoverdin was required for virulence and was sufficient to induce a hypoxic response and death in the absence of bacteria. Loss of the C. elegans hypoxia-inducing factor HIF-1, which regulates iron homeostasis, exacerbated P. aeruginosa pathogenesis, further linking hypoxia and killing. As pyoverdin is indispensable for virulence in mice, pyoverdin-mediated hypoxia is likely to be relevant in human pathogenesis. Copyright © 2013 Elsevier Inc. All rights reserved.

  19. Biofilm attachment reduction on bioinspired, dynamic, micro-wrinkling surfaces

    International Nuclear Information System (INIS)

    Epstein, Alexander K; Hong, Donggyoon; Kim, Philseok; Aizenberg, Joanna

    2013-01-01

    Most bacteria live in multicellular communities known as biofilms that are adherent to surfaces in our environment, from sea beds to plumbing systems. Biofilms are often associated with clinical infections, nosocomial deaths and industrial damage such as bio-corrosion and clogging of pipes. As mature biofilms are extremely challenging to eradicate once formed, prevention is advantageous over treatment. However, conventional surface chemistry strategies are either generally transient, due to chemical masking, or toxic, as in the case of leaching marine antifouling paints. Inspired by the nonfouling skins of echinoderms and other marine organisms, which possess highly dynamic surface structures that mechanically frustrate bio-attachment, we have developed and tested a synthetic platform based on both uniaxial mechanical strain and buckling-induced elastomer microtopography. Bacterial biofilm attachment to the dynamic substrates was studied under an array of parameters, including strain amplitude and timescale (1–100 mm s −1 ), surface wrinkle length scale, bacterial species and cell geometry, and growth time. The optimal conditions for achieving up to ∼ 80% Pseudomonas aeruginosa biofilm reduction after 24 h growth and ∼ 60% reduction after 48 h were combinatorially elucidated to occur at 20% strain amplitude, a timescale of less than ∼ 5 min between strain cycles and a topography length scale corresponding to the cell dimension of ∼ 1 μm. Divergent effects on the attachment of P. aeruginosa, Staphylococcus aureus and Escherichia coli biofilms showed that the dynamic substrate also provides a new means of species-specific biofilm inhibition, or inversely, selection for a desired type of bacteria, without reliance on any toxic or transient surface chemical treatments. (paper)

  20. Biofilm attachment reduction on bioinspired, dynamic, micro-wrinkling surfaces

    Science.gov (United States)

    Epstein, Alexander K.; Hong, Donggyoon; Kim, Philseok; Aizenberg, Joanna

    2013-09-01

    Most bacteria live in multicellular communities known as biofilms that are adherent to surfaces in our environment, from sea beds to plumbing systems. Biofilms are often associated with clinical infections, nosocomial deaths and industrial damage such as bio-corrosion and clogging of pipes. As mature biofilms are extremely challenging to eradicate once formed, prevention is advantageous over treatment. However, conventional surface chemistry strategies are either generally transient, due to chemical masking, or toxic, as in the case of leaching marine antifouling paints. Inspired by the nonfouling skins of echinoderms and other marine organisms, which possess highly dynamic surface structures that mechanically frustrate bio-attachment, we have developed and tested a synthetic platform based on both uniaxial mechanical strain and buckling-induced elastomer microtopography. Bacterial biofilm attachment to the dynamic substrates was studied under an array of parameters, including strain amplitude and timescale (1-100 mm s-1), surface wrinkle length scale, bacterial species and cell geometry, and growth time. The optimal conditions for achieving up to ˜ 80% Pseudomonas aeruginosa biofilm reduction after 24 h growth and ˜ 60% reduction after 48 h were combinatorially elucidated to occur at 20% strain amplitude, a timescale of less than ˜ 5 min between strain cycles and a topography length scale corresponding to the cell dimension of ˜ 1 μm. Divergent effects on the attachment of P. aeruginosa, Staphylococcus aureus and Escherichia coli biofilms showed that the dynamic substrate also provides a new means of species-specific biofilm inhibition, or inversely, selection for a desired type of bacteria, without reliance on any toxic or transient surface chemical treatments.

  1. Tolerance to the antimicrobial peptide colistin in Pseudomonas aeruginosa biofilms is linked to metabolically active cells, and depends on the pmr and mexAB-oprM genes

    DEFF Research Database (Denmark)

    Pamp, Sünje Johanna; Gjermansen, Morten; Johansen, Helle Krogh

    2008-01-01

    -mediated killing in biofilms, conventional antimicrobial compounds such as ciprofloxacin and tetracycline were found to specifically kill the subpopulation of metabolically active biofilm cells, whereas the subpopulation exhibiting low metabolic activity survived the treatment. Consequently, targeting the two...... physiologically distinct subpopulations by combined antimicrobial treatment with either ciprofloxacin and colistin or tetracycline and colistin almost completely eradicated all biofilm cells....

  2. Biofilm formation by clinical isolates and the implications in chronic infections

    Directory of Open Access Journals (Sweden)

    Sanchez Carlos J

    2013-01-01

    Full Text Available Abstract Background Biofilm formation is a major virulence factor contributing to the chronicity of infections. To date few studies have evaluated biofilm formation in infecting isolates of patients including both Gram-positive and Gram-negative multidrug-resistant (MDR species in the context of numerous types of infectious syndromes. Herein, we investigated the biofilm forming capacity in a large collection of single patient infecting isolates and compared the relationship between biofilm formation to various strain characteristics. Methods The biofilm-forming capacity of 205 randomly sampled clinical isolates from patients, collected from various anatomical sites, admitted for treatment at Brooke Army Medical Center (BAMC from 2004–2011, including methicillin-resistant/methicillin susceptible Staphylococcus aureus (MRSA/MSSA (n=23, Acinetobacter baumannii (n=53, Pseudomonas aeruginosa (n=36, Klebsiella pneumoniae (n=54, and Escherichia coli (n=39, were evaluated for biofilm formation using the high-throughput microtiter plate assay and scanning electron microscopy (SEM. Relationships between biofilm formation to clonal type, site of isolate collection, and MDR phenotype were evaluated. Furthermore, in patients with relapsing infections, serial strains were assessed for their ability to form biofilms in vitro. Results Of the 205 clinical isolates tested, 126 strains (61.4% were observed to form biofilms in vitro at levels greater than or equal to the Staphylococcus epidermidis, positive biofilm producing strain, with P. aeruginosa and S. aureus having the greatest number of biofilm producing strains. Biofilm formation was significantly associated with specific clonal types, the site of isolate collection, and strains positive for biofilm formation were more frequently observed to be MDR. In patients with relapsing infections, the majority of serial isolates recovered from these individuals were observed to be strong biofilm producers in vitro

  3. BACTERIAL BIOFILM FORMATION UNDER MICROGRAVITY CONDITIONS. (R825503)

    Science.gov (United States)

    Although biofilm formation is widely documented on Earth, it has not been demonstrated in the absence of gravity. To explore this possibility, Pseudomonas aeruginosa, suspended in sterile buffer, was flown in a commercial payload on space shuttle flight STS-95. During earth or...

  4. Pseudomonas biofilm matrix composition and niche biology

    Science.gov (United States)

    Mann, Ethan E.; Wozniak, Daniel J.

    2014-01-01

    Biofilms are a predominant form of growth for bacteria in the environment and in the clinic. Critical for biofilm development are adherence, proliferation, and dispersion phases. Each of these stages includes reinforcement by, or modulation of, the extracellular matrix. Pseudomonas aeruginosa has been a model organism for the study of biofilm formation. Additionally, other Pseudomonas species utilize biofilm formation during plant colonization and environmental persistence. Pseudomonads produce several biofilm matrix molecules, including polysaccharides, nucleic acids, and proteins. Accessory matrix components shown to aid biofilm formation and adaptability under varying conditions are also produced by pseudomonads. Adaptation facilitated by biofilm formation allows for selection of genetic variants with unique and distinguishable colony morphology. Examples include rugose small-colony variants and wrinkly spreaders (WS), which over produce Psl/Pel or cellulose, respectively, and mucoid bacteria that over produce alginate. The well-documented emergence of these variants suggests that pseudomonads take advantage of matrix-building subpopulations conferring specific benefits for the entire population. This review will focus on various polysaccharides as well as additional Pseudomonas biofilm matrix components. Discussions will center on structure–function relationships, regulation, and the role of individual matrix molecules in niche biology. PMID:22212072

  5. Synergistic Effect of 14-Alpha-Lipoyl Andrographolide and Various Antibiotics on the Formation of Biofilms and Production of Exopolysaccharide and Pyocyanin by Pseudomonas aeruginosa▿

    OpenAIRE

    Zeng, Xiangping; Liu, Xiangyang; Bian, Jiang; Pei, Gang; Dai, Huanqin; Polyak, Steven W.; Song, Fuhang; Ma, Li; Wang, Yuqiang; Zhang, Lixin

    2011-01-01

    Pseudomonas aeruginosa produces a biofilm that provides the bacteria with an effective barrier against antibiotics. Here, we investigated the synergy of various antibiotics with 14-alpha-lipoyl andrographolide (AL-1), focusing upon synthesis of the biofilm. AL-1 also inhibited the production of the exopolysaccharide and pyocyanin components. We propose that AL-1 may potentially serve as a cotherapy to combat P. aeruginosa.

  6. The contribution of cell-cell signaling and motility to bacterial biofilm formation

    DEFF Research Database (Denmark)

    Shrout, Joshua D; Tolker-Nielsen, Tim; Givskov, Michael

    2011-01-01

    Many bacteria grow attached to a surface as biofilms. Several factors dictate biofilm formation, including responses by the colonizing bacteria to their environment. Here we review how bacteria use cell-cell signaling (also called quorum sensing) and motility during biofilm formation. Specifically...... gene expression important to the production of polysaccharides, rhamnolipid, and other virulence factors. Surface motility affects the assembly and architecture of biofilms, and some aspects of motility are also influenced by quorum sensing. While some genes and their function are specific to P....... aeruginosa, many aspects of biofilm development can be used as a model system to understand how bacteria differentially colonize surfaces....

  7. Next generation in vitro systems for biofilm studies - a cystic fibrosis patient airway perspective

    DEFF Research Database (Denmark)

    Weiss Nielsen, Martin

    proven essential aspects on biofilm formations, however generates highly artificial biofilms that lack several CF airway scenarios. The driving force and the heart of this project has its origin in the study of the role played by P. aeruginosa in the CF airways. One of the aims of this thesis...

  8. Extracts of Cordia gilletii de wild (Boraginaceae) quench the quorum sensing of Pseudomonas aeruginosa PAO1.

    Science.gov (United States)

    Okusa, Philippe N; Rasamiravaka, Tsiry; Vandeputte, Olivier; Stévigny, Caroline; Jaziri, Mondher El; Duez, Pierre

    2014-01-01

    The fight against infectious diseases and antimicrobial resistances needs the exploration of new active compounds with new proprieties like disrupting quorum sensing (QS) mechanisms, which is a cell-to-cell communication that regulates bacterial virulence factors. In this work, leaves and root barks extracts of a Congolese medicinal plant, Cordia gilletii, were investigated for their effect on the production of Pseudomonas aeruginosa major virulence factors regulated by QS. The effect of C. gilletii extracts on virulence factors of P. aeruginosa PAO1 was studied by the evaluation of the production of pyocyanine, elastase and biofilm; and by the measurement of the expression of QS-related genes. The dichloromethane extract from root barks was found to quench the production of pyocyanin, a QS-dependent virulence factor in P. aeruginosa PAO1. Moreover, this extract specifically inhibits the expression of several QS-regulated genes (i.e. lasB, rhlA, lasI, lasR, rhlI, and rhlR) and reduces biofilm formation by PAO1. This study contributes to explain the efficacy of C. gilletii in the traditional treatment of infectious diseases caused by P. aeruginosa.

  9. Extracts of Cordia gilletii de wild (Boraginaceae) quench the quorum sensing of Pseudomonas aeruginosa PAO1

    Science.gov (United States)

    Okusa, Philippe N.; Rasamiravaka, Tsiry; Vandeputte, Olivier; Stévigny, Caroline; Jaziri, Mondher El; Duez, Pierre

    2014-01-01

    Aim: The fight against infectious diseases and antimicrobial resistances needs the exploration of new active compounds with new proprieties like disrupting quorum sensing (QS) mechanisms, which is a cell-to-cell communication that regulates bacterial virulence factors. In this work, leaves and root barks extracts of a Congolese medicinal plant, Cordia gilletii, were investigated for their effect on the production of Pseudomonas aeruginosa major virulence factors regulated by QS. Materials and Methods: The effect of C. gilletii extracts on virulence factors of P. aeruginosa PAO1 was studied by the evaluation of the production of pyocyanine, elastase and biofilm; and by the measurement of the expression of QS-related genes. Results: The dichloromethane extract from root barks was found to quench the production of pyocyanin, a QS-dependent virulence factor in P. aeruginosa PAO1. Moreover, this extract specifically inhibits the expression of several QS-regulated genes (i.e. lasB, rhlA, lasI, lasR, rhlI, and rhlR) and reduces biofilm formation by PAO1. Conclusion: This study contributes to explain the efficacy of C. gilletii in the traditional treatment of infectious diseases caused by P. aeruginosa. PMID:26401363

  10. Initial development and structure of biofilms on microbial fuel cell anodes

    Directory of Open Access Journals (Sweden)

    Keller Jürg

    2010-04-01

    Full Text Available Abstract Background Microbial fuel cells (MFCs rely on electrochemically active bacteria to capture the chemical energy contained in organics and convert it to electrical energy. Bacteria develop biofilms on the MFC electrodes, allowing considerable conversion capacity and opportunities for extracellular electron transfer (EET. The present knowledge on EET is centred around two Gram-negative models, i.e. Shewanella and Geobacter species, as it is believed that Gram-positives cannot perform EET by themselves as the Gram-negatives can. To understand how bacteria form biofilms within MFCs and how their development, structure and viability affects electron transfer, we performed pure and co-culture experiments. Results Biofilm viability was maintained highest nearer the anode during closed circuit operation (current flowing, in contrast to when the anode was in open circuit (soluble electron acceptor where viability was highest on top of the biofilm, furthest from the anode. Closed circuit anode Pseudomonas aeruginosa biofilms were considerably thinner compared to the open circuit anode (30 ± 3 μm and 42 ± 3 μm respectively, which is likely due to the higher energetic gain of soluble electron acceptors used. The two Gram-positive bacteria used only provided a fraction of current produced by the Gram-negative organisms. Power output of co-cultures Gram-positive Enterococcus faecium and either Gram-negative organisms, increased by 30-70% relative to the single cultures. Over time the co-culture biofilms segregated, in particular, Pseudomonas aeruginosa creating towers piercing through a thin, uniform layer of Enterococcus faecium. P. aeruginosa and E. faecium together generated a current of 1.8 ± 0.4 mA while alone they produced 0.9 ± 0.01 and 0.2 ± 0.05 mA respectively. Conclusion We postulate that this segregation may be an essential difference in strategy for electron transfer and substrate capture between the Gram-negative and the Gram

  11. Garlic blocks quorum sensing and promotes rapid clearing of pulmonary Pseudomonas aeruginosa infections

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Jensen, Peter Østrup; Rasmussen, Thomas B

    2005-01-01

    to otherwise lethal doses of antibiotics, and protects against the bactericidal activity of polymorphonuclear leukocytes (PMNs). It has been previously demonstrated that QS is inhibited by garlic extract. In this study, the synergistic effects of garlic and tobramycin, and PMNs activities have been evaluated....... P. aeruginosa was grown in vitro in continuous-culture once-through flow chambers with and without garlic extract. The garlic-treated biofilms were susceptible to both tobramycin and PMN grazing. Furthermore, the PMNs showed an increase in respiratory burst activation, when incubated with the garlic......-treated biofilm. Garlic extract was administered as treatment for a mouse pulmonary infection model. Mice were treated with garlic extract or placebo for 7 days, with the initial 2 days being prophylactic before P. aeruginosa was instilled in the left lung of the mice. Bacteriology, mortality, histopathology...

  12. Anti-biofilm and cytotoxicity activity of impregnated dressings with silver nanoparticles

    International Nuclear Information System (INIS)

    Velázquez-Velázquez, Jorge Luis; Santos-Flores, Andrés; Araujo-Meléndez, Javier; Sánchez-Sánchez, Roberto; Velasquillo, Cristina; González, Carmen; Martínez-Castañon, Gabriel; Martinez-Gutierrez, Fidel

    2015-01-01

    Infections arising from bacterial adhesion and colonization on chronic wounds are a significant healthcare problem. Silver nanoparticles (AgNPs) impregnated in dressing have attracted a great deal of attention as a potential solution. The goal of the present study was to evaluate the anti-biofilm activities of AgNPs impregnated in commercial dressings against Pseudomonas aeruginosa, bacteria isolated of chronic wounds from a hospital patient. The antimicrobial activity of AgNPs was tested within biofilms generated under slow fluid shear conditions using a standard bioreactor. A 2-log reduction in the number of colony-forming units of P. aeruginosa was recorded in the reactor on exposure to dressing impregnated with 250 ppm of AgNPs, diameter 9.3 ± 1.1 nm, and also showed compatibility to mammalian cells (human fibroblasts). Our study suggests that the use of dressings with AgNPs may either prevent or reduce microbial growth in the wound environment, and reducing wound bioburden may improve wound-healing outcomes. - Highlights: • Biological activities of silver nanoparticles for wound-healing purposes • Characterization of the silver nanoparticles impregnated in dressings • Reduction in the P. aeruginosa biofilm formation was statistically significant. • Compatibility to human dermal fibroblasts as the main cell type involved in the reparation • AgNPs covering the surfaces would provide great potential for prevention and treatment

  13. Anti-biofilm and cytotoxicity activity of impregnated dressings with silver nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Velázquez-Velázquez, Jorge Luis [Laboratorio de Microbiología, Facultad de Ciencias Químicas, UASLP (Mexico); Santos-Flores, Andrés; Araujo-Meléndez, Javier [Servicio de Epidemiología del Hospital Central “Dr. Ignacio Morones Prieto”, San Luis Potosí (Mexico); Sánchez-Sánchez, Roberto; Velasquillo, Cristina [Laboratorio de Biotecnología, Instituto Nacional de Rehabilitación (Mexico); González, Carmen [Laboratorio de Fisiología Celular, Facultad de Ciencias Químicas, UASLP (Mexico); Martínez-Castañon, Gabriel [Maestría en Ciencias Odontológicas Facultad de Estomatología, UASLP (Mexico); Martinez-Gutierrez, Fidel, E-mail: fidel@uaslp.mx [Laboratorio de Microbiología, Facultad de Ciencias Químicas, UASLP (Mexico)

    2015-04-01

    Infections arising from bacterial adhesion and colonization on chronic wounds are a significant healthcare problem. Silver nanoparticles (AgNPs) impregnated in dressing have attracted a great deal of attention as a potential solution. The goal of the present study was to evaluate the anti-biofilm activities of AgNPs impregnated in commercial dressings against Pseudomonas aeruginosa, bacteria isolated of chronic wounds from a hospital patient. The antimicrobial activity of AgNPs was tested within biofilms generated under slow fluid shear conditions using a standard bioreactor. A 2-log reduction in the number of colony-forming units of P. aeruginosa was recorded in the reactor on exposure to dressing impregnated with 250 ppm of AgNPs, diameter 9.3 ± 1.1 nm, and also showed compatibility to mammalian cells (human fibroblasts). Our study suggests that the use of dressings with AgNPs may either prevent or reduce microbial growth in the wound environment, and reducing wound bioburden may improve wound-healing outcomes. - Highlights: • Biological activities of silver nanoparticles for wound-healing purposes • Characterization of the silver nanoparticles impregnated in dressings • Reduction in the P. aeruginosa biofilm formation was statistically significant. • Compatibility to human dermal fibroblasts as the main cell type involved in the reparation • AgNPs covering the surfaces would provide great potential for prevention and treatment.

  14. Antibiofilm Effect of DNase against Single and Mixed Species Biofilm

    Science.gov (United States)

    Sharma, Komal

    2018-01-01

    Biofilms are aggregates of microorganisms that coexist in socially coordinated micro-niche in a self-produced polymeric matrix on pre-conditioned surfaces. The biofilm matrix reduces the efficacy of antibiofilm strategies. DNase degrades the extracellular DNA (e-DNA) present in the matrix, rendering the matrix weak and susceptible to antimicrobials. In the current study, the effect of DNase I was evaluated during biofilm formation (pre-treatment), on preformed biofilms (post-treatment) and both (dual treatment). The DNase I pre-treatment was optimized for P. aeruginosa PAO1 (model biofilm organism) at 10 µg/mL and post-treatment at 10 µg/mL with 15 min of contact duration. Inclusion of Mg2+ alongside DNase I post-treatment resulted in 90% reduction in biofilm within only 5 min of contact time (irrespective of age of biofilm). On extension of these findings, DNase I was found to be less effective against mixed species biofilm than individual biofilms. DNase I can be used as potent antibiofilm agent and with further optimization can be effectively used for biofilm prevention and reduction in situ. PMID:29562719

  15. Sterilization of Biofilm on a Titanium Surface Using a Combination of Nonthermal Plasma and Chlorhexidine Digluconate

    Directory of Open Access Journals (Sweden)

    Tripti Thapa Gupta

    2017-01-01

    Full Text Available Nosocomial infections caused by opportunistic bacteria pose major healthcare problem worldwide. Out of the many microorganisms responsible for such infections, Pseudomonas aeruginosa is a ubiquitous bacterium that accounts for 10–20% of hospital-acquired infections. These infections have mortality rates ranging from 18 to 60% and the cost of treatment ranges from $20,000 to $80,000 per infection. The formation of biofilms on medical devices and implants is responsible for the majority of those infections. Only limited progress has been made to prevent this issue in a safe and cost-effective manner. To address this, we propose employing jet plasma to break down and inactivate biofilms in vitro. Moreover, to improve the antimicrobial effect on the biofilm, a treatment method using a combination of jet plasma and a biocide known as chlorhexidine (CHX digluconate was investigated. We found that complete sterilization of P. aeruginosa biofilms can be achieved after combinatorial treatment using plasma and CHX. A decrease in biofilm viability was also observed using confocal laser scanning electron microscopy (CLSM. This treatment method sterilized biofilm-contaminated surfaces in a short treatment time, indicating it to be a potential tool for the removal of biofilms present on medical devices and implants.

  16. Srv mediated dispersal of streptococcal biofilms through SpeB is observed in CovRS+ strains.

    Directory of Open Access Journals (Sweden)

    Kristie L Connolly

    Full Text Available Group A Streptococcus (GAS is a human specific pathogen capable of causing both mild infections and severe invasive disease. We and others have shown that GAS is able to form biofilms during infection. That is to say, they form a three-dimensional, surface attached structure consisting of bacteria and a multi-component extracellular matrix. The mechanisms involved in regulation and dispersal of these GAS structures are still unclear. Recently we have reported that in the absence of the transcriptional regulator Srv in the MGAS5005 background, the cysteine protease SpeB is constitutively produced, leading to increased tissue damage and decreased biofilm formation during a subcutaneous infection in a mouse model. This was interesting because MGAS5005 has a naturally occurring mutation that inactivates the sensor kinase domain of the two component regulatory system CovRS. Others have previously shown that strains lacking covS are associated with decreased SpeB production due to CovR repression of speB expression. Thus, our results suggest the inactivation of srv can bypass CovR repression and lead to constitutive SpeB production. We hypothesized that Srv control of SpeB production may be a mechanism to regulate biofilm dispersal and provide a mechanism by which mild infection can transition to severe disease through biofilm dispersal. The question remained however, is this mechanism conserved among GAS strains or restricted to the unique genetic makeup of MGAS5005. Here we show that Srv mediated control of SpeB and biofilm dispersal is conserved in the invasive clinical isolates RGAS053 (serotype M1 and MGAS315 (serotype M3, both of which have covS intact. This work provides additional evidence that Srv regulated control of SpeB may mediate biofilm formation and dispersal in diverse strain backgrounds.

  17. Correlative Imaging of Structural and Elemental Composition of Bacterial Biofilms

    International Nuclear Information System (INIS)

    Yang, Y; Heine, R; Xu, F; Helfen, L; Baumbach, T; Suhonen, H; Rosenhahn, A; Gorniak, T; Kirchen, S; Schwartz, T

    2013-01-01

    Synchrotron-based phase contrast tomography (holotomography) and scanning hard X-ray fluorescence microscopy (SXFM) are combined to characterize the three-dimensional (3D) structural and corresponding elemental distribution of bacterial biofilms of Pseudomonas aeruginosa. Samples were fixed without contrast agents or microtomal sectioning. Within an intact microbial community single bacteria are clearly resolved, and their morphology can be directly visualized together with the elemental content. Such 3D set of complementary information at cellular level is essential for gaining a deeper understanding of biofilm evolution aiming to develop potential strategies on biofilm growth control and prevention

  18. The Yin and Yang of SagS: Distinct Residues in the HmsP Domain of SagS Independently Regulate Biofilm Formation and Biofilm Drug Tolerance

    Science.gov (United States)

    Dingemans, Jozef; Poudyal, Bandita

    2018-01-01

    ABSTRACT The formation of inherently drug-tolerant biofilms by the opportunistic pathogen Pseudomonas aeruginosa requires the sensor-regulator hybrid SagS, with ΔsagS biofilms being unstructured and exhibiting increased antimicrobial susceptibility. Recent findings indicated SagS to function as a switch to control biofilm formation and drug tolerance independently. Moreover, findings suggested the periplasmic sensory HmsP domain of SagS is likely to be the control point in the regulation of biofilm formation and biofilm cells transitioning to a drug-tolerant state. We thus asked whether specific amino acid residues present in the HmsP domain contribute to the switch function of SagS. HmsP domain residues were therefore subjected to alanine replacement mutagenesis to identify substitutions that block the sensory function(s) of SagS, which is apparent by attached cells being unable to develop mature biofilms and/or prevent transition to an antimicrobial-resistant state. Mutant analyses revealed 32 residues that only contribute to blocking one sensory function. Moreover, amino acid residues affecting attachment and subsequent biofilm formation but not biofilm tolerance also impaired histidine kinase signaling via BfiS. In contrast, residues affecting biofilm drug tolerance but not attachment and subsequent biofilm formation negatively impacted BrlR transcription factor levels. Structure prediction suggested the two sets of residues affecting sensory functions are located in distinct areas that were previously described as being involved in ligand binding interactions. Taken together, these studies identify the molecular basis for the dual regulatory function of SagS. IMPORTANCE The membrane-bound sensory protein SagS plays a pivotal role in P. aeruginosa biofilm formation and biofilm cells gaining their heightened resistance to antimicrobial agents, with SagS being the control point at which both pathways diverge. Here, we demonstrate for the first time that the two

  19. In vitro antibiofilm efficacy of Piper betle against quorum sensing mediated biofilm formation of luminescent Vibrio harveyi.

    Science.gov (United States)

    Srinivasan, Ramanathan; Santhakumari, Sivasubramanian; Ravi, Arumugam Veera

    2017-09-01

    Vibrio harveyi is a potent biofilm former, which confers resistance to multiple antimicrobials, disinfectants, chemicals and biocides. The prevalence of biofilm mediated antibiotic resistance among aquatic bacterial pathogens stresses the search for novel alternative approach to treat vibriosis in aquaculture. Exploring suitable therapeutics from natural resources could be a novel area of research. Therefore, this work was executed to evaluate the inhibitory effect of Piper betle ethyl acetate extract (PBE) on bioluminescence production and biofilm formation of V. harveyi. Minimal inhibitory concentration (MIC) of PBE against planktonic V. harveyi was found to be 1600 μg ml -1 ; furthermore, PBE inhibited the quorum sensing (QS) mediated bioluminescence production and biofilm formation in V. harveyi upto 98 and 74% respectively, at its sub-MIC concentration of 400 μg ml -1 without affecting their cell viability. Similar results were obtained for exopolysaccharides production and swimming motility related to biofilm formation of V. harveyi, where PBE reduced EPS production upto 64%. Light and confocal laser scanning microscopic analyses further confirmed that the PBE effectively prevented the initial attachment as well as microcolonies formation of V. harveyi biofilm, when compared to their untreated controls. This study demonstrates the promising antibiofilm activity of PBE and confirms the ethnopharmacological potential of this plant against V. harveyi infections. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Interactions of methicillin resistant Staphylococcus aureus USA300 and Pseudomonas aeruginosa in polymicrobial wound infection.

