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Sample records for aeruginosa forms biofilms

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

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    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.

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

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    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...... from biofilms formed by mucoid P. aeruginosa were investigated. Alginate is not an essential structure component for mucoid P. aeruginosa biofilms. Genetic studies revealed that Pel and Psl polysaccharides serve as essential scaffold and mediate macrocolony formation in mucoid P. aeruginosa biofilms...

  3. Pseudomonas aeruginosa biofilm infections

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    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...

  4. Interactions of Pseudomonas aeruginosa in predominant biofilm or planktonic forms of existence in mixed culture with Escherichia coli in vitro.

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    Kuznetsova, Marina V; Maslennikova, Irina L; Karpunina, Tamara I; Nesterova, Larisa Yu; Demakov, Vitaly A

    2013-09-01

    Pseudomonas aeruginosa and Escherichia coli are known to be involved in mixed communities in diverse niches. In this study we examined the influence of the predominant form of cell existence of and the exometabolite production by P. aeruginosa strains on interspecies interactions, in vitro. Bacterial numbers of P. aeruginosa and E. coli in mixed plankton cultures and biofilms compared with their numbers in single plankton cultures and biofilms changed in a different way, but were in accordance with the form of P. aeruginosa cell existence. The mass of a mixed-species biofilm was greater than the mass of a single-species biofilm. Among the mixed biofilms, the one with the "planktonic" P. aeruginosa strain had the least biomass. The total pyocyanin and pyoverdin levels were found to be lower in all mixed plankton cultures. Despite this, clinical P. aeruginosa strains irrespective of the predominant form of existence ("biofilm" or "planktonic") had a higher total concentration of exometabolites than did the reference strain in 12-24 h mixed cultures. The metabolism of E. coli, according to its bioluminescence, was reduced in mixed cultures, and the decrease was by 20- to 100-fold greater with the clinical Pseudomonas strains than the reference Pseudomonas strain. Thus, both the predominant form of existence of and the exometabolite production by distinct P. aeruginosa strains should be considered to fully understand the interspecies relationship and bacteria survival in natural communities.

  5. Characterization of biofilm-like structures formed by Pseudomonas aeruginosa in a synthetic mucus medium

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    Haley Cecily L

    2012-08-01

    Full Text Available Abstract Background The accumulation of thick stagnant mucus provides a suitable environment for the growth of Pseudomonas aeruginosa and Staphylococcus aureus within the lung alveoli of cystic fibrosis (CF patients. These infections cause significant lung damage, leading to respiratory failure and death. In an artificial mucin containing medium ASM+, P. aeruginosa forms structures that resemble typical biofilms but are not attached to any surface. We refer to these structures as biofilm like structures (BLS. Using ASM+ in a static microtiter plate culture system, we examined the roles of mucin, extracellular DNA, environmental oxygen (EO2, and quorum sensing (QS in the development of biofilm-like structures (BLS by P. aeruginosa; and the effect of EO2 and P. aeruginosa on S. aureus BLS. Results Under 20% EO2, P. aeruginosa strain PAO1 produced BLS that resemble typical biofilms but are confined to the ASM+ and not attached to the surface. Levels of mucin and extracellular DNA within the ASM+ were optimized to produce robust well developed BLS. At 10% EO2, PAO1 produced thicker, more developed BLS, while under 0% EO2, BLS production was diminished. In contrast, the S. aureus strain AH133 produced well-developed BLS only under 20% EO2. In PAO1, loss of the QS system genes rhlI and rhlR affected the formation of BLS in ASM+ in terms of both structure and architecture. Whether co-inoculated into ASM+ with AH133, or added to established AH133 BLS, PAO1 eliminated AH133 within 48–56 h. Conclusions The thick, viscous ASM+, which contains mucin and extracellular DNA levels similar to those found in the CF lung, supports the formation of biofilm-like structures similar to the aggregates described within CF airways. Alterations in environmental conditions or in the QS genes of P. aeruginosa, as occurs naturally during the progression of CF lung infection, affect the architecture and quantitative structural features of these BLS. Thus, ASM+ provides an

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

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

    2005-01-01

    Biofilms are thought to play a key role in the occurrence of lung infections by Pseudomonas aeruginosa in patients with cystic fibrosis (CF). In this study, 20 nonmucoid P. aeruginosa isolates collected during different periods of chronic infection from eight CF patients were assessed with respect...

  7. Pattern differentiation in co-culture biofilms formed by Staphylococcus aureus and Pseudomonas aeruginosa

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    Yang, Liang; Liu, Yang; Markussen, Trine;

    2011-01-01

    important for understanding of biofilm physiology and the treatment of biofilm-related infectious diseases. Here, we have investigated interactions of two of the major bacterial species of cystic fibrosis lung microbial communities -Pseudomonas aeruginosa and Staphylococcus aureus- when grown in co...

  8. Chemical analysis of cellular and extracellular carbohydrates of a biofilm-forming strain Pseudomonas aeruginosa PA14.

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    Charlène Coulon

    Full Text Available 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

  9. Silver against Pseudomonas aeruginosa biofilms

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

    2007-01-01

    Silver has been recognized for its antimicrobial properties for centuries. Most studies on the antibacterial efficacy of silver, with particular emphasis on wound healing, have been performed on planktonic bacteria. Our recent studies, however, strongly suggest that colonization of wounds involves...... 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...

  10. Pseudomonas aeruginosa Biofilm Infections

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    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...

  11. Photodynamic antibacterial and antibiofilm activity of RLP068/Cl against Staphylococcus aureus and Pseudomonas aeruginosa forming biofilms on prosthetic material.

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    Vassena, Christian; Fenu, Simone; Giuliani, Francesco; Fantetti, Lia; Roncucci, Gabrio; Simonutti, Giulio; Romanò, Carlo Luca; De Francesco, Raffaele; Drago, Lorenzo

    2014-07-01

    Prosthetic joint infections (PJIs) are becoming a growing public health concern in developed countries as more people undergo arthroplasty for bone fixation or joint replacement. Because a wide range of bacterial strains responsible for PJIs can produce biofilms on prosthetic implants and because the biofilm structure confers elevated bacterial resistance to antibiotic therapy, new drugs and therapies are needed to improve the clinical outcome of treatment of PJIs. Antimicrobial photodynamic therapy (APDT), a non-antibiotic broad-spectrum antimicrobial treatment, is also active against multidrug-resistant micro-organisms such as meticillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa. APDT uses a photosensitiser that targets bacterial cells following exposure to visible light. APDT with RLP068/Cl, a novel photosensitiser, was studied by confocal laser scanning microscopy (CLSM) to evaluate the disruption of MRSA and P. aeruginosa biofilms on prosthetic material. Quantitative CLSM studies showed a reduction in biofilm biomass (biofilm disruption) and a decrease in viable cell numbers, as determined by standard plate counting, in the S. aureus and P. aeruginosa biofilms exposed to APDT with the photosensitiser RLP068/Cl. APDT with RLP068/Cl may be a useful approach to the treatment of PJI-associated biofilms.

  12. Spaceflight promotes biofilm formation by Pseudomonas aeruginosa.

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    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. Pseudomonas aeruginosa Biofilms

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    Alhede, Maria; Bjarnsholt, Thomas; Givskov, Michael Christian;

    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...

  14. The roles of biofilm matrix polysaccharide Psl in mucoid Pseudomonas aeruginosa biofilms.

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    Ma, Luyan; Wang, Shiwei; Wang, Di; Parsek, Matthew R; Wozniak, Daniel J

    2012-07-01

    The opportunistic pathogen Pseudomonas aeruginosa causes life-threatening, persistent infections in patients with cystic fibrosis (CF). Persistence is attributed to the ability of these bacteria to form structured communities (biofilms). Biofilms rely on an extracellular polymeric substances matrix to maintain structure. Psl exopolysaccharide is a key matrix component of nonmucoid biofilms, yet the role of Psl in mucoid biofilms is unknown. In this report, using a variety of mutants in a mucoid P. aeruginosa background, we found that deletion of Psl-encoding genes dramatically decreased their biofilm formation ability, indicating that Psl is also a critical matrix component of mucoid biofilms. Our data also suggest that the overproduction of alginate leads to mucoid biofilms, which occupy more space, whereas Psl-dependent biofilms are densely packed. These data suggest that Psl polysaccharide may have significant contributions in biofilm persistence in patients with CF and may be helpful for designing therapies for P. aeruginosa CF infection.

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

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    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.

  16. Pseudomonas aeruginosa promotes Escherichia coli biofilm formation in nutrient-limited medium.

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    Alessandro Culotti

    Full Text Available Biofilms have been implicated as an important reservoir for pathogens and commensal enteric bacteria such as Escherichia coli in natural and engineered water systems. However, the processes that regulate the survival of E. coli in aquatic biofilms have not been thoroughly studied. We examined the effects of hydrodynamic shear and nutrient concentrations on E. coli colonization of pre-established Pseudomonas aeruginosa biofilms, co-inoculation of E. coli and P. aeruginosa biofilms, and P. aeruginosa colonization of pre-established E. coli biofilms. In nutritionally-limited R2A medium, E. coli dominated biofilms when co-inoculated with P. aeruginosa, and successfully colonized and overgrew pre-established P. aeruginosa biofilms. In more enriched media, P. aeruginosa formed larger clusters, but E. coli still extensively overgrew and colonized the interior of P. aeruginosa clusters. In mono-culture, E. coli formed sparse and discontinuous biofilms. After P. aeruginosa was introduced to these biofilms, E. coli growth increased substantially, resulting in patterns of biofilm colonization similar to those observed under other sequences of organism introduction, i.e., E. coli overgrew P. aeruginosa and colonized the interior of P. aeruginosa clusters. These results demonstrate that E. coli not only persists in aquatic biofilms under depleted nutritional conditions, but interactions with P. aeruginosa can greatly increase E. coli growth in biofilms under these experimental conditions.

  17. Novel Targets for Treatment of Pseudomonas aeruginosa Biofilms

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    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’s suscep......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......’s susceptibility to antibiotics. The presence of such biofilms is acknowledged to equal a persistent infection due to their inherent high tolerance to all antimicrobials and immune cells. In this chapter we discuss the mechanisms of biofilm tolerance. The latest biofilm research is reviewed and future treatment...

  18. Stratified growth in Pseudomonas aeruginosa biofilms

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    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...... carried an isopropyl-beta-D-thiogalactopyranoside (IPTG)-inducible gfpmut2 gene encoding a stable GFP. The second construct carried a GFP derivative, gfp-AGA, encoding an unstable GFP under the control of the growth-rate-dependent rrnBp(1) promoter. Both GFP reporters indicated that active protein...... of oxygen limitation in the biofilm. Oxygen microelectrode measurements showed that oxygen only penetrated approximately 50 mum into the biofilm. P. aeruginosa was incapable of anaerobic growth in the medium used for this investigation. These results show that while mature P. aeruginosa biofilms contain...

  19. Effects of ambroxol on alginate of mature Pseudomonas aeruginosa biofilms.

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    Li, Fang; Yu, Jialin; Yang, Hua; Wan, Zhenyan; Bai, Dan

    2008-07-01

    Biofilm-forming bacteria Pseudomonas aeruginosa is a common pathogen in mechanically ventilated newborns, which can cause life-threatening infections. Alginate of mucoid Pseudomonas aeruginosa biofilms is considered an important virulence factor which contributes to the resistance to antibiotics. Traditionally, ambroxol is widely used in newborns with lung problems as a mucolytic agent and antioxidant agent as well. And there are few studies that demonstrated the anti-biofilm activity of ambroxol. In this study, we found that ambroxol can affect the structure of mucoid Pseudomonas aeruginosa biofilms. Further, we found that ambroxol reduces the production of alginate, the expression of the important genes and the activity of key enzyme guanosine diphospho-D-mannose dehydrogenase (GDP-mannose dehydrogenase; GMD) which were involved in alginate biosynthesis.