    Directory of Open Access Journals (Sweden)

    Irena Pastar

    Full Text Available Understanding the pathology resulting from Staphylococcus aureus and Pseudomonas aeruginosa polymicrobial wound infections is of great importance due to their ubiquitous nature, increasing prevalence, growing resistance to antimicrobial agents, and ability to delay healing. Methicillin-resistant S. aureus USA300 is the leading cause of community-associated bacterial infections resulting in increased morbidity and mortality. We utilized a well-established porcine partial thickness wound healing model to study the synergistic effects of USA300 and P. aeruginosa on wound healing. Wound re-epithelialization was significantly delayed by mixed-species biofilms through suppression of keratinocyte growth factor 1. Pseudomonas showed an inhibitory effect on USA300 growth in vitro while both species co-existed in cutaneous wounds in vivo. Polymicrobial wound infection in the presence of P. aeruginosa resulted in induced expression of USA300 virulence factors Panton-Valentine leukocidin and α-hemolysin. These results provide evidence for the interaction of bacterial species within mixed-species biofilms in vivo and for the first time, the contribution of virulence factors to the severity of polymicrobial wound infections.

  1. Pseudomonas aeruginosa uses type III secretion system to kill biofilm-associated amoebae

    DEFF Research Database (Denmark)

    Matz, Carsten; Moreno, Ana Maria; Alhede, Morten

    2008-01-01

    Bacteria and protozoa coexist in a wide range of biofilm communities of natural, technical and medical importance. Generally, this interaction is characterized by the extensive grazing activity of protozoa on bacterial prey populations. We hypothesized that the close spatial coexistence in biofilms...... suggest that conserved virulence pathways and specifically the T3SS play a central role in bacteria- protozoa interactions in biofilms and may be instrumental for the environmental persistence and evolution of opportunistic bacterial pathogens....

  2. Polymorphonuclear leucocytes consume oxygen in sputum from chronic Pseudomonas aeruginosa pneumonia in cystic fibrosis

    DEFF Research Database (Denmark)

    Kolpen, Mette; Hansen, C. R.; Bjarnsholt, Thomas

    2010-01-01

    BACKGROUND: Chronic lung infection with Pseudomonas aeruginosa is the most severe complication for patients with cystic fibrosis (CF). This infection is characterised by endobronchial mucoid biofilms surrounded by numerous polymorphonuclear leucocytes (PMNs). The mucoid phenotype offers protection...

  3. The Effect of Cryopreserved Human Placental Tissues on Biofilm Formation of Wound-Associated Pathogens.

    Science.gov (United States)

    Mao, Yong; Singh-Varma, Anya; Hoffman, Tyler; Dhall, Sandeep; Danilkovitch, Alla; Kohn, Joachim

    2018-01-08

    Biofilm, a community of bacteria, is tolerant to antimicrobial agents and ubiquitous in chronic wounds. In a chronic DFU (Diabetic Foot Ulcers) clinical trial, the use of a human cryopreserved viable amniotic membrane (CVAM) resulted in a high rate of wound closure and reduction of wound-related infections. Our previous study demonstrated that CVAM possesses intrinsic antimicrobial activity against a spectrum of wound-associated bacteria under planktonic culture conditions. In this study, we evaluated the effect of CVAM and cryopreserved viable umbilical tissue (CVUT) on biofilm formation of S. aureus and P. aeruginosa , the two most prominent pathogens associated with chronic wounds. Firstly, we showed that, like CVAM, CVUT released antibacterial activity against multiple bacterial pathogens and the devitalization of CVUT reduced its antibacterial activity. The biofilm formation was then measured using a high throughput method and an ex vivo porcine dermal tissue model. We demonstrate that the formation of biofilm was significantly reduced in the presence of CVAM- or CVUT-derived conditioned media compared to control assay medium. The formation of P. aeruginosa biofilm on CVAM-conditioned medium saturated porcine dermal tissues was reduced 97% compared with the biofilm formation on the control medium saturated dermal tissues. The formation of S. auerus biofilm on CVUT-conditioned medium saturated dermal tissues was reduced 72% compared with the biofilm formation on the control tissues. This study is the first to show that human cryopreserved viable placental tissues release factors that inhibit biofilm formation. Our results provide an explanation for the in vivo observation of their ability to support wound healing.

  4. The Effect of Cryopreserved Human Placental Tissues on Biofilm Formation of Wound-Associated Pathogens

    Directory of Open Access Journals (Sweden)

    Yong Mao

    2018-01-01

    Full Text Available Biofilm, a community of bacteria, is tolerant to antimicrobial agents and ubiquitous in chronic wounds. In a chronic DFU (Diabetic Foot Ulcers clinical trial, the use of a human cryopreserved viable amniotic membrane (CVAM resulted in a high rate of wound closure and reduction of wound-related infections. Our previous study demonstrated that CVAM possesses intrinsic antimicrobial activity against a spectrum of wound-associated bacteria under planktonic culture conditions. In this study, we evaluated the effect of CVAM and cryopreserved viable umbilical tissue (CVUT on biofilm formation of S. aureus and P. aeruginosa, the two most prominent pathogens associated with chronic wounds. Firstly, we showed that, like CVAM, CVUT released antibacterial activity against multiple bacterial pathogens and the devitalization of CVUT reduced its antibacterial activity. The biofilm formation was then measured using a high throughput method and an ex vivo porcine dermal tissue model. We demonstrate that the formation of biofilm was significantly reduced in the presence of CVAM- or CVUT-derived conditioned media compared to control assay medium. The formation of P. aeruginosa biofilm on CVAM-conditioned medium saturated porcine dermal tissues was reduced 97% compared with the biofilm formation on the control medium saturated dermal tissues. The formation of S. auerus biofilm on CVUT-conditioned medium saturated dermal tissues was reduced 72% compared with the biofilm formation on the control tissues. This study is the first to show that human cryopreserved viable placental tissues release factors that inhibit biofilm formation. Our results provide an explanation for the in vivo observation of their ability to support wound healing.

  5. Effect of Mono and Di-rhamnolipids on Biofilms Pre-formed by Bacillus subtilis BBK006.

    Science.gov (United States)

    De Rienzo, Mayri A Díaz; Martin, Peter J

    2016-08-01

    Different microbial inhibition strategies based on the planktonic bacterial physiology have been known to have limited efficacy on the growth of biofilms communities. This problem can be exacerbated by the emergence of increasingly resistant clinical strains. Biosurfactants have merited renewed interest in both clinical and hygienic sectors due to their potential to disperse microbial biofilms. In this work, we explore the aspects of Bacillus subtilis BBK006 biofilms and examine the contribution of biologically derived surface-active agents (rhamnolipids) to the disruption or inhibition of microbial biofilms produced by Bacillus subtilis BBK006. The ability of mono-rhamnolipids (Rha-C10-C10) produced by Pseudomonas aeruginosa ATCC 9027 and the di-rhamnolipids (Rha-Rha-C14-C14) produced by Burkholderia thailandensis E264, and phosphate-buffered saline to disrupt biofilm of Bacillus subtilis BBK006 was evaluated. The biofilm produced by Bacillus subtilis BBK006 was more sensitive to the di-rhamnolipids (0.4 g/L) produced by Burkholderia thailandensis than the mono-rhamnolipids (0.4 g/L) produced by Pseudomonas aeruginosa ATCC 9027. Rhamnolipids are biologically produced compounds safe for human use. This makes them ideal candidates for use in new generations of bacterial dispersal agents and useful for use as adjuvants for existing microbial suppression or eradication strategies.

  6. Biofilm inhibition activity of compounds isolated from two Eunicea species collected at the Caribbean Sea

    Directory of Open Access Journals (Sweden)

    Yenny Martínez Díaz

    Full Text Available Abstract Biofilm has a primary role in the pathogenesis of diseases and in the attachment of multicellular organisms to a fouled surface. Because of that, the control of bacterial biofilms has been identified as an important target. In the present study, five lipid compounds isolated from soft coral Eunicea sp. and three terpenoids together with a mixture of sterols from Eunicea fusca collected at the Colombian Caribbean Sea showed different effectiveness against biofilm formation by three marine bacteria associated with immersed fouled surfaces, Ochrobactrum pseudogringnonense,Alteromona macleodii and Vibrio harveyi, and against two known biofilm forming bacteria, Pseudomonas aeruginosa ATCC 27853 and Staphylococcus aureus ATCC 25923. The pure compounds were characterized by NMR, HRESI-MS, HRGC-MS and optical rotation. The most effective compounds were batyl alcohol (1 and fuscoside E peracetate (6, acting against four strains without affecting their microbial growth. Compound 1 showed biofilm inhibition greater than 30% against A. macleodii, and up to 60% against O. pseudogringnonense,V. harveyi and S. aureus. Compound 6 inhibited O. pseudogringnonense and V. harveyi between 25 and 50%, and P. aeruginosa or S. aureus up to 60% at 0.5 mg/ml. The results suggest that these compounds exhibit specific biofilm inhibition with lower antimicrobial effect against the bacterial species assayed.

  7. The Extra-Cytoplasmic Function Sigma Factor SigX Modulates Biofilm and Virulence-Related Properties in Pseudomonas aeruginosa

    Science.gov (United States)

    Gicquel, Gwendoline; Bouffartigues, Emeline; Bains, Manjeet; Oxaran, Virginie; Rosay, Thibaut; Lesouhaitier, Olivier; Connil, Nathalie; Bazire, Alexis; Maillot, Olivier; Bénard, Magalie; Cornelis, Pierre; Hancock, Robert E. W.; Dufour, Alain; Feuilloley, Marc G. J.; Orange, Nicole; Déziel, Eric; Chevalier, Sylvie

    2013-01-01

    SigX, one of the 19 extra-cytoplasmic function sigma factors of P. aeruginosa, was only known to be involved in transcription of the gene encoding the major outer membrane protein OprF. We conducted a comparative transcriptomic study between the wildtype H103 strain and its sigX mutant PAOSX, which revealed a total of 307 differentially expressed genes that differed by more than 2 fold. Most dysregulated genes belonged to six functional classes, including the “chaperones and heat shock proteins”, “antibiotic resistance and susceptibility”, “energy metabolism”, “protein secretion/export apparatus”, and “secreted factors”, and “motility and attachment” classes. In this latter class, the large majority of the affected genes were down-regulated in the sigX mutant. In agreement with the array data, the sigX mutant was shown to demonstrate substantially reduced motility, attachment to biotic and abiotic surfaces, and biofilm formation. In addition, virulence towards the nematode Caenorhabditis elegans was reduced in the sigX mutant, suggesting that SigX is involved in virulence-related phenotypes. PMID:24260387

  8. Proteomic Response of Pseudomonas aeruginosa PAO1 Adhering to Solid Surfaces

    Directory of Open Access Journals (Sweden)

    Morgan Guilbaud

    2017-08-01

    Full Text Available Pseudomonas aeruginosa is a pathogenic micro-organism responsible for many hospital-acquired infections. It is able to adhere to solid surfaces and develop an immobilized community or so-called biofilm. Many studies have been focusing on the use of specific materials to prevent the formation of these biofilms, but the reactivity of the bacteria in contact to surfaces remains unknown. The aim of this study was to evaluate the impact of the abiotic surface on the physiology of adherent bacteria. Three different materials, stainless steel (SS, glass (G, and polystyrene (PS that were relevant to industrial or medical environments were characterized at the physicochemical level in terms of their hydrophobicity and roughness. We showed that SS was moderately hydrophilic and rough, potentially containing crevices, G was hydrophilic and smooth while PS was hydrophobic and smooth. We further showed that P. aeruginosa cells were more likely able to adhere to SS and G rather than PS surfaces under our experimental conditions. The physiological response of P. aeruginosa when adhering to each of these materials was then evaluated by global proteomic analysis. The abundance of 70 proteins was shown to differ between the materials suggesting that their abundance was modified as a function of the material to which bacteria adhered. Our data lead to enabling the identification of abundance patterns that appeared to be specific to a given surface. Taken together, our data showed that P. aeruginosa is capable of sensing and responding to a surface probably via specific programmes to adapt its physiological response accordingly.

  9. Effects of substrates on biofilm formation observed by atomic force microscopy

    International Nuclear Information System (INIS)

    Oh, Y.J.; Lee, N.R.; Jo, W.; Jung, W.K.; Lim, J.S.

    2009-01-01

    Formation of biofilm is known to be strongly dependent on substrates including topography, materials, and chemical treatment. In this study, a variety of substrates are tested for understanding biofilm formation. Sheets of aluminum, steel, rubber, and polypropylene have been used to examine their effects on formation of Pseudomonas aeruginosa biofilm. In particular, the morphological variation, transition, and adhesiveness of biofilm were investigated through local measurement by atomic force microscopy (AFM). Mechanism of removing biofilm from adhering to substrate is also analyzed, thus the understanding of the mechanism can be potentially useful to prevent the biofilm formation. The results reveal that formation of biofilm can remain on rough surface regardless of substrates in hot water, which may easily induce extra-polymeric substances detachment from bacterial surface. By probing using AFM, local force-distance characterization of extra-cellular materials extracted from the bacteria can exhibit the progress of the biofilm formation and functional complexities.

  10. Antivirulence activity of azithromycin in Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Francesco eImperi

    2014-04-01

    Full Text Available Antibiotics represent our bulwark to combat bacterial infections, but the spread of antibiotic resistance compromises their clinical efficacy. Alternatives to conventional antibiotics are urgently needed in order to complement the existing antibacterial arsenal. The macrolide antibiotic azithromycin (AZM provides a paradigmatic example of an unconventional antibacterial drug. Besides its growth-inhibiting activity, AZM displays potent anti-inflammatory properties, as well as antivirulence activity on some intrinsically resistant bacteria, such as Pseudomonas aeruginosa. In this bacterium, the antivirulence activity of AZM mainly relies on its ability to interact with the ribosome, resulting in direct and/or indirect repression of specific subsets of genes involved in virulence, quorum sensing, biofilm formation and intrinsic antibiotic resistance. Both clinical experience and clinical trials have shown the efficacy of AZM in the treatment of chronic pulmonary infections caused by P. aeruginosa. The aim of this review is to combine results from laboratory studies with evidence from clinical trials in order to unify the information on the in vivo mode of action of AZM in P. aeruginosa infection.

  11. Propolis-Sahara honeys preparation exhibits antibacterial and anti-biofilm activity against bacterial biofims formed on urinary catheters

    Directory of Open Access Journals (Sweden)

    Saad Aissat

    2016-11-01

    Full Text Available Objective: To evaluate the antibacterial effect of Sahara honeys (SHs against bacterial biofilms formed on urinary catheters in combination with propolis-Sahara honeys (P-SHs. Methods: Three clinical isolates were subjected to biofilm detection methods. The antibacterial and anti-biofilm activity for SHs and P-SHs were determined using agar well diffusion and the percentage of biofilm inhibition (PBI methods. Results: The PBI for Gram-positive bacteria [Staphylococcus aureus (S. aureus] was in the range of 0%–20%, while PBI for Gram-negative bacteria [Pseudomonas aeruginosa and Escherichia coli (E. coli] were in range of 17%–57% and 16%–65%, respectively. The highest PBI (65% was produced by SH2 only on E. coli. In agar well diffusion assay, zones of inhibition ranged from 11–20 mm (S. aureus, 9–19 mm (Pseudomonas aeruginosa and 11–19 mm (E. coli. The highest inhibition (20 mm was produced by SH1 only on S. aureus. In addition, the treatment of SHs and P-SHs catheters with a polymicrobial biofilms reduced biofilm formation after 48 h exposure period. Conclussions: SHs and P-SHs applied as a natural agent can be used as a prophylactic agent to prevent the formation of in vitro biofilm.

  12. Phosphate limitation induces the intergeneric inhibition of Pseudomonas aeruginosa by Serratia marcescens isolated from paper machines

    Science.gov (United States)

    Kuo, Pei-An; Kuo, Chih-Horng; Lai, Yiu-Kay; Graumann, Peter L; Tu, Jenn

    2013-01-01

    Phosphate is an essential nutrient for heterotrophic bacteria, affecting bacterioplankton in aquatic ecosystems and bacteria in biofilms. However, the influence of phosphate limitation on bacterial competition and biofilm development in multispecies populations has received limited attention in existing studies. To address this issue, we isolated 13 adhesive bacteria from paper machine aggregates. Intergeneric inhibition of Pseudomonas aeruginosa WW5 by Serratia marcescens WW4 was identified under phosphate-limited conditions, but not in Luria–Bertani medium or M9 minimal medium. The viable numbers of the pure S. marcescens WW4 culture decreased over 3 days in the phosphate-limited medium; however, the mortality of S. marcescens WW4 was significantly reduced when it was co-cultured with P. aeruginosa WW5, which appeared to sustain the S. marcescens WW4 biofilm. In contrast, viable P. aeruginosa WW5 cells immediately declined in the phosphate-limited co-culture. To identify the genetic/inhibitory element(s) involved in this process, we inserted a mini-Tn5 mutant of S. marcescens WW4 that lacked inhibitory effect. The results showed that an endonuclease bacteriocin was involved in this intergeneric inhibition by S. marcescens WW4 under phosphate limitation. In conclusion, this study highlights the importance of nutrient limitation in bacterial interactions and provides a strong candidate gene for future functional characterisation. PMID:23398522

  13. Investigation of Pseudomonas aeruginosa quorum-sensing signaling system for identifying multiple inhibitors using molecular docking and structural analysis methodology.

    Science.gov (United States)

    Soheili, Vahid; Bazzaz, Bibi Sedigheh Fazly; Abdollahpour, Nooshin; Hadizadeh, Farzin

    2015-12-01

    Pseudomonas aeruginosa is an opportunistic human pathogen and a common Gram-negative bacterium in hospital-acquired infections. It causes death in many burn victims, cystic-fibrosis and neutropenic-cancer patients. It is known that P. aeruginosa biofilm maturation and production of cell-associated and extracellular virulence factors such as pyocyanin, elastase and rhamnolipids are under the control of a quorum-sensing (QS) system. Among several proteins involved in the Pseudomonas QS mechanism, LasR and PqsE play an important role in its cascade signaling system. They can cause increases in QS factors, biofilm maturation, and the production of virulence factors. Therefore, inhibition of these proteins can reduce the pathogenicity of P. aeruginosa. According to the structure of corresponding auto-inducers bound to these proteins, in silico calculations were performed with some non-steroidal anti-inflammatory drugs (NSAIDs) to estimate possible interactions and find the co-inhibitors of LasR and PqsE. The results showed that oxicams (Piroxicam and Meloxicam) can interact well with active sites of both proteins with the Ki of 119.43 nM and 4.0 μM for Meloxicam and 201.39 nM and 4.88 μM against LasR and PqsE, respectively. These findings suggested that Piroxicam and Meloxicam can be used as potential inhibitors for control of the P. aeruginosa QS signaling system and biofilm formation, and may be used in the design of multiple inhibitors. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. Evaluation of Enoyl-Acyl Carrier Protein Reductase Inhibitors as Pseudomonas aeruginosa Quorum-Quenching Reagents

    DEFF Research Database (Denmark)

    Yang, Liang; Liu, Yang; Sternberg, Claus

    2010-01-01

    Pseudomonas aeruginosa is an opportunistic pathogen which is responsible for a wide range of infections. Production of virulence factors and biofilm formation by P. aeruginosa are partly regulated by cell-to-cell communication quorum-sensing systems. Identification of quorum-quenching reagents...... which block the quorum-sensing process can facilitate development of novel treatment strategies for P. aeruginosa infections. We have used molecular dynamics simulation and experimental studies to elucidate the efficiencies of two potential quorum-quenching reagents, triclosan and green tea...... epigallocatechin gallate (EGCG), which both function as inhibitors of the enoyl-acyl carrier protein (ACP) reductase (ENR) from the bacterial type II fatty acid synthesis pathway. Our studies suggest that EGCG has a higher binding affinity towards ENR of P. aeruginosa and is an efficient quorum-quenching reagent...

  15. Multiple sensors control reciprocal expression of Pseudomonas aeruginosa regulatory RNA and virulence genes

    DEFF Research Database (Denmark)

    Ventre, I.; Goodman, A.L.; Vallet-Gely, I.

    2006-01-01

    The opportunistic pathogen Pseudomonas aeruginosa is responsible for a wide range of acute and chronic infections. The transition to chronic infections is accompanied by physiological changes in the bacteria favoring formation of biofilm communities. Here we report the identification of LadS, a h...

  16. Nitrous oxide production in sputum from cystic fibrosis patients with chronic Pseudomonas aeruginosa lung infection.

    Directory of Open Access Journals (Sweden)

    Mette Kolpen

    Full Text Available Chronic lung infection by Pseudomonas aeruginosa is the major severe complication in cystic fibrosis (CF patients, where P. aeruginosa persists and grows in biofilms in the endobronchial mucus under hypoxic conditions. Numerous polymorphonuclear leukocytes (PMNs surround the biofilms and create local anoxia by consuming the majority of O2 for production of reactive oxygen species (ROS. We hypothesized that P. aeruginosa acquires energy for growth in anaerobic endobronchial mucus by denitrification, which can be demonstrated by production of nitrous oxide (N2O, an intermediate in the denitrification pathway. We measured N2O and O2 with electrochemical microsensors in 8 freshly expectorated sputum samples from 7 CF patients with chronic P. aeruginosa infection. The concentrations of NO3(- and NO2(- in sputum were estimated by the Griess reagent. We found a maximum median concentration of 41.8 µM N2O (range 1.4-157.9 µM N2O. The concentration of N2O in the sputum was higher below the oxygenated layers. In 4 samples the N2O concentration increased during the initial 6 h of measurements before decreasing for approximately 6 h. Concomitantly, the concentration of NO3(- decreased in sputum during 24 hours of incubation. We demonstrate for the first time production of N2O in clinical material from infected human airways indicating pathogenic metabolism based on denitrification. Therefore, P. aeruginosa may acquire energy for growth by denitrification in anoxic endobronchial mucus in CF patients. Such ability for anaerobic growth may be a hitherto ignored key aspect of chronic P. aeruginosa infections that can inform new strategies for treatment and prevention.

  17. Nitrous oxide production in sputum from cystic fibrosis patients with chronic Pseudomonas aeruginosa lung infection.

    Science.gov (United States)

    Kolpen, Mette; Kühl, Michael; Bjarnsholt, Thomas; Moser, Claus; Hansen, Christine Rønne; Liengaard, Lars; Kharazmi, Arsalan; Pressler, Tanja; Høiby, Niels; Jensen, Peter Østrup

    2014-01-01

    Chronic lung infection by Pseudomonas aeruginosa is the major severe complication in cystic fibrosis (CF) patients, where P. aeruginosa persists and grows in biofilms in the endobronchial mucus under hypoxic conditions. Numerous polymorphonuclear leukocytes (PMNs) surround the biofilms and create local anoxia by consuming the majority of O2 for production of reactive oxygen species (ROS). We hypothesized that P. aeruginosa acquires energy for growth in anaerobic endobronchial mucus by denitrification, which can be demonstrated by production of nitrous oxide (N2O), an intermediate in the denitrification pathway. We measured N2O and O2 with electrochemical microsensors in 8 freshly expectorated sputum samples from 7 CF patients with chronic P. aeruginosa infection. The concentrations of NO3(-) and NO2(-) in sputum were estimated by the Griess reagent. We found a maximum median concentration of 41.8 µM N2O (range 1.4-157.9 µM N2O). The concentration of N2O in the sputum was higher below the oxygenated layers. In 4 samples the N2O concentration increased during the initial 6 h of measurements before decreasing for approximately 6 h. Concomitantly, the concentration of NO3(-) decreased in sputum during 24 hours of incubation. We demonstrate for the first time production of N2O in clinical material from infected human airways indicating pathogenic metabolism based on denitrification. Therefore, P. aeruginosa may acquire energy for growth by denitrification in anoxic endobronchial mucus in CF patients. Such ability for anaerobic growth may be a hitherto ignored key aspect of chronic P. aeruginosa infections that can inform new strategies for treatment and prevention.

  18. Lipopeptide biosurfactants from Paenibacillus polymyxa inhibit single and mixed species biofilms.

    Science.gov (United States)

    Quinn, Gerry A; Maloy, Aaron P; McClean, Stephen; Carney, Brian; Slater, John W

    2012-01-01

    Although biofilms are recognised as important in microbial colonisation, solutions to their inhibition are predominantly based on planktonic assays. These solutions have limited efficacy against biofilms. Here, a series of biofilm-orientated tests were used to identify anti-biofilm compounds from marine micro-flora. This led to the isolation of a complex of anti-biofilm compounds from an extract of Paenibacillus polymyxa (PPE). A combination of rpHPLC and mass spectrometry identified the principle components of PPE as fusaricidin B (LI-FO4b) and polymyxin D1, with minor contributions from surfactins. This complex (PPE) reduced the biofilm biomass of Bacillus subtilis, Micrococcus luteus, Pseudomonas aeruginosa, Staphylococcus aureus and Streptococcus bovis. In contrast, ampicillin was only effective against S. aureus. PPE also inhibited a self-assembling marine biofilm (SAMB) in co-incubation assays by 99.3% ± 1.9 and disrupted established SAMB by 72.4% ± 4.4, while ampicillin showed no significant reduction. The effectiveness of this complex of lipopeptides against single and multispecies biofilms suggests a future role in biofilm prevention strategies.

  19. Azithromycin blocks quorum sensing and alginate polymer formation and increases the sensitivity to serum and stationary growth phase killing of P. aeruginosa and attenuates chronic P. aeruginosa lung infection in Cftr -/--mice

    DEFF Research Database (Denmark)

    Hoffmann, N.; Lee, Bao le ri; Hentzer, Morten

    2007-01-01

    The consequences of O-acetylated alginate-producing Pseudomonas aeruginosa biofilms in the lungs of chronically infected cystic fibrosis (CF) patients are tolerance to both antibiotic treatments and effects on the innate and the adaptive defense mechanisms. In clinical trials, azithromycin (AZM...... and the complement system. Moreover, we show that AZM may affect the polymerization of P. aeruginosa alginate by the incomplete precipitation of polymerized alginate and high levels of readily dialyzable uronic acids. In addition, we find that mucoid bacteria in the stationary growth phase became sensitive to AZM......, whereas cells in the exponential phase did not. Interestingly, AZM-treated P. aeruginosa lasI mutants appeared to be particularly resistant to serum, whereas bacteria with a functional QS system did not. We show in a CF mouse model of chronic P. aeruginosa lung infection that AZM treatment results...

  20. Gauging and visualizing c-di-GMP levels in pseudomonas aeruginosa using fluorescence-based biosensors

    DEFF Research Database (Denmark)

    Rybtke, Morten; Chua, Song Lin; Yam, Joey Kuok Hoong

    2017-01-01

    Recent research has shown that the molecule c-di-GMP is an important second messenger regulating various functions in bacteria. In particular, the implication of c-di-GMP as a positive regulator of adhesion and biofilm formation has gained momentum as a highly relevant research topic, as detailed...... knowledge about the underlying regulatory mechanisms may enable the development of measures to control biofilms in both industrial and medical settings. Accordingly, it is in many cases of interest to measure the c-di-GMP level in bacteria under specific conditions or in specific mutant strains. We have...... developed a collection of fluorescence-based c-di-GMP biosensors capable of gauging the c-di-GMP level in Pseudomonas aeruginosa and closely related bacteria. Here, we describe protocols for the use of these biosensors in gauging and visualizing cellular c-di-GMP levels of P. aeruginosa both in in vitro...