  20. Pseudomonas aeruginosa facilitates Campylobacter jejuni growth in biofilms under oxic flow conditions.

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    Culotti, Alessandro; Packman, Aaron I

    2015-12-01

    We investigated the growth of Campylobacter jejuni in biofilms with Pseudomonas aeruginosa under oxic flow conditions. We observed the growth of C. jejuni in mono-culture, deposited on pre-established P. aeruginosa biofilms, and co-inoculated with P. aeruginosa. In mono-culture, C. jejuni was unable to form biofilms. However, deposited C. jejuni continuously grew on pre-established P. aeruginosa biofilms for a period of 3 days. The growth of scattered C. jejuni clusters was strictly limited to the P. aeruginosa biofilm surface, and no intergrowth was observed. Co-culturing of C. jejuni and P. aeruginosa also enabled the growth of both organisms in biofilms, with C. jejuni clusters developing on the surface of the P. aeruginosa biofilm. Dissolved oxygen (DO) measurements in the medium showed that P. aeruginosa biofilms depleted the effluent DO from 9.0 to 0.5 mg L(-1) 24 hours after inoculation. The localized microaerophilic environment generated by P. aeruginosa promoted the persistence and growth of C. jejuni. Our findings show that P. aeruginosa not only prolongs the survival of C. jejuni under oxic conditions, but also enables the growth of C. jejuni on the surface of P. aeruginosa biofilms.

  1. Glycopeptide dendrimers as Pseudomonas aeruginosa biofilm inhibitors.

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    Reymond, Jean-Louis; Bergmann, Myriam; Darbre, Tamis

    2013-06-01

    Synthetic glycopeptide dendrimers composed of a branched oligopeptide tree structure appended with glycosidic groups at its multiple N-termini were investigated for binding to the Pseudomonas aeruginosa lectins LecB and LecA. These lectins are partly responsible for the formation of antibiotic resistant biofilms in the human pathogenic bacterium P. aeruginosa, which causes lethal airway infections in immune-compromised and cystic fibrosis patients. Glycopeptide dendrimers with high affinity to the lectins were identified by screening of combinatorial libraries. Several of these dendrimers, in particular the LecB specific glycopeptide dendrimers FD2 and D-FD2 and the LecA specific glycopeptide dendrimers GalAG2 and GalBG2, also efficiently block P. aeruginosa biofilm formation and induce biofilm dispersal in vitro. Structure-activity relationship and structural studies are reviewed, in particular the observation that multivalency is essential to the anti-biofilm effect in these dendrimers.

  2. Pseudomonas aeruginosa biofilms in cystic fibrosis

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    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. The immune system vs. Pseudomonas aeruginosa biofilms

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

    2010-01-01

    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...... established. However, because biofilm infections are often persistent (or chronic), an odd situation appears with the simultaneous activation of both arms of the host immune response, neither of which can eliminate the biofilm pathogen, but instead, in synergy, causes collateral tissue damage. Although...... the present review on the immune system vs. biofilm bacteria is focused on Pseudomonas aeruginosa (mainly because this is the most thoroughly studied), many of the same mechanisms are also seen with biofilm infections generated by other microorganisms....

  4. Some bacterial parameters influencing the neutrophil oxidative burst response to Pseudomonas aeruginosa biofilms

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    Jensen, E T; Kharazmi, A; Høiby, N;

    1992-01-01

    or biofilms is an important protective mechanism of the microorganisms. We examined the human PMN oxidative burst response to P. aeruginosa in biofilm and in planktonic form. The PMN chemiluminescence response to P. aeruginosa in biofilms was reduced to 30.5-47.5% (p less than 0.04) and the superoxide...... conclude that biofilm bacteria, although able to stimulate the PMN, result in a reduced, suboptimal response leading to lack of efficient eradication of the bacteria in the chronic infection....

  5. Anionic fluoroquinolones as antibacterials against biofilm-producing Pseudomonas aeruginosa.

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    Long, Timothy E; Keding, Lexie C; Lewis, Demetria D; Anstead, Michael I; Withers, T Ryan; Yu, Hongwei D

    2016-02-15

    Pseudomonas aeruginosa is a common biofilm-forming bacterial pathogen implicated in diseases of the lungs. The extracellular polymeric substances (EPS) of respiratory Pseudomonas biofilms are largely comprised of anionic molecules such as rhamnolipids and alginate that promote a mucoid phenotype. In this Letter, we examine the ability of negatively-charged fluoroquinolones to transverse the EPS and inhibit the growth of mucoid P. aeruginosa. Anionic fluoroquinolones were further compared with standard antibiotics via a novel microdiffusion assay to evaluate drug penetration through pseudomonal alginate and respiratory mucus from a patient with cystic fibrosis.

  6. Complement activation by Pseudomonas aeruginosa biofilms

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    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...... 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...... influx of neutrophils are known to cause inflammatory changes in the lungs. P. aeruginosa persisting in biofilms may contribute to the constant inflammation taking place in the lungs of CF patients....

  7. Ginger extract inhibits biofilm formation by Pseudomonas aeruginosa PA14.

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    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.

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

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    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.

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

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    Pamp, Sünje Johanna; Tolker-Nielsen, Tim

    2007-01-01

    . aeruginosa rhl4 mutants were defective in migration-dependent development of mushroom-shaped multicellular structures in the later phase of biofilm formation. Experiments involving three-color-coded mixed-strain P. aeruginosa biofilms demonstrated that the wild-type and rhl4 and pil4 mutant strains formed...

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

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

    2004-01-01

    Extracellular polymers can facilitate the non-specific attachment of bacteria to surfaces and hold together developing biofilms. This study was undertaken to qualitatively and quantitatively compare the architecture of biofilms produced by Pseudomonas aeruginosa strain PAO1 and its alginate......-overproducing (mucA22) and alginate-defective (algD) variants in order to discern the role of alginate in biofilm formation. These strains, PAO1, Alg(+) PAOmucA22 and Alg(-) PAOalgD, tagged with green fluorescent protein, were grown in a continuous flow cell system to characterize the developmental cycles...... 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...

  11. Functionalized polyanilines disrupt Pseudomonas aeruginosa and Staphylococcus aureus biofilms.

    Science.gov (United States)

    Gizdavic-Nikolaidis, Marija R; Pagnon, Joanne C; Ali, Naseem; Sum, Reuben; Davies, Noel; Roddam, Louise F; Ambrose, Mark

    2015-12-01

    The purpose of the present study was to investigate the antimicrobial effects of functionalized polyanilines (fPANIs) against stationary phase cells and biofilms of Pseudomonas aeruginosa and Staphylococcus aureus using homopolymer of sulfanilic acid (poly-SO3H) as a model. The chemically synthesized poly-SO3H was characterized using Fourier Transform Infra-Red (FTIR) and Ultraviolet-Visible (UV-Vis) spectroscopies. The molecular weight (Mw) and elemental analysis of homopolymer poly-SO3H were also examined. We found that poly-SO3H was bactericidal against stationary phase cells of P. aeruginosa and S. aureus at a concentration of 20 mgml(-1). Surprisingly, we discovered that the same concentration (20 mgml(-1)) of poly-SO3H significantly disrupted and killed bacterial cells present in pre-established forty-eight hour static biofilms of these organisms, as shown by crystal violet and bacterial live/dead fluorescence staining assays. In support of these data, poly-SO3H extensively diminished the expression of bacterial genes related to biofilm formation in stationary phase cells of P. aeruginosa, and seemed to greatly reduce the amount of the quorum sensing molecule N-(3-oxododecanoyl)-l-homoserine lactone (3OC12-HSL) able to be recovered from biofilms of this organism. Furthermore, we found that poly-SO3H was able to effectively penetrate and kill cells in biofilms formed by the P. aeruginosa (AESIII) isolate derived from the sputum of a cystic fibrosis patient. Taken together, the results of the present study emphasise the broad antimicrobial activities of fPANI, and suggest that they could be developed further and used in some novel ways to construct medical devices and/or industrial equipment that are refractory to colonization by biofilm-forming bacteria.

  12. Oxylipins produced by Pseudomonas aeruginosa promote biofilm formation and virulence

    Science.gov (United States)

    Martínez, Eriel; Campos-Gómez, Javier

    2016-01-01

    The oxygenation of unsaturated fatty acids by dioxygenases occurs in all kingdoms of life and produces physiologically important lipids called oxylipins. The biological roles of oxylipins have been extensively studied in animals, plants, algae and fungi, but remain largely unidentified in prokaryotes. The bacterium Pseudomonas aeruginosa displays a diol synthase activity that transforms several monounsaturated fatty acids into mono- and di-hydroxylated derivatives. Here we show that oxylipins derived from this activity inhibit flagellum-driven motility and upregulate type IV pilus-dependent twitching motility of P. aeruginosa. Consequently, these oxylipins promote bacterial organization in microcolonies, increasing the ability of P. aeruginosa to form biofilms in vitro and in vivo (in Drosophila flies). We also demonstrate that oxylipins produced by P. aeruginosa promote virulence in Drosophila flies and lettuce. Our study thus uncovers a role for prokaryotic oxylipins in the physiology and pathogenicity of bacteria. PMID:27929111

  13. 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.

  14. 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....

  15. Cell death in Pseudomonas aeruginosa biofilm development

    DEFF Research Database (Denmark)

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

    2003-01-01

    . 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....... We propose that prophage-mediated cell death is an important mechanism of differentiation inside microcolonies that facilitates dispersal of a subpopulation of surviving cells....

  16. In vitro prevention of Pseudomonas aeruginosa early biofilm formation with antibiotics used in cystic fibrosis patients.

    Science.gov (United States)

    Fernández-Olmos, Ana; García-Castillo, María; Maiz, Luis; Lamas, Adelaida; Baquero, Fernando; Cantón, Rafael

    2012-08-01

    The ability of antibiotics used in bronchopulmonary infections in cystic fibrosis (CF) patients to prevent Pseudomonas aeruginosa early biofilm formation was studied using a biofilm microtitre assay with 57 non-mucoid P. aeruginosa isolates (44 first colonisers and 13 recovered during the initial intermittent colonisation stage) obtained from 35 CF patients. Minimum biofilm inhibitory concentrations (BICs) of levofloxacin, ciprofloxacin, imipenem, ceftazidime, tobramycin, colistin and azithromycin were determined by placing a peg lid with a formed biofilm onto microplates containing antibiotics. A modification of this protocol consisting of antibiotic challenge during biofilm formation was implemented in order to determine the biofilm prevention concentration (BPC), i.e. the minimum concentration able to prevent biofilm formation. The lowest BPCs were for fluoroquinolones, tobramycin and colistin and the highest for ceftazidime and imipenem. The former antibiotics had BPCs identical to or only slightly higher than their minimum inhibitory concentrations (MICs) determined by standard Clinical and Laboratory Standards Institute (CLSI) microdilution and were also active on formed biofilms as reflected by their low BIC values. In contrast, ceftazidime and imipenem were less effective for prevention of biofilm formation and on formed biofilms. In conclusion, the new BPC parameter determined in non-mucoid P. aeruginosa isolates recovered during early colonisation stages in CF patients supports early aggressive antimicrobial treatment guidelines in first P. aeruginosa-colonised CF patients.

  17. 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.

  18. Biofilm formation by Pseudomonas aeruginosa wild type, flagella and type IV pili mutants

    DEFF Research Database (Denmark)

    Klausen, M.; Heydorn, Arne; Ragas, Paula Cornelia;

    2003-01-01

    Biofilm formation by Gfp-tagged Pseudomonas aeruginosa PAO1 wild type, flagella and type IV pili mutants in flow chambers irrigated with citrate minimal medium was characterized by the use of confocal laser scanning microscopy and comstat image analysis. Flagella and type IV pili were not necessary...... for P. aeruginosa initial attachment or biofilm formation, but the cell appendages had roles in biofilm development, as wild type, flagella and type IV pili mutants formed biofilms with different structures. Dynamics and selection during biofilm formation were investigated by tagging the wild type...... and flagella/type IV mutants with Yfp and Cfp and performing time-lapse confocal laser scanning microscopy in mixed colour biofilms. The initial microcolony formation occurred by clonal growth, after which wild-type P. aeruginosa bacteria spread over the substratum by means of twitching motility. The wild-type...

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

    DEFF Research Database (Denmark)

    Wu, Hong; Lee, Baoleri; Yang, Liang

    2011-01-01

    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......-associated chronic infections caused by P. aeruginosa....

  20. Glycerol metabolism promotes biofilm formation by Pseudomonas aeruginosa.

    Science.gov (United States)

    Scoffield, Jessica; Silo-Suh, Laura

    2016-08-01

    Pseudomonas aeruginosa causes persistent infections in the airways of cystic fibrosis (CF) patients. Airway sputum contains various host-derived nutrients that can be utilized by P. aeruginosa, including phosphotidylcholine, a major component of host cell membranes. Phosphotidylcholine can be degraded by P. aeruginosa to glycerol and fatty acids to increase the availability of glycerol in the CF lung. In this study, we explored the role that glycerol metabolism plays in biofilm formation by P. aeruginosa. We report that glycerol metabolism promotes biofilm formation by both a chronic CF isolate (FRD1) and a wound isolate (PAO1) of P. aeruginosa. Moreover, loss of the GlpR regulator, which represses the expression of genes involved in glycerol metabolism, enhances biofilm formation in FRD1 through the upregulation of Pel polysaccharide. Taken together, our results suggest that glycerol metabolism may be a key factor that contributes to P. aeruginosa persistence by promoting biofilm formation.

  1. 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...

  2. Anthranilate deteriorates the structure of Pseudomonas aeruginosa biofilms and antagonizes the biofilm-enhancing indole effect.

    Science.gov (United States)

    Kim, Soo-Kyoung; Park, Ha-Young; Lee, Joon-Hee

    2015-04-01

    Anthranilate and indole are alternative degradation products of tryptophan, depending on the bacterial species. While indole enhances the biofilm formation of Pseudomonas aeruginosa, we found that anthranilate, the tryptophan degradation product of P. aeruginosa, had an opposite effect on P. aeruginosa biofilm formation, in which anthranilate deteriorated the mushroom structure of biofilm. The anthranilate effect on biofilm formation was differentially exerted depending on the developmental stage and the presence of shear force. Anthranilate slightly accelerated the initial attachment of P. aeruginosa at the early stage of biofilm development and appeared to build more biofilm without shear force. But anthranilate weakened the biofilm structure in the late stage, deteriorating the mushroom structure of biofilms with shear force to make a flat biofilm. To investigate the interplay of anthranilate with indole in biofilm formation, biofilms were cotreated with anthranilate and indole, and the results showed that anthranilate antagonized the biofilm-enhancing effect of indole. Anthranilate was able to deteriorate the preformed biofilm. The effect of anthranilate and indole on biofilm formation was quorum sensing independent. AntR, a regulator of anthranilate-degrading metabolism was synergistically activated by cotreatment with anthranilate and indole, suggesting that indole might enhance biofilm formation by facilitating the degradation of anthranilate. Anthranilate slightly but significantly affected the cyclic diguaniylate (c-di-GMP) level and transcription of major extracellular polysaccharide (Psl, Pel, and alginate) operons. These results suggest that anthranilate may be a promising antibiofilm agent and antagonize the effect of indole on P. aeruginosa biofilm formation.

  3. 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....

  4. Extracellular DNA Shields against Aminoglycosides in Pseudomonas aeruginosa Biofilms

    DEFF Research Database (Denmark)

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

    2013-01-01

    , which are thought to be a source of extracellular DNA at sites of infections, increases the tolerance of P. aeruginosa biofilms toward aminoglycosides. Although biofilm-associated aminoglycoside tolerance recently has been linked to extracellular DNA-mediated activation of the pmr genes, we demonstrate...... that the aminoglycoside tolerance mediated by the presence of extracellular DNA is not caused by activation of the pmr genes in our P. aeruginosa biofilms but rather by a protective shield effect of the extracellular DNA....

  5. 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...

  6. Antipseudomonal agents exhibit differential pharmacodynamic interactions with human polymorphonuclear leukocytes against established biofilms of Pseudomonas aeruginosa.

    Science.gov (United States)

    Chatzimoschou, Athanasios; Simitsopoulou, Maria; Antachopoulos, Charalampos; Walsh, Thomas J; Roilides, Emmanuel

    2015-04-01

    Pseudomonas aeruginosa is the most common pathogen infecting the lower respiratory tract of cystic fibrosis (CF) patients, where it forms tracheobronchial biofilms. Pseudomonas biofilms are refractory to antibacterials and to phagocytic cells with innate immunity, leading to refractory infection. Little is known about the interaction between antipseudomonal agents and phagocytic cells in eradication of P. aeruginosa biofilms. Herein, we investigated the capacity of three antipseudomonal agents, amikacin (AMK), ceftazidime (CAZ), and ciprofloxacin (CIP), to interact with human polymorphonuclear leukocytes (PMNs) against biofilms and planktonic cells of P. aeruginosa isolates recovered from sputa of CF patients. Three of the isolates were resistant and three were susceptible to each of these antibiotics. The concentrations studied (2, 8, and 32 mg/liter) were subinhibitory for biofilms of resistant isolates, whereas for biofilms of susceptible isolates, they ranged between sub-MIC and 2 × MIC values. The activity of each antibiotic alone or in combination with human PMNs against 48-h mature biofilms or planktonic cells was determined by XTT [2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide] assay. All combinations of AMK with PMNs resulted in synergistic or additive effects against planktonic cells and biofilms of P. aeruginosa isolates compared to each component alone. More than 75% of CAZ combinations exhibited additive interactions against biofilms of P. aeruginosa isolates, whereas CIP had mostly antagonistic interaction or no interaction with PMNs against biofilms of P. aeruginosa. Our findings demonstrate a greater positive interaction between AMK with PMNs than that observed for CAZ and especially CIP against isolates of P. aeruginosa from the respiratory tract of CF patients.

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

    Science.gov (United States)

    Vital-Lopez, Francisco G; Reifman, Jaques; Wallqvist, Anders

    2015-10-01

    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 cells. Developing treatments against biofilms requires an understanding of bacterial biofilm-specific physiological traits. Research efforts have started to elucidate the intricate mechanisms underlying biofilm development. However, many aspects of these mechanisms are still poorly understood. Here, we addressed questions regarding biofilm metabolism using a genome-scale kinetic model of the P. aeruginosa metabolic network and gene expression profiles. Specifically, we computed metabolite concentration differences between known mutants with altered biofilm formation and the wild-type strain to predict drug targets against P. aeruginosa biofilms. We also simulated the altered metabolism driven by gene expression changes between biofilm and stationary growth-phase planktonic cultures. Our analysis suggests that the synthesis of important biofilm-related molecules, such as the quorum-sensing molecule Pseudomonas quinolone signal and the exopolysaccharide Psl, is regulated not only through the expression of genes in their own synthesis pathway, but also through the biofilm-specific expression of genes in pathways competing for precursors to these molecules. Finally, we investigated why mutants defective in anthranilate degradation have an impaired ability to form biofilms. Alternative to a previous hypothesis that this biofilm reduction is caused by a decrease in energy production, we proposed that the dysregulation of the synthesis of secondary metabolites derived from anthranilate and chorismate is what impaired the biofilms of these mutants. Notably, these insights generated through our kinetic model-based approach are not accessible from previous constraint-based model analyses of P. aeruginosa biofilm

  8. Effects of Chlorine Stress on Pseudomonas aeruginosa Biofilm and Analysis of Related Gene Expressions.

    Science.gov (United States)

    Kekeç, Özge; Gökalsın, Barış; Karaltı, İskender; Kayhan, Figen Esin; Sesal, Nüzhet Cenk

    2016-08-01

    Chlorine is deployed worldwide to clean waters and prevent water-originated illnesses. However, chlorine has a limited disinfection capacity against biofilms. Microorganisms form biofilms to protect themselves from biological threats such as disinfectant chemicals. Pseudomonas aeruginosa is an opportunistic pathogen and its biofilm form attaches to surfaces, living buried into exopolysaccharides, can be present in all watery environments including tap water and drinking water. This research aimed to study the biofilm trigger mechanism of the opportunistic pathogen P. aeruginosa PAO1 strain, which is known to form biofilm in water supply systems and human body, under chlorine stress levels. In addition to biofilm staining, certain genes that are relevant to the stress condition were selected for gene expression analysis. The bacteria cultures were grown under chlorine stress with concentrations of 0.5, 0.7 and 1 mg/l. Six gene regions were determined related to biofilm and stress response: rpoS, bifA, migA, katB, soxR, and algC. Biofilm formation was analyzed by basic fuchsin staining, and gene expressions were quantified by quantitative real-time PCR. According to the results, highest biofilm production was observed in P. aeruginosa PAO1 wild strain under no stress conditions. Higher biofilm amounts were observed for bacteria under 0.5 and 0.7 mg/l chlorine stress compared to 1 mg/l chlorine stress.

  9. Magnetic fields suppress Pseudomonas aeruginosa biofilms and enhance ciprofloxacin activity.

    Science.gov (United States)

    Bandara, H M H N; Nguyen, D; Mogarala, S; Osiñski, M; Smyth, H D C

    2015-01-01

    Due to the refractory nature of pathogenic microbial biofilms, innovative biofilm eradication strategies are constantly being sought. Thus, this study addresses a novel approach to eradicate Pseudomonas aeruginosa biofilms. Magnetic nanoparticles (MNP), ciprofloxacin (Cipro), and magnetic fields were systematically evaluated in vitro for their relative anti-biofilm contributions. Twenty-four-hour biofilms exposed to aerosolized MNPs, Cipro, or a combination of both, were assessed in the presence or absence of magnetic fields (Static one-sided, Static switched, Oscillating, Static + oscillating) using changes in bacterial metabolism, biofilm biomass, and biofilm imaging. The biofilms exposed to magnetic fields alone exhibited significant metabolic and biomass reductions (p biofilms were treated with a MNP/Cipro combination, the most significant metabolic and biomass reductions were observed when exposed to static switched magnetic fields (p biofilms to a static switched magnetic field alone, or co-administration with MNP/Cipro/MNP + Cipro appears to be a promising approach to eradicate biofilms of this bacterium.

  10. Biofilm assembly becomes crystal clear – filamentous bacteriophage organize the Pseudomonas aeruginosa biofilm matrix into a liquid crystal

    Science.gov (United States)

    Secor, Patrick R.; Jennings, Laura K.; Michaels, Lia A.; Sweere, Johanna M.; Singh, Pradeep K.; Parks, William C.; Bollyky, Paul L.

    2015-01-01

    Pseudomonas aeruginosa is an opportunistic bacterial pathogen associated with many types of chronic infection. At sites of chronic infection, such as the airways of people with cystic fibrosis (CF), P. aeruginosa forms biofilm-like aggregates. These are clusters of bacterial cells encased in a polymer-rich matrix that shields bacteria from environmental stresses and antibiotic treatment. When P. aeruginosa forms a biofilm, large amounts of filamentous Pf bacteriophage (phage) are produced. Unlike most phage that typically lyse and kill their bacterial hosts, filamentous phage of the genus Inovirus, which includes Pf phage, often do not, and instead are continuously extruded from the bacteria. Here, we discuss the implications of the accumulation of filamentous Pf phage in the biofilm matrix, where they interact with matrix polymers to organize the biofilm into a highly ordered liquid crystal. This structural configuration promotes bacterial adhesion, desiccation survival, and antibiotic tolerance – all features typically associated with biofilms. We propose that Pf phage make structural contributions to P. aeruginosa biofilms and that this constitutes a novel form of symbiosis between bacteria and bacteriophage.