  1. Biofilm mediated decontamination of pollutants from the environment

    Directory of Open Access Journals (Sweden)

    Arindam Mitra

    2016-01-01

    Full Text Available In this review, we highlight beneficial use of microbial biofilms in remediation of environmental pollutants by bioremediation. Bioremediation is an environment friendly, cost effective, sustainable technology that utilizes microbes to decontaminate and degrade a wide variety of pollutants into less harmful products. Relative to free-floating planktonic cells, microbes existing in biofilm mode are advantageous for bioremediation because of greater tolerance to pollutants, environmental stress and ability to degrade varied harsh pollutants via diverse catabolic pathways. In biofilm mode, microbes are immobilized in a self-synthesized matrix which offers protection from stress, contaminants and predatory protozoa. Contaminants ranging from heavy metals, petroleum, explosives, pesticides have been remediated using microbial consortia of biofilms. In the industry, biofilm based bioremediation is used to decontaminate polluted soil and groundwater. Here we discuss conventional and newer strategies utilizing biofilms in environmental remediation.

  2. Comparative genome and transcriptome analysis reveals distinctive surface characteristics and unique physiological potentials of Pseudomonas aeruginosa ATCC 27853

    KAUST Repository

    Cao, Huiluo

    2017-06-12

    Pseudomonas aeruginosa ATCC 27853 was isolated from a hospital blood specimen in 1971 and has been widely used as a model strain to survey antibiotics susceptibilities, biofilm development, and metabolic activities of Pseudomonas spp.. Although four draft genomes of P. aeruginosa ATCC 27853 have been sequenced, the complete genome of this strain is still lacking, hindering a comprehensive understanding of its physiology and functional genome.Here we sequenced and assembled the complete genome of P. aeruginosa ATCC 27853 using the Pacific Biosciences SMRT (PacBio) technology and Illumina sequencing platform. We found that accessory genes of ATCC 27853 including prophages and genomic islands (GIs) mainly contribute to the difference between P. aeruginosa ATCC 27853 and other P. aeruginosa strains. Seven prophages were identified within the genome of P. aeruginosa ATCC 27853. Of the predicted 25 GIs, three contain genes that encode monoxoygenases, dioxygenases and hydrolases that could be involved in the metabolism of aromatic compounds. Surveying virulence-related genes revealed that a series of genes that encode the B-band O-antigen of LPS are lacking in ATCC 27853. Distinctive SNPs in genes of cellular adhesion proteins such as type IV pili and flagella biosynthesis were also observed in this strain. Colony morphology analysis confirmed an enhanced biofilm formation capability of ATCC 27853 on solid agar surface compared to Pseudomonas aeruginosa PAO1. We then performed transcriptome analysis of ATCC 27853 and PAO1 using RNA-seq and compared the expression of orthologous genes to understand the functional genome and the genomic details underlying the distinctive colony morphogenesis. These analyses revealed an increased expression of genes involved in cellular adhesion and biofilm maturation such as type IV pili, exopolysaccharide and electron transport chain components in ATCC 27853 compared with PAO1. In addition, distinctive expression profiles of the

  3. Antibacterial and anti-biofilm activity of ginger (Zingiber officinale (Roscoe ethanolic extract

    Directory of Open Access Journals (Sweden)

    Nikolić Miloš

    2014-01-01

    Full Text Available The antibacterial and anti-biofilm activity of ethanolic extract from the rhizome of Zingiber officinale were evaluated. In vitro antibacterial activity was investigated by microdilution method. Minimum inhibitory concentration (MIC and minimum bactericidal concentration (MBC have been determined. The values were in the range from 0.0024 to > 20 mg/ml. The most sensitive bacteria were Gram-positive bacteria: Staphylococcus aureus and Staphylococcus aureus ATCC 25923. Anti-biofilm activity was tested by crystal violet assay. Pseudomonas aeruginosa ATCC 27853, Proteus mirabilis and Escherichia coli ATCC 25922 were used as the test organisms. Ethanolic extract showed the best result on Proteus mirabilis biofilm where biofilm inhibitory concentration (BIC50 was 19 mg/ml.

  4. Phosphorylcholine impairs susceptibility to biofilm formation of hydrogel contact lenses.

    Science.gov (United States)

    Selan, Laura; Palma, Stefano; Scoarughi, Gian Luca; Papa, Rosanna; Veeh, Richard; Di Clemente, Daniele; Artini, Marco

    2009-01-01

    To compare silicone-hydrogel, poly(2-hydroxyethyl methacrylate) (pHEMA), and phosphorylcholine-coated (PC-C) contact lenses in terms of their susceptibility to biofilm formation by Staphylococcus epidermidis and Pseudomonas aeruginosa. Laboratory investigation. Biofilm formation on colonized test lenses was evaluated with confocal microscopy and in vitro antibiotic susceptibility assays. The results of the latter assays were compared with those performed on planktonic cultures of the same organism. For both microorganisms, sessile colonies on silicone-hydrogel and pHEMA lenses displayed lower antibiotic susceptibility than their planktonic counterparts. In contrast, the susceptibility of cultures growing on PC-C lenses was comparable with that for planktonic cultures. In particular, minimum inhibitory concentration for Tazocin (piperacillin plus tazobactam; Wyeth Pharmaceuticals, Aprilia, Italy; S. epidermidis) and gentamicin (P. aeruginosa) was identical, either in the presence of PC-C support or in planktonic cultures (Tazocin, aeruginosa) was two-fold higher for PC-C lenses (0.4 mug/ml) with respect to planktonic cultures (0.2 mug/ml). Confocal microscopy of lenses colonized for 24 hours with P. aeruginosa green fluorescent protein-expressing cells revealed a sessile colonization on silicone-hydrogel lens and a few isolated bacterial cells scattered widely over the surface of the PC-C lens. An increase in antibiotic susceptibility of bacterial cultures was associated with diminished bacterial adhesion. Our results indicate that PC-C lenses seem to be more resistant than silicone-hydrogel and pHEMA lenses to bacterial adhesion and colonization. This feature may facilitate their disinfection.

  5. Physics of biofilms: the initial stages of biofilm formation and dynamics

    International Nuclear Information System (INIS)

    Lambert, Guillaume; Bergman, Andrew; Zhang, Qiucen; Bortz, David; Austin, Robert

    2014-01-01

    One of the physiological responses of bacteria to external stress is to assemble into a biofilm. The formation of a biofilm greatly increases a bacterial population's resistance to a hostile environment by shielding cells, for example, from antibiotics. In this paper, we describe the conditions necessary for the emergence of biofilms in natural environments and relate them to the emergence of biofilm formation inside microfluidic devices. We show that competing species of Escherichia coli bacteria form biofilms to spatially segregate themselves in response to starvation stress, and use in situ methods to characterize the physical properties of the biofilms. Finally, we develop a microfluidic platform to study the inter-species interactions and show how biofilm-mediated genetic interactions can improve a species’ resistance to external stress. (paper)

  6. Staphylococcus aureus biofilms: recent developments in biofilm dispersal.

    Science.gov (United States)

    Lister, Jessica L; Horswill, Alexander R

    2014-01-01

    Staphylococcus aureus is a major cause of nosocomial and community-acquired infections and represents a significant burden on the healthcare system. S. aureus attachment to medical implants and host tissue, and the establishment of a mature biofilm, play an important role in the persistence of chronic infections. The formation of a biofilm, and encasement of cells in a polymer-based matrix, decreases the susceptibility to antimicrobials and immune defenses, making these infections difficult to eradicate. During infection, dispersal of cells from the biofilm can result in spread to secondary sites and worsening of the infection. In this review, we discuss the current understanding of the pathways behind biofilm dispersal in S. aureus, with a focus on enzymatic and newly described broad-spectrum dispersal mechanisms. Additionally, we explore potential applications of dispersal in the treatment of biofilm-mediated infections.

  7. Effectiveness of current disinfection procedures against biofilm on contaminated GI endoscopes.

    Science.gov (United States)

    Neves, Marcelo S; da Silva, Marlei Gomes; Ventura, Grasiella M; Côrtes, Patrícia Barbur; Duarte, Rafael Silva; de Souza, Heitor S

    2016-05-01

    Attention to patient safety has increased recently due to outbreaks of nosocomial infections associated with GI endoscopy. The aim of this study was to evaluate current cleaning and disinfection procedures of endoscope channels with high bioburden and biofilm analysis, including the use of resistant mycobacteria associated with postsurgical infections in Brazil. Twenty-seven original endoscope channels were contaminated with organic soil containing 10(8) colony-forming units/mL of Pseudomonas aeruginosa, Staphylococcus aureus, or Mycobacterium abscessus subsp bolletii. Biofilms with the same microorganisms were developed on the inner surface of channels with the initial inoculum of 10(5) colony-forming units/mL. Channels were reprocessed following current protocol, and samples from cleaning and disinfection steps were analyzed by bioluminescence for adenosine triphosphate, cultures for viable microorganisms, and confocal microscopy. After contamination, adenosine triphosphate levels increased dramatically, and high bacterial growth was observed in all cultures. After cleaning, adenosine triphosphate levels decreased to values comparable to precontamination levels, and bacterial growth was demonstrated in 5 of 27 catheters, 2 with P aeruginosa and 3 with M abscessus. With regard to induced biofilm, a remarkable reduction occurred after cleaning, but significant microbial growth inhibition occurred only after disinfection. Nevertheless, viable microorganisms within the biofilm were still detected by confocal microscopy, more so with glutaraldehyde than with peracetic acid or O-phataladehyde. After the complete disinfection procedure, viable microorganisms could still be detected within the biofilm on endoscope channels. Prevention of biofilm development within endoscope channels should be a priority in disinfection procedures, particularly for ERCP and EUS. Copyright © 2016 American Society for Gastrointestinal Endoscopy. Published by Elsevier Inc. All rights

  8. PAMDB: a comprehensive Pseudomonas aeruginosa metabolome database.

    Science.gov (United States)

    Huang, Weiliang; Brewer, Luke K; Jones, Jace W; Nguyen, Angela T; Marcu, Ana; Wishart, David S; Oglesby-Sherrouse, Amanda G; Kane, Maureen A; Wilks, Angela

    2018-01-04

    The Pseudomonas aeruginosaMetabolome Database (PAMDB, http://pseudomonas.umaryland.edu) is a searchable, richly annotated metabolite database specific to P. aeruginosa. P. aeruginosa is a soil organism and significant opportunistic pathogen that adapts to its environment through a versatile energy metabolism network. Furthermore, P. aeruginosa is a model organism for the study of biofilm formation, quorum sensing, and bioremediation processes, each of which are dependent on unique pathways and metabolites. The PAMDB is modelled on the Escherichia coli (ECMDB), yeast (YMDB) and human (HMDB) metabolome databases and contains >4370 metabolites and 938 pathways with links to over 1260 genes and proteins. The database information was compiled from electronic databases, journal articles and mass spectrometry (MS) metabolomic data obtained in our laboratories. For each metabolite entered, we provide detailed compound descriptions, names and synonyms, structural and physiochemical information, nuclear magnetic resonance (NMR) and MS spectra, enzymes and pathway information, as well as gene and protein sequences. The database allows extensive searching via chemical names, structure and molecular weight, together with gene, protein and pathway relationships. The PAMBD and its future iterations will provide a valuable resource to biologists, natural product chemists and clinicians in identifying active compounds, potential biomarkers and clinical diagnostics. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  9. Copper-Containing Anti-Biofilm Nanofiber Scaffolds as a Wound Dressing Material.

    Directory of Open Access Journals (Sweden)

    Jayesh J Ahire

    Full Text Available Copper particles were incorporated into nanofibers during the electrospinning of poly-D,L-lactide (PDLLA and poly(ethylene oxide (PEO. The ability of the nanofibers to prevent Pseudomonas aeruginosa PA01 and Staphylococcus aureus (strain Xen 30 to form biofilms was tested. Nanofibers containing copper particles (Cu-F were thinner (326 ± 149 nm in diameter, compared to nanofibers without copper (CF; 445 ± 93 nm in diameter. The crystalline structure of the copper particles in Cu-F was confirmed by X-ray diffraction (XRD. Copper crystals were encapsulated, but also attached to the surface of Cu-F, as shown scanning transmission electron microscopy (STEM and transmission electron microscopy (TEM, respectively. The copper particles had no effect on the thermal degradation and thermal behaviour of Cu-F, as shown by thermogravimetric analysis (TGA and differential scanning calorimeter (DSC. After 48 h in the presence of Cu-F, biofilm formation by P. aeruginosa PA01 and S. aureus Xen 30 was reduced by 41% and 50%, respectively. Reduction in biofilm formation was ascribed to copper released from the nanofibers. Copper-containing nanofibers may be incorporated into wound dressings.

  10. Evaluation of various metallic coatings on steel to mitigate biofilm formation.

    Science.gov (United States)

    Kanematsu, Hideyuki; Ikigai, Hajime; Yoshitake, Michiko

    2009-02-01

    In marine environments and water systems, it is easy for many structures to form biofilms on their surfaces and to be deteriorated due to the corrosion caused by biofilm formation by bacteria. The authors have investigated the antibacterial effects of metallic elements in practical steels so far to solve food-related problems, using Escherichia coli and Staphylococcus aureus. However, from the viewpoint of material deterioration caused by bacteria and their antifouling measures, we should consider the biofilm behavior as aggregate rather than individual bacterium. Therefore, we picked up Pseudomonas aeruginosa and Pseudoalteromonas carageenovara in this study, since they easily form biofilms in estuarine and marine environments. We investigated what kind of metallic elements could inhibit the biofilm formation at first and then discussed how the thin films of those inhibitory elements on steels could affect biofilm formation. The information would lead to the establishment of effective antifouling measures against corrosion in estuarine and marine environments.

  11. Evaluation of Various Metallic Coatings on Steel to Mitigate Biofilm Formation

    Directory of Open Access Journals (Sweden)

    Hajime Ikigai

    2009-02-01

    Full Text Available In marine environments and water systems, it is easy for many structures to form biofilms on their surfaces and to be deteriorated due to the corrosion caused by biofilm formation by bacteria. The authors have investigated the antibacterial effects of metallic elements in practical steels so far to solve food-related problems, using Escherichia coli and Staphylococcus aureus. However, from the viewpoint of material deterioration caused by bacteria and their antifouling measures, we should consider the biofilm behavior as aggregate rather than individual bacterium. Therefore, we picked up Pseudomonas aeruginosa and Pseudoalteromonas carageenovara in this study, since they easily form biofilms in estuarine and marine environments. We investigated what kind of metallic elements could inhibit the biofilm formation at first and then discussed how the thin films of those inhibitory elements on steels could affect biofilm formation. The information would lead to the establishment of effective antifouling measures against corrosion in estuarine and marine environments.

  12. Management of dental unit waterline biofilms in the 21st century.

    LENUS (Irish Health Repository)

    O'Donnell, Mary J

    2011-10-01

    Dental chair units (DCUs) use water to cool and irrigate DCU-supplied instruments and tooth surfaces, and provide rinsewater during dental treatment. A complex network of interconnected plastic dental unit waterlines (DUWLs) supply water to these instruments. DUWLs are universally prone to microbial biofilm contamination seeded predominantly from microorganisms in supply water. Consequently, DUWL output water invariably becomes contaminated by high densities of microorganisms, principally Gram-negative environmental bacteria including Pseudomonas aeruginosa and Legionella species, but sometimes contain human-derived pathogens such as Staphylococcus aureus. Patients and staff are exposed to microorganisms from DUWL output water and to contaminated aerosols generated by DCU instruments. A wide variety of approaches, many unsuccessful, have been proposed to control DUWL biofilm. More recently, advances in biofilm science, chemical DUWL biofilm treatment agents, DCU design, supply water treatment and development of automated DUWL biofilm control systems have provided effective long-term solutions to DUWL biofilm control.

  13. Novel approaches to mitigating bacterial biofilm formation and intercellular communication

    Science.gov (United States)

    Kasper, Stephen H.

    RNA-sequencing analysis and various other genetic and biochemical assays were performed to uncover the molecular target of our lead inhibitor in P. aeruginosa. Our results support the idea that the lead compound, S-phenyl-L-cysteine sulfoxide, is a competitive inhibitor of the enzyme kynureninase (KynU). KynU is a critical source of anthranilate, which is a precursor to a QS signaling molecule known as PQS (Pseudomonas quinolone signal). Previous research showed that kynU-knockout mutants have attenuated virulence; however, this was the first report to demonstrate a chemical probe could achieve KynU-mediated virulence attenuation. Because biofilm/QS inhibitors target cell signaling, as opposed to cell viability, novel approaches must be developed to enhance bioavailability and maximize their efficacy in complex and dynamic environments, such as the oral cavity. Therefore, a nanocapsule-based drug delivery system prepared from a natural plant protein was designed, fabricated, and characterized. The loading properties, release profiles, and ability to adsorb to and form films on hydroxyapatite (i.e. material of tooth surface), and ability to deter Streptococcus mutans biofilm was studied. To enhance performance of this drug delivery system, a synthetic biology approach was used to genetically fuse an oligopeptide hydroxyapatite-affinity tag to the zein peptide sequence. As observed antibiotic resistance is occurring at a faster pace than the approval of new antibiotics, it has become apparent that antimicrobial treatment is not a sustainable method to fight pathogenic infection. If the treatment paradigm of infectious disease is to transform to a more sustainable approach by mitigating bacterial virulence, significant advances must be made in this field. My dissertation explores several avenues of infectious disease research: (1) identification of new compounds for disarming pathogens, (2) discovery of the mechanism of action of a lead inhibitor, and (3) the design of a

  14. Quorum-sensing and cheating in bacterial biofilms

    Science.gov (United States)

    Popat, Roman; Crusz, Shanika A.; Messina, Marco; Williams, Paul; West, Stuart A.; Diggle, Stephen P.

    2012-01-01

    The idea from human societies that self-interest can lead to a breakdown of cooperation at the group level is sometimes termed the public goods dilemma. We tested this idea in the opportunistic bacterial pathogen, Pseudomonas aeruginosa, by examining the influence of putative cheats that do not cooperate via cell-to-cell signalling (quorum-sensing, QS). We found that: (i) QS cheating occurs in biofilm populations owing to exploitation of QS-regulated public goods; (ii) the thickness and density of biofilms was reduced by the presence of non-cooperative cheats; (iii) population growth was reduced by the presence of cheats, and this reduction was greater in biofilms than in planktonic populations; (iv) the susceptibility of biofilms to antibiotics was increased by the presence of cheats; and (v) coercing cooperator cells to increase their level of cooperation decreases the extent to which the presence of cheats reduces population productivity. Our results provide clear support that conflict over public goods reduces population fitness in bacterial biofilms, and that this effect is greater than in planktonic populations. Finally, we discuss the clinical implications that arise from altering the susceptibility to antibiotics. PMID:23034707

  15. Adhesion of Pseudomonas aeruginosa and Staphylococcus epidermidis to silicone-hydrogel contact lenses.

    Science.gov (United States)

    Henriques, Mariana; Sousa, Cláudia; Lira, Madalena; Elisabete, M; Oliveira, Real; Oliveira, Rosário; Azeredo, Joana

    2005-06-01

    The purpose of this study is to compare the adhesion capabilities of the most important etiologic agents of microbial ocular infection to the recently available silicone-hydrogel lenses with those to a conventional hydrogel lens. In vitro static adhesion assays of Pseudomonas aeruginosa 10,145, Staphylococcus epidermidis 9142 (biofilm-positive), and 12,228 (biofilm-negative) to two extended-wear silicone-hydrogel lenses (balafilcon A and lotrafilcon A), a daily wear silicone-hydrogel lens (galyfilcon A) and a conventional hydrogel (etafilcon A) were performed. To interpret the adhesion results, lens surface relative hydrophobicity was assessed by water contact angle measurements. P. aeruginosa and S. epidermidis 9142 exhibited greater adhesion capabilities to the extended wear silicone-hydrogel lenses than to the daily wear silicone- and conventional hydrogel lenses (p adhesion extent of these strains to galyfilcon A and etafilcon A. The biofilm negative strain of S. epidermidis adhered in larger extents to the silicone-hydrogel lenses than to the conventional hydrogel (p contact angle measurements revealed that the extended wear silicone-hydrogel lenses are hydrophobic, whereas the daily wear silicone- and conventional hydrogel lenses are hydrophilic. As a result of their hydrophobicity, the extended wear silicone-hydrogel lenses (lotrafilcon A and balafilcon A) may carry higher risk of microbial contamination than both the hydrophilic daily wear silicone-hydrogel lens, galyfilcon A and the conventional hydrogel lens, etafilcon A.

  16. Bacteria of the Burkholderia cepacia complex are cyanogenic under biofilm and colonial growth conditions

    Directory of Open Access Journals (Sweden)

    Hoshino Saiko

    2008-06-01

    Full Text Available Abstract Background The Burkholderia cepacia complex (Bcc is a collection of nine genotypically distinct but phenotypically similar species. They show wide ecological diversity and include species that are used for promoting plant growth and bio-control as well species that are opportunistic pathogens of vulnerable patients. Over recent years the Bcc have emerged as problematic pathogens of the CF lung. Pseudomonas aeruginosa is another important CF pathogen. It is able to synthesise hydrogen cyanide (HCN, a potent inhibitor of cellular respiration. We have recently shown that HCN production by P. aeruginosa may have a role in CF pathogenesis. This paper describes an investigation of the ability of bacteria of the Bcc to make HCN. Results The genome of Burkholderia cenocepacia has 3 putative HCN synthase encoding (hcnABC gene clusters. B. cenocepacia and all 9 species of the Bcc complex tested were able to make cyanide at comparable levels to P. aeruginosa, but only when grown surface attached as colonies or during biofilm growth on glass beads. In contrast to P. aeruginosa and other cyanogenic bacteria, cyanide was not detected during planktonic growth of Bcc strains. Conclusion All species in the Bcc are cyanogenic when grown as surface attached colonies or as biofilms.

  17. AtlA Mediates Extracellular DNA Release, Which Contributes to Streptococcus mutans Biofilm Formation in an Experimental Rat Model of Infective Endocarditis.

    Science.gov (United States)

    Jung, Chiau-Jing; Hsu, Ron-Bin; Shun, Chia-Tung; Hsu, Chih-Chieh; Chia, Jean-San

    2017-09-01

    Host factors, such as platelets, have been shown to enhance biofilm formation by oral commensal streptococci, inducing infective endocarditis (IE), but how bacterial components contribute to biofilm formation in vivo is still not clear. We demonstrated previously that an isogenic mutant strain of Streptococcus mutans deficient in autolysin AtlA (Δ atlA ) showed a reduced ability to cause vegetation in a rat model of bacterial endocarditis. However, the role of AtlA in bacterial biofilm formation is unclear. In this study, confocal laser scanning microscopy analysis showed that extracellular DNA (eDNA) was embedded in S. mutans GS5 floes during biofilm formation on damaged heart valves, but an Δ atlA strain could not form bacterial aggregates. Semiquantification of eDNA by PCR with bacterial 16S rRNA primers demonstrated that the Δ atlA mutant strain produced dramatically less eDNA than the wild type. Similar results were observed with in vitro biofilm models. The addition of polyanethol sulfonate, a chemical lysis inhibitor, revealed that eDNA release mediated by bacterial cell lysis is required for biofilm initiation and maturation in the wild-type strain. Supplementation of cultures with calcium ions reduced wild-type growth but increased eDNA release and biofilm mass. The effect of calcium ions on biofilm formation was abolished in Δ atlA cultures and by the addition of polyanethol sulfonate. The VicK sensor, but not CiaH, was found to be required for the induction of eDNA release or the stimulation of biofilm formation by calcium ions. These data suggest that calcium ion-regulated AtlA maturation mediates the release of eDNA by S. mutans , which contributes to biofilm formation in infective endocarditis. Copyright © 2017 American Society for Microbiology.

  18. Combination of cupric ion with hydroxylamine and hydrogen peroxide for the control of bacterial biofilms on RO membranes.

    Science.gov (United States)

    Lee, Hye-Jin; Kim, Hyung-Eun; Lee, Changha

    2017-03-01

    Combinations of Cu(II) with hydroxylamine (HA) and hydrogen peroxide (H 2 O 2 ) (i.e., Cu(II)/HA, Cu(II)/H 2 O 2 , and Cu(II)/HA/H 2 O 2 systems) were investigated for the control of P. aeruginosa biofilms on reverse osmosis (RO) membranes. These Cu(II)-based disinfection systems effectively inactivated P. aeruginosa cells, exhibiting different behaviors depending on the state of bacterial cells (planktonic or biofilm) and the condition of biofilm growth and treatment (normal or pressurized condition). The Cu(II)/HA and Cu(II)/HA/H 2 O 2 systems were the most effective reagents for the inactivation of planktonic cells. However, these systems were not effective in inactivating cells in biofilms on the RO membranes possibly due to the interactions of Cu(I) with extracellular polymeric substances (EPS), where biofilms were grown and treated in center for disease control (CDC) reactors. Different from the results using CDC reactors, in a pressurized cross-flow RO filtration unit, the Cu(II)/HA/H 2 O 2 treatment significantly inactivated biofilm cells formed on the RO membranes, successfully recovering the permeate flux reduced by the biofouling. The pretreatment of feed solutions by Cu(II)/HA and Cu(II)/HA/H 2 O 2 systems (applied before the biofilm formation) effectively mitigated the permeate flux decline by preventing the biofilm growth on the RO membranes. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Evaluation of bactericidal and anti-biofilm properties of a novel surface-active organosilane biocide against healthcare associated pathogens and Pseudomonas aeruginosa biolfilm.

    Directory of Open Access Journals (Sweden)

    Jason Murray

    Full Text Available Healthcare acquired infections (HAI pose a great threat in hospital settings and environmental contamination can be attributed to the spread of these. De-contamination and, significantly, prevention of re-contamination of the environment could help in preventing/reducing this threat. Goldshield (GS5 is a novel organosilane biocide marketed as a single application product with residual biocidal activity. We tested the hypothesis that GS5 could provide longer-term residual antimicrobial activity than existing disinfectants once applied to surfaces. Thus, the residual bactericidal properties of GS5, Actichlor and Distel against repeated challenge with Staphylococcus aureus ATCC43300 were tested, and showed that GS5 alone exhibited longer-term bactericidal activity for up to 6 days on 316I stainless steel surfaces. Having established efficacy against S. aureus, we tested GS5 against common healthcare acquired pathogens, and demonstrated that, on average, a 1 log10 bactericidal effect was exhibited by GS5 treated surfaces, although biocidal activity varied depending upon the surface type and the species of bacteria. The ability of GS5 to prevent Pseudomonas aeruginosa biofilm formation was measured in standard microtitre plate assays, where it had no significant effect on either biofilm formation or development. Taken together the data suggests that GS5 treatment of surfaces may be a useful means to reducing bacterial contamination in the context of infection control practices.

  20. Contamination potential of drinking water distribution network biofilms.

    Science.gov (United States)

    Wingender, J; Flemming, H C

    2004-01-01

    Drinking water distribution system biofilms were investigated for the presence of hygienically relevant microorganisms. Early biofilm formation was evaluated in biofilm reactors on stainless steel, copper, polyvinyl chloride (PVC) and polyethylene coupons exposed to unchlorinated drinking water. After 12 to 18 months, a plateau phase of biofilm development was reached. Surface colonization on the materials ranged between 4 x 10(6) and 3 x 10(7) cells/cm2, with heterotrophic plate count (HPC) bacteria between 9 x 10(3) and 7 x 10(5) colony-forming units (cfu)/cm2. Established biofilms were investigated in 18 pipe sections (2 to 99 years old) cut out from distribution pipelines. Materials included cast iron, galvanized steel, cement and PVC. Colonization ranged from 4 x 10(5) to 2 x 10(8) cells/cm2, HPC levels varied between 1 and 2 x 10(5) cfu/cm2. No correlation was found between extent of colonization and age of the pipes. Using cultural detection methods, coliform bacteria were rarely found, while Escherichia coli, Pseudomonas aeruginosa and Legionella spp. were not detected in the biofilms. In regular operation, distribution system biofilms do not seem to be common habitats for pathogens. However, nutrient-leaching materials like rubber-coated valves were observed with massive biofilms which harboured coliform bacteria contaminating drinking water.

  1. From Nanowires to Biofilms: An Exploration of Novel Mechanisms of Uranium Transformation Mediated by Geobacter Bacteria

    Energy Technology Data Exchange (ETDEWEB)

    REGUERA, GEMMA [Michigan State University

    2014-01-16

    One promising strategy for the in situ bioremediation of radioactive groundwater contaminants that has been identified by the SBR Program is to stimulate the activity of dissimilatory metal-reducing microorganisms to reductively precipitate uranium and other soluble toxic metals. The reduction of U(VI) and other soluble contaminants by Geobacteraceae is directly dependent on the reduction of Fe(III) oxides, their natural electron acceptor, a process that requires the expression of Geobacter’s conductive pili (pilus nanowires). Expression of conductive pili by Geobacter cells leads to biofilm development on surfaces and to the formation of suspended biogranules, which may be physiological closer to biofilms than to planktonic cells. Biofilm development is often assumed in the subsurface, particularly at the matrix-well screen interface, but evidence of biofilms in the bulk aquifer matrix is scarce. Our preliminary results suggest, however, that biofilms develop in the subsurface and contribute to uranium transformations via sorption and reductive mechanisms. In this project we elucidated the mechanism(s) for uranium immobilization mediated by Geobacter biofilms and identified molecular markers to investigate if biofilm development is happening in the contaminated subsurface. The results provided novel insights needed in order to understand the metabolic potential and physiology of microorganisms with a known role in contaminant transformation in situ, thus having a significant positive impact in the SBR Program and providing novel concept to monitor, model, and predict biological behavior during in situ treatments.

  2. Pattern differentiation in co-culture biofilms formed by Staphylococcus aureus and Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Yang, Liang; Liu, Yang; Markussen, Trine

    2011-01-01

    Biofilm infections may not simply be the result of colonization by one bacterium, but rather the consequence of pathogenic contributions from several bacteria. Interspecies interactions of different organisms in mixed-species biofilms remain largely unexplained, but knowledge of these is very imp...

  3. The Effect of NIR Light and the Light-Activated Antimicrobial Agent on Wound Pathogenic Biofilms; Implication for Nonpharmacologic Chronic Wound Treatment

    DEFF Research Database (Denmark)

    Omar, Ghada Said Mohammed

    2015-01-01

    to the increase of lifestyle diseases, as diabetes, obesity, and cardiovascular diseases. The presence of bacterial biofilms is considered an important factor responsible for wounds chronicity. Therefore, this study investigates the efficacy of near-infrared (NIR) laser in vitro, in disrupting wound pathogenic...... biofilms. Metoder / Methods Biofilms of Staphylococcus aureus and Pseudomonas aeruginosa were grown in a 96-well microtiter plate for 18-22 h. The study included 4 arms: (a) control; (b) 200 µg/mL Indocyanin green (ICG) kept in the dark, (c) NIR laser alone; (d) NIR laser combined with 200 µg/mL. ICG....... The biofilms were exposed to different light doses from the 808 nm NIR laser at a fluence rate of 0.3 W/cm2. Crystal violate assay and viable count were used to detect the effect. (Foreløbige) resultater / (Preliminary) Results A light dose of 90 J/cm2 killed approximately 99.9% of P. aeruginosa bacterial...

  4. Comparative genome and transcriptome analysis reveals distinctive surface characteristics and unique physiological potentials of Pseudomonas aeruginosa ATCC 27853

    KAUST Repository

    Cao, Huiluo; Lai, Yong; Bougouffa, Salim; Xu, Zeling; Yan, Aixin

    2017-01-01

    Pseudomonas aeruginosa ATCC 27853 was isolated from a hospital blood specimen in 1971 and has been widely used as a model strain to survey antibiotics susceptibilities, biofilm development, and metabolic activities of Pseudomonas spp.. Although four

  5. A Pseudomonas aeruginosa toxin that hijacks the host ubiquitin proteolytic system.

    Directory of Open Access Journals (Sweden)

    Jennifer M Bomberger

    2011-03-01

    Full Text Available Pseudomonas aeruginosa (P. aeruginosa is an opportunistic pathogen chronically infecting the lungs of patients with chronic obstructive pulmonary disease (COPD, pneumonia, cystic fibrosis (CF, and bronchiectasis. Cif (PA2934, a bacterial toxin secreted in outer membrane vesicles (OMV by P. aeruginosa, reduces CFTR-mediated chloride secretion by human airway epithelial cells, a key driving force for mucociliary clearance. The aim of this study was to investigate the mechanism whereby Cif reduces CFTR-mediated chloride secretion. Cif redirected endocytosed CFTR from recycling endosomes to lysosomes by stabilizing an inhibitory effect of G3BP1 on the deubiquitinating enzyme (DUB, USP10, thereby reducing USP10-mediated deubiquitination of CFTR and increasing the degradation of CFTR in lysosomes. This is the first example of a bacterial toxin that regulates the activity of a host DUB. These data suggest that the ability of P. aeruginosa to chronically infect the lungs of patients with COPD, pneumonia, CF, and bronchiectasis is due in part to the secretion of OMV containing Cif, which inhibits CFTR-mediated chloride secretion and thereby reduces the mucociliary clearance of pathogens.

  6. Phenotypic shift in Pseudomonas aeruginosa populations from cystic fibrosis lungs after 2-week antipseudomonal treatment

    DEFF Research Database (Denmark)

    Fernandez-Barat, Laia; Ciofu, Oana; Kragh, Kasper N.

    2017-01-01

    ) colony-forming units (CFU/mL), b) frequency of mucoids and non-mucoids, c) resistance pattern to anti-pseudomonal drugs, d) hypermutability, e) transcriptomic profiles, and f) presence of biofilms. Results We collected 23 sputum samples (12 before antibiotic treatment and 11 after) and 77 P. aeruginosa...

  7. Investigating electrochemical removal of bacterial biofilms from stainless steel substrates.

    Science.gov (United States)

    Dargahi, Mahdi; Hosseinidoust, Zeinab; Tufenkji, Nathalie; Omanovic, Sasha

    2014-05-01

    Electrochemical removal of biofilms deserves attention because of its ease of use and environmentally friendly nature. We investigated the influence of electrode potential and treatment time on the removal of a 10-day old Pseudomonas aeruginosa biofilm formed on stainless steel 316 L substrates. At electrode potentials more positive than -1.5 V vs. Ag/AgCl, lower removal rates were observed and only partial removal of the biofilm was achieved during a 1-min time interval. Electrostatic repulsion between the film and electrode surface is believed to drive biofilm detachment under these conditions. However, when the biofilm-coated substrates were treated at potentials negative of -1.5 V vs. Ag/AgCl, complete removal of a biofilm was achieved within seconds. Under these conditions, vigorous evolution of hydrogen gas is believed to be responsible for the film removal, mechanically detaching the bacteria and extracellular polymeric matrix from the substrate. Stainless steel substrates were also subjected to repeated cycles of biofilm formation and electrochemical removal. High removal efficiencies were maintained throughout this process suggesting the potential of the proposed technology for application on conductive surfaces in various industrial settings. Copyright © 2014 Elsevier B.V. All rights reserved.

  8. Bacterial Biofilm Control by Perturbation of Bacterial Signaling Processes

    Directory of Open Access Journals (Sweden)

    Tim Holm Jakobsen

    2017-09-01

    Full Text Available The development of effective strategies to combat biofilm infections by means of either mechanical or chemical approaches could dramatically change today’s treatment procedures for the benefit of thousands of patients. Remarkably, considering the increased focus on biofilms in general, there has still not been invented and/or developed any simple, efficient and reliable methods with which to “chemically” eradicate biofilm infections. This underlines the resilience of infective agents present as biofilms and it further emphasizes the insufficiency of today’s approaches used to combat chronic infections. A potential method for biofilm dismantling is chemical interception of regulatory processes that are specifically involved in the biofilm mode of life. In particular, bacterial cell to cell signaling called “Quorum Sensing” together with intracellular signaling by bis-(3′-5′-cyclic-dimeric guanosine monophosphate (cyclic-di-GMP have gained a lot of attention over the last two decades. More recently, regulatory processes governed by two component regulatory systems and small non-coding RNAs have been increasingly investigated. Here, we review novel findings and potentials of using small molecules to target and modulate these regulatory processes in the bacterium Pseudomonas aeruginosa to decrease its pathogenic potential.

  9. Phenotypic and Genotypic Comparison of Epidemic and Non-Epidemic Strains of Pseudomonas aeruginosa from Individuals with Cystic Fibrosis.

    Directory of Open Access Journals (Sweden)

    Jessica Duong

    Full Text Available Epidemic strains of Pseudomonas aeruginosa have been found worldwide among the cystic fibrosis (CF patient population. Using pulse-field gel electrophoresis, the Prairie Epidemic Strain (PES has recently been found in one-third of patients attending the Calgary Adult CF Clinic in Canada. Using multi-locus sequence typing, PES isolates from unrelated patients were found to consistently have ST192. Though most patients acquired PES prior to enrolling in the clinic, some patients were observed to experience strain replacement upon transitioning to the clinic whereby local non-epidemic P. aeruginosa isolates were displaced by PES. Here we genotypically and phenotypically compared PES to other P. aeruginosa epidemic strains (OES found around the world as well as local non-epidemic CF P. aeruginosa isolates in order to characterize PES. Since some epidemic strains are associated with worse clinical outcomes, we assessed the pathogenic potential of PES to determine if these isolates are virulent, shared properties with OES, and if its phenotypic properties may offer a competitive advantage in displacing local non-epidemic isolates during strain replacement. As such, we conducted a comparative analysis using fourteen phenotypic traits, including virulence factor production, biofilm formation, planktonic growth, mucoidy, and antibiotic susceptibility to characterize PES, OES, and local non-epidemic isolates. We observed that PES and OES could be differentiated from local non-epidemic isolates based on biofilm growth with PES isolates being more mucoid. Pairwise comparisons indicated that PES produced significantly higher levels of proteases and formed better biofilms than OES but were more susceptible to antibiotic treatment. Amongst five patients experiencing strain replacement, we found that super-infecting PES produced lower levels of proteases and elastases but were more resistant to antibiotics compared to the displaced non-epidemic isolates. This

  10. The LasB Elastase of Pseudomonas aeruginosa Acts in Concert with Alkaline Protease AprA To Prevent Flagellin-Mediated Immune Recognition.

    Science.gov (United States)

    Casilag, Fiordiligie; Lorenz, Anne; Krueger, Jonas; Klawonn, Frank; Weiss, Siegfried; Häussler, Susanne

    2016-01-01

    The opportunistic pathogen Pseudomonas aeruginosa is capable of establishing severe and persistent infections in various eukaryotic hosts. It encodes a wide array of virulence factors and employs several strategies to evade immune detection. In the present study, we screened the Harvard Medical School transposon mutant library of P. aeruginosa PA14 for bacterial factors that modulate interleukin-8 responses in A549 human airway epithelial cells. We found that in addition to the previously identified alkaline protease AprA, the elastase LasB is capable of degrading exogenous flagellin under calcium-replete conditions and prevents flagellin-mediated immune recognition. Our results indicate that the production of two proteases with anti-flagellin activity provides a failsafe mechanism for P. aeruginosa to ensure the maintenance of protease-dependent immune-modulating functions. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  11. Elimination of the formation of biofilm in industrial pipes using enzyme cleaning technique

    Directory of Open Access Journals (Sweden)

    Xiaobo Liu

    2014-01-01

    Lastly, the terminal water was tested with SLYM-BART™ (HACH Corporation to find out whether there were biofilm-forming bacteria, such as Pseudomonas aeruginosa (Lakretz et al. (2011 [1], Pseudomonas fluorescens (O’Toole and Kolter (1998 [2], iron bacterium, etc.

  12. Komodo dragon-inspired synthetic peptide DRGN-1 promotes wound-healing of a mixed-biofilm infected wound.

    Science.gov (United States)

    M C Chung, Ezra; Dean, Scott N; Propst, Crystal N; Bishop, Barney M; van Hoek, Monique L

    2017-01-01

    Cationic antimicrobial peptides are multifunctional molecules that have a high potential as therapeutic agents. We have identified a histone H1-derived peptide from the Komodo dragon ( Varanus komodoensis) , called VK25. Using this peptide as inspiration, we designed a synthetic peptide called DRGN-1. We evaluated the antimicrobial and anti-biofilm activity of both peptides against Pseudomonas aeruginosa and Staphylococcus aureus . DRGN-1, more than VK25, exhibited potent antimicrobial and anti-biofilm activity, and permeabilized bacterial membranes. Wound healing was significantly enhanced by DRGN-1 in both uninfected and mixed biofilm ( Pseudomonas aeruginosa and Staphylococcus aureus )-infected murine wounds. In a scratch wound closure assay used to elucidate the wound healing mechanism, the peptide promoted the migration of HEKa keratinocyte cells, which was inhibited by mitomycin C (proliferation inhibitor) and AG1478 (epidermal growth factor receptor inhibitor). DRGN-1 also activated the EGFR-STAT1/3 pathway. Thus, DRGN-1 is a candidate for use as a topical wound treatment. Wound infections are a major concern; made increasingly complicated by the emerging, rapid spread of bacterial resistance. The novel synthetic peptide DRGN-1 (inspired by a peptide identified from Komodo dragon) exhibits pathogen-directed and host-directed activities in promoting the clearance and healing of polymicrobial ( Pseudomonas aeruginosa & Staphylococcus aureus ) biofilm infected wounds. The effectiveness of this peptide cannot be attributed solely to its ability to act upon the bacteria and disrupt the biofilm, but also reflects the peptide's ability to promsote keratinocyte migration. When applied in a murine model, infected wounds treated with DRGN-1 healed significantly faster than did untreated wounds, or wounds treated with other peptides. The host-directed mechanism of action was determined to be via the EGFR-STAT1/3 pathway. The pathogen-directed mechanism of action was

  13. Plant-expressed pyocins for control of Pseudomonas aeruginosa.

    Directory of Open Access Journals (Sweden)

    Šarūnas Paškevičius

    Full Text Available The emergence, persistence and spread of antibiotic-resistant human pathogenic bacteria heralds a growing global health crisis. Drug-resistant strains of gram-negative bacteria, such as Pseudomonas aeruginosa, are especially dangerous and the medical and economic burden they impose underscore the critical need for finding new antimicrobials. Recent studies have demonstrated that plant-expressed bacteriocins of the colicins family can be efficient antibacterials against all major enteropathogenic strains of E. coli. We extended our studies of colicin-like bacteriocins to pyocins, which are produced by strains of P. aeruginosa for ecological advantage against other strains of the same species. Using a plant-based transient expression system, we expressed six different pyocins, namely S5, PaeM, L1, L2, L3 and one new pyocin, PaeM4, and purified them to homogeneity. Among these pyocins, PaeM4 demonstrated the broadest spectrum of activity by controlling 53 of 100 tested clinical isolates of P. aeruginosa. The activity of plant-made pyocins was confirmed in the agar drop, liquid culture susceptibility and biofilm assays, and in the Galleria mellonella animal infection model.

  14. In-vivo expression profiling of Pseudomonas aeruginosa infections reveals niche-specific and strain-independent transcriptional programs.

    Directory of Open Access Journals (Sweden)

    Piotr Bielecki

    Full Text Available Pseudomonas aeruginosa is a threatening, opportunistic pathogen causing disease in immunocompromised individuals. The hallmark of P. aeruginosa virulence is its multi-factorial and combinatorial nature. It renders such bacteria infectious for many organisms and it is often resistant to antibiotics. To gain insights into the physiology of P. aeruginosa during infection, we assessed the transcriptional programs of three different P. aeruginosa strains directly after isolation from burn wounds of humans. We compared the programs to those of the same strains using two infection models: a plant model, which consisted of the infection of the midrib of lettuce leaves, and a murine tumor model, which was obtained by infection of mice with an induced tumor in the abdomen. All control conditions of P. aeruginosa cells growing in suspension and as a biofilm were added to the analysis. We found that these different P. aeruginosa strains express a pool of distinct genetic traits that are activated under particular infection conditions regardless of their genetic variability. The knowledge herein generated will advance our understanding of P. aeruginosa virulence and provide valuable cues for the definition of prospective targets to develop novel intervention strategies.

  15. Diffusion Retardation by Binding of Tobramycin in an Alginate Biofilm Model

    DEFF Research Database (Denmark)

    Cao, Bao; Christophersen, Lars; Kolpen, Mette

    2016-01-01

    Microbial cells embedded in a self-produced extracellular biofilm matrix cause chronic infections, e. g. by Pseudomonas aeruginosa in the lungs of cystic fibrosis patients. The antibiotic killing of bacteria in biofilms is generally known to be reduced by 100–1000 times relative to planktonic...... bacteria. This makes such infections difficult to treat. We have therefore proposed that biofilms can be regarded as an independent compartment with distinct pharmacokinetics. To elucidate this pharmacokinetics we have measured the penetration of the tobramycin into seaweed alginate beads which serve...... to be uniformly distributed throughout the volume of the alginate bead. The power-law appears to be a consequence of binding to a multitude of different binding sites. In a diffusion model these results are shown to produce pronounced retardation of the penetration of tobramycin into the biofilm. This filtering...

  16. Role of bacterial adhesion in the microbial ecology of biofilms in cooling tower systems.

    Science.gov (United States)

    Liu, Yang; Zhang, Wei; Sileika, Tadas; Warta, Richard; Cianciotto, Nicholas P; Packman, Aaron

    2009-01-01

    The fate of the three heterotrophic biofilm forming bacteria, Pseudomonas aeruginosa, Klebsiella pneumoniae and Flavobacterium sp. in pilot scale cooling towers was evaluated both by observing the persistence of each species in the recirculating water and the formation of biofilms on steel coupons placed in each cooling tower water reservoir. Two different cooling tower experiments were performed: a short-term study (6 days) to observe the initial bacterial colonization of the cooling tower, and a long-term study (3 months) to observe the ecological dynamics with repeated introduction of the test strains. An additional set of batch experiments (6 days) was carried out to evaluate the adhesion of each strain to steel surfaces under similar conditions to those found in the cooling tower experiments. Substantial differences were observed in the microbial communities that developed in the batch systems and cooling towers. P. aeruginosa showed a low degree of adherence to steel surfaces both in batch and in the cooling towers, but grew much faster than K. pneumoniae and Flavobacterium in mixed-species biofilms and ultimately became the dominant organism in the closed batch systems. However, the low degree of adherence caused P. aeruginosa to be rapidly washed out of the open cooling tower systems, and Flavobacterium became the dominant microorganism in the cooling towers in both the short-term and long-term experiments. These results indicate that adhesion, retention and growth on solid surfaces play important roles in the bacterial community that develops in cooling tower systems.

  17. Pseudomonas aeruginosa sabotages the generation of host proresolving lipid mediators

    Energy Technology Data Exchange (ETDEWEB)

    Flitter, Becca A.; Hvorecny, Kelli L.; Ono, Emiko; Eddens, Taylor; Yang, Jun; Kwak, Daniel H.; Bahl, Christopher D.; Hampton, Thomas H.; Morisseau, Christophe; Hammock, Bruce D.; Liu, Xinyu; Lee, Janet S.; Kolls, Jay K.; Levy, Bruce D.; Madden, Dean R.; Bomberger, Jennifer M.

    2016-12-15

    Recurrent Pseudomonas aeruginosa infections coupled with robust, damaging neutrophilic inflammation characterize the chronic lung disease cystic fibrosis (CF). The proresolving lipid mediator, 15-epi lipoxin A4 (15-epi LXA4), plays a critical role in limiting neutrophil activation and tissue inflammation, thus promoting the return to tissue homeostasis. Here, we show that a secreted P. aeruginosa epoxide hydrolase, cystic fibrosis transmembrane conductance regulator inhibitory factor (Cif), can disrupt 15-epi LXA4 transcellular biosynthesis and function. In the airway, 15-epi LXA4 production is stimulated by the epithelial-derived eicosanoid 14,15-epoxyeicosatrienoic acid (14,15-EET). Cif sabotages the production of 15-epi LXA4 by rapidly hydrolyzing 14,15-EET into its cognate diol, eliminating a proresolving signal that potently suppresses IL-8–driven neutrophil transepithelial migration in vitro. Retrospective analyses of samples from patients with CF supported the translational relevance of these preclinical findings. Elevated levels of Cif in bronchoalveolar lavage fluid were correlated with lower levels of 15-epi LXA4, increased IL-8 concentrations, and impaired lung function. Together, these findings provide structural, biochemical, and immunological evidence that the bacterial epoxide hydrolase Cif disrupts resolution pathways during bacterial lung infections. The data also suggest that Cif contributes to sustained pulmonary inflammation and associated loss of lung function in patients with CF.

  18. Why Does the Healthy Cornea Resist Pseudomonas aeruginosa Infection?

    Science.gov (United States)

    Evans, David J.; Fleiszig, Suzanne M. J.

    2013-01-01

    Purpose To provide our perspective on why the cornea is resistant to infection based on our research results with Pseudomonas aeruginosa. Perspective We focus on our current understanding of the interplay between bacteria, tear fluid and the corneal epithelium that determine health as the usual outcome, and propose a theoretical model for how contact lens wear might change those interactions to enable susceptibility to P. aeruginosa infection. Methods Use of “null-infection” in vivo models, cultured human corneal epithelial cells, contact lens-wearing animal models, and bacterial genetics help to elucidate mechanisms by which P. aeruginosa survive at the ocular surface, adheres, and traverses multilayered corneal epithelia. These models also help elucidate the molecular mechanisms of corneal epithelial innate defense. Results and Discussion Tear fluid and the corneal epithelium combine to make a formidable defense against P. aeruginosa infection of the cornea. Part of that defense involves the expression of antimicrobials such as β-defensins, the cathelicidin LL-37, cytokeratin-derived antimicrobial peptides, and RNase7. Immunomodulators such as SP-D and ST2 also contribute. Innate defenses of the cornea depend in part on MyD88, a key adaptor protein of TLR and IL-1R signaling, but the basal lamina represents the final barrier to bacterial penetration. Overcoming these defenses involves P. aeruginosa adaptation, expression of the type three secretion system, proteases, and P. aeruginosa biofilm formation on contact lenses. Conclusion After more than two decades of research focused on understanding how contact lens wear predisposes to P. aeruginosa infection, our working hypothesis places blame for microbial keratitis on bacterial adaptation to ocular surface defenses, combined with changes to the biochemistry of the corneal surface caused by trapping bacteria and tear fluid against the cornea under the lens. PMID:23601656

  19. Reduced intracellular c-di-GMP content increases expression of quorum sensing-regulated genes in Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Chua, Song Lin; Liu, Yang; Li, Yingying

    2017-01-01

    Cyclic-di-GMP (c-di-GMP) is an intracellular secondary messenger which controls the biofilm life cycle in many bacterial species. High intracellular c-di-GMP content enhances biofilm formation via the reduction of motility and production of biofilm matrix, while low c-di-GMP content in biofilm...... cells leads to increased motility and biofilm dispersal. While the effect of high c-di-GMP levels on bacterial lifestyles is well studied, the physiology of cells at low c-di-GMP levels remains unclear. Here, we showed that Pseudomonas aeruginosa cells with high and low intracellular c-di-GMP contents...... possessed distinct transcriptome profiles. There were 535 genes being upregulated and 432 genes downregulated in cells with low c-di-GMP, as compared to cells with high c-di-GMP. Interestingly, both rhl and pqs quorum-sensing (QS) operons were expressed at higher levels in cells with low intracellular c-di-GMP...

  20. Trigonella foenum-graceum (Seed Extract Interferes with Quorum Sensing Regulated Traits and Biofilm Formation in the Strains of Pseudomonas aeruginosa and Aeromonas hydrophila

    Directory of Open Access Journals (Sweden)

    Fohad Mabood Husain

    2015-01-01

    Full Text Available Trigonella foenum-graecum L. (Fenugreek is an important plant of the Leguminosae family known to have medicinal properties. However, fraction based antiquorum sensing and antibiofilm activities have not been reported from this plant. In the present study T. foenum-graecum seed extract was sequentially fractionated and sub-MICs were tested for above activities. The methanol fraction of the extract demonstrated significant inhibition of AHL regulated virulence factors: protease, LasB elastase, pyocyanin production, chitinase, EPS, and swarming motility in Pseudomonas aeruginosa PAO1 and PAF79. Further, QS dependent virulence factor in the aquatic pathogen Aeromonas hydrophila WAF38 was also reduced. Application of T. foenum-graecum seed extract to PAO1, PAF79, and WAF38 decreased the biofilm forming abilities of the pathogens by significant levels. The extract also exhibited reduced AHL levels and subsequent downregulation of lasB gene. In vivo study showed an enhanced survival of PAO1-preinfected C. elegans after treatment with extract at 1 mg/mL. Further, the major compound detected by GC-MS, caffeine, reduced the production of QS regulated virulence factors and biofilm at 200 µg/mL concentration indicating its role in the activity of the methanol extract. The results of the present study reveal the potential anti-QS and antibiofilm property of T. foenum-graceum extract and caffeine.

  1. Role of Multicellular Aggregates in Biofilm Formation

    Directory of Open Access Journals (Sweden)

    Kasper N. Kragh

    2016-03-01

    Full Text Available In traditional models of in vitro biofilm development, individual bacterial cells seed a surface, multiply, and mature into multicellular, three-dimensional structures. Much research has been devoted to elucidating the mechanisms governing the initial attachment of single cells to surfaces. However, in natural environments and during infection, bacterial cells tend to clump as multicellular aggregates, and biofilms can also slough off aggregates as a part of the dispersal process. This makes it likely that biofilms are often seeded by aggregates and single cells, yet how these aggregates impact biofilm initiation and development is not known. Here we use a combination of experimental and computational approaches to determine the relative fitness of single cells and preformed aggregates during early development of Pseudomonas aeruginosa biofilms. We find that the relative fitness of aggregates depends markedly on the density of surrounding single cells, i.e., the level of competition for growth resources. When competition between aggregates and single cells is low, an aggregate has a growth disadvantage because the aggregate interior has poor access to growth resources. However, if competition is high, aggregates exhibit higher fitness, because extending vertically above the surface gives cells at the top of aggregates better access to growth resources. Other advantages of seeding by aggregates, such as earlier switching to a biofilm-like phenotype and enhanced resilience toward antibiotics and immune response, may add to this ecological benefit. Our findings suggest that current models of biofilm formation should be reconsidered to incorporate the role of aggregates in biofilm initiation.

  2. Methods and Guidance for Testing the Efficacy of Antimicrobials against Biofilm Bacteria on Hard, Non-Porous Surfaces

    Science.gov (United States)

    EPA is announcing the availability of two test methods (MB-19 and MB-20) for evaluating the efficacy of antimicrobial pesticides against two biofilm bacteria, Pseudomonas aeruginosa and Staphylococcus aureus.