  11. Biofilm assembly becomes crystal clear – filamentous bacteriophage organize the Pseudomonas aeruginosa biofilm matrix into a liquid crystal

    Directory of Open Access Journals (Sweden)

    Patrick R. Secor

    2015-12-01

    Full Text Available Pseudomonas aeruginosa is an opportunistic bacterial pathogen associated with many types of chronic infection. At sites of chronic infection, such as the airways of people with cystic fibrosis (CF, P. aeruginosa forms biofilm-like aggregates. These are clusters of bacterial cells encased in a polymer-rich matrix that shields bacteria from environmental stresses and antibiotic treatment. When P. aeruginosa forms a biofilm, large amounts of filamentous Pf bacteriophage (phage are produced. Unlike most phage that typically lyse and kill their bacterial hosts, filamentous phage of the genus Inovirus, which includes Pf phage, often do not, and instead are continuously extruded from the bacteria. Here, we discuss the implications of the accumulation of filamentous Pf phage in the biofilm matrix, where they interact with matrix polymers to organize the biofilm into a highly ordered liquid crystal. This structural configuration promotes bacterial adhesion, desiccation survival, and antibiotic tolerance – all features typically associated with biofilms. We propose that Pf phage make structural contributions to P. aeruginosa biofilms and that this constitutes a novel form of symbiosis between bacteria and bacteriophage.

  12. In vitro management of hospital Pseudomonas aeruginosa biofilm using indigenous T7-like lytic phage.

    Science.gov (United States)

    Ahiwale, Sangeeta; Tamboli, Nilofer; Thorat, Kiran; Kulkarni, Rajendra; Ackermann, Hans; Kapadnis, Balasaheb

    2011-02-01

    Pseudomonas aeruginosa, a human pathogen capable of forming biofilm and contaminating medical settings, is responsible for 65% mortality in the hospitals all over the world. This study was undertaken to isolate lytic phages against biofilm forming Ps. aeruginosa hospital isolates and to use them for in vitro management of biofilms in the microtiter plate. Multidrug resistant strains of Ps. aeruginosa were isolated from the hospital environment in and around Pimpri-Chinchwad, Maharashtra by standard microbiological methods. Lytic phages against these strains were isolated from the Pavana river water by double agar layer plaque assay method. A wide host range phage bacterial virus Ps. aeruginosa phage (BVPaP-3) was selected. Electron microscopy revealed that BVPaP-3 phage is a T7-like phage and is a relative of phage species gh-1. A phage at MOI-0.001 could prevent biofilm formation by Ps. aeruginosa hospital strain-6(HS6) on the pegs within 24 h. It could also disperse pre-formed biofilms of all hospital isolates (HS1-HS6) on the pegs within 24 h. Dispersion of biofilm was studied by monitoring log percent reduction in cfu and log percent increase in pfu of respective bacterium and phage on the peg as well as in the well. Scanning electron microscopy confirmed that phage BVPaP-3 indeed causes biofilm reduction and bacterial cell killing. Laboratory studies prove that BVPaP-3 is a highly efficient phage in preventing and dispersing biofilms of Ps. aeruginosa. Phage BVPaP-3 can be used as biological disinfectant to control biofilm problem in medical devices.

  13. 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...

  14. 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

    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...

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

    DEFF Research Database (Denmark)

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

    2010-01-01

    . 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......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...... 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....

  16. The action of Pseudomonas aeruginosa biofilms in intrinsic drug resistance

    Institute of Scientific and Technical Information of China (English)

    XIE Yi; JIA Wen-xiang; ZENG Wei; YANG Wei-qing; CHENG Xi; LI Xue-ru; WANG Lan-lan; KANG Mei; ZHANG Zai-rong

    2005-01-01

    Background There is a growing interest in studying the relationship between intrinsic resistance and biofilms resistance to drugs. However, the relationship still remains unclear in the macroscopic bacterial growth. Our study is to illuminate the change of bacterial drug resistance of gyrA mutant and active efflux pump during the development of Pseudomonas aeruginosa (P. aeruginosa) biofilms. Methods The strains of type Ⅱ topoisomerase gene mutant (gyrA mutant) and multidrug resistance (MDR) efflux pump were clinical isolates and detected by polymerase chain reaction (PCR). The process of bacterial biofilms development was observed by scanning electron microscope. Triparental mating experiments were performed to transfer report gene of green fluorescent protein (GFP) into P. aeruginosa biofilms strains and followed by analysis of bacterial survival rate between intrinsic resistance and biofilms resistance.Results The fluorescent strains with pGFPuv could develop mature biofilms on Teflon surface. Before a period of 72 hours, the survival rate of biofilms bacteria and intrinsic resistance strains in ciprofloxacin solution was significantly different (P0.05). The carbonyl cyanide m-chlorophenylhydrazone and azithromycin could significantly reduce the drug resistance of biofilm strains and efflux pump strains.Conclusions In the development of P. aeruginosa biofilms, the strains of gyrA mutation and MDR efflux could be conferred with new level of drug resistance. When co-cultured mutated strains with biofilm strains, biofilms may play a major role in bacterial resistance. But after 72 hours incubation (a mature biofilms had been developed), there was no clearly difference between the number of mutant strains and biofilm strains.

  17. 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.

  18. Inhibition of Pseudomonas aeruginosa biofilm formation on wound dressings.

    Science.gov (United States)

    Brandenburg, Kenneth S; Calderon, Diego F; Kierski, Patricia R; Brown, Amanda L; Shah, Nihar M; Abbott, Nicholas L; Schurr, Michael J; Murphy, Christopher J; McAnulty, Jonathan F; Czuprynski, Charles J

    2015-01-01

    Chronic nonhealing skin wounds often contain bacterial biofilms that prevent normal wound healing and closure and present challenges to the use of conventional wound dressings. We investigated inhibition of Pseudomonas aeruginosa biofilm formation, a common pathogen of chronic skin wounds, on a commercially available biological wound dressing. Building on prior reports, we examined whether the amino acid tryptophan would inhibit P. aeruginosa biofilm formation on the three-dimensional surface of the biological dressing. Bacterial biomass and biofilm polysaccharides were quantified using crystal violet staining or an enzyme linked lectin, respectively. Bacterial cells and biofilm matrix adherent to the wound dressing were visualized through scanning electron microscopy. D-/L-tryptophan inhibited P. aeruginosa biofilm formation on the wound dressing in a dose dependent manner and was not directly cytotoxic to immortalized human keratinocytes although there was some reduction in cellular metabolism or enzymatic activity. More importantly, D-/L-tryptophan did not impair wound healing in a splinted skin wound murine model. Furthermore, wound closure was improved when D-/L-tryptophan treated wound dressing with P. aeruginosa biofilms were compared with untreated dressings. These findings indicate that tryptophan may prove useful for integration into wound dressings to inhibit biofilm formation and promote wound healing.

  19. Alginate overproduction affects Pseudomonas aeruginosa biofilm structure and function

    DEFF Research Database (Denmark)

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

    2001-01-01

    During the course of chronic cystic fibrosis (CF) infections, Pseudomonas aeruginosa undergoes a conversion to a mucoid phenotype, which is characterized by overproduction of the exopolysaccharide alginate. Chronic P. aeruginosa infections involve surface-attached, highly antibiotic-resistant com......During the course of chronic cystic fibrosis (CF) infections, Pseudomonas aeruginosa undergoes a conversion to a mucoid phenotype, which is characterized by overproduction of the exopolysaccharide alginate. Chronic P. aeruginosa infections involve surface-attached, highly antibiotic......-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...

  20. Role of mutation in Pseudomonas aeruginosa biofilm development.

    Directory of Open Access Journals (Sweden)

    Tim C R Conibear

    Full Text Available The survival of bacteria in nature is greatly enhanced by their ability to grow within surface-associated communities called biofilms. Commonly, biofilms generate proliferations of bacterial cells, called microcolonies, which are highly recalcitrant, 3-dimensional foci of bacterial growth. Microcolony growth is initiated by only a subpopulation of bacteria within biofilms, but processes responsible for this differentiation remain poorly understood. Under conditions of crowding and intense competition between bacteria within biofilms, microevolutionary processes such as mutation selection may be important for growth; however their influence on microcolony-based biofilm growth and architecture have not previously been explored. To study mutation in-situ within biofilms, we transformed Pseudomonas aeruginosa cells with a green fluorescent protein gene containing a +1 frameshift mutation. Transformed P. aeruginosa cells were non-fluorescent until a mutation causing reversion to the wildtype sequence occurs. Fluorescence-inducing mutations were observed in microcolony structures, but not in other biofilm cells, or in planktonic cultures of P. aeruginosa cells. Thus microcolonies may represent important foci for mutation and evolution within biofilms. We calculated that microcolony-specific increases in mutation frequency were at least 100-fold compared with planktonically grown cultures. We also observed that mutator phenotypes can enhance microcolony-based growth of P. aeruginosa cells. For P. aeruginosa strains defective in DNA fidelity and error repair, we found that microcolony initiation and growth was enhanced with increased mutation frequency of the organism. We suggest that microcolony-based growth can involve mutation and subsequent selection of mutants better adapted to grow on surfaces within crowded-cell environments. This model for biofilm growth is analogous to mutation selection that occurs during neoplastic progression and tumor

  1. The effects of D-Tyrosine combined with amikacin on the biofilms of Pseudomonas aeruginosa.

    Science.gov (United States)

    She, Pengfei; Chen, Lihua; Liu, Hongbo; Zou, Yaru; Luo, Zhen; Koronfel, Asmaa; Wu, Yong

    2015-09-01

    The biofilm formation of microorganisms causes persistent tissue infections resistant to treatment with antimicrobial agents. Pseudomonas aeruginosa is commonly isolated from the airways of patients with chronic fibrosis (CF) and often forms biofilms, which are extremely hard to eradicate and a major cause of mortality and morbidity. Recent studies have shown that D-amino acids (D-AAs) inhibited and disrupted biofilm formation by causing the release of the protein component of the polymeric matrix. However, the effects of D-AAs combined with common antibiotics on biofilms have rarely been studied. The current study first determined whether D-AAs disrupted the biofilms of PAO1 and the clinical airway isolates of P. aeruginosa. It was then determined whether combinations of D-Tyr (the most effective one) and the antibiotic amikacin (AMK) enhanced the activity against these biofilms. The results of the current study showed that D-Tyr is the most effective among those that disassemble the D-amino acids (D-leucine, D-methionine, D-Tyrptophan, and D-tryptophan), and D-Tyr at concentrations higher than 5 mM significantly reduced the biofilm biomass of P. aeruginosa (p biofilms, as indicated by a reduction in the minimal biofilm-inhibiting concentration (MBIC50 and MBIC90) without a change in the minimal inhibitory concentration (MIC) of planktonic bacteria. Thus, the findings indicated that D-Tyr supplementation overcame the resistance of P. aeruginosa biofilms to AMK, which might be helpful for preventing AMK overuse when this specific D-Tyr is recommended for combatting these biofilms. Also, toxicity of the liver and kidney from AMK could be potentially mitigated by co-delivery with D-Tyr.

  2. Pseudomonas aeruginosa biofilm growth inhibition on medical plastic materials by immobilized esterases and acylase.

    Science.gov (United States)

    Kisch, Johannes Martin; Utpatel, Christian; Hilterhaus, Lutz; Streit, Wolfgang R; Liese, Andreas

    2014-09-01

    Biofilms are matrix-encapsulated cell aggregates that cause problems in technical and health-related areas; for example, 65 % of all human infections are biofilm associated. This is mainly due to their ameliorated resistance against antimicrobials and immune systems. Pseudomonas aeruginosa, a biofilm-forming organism, is commonly responsible for nosocomial infections. Biofilm development is partly mediated by signal molecules, such as acyl-homoserine lactones (AHLs) in Gram-negative bacteria. We applied horse liver esterase, porcine kidney acylase, and porcine liver esterase; these can hydrolyze AHLs, thereby inhibiting biofilm formation. As biofilm infections are often related to foreign material introduced into the human body, we immobilized the enzymes on medical plastic materials. Biofilm formation was quantified by Crystal Violet staining and confocal laser scanning microscopy, revealing up to 97 % (on silicone), 54 % (on polyvinyl chloride), and 77 % (on polyurethane) reduced biomass after 68 h growth.

  3. The impact of quorum sensing and swarming motility on Pseudomonas aeruginosa biofilm formation is nutritionally conditional

    DEFF Research Database (Denmark)

    Shrout, J.D.; Chopp, D.L.; Just, C.L.

    2006-01-01

    The role of quorum sensing in Pseudomonas aeruginosa biofilm formation is unclear. Some researchers have shown that quorum sensing is important for biofilm development, while others have indicated it has little or no role. In this study, the contribution of quorum sensing to biofilm development...... was found to depend upon the nutritional environment. Depending upon the carbon source, quorum-sensing mutant strains (lasIrhlI and lasRrhlR) either exhibited a pronounced defect early in biofilm formation or formed biofilms identical to the wild-type strain. Quorum sensing was then shown to exert its...... nutritionally conditional control of biofilm development through regulation of swarming motility. Examination of pilA and fliM mutant strains further supported the role of swarming motility in biofilm formation. These data led to a model proposing that the prevailing nutritional conditions dictate...

  4. Ambroxol inhibits mucoid conversion of Pseudomonas aeruginosa and contributes to the bactericidal activity of ciprofloxacin against mucoid P. aeruginosa biofilms.

    Science.gov (United States)

    Wang, Wenlei; Yu, Jialin; He, Yu; Wang, Zhengli; Li, Fang

    2016-07-01

    Pseudomonas aeruginosa is an opportunistic human pathogen that can cause severe infections in immunocompromised individuals. Because it forms biofilms, which protect against host immune attack and increase resistance to conventional antibiotics, mucoid P. aeruginosa is nearly impossible to eradicate. Moreover, mucoid conversion of P. aeruginosa in cystic fibrosis (CF) patients leads to poor outcomes. This conversion is mainly due to mucA gene mutation, which is thought to be induced by polymorphonuclear leukocytes (PMNs) and the reactive oxygen species they release. Ambroxol, a mucolytic agent with antioxidant characteristics, is used clinically, and this compound has recently been demonstrated to possess anti-biofilm properties. In this study, we found that ambroxol inhibits the H2 O2 -mediated conversion of P. aeruginosa from a non-mucoid to a mucoid phenotype, an effect that is due to its antioxidant property against H2 O2 . Furthermore, the bactericidal activity of ciprofloxacin against mucoid P. aeruginosa biofilms was increased in vitro when used in combination with ambroxol.

  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. Synergistic effect of fosfomycin and fluoroquinolones against Pseudomonas aeruginosa growing in a biofilm.

    Directory of Open Access Journals (Sweden)

    Mikuniya,Takeshi

    2005-10-01

    Full Text Available

    Ulifloxacin is the active form of the prodrug prulifloxacin and shows a highly potent antipseudomonal activity. In this study, we examined the combined effect of fosfomycin and ulifloxacin against Pseudomonas aeruginosa (P. aeruginosa growing in a biofilm using a modified Robbins device with artificial urine, and compared it to that of the combination of fosfomycin and ciprofloxacin or levofloxacin. An ATP bioluminescence assay was used to evaluate the antibacterial activity of the agents against sessile cells in a mature biofilm developed on a silicon disk. The total bioactivity of P. aeruginosa growing in a biofilm that had not been fully eradicated by fosfomycin or any of the fluoroquinolones alone at 10 times the MIC decreased after combination treatment with fosfomycin and fluoroquinolones. Morphological changes occurred in a time-dependent fashion; namely, swollen and/or rounding cells emerged within a couple of hours after combination treatment, marking the initial stage in the process leading to the destruction of the biofilms. We could not find any difference among the 3 fluoroquinolones with regard to their synergistic effects when administered with fosfomycin. The combination treatment of fosfomycin and fluoroquinolones with highly potent antipseudomonal activities was effective in eradicating sessile cells of P. aeruginosa in the biofilm and promises to be beneficial against biofilm-associated infectious diseases.

  7. An Integrated Modeling and Experimental Approach to Study the Influence of Environmental Nutrients on Biofilm Formation of Pseudomonas aeruginosa.

    Science.gov (United States)

    Xu, Zhaobin; Islam, Sabina; Wood, Thomas K; Huang, Zuyi

    2015-01-01

    The availability of nutrient components in the environment was identified as a critical regulator of virulence and biofilm formation in Pseudomonas aeruginosa. This work proposes the first systems-biology approach to quantify microbial biofilm formation upon the change of nutrient availability in the environment. Specifically, the change of fluxes of metabolic reactions that were positively associated with P. aeruginosa biofilm formation was used to monitor the trend for P. aeruginosa to form a biofilm. The uptake rates of nutrient components were changed according to the change of the nutrient availability. We found that adding each of the eleven amino acids (Arg, Tyr, Phe, His, Iso, Orn, Pro, Glu, Leu, Val, and Asp) to minimal medium promoted P. aeruginosa biofilm formation. Both modeling and experimental approaches were further developed to quantify P. aeruginosa biofilm formation for four different availability levels for each of the three ions that include ferrous ions, sulfate, and phosphate. The developed modeling approach correctly predicted the amount of biofilm formation. By comparing reaction flux change upon the change of nutrient concentrations, metabolic reactions used by P. aeruginosa to regulate its biofilm formation are mainly involved in arginine metabolism, glutamate production, magnesium transport, acetate metabolism, and the TCA cycle.

  8. An Integrated Modeling and Experimental Approach to Study the Influence of Environmental Nutrients on Biofilm Formation of Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Zhaobin Xu

    2015-01-01

    Full Text Available The availability of nutrient components in the environment was identified as a critical regulator of virulence and biofilm formation in Pseudomonas aeruginosa. This work proposes the first systems-biology approach to quantify microbial biofilm formation upon the change of nutrient availability in the environment. Specifically, the change of fluxes of metabolic reactions that were positively associated with P. aeruginosa biofilm formation was used to monitor the trend for P. aeruginosa to form a biofilm. The uptake rates of nutrient components were changed according to the change of the nutrient availability. We found that adding each of the eleven amino acids (Arg, Tyr, Phe, His, Iso, Orn, Pro, Glu, Leu, Val, and Asp to minimal medium promoted P. aeruginosa biofilm formation. Both modeling and experimental approaches were further developed to quantify P. aeruginosa biofilm formation for four different availability levels for each of the three ions that include ferrous ions, sulfate, and phosphate. The developed modeling approach correctly predicted the amount of biofilm formation. By comparing reaction flux change upon the change of nutrient concentrations, metabolic reactions used by P. aeruginosa to regulate its biofilm formation are mainly involved in arginine metabolism, glutamate production, magnesium transport, acetate metabolism, and the TCA cycle.

  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. 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.

  11. 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

    were used to visualize AHL-mediated communication in mixed biofilms, which were cultivated either in artificial flow chambers or in alginate beads in mouse lung tissue. In both model systems B. cepacia was capable of perceiving the AHL signals produced by P. aeruginosa, while the latter strain did......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...

  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. Potential of Ocimum basilicum L. and Salvia officinalis L. essential oils against biofilms of P. aeruginosa clinical isolates.

    Science.gov (United States)

    Stojanović-Radić, Z; Pejcić, M; Stojanović, N; Sharifi-Rad, J; Stanković, N

    2016-08-29

    Biofilms are complex communities of microorganisms, responsible for more than 60% of the chronic human infections and they represent one of the leading concerns in medicine. Pseudomonas aeruginosa is human pathogenic bacteria which causes numerous diseases and is known for its ability to produce biofilm. Ocimum basilicum L. (basil) and Salvia officinalis L. (sage) are widely used plants in traditional medicine for the treatment of different conditions. Therefore, the aim of this study was to investigate the potential of basil and sage essential oils against P. aeruginosa biofilm producing strains. The efficacy of two essential oils on P. aeruginosa biofilm forming ability was determined using crystal violet method. Out of 15 strains isolated from different clinical biological samples, two were strong, 11 moderate and one weak biofilm producer. Good efficacy of sage essential oil towards strong and weak biofilm producers, but not of basil essential oil, was observed. In the case of moderate biofilm producers, 81.8% showed lower biofilm production after incubation with the sage oil, while 63.6% showed the reduction of biofilm production after basil essential oil treatment. The obtained results showed high potential of both oils for the treatment of persistent infections caused by Pseudomonas aeruginosa biofilms.

  14. 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...

  15. Attenuation of Pseudomonas aeruginosa biofilm formation by Vitexin: A combinatorial study with azithromycin and gentamicin

    Science.gov (United States)

    Das, Manash C.; Sandhu, Padmani; Gupta, Priya; Rudrapaul, Prasenjit; de, Utpal C.; Tribedi, Prosun; Akhter, Yusuf; Bhattacharjee, Surajit

    2016-03-01

    Microbial biofilm are communities of surface-adhered cells enclosed in a matrix of extracellular polymeric substances. Extensive use of antibiotics to treat biofilm associated infections has led to the emergence of multiple drug resistant strains. Pseudomonas aeruginosa is recognised as a model biofilm forming pathogenic bacterium. Vitexin, a polyphenolic group of phytochemical with antimicrobial property, has been studied for its antibiofilm potential against Pseudomonas aeruginosa in combination with azithromycin and gentamicin. Vitexin shows minimum inhibitory concentration (MIC) at 260 μg/ml. It’s antibiofilm activity was evaluated by safranin staining, protein extraction, microscopy methods, quantification of EPS and in vivo models using several sub-MIC doses. Various quorum sensing (QS) mediated phenomenon such as swarming motility, azocasein degrading protease activity, pyoverdin and pyocyanin production, LasA and LasB activity of the bacteria were also evaluated. Results showed marked attenuation in biofilm formation and QS mediated phenotype of Pseudomonas aeruginosa in presence of 110 μg/ml vitexin in combination with azithromycin and gentamicin separately. Molecular docking of vitexin with QS associated LuxR, LasA, LasI and motility related proteins showed high and reasonable binding affinity respectively. The study explores the antibiofilm potential of vitexin against P. aeruginosa which can be used as a new antibiofilm agent against microbial biofilm associated pathogenesis.

  16. Characterization of temporal protein production in Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Southey-Pillig, Christopher J; Davies, David G; Sauer, Karin

    2005-12-01

    Phenotypic and genetic evidence supporting the notion of biofilm formation as a developmental process is growing. In the present work, we provide additional support for this hypothesis by identifying the onset of accumulation of biofilm-stage specific proteins during Pseudomonas aeruginosa biofilm maturation and by tracking the abundance of these proteins in planktonic and three biofilm developmental stages. The onset of protein production was found to correlate with the progression of biofilms in developmental stages. Protein identification revealed that proteins with similar function grouped within similar protein abundance patterns. Metabolic and housekeeping proteins were found to group within a pattern separate from virulence, antibiotic resistance, and quorum-sensing-related proteins. The latter were produced in a progressive manner, indicating that attendant features that are characteristic of biofilms such as antibiotic resistance and virulence may be part of the biofilm developmental process. Mutations in genes for selected proteins from several protein production patterns were made, and the impact of these mutations on biofilm development was evaluated. The proteins cytochrome c oxidase, a probable chemotaxis transducer, a two-component response regulator, and MexH were produced only in mature and late-stage biofilms. Mutations in the genes encoding these proteins did not confer defects in growth, initial attachment, early biofilm formation, or twitching motility but were observed to arrest biofilm development at the stage of cell cluster formation we call the maturation-1 stage. The results indicated that expression of theses genes was required for the progression of biofilms into three-dimensional structures on abiotic surfaces and the completion of the biofilm developmental cycle. Reverse transcription-PCR analysis confirmed the detectable change in expression of the respective genes ccoO, PA4101, and PA4208. We propose a possible mechanism for the

  17. Flagellum-Mediated Biofilm Defense Mechanisms of Pseudomonas aeruginosa against Host-Derived Lactoferrin ▿

    Science.gov (United States)

    Leid, Jeff G.; Kerr, Mathias; Selgado, Candice; Johnson, Chelsa; Moreno, Gabriel; Smith, Alyssa; Shirtliff, Mark E.; O'Toole, George A.; Cope, Emily K.