  3. Pyoverdine, the Major Siderophore in Pseudomonas aeruginosa, Evades NGAL Recognition

    Directory of Open Access Journals (Sweden)

    Mary E. Peek

    2012-01-01

    Full Text Available Pseudomonas aeruginosa is the most common pathogen that persists in the cystic fibrosis lungs. Bacteria such as P. aeruginosa secrete siderophores (iron-chelating molecules and the host limits bacterial growth by producing neutrophil-gelatinase-associated lipocalin (NGAL that specifically scavenges bacterial siderophores, therefore preventing bacteria from establishing infection. P. aeruginosa produces a major siderophore known as pyoverdine, found to be important for bacterial virulence and biofilm development. We report that pyoverdine did not bind to NGAL, as measured by tryptophan fluorescence quenching, while enterobactin bound to NGAL effectively causing a strong response. The experimental data indicate that pyoverdine evades NGAL recognition. We then employed a molecular modeling approach to simulate the binding of pyoverdine to human NGAL using NGAL’s published crystal structures. The docking of pyoverdine to NGAL predicted nine different docking positions; however, neither apo- nor ferric forms of pyoverdine docked into the ligand-binding site in the calyx of NGAL where siderophores are known to bind. The molecular modeling results offer structural support that pyoverdine does not bind to NGAL, confirming the results obtained in the tryptophan quenching assay. The data suggest that pyoverdine is a stealth siderophore that evades NGAL recognition allowing P. aeruginosa to establish chronic infections in CF lungs.

  4. Intrinsic and Extrinsic Aspects on Campylobacter jejuni Biofilms

    Directory of Open Access Journals (Sweden)

    Roberta T. Melo

    2017-07-01

    Full Text Available Biofilm represents a way of life that allows greater survival of microorganisms in hostile habitats. Campylobacter jejuni is able to form biofilms in vitro and on surfaces at several points in the poultry production chain. Genetic determinants related to their formation are expressed differently between strains and external conditions are decisive in this respect. Our approach combines phylogenetic analysis and the presence of seven specific genes linked to biofilm formation in association with traditional microbiology techniques, using Mueller Hinton and chicken juice as substrates in order to quantify, classify, determine the composition and morphology of the biomass of simple and mixed biofilms of 30 C. jejuni strains. It also evaluates the inhibition of its formation by biocides commonly used in industry and also by zinc oxide nanoparticles. Genetic analysis showed high heterogeneity with the identification of 23 pulsotypes. Despite the diversity, the presence of flaA, cadF, luxS, dnaJ, htrA, cbrA, and sodB genes in all strains shows the high potential for biofilm formation. This ability was only expressed in chicken juice, where they presented phenotype of a strong biofilm producer, with a mean count of 7.37 log CFU/mL and an ultrastructure characteristic of mature biofilm. The composition of simple and mixed biofilms was predominantly composed by proteins. The exceptions were found in mixed biofilms with Pseudomonas aeruginosa, which includes a carbohydrate-rich matrix, lower ability to sessile form in chicken juice and compact architecture of the biofilm, this aspects are intrinsic to this species. Hypochlorite, chlorhexidine, and peracetic acid were more effective in controlling viable cells of C. jejuni in biofilm, but the existence of tolerant strains indicates exposure to sublethal concentrations and development of adaptation mechanisms. This study shows that in chicken juice C. jejuni presents greater potential in producing mature

  5. Biofilm-mediated Antibiotic-resistant Oral Bacterial Infections: Mechanism and Combat Strategies.

    Science.gov (United States)

    Kanwar, Indulata; Sah, Abhishek K; Suresh, Preeti K

    2017-01-01

    Oral diseases like dental caries and periodontal disease are directly associated with the capability of bacteria to form biofilm. Periodontal diseases have been associated to anaerobic Gram-negative bacteria forming a subgingival plaque (Porphyromonas gingivalis, Actinobacillus, Prevotella and Fusobacterium). Biofilm is a complex bacterial community that is highly resistant to antibiotics and human immunity. Biofilm communities are the causative agents of biological developments such as dental caries, periodontitis, peri-implantitis and causing periodontal tissue breakdown. The review recapitulates the latest advancements in treatment of clinical biofilm infections and scientific investigations, while these novel anti-biofilm strategies are still in nascent phases of development, efforts dedicated to these technologies could ultimately lead to anti-biofilm therapies that are superior to the current antibiotic treatment. This paper provides a review of the literature focusing on the studies on biofilm in the oral cavity, formation of dental plaque biofilm, drug resistance of bacterial biofilm and the antibiofilm approaches as biofilm preventive agents in dentistry, and their mechanism of biofilm inhibition. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  6. Advances toward the elucidation of hypertonic saline effects on Pseudomonas aeruginosa from cystic fibrosis patients.

    Directory of Open Access Journals (Sweden)

    Anne-Laure Michon

    Full Text Available OBJECTIVES: Nebulized hypertonic saline (HTS has beneficial effects including reducing pulmonary exacerbations in Cystic Fibrosis (CF patients. Several mechanisms may explain these effects but antimicrobial activity of NaCl remains largely unexplored. We aimed to measure the antimicrobial effect of NaCl on Pseudomonas aeruginosa isolated from the respiratory tract in CF patients. METHODS: NaCl minimal inhibitory concentration (MIC and minimal bactericidal concentration (MBC were determined for strains characterized for mucoidy, antimicrobial resistance, and ability to form biofilm using 0,9% to 15% NaCl solutions. NaCl effects on biofilm formation, preformed biofilm, and mobility were evaluated. Kinetics of antimicrobial effects was studied. RESULTS: The growth of all isolates (n = 85 from 34 patients was inhibited by 6% NaCl solution. A 10% concentration had a bactericidal activity on 90% of the isolates. Mucoid and multidrug resistant (MDR isolates displayed lower MICs compared to non-mucoid and to non-MDR isolates, respectively. Time-kill kinetics showed that NaCl exhibited a rapid, dose and growth phase dependent bactericidal effect. Three percent or more of NaCl inhibited biofilm formation for 69% of strongly adherent isolates. A dose-dependent decrease of preformed biofilm viability and an inhibitory activity on bacterial motility were observed. CONCLUSIONS: NaCl inhibited the growth of all isolates and killed 38% of tested isolates within concentration range currently used in therapeutics. Our results suggest that anti-pseudomonal activity is another mechanism of action of HTS to add to those already established. Clinical trials are needed to compare diverse HTS conditions of use (rhythm, dose and mode of delivery to obtain efficient and optimized anti-P. aeruginosa effects. More generally, NaCl effect on other opportunistic pathogens as well as on global microbiotae recovered during polymicrobial diseases warrants further investigations.

  7. Control of Biofilms with the Fatty Acid Signaling Molecule cis-2-Decenoic Acid

    Directory of Open Access Journals (Sweden)

    Cláudia N. H. Marques

    2015-11-01

    Full Text Available Biofilms are complex communities of microorganisms in organized structures attached to surfaces. Importantly, biofilms are a major cause of bacterial infections in humans, and remain one of the most significant challenges to modern medical practice. Unfortunately, conventional therapies have shown to be inadequate in the treatment of most chronic biofilm infections based on the extraordinary innate tolerance of biofilms to antibiotics. Antagonists of quorum sensing signaling molecules have been used as means to control biofilms. QS and other cell-cell communication molecules are able to revert biofilm tolerance, prevent biofilm formation and disrupt fully developed biofilms, albeit with restricted effectiveness. Recently however, it has been demonstrated that Pseudomonas aeruginosa produces a small messenger molecule cis-2-decenoic acid (cis-DA that shows significant promise as an effective adjunctive to antimicrobial treatment of biofilms. This molecule is responsible for induction of the native biofilm dispersion response in a range of Gram-negative and Gram-positive bacteria and in yeast, and has been shown to reverse persistence, increase microbial metabolic activity and significantly enhance the cidal effects of conventional antimicrobial agents. In this manuscript, the use of cis-2-decenoic acid as a novel agent for biofilm control is discussed. Stimulating the biofilm dispersion response as a novel antimicrobial strategy holds significant promise for enhanced treatment of infections and in the prevention of biofilm formation.

  8. Pharmacokinetics and pharmacodynamics of antibiotics in biofilm infections of Pseudomonas aeruginosa in vitro and in vivo

    DEFF Research Database (Denmark)

    Hengzhuang, Wang; Høiby, Niels; Ciofu, Oana

    2014-01-01

    Although progress on biofilm research has been obtained during the past decades, the treatment of biofilm infections with antibiotics remains a riddle. The pharmacokinetic (PK) and pharmacodynamic (PD) profiles of an antimicrobial agent provide important information helping to establish an effici......Although progress on biofilm research has been obtained during the past decades, the treatment of biofilm infections with antibiotics remains a riddle. The pharmacokinetic (PK) and pharmacodynamic (PD) profiles of an antimicrobial agent provide important information helping to establish...

  9. The efficacy of sewage influent-isolated bacteriophages on Pseudomonas aeruginosa in a mixed-species biofilm

    KAUST Repository

    Yap, Scott

    2016-01-01

    The growth of environmentally persistent biofilms in cooling towers causes several associated problems, including microbiologically-induced corrosion (MIC) and biofouling. Current chemical control methods are not only ineffective against biofilms

  10. Conjugative type IVb pilus recognizes lipopolysaccharide of recipient cells to initiate PAPI-1 pathogenicity island transfer in Pseudomonas aeruginosa

    Science.gov (United States)

    Pseudomonas aeruginosa pathogenicity island 1 (PAPI-1) is one of the largest genomic islands of this important opportunistic human pathogen. Previous studies have shown that PAPI-1 encodes several putative virulence factors, a major regulator of biofilm formation, and antibiotic-resistance traits, a...

  11. Efficacy of Poly-Lactic-Co-Glycolic Acid Micro- and Nanoparticles of Ciprofloxacin Against Bacterial Biofilms.

    Science.gov (United States)

    Thomas, Nicky; Thorn, Chelsea; Richter, Katharina; Thierry, Benjamin; Prestidge, Clive

    2016-10-01

    Bacterial biofilms are associated with a number of recurring infectious diseases and are a major cause for antibiotic resistance. Despite the broad use of polymeric microparticles and nanoparticles in biomedical research, it is not clear which particle size is more effective against biofilms. The purpose of this study was to evaluate the efficacy of sustained release poly-lactic-co-glycolic acid (PLGA) micro- and nanoparticles containing ciprofloxacin against biofilms of Staphylococcus aureus and Pseudomonas aeruginosa. The PLGA particles were prepared by the double emulsion solvent evaporation method. The resulting microparticles (12 μm) and nanoparticles (300 nm) contained drug loads of 7.3% and 4.5% (wt/wt) ciprofloxacin, respectively. Drug release was complete within 1 week following comparable release profiles for both particle sizes. Micro- and nanoparticles demonstrated a similar in vitro antibiofilm performance against mature P aeruginosa and S aureus with marked differences between the 2 strains. The sustained release of ciprofloxacin from micro- and nanoparticles over 6 days was equally effective as the continuous treatment with ciprofloxacin solution over the same period resulting in the eradication of culturable S aureus suggesting that reformulation of ciprofloxacin as sustained release PLGA micro- and nanoparticles might be valuable formulation approaches for the treatment of biofilms. Copyright © 2016 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

  12. Visualizing Antimicrobials in Bacterial Biofilms: Three-Dimensional Biochemical Imaging Using TOF-SIMS.

    Science.gov (United States)

    Davies, Sarah K; Fearn, Sarah; Allsopp, Luke P; Harrison, Freya; Ware, Ecaterina; Diggle, Stephen P; Filloux, Alain; McPhail, David S; Bundy, Jacob G

    2017-01-01

    Bacterial biofilms are groups of bacteria that exist within a self-produced extracellular matrix, adhering to each other and usually to a surface. They grow on medical equipment and inserts such as catheters and are responsible for many persistent infections throughout the body, as they can have high resistance to many antimicrobials. Pseudomonas aeruginosa is an opportunistic pathogen that can cause both acute and chronic infections and is used as a model for research into biofilms. Direct biochemical methods of imaging of molecules in bacterial biofilms are of high value in gaining a better understanding of the fundamental biology of biofilms and biochemical gradients within them. Time of flight-secondary-ion mass spectrometry (TOF-SIMS) is one approach, which combines relatively high spatial resolution and sensitivity and can perform depth profiling analysis. It has been used to analyze bacterial biofilms but has not yet been used to study the distribution of antimicrobials (including antibiotics and the antimicrobial metal gallium) within biofilms. Here we compared two methods of imaging of the interior structure of P. aeruginosa in biological samples using TOF-SIMS, looking at both antimicrobials and endogenous biochemicals: cryosectioning of tissue samples and depth profiling to give pseudo-three-dimensional (pseudo-3D) images. The sample types included both simple biofilms grown on glass slides and bacteria growing in tissues in an ex vivo pig lung model. The two techniques for the 3D imaging of biofilms are potentially valuable complementary tools for analyzing bacterial infection. IMPORTANCE Modern analytical techniques are becoming increasingly important in the life sciences; imaging mass spectrometry offers the opportunity to gain unprecedented amounts of information on the distribution of chemicals in samples-both xenobiotics and endogenous compounds. In particular, simultaneous imaging of antibiotics (and other antimicrobial compounds) and bacterium

  13. Bioprinting Living Biofilms through Optogenetic Manipulation.

    Science.gov (United States)

    Huang, Yajia; Xia, Aiguo; Yang, Guang; Jin, Fan

    2018-04-18

    In this paper, we present a new strategy for microprinting dense bacterial communities with a prescribed organization on a substrate. Unlike conventional bioprinting techniques that require bioinks, through optogenetic manipulation, we directly manipulated the behaviors of Pseudomonas aeruginosa to allow these living bacteria to autonomically form patterned biofilms following prescribed illumination. The results showed that through optogenetic manipulation, patterned bacterial communities with high spatial resolution (approximately 10 μm) could be constructed in 6 h. Thus, optogenetic manipulation greatly increases the range of available bioprinting techniques.

  14. Attenuation of Pseudomonas aeruginosa quorum sensing, virulence and biofilm formation by extracts of Andrographis paniculata.

    Science.gov (United States)

    Banerjee, Malabika; Moulick, Soumitra; Bhattacharya, Kunal Kumar; Parai, Debaprasad; Chattopadhyay, Subrata; Mukherjee, Samir Kumar

    2017-12-01

    Quorum-sensing (QS) is known to play an essential role in regulation of virulence factors and toxins during Pseudomonas aeruginosa infection which may frequently cause antibiotic resistance and hostile outcomes of inflammatory injury. Therefore, it is an urgent need to search for a novel agent with low risk of resistance development that can target QS and inflammatory damage prevention as well. Andrographis paniculata, a herbaceous plant under the family Acanthaceae, native to Asian countries and also cultivated in Scandinavia and some parts of Europe, has a strong traditional usage with its known antibacterial, anti-inflammatory, antipyretic, antiviral and antioxidant properties. In this study, three different solvent extracts (viz., chloroform, methanol and aqueous) of A. paniculata were examined for their anti-QS and anti-inflammatory activities. Study was carried out to assess the effect on some selected QS-regulatory genes at transcriptional level using Real Time-PCR. In addition, ability to attenuate MAPK pathways upon P. aeruginosa infection was performed to check its potential anti-inflammatory activity. Chloroform and methanol extracts showed significant reduction (p paniculata extracts inhibit QS in P. aeruginosa and exhibit anti-inflammatory activities, therefore it represents itself as a prospective therapeutic agent against P. aeruginosa infection. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. SuhB Is a Regulator of Multiple Virulence Genes and Essential for Pathogenesis of Pseudomonas aeruginosa

    Science.gov (United States)

    Li, Kewei; Xu, Chang; Jin, Yongxin; Sun, Ziyu; Liu, Chang; Shi, Jing; Chen, Gukui; Chen, Ronghao; Jin, Shouguang; Wu, Weihui

    2013-01-01

    ABSTRACT During initial colonization and chronic infection, pathogenic bacteria encounter distinct host environments. Adjusting gene expression accordingly is essential for the pathogenesis. Pseudomonas aeruginosa has evolved complicated regulatory networks to regulate different sets of virulence factors to facilitate colonization and persistence. The type III secretion system (T3SS) and motility are associated with acute infections, while biofilm formation and the type VI secretion system (T6SS) are associated with chronic persistence. To identify novel regulatory genes required for pathogenesis, we screened a P. aeruginosa transposon (Tn) insertion library and found suhB to be an essential gene for the T3SS gene expression. The expression of suhB was upregulated in a mouse acute lung infection model, and loss of suhB resulted in avirulence. Suppression of T3SS gene expression in the suhB mutant is linked to a defective translation of the T3SS master regulator, ExsA. Further studies demonstrated that suhB mutation led to the upregulation of GacA and its downstream small RNAs, RsmY and RsmZ, triggering T6SS expression and biofilm formation while inhibiting the T3SS. Our results demonstrate that an in vivo-inducible gene, suhB, reciprocally regulates genes associated with acute and chronic infections and plays an essential role in the pathogenesis of P. aeruginosa. PMID:24169572

  16. Pseudomonas aeruginosa biofilm formation and slime excretion on antibiotic-loaded bone cement

    NARCIS (Netherlands)

    Neut, D; Hendriks, JGE; van Horn, Jim R.; van der Mei, HC; Busscher, HJ

    Background Infection is an infrequent but serious complication of prosthetic joint surgery. These infections will usually not clear until the implant is removed and re-implantation has a high failure rate, especially when Pseudomonas aeruginosa is involved. Material and methods We examined

  17. Expansion of plasmid mediated blaACT-2 among Pseudomonas aeruginosa associated with postoperative infection and its transcriptional response under cephalosporin stress.

    Directory of Open Access Journals (Sweden)

    Birson Ingti, Deepjyoti Paul, Anand Prakash Maurya

    2017-06-01

    Full Text Available Objectives: Organisms harboring multiple plasmid mediated β-lactamases are major concerns in nosocomial infections. Among these plasmid mediated β-lactamases, ACT (EBC family is a clinically important enzyme capable of hydrolyzing broad spectrum cephalosporins. Therefore, the present study was undertaken to determine the prevalence of ACT determinant along with other co-existing β-lactamase genes in P. aeruginosa strains. Methods: A total of 176 Pseudomonas isolates were phenotypically screened for the presence of AmpC β-lactamase by M3DET Method followed by Molecular detection using PCR assay. Transcriptional evaluation of blaACT-2 gene was analyzed by RT-PCR and its transferability was performed by transformation and conjugation. Results: Present study demonstrates the presence of ACT-2 allele among 12 strains of P. aeruginosa. Co-existence of other β-lactamase genes were encountered among ACT-2 harboring strains which includes CTX-M (n=2, SHV (n=3, TEM (n=2, VEB (n=2, OXA-10 (n=1, CIT (n=2 and DHA (n=3. Fingerprinting by REP PCR revealed the isolates harboring ACT-2 to be distinct and these isolates showed high resistance to expanded-spectrum cephalosporins and even to carbapenem group of drugs. This ACT-2 allele was encoded in the plasmid (L/M, FIA, FIB Inc. Group and conjugatively transferable. Transcriptional analysis revealed a significant increase in ACT-2 expression (483 fold when induced by ceftriaxone at 4 µg/ml followed by ceftazidime at 8 µg/ml (31 fold and cefotaxime 4 µg/ml (8 fold. Conclusion: In this study detection of ACT-2 plasmid mediated AmpC β-lactamase along with other β-lactamase genes in clinical isolates of P. aeruginosa represents a serious therapeutic challenge. Therefore, revision in antimicrobial policy is required for effective treatment of patients infected with pathogen expressing this mechanism. J Microbiol Infect Dis 2017; 7(2: 75-82

  18. Cell surface physico chemistry alters biofilm development of Pseudomonas aeruginosa lipopolysaccharide mutants

    NARCIS (Netherlands)

    Flemming, CA; Palmer, RJ; Arrage, AA; van der Mei, H.C.; White, DC

    1999-01-01

    The hydrophobic and electrostatic characteristics of bacterial cell surfaces were compared with attachment proclivity and biomass accumulation over time between wildtype Pseudomonas aeruginosa serotype O6 (possesses A and B band LPS), and three LPS-deficient mutants, vi;. A28 (A(+)B(-)), R5

  19. Mechanics governs single-cell signaling and multi-cell robustness in biofilm infections

    Science.gov (United States)

    Gordon, Vernita

    In biofilms, bacteria and other microbes are embedded in extracellular polymers (EPS). Multiple types of EPS can be produced by a single bacterial strain - the reasons for this redundancy are not well-understood. Our work suggests that different polymers may confer distinct mechanical benefits. Our model organism is Pseudomonas aeruginosa, an opportunistic human pathogen that forms chronic biofilm infections associated with increased antibiotic resistance and evasion of the immune defense. Biofilms initiate when bacteria attach to a surface, sense the surface, and change their gene expression. Changes in gene expression are regulated by a chemical signal, cyclic-di-GMP. We find that one EPS material, called ``PEL,'' enhances surface sensing by increasing mechanical coupling of single bacteria to the surface. Measurements of bacterial motility suggest that PEL may increase frictional interactions between the surface and the bacteria. Consistent with this, we show that bacteria increase cyclic-di-GMP signaling in response to mechanical shear stress. Mechanosensing has long been known to be important to the function of cells in higher eukaryotes, but this is one of only a handful of studies showing that bacteria can sense and respond to mechanical forces. For the mature biofilm, the embedding polymer matrix can protect bacteria both chemically and mechanically. P. aeruginosa infections in the cystic fibrosis (CF) lung often last for decades, ample time for the infecting strain(s) to evolve. Production of another EPS material, alginate, is well-known to tend to increase over time in CF infections. Alginate chemically protects biofilms, but also makes them softer and weaker. Recently, it is being increasingly recognized that bacteria in chronic CF infections also evolve to increase PSL production. We use oscillatory bulk rheology to determine the unique contributions of EPS materials to biofilm mechanics. Unlike alginate, increased PSL stiffens biofilms. Increasing both

  20. Mutation of the Streptococcus gordonii Thiol-Disulfide Oxidoreductase SdbA Leads to Enhanced Biofilm Formation Mediated by the CiaRH Two-Component Signaling System.

    Directory of Open Access Journals (Sweden)

    Lauren Davey

    Full Text Available Streptococcus gordonii is a commensal inhabitant of human oral biofilms. Previously, we identified an enzyme called SdbA that played an important role in biofilm formation by S. gordonii. SdbA is thiol-disulfide oxidoreductase that catalyzes disulfide bonds in secreted proteins. Surprisingly, inactivation of SdbA results in enhanced biofilm formation. In this study we investigated the basis for biofilm formation by the ΔsdbA mutant. The results revealed that biofilm formation was mediated by the interaction between the CiaRH and ComDE two-component signalling systems. Although it did not affect biofilm formation by the S. gordonii parent strain, CiaRH was upregulated in the ΔsdbA mutant and it was essential for the enhanced biofilm phenotype. The biofilm phenotype was reversed by inactivation of CiaRH or by the addition of competence stimulating peptide, the production of which is blocked by CiaRH activity. Competition assays showed that the enhanced biofilm phenotype also corresponded to increased oral colonization in mice. Thus, the interaction between SdbA, CiaRH and ComDE affects biofilm formation both in vitro and in vivo.

  1. Development of molecularly imprinted polymers to block quorum sensing and inhibit bacterial biofilm formation.

    Science.gov (United States)

    Ma, Luyao; Feng, Shaolong; de la Fuente-Nunez, Cesar; Hancock, Robert E W; Lu, Xiaonan

    2018-05-16

    Bacterial biofilms are responsible for most clinical infections and show increased antimicrobial resistance. In this study, molecularly imprinted polymers (MIPs) were developed to specifically capture prototypical quorum sensing autoinducers [i.e., N-(3-oxododecanoyl)-L-homoserine lactone (3-oxo-C12AHL)], interrupt quorum sensing, and subsequently inhibit biofilm formation of Pseudomonas aeruginosa, an important human nosocomial pathogen. The synthesis of MIPs was optimized by considering the amount and type of the functional monomers itaconic acid (IA) and 2-hydroxyethyl methacrylate (HEMA). IA-based MIPs showed high adsorption affinity towards 3-oxo-C12AHL with an imprinting factor of 1.68. Compared to IA-based MIPs, the adsorption capacity of HEMA-based MIPs was improved 5-fold. HEMA-based MIPs significantly reduced biofilm formation (by ~65%), while biofilm suppression by IA-based MIPs was neutralized due to increased bacterial attachment. The developed MIPs represent promising alternative biofilm intervention agents that can be applied to surfaces relevant to clinical settings and food processing equipment.

  2. Control of bacterial biofilm growth on surfaces by nanostructural mechanics and geometry

    International Nuclear Information System (INIS)

    Epstein, A K; Hochbaum, A I; Kim, Philseok; Aizenberg, J

    2011-01-01

    Surface-associated communities of bacteria, called biofilms, pervade natural and anthropogenic environments. Mature biofilms are resistant to a wide range of antimicrobial treatments and therefore pose persistent pathogenic threats. The use of surface chemistry to inhibit biofilm growth has been found to only transiently affect initial attachment. In this work, we investigate the tunable effects of physical surface properties, including high-aspect-ratio (HAR) surface nanostructure arrays recently reported to induce long-range spontaneous spatial patterning of bacteria on the surface. The functional parameters and length scale regimes that control such artificial patterning for the rod-shaped pathogenic species Pseudomonas aeruginosa are elucidated through a combinatorial approach. We further report a crossover regime of biofilm growth on a HAR nanostructured surface versus the nanostructure effective stiffness. When the 'softness' of the hair-like nanoarray is increased beyond a threshold value, biofilm growth is inhibited as compared to a flat control surface. This result is consistent with the mechanoselective adhesion of bacteria to surfaces. Therefore by combining nanoarray-induced bacterial patterning and modulating the effective stiffness of the nanoarray—thus mimicking an extremely compliant flat surface—bacterial mechanoselective adhesion can be exploited to control and inhibit biofilm growth.

  3. Microbial-Influenced Corrosion of Corten Steel Compared with Carbon Steel and Stainless Steel in Oily Wastewater by Pseudomonas aeruginosa

    Science.gov (United States)

    Mansouri, Hamidreza; Alavi, Seyed Abolhasan; Fotovat, Meysam

    2015-07-01

    The microbial corrosion behavior of three important steels (carbon steel, stainless steel, and Corten steel) was investigated in semi petroleum medium. This work was done in modified nutrient broth (2 g nutrient broth in 1 L oily wastewater) in the presence of Pseudomonas aeruginosa and mixed culture (as a biotic media) and an abiotic medium for 2 weeks. The behavior of corrosion was analyzed by spectrophotometric and electrochemical methods and at the end was confirmed by scanning electron microscopy. The results show that the degree of corrosion of Corten steel in mixed culture, unlike carbon steel and stainless steel, is less than P. aeruginosa inoculated medium because some bacteria affect Corten steel less than other steels. According to the experiments, carbon steel had less resistance than Corten steel and stainless steel. Furthermore, biofilm inhibits separated particles of those steels to spread to the medium; in other words, particles get trapped between biofilm and steel.

  4. High-resolution visualization of Pseudomonas aeruginosa PAO1 biofilms by freeze-substitution transmission electron microscopy.

    Science.gov (United States)

    Hunter, Ryan C; Beveridge, Terry J

    2005-11-01

    High-pressure freeze-substitution and transmission electron microscopy have been used for high-resolution imaging of the natural structure of a gram-negative biofilm. Unlike more conventional embedding techniques, this method confirms many of the observations seen by confocal microscopy but with finer structural detail. It further reveals that there is a structural complexity to biofilms at both the cellular and extracellular matrix levels that has not been seen before. Different domains of healthy and lysed cells exist randomly dispersed within a single biofilm as well as different structural organizations of exopolymers. Particulate matter is suspended within this network of fibers and appears to be an integral part of the exopolymeric substance (EPS). O-side chains extending from the outer membrane are integrated into EPS polymers so as to form a continuum. Together, the results support the concept of physical microenvironments within biofilms and show a complexity that was hitherto unknown.

  5. OligoG CF-5/20 disruption of mucoid Pseudomonas aeruginosa biofilm in a murine lung infection model

    DEFF Research Database (Denmark)

    Wang, Hengzhuang; Song, Zhijun; Ciofu, Oana

    2016-01-01

    Biofilm growth is a universal survival strategy for bacteria, providing an effective and resilient approach for survival in an otherwise hostile environment. In the context of an infection, a biofilm provides resistance and tolerance to host immune defenses and antibiotics, allowing the biofilm p...