    2009-01-01

    Chronic infection with the gram-negative organism Pseudomonas aeruginosa is a leading cause of morbidity and mortality in human patients, despite high doses of antibiotics used to treat the various diseases this organism causes. These infections are chronic because P. aeruginosa readily forms biofilms, which are inherently resistant to antibiotics as well as the host's immune system. Our laboratory has been investigating specific mutations in P. aeruginosa that regulate biofilm bacterial susceptibility to the host. To continue our investigation of the role of genetics in bacterial biofilm host resistance, we examined P. aeruginosa biofilms that lack the flgK gene. This mutant lacks flagella, which results in defects in early biofilm development (up to 36 h). For these experiments, the flgK-disrupted strain and the parental strain (PA14) were used in a modified version of the 96-well plate microtiter assay. Biofilms were challenged with freshly isolated human leukocytes for 4 to 6 h and viable bacteria enumerated by CFU. Subsequent to the challenge, both mononuclear cells (monocytes and lymphocytes) and neutrophils, along with tumor necrosis factor alpha (TNF-α), were required for optimal killing of the flgK biofilm bacteria. We identified a cytokine cross talk network between mononuclear cells and neutrophils that was essential to the production of lactoferrin and bacterial killing. Our data suggest that TNF-α is secreted from mononuclear cells, causing neutrophil activation, resulting in the secretion of bactericidal concentrations of lactoferrin. These results extend previous studies of the importance of lactoferrin in the innate immune defense against bacterial biofilms. PMID:19651866

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

    Science.gov (United States)

    Maderova, Zdenka; Horska, Katerina; Kim, Sang-Ryoung; Lee, Chung-Hak; Pospiskova, Kristyna; Safarikova, Mirka; Safarik, Ivo

    2016-01-01

    The formation of bacterial biofilm on various surfaces has significant negative economic effects. The aim of this study was to find a simple procedure to decrease the Pseudomonas aeruginosa biofilm formation in a water environment by using different food waste biological materials as signal molecule adsorbents. The selected biomaterials did not reduce the cell growth but affected biofilm formation. Promising biomaterials were magnetically modified in order to simplify manipulation and facilitate their magnetic separation. The best biocomposite, magnetically modified spent grain, exhibited substantial adsorption of signal molecules and decreased the biofilm formation. These results suggest that selected food waste materials and their magnetically responsive derivatives could be applied to solve biofilm problems in water environment.

  19. The implication of Pseudomonas aeruginosa biofilms in infections

    DEFF Research Database (Denmark)

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

    2011-01-01

    of infection in the lungs of cystic fibrosis patients and in chronic wounds. In this review we address the molecular basis of biofilm development by P. aeruginosa as well as the mechanisms employed by this bacterium in the increased tolerance displayed against antimicrobials. The complex build......-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....... The immune response leading to this chronic inflammation is described. Finally, novel treatment strategies against P. aeruginosa are described including, quorum-sensing inhibition and induced biofilm-dispersion. The tolerance towards currently available antimicrobials calls for development of alternative...

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

    Science.gov (United States)

    de la Fuente-Núñez, César; Mansour, Sarah C; Wang, Zhejun; Jiang, Lucy; Breidenstein, Elena B M; Elliott, Melissa; Reffuveille, Fany; Speert, David P; Reckseidler-Zenteno, Shauna L; Shen, Ya; Haapasalo, Markus; Hancock, Robert E W

    2014-01-01

    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.

  1. 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.

  2. Influence of Melaleuca alternifolia oil nanoparticles on aspects of Pseudomonas aeruginosa biofilm.

    Science.gov (United States)

    Comin, Vanessa M; Lopes, Leonardo Q S; Quatrin, Priscilla M; de Souza, Márcia E; Bonez, Pauline C; Pintos, Francieli G; Raffin, Renata P; Vaucher, Rodrigo de A; Martinez, Diego S T; Santos, Roberto C V

    2016-04-01

    The Pseudomonas aeruginosa is a gram-negative bacillus and frequent cause of infection. This microorganism is resistant intrinsically to various drugs. The P. aeruginosa is associated with the biofilm formation, which causes worsen the prognosis and difficulty the treatment. The influence of Melaleuca alternifolia oil or "tree of tee" oil (TTO) and TTO nanoparticles on adhesion of P. aeruginosa in buccal epithelial cells was investigated. Also was determined the antimicrobial and antibiofilm activity against this microorganism. The TTO nanoparticles were produced by deposition of preformed polymer and the physic-chemical properties of nanoparticles were measured by electrophoresis and dynamic light scattering. The characterization of nanoparticle showed acceptable values for diameter and zeta potential. The evaluation of antimicrobial and antibiofilm activity against P. aeruginosa PAO1 was performed by microdilution indicating the minimal inhibitory concentration, and the potential antibiofilm. It was verified the action on virulence factors such the motility, besides the influence on adhesion in buccal epithelial cells. Both oil and nanoparticles showed a decrease in adhesion of microorganisms to buccal cells, decrease of biofilm and interfering on P. aeruginosa PAO1 motility. The nanostructuration of TTO, shows be a viable alternative against formed biofilm microorganisms.

  3. Inactivation of Pseudomonas aeruginosa biofilm by dense phase carbon dioxide.

    Science.gov (United States)

    Mun, Sungmin; Jeong, Jin-Seong; Kim, Jaeeun; Lee, Youn-Woo; Yoon, Jeyong

    2009-01-01

    Dense phase carbon dioxide (DPCD) is one of the most promising techniques available to control microorganisms as a non-thermal disinfection method. However, no study on the efficiency of biofilm disinfection using DPCD has been reported. The efficiency of DPCD in inactivating Pseudomonas aeruginosa biofilm, which is known to have high antimicrobial resistance, was thus investigated. P. aeruginosa biofilm, which was not immersed in water but was completely wet, was found to be more effectively inactivated by DPCD treatment, achieving a 6-log reduction within 7 min. The inactivation efficiency increased modestly with increasing pressure and temperature. This study also reports that the water-unimmersed condition is one of the most important operating parameters in achieving efficient biofilm control by DPCD treatment. In addition, observations by confocal laser scanning microscopy revealed that DPCD treatment not only inactivated biofilm cells on the glass coupons but also caused detachment of the biofilm following weakening of its structure as a result of the DPCD treatment; this is an added benefit of DPCD treatment.

  4. Distribution and Inhibition of Liposomes on Staphylococcus aureus and Pseudomonas aeruginosa Biofilm.

    Directory of Open Access Journals (Sweden)

    Dong Dong

    Full Text Available Staphylococcus aureus and Pseudomonas aeruginosa are major pathogens in chronic rhinosinusitis (CRS and their biofilms have been associated with poorer postsurgical outcomes. This study investigated the distribution and anti-biofilm effect of cationic (+ and anionic (- phospholipid liposomes with different sizes (unilamellar and multilamellar vesicle, ULV and MLV respectively on S. aureus and P. aeruginosa biofilms.Specific biofilm models for S. aureus ATCC 25923 and P. aeruginosa ATCC 15692 were established. Liposomal distribution was determined by observing SYTO9 stained biofilm exposed to DiI labeled liposomes using confocal scanning laser microscopy, followed by quantitative image analysis. The anti-biofilm efficacy study was carried out by using the alamarBlue assay to test the relative viability of biofilm treated with various liposomes for 24 hours and five minutes.The smaller ULVs penetrated better than larger MLVs in both S. aureus and P. aeruginosa biofilm. Except that +ULV and -ULV displayed similar distribution in S. aureus biofilm, the cationic liposomes adhered better than their anionic counterparts. Biofilm growth was inhibited at 24-hour and five-minute exposure time, although the decrease of viability for P. aeruginosa biofilm after liposomal treatment did not reach statistical significance.The distribution and anti-biofilm effects of cationic and anionic liposomes of different sizes differed in S. aureus and P. aeruginosa biofilms. Reducing the liposome size and formulating liposomes as positively charged enhanced the penetration and inhibition of S. aureus and P. aeruginosa biofilms.

  5. Activity of ozonated water and ozone against Staphylococcus aureus and Pseudomonas aeruginosa biofilms

    Science.gov (United States)

    Bialoszewski, Dariusz; Pietruczuk-Padzik, Anna; Kalicinska, Agnieszka; Bocian, Ewa; Czajkowska, Magdalena; Bukowska, Bozena; Tyski, Stefan

    2011-01-01

    Summary Background The known bactericidal properties of ozone have not been checked in relation to its action on bacterial biofilms. This is especially true of ozonated fluids. The aim of this study was to investigate the bactericidal activity of ozonated water and that of a mixture of ozone and oxygen against biofilms. Material/Methods Eighteen clinical strains of Staphylococcus aureus and Pseudomonas aeruginosa exhibiting various levels of antibiotic sensitivity were investigated. Bacteria were cultured in biofilm form on polystyrene titration plates for periods of 2 to 72 hours. The biofilms formed in this way were exposed to in statu nascendi ozonated water produced in a prototype device that had been tested in clinical conditions, or to a mixture of oxygen and ozone generated in the same device. Live cells in the biofilm were stained with a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) bromide solution. The degree of reduction of viable bacteria following ozone exposure was determined. Results Ozonated water was found to be an effective bactericidal agent against biofilms after as little as 30 seconds of exposure, while the bactericidal activity of the ozone-oxygen solution was much lower. Prolongation of the duration of biofilm exposure to the gaseous disinfectant to 40 minutes led to a reduction in the viable cell count, which nevertheless remained high. Conclusions Unlike the ozone-oxygen mixture, ozonated water effectively destroys bacterial biofilms in vitro. PMID:22037737

  6. 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.

  7. 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.

  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. 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 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...... with low iron concentrations (5 mu M FeCIA and decreased with increasing iron concentrations. Experiments involving cultivation of P. aeruginosa in a flow-chamber system suggested that a high level of iron (1100 mu M FeCl3) in the medium suppressed DNA release, structural biofilm development...

  10. Pseudomonas aeruginosa and Saccharomyces cerevisiae Biofilm in Flow Cells

    DEFF Research Database (Denmark)

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

    2011-01-01

    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). ...

  11. 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.

  12. Pseudomonas aeruginosa and the in vitro and in vivo biofilm mode of growth

    DEFF Research Database (Denmark)

    Høiby, N; Krogh Johansen, H; Moser, C

    2001-01-01

    The biofilm mode of growth is the survival strategy of environmental bacteria like Pseudomonas aeruginosa. Such P. aeruginosa biofilms also occur in the lungs of chronically infected cystic fibrosis patients, where they protect the bacteria against antibiotics and the immune response. The lung...

  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...... organisms do not possess comprehensive genetic programs for biofilm development. Instead the bacteria appear to have evolved a number of different mechanisms to optimize surface colonization, of which they express a subset in response to the prevailing environmental conditions. These mechanisms include...

  14. 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

    The development of resistance to beta-lactam antibiotics is a problem in the treatment of chronic Pseudomonas aeruginosa infection in the lungs of patients with cystic fibrosis. The main resistance mechanism is high-level expression of the chromosomally encoded AmpC beta-lactamase of P. aeruginosa...... cells growing in biofilms. Several genes have been shown to influence the level of ampC expression, but little is known about the regulation of ampC expression in P. aeruginosa biofilms. To study the expression of ampC in P. aeruginosa biofilms, we constructed a reporter that consisted of the fusion...... of the ampC promoter to gfp(ASV) encoding an unstable version of the green fluorescent protein. In vitro biofilms of P. aeruginosa were exposed to the beta-lactam antibiotics imipenem and ceftazidime. Sub-MICs of imipenem significantly induced the monitor system of the biofilm bacteria in the peripheries...

  15. Candida biofilm disrupting ability of di-rhamnolipid (RL-2) produced from Pseudomonas aeruginosa DSVP20.

    Science.gov (United States)

    Singh, Nivedita; Pemmaraju, Suma C; Pruthi, Parul A; Cameotra, Swaranjit S; Pruthi, Vikas

    2013-04-01

    Biosurfactant produced from Pseudomonas aeruginosa DSVP20 was evaluated for its potential to disrupt Candida albicans biofilm formed on polystyrene (PS) surfaces in this investigation. P. aeruginosa DSVP20 exhibited optimum production of biosurfactant (5.8 g L(-1)) after 96 h of growth with an ability to reduce surface tension of the aqueous solution from 72 to 28 mN m(-1). Analysis of purified biosurfactant with FT-IR, (1)H and (13)C NMR and MALDI-TOF MS revealed it to be di-rhamnolipid (RL-2) in nature. Biofilm disrupting ability of RL-2 (0.16 mg mL(-1)) on Candida cells when checked using XTT reduction assay revealed that about 50 % of the cells remain adhered to 96-well plate after 2 h of treatment, while up to 90 % reduction in pre-formed C. albicans biofilm on PS surface was observed with RL-2 (5.0 mg mL(-1)) in a dose-dependent manner. Microscopic analyses (SEM and CLSM) further confirm the influence of RL-2 on disruption of Candida biofilm extracellular matrix on PS surface which can be exploited as a potential alternative to the available conventional therapies.

  16. 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...

  17. The implication of Pseudomonas aeruginosa biofilms in infections

    DEFF Research Database (Denmark)

    Rybtke, Morten Theil; Jensen, Peter Ø; Høiby, Niels

    2011-01-01

    of infection in the lungs of cystic fibrosis patients and in chronic wounds. In this review we address the molecular basis of biofilm development by P. aeruginosa as well as the mechanisms employed by this bacterium in the increased tolerance displayed against antimicrobials. The complex build......-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...... 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....

  18. Phenotypes selected during chronic lung infection in cystic fibrosis patients: implications for the treatment of Pseudomonas aeruginosa biofilm infections.

    Science.gov (United States)

    Ciofu, Oana; Mandsberg, Lotte F; Wang, Hengzhuang; Høiby, Niels

    2012-07-01

    During chronic lung infection of patients with cystic fibrosis, Pseudomonas aeruginosa can survive for long periods of time under the challenging selective pressure imposed by the immune system and antibiotic treatment as a result of its biofilm mode of growth and adaptive evolution mediated by genetic variation. Mucoidy, hypermutability and acquirement of mutational antibiotic resistance are important adaptive phenotypes that are selected during chronic P. aeruginosa infection. This review dicsusses the role played by these phenotypes for the tolerance of biofilms to antibiotics and show that mucoidy and hypermutability change the architecture of in vitro formed biofilms and lead to increase tolerance to antibiotics. Production of high levels of beta-lactamase impairs penetration of beta-lactam antibiotics due to inactivation of the antibiotic. In conclusion, these data underline the importance of biofilm prevention strategies by early aggressive antibiotic prophylaxis or therapy before phenotypic diversification during chronic lung infection of patients with cystic fibrosis.

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

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    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.

  20. An examination of potential differences in biofilm production among different genotypes of Pseudomonas aeruginosa

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    Vasiljević Zorica

    2014-01-01

    Full Text Available In the present study, we have examined if there is any difference in biofilm production among different genotypes of Pseudomonas aeruginosa. The study investigated 526 non-duplicate P. aeruginosa isolated from clinical specimens and from a hospital environment. Isolates were grouped into thirty-five genotypes based on an identical ERIC2-band pattern. Biofilm formation was quantified by the microtiter plate test and all strains were classified into the following categories: no biofilm producers (0, weak (+, moderate (+, or strong (+++ biofilm producers. Only 2.45% of examined strains were not biofilm producers. Among biofilm producers, 39.26% were strong biofilm producers, 34.36% were moderate biofilm producers, while 23.93% were weak biofilm producers. Although the majority of strong biofilm producers were in genotype groups 2 and 3, the degree of in vitro biofilm formation in our study was not significantly affected by the genotype of Pseudomonas aeruginosa. In this study, we demonstrated that the degree of in vitro biofilm formation is not significantly affected by the genotype of Pseudomonas aeruginosa. [Projekat Ministarstva nauke Republike Srbije, br. 175039 i br. 143036

  1. 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

    , 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...

  2. Characterization of the Newly Isolated Lytic Bacteriophages KTN6 and KT28 and Their Efficacy against Pseudomonas aeruginosa Biofilm.

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    Katarzyna Danis-Wlodarczyk

    Full Text Available We here describe two novel lytic phages, KT28 and KTN6, infecting Pseudomonas aeruginosa, isolated from a sewage sample from an irrigated field near Wroclaw, in Poland. Both viruses show characteristic features of Pbunalikevirus genus within the Myoviridae family with respect to shape and size of head/tail, as well as LPS host receptor recognition. Genome analysis confirmed the similarity to other PB1-related phages, ranging between 48 and 96%. Pseudomonas phage KT28 has a genome size of 66,381 bp and KTN6 of 65,994 bp. The latent period, burst size, stability and host range was determined for both viruses under standard laboratory conditions. Biofilm eradication efficacy was tested on peg-lid plate assay and PET membrane surface. Significant reduction of colony forming units was observed (70-90% in 24 h to 72 h old Pseudomonas aeruginosa PAO1 biofilm cultures for both phages. Furthermore, a pyocyanin and pyoverdin reduction tests reveal that tested phages lowers the amount of both secreted dyes in 48-72 h old biofilms. Diffusion and goniometry experiments revealed the increase of diffusion rate through the biofilm matrix after phage application. These characteristics indicate these phages could be used to prevent Pseudomonas aeruginosa infections and biofilm formation. It was also shown, that PB1-related phage treatment of biofilm caused the emergence of stable phage-resistant mutants growing as small colony variants.

  3. Characterization of the Newly Isolated Lytic Bacteriophages KTN6 and KT28 and Their Efficacy against Pseudomonas aeruginosa Biofilm.

    Science.gov (United States)

    Danis-Wlodarczyk, Katarzyna; Olszak, Tomasz; Arabski, Michal; Wasik, Slawomir; Majkowska-Skrobek, Grazyna; Augustyniak, Daria; Gula, Grzegorz; Briers, Yves; Jang, Ho Bin; Vandenheuvel, Dieter; Duda, Katarzyna Anna; Lavigne, Rob; Drulis-Kawa, Zuzanna

    2015-01-01

    We here describe two novel lytic phages, KT28 and KTN6, infecting Pseudomonas aeruginosa, isolated from a sewage sample from an irrigated field near Wroclaw, in Poland. Both viruses show characteristic features of Pbunalikevirus genus within the Myoviridae family with respect to shape and size of head/tail, as well as LPS host receptor recognition. Genome analysis confirmed the similarity to other PB1-related phages, ranging between 48 and 96%. Pseudomonas phage KT28 has a genome size of 66,381 bp and KTN6 of 65,994 bp. The latent period, burst size, stability and host range was determined for both viruses under standard laboratory conditions. Biofilm eradication efficacy was tested on peg-lid plate assay and PET membrane surface. Significant reduction of colony forming units was observed (70-90%) in 24 h to 72 h old Pseudomonas aeruginosa PAO1 biofilm cultures for both phages. Furthermore, a pyocyanin and pyoverdin reduction tests reveal that tested phages lowers the amount of both secreted dyes in 48-72 h old biofilms. Diffusion and goniometry experiments revealed the increase of diffusion rate through the biofilm matrix after phage application. These characteristics indicate these phages could be used to prevent Pseudomonas aeruginosa infections and biofilm formation. It was also shown, that PB1-related phage treatment of biofilm caused the emergence of stable phage-resistant mutants growing as small colony variants.

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

    Science.gov (United States)

    Luján, Adela M; Maciá, María D; Yang, Liang; Molin, Søren; Oliver, Antonio; Smania, Andrea M

    2011-01-01

    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.

  5. 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.

  6. Susceptibility of Pseudomonas aeruginosa biofilm to alpha-helical peptides: D-enantiomer of LL-37

    Directory of Open Access Journals (Sweden)

    Scott N. Dean

    2011-07-01

    Full Text Available Pseudomonas aeruginosa is a highly versatile opportunistic pathogen and its ability to produce biofilms is a direct impediment to the healing of wounds and recovery from infection. Interest in anti-microbial peptides has grown due to their potential therapeutic applications and their possible use against antibiotic resistant bacteria. LL-37 is the only cathelicidin expressed by humans. In this study, we tested LL-37 and the effect of a protease-resistant LL-37 peptide mimetic, the peptide enantiomer D-LL-37, for anti-microbial and anti-biofilm activity against P. aeruginosa. Both forms of the peptide were equally effective as anti-microbial peptides with similar killing kinetics. CD spectra were obtained to demonstrate the chirality of D- and L-LL-37, and the trypsin resistance of D-LL-37 was confirmed. The helical cathelicidin from the cobra Naja atra (NA-CATH, and synthetic peptide variations (ATRA-1, ATRA-2, NA-CATH:ATRA1-ATRA1 were also tested. Although the cobra cathelicidin and related peptides had strong anti-microbial activity, those tested did not inhibit Pseudomonas biofilm formation, neither did control peptides. Both D- and L-LL-37 inhibited the attachment of Pseudomonas to a 96-well plate and decreased the amount of pre-formed (established biofilm. D-LL-37 is able to promote Pseudomonas motility and decrease biofilm formation by altering the rate of twitching as well as by downregulating the expression of the biofilm-related genes, rhlA and rhlB, similar to L-LL-37. Both L- and D-LL-37 protected Galleria mellonella in vivo against Pseudomonas infection, while NA-CATH:ATRA1-ATRA1 peptide did not. This study demonstrates the ability and equivalence of D-LL-37 compared to L-LL-37 to promote bacterial twitching motility and inhibit biofilm formation, and protect against in vivo infection, and suggests that this peptide could be a critical advancement in the development of new treatments for P. aeruginosa infection.

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

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    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.

  8. The urinary antibiotic 5-nitro-8-hydroxyquinoline (Nitroxoline) reduces the formation and induces the dispersal of Pseudomonas aeruginosa biofilms by chelation of iron and zinc.

    Science.gov (United States)

    Sobke, A; Klinger, M; Hermann, B; Sachse, S; Nietzsche, S; Makarewicz, O; Keller, P M; Pfister, W; Straube, E

    2012-11-01

    Since cations have been reported as essential regulators of biofilm, we investigated the potential of the broad-spectrum antimicrobial and cation-chelator nitroxoline as an antibiofilm agent. Biofilm mass synthesis was reduced by up to 80% at sub-MIC nitroxoline concentrations in Pseudomonas aeruginosa, and structures formed were reticulate rather than compact. In preformed biofilms, viable cell counts were reduced by 4 logs at therapeutic concentrations. Complexation of iron and zinc was demonstrated to underlie nitroxoline's potent antibiofilm activity.

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

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    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

  10. Sound waves effectively assist tobramycin in elimination of Pseudomonas aeruginosa biofilms in vitro.

    Science.gov (United States)

    Bandara, H M H N; Harb, A; Kolacny, D; Martins, P; Smyth, H D C

    2014-12-01

    Microbial biofilms are highly refractory to antimicrobials. The aim of this study was to investigate the use of low-frequency vibration therapy (20-20 kHz) on antibiotic-mediated Pseudomonas aeruginosa biofilm eradication. In screening studies, low-frequency vibrations were applied on model biofilm compositions to identify conditions in which surface standing waves were observed. Alginate surface tension and viscosity were also measured. The effect of vibration on P. aeruginosa biofilms was studied using a standard biofilm assay. Subminimal inhibitory concentrations (sub-MIC) of tobramycin (5 μg/ml) were added to biofilms 3 h prior, during, and immediately after vibration and quantitatively assessed by (2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide) reduction assay (XTT) and, qualitatively, by confocal laser scanning microscopy (CLSM). The standing waves occurred at frequencies vibrated without sub-MIC tobramycin showed a significantly reduced metabolism compared to untreated controls (p vibrated simultaneously (450, 530, 610, and 650 Hz), or vibrated (450 and 650 Hz) then treated with tobramycin subsequently, or vibrated (610 Hz, 650 Hz) after 3 h of tobramycin treatment showed significantly lower metabolism compared to P. aeruginosa biofilm treated with tobramycin alone (p vibrations assisted tobramycin in killing P. aeruginosa biofilms at sub-MIC. Thus, sound waves together with antibiotics are a promising approach in eliminating pathogenic biofilms.

  11. 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.