  6. Origanum vulgare subsp. hirtum essential oil prevented biofilm formation and showed antibacterial activity against planktonic and sessile bacterial cells.

    Science.gov (United States)

    Schillaci, Domenico; Napoli, Edoardo Marco; Cusimano, Maria Grazia; Vitale, Maria; Ruberto, Andgiuseppe

    2013-10-01

    Essential oils from six different populations of Origanum vulgare subsp. hirtum were compared for their antibiofilm properties. The six essential oils (A to F) were characterized by a combination of gas chromatography with flame ionization detector and gas chromatography with mass spectrometer detector analyses. All oils showed weak activity against the planktonic form of a group of Staphylococcus aureus strains and against a Pseudomonas aeruginosa ATCC 15442 reference strain. The ability to inhibit biofilm formation was investigated at sub-MIC levels of 200, 100, and 50 m g/ml by staining sessile cells with safranin. Sample E showed the highest average effectiveness against all tested strains at 50 m g/ml and had inhibition percentages ranging from 30 to 52%. In the screening that used preformed biofilm from the reference strain P. aeruginosa, essential oils A through E were inactive at 200 m g/ml; F was active with a percentage of inhibition equal to 53.2%. Oregano essential oil can inhibit the formation of biofilms of various food pathogens and food spoilage organisms.

  7. Killing of Pseudomonas aeruginosa by Chicken Cathelicidin-2 Is Immunogenically Silent, Preventing Lung Inflammation In Vivo

    Science.gov (United States)

    Coorens, Maarten; Banaschewski, Brandon J. H.; Baer, Brandon J.; Yamashita, Cory; van Dijk, Albert; Veldhuizen, Ruud A. W.; Veldhuizen, Edwin J. A.

    2017-01-01

    ABSTRACT The development of antibiotic resistance by Pseudomonas aeruginosa is a major concern in the treatment of bacterial pneumonia. In the search for novel anti-infective therapies, the chicken-derived peptide cathelicidin-2 (CATH-2) has emerged as a potential candidate, with strong broad-spectrum antimicrobial activity and the ability to limit inflammation by inhibiting Toll-like receptor 2 (TLR2) and TLR4 activation. However, as it is unknown how CATH-2 affects inflammation in vivo, we investigated how CATH-2-mediated killing of P. aeruginosa affects lung inflammation in a murine model. First, murine macrophages were used to determine whether CATH-2-mediated killing of P. aeruginosa reduced proinflammatory cytokine production in vitro. Next, a murine lung model was used to analyze how CATH-2-mediated killing of P. aeruginosa affects neutrophil and macrophage recruitment as well as cytokine/chemokine production in the lung. Our results show that CATH-2 kills P. aeruginosa in an immunogenically silent manner both in vitro and in vivo. Treatment with CATH-2-killed P. aeruginosa showed reduced neutrophil recruitment to the lung as well as inhibition of cytokine and chemokine production, compared to treatment with heat- or gentamicin-killed bacteria. Together, these results show the potential for CATH-2 as a dual-activity antibiotic in bacterial pneumonia, which can both kill P. aeruginosa and prevent excessive inflammation. PMID:28947647

  8. Catheter-related infections caused by Pseudomonas aeruginosa: virulence factors involved and their relationships.

    Science.gov (United States)

    Olejnickova, Katerina; Hola, Veronika; Ruzicka, Filip

    2014-11-01

    The nosocomial pathogen Pseudomonas aeruginosa is equipped with a large arsenal of cell-associated and secreted virulence factors which enhance its invasive potential. The complex relationships among virulence determinants have hitherto not been fully elucidated. In the present study, 175 catheter-related isolates were observed for the presence of selected virulence factors, namely extracellular enzymes and siderophore production, biofilm formation, resistance to antibiotics, and motility. A high percentage of the strains produced most of the tested virulence factors. A positive correlation was identified between the production of several exoproducts, and also between the formation of both types of biofilm. An opposite trend was observed between the two types of biofilm and the production of siderophores. Whereas the relationship between the submerged biofilm production (i.e. the biofilm formed on the solid surface below the water level) and the siderophore secretion was negative, the production of air-liquid interface (A-L) biofilm (i.e. the biofilm floating on the surface of the cultivation medium) and the siderophore secretion were positively correlated. All correlations were statistically significant at the level P = 0.05 with the correlation coefficient γ ≥ 0.50. Our results suggest that: (1) the co-production of the lytic enzymes and siderophores can play an important role in the pathogenesis of the catheter-related infections and should be taken into account when the virulence potential is assessed; (2) biofilm-positive strains are capable of forming both submerged and non-attached A-L biofilms; and (3) the different micro-environment in the submerged biofilm and A-L biofilm layers have opposite consequences for the production of other virulence factors. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  9. Ciprofloxacin-loaded PLGA nanoparticles against cystic fibrosis P. aeruginosa lung infections.

    Science.gov (United States)

    Günday Türeli, Nazende; Torge, Afra; Juntke, Jenny; Schwarz, Bianca C; Schneider-Daum, Nicole; Türeli, Akif Emre; Lehr, Claus-Michael; Schneider, Marc

    2017-08-01

    Current pulmonary treatments against Pseudomonas aeruginosa infections in cystic fibrosis (CF) lung suffer from deactivation of the drug and immobilization in thick and viscous biofilm/mucus blend, along with the general antibiotic resistance. Administration of nanoparticles (NPs) with high antibiotic load capable of penetrating the tight mesh of biofilm/mucus can be an advent to overcome the treatment bottlenecks. Biodegradable and biocompatible polymer nanoparticles efficiently loaded with ciprofloxacin complex offer a solution for emerging treatment strategies. NPs were prepared under controlled conditions by utilizing MicroJet Reactor (MJR) to yield a particle size of 190.4±28.6nm with 0.089 PDI. Encapsulation efficiency of the drug was 79% resulting in a loading of 14%. Release was determined to be controlled and medium-independent in PBS, PBS+0.2% Tween 80 and simulated lung fluid. Cytotoxicity assays with Calu-3 cells and CF bronchial epithelial cells (CFBE41o - ) indicated that complex-loaded PLGA NPs were non-toxic at concentrations ≫ MIC cipro against lab strains of the bacteria. Antibacterial activity tests revealed enhanced activity when applied as nanoparticles. NPs' colloidal stability in mucus was proven. Notably, a decrease in mucus turbidity was observed upon incubation with NPs. Herewith, ciprofloxacin complex-loaded PLGA NPs are introduced as promising pulmonary nano drug delivery systems against P.aeruginosa infections in CF lung. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. Antimicrobial resistance, respiratory tract infections and role of biofilms in lung infections in cystic fibrosis patients

    DEFF Research Database (Denmark)

    Ciofu, Oana; Tolker-Nielsen, Tim; Jensen, Peter Østrup

    2015-01-01

    Lung infection is the main cause of morbidity and mortality in patients with cystic fibrosis and is mainly dominated by Pseudomonas aeruginosa. The biofilm mode of growth makes eradication of the infection impossible, and it causes a chronic inflammation in the airways. The general mechanisms...

  11. Genome-Wide Survey of Pseudomonas aeruginosa PA14 Reveals a Role for the Glyoxylate Pathway and Extracellular Proteases in the Utilization of Mucin

    Science.gov (United States)

    Flynn, Jeffrey M.; Phan, Chi

    2017-01-01

    ABSTRACT Chronic airway infections by the opportunistic pathogen Pseudomonas aeruginosa are a major cause of mortality in cystic fibrosis (CF) patients. Although this bacterium has been extensively studied for its virulence determinants, biofilm growth, and immune evasion mechanisms, comparatively little is known about the nutrient sources that sustain its growth in vivo. Respiratory mucins represent a potentially abundant bioavailable nutrient source, although we have recently shown that canonical pathogens inefficiently use these host glycoproteins as a growth substrate. However, given that P. aeruginosa, particularly in its biofilm mode of growth, is thought to grow slowly in vivo, the inefficient use of mucin glycoproteins may be relevant to its persistence within the CF airways. To this end, we used whole-genome fitness analysis, combining transposon mutagenesis with high-throughput sequencing, to identify genetic determinants required for P. aeruginosa growth using intact purified mucins as a sole carbon source. Our analysis reveals a biphasic growth phenotype, during which the glyoxylate pathway and amino acid biosynthetic machinery are required for mucin utilization. Secondary analyses confirmed the simultaneous liberation and consumption of acetate during mucin degradation and revealed a central role for the extracellular proteases LasB and AprA. Together, these studies describe a molecular basis for mucin-based nutrient acquisition by P. aeruginosa and reveal a host-pathogen dynamic that may contribute to its persistence within the CF airways. PMID:28507068

  12. New method for characterizing electron mediators in microbial systems using a thin-layer twin-working electrode cell.

    Science.gov (United States)

    Hassan, Md Mahamudul; Cheng, Ka Yu; Ho, Goen; Cord-Ruwisch, Ralf

    2017-01-15

    Microbial biofilms are significant ecosystems where the existence of redox gradients drive electron transfer often via soluble electron mediators. This study describes the use of two interfacing working electrodes (WEs) to simulate redox gradients within close proximity (250µm) for the detection and quantification of electron mediators. By using a common counter and reference electrode, the potentials of the two WEs were independently controlled to maintain a suitable "voltage window", which enabled simultaneous oxidation and reduction of electron mediators as evidenced by the concurrent anodic and cathodic currents, respectively. To validate the method, the electrochemical properties of different mediators (hexacyanoferrate, HCF, riboflavin, RF) were characterized by stepwise shifting the "voltage window" (ranging between 25 and 200mV) within a range of potentials after steady equilibrium current of both WEs was established. The resulting differences in electrical currents between the two WEs were recorded across a defined potential spectrum (between -1V and +0.5V vs. Ag/AgCl). Results indicated that the technique enabled identification (by the distinct peak locations at the potential scale) and quantification (by the peak of current) of the mediators for individual species as well as in an aqueous mixture. It enabled a precise determination of mid-potentials of the externally added mediators (HCF, RF) and mediators produced by pyocyanin-producing Pseudomonas aeruginosa (WACC 91) culture. The twin working electrode described is particularly suitable for studying mediator-dependent microbial electron transfer processes or simulating redox gradients as they exist in microbial biofilms. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Mechanism of action and in vitro activity of short hybrid antimicrobial peptide PV3 against Pseudomonas aeruginosa

    International Nuclear Information System (INIS)

    Memariani, Hamed; Shahbazzadeh, Delavar; Sabatier, Jean-Marc; Memariani, Mojtaba; Karbalaeimahdi, Ali; Bagheri, Kamran Pooshang

    2016-01-01

    Antimicrobial peptides are attractive candidates for developing novel therapeutic agents, since they are lethal to a broad spectrum of pathogens and have a unique low tendency for resistance development. In this study, mechanism of action and in vitro anti-pseudomonal activity of previously designed short hybrid antimicrobial peptide PV3 were investigated. Compared to ceftazidime, PV3 had not only higher antibacterial activity but also faster bactericidal activity. PV3 reduced biofilm biomass and viability of biofilm embedded bacteria in a concentration-dependent manner. Although the antimicrobial activity of PV3 was reduced in Mueller-Hinton broth (MHB) containing human serum, it was still active enough to eradication of bacteria at low concentrations. Compared with standard condition (MHB only), there was no significant decrease in antibacterial activity of PV3 against P. aeruginosa strains under 150 mM NaCl (p = 0.615) and 1 mM MgCl 2 (p = 0.3466). Fluorescence microscopy and field emission scanning electron microscopy further indicated that PV3 killed bacteria by disrupting the cell membrane. Since PV3 has potent anti-pseudomonal activity and has little cytotoxicity in vitro, it seems plausible that the peptide should be further investigated with animal studies to support future pharmacological formulations and potential topical applications. - Highlights: • PV3 killed Pseudomonas aeruginosa by membrane-disrupting mechanism. • PV3 reduced biofilm biomass and viability of biofilm embedded bacteria in a concentration-dependent manner. • Short hybrid antimicrobial peptide PV3 exhibited higher and faster bactericidal activity comparing to ceftazidime.

  14. Feeding Behavior Modulates Biofilm-Mediated Transmission of Yersinia pestis by the Cat Flea, Ctenocephalides felis

    Science.gov (United States)

    Bland, David M.; Hinnebusch, B. Joseph

    2016-01-01

    Background The cat flea, Ctenocephalides felis, is prevalent worldwide, will parasitize animal reservoirs of plague, and is associated with human habitations in known plague foci. Despite its pervasiveness, limited information is available about the cat flea’s competence as a vector for Yersinia pestis. It is generally considered to be a poor vector, based on studies examining early-phase transmission during the first week after infection, but transmission potential by the biofilm-dependent proventricular-blocking mechanism has never been systematically evaluated. In this study, we assessed the vector competence of cat fleas by both mechanisms. Because the feeding behavior of cat fleas differs markedly from important rat flea vectors, we also examined the influence of feeding behavior on transmission dynamics. Methodology/Principal Findings Groups of cat fleas were infected with Y. pestis and subsequently provided access to sterile blood meals twice-weekly, 5 times per week, or daily for 4 weeks and monitored for infection, the development of proventricular biofilm and blockage, mortality, and the ability to transmit. In cat fleas allowed prolonged, daily access to blood meals, mimicking their natural feeding behavior, Y. pestis did not efficiently colonize the digestive tract and could only be transmitted during the first week after infection. In contrast, cat fleas that were fed intermittently, mimicking the feeding behavior of the efficient vector Xenopsylla cheopis, could become blocked and regularly transmitted Y. pestis for 3–4 weeks by the biofilm-mediated mechanism, but early-phase transmission was not detected. Conclusions The normal feeding behavior of C. felis, more than an intrinsic resistance to infection or blockage by Y. pestis, limits its vector competence. Rapid turnover of midgut contents results in bacterial clearance and disruption of biofilm accumulation in the proventriculus. Anatomical features of the cat flea foregut may also restrict

  15. Feeding Behavior Modulates Biofilm-Mediated Transmission of Yersinia pestis by the Cat Flea, Ctenocephalides felis.

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    David M Bland

    2016-02-01

    Full Text Available The cat flea, Ctenocephalides felis, is prevalent worldwide, will parasitize animal reservoirs of plague, and is associated with human habitations in known plague foci. Despite its pervasiveness, limited information is available about the cat flea's competence as a vector for Yersinia pestis. It is generally considered to be a poor vector, based on studies examining early-phase transmission during the first week after infection, but transmission potential by the biofilm-dependent proventricular-blocking mechanism has never been systematically evaluated. In this study, we assessed the vector competence of cat fleas by both mechanisms. Because the feeding behavior of cat fleas differs markedly from important rat flea vectors, we also examined the influence of feeding behavior on transmission dynamics.Groups of cat fleas were infected with Y. pestis and subsequently provided access to sterile blood meals twice-weekly, 5 times per week, or daily for 4 weeks and monitored for infection, the development of proventricular biofilm and blockage, mortality, and the ability to transmit. In cat fleas allowed prolonged, daily access to blood meals, mimicking their natural feeding behavior, Y. pestis did not efficiently colonize the digestive tract and could only be transmitted during the first week after infection. In contrast, cat fleas that were fed intermittently, mimicking the feeding behavior of the efficient vector Xenopsylla cheopis, could become blocked and regularly transmitted Y. pestis for 3-4 weeks by the biofilm-mediated mechanism, but early-phase transmission was not detected.The normal feeding behavior of C. felis, more than an intrinsic resistance to infection or blockage by Y. pestis, limits its vector competence. Rapid turnover of midgut contents results in bacterial clearance and disruption of biofilm accumulation in the proventriculus. Anatomical features of the cat flea foregut may also restrict transmission by both early-phase and

  16. Feeding Behavior Modulates Biofilm-Mediated Transmission of Yersinia pestis by the Cat Flea, Ctenocephalides felis.

    Science.gov (United States)

    Bland, David M; Hinnebusch, B Joseph

    2016-02-01

    The cat flea, Ctenocephalides felis, is prevalent worldwide, will parasitize animal reservoirs of plague, and is associated with human habitations in known plague foci. Despite its pervasiveness, limited information is available about the cat flea's competence as a vector for Yersinia pestis. It is generally considered to be a poor vector, based on studies examining early-phase transmission during the first week after infection, but transmission potential by the biofilm-dependent proventricular-blocking mechanism has never been systematically evaluated. In this study, we assessed the vector competence of cat fleas by both mechanisms. Because the feeding behavior of cat fleas differs markedly from important rat flea vectors, we also examined the influence of feeding behavior on transmission dynamics. Groups of cat fleas were infected with Y. pestis and subsequently provided access to sterile blood meals twice-weekly, 5 times per week, or daily for 4 weeks and monitored for infection, the development of proventricular biofilm and blockage, mortality, and the ability to transmit. In cat fleas allowed prolonged, daily access to blood meals, mimicking their natural feeding behavior, Y. pestis did not efficiently colonize the digestive tract and could only be transmitted during the first week after infection. In contrast, cat fleas that were fed intermittently, mimicking the feeding behavior of the efficient vector Xenopsylla cheopis, could become blocked and regularly transmitted Y. pestis for 3-4 weeks by the biofilm-mediated mechanism, but early-phase transmission was not detected. The normal feeding behavior of C. felis, more than an intrinsic resistance to infection or blockage by Y. pestis, limits its vector competence. Rapid turnover of midgut contents results in bacterial clearance and disruption of biofilm accumulation in the proventriculus. Anatomical features of the cat flea foregut may also restrict transmission by both early-phase and proventricular biofilm

  17. Patterned biofilm formation reveals a mechanism for structural heterogeneity in bacterial biofilms.

    Science.gov (United States)

    Gu, Huan; Hou, Shuyu; Yongyat, Chanokpon; De Tore, Suzanne; Ren, Dacheng

    2013-09-03

    Bacterial biofilms are ubiquitous and are the major cause of chronic infections in humans and persistent biofouling in industry. Despite the significance of bacterial biofilms, the mechanism of biofilm formation and associated drug tolerance is still not fully understood. A major challenge in biofilm research is the intrinsic heterogeneity in the biofilm structure, which leads to temporal and spatial variation in cell density and gene expression. To understand and control such structural heterogeneity, surfaces with patterned functional alkanthiols were used in this study to obtain Escherichia coli cell clusters with systematically varied cluster size and distance between clusters. The results from quantitative imaging analysis revealed an interesting phenomenon in which multicellular connections can be formed between cell clusters depending on the size of interacting clusters and the distance between them. In addition, significant differences in patterned biofilm formation were observed between wild-type E. coli RP437 and some of its isogenic mutants, indicating that certain cellular and genetic factors are involved in interactions among cell clusters. In particular, autoinducer-2-mediated quorum sensing was found to be important. Collectively, these results provide missing information that links cell-to-cell signaling and interaction among cell clusters to the structural organization of bacterial biofilms.

  18. Diarrhea-associated biofilm formed by enteroaggregative Escherichia coli and aggregative Citrobacter freundii: a consortium mediated by putative F pili

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    Araújo Ana CG

    2010-02-01

    Full Text Available Abstract Background Enteroaggregative Escherichia coli (EAEC are enteropathogenic strains identified by the aggregative adhesion (AA pattern that share the capability to form biofilms. Citrobacter freundii is classically considered as an indigenous intestinal species that is sporadically associated with diarrhea. Results During an epidemiologic study focusing on infantile diarrhea, aggregative C. freundii (EACF and EAEC strains were concomitantly recovered from a severe case of mucous diarrhea. Thereby, the occurrence of synergic events involving these strains was investigated. Coinfection of HeLa cells with EACF and EAEC strains showed an 8-fold increase in the overall bacterial adhesion compared with single infections (P traA were capable of forming bacterial aggregates only in the presence of EACF. Scanning electronic microscopy analyses revealed that bacterial aggregates as well as enhanced biofilms formed by EACF and traA-positive EAEC were mediated by non-bundle forming, flexible pili. Moreover, mixed biofilms formed by EACF and traA-positive EAEC strains were significantly reduced using nonlethal concentration of zinc, a specific inhibitor of F pili. In addition, EAEC strains isolated from diarrheic children frequently produced single biofilms sensitive to zinc. Conclusions Putative F pili expressed by EAEC strains boosted mixed biofilm formation when in the presence of aggregative C. freundii.

  19. FpvA receptor involvement in pyoverdine biosynthesis in Pseudomonas aeruginosa.

    Science.gov (United States)

    Shen, Jiangsheng; Meldrum, Allison; Poole, Keith

    2002-06-01

    Alignment of the Pseudomonas aeruginosa ferric pyoverdine receptor, FpvA, with similar ferric-siderophore receptors revealed that the mature protein carries an extension of ca. 70 amino acids at its N terminus, an extension shared by the ferric pseudobactin receptors of P. putida. Deletion of fpvA from the chromosome of P. aeruginosa reduced pyoverdine production in this organism, as a result of a decline in expression of genes (e.g., pvdD) associated with the biosynthesis of the pyoverdine peptide moiety. Wild-type fpvA restored pvd expression in the mutant, thereby complementing its pyoverdine deficiency, although a deletion derivative of fpvA encoding a receptor lacking the N terminus of the mature protein did not. The truncated receptor was, however, functional in pyoverdine-mediated iron uptake, as evidenced by its ability to promote pyoverdine-dependent growth in an iron-restricted medium. These data are consistent with the idea that the N-terminal extension plays a role in FpvA-mediated pyoverdine biosynthesis in P. aeruginosa.

  20. Rechargeable biofilm-controlling tubing materials for use in dental unit water lines.

    Science.gov (United States)

    Luo, Jie; Porteous, Nuala; Sun, Yuyu

    2011-08-01

    A simple and practical surface grafting approach was developed to introduce rechargeable N-halamine-based antimicrobial functionality onto the inner surfaces of continuous small-bore polyurethane (PU) dental unit waterline (DUWL) tubing. In this approach, tetrahydrofuran (THF) solution of a free-radical initiator, dicumyl peroxide (DCP), flowed through the PU tubing (inner diameter of 1/16 in., or 1.6 mm) to diffuse DCP into the tubing's inner walls, which was used as initiator in the subsequent grafting polymerization of methacrylamide (MAA) onto the tubing. Upon chlorine bleach treatment, the amide groups of the grafted MAA side chains were transformed into acyclic N-halamines. The reactions were confirmed with attenuated total reflectance infrared (ATR) spectra and iodometric titration. The mechanical properties of the tubing were not significantly affected by the grafting reactions. The biofilm-controlling function of the new N-halamine-based PU tubing was evaluated with Pseudomonas aeruginosa (P. aeruginosa), one of the most isolated water bacteria from DUWLs, in a continuous bacterial flow model. Bacteria culturing and SEM studies showed that the inner surfaces of the new N-halamine-based PU tubing completely prevented bacterial biofilm formation for at least three to four weeks. After that, bacteria began to colonize the tubing surface. However, the lost function was fully regenerated by exposing the tubing inner surfaces to diluted chlorine bleach. The recharging process could be repeated periodically to further extend the biofilm-controlling duration for long-term applications.

  1. Characterization of the effect of serum and chelating agents on Staphylococcus aureus biofilm formation; chelating agents augment biofilm formation through clumping factor B

    Science.gov (United States)

    Abraham, Nabil Mathew

    Staphylococcus aureus is the causative agent of a diverse array of acute and chronic infections, and some these infections, including infective endocarditis, joint infections, and medical device-associated bloodstream infections, depend upon its capacity to form tenacious biofilms on surfaces. Inserted medical devices such as intravenous catheters, pacemakers, and artificial heart valves save lives, but unfortunately, they can also serve as a substrate on which S. aureus can form a biofilm, attributing S. aureus as a leading cause of medical device-related infections. The major aim of this work was take compounds to which S. aureus would be exposed during infection and to investigate their effects on its capacity to form a biofilm. More specifically, the project investigated the effects of serum, and thereafter of catheter lock solutions on biofilm formation by S. aureus. Pre-coating polystyrene with serum is frequently used as a method to augment biofilm formation. The effect of pre-coating with serum is due to the deposition of extracellular matrix components onto the polystyrene, which are then recognized by MSCRAMMs. We therefore hypothesized that the major component of blood, serum, would induce biofilm formation. Surprisingly, serum actually inhibited biofilm formation. The inhibitory activity was due to a small molecular weight, heat-stable, non-proteinaceous component/s of serum. Serum-mediated inhibition of biofilm formation may represent a previously uncharacterized aspect of host innate immunity that targets the expression of a key bacterial virulence factor: the ability to establish a resistant biofilm. Metal ion chelators like sodium citrate are frequently chosen to lock intravenous catheters because they are regarded as potent inhibitors of bacterial biofilm formation and viability. We found that, while chelating compounds abolished biofilm formation in most strains of S. aureus, they actually augmented the phenotype in a subset of strains. We

  2. Candida albicans mannans mediate Streptococcus mutans exoenzyme GtfB binding to modulate cross-kingdom biofilm development in vivo.

    Science.gov (United States)

    Hwang, Geelsu; Liu, Yuan; Kim, Dongyeop; Li, Yong; Krysan, Damian J; Koo, Hyun

    2017-06-01

    Candida albicans is frequently detected with heavy infection by Streptococcus mutans in plaque-biofilms from children with early-childhood caries (ECC). This cross-kingdom biofilm contains an extensive matrix of extracellular α-glucans that is produced by an exoenzyme (GtfB) secreted by S. mutans. Here, we report that mannans located on the outer surface of C. albicans cell-wall mediates GtfB binding, enhancing glucan-matrix production and modulating bacterial-fungal association within biofilms formed in vivo. Using single-molecule atomic force microscopy, we determined that GtfB binds with remarkable affinity to mannans and to the C. albicans surface, forming a highly stable and strong bond (1-2 nN). However, GtfB binding properties to C. albicans was compromised in strains defective in O-mannan (pmt4ΔΔ) or N-mannan outer chain (och1ΔΔ). In particular, the binding strength of GtfB on och1ΔΔ strain was severely disrupted (>3-fold reduction vs. parental strain). In turn, the GtfB amount on the fungal surface was significantly reduced, and the ability of C. albicans mutant strains to develop mixed-species biofilms with S. mutans was impaired. This phenotype was independent of hyphae or established fungal-biofilm regulators (EFG1, BCR1). Notably, the mechanical stability of the defective biofilms was weakened, resulting in near complete biomass removal by shear forces. In addition, these in vitro findings were confirmed in vivo using a rodent biofilm model. Specifically, we observed that C. albicans och1ΔΔ was unable to form cross-kingdom biofilms on the tooth surface of rats co-infected with S. mutans. Likewise, co-infection with S. mutans defective in GtfB was also incapable of forming mixed-species biofilms. Taken together, the data support a mechanism whereby S. mutans-secreted GtfB binds to the mannan layer of C. albicans to promote extracellular matrix formation and their co-existence within biofilms. Enhanced understanding of GtfB-Candida interactions

  3. A Marine Actinomycete Rescues Caenorhabditis elegans from Pseudomonas aeruginosa Infection through Restitution of Lysozyme 7

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    Siti N. Fatin

    2017-11-01

    Full Text Available The resistance of Pseudomonas aeruginosa to conventional antimicrobial treatment is a major scourge in healthcare. Therefore, it is crucial that novel potent anti-infectives are discovered. The aim of the present study is to screen marine actinomycetes for chemical entities capable of overcoming P. aeruginosa infection through mechanisms involving anti-virulence or host immunity activities. A total of 18 actinomycetes isolates were sampled from marine sediment of Songsong Island, Kedah, Malaysia. Upon confirming that the methanolic crude extract of these isolates do not display direct bactericidal activities, they were tested for capacity to rescue Caenorhabditis elegans infected with P. aeruginosa strain PA14. A hexane partition of the extract from one isolate, designated as Streptomyces sp. CCB-PSK207, could promote the survival of PA14 infected worms by more than 60%. Partial 16S sequence analysis on this isolate showed identity of 99.79% with Streptomyces sundarbansensis. This partition did not impair feeding behavior of C. elegans worms. Tested on PA14, the partition also did not affect bacterial growth or its ability to colonize host gut. The production of biofilm, protease, and pyocyanin in PA14 were uninterrupted, although there was an increase in elastase production. In lys-7::GFP worms, this partition was shown to induce the expression of lysozyme 7, an important innate immunity defense molecule that was repressed during PA14 infection. GC-MS analysis of the bioactive fraction of Streptomyces sp. CCB-PSK207 revealed the presence of methyl esters of branched saturated fatty acids. In conclusion, this is the first report of a marine actinomycete producing metabolites capable of rescuing C. elegans from PA14 through a lys-7 mediated activity.

  4. Facile biofunctionalization of silver nanoparticles for enhanced antibacterial properties, endotoxin removal, and biofilm control

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    Lambadi PR

    2015-03-01

    Full Text Available Paramesh Ramulu Lambadi,1,* Tarun Kumar Sharma,1,* Piyush Kumar,1 Priyanka Vasnani,2 Sitaramanjaneya Mouli Thalluri,2 Neha Bisht,1 Ranjana Pathania,1,2 Naveen Kumar Navani1,21Department of Biotechnology, 2Centre of Nanotechnology, Indian Institute of Technology, Roorkee, Uttarakhand, India*These authors contributed equally to this workAbstract: Infectious diseases cause a huge burden on healthcare systems worldwide. Pathogenic bacteria establish infection by developing antibiotic resistance and modulating the host’s immune system, whereas opportunistic pathogens like Pseudomonas aeruginosa adapt to adverse conditions owing to their ability to form biofilms. In the present study, silver nanoparticles were biofunctionalized with polymyxin B, an antibacterial peptide using a facile method. The biofunctionalized nanoparticles (polymyxin B-capped silver nanoparticles, PBSNPs were assessed for antibacterial activity against multiple drug-resistant clinical strain Vibrio fluvialis and nosocomial pathogen P. aeruginosa. The results of antibacterial assay revealed that PBSNPs had an approximately 3-fold higher effect than the citrate-capped nanoparticles (CSNPs. Morphological damage to the cell membrane was followed by scanning electron microscopy, testifying PBSNPs to be more potent in controlling the bacterial growth as compared with CSNPs. The bactericidal effect of PBSNPs was further confirmed by Live/Dead staining assays. Apart from the antibacterial activity, the biofunctionalized nanoparticles were found to resist biofilm formation. Electroplating of PBSNPs onto stainless steel surgical blades retained the antibacterial activity against P. aeruginosa. Further, the affinity of polymyxin for endotoxin was exploited for its removal using PBSNPs. It was found that the prepared nanoparticles removed 97% of the endotoxin from the solution. Such multifarious uses of metal nanoparticles are an attractive means of enhancing the potency of antimicrobial

  5. Synthetic Peptides to Target Stringent Response-Controlled Virulence in a Pseudomonas aeruginosa Murine Cutaneous Infection Model

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    Daniel Pletzer

    2017-09-01

    Full Text Available Microorganisms continuously monitor their surroundings and adaptively respond to environmental cues. One way to cope with various stress-related situations is through the activation of the stringent stress response pathway. In Pseudomonas aeruginosa this pathway is controlled and coordinated by the activity of the RelA and SpoT enzymes that metabolize the small nucleotide secondary messenger molecule (pppGpp. Intracellular ppGpp concentrations are crucial in mediating adaptive responses and virulence. Targeting this cellular stress response has recently been the focus of an alternative approach to fight antibiotic resistant bacteria. Here, we examined the role of the stringent response in the virulence of P. aeruginosa PAO1 and the Liverpool epidemic strain LESB58. A ΔrelA/ΔspoT double mutant showed decreased cytotoxicity toward human epithelial cells, exhibited reduced hemolytic activity, and caused down-regulation of the expression of the alkaline protease aprA gene in stringent response mutants grown on blood agar plates. Promoter fusions of relA or spoT to a bioluminescence reporter gene revealed that both genes were expressed during the formation of cutaneous abscesses in mice. Intriguingly, virulence was attenuated in vivo by the ΔrelA/ΔspoT double mutant, but not the relA mutant nor the ΔrelA/ΔspoT complemented with either gene. Treatment of a cutaneous P. aeruginosa PAO1 infection with anti-biofilm peptides increased animal welfare, decreased dermonecrotic lesion sizes, and reduced bacterial numbers recovered from abscesses, resembling the phenotype of the ΔrelA/ΔspoT infection. It was previously demonstrated by our lab that ppGpp could be targeted by synthetic peptides; here we demonstrated that spoT promoter activity was suppressed during cutaneous abscess formation by treatment with peptides DJK-5 and 1018, and that a peptide-treated relA complemented stringent response double mutant strain exhibited reduced peptide

  6. Identification of multi-drug resistant Pseudomonas aeruginosa clinical isolates that are highly disruptive to the intestinal epithelial barrier

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    Shevchenko Olga

    2006-06-01

    Full Text Available Abstract Background Multi-drug resistant Pseudomonas aeruginosa nosocomial infections are increasingly recognized worldwide. In this study, we focused on the virulence of multi-drug resistant clinical strains P. aeruginosa against the intestinal epithelial barrier, since P. aeruginosa can cause lethal sepsis from within the intestinal tract of critically ill and immuno-compromised patients via mechanisms involving disruption of epithelial barrier function. Methods We screened consecutively isolated multi-drug resistant P. aeruginosa clinical strains for their ability to disrupt the integrity of human cultured intestinal epithelial cells (Caco-2 and correlated these finding to related virulence phenotypes such as adhesiveness, motility, biofilm formation, and cytotoxicity. Results Results demonstrated that the majority of the multi-drug resistant P. aeruginosa clinical strains were attenuated in their ability to disrupt the barrier function of cultured intestinal epithelial cells. Three distinct genotypes were found that displayed an extreme epithelial barrier-disrupting phenotype. These strains were characterized and found to harbor the exoU gene and to display high swimming motility and adhesiveness. Conclusion These data suggest that detailed phenotypic analysis of the behavior of multi-drug resistant P. aeruginosa against the intestinal epithelium has the potential to identify strains most likely to place patients at risk for lethal gut-derived sepsis. Surveillance of colonizing strains of P. aeruginosa in critically ill patients beyond antibiotic sensitivity is warranted.

  7. Quantum dots conjugated zinc oxide nanosheets: Impeder of microbial growth and biofilm

    Science.gov (United States)

    Patil, Rajendra; Gholap, Haribhau; Warule, Sambhaji; Banpurkar, Arun; Kulkarni, Gauri; Gade, Wasudeo

    2015-01-01

    The grieving problem of the 21st century has been the antimicrobial resistance in pathogenic microorganisms to conventional antibiotics. Therefore, developments of novel antibacterial materials which effectively inhibit or kill such resistant microorganisms have become the need of the hour. In the present study, we communicate the synthesis of quantum dots conjugated zinc oxide nanostructures (ZnO/CdTe) as an impeder of microbial growth and biofilm. The as-synthesized nanostructures were characterized by X-ray diffraction, ultraviolet-visible spectroscopy, photoluminescence spectroscopy, field emission scanning electron microscopy and high resolution transmission electron microscopy. The growth impedance property of ZnO and ZnO/CdTe on Gram positive organism, Bacillus subtilis NCIM 2063 and Gram negative, Escherichia coli NCIM 2931 and biofilm impedance activity in Pseudomonas aeruginosa O1 was found to occur due to photocatalytical action on the cell biofilm surfaces. The impedance in microbial growth and biofilm formation was further supported by ruptured appearances of cells and dettrered biofilm under field emission scanning electron and confocal laser scanning microscope. The ZnO/CdTe nanostructures array synthesized by hydrothermal method has an advantage of low growth temperature, and opportunity to fabricate inexpensive material for nano-biotechnological applications.

  8. Disinfection byproduct formation from chlorination of pure bacterial cells and pipeline biofilms.

    Science.gov (United States)

    Wang, Jun-Jian; Liu, Xin; Ng, Tsz Wai; Xiao, Jie-Wen; Chow, Alex T; Wong, Po Keung

    2013-05-15

    Disinfection byproduct (DBP) formation is commonly attributed to the reaction between natural organic matters and disinfectants, yet few have considered the contribution from disinfecting bacterial materials - the essential process of water disinfection. Here, we explored the DBP formation from chlorination and chloramination of Escherichia coli and found that most selected DBPs were detectable, including trihalomethanes, haloacetonitriles, chloral hydrate, chloropicrin, and 1,1,1-trichloro-2-propanone. A positive correlation (P = 0.08-0.09) between DBP formation and the log reduction of E. coli implied that breaking down of bacterial cells released precursors for DBP formation. As Pseudomonas aeruginosa is a dominant bacterial species in pipeline biofilms, the DBP formation potentials (DBPFPs) from its planktonic cells and biofilms were characterized. Planktonic cells formed 7-11 times greater trihalomethanes per carbon of those from biofilms but significantly lower (P biofilms on polyvinyl chloride compared to that on galvanized zinc. This study revealed both the in situ disinfection of bacterial planktonic cells in source water and ex situ reaction between biofilms and residual chlorine in pipeline networks as hitherto unknown DBP sources in drinking water. Copyright © 2013 Elsevier Ltd. All rights reserved.

  9. Anti-biofilm efficacy of low temperature processed AgCl–TiO2 nanocomposite coating

    International Nuclear Information System (INIS)

    Naik, Kshipra; Kowshik, Meenal

    2014-01-01

    Biofilms are a major concern in the medical settings and food industries due to their high tolerance to antibiotics, biocides and mechanical stress. Currently, the development of novel methods to control biofilm formation is being actively pursued. In the present study, sol–gel coatings of AgCl–TiO 2 nanoparticles are presented as potential anti-biofilm agents, wherein TiO 2 acts as a good supporting matrix to prevent aggregation of silver and facilitates its controlled release. Low-temperature processed AgCl–TiO 2 nanocomposite coatings inhibit biofilm formation by Escherichia coli, Staphylococcus epidermidis and Pseudomonas aeruginosa. In vitro biofilm assay experiments demonstrated that AgCl–TiO 2 nanocomposite coated surfaces, inhibited the development of biofilms over a period of 10 days as confirmed by scanning electron microscopy. The silver release kinetics exhibited an initial high release, followed by a slow and sustained release. The anti-biofilm efficacy of the coatings could be attributed to the release of silver, which prevents the initial bacterial adhesion required for biofilm formation. - Highlights: • Potential of AgCl–TiO 2 nanocomposite coating to inhibit biofilm formation is exhibited. • Initial rapid release followed by later slow and sustained release of silver obtained. • TiO 2 being porous and inorganic in nature acts as a good supporting matrix

  10. In vitro and in vivo generation and characterization of Pseudomonas aeruginosa biofilm-dispersed cells via c-di-GMP manipulation

    DEFF Research Database (Denmark)

    Chua, Song Lin; Hultqvist, Louise D; Yuan, Mingjun

    2015-01-01

    Bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP) is a global secondary bacterial messenger that controls the formation of drug-resistant multicellular biofilms. Lowering the intracellular c-di-GMP content can disperse biofilms, and it is proposed as a biofilm eradication strategy...... biofilms by reducing the intracellular c-di-GMP content through modulation of phosphodiesterases (PDEs). Unlike conventional protocols that demonstrate biofilm dispersal by biomass quantification, our protocols enable physiological characterization of the dispersed cells. Biomarkers of dispersed cells...

  11. Caratterizzazione di isolati di Pseudomonas aeruginosa provenienti da pazienti affetti da fibrosi cistica

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    Giovanna Pulcrano

    2007-06-01

    Full Text Available Pseudomonas aeruginosa is the major pulmonary pathogen that causes morbidity and mortality in burned, immunocompromised and cystic fibrosis patients. Among the various virulence factors, type IV-pili, play a major role in mediating bacteria-host cells interactions, in formation of biofilm and for twiching motility.These pili are composed of pilin, 15000-16000 molecular weight monomeric subunit, synthesized from pilA gene. The N-terminal region of pilin protein is strong conserved and is important for the oligomerization.The C-terminal region is less conserved and contains a disulfide-bonded loop (DSL structure that is thought to interact with the eukaryotic glycolipid receptor “asialo GM1”. Analysis of pilin allele distribution among isolates from various sources revealed the presence of six groups of pilin allels characterized by different DSL sequence and different associated accessory genes in pilA chromosomal locus. 81 P. aeruginosa isolates were recovered from cystic fibrosis patients during a 3 years period. 30 of these strains were grown and their genomic DNA was prepared using a rapid method for gram-negative bacteria. PCR primers were used for amplification of pilA and adjacent sequences revealing the presence of three different amplification products. One of these is highly homologous with pilA gene of PA14 strain, the others are identical to PA103 and PAK pilA genes. Our study revealed in the prevalence of isolates with group II pilin genes from Cystic Fibrosis compared with other groups that are predominant in previous studies.

  12. Impact of Pseudomonas aeruginosa quorum sensing on biofilm persistence in an in vivo intraperitoneal foreign-body infection model

    DEFF Research Database (Denmark)

    Christensen, Louise Dahl; Moser, Claus; Jensen, Peter Østrup

    2007-01-01

    . aeruginosa are cleared more rapidly from silicone implants as compared to their wild-type counterparts. Concurrently, treatment with the QS inhibitor furanone C-30 of mice harbouring implants colonized with the wild-type P. aeruginosa resulted in a significantly faster clearing of the implants as compared...

  13. Clinical implications of microbial biofilms in chronic rhinosinusitis and orbital cellulitis.

    Science.gov (United States)

    Nayak, Niranjan; Satpathy, Gita; Prasad, Sujata; Thakar, Alok; Chandra, Mahesh; Nag, T C

    2016-09-21

    Discovery of sessile mode of microbial existence (Biofilm state) focussed much interest, during the recent years, on the study of biofilms in many recurring and chronic infections. However, the exact role of microbial biofilms in chronic rhinosinusitis and orbital cellulitis were not elucidated earlier. The purpose of the present study was to look for the adherent property and biofilm producing ability of the clinical isolates in chronic rhinosinusitis and orbital cellulitis, and to look for the effects of antimicrobial agents on these biofilms by colorimetric assay and ultrastructural analysis. Organisms were isolated and identified from various clinical samples in patients with chronic sinusitis and orbital cellulitis. Antimicrobial sensitivity testing was carried out by the standard protocol. Biofilms were developed; quantified and antimicrobial drug perfusion through the biofilm model was evaluated by the earlier devised procedure. Electronmicroscopic study of the biofilm was performed by the recommended technique. Of the total of 70 clinical samples processed, 48 i.e. 68.5 % grew bacteria and 13 i.e.(18.6 %) fungi. Staphylococcus aureus (20), S epidermidis (16) and Pseudomonas aeruginosa (6) accounted for the majority of the bacterial isolates. Aspergillus flavus (8), however was the commonest amongst the fungi. A total of 40 bacteria and 8 fungi could be tested for biofilm production. Eighteen (45 %) of the 40 bacterial isolates and 4(50 %) out of the 8 A flavus isolates were found to be biofilm producers. In vitro adherence testing revealed that majority i.e. 16 (88.8 %) of the 18 biofilm positive bacteria were adherent to artificial surfaces. Antimicrobial drug perfusion through the biofilm model was poor. Antimicrobial treatment was totally ineffective against strong biofilm producers, whose electron microscopic picture was quite similar to that observed for biofilm producers without any antimicrobial pre-treatment. Filamentous fungi, like bacteria

  14. Chemical Inhibition of Kynureninase Reduces Pseudomonas aeruginosa Quorum Sensing and Virulence Factor Expression.

    Science.gov (United States)

    Kasper, Stephen H; Bonocora, Richard P; Wade, Joseph T; Musah, Rabi Ann; Cady, Nathaniel C

    2016-04-15

    The opportunistic pathogen Pseudomonas aeruginosa utilizes multiple quorum sensing (QS) pathways to coordinate an arsenal of virulence factors. We previously identified several cysteine-based compounds inspired by natural products from the plant Petiveria alliacea which are capable of antagonizing multiple QS circuits as well as reducing P. aeruginosa biofilm formation. To understand the global effects of such compounds on virulence factor production and elucidate their mechanism of action, RNA-seq transcriptomic analysis was performed on P. aeruginosa PAO1 exposed to S-phenyl-l-cysteine sulfoxide, the most potent inhibitor from the prior study. Exposure to this inhibitor down-regulated expression of several QS-regulated virulence operons (e.g., phenazine biosynthesis, type VI secretion systems). Interestingly, many genes that were differentially regulated pertain to the related metabolic pathways that yield precursors of pyochelin, tricarboxylic acid cycle intermediates, phenazines, and Pseudomonas quinolone signal (PQS). Activation of the MexT-regulon was also indicated, including the multidrug efflux pump encoded by mexEF-oprN, which has previously been shown to inhibit QS and pathogenicity. Deeper investigation of the metabolites involved in these systems revealed that S-phenyl-l-cysteine sulfoxide has structural similarity to kynurenine, a precursor of anthranilate, which is critical for P. aeruginosa virulence. By supplementing exogenous anthranilate, the QS-inhibitory effect was reversed. Finally, it was shown that S-phenyl-l-cysteine sulfoxide competitively inhibits P. aeruginosa kynureninase (KynU) activity in vitro and reduces PQS production in vivo. The kynurenine pathway has been implicated in P. aeruginosa QS and virulence factor expression; however, this is the first study to show that targeted inhibition of KynU affects P. aeruginosa gene expression and QS, suggesting a potential antivirulence strategy.

  15. Chitosanase purified from bacterial isolate Bacillus licheniformis of ruined vegetables displays broad spectrum biofilm inhibition.

    Science.gov (United States)

    Muslim, Sahira Nsayef; Al-Kadmy, Israa M S; Hussein, Nadheema Hammood; Mohammed Ali, Alaa Naseer; Taha, Buthainah Mohammed; Aziz, Sarah Naji; Kheraif, Abdulaziz Abdullah Al; Divakar, Darshan Devang; Ramakrishnaiah, Ravikumar

    2016-11-01

    A number of bacterial species produces chitosanases which has variety of applications because of its high biodegradability, non-toxicity and antimicrobial assets. In the present study chitosanase is purified from new bacterial species Bacillus licheniformis from spoiled vegetable. This novel strain of Bacillus licheniformis isolated from spoilt cucumber and pepper samples has the ability to produce the chitosanase enzyme when grown on chitosan substrate. Study also examined its antibiofilm properties against diverse bacterial species with biofilm forming ability. The purified chitosanase inhibited the biofilm formation ability for all Gram-negative and Gram-positive biofilm-forming bacteria [biofilm producers] tested in this study in congo red agar and microtiter plate's methods. Highly antibiofilm activity of chitosanase was recorded against Pseudomonas aeruginosa followed by Klebsiella pneumoniae with reduction of biofilm formation upto 22 and 29%, respectively compared with [100] % of control. Biofilm formation has multiple role including ability to enhance resistance and self-protection from external stress. This chitosanase has promising benefit as antibiofilm agent against biofilm forming pathogenic bacteria and has promising application as alternative antibiofilm agents to combat the growing number of multidrug resistant pathogen-associated infections, especially in situation where biofilms are involved. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. P. aeruginosa in the paranasal sinuses and transplanted lungs have similar adaptive mutations as isolates from chronically infected CF lungs

    DEFF Research Database (Denmark)

    Ciofu, Oana; Johansen, Helle Krogh; Aanaes, Kasper

    2013-01-01

    BACKGROUND: Pseudomonas aeruginosa cells are present as biofilms in the paranasal sinuses and the lungs of chronically infected cystic fibrosis (CF) patients. Since different inflammatory responses and selective antibiotic pressures are acting in the sinuses compared with the lungs, we compared...... the adaptive profiles of mucoid and non-mucoid isolates from the two locations. METHODS: We studied the genetic basis of phenotypic diversification and gene expression profiles in sequential lung and sinus P. aeruginosa isolates from four chronically infected CF patients, including pre- and post-lung...... transplantation isolates. RESULTS: The same phenotypes caused by similar mutations and similar gene expression profiles were found in mucoid and non-mucoid isolates from the paranasal sinuses and from the lungs before and after transplantation. CONCLUSION: Bilateral exchange of P. aeruginosa isolates between...

  17. UVC fluencies for preventative treatment of Pseudomonas aeruginosa contaminated polymer tubes

    DEFF Research Database (Denmark)

    Bak, Jimmy; Ladefoged, Søren D; Begovic, Tanja

    2010-01-01

    Exposing Pseudomonas aeruginosa biofilm grown on the inner surface of Teflon and silicone tubes to UVC light (265 nm) from light emitting diodes (LED) has previously been shown to substantially reduce biofilm growth. Smaller UVC fluencies were required to disinfect Teflon tubes compared to silicone...... tubes. Light propagation enhancement in tubes can be obtained if the refractive index of the intra-luminal saline solution is higher than that of the polymer. This condition is achieved by using Teflon tubes with a low refractive index (1.34) instead of the polymers with a high refractive index (1...... is demonstrated to be a preventative disinfection treatment on tubes made of Teflon, which enhances the UVC light propagation, and on tubes made of a softer material, ethylene vinyl acetate (EVA), which is suitable for catheters but much less suitable for UVC light propagation. Simulating an aseptic breach (~10...

  18. Anti-biofilm activity of chitosan gels formulated with silver nanoparticles and their cytotoxic effect on human fibroblasts

    International Nuclear Information System (INIS)

    Pérez-Díaz, M.; Alvarado-Gomez, E.; Magaña-Aquino, M.; Sánchez-Sánchez, R.; Velasquillo, C.; Gonzalez, C.; Ganem-Rondero, A.; Martínez-Castañon, G.; Zavala-Alonso, N.; Martinez-Gutierrez, F.

    2016-01-01

    The development of multi-species biofilms in chronic wounds is a serious health problem that primarily generates strong resistance mechanisms to antimicrobial therapy. The use of silver nanoparticles (AgNPs) as a broad-spectrum antimicrobial agent has been studied previously. However, their cytotoxic effects limit its use within the medical area. The purpose of this study was to evaluate the anti-biofilm capacity of chitosan gel formulations loaded with AgNPs, using silver sulfadiazine (SSD) as a standard treatment, on strains of clinical isolates, as well as their cytotoxic effect on human primary fibroblasts. Multi-species biofilm of Staphylococcus aureus oxacillin resistant (MRSA) and Pseudomonas aeruginosa obtained from a patient with chronic wound infection were carried out using a standard Drip Flow Reactor (DFR) under conditions that mimic the flow of nutrients in the human skin. Anti-biofilm activity of chitosan gels and SSD showed a log-reduction of 6.0 for MRSA when chitosan gel with AgNPs at a concentration of 100 ppm was used, however it was necessary to increase the concentration of the chitosan gel with AgNPs to 1000 ppm to get a log-reduction of 3.3, while the SSD showed a total reduction of both bacteria in comparison with the negative control. The biocompatibility evaluation on primary fibroblasts showed better results when the chitosan gels with AgNPs were tested even in the high concentration, in contrast with SSD, which killed all the primary fibroblasts. In conclusion, chitosan gel formulations loaded with AgNPs effectively prevent the formation of biofilm and kill bacteria in established biofilm, which suggest that chitosan gels with AgNPs could be used for prevention and treatment of infections in chronic wounds. The statistic significance of the biocompatibility of chitosan gel formulations loaded with AgNPs represents an advance; however further research and development are necessary to translate this technology into therapeutic and

  19. Anti-biofilm activity of chitosan gels formulated with silver nanoparticles and their cytotoxic effect on human fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Pérez-Díaz, M.; Alvarado-Gomez, E. [Facultad de Ciencias Quimicas, Universidad Autonoma de San Luis Potosi (Mexico); Magaña-Aquino, M. [Servicio de Epidemiologia del Hospital Central “Dr. Ignacio Morones Prieto”, San Luis Potosi (Mexico); Sánchez-Sánchez, R.; Velasquillo, C. [Laboratorio de Biotecnologia, Instituto Nacional de Rehabilitacion, Mexico, D.F. (Mexico); Gonzalez, C. [Facultad de Ciencias Quimicas, Universidad Autonoma de San Luis Potosi (Mexico); Ganem-Rondero, A. [Division de Estudios de Posgrado (Tecnologia Farmaceutica), Facultad de Estudios Superiores Cuautitlan, Universidad Nacional Autonoma de Mexico, Cuautitlan Izcalli, Estado de Mexico (Mexico); Martínez-Castañon, G.; Zavala-Alonso, N. [Doctorado en Ciencias Odontológicas Facultad de Estomatologia, UASLP (Mexico); Martinez-Gutierrez, F. [Facultad de Ciencias Quimicas, Universidad Autonoma de San Luis Potosi (Mexico)

    2016-03-01

    The development of multi-species biofilms in chronic wounds is a serious health problem that primarily generates strong resistance mechanisms to antimicrobial therapy. The use of silver nanoparticles (AgNPs) as a broad-spectrum antimicrobial agent has been studied previously. However, their cytotoxic effects limit its use within the medical area. The purpose of this study was to evaluate the anti-biofilm capacity of chitosan gel formulations loaded with AgNPs, using silver sulfadiazine (SSD) as a standard treatment, on strains of clinical isolates, as well as their cytotoxic effect on human primary fibroblasts. Multi-species biofilm of Staphylococcus aureus oxacillin resistant (MRSA) and Pseudomonas aeruginosa obtained from a patient with chronic wound infection were carried out using a standard Drip Flow Reactor (DFR) under conditions that mimic the flow of nutrients in the human skin. Anti-biofilm activity of chitosan gels and SSD showed a log-reduction of 6.0 for MRSA when chitosan gel with AgNPs at a concentration of 100 ppm was used, however it was necessary to increase the concentration of the chitosan gel with AgNPs to 1000 ppm to get a log-reduction of 3.3, while the SSD showed a total reduction of both bacteria in comparison with the negative control. The biocompatibility evaluation on primary fibroblasts showed better results when the chitosan gels with AgNPs were tested even in the high concentration, in contrast with SSD, which killed all the primary fibroblasts. In conclusion, chitosan gel formulations loaded with AgNPs effectively prevent the formation of biofilm and kill bacteria in established biofilm, which suggest that chitosan gels with AgNPs could be used for prevention and treatment of infections in chronic wounds. The statistic significance of the biocompatibility of chitosan gel formulations loaded with AgNPs represents an advance; however further research and development are necessary to translate this technology into therapeutic and

  20. Anti-biofilm efficacy of low temperature processed AgCl–TiO{sub 2} nanocomposite coating

    Energy Technology Data Exchange (ETDEWEB)

    Naik, Kshipra, E-mail: kshipra_naik21@yahoo.co.in; Kowshik, Meenal, E-mail: meenal@goa.bits-pilani.ac.in

    2014-01-01

    Biofilms are a major concern in the medical settings and food industries due to their high tolerance to antibiotics, biocides and mechanical stress. Currently, the development of novel methods to control biofilm formation is being actively pursued. In the present study, sol–gel coatings of AgCl–TiO{sub 2} nanoparticles are presented as potential anti-biofilm agents, wherein TiO{sub 2} acts as a good supporting matrix to prevent aggregation of silver and facilitates its controlled release. Low-temperature processed AgCl–TiO{sub 2} nanocomposite coatings inhibit biofilm formation by Escherichia coli, Staphylococcus epidermidis and Pseudomonas aeruginosa. In vitro biofilm assay experiments demonstrated that AgCl–TiO{sub 2} nanocomposite coated surfaces, inhibited the development of biofilms over a period of 10 days as confirmed by scanning electron microscopy. The silver release kinetics exhibited an initial high release, followed by a slow and sustained release. The anti-biofilm efficacy of the coatings could be attributed to the release of silver, which prevents the initial bacterial adhesion required for biofilm formation. - Highlights: • Potential of AgCl–TiO{sub 2} nanocomposite coating to inhibit biofilm formation is exhibited. • Initial rapid release followed by later slow and sustained release of silver obtained. • TiO{sub 2} being porous and inorganic in nature acts as a good supporting matrix.