  12. 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 planktonic cultures were challenged with P. aeruginosa supernatant cultures overnight. Results indicated that quorum-sensing-controlled factors from P. aeruginosa supernatant inhibited S. epidermidis growth in planktonic cultures. We also found that P. aeruginosa extracellular products, mainly...... in overnight cultures had no effect on established P. aeruginosa biofilms and planktonic growth. These findings reveal that P. aeruginosa extracellular products are important microbial competition factors that overcome competition with S. epidermidis, and the results may provide clues for the development...

  13. Pseudomonas aeruginosa biofilm infections in cystic fibrosis: insights into pathogenic processes and treatment strategies

    DEFF Research Database (Denmark)

    Hassett, Daniel J; Korfhagen, Thomas R; Irvin, Randall T;

    2010-01-01

    CF airway mucus can be infected by opportunistic microorganisms, notably Pseudomonas aeruginosa. Once organisms are established as biofilms, even the most potent antibiotics have little effect on their viability, especially during late-stage chronic infections. Better understanding of the mechani...... of the mechanisms used by P. aeruginosa to circumvent host defenses and therapeutic intervention strategies is critical for advancing novel treatment strategies....

  14. Synergistic Activities of an Efflux Pump Inhibitor and Iron Chelators against Pseudomonas aeruginosa Growth and Biofilm Formation

    DEFF Research Database (Denmark)

    Liu, Yang; Yang, Liang; Molin, Søren

    2010-01-01

    The efflux pump inhibitor phenyl-arginine-beta-naphthylamide (PA beta N) was paired with iron chelators 2,2'-dipyridyl, acetohydroxamic acid, and EDTA to assess synergistic activities against Pseudomonas aeruginosa growth and biofilm formation. All of the tested iron chelators synergistically...... inhibited P. aeruginosa growth and biofilm formation with PA beta N. PA beta N-EDTA showed the most promising activity against P. aeruginosa growth and biofilm formation....

  15. 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...

  16. 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...

  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

    of O2-depleted biofilms may thus improve susceptibility to ciprofloxacin possibly by restoring aerobic respiration. We tested such a strategy using reoxygenation of O2-depleted P. aeruginosa strain PAO1 agarose-embedded biofilms by hyperbaric oxygen treatment (HBOT) (100% O2, 2.8bar), enhancing...

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

    DEFF Research Database (Denmark)

    Mette, Kolpen; 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 wildtype PAO1, a catalase deficient mutant (ΔkatA) and a colistin resistant CF isolate cultured in microtiter plates...

  19. Pseudomonas aeruginosa biofilm-associated homoserine lactone C12 rapidly activates apoptosis in airway epithelia.

    Science.gov (United States)

    Schwarzer, Christian; Fu, Zhu; Patanwala, Maria; Hum, Lauren; Lopez-Guzman, Mirielle; Illek, Beate; Kong, Weidong; Lynch, Susan V; Machen, Terry E

    2012-05-01

    Pseudomonas aeruginosa (PA) forms biofilms in lungs of cystic fibrosis (CF) patients, a process regulated by quorum-sensing molecules including N-(3-oxododecanoyl)-l-homoserine lactone (C12). C12 (10-100 µM) rapidly triggered events commonly associated with the intrinsic apoptotic pathway in JME (CF ΔF508CFTR, nasal surface) epithelial cells: depolarization of mitochondrial (mito) membrane potential (Δψ(mito)) and release of cytochrome C (cytoC) from mitos into cytosol and activation of caspases 3/7, 8 and 9. C12 also had novel effects on the endoplasmic reticulum (release of both Ca(2+) and ER-targeted GFP and oxidized contents into the cytosol). Effects began within 5 min and were complete in 1-2 h. C12 caused similar activation of caspases and release of cytoC from mitos in Calu-3 (wtCFTR, bronchial gland) cells, showing that C12-triggered responses occurred similarly in different airway epithelial types. C12 had nearly identical effects on three key aspects of the apoptosis response (caspase 3/7, depolarization of Δψ(mito) and reduction of redox potential in the ER) in JME and CFTR-corrected JME cells (adenoviral expression), showing that CFTR was likely not an important regulator of C12-triggered apoptosis in airway epithelia. Exposure of airway cultures to biofilms from PAO1wt caused depolarization of Δψ(mito) and increases in Ca(cyto) like 10-50 µM C12. In contrast, biofilms from PAO1ΔlasI (C12 deficient) had no effect, suggesting that C12 from P. aeruginosa biofilms may contribute to accumulation of apoptotic cells that cannot be cleared from CF lungs. A model to explain the effects of C12 is proposed.

  20. 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

    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...... significantly lower after 7 days in animals receiving antibiotic combination than in animals receiving single antibiotics. In patients with cystic fibrosis, inhaled colistin-tobramycin was well tolerated and resulted in a mean decrease of 2.52 +/- 2.5 cfu of P. aeruginosa per milliliter of sputum (P = .027...

  1. 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

    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...... 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...

  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. Characterization of biofilm formed on intrauterine devices

    Directory of Open Access Journals (Sweden)

    Pruthi V

    2003-01-01

    Full Text Available PURPOSE: Intrauterine device (IUD is one of the most convenient contraceptive procedures used by women of Asian and African countries. Previous surveys have revealed that 75% of the IUDs recovered from patients suffering from reproductive tract infections (RTIs were covered with a consortium of microbes. This study was designed to characterize these microbes and recommend remedial measures. METHODS: Quantitative measurement of biofilm formation was assessed by a microtitre plate assay on 86 samples of microorganisms dislodged from IUDs of patients with RTIs. Susceptibility of biofilm to various antimicrobial agents was also quantified. Scanning electron microscopy (SEM was used to scrutinize the microorganisms adherent to IUDs. RESULTS: The organisms associated with IUDs were predominantly composed of Staphylococcus aureus (16%, Staphylococcus epidermidis (18%, Pseudomonas aeruginosa (5%, Escherichia coli (27%, Neisseria gonorrhoeae (2%, Candida albicans (20% and Candida dubliniesis (12%. SEM studies indicated that these organisms were organized into biofilms. Studies on the in vitro adherence pattern by crystal violet staining on 96 well microtitre plates revealed that the biofilms were stably established after 60 hours. These biofilms are resistant to an array of antibiotics tested. CONCLUSION: Biofilm formation may be one of the major causes for persistent infection and antibiotic resistance in IUD users.

  4. Flagellin FliC Phosphorylation Affects Type 2 Protease Secretion and Biofilm Dispersal in Pseudomonas aeruginosa PAO1

    Science.gov (United States)

    Suriyanarayanan, Tanujaa; Periasamy, Saravanan; Lin, Miao-Hsia; Ishihama, Yasushi; Swarup, Sanjay

    2016-01-01

    Protein phosphorylation has a major role in controlling the life-cycle and infection stages of bacteria. Proteome-wide occurrence of S/T/Y phosphorylation has been reported for many prokaryotic systems. Previously, we reported the phosphoproteome of Pseudomonas aeruginosa and Pseudomonas putida. In this study, we show the role of S/T phosphorylation of one motility protein, FliC, in regulating multiple surface-associated phenomena of P. aeruginosa PAO1. This is the first report of occurrence of phosphorylation in the flagellar protein, flagellin FliC in its highly conserved N-terminal NDO domain across several Gram negative bacteria. This phosphorylation is likely a well-regulated phenomenon as it is growth phase dependent in planktonic cells. The absence of phosphorylation in the conserved T27 and S28 residues of FliC, interestingly, did not affect swimming motility, but affected the secretome of type 2 secretion system (T2SS) and biofilm formation of PAO1. FliC phosphomutants had increased levels and activities of type 2 secretome proteins. The secretion efficiency of T2SS machinery is associated with flagellin phosphorylation. FliC phosphomutants also formed reduced biofilms at 24 h under static conditions and had delayed biofilm dispersal under dynamic flow conditions, respectively. The levels of type 2 secretome and biofilm formation under static conditions had an inverse correlation. Hence, increase in type 2 secretome levels was accompanied by reduced biofilm formation in the FliC phosphomutants. As T2SS is involved in nutrient acquisition and biofilm dispersal during survival and spread of P. aeruginosa, we propose that FliC phosphorylation has a role in ecological adaptation of this opportunistic environmental pathogen. Altogether, we found a system of phosphorylation that affects key surface related processes such as proteases secretion by T2SS, biofilm formation and dispersal. PMID:27701473

  5. Pseudomonas aeruginosa produces aspirin insensitive eicosanoids and contributes to the eicosanoid profile of polymicrobial biofilms with Candida albicans.

    Science.gov (United States)

    Fourie, Ruan; Ells, Ruan; Kemp, Gabré; Sebolai, Olihile M; Albertyn, Jacobus; Pohl, Carolina H

    2017-02-01

    The interaction of clinically relevant microorganisms is the focus of various studies, e.g. the interaction between the pathogenic yeast, Candida albicans, and the bacterium, Pseudomonas aeruginosa. During infection both release arachidonic acid, which they can transform into eicosanoids. This study evaluated the production of prostaglandin E2, prostaglandin F2α and 15-hydroxyeicosatetraenoic acid by biofilms of P. aeruginosa and C. albicans. The influence of co-incubation, acetylsalicylic acid and nordihydroguaiaretic acid on biofilm formation and eicosanoid production was evaluated. Acetylsalicylic acid decreased colony forming units of P. aeruginosa, but increased metabolic activity and eicosanoid production of the cells. In contrast to prostaglandin E2, prostaglandin F2a production by C. albicans was insensitive to acetylsalicylic acid, indicating that different enzymes are responsible for their production in this yeast. Nordihydroguaiaretic acid inhibited biofilm formation by P. aeruginosa, however co-incubation provided protection against this inhibitor. Production of these eicosanoids could affect pathogen-clearance and infection dynamics and this previously uncharacterized facet of interaction could facilitate novel therapeutic intervention against polymicrobial infection.

  6. Phenolic compounds affect production of pyocyanin, swarming motility and biofilm formation of Pseudomonas aeruginosa

    Institute of Scientific and Technical Information of China (English)

    Aylin Ugurlu; Aysegul Karahasan Yagci; Seyhan Ulusoy; Burak Aksu; Gulgun Bosgelmez-Tinaz

    2016-01-01

    Objective: To investigate the effects of plant-derived phenolic compounds (i.e. caffeic acid, cinnamic acid, ferulic acid and vanillic acid) on the production of quorum sensing regulated virulence factors such as pyocyanin, biofilm formation and swarming motility of Pseudomonas aeruginosa (P. aeruginosa) isolates. Methods: Fourteen clinical P. aeruginosa isolates obtained from urine samples and P. aeruginosa PA01 strain were included in the study. The antibacterial effects of phenolic compounds were screened by well diffusion assay. Pyocyanin and biofilm ac-tivity were measured from culture supernatants and the absorbance values were measured using a spectrophotometer. Swarming plates supplemented with phenolic acids were point inoculated with P. aeruginosa strains and the ability to swarm was determined by measuring the distance of swarming from the central inoculation site. Results: Tested phenolic compounds reduced the production of pyocyanin and biofilm formation without affecting growth compared to untreated cultures. Moreover, these compounds blocked about 50% of biofilm production and swarming motility in P. aeruginosa isolates. Conclusions: We may suggest that if swarming and consecutive biofilm formation could be inhibited by the natural products as shown in our study, the bacteria could not attach to the surfaces and produce chronic infections. Antimicrobials and natural products could be combined and the dosage of antimicrobials could be reduced to overcome antimicrobial resistance and drug side effects.

  7. Phenolic compounds affect production of pyocyanin, swarming motility and biofilm formation of Pseudomonas aeruginosa

    Institute of Scientific and Technical Information of China (English)

    Aylin Ugurlu; Aysegul Karahasan Yagci; Seyhan Ulusoy; Burak Aksu; Gulgun Bosgelmez-Tinaz

    2016-01-01

    Objective: To investigate the effects of plant-derived phenolic compounds(i.e. caffeic acid, cinnamic acid, ferulic acid and vanillic acid) on the production of quorum sensing regulated virulence factors such as pyocyanin, biofilm formation and swarming motility of Pseudomonas aeruginosa(P. aeruginosa