  1. Biofilms and type III secretion are not mutually exclusive in Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Mikkelsen, H; Bond, N J; Skindersoe, M E

    2009-01-01

    in exponential phase than to those in stationary phase. In the current study, we investigated how these conditions influence the production of virulence factors using a transcriptomic approach. Our results show that biofilms express the type III secretion system, whereas planktonic cells do not...

  2. Dynamics and spatial distribution of beta-lactamase expression in Pseudomonas aeruginosa biofilms

    DEFF Research Database (Denmark)

    Bagge, N.; Hentzer, Morten; Andersen, Jens Bo

    2004-01-01

    of increased imipenem concentrations. Ceftazidime induced the monitor system of the biofilm bacteria as well, but only bacteria in the peripheries of the microcolonies were induced in the presence of even very high concentrations. The experiments illustrate for the first time the dynamic and spatial...

  3. Curcumin Quantum Dots Mediated Degradation of Bacterial Biofilms

    Directory of Open Access Journals (Sweden)

    Ashish K. Singh

    2017-08-01

    Full Text Available Bacterial biofilm has been reported to be associated with more than 80% of bacterial infections. Curcumin, a hydrophobic polyphenol compound, has anti-quorum sensing activity apart from having antimicrobial action. However, its use is limited by its poor aqueous solubility and rapid degradation. In this study, we attempted to prepare quantum dots of the drug curcumin in order to achieve enhanced solubility and stability and investigated for its antimicrobial and antibiofilm activity. We utilized a newer two-step bottom up wet milling approach to prepare Curcumin Quantum Dots (CurQDs using acetone as a primary solvent. Minimum inhibitory concentration against select Gram-positive and Gram-negative bacteria was performed. The antibiofilm assay was performed at first using 96-well tissue culture plate and subsequently validated by Confocal Laser Scanning Microscopy. Further, biofilm matrix protein was isolated using formaldehyde sludge and TCA/Acetone precipitation method. Protein extracted was incubated with varying concentration of CurQDs for 4 h and was subjected to SDS–PAGE. Molecular docking study was performed to observe interaction between curcumin and phenol soluble modulins as well as curli proteins. The biophysical evidences obtained from TEM, SEM, UV-VIS, fluorescence, Raman spectroscopy, and zeta potential analysis confirmed the formation of curcumin quantum dots with increased stability and solubility. The MICs of curcumin quantum dots, as observed against both select gram positive and negative bacterial isolates, was observed to be significantly lower than native curcumin particles. On TCP assay, Curcumin observed to be having antibiofilm as well as biofilm degrading activity. Results of SDS–PAGE and molecular docking have shown interaction between biofilm matrix proteins and curcumin. The results indicate that aqueous solubility and stability of Curcumin can be achieved by preparing its quantum dots. The study also demonstrates

  4. Application of bacteriophages to selectively remove Pseudomonas aeruginosa in water and wastewater filtration systems.

    Science.gov (United States)

    Zhang, Yanyan; Hunt, Heather K; Hu, Zhiqiang

    2013-09-01

    Water and wastewater filtration systems often house pathogenic bacteria, which must be removed to ensure clean, safe water. Here, we determine the persistence of the model bacterium Pseudomonas aeruginosa in two types of filtration systems, and use P. aeruginosa bacteriophages to determine their ability to selectively remove P. aeruginosa. These systems used beds of either anthracite or granular activated carbon (GAC), which were operated at an empty bed contact time (EBCT) of 45 min. The clean bed filtration systems were loaded with an instantaneous dose of P. aeruginosa at a total cell number of 2.3 (± 0.1 [standard deviation]) × 10(7) cells. An immediate dose of P. aeruginosa phages (1 mL of phage stock at the concentration of 2.7 × 10(7) PFU (Plaque Forming Units)/mL) resulted in a reduction of 50% (± 9%) and >99.9% in the effluent P. aeruginosa concentrations in the clean anthracite and GAC filters, respectively. To further evaluate the effects of P. aeruginosa phages, synthetic stormwater was run through anthracite and GAC biofilters where mixed-culture biofilms were present. Eighty five days after an instantaneous dose of P. aeruginosa (2.3 × 10(7) cells per filter) on day 1, 7.5 (± 2.8) × 10(7) and 1.1 (± 0.5) × 10(7) P. aeruginosa cells/g filter media were detected in the top layer (close to the influent port) of the anthracite and GAC biofilters, respectively, demonstrating the growth and persistence of pathogenic bacteria in the biofilters. A subsequent 1-h dose of phages, at the concentration of 5.1 × 10(6) PFU/mL and flow rate of 1.6 mL/min, removed the P. aeruginosa inside the GAC biofilters and the anthracite biofilters by 70% (± 5%) and 56% (± 1%), respectively, with no P. aeruginosa detected in the effluent, while not affecting ammonia oxidation or the ammonia-oxidizing bacterial community inside the biofilters. These results suggest that phage treatment can selectively remove pathogenic bacteria with minimal impact on beneficial

  5. Positive signature-tagged mutagenesis in Pseudomonas aeruginosa: tracking patho-adaptive mutations promoting airways chronic infection.

    Directory of Open Access Journals (Sweden)

    Irene Bianconi

    2011-02-01

    Full Text Available The opportunistic pathogen Pseudomonas aeruginosa can establish life-long chronic infections in the airways of cystic fibrosis (CF patients. Persistent lifestyle is established with P. aeruginosa patho-adaptive variants, which are clonal with the initially-acquired strains. Several reports indicated that P. aeruginosa adapts by loss-of-function mutations which enhance fitness in CF airways and sustain its clonal expansion during chronic infection. To validate this model of P. aeruginosa adaptation to CF airways and to identify novel genes involved in this microevolution, we designed a novel approach of positive-selection screening by PCR-based signature-tagged mutagenesis (Pos-STM in a murine model of chronic airways infection. A systematic positive-selection scheme using sequential rounds of in vivo screenings for bacterial maintenance, as opposed to elimination, generated a list of genes whose inactivation increased the colonization and persistence in chronic airways infection. The phenotypes associated to these Pos-STM mutations reflect alterations in diverse aspects of P. aeruginosa biology which include lack of swimming and twitching motility, lack of production of the virulence factors such as pyocyanin, biofilm formation, and metabolic functions. In addition, Pos-STM mutants showed altered invasion and stimulation of immune response when tested in human respiratory epithelial cells, indicating that P. aeruginosa is prone to revise the interaction with its host during persistent lifestyle. Finally, sequence analysis of Pos-STM genes in longitudinally P. aeruginosa isolates from CF patients identified signs of patho-adaptive mutations within the genome. This novel Pos-STM approach identified bacterial functions that can have important clinical implications for the persistent lifestyle and disease progression of the airway chronic infection.

  6. Effect of bacterial components of mixed culture supernatants of planktonic and biofilm Pseudomonas aeruginosa with commensal Escherichia coli on the neutrophil response in vitro.

    Science.gov (United States)

    Maslennikova, Irina L; Kuznetsova, Marina V; Nekrasova, Irina V; Shirshev, Sergei V

    2017-11-30

    Pseudomonas aeruginosa (PA) responsible for acute and chronic infections often forms a well-organized bacterial population with different microbial species including commensal strains of Escherichia coli. Bacterial extracellular components of mixed culture can modulate the influence of bacteria on the neutrophil functions. The objective of this study was to compare the effect of pyocyanin, pyoverdine, LPS, exopolysaccharide of single species and mixed culture supernatants of PA strains and E. coli K12 on microbicidal, secretory activity of human neutrophils in vitro. Bacterial components of E. coli K12 in mixed supernatants with 'biofilm' PA strains (PA ATCC, PA BALG) enhanced short-term microbicidal mechanisms and inhibited neutrophil secretion delayed in time. The influence of 'planktonic' PA (PA 9-3) exometabolites in mixed culture is almost mimicked by E. coli K12 effect on functional neutrophil changes. This investigation may help to understand some of the mechanisms of neutrophil response to mixed infections of different PA with other bacteria species. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  7. Extracellular-matrix-mediated osmotic pressure drives Vibrio cholerae biofilm expansion and cheater exclusion.

    Science.gov (United States)

    Yan, Jing; Nadell, Carey D; Stone, Howard A; Wingreen, Ned S; Bassler, Bonnie L

    2017-08-23

    Biofilms, surface-attached communities of bacteria encased in an extracellular matrix, are a major mode of bacterial life. How the material properties of the matrix contribute to biofilm growth and robustness is largely unexplored, in particular in response to environmental perturbations such as changes in osmotic pressure. Here, using Vibrio cholerae as our model organism, we show that during active cell growth, matrix production enables biofilm-dwelling bacterial cells to establish an osmotic pressure difference between the biofilm and the external environment. This pressure difference promotes biofilm expansion on nutritious surfaces by physically swelling the colony, which enhances nutrient uptake, and enables matrix-producing cells to outcompete non-matrix-producing cheaters via physical exclusion. Osmotic pressure together with crosslinking of the matrix also controls the growth of submerged biofilms and their susceptibility to invasion by planktonic cells. As the basic physicochemical principles of matrix crosslinking and osmotic swelling are universal, our findings may have implications for other biofilm-forming bacterial species.Most bacteria live in biofilms, surface-attached communities encased in an extracellular matrix. Here, Yan et al. show that matrix production in Vibrio cholerae increases the osmotic pressure within the biofilm, promoting biofilm expansion and physical exclusion of non-matrix producing cheaters.

  8. Biofilm in endodontics: A review

    Science.gov (United States)

    Jhajharia, Kapil; Parolia, Abhishek; Shetty, K Vikram; Mehta, Lata Kiran

    2015-01-01

    Endodontic disease is a biofilm-mediated infection, and primary aim in the management of endodontic disease is the elimination of bacterial biofilm from the root canal system. The most common endodontic infection is caused by the surface-associated growth of microorganisms. It is important to apply the biofilm concept to endodontic microbiology to understand the pathogenic potential of the root canal microbiota as well as to form the basis for new approaches for disinfection. It is foremost to understand how the biofilm formed by root canal bacteria resists endodontic treatment measures. Bacterial etiology has been confirmed for common oral diseases such as caries and periodontal and endodontic infections. Bacteria causing these diseases are organized in biofilm structures, which are complex microbial communities composed of a great variety of bacteria with different ecological requirements and pathogenic potential. The biofilm community not only gives bacteria effective protection against the host's defense system but also makes them more resistant to a variety of disinfecting agents used as oral hygiene products or in the treatment of infections. Successful treatment of these diseases depends on biofilm removal as well as effective killing of biofilm bacteria. So, the fundamental to maintain oral health and prevent dental caries, gingivitis, and periodontitis is to control the oral biofilms. From these aspects, the formation of biofilms carries particular clinical significance because not only host defense mechanisms but also therapeutic efforts including chemical and mechanical antimicrobial treatment measures have the most difficult task of dealing with organisms that are gathered in a biofilm. The aim of this article was to review the mechanisms of biofilms’ formation, their roles in pulpal and periapical pathosis, the different types of biofilms, the factors influencing biofilm formation, the mechanisms of their antimicrobial resistance, techniques to

  9. Biofilm is a Major Virulence Determinant in Bacterial Colonization of Chronic Skin Ulcers Independently from the Multidrug Resistant Phenotype

    Directory of Open Access Journals (Sweden)

    Enea Gino Di Domenico

    2017-05-01

    Full Text Available Bacterial biofilm is a major factor in delayed wound healing and high levels of biofilm production have been repeatedly described in multidrug resistant organisms (MDROs. Nevertheless, a quantitative correlation between biofilm production and the profile of antimicrobial drug resistance in delayed wound healing remains to be determined. Microbial identification, antibiotic susceptibility and biofilm production were assessed in 135 clinical isolates from 87 patients. Gram-negative bacteria were the most represented microorganisms (60.8% with MDROs accounting for 31.8% of the total isolates. Assessment of biofilm production revealed that 80% of the strains were able to form biofilm. A comparable level of biofilm production was found with both MDRO and not-MDRO with no significant differences between groups. All the methicillin-resistant Staphylococcus aureus (MRSA and 80% of Pseudomonas aeruginosa MDR strains were found as moderate/high biofilm producers. Conversely, less than 17% of Klebsiella pneumoniae extended-spectrum beta-lactamase (ESBL, Escherichia coli-ESBL and Acinetobacter baumannii were moderate/high biofilm producers. Notably, those strains classified as non-biofilm producers, were always associated with biofilm producer bacteria in polymicrobial colonization. This study shows that biofilm producers were present in all chronic skin ulcers, suggesting that biofilm represents a key virulence determinant in promoting bacterial persistence and chronicity of ulcerative lesions independently from the MDRO phenotype.

  10. Biofilms in Endodontics—Current Status and Future Directions

    Science.gov (United States)

    Neelakantan, Prasanna; Romero, Monica; Vera, Jorge; Daood, Umer; Khan, Asad U.; Yan, Aixin; Cheung, Gary Shun Pan

    2017-01-01

    Microbiota are found in highly organized and complex entities, known as biofilms, the characteristics of which are fundamentally different from microbes in planktonic suspensions. Root canal infections are biofilm mediated. The complexity and variability of the root canal system, together with the multi-species nature of biofilms, make disinfection of this system extremely challenging. Microbial persistence appears to be the most important factor for failure of root canal treatment and this could further have an impact on pain and quality of life. Biofilm removal is accomplished by a chemo-mechanical process, using specific instruments and disinfecting chemicals in the form of irrigants and/or intracanal medicaments. Endodontic research has focused on the characterization of root canal biofilms and the clinical methods to disrupt the biofilms in addition to achieving microbial killing. In this narrative review, we discuss the role of microbial biofilms in endodontics and review the literature on the role of root canal disinfectants and disinfectant-activating methods on biofilm removal. PMID:28800075

  11. Biofilms in Endodontics-Current Status and Future Directions.

    Science.gov (United States)

    Neelakantan, Prasanna; Romero, Monica; Vera, Jorge; Daood, Umer; Khan, Asad U; Yan, Aixin; Cheung, Gary Shun Pan

    2017-08-11

    Microbiota are found in highly organized and complex entities, known as biofilms, the characteristics of which are fundamentally different from microbes in planktonic suspensions. Root canal infections are biofilm mediated. The complexity and variability of the root canal system, together with the multi-species nature of biofilms, make disinfection of this system extremely challenging. Microbial persistence appears to be the most important factor for failure of root canal treatment and this could further have an impact on pain and quality of life. Biofilm removal is accomplished by a chemo-mechanical process, using specific instruments and disinfecting chemicals in the form of irrigants and/or intracanal medicaments. Endodontic research has focused on the characterization of root canal biofilms and the clinical methods to disrupt the biofilms in addition to achieving microbial killing. In this narrative review, we discuss the role of microbial biofilms in endodontics and review the literature on the role of root canal disinfectants and disinfectant-activating methods on biofilm removal.

  12. Pseudomonas aeruginosa PA14 pathogenesis in Caenorhabditis elegans.

    Science.gov (United States)

    Kirienko, Natalia V; Cezairliyan, Brent O; Ausubel, Frederick M; Powell, Jennifer R

    2014-01-01

    The nematode Caenorhabditis elegans is a simple model host for studying the interaction between bacterial pathogens such as Pseudomonas aeruginosa and the metazoan innate immune system. Powerful genetic and molecular tools in both C. elegans and P. aeruginosa facilitate the identification and analysis of bacterial virulence factors as well as host defense factors. Here we describe three different assays that use the C. elegans-P. aeruginosa strain PA14 host-pathogen system. Fast Killing is a toxin-mediated death that depends on a diffusible toxin produced by PA14 but not on live bacteria. Slow Killing is due to an active infection in which bacteria colonize the C. elegans intestinal lumen. Liquid Killing is designed for high-throughput screening of chemical libraries for anti-infective compounds. Each assay has unique features and, interestingly, the PA14 virulence factors involved in killing are different in each assay.

  13. Regulation of Nicotine Tolerance by Quorum Sensing and High Efficiency of Quorum Quenching Under Nicotine Stress in Pseudomonas aeruginosa PAO1

    Directory of Open Access Journals (Sweden)

    Huiming Tang

    2018-03-01

    Full Text Available Quorum sensing (QS regulates the behavior of bacterial populations and promotes their adaptation and survival under stress. As QS is responsible for the virulence of vast majority of bacteria, quorum quenching (QQ, the interruption of QS, has become an attractive therapeutic strategy. However, the role of QS in stress tolerance and the efficiency of QQ under stress in bacteria are seldom explored. In this study, we demonstrated that QS-regulated catalase (CAT expression and biofilm formation help Pseudomonas aeruginosa PAO1 resist nicotine stress. CAT activity and biofilm formation in wild type (WT and ΔrhlR strains are significantly higher than those in the ΔlasR strain. Supplementation of ΔlasI strain with 3OC12-HSL showed similar CAT activity and biofilm formation as those of the WT strain. LasIR circuit rather than RhlIR circuit is vital to nicotine tolerance. Acylase I significantly decreased the production of virulence factors, namely elastase, pyocyanin, and pyoverdine under nicotine stress compared to the levels observed in the absence of nicotine stress. Thus, QQ is more efficient under stress. To our knowledge, this is the first study to report that QS contributes to nicotine tolerance in P. aeruginosa. This work facilitates a better application of QQ for the treatment of bacterial infections, especially under stress.

  14. Regulation of Nicotine Tolerance by Quorum Sensing and High Efficiency of Quorum Quenching Under Nicotine Stress in Pseudomonas aeruginosa PAO1.

    Science.gov (United States)

    Tang, Huiming; Zhang, Yunyun; Ma, Yifan; Tang, Mengmeng; Shen, Dongsheng; Wang, Meizhen

    2018-01-01

    Quorum sensing (QS) regulates the behavior of bacterial populations and promotes their adaptation and survival under stress. As QS is responsible for the virulence of vast majority of bacteria, quorum quenching (QQ), the interruption of QS, has become an attractive therapeutic strategy. However, the role of QS in stress tolerance and the efficiency of QQ under stress in bacteria are seldom explored. In this study, we demonstrated that QS-regulated catalase (CAT) expression and biofilm formation help Pseudomonas aeruginosa PAO1 resist nicotine stress. CAT activity and biofilm formation in wild type (WT) and Δ rhlR strains are significantly higher than those in the Δ lasR strain. Supplementation of Δ lasI strain with 3OC12-HSL showed similar CAT activity and biofilm formation as those of the WT strain. LasIR circuit rather than RhlIR circuit is vital to nicotine tolerance. Acylase I significantly decreased the production of virulence factors, namely elastase, pyocyanin, and pyoverdine under nicotine stress compared to the levels observed in the absence of nicotine stress. Thus, QQ is more efficient under stress. To our knowledge, this is the first study to report that QS contributes to nicotine tolerance in P. aeruginosa . This work facilitates a better application of QQ for the treatment of bacterial infections, especially under stress.

  15. Cold Plasma Inactivation of Bacterial Biofilms and Reduction of Quorum Sensing Regulated Virulence Factors.

    Directory of Open Access Journals (Sweden)

    Dana Ziuzina

    Full Text Available The main objectives of this work were to investigate the effect of atmospheric cold plasma (ACP against a range of microbial biofilms commonly implicated in foodborne and healthcare associated human infections and against P. aeruginosa quorum sensing (QS-regulated virulence factors, such as pyocyanin, elastase (Las B and biofilm formation capacity post-ACP treatment. The effect of processing factors, namely treatment time and mode of plasma exposure on antimicrobial activity of ACP were also examined. Antibiofilm activity was assessed for E. coli, L. monocytogenes and S. aureus in terms of reduction of culturability and retention of metabolic activity using colony count and XTT assays, respectively. All samples were treated 'inpack' using sealed polypropylene containers with a high voltage dielectric barrier discharge ACP generated at 80 kV for 0, 60, 120 and 300 s and a post treatment storage time of 24 h. According to colony counts, ACP treatment for 60 s reduced populations of E. coli to undetectable levels, whereas 300 s was necessary to significantly reduce populations of L. monocytogenes and S. aureus biofilms. The results obtained from XTT assay indicated possible induction of viable but non culturable state of bacteria. With respect to P. aeruginosa QS-related virulence factors, the production of pyocyanin was significantly inhibited after short treatment times, but reduction of elastase was notable only after 300 s and no reduction in actual biofilm formation was achieved post-ACP treatment. Importantly, reduction of virulence factors was associated with reduction of the cytotoxic effects of the bacterial supernatant on CHO-K1 cells, regardless of mode and duration of treatment. The results of this study point to ACP technology as an effective strategy for inactivation of established biofilms and may play an important role in attenuation of virulence of pathogenic bacteria. Further investigation is warranted to propose direct evidence

  16. Biofilm Formation on Stainless Steel by Streptococcus thermophilus UC8547 in Milk Environments Is Mediated by the Proteinase PrtS.

    Science.gov (United States)

    Bassi, D; Cappa, F; Gazzola, S; Orrù, L; Cocconcelli, P S

    2017-04-15

    In Streptococcus thermophilus , gene transfer events and loss of ancestral traits over the years contribute to its high level of adaptation to milk environments. Biofilm formation capacity, a phenotype that is lost in the majority of strains, plays a role in persistence in dairy environments, such as milk pasteurization and cheese manufacturing plants. To investigate this property, we have studied S. thermophilus UC8547, a fast-acidifying dairy starter culture selected for its high capacity to form biofilm on stainless steel under environmental conditions resembling the dairy environment. Using a dynamic flow cell apparatus, it was shown that S. thermophilus UC8547 biofilm formation on stainless steel depends on the presence of milk proteins. From this strain, which harbors the prtS gene for the cell wall protease and shows an aggregative phenotype, spontaneous mutants with impaired biofilm capacity can be isolated at high frequency. These mutants lack the PrtS expendable island, as confirmed by comparison of the genome sequence of UC8547Δ3 with that of the parent strain. The prtS island excision occurs between two 26-bp direct repeats located in the two copies of the IS Sth1 flanking this genomic island. The central role of PrtS was confirmed by analyzing the derivative strain UC8547Δ16, whose prtS gene was interrupted by an insertional mutation, thereby making it incapable of biofilm formation. PrtS, acting as a binding substance between the milk proteins adhered to stainless steel and S. thermophilus cell envelopes, mediates biofilm formation in dairy environments. This feature provides S. thermophilus with an ecological benefit for its survival and persistence in this environment. IMPORTANCE The increased persistence of S. thermophilus biofilm has consequences in the dairy environment: if, on the one hand, the release of this microorganism from biofilm can promote the fermentation of artisanal cheeses, under industrial conditions it may lead to undesirable

  17. Host defense peptides of thrombin modulate inflammation and coagulation in endotoxin-mediated shock and Pseudomonas aeruginosa sepsis.

    Science.gov (United States)

    Kalle, Martina; Papareddy, Praveen; Kasetty, Gopinath; Mörgelin, Matthias; van der Plas, Mariena J A; Rydengård, Victoria; Malmsten, Martin; Albiger, Barbara; Schmidtchen, Artur

    2012-01-01

    Gram-negative sepsis is accompanied by a disproportionate innate immune response and excessive coagulation mainly induced by endotoxins released from bacteria. Due to rising antibiotic resistance and current lack of other effective treatments there is an urgent need for new therapies. We here present a new treatment concept for sepsis and endotoxin-mediated shock, based on host defense peptides from the C-terminal part of human thrombin, found to have a broad and inhibitory effect on multiple sepsis pathologies. Thus, the peptides abrogate pro-inflammatory cytokine responses to endotoxin in vitro and in vivo. Furthermore, they interfere with coagulation by modulating contact activation and tissue factor-mediated clotting in vitro, leading to normalization of coagulation responses in vivo, a previously unknown function of host defense peptides. In a mouse model of Pseudomonas aeruginosa sepsis, the peptide GKY25, while mediating a modest antimicrobial effect, significantly inhibited the pro-inflammatory response, decreased fibrin deposition and leakage in the lungs, as well as reduced mortality. Taken together, the capacity of such thrombin-derived peptides to simultaneously modulate bacterial levels, pro-inflammatory responses, and coagulation, renders them attractive therapeutic candidates for the treatment of invasive infections and sepsis.

  18. Host defense peptides of thrombin modulate inflammation and coagulation in endotoxin-mediated shock and Pseudomonas aeruginosa sepsis.

    Directory of Open Access Journals (Sweden)

    Martina Kalle

    Full Text Available Gram-negative sepsis is accompanied by a disproportionate innate immune response and excessive coagulation mainly induced by endotoxins released from bacteria. Due to rising antibiotic resistance and current lack of other effective treatments there is an urgent need for new therapies. We here present a new treatment concept for sepsis and endotoxin-mediated shock, based on host defense peptides from the C-terminal part of human thrombin, found to have a broad and inhibitory effect on multiple sepsis pathologies. Thus, the peptides abrogate pro-inflammatory cytokine responses to endotoxin in vitro and in vivo. Furthermore, they interfere with coagulation by modulating contact activation and tissue factor-mediated clotting in vitro, leading to normalization of coagulation responses in vivo, a previously unknown function of host defense peptides. In a mouse model of Pseudomonas aeruginosa sepsis, the peptide GKY25, while mediating a modest antimicrobial effect, significantly inhibited the pro-inflammatory response, decreased fibrin deposition and leakage in the lungs, as well as reduced mortality. Taken together, the capacity of such thrombin-derived peptides to simultaneously modulate bacterial levels, pro-inflammatory responses, and coagulation, renders them attractive therapeutic candidates for the treatment of invasive infections and sepsis.

  19. The Pseudomonas aeruginosa PSL polysaccharide is a social but noncheatable trait in biofilms

    DEFF Research Database (Denmark)

    Irie, Yasuhiko; Roberts, Aled E.L.; Kragh, Kasper N.

    2017-01-01

    Extracellular polysaccharides are compounds secreted by microorganisms into the surrounding environment, and they are important for surface attachment and maintaining structural integrity within biofilms. The social nature of many extracellular polysaccharides remains unclear, and it has been sug...

  20. Characterization of the Pseudomonas aeruginosa recA analog and its protein product: rec-102 is a mutant allele of the P. aeruginosa PAO recA gene

    Energy Technology Data Exchange (ETDEWEB)

    Kokjohn, T.A.; Miller, R.V.

    1987-04-01

    We cloned a 2.3-kilobase-pair fragment of the Pseudomonas aeruginosa PAO chromosome which is capable of complementing recA mutations of Escherichia coli. The recA-complementing activity was further localized to a 1.5-kilobase-pair PvuII-HindIII fragment. Southern blot analysis under conditions of high stringency indicated that DNA sequence homology is shared by the E. coli recA gene and the P. aeruginosa recA analog. The cloned recA analog was shown to restore resistance to methyl methanesulfonate, nitrofurantoin, and UV irradiation to E. coli recA mutants. Upon introduction of the cloned P. aeruginosa gene, these mutants regained recombination proficiency in HfrH-mediated conjugation and the ability to induce lambda prophages and SOS functions (din gene transcription) after exposure to DNA-damaging agents. Lambda prophage carrying a cI ind mutation was not inducible, suggesting that the mechanism of induction of these SOS functions by the P. aeruginosa RecA analog is similar to that by the activated E. coli RecA protein. The product of the recA analog was identified in minicells as a protein of approximately 47,000 daltons. Western blot analysis using anti-E. coli RecA antibody demonstrated that this protein is antigenically cross-reactive with the E. coli recA protein. The recA-containing fragment was cloned into the broad-host-range vector pCP13 and introduced into Rec- strains of P. aeruginosa containing the rec-102 allele. The plasmid was shown to restore recombination proficiency in FP5-mediated conjugations and to restore resistance to UV irradiation and methyl methanesulfonate to these Rec- mutants. It was shown that a wild-type allele of rec-102 is necessary for UV-mediated induction of D3 and F116 prophages. The cloned recA analog restored the UV inducibility of these prophages in rec-102 mutants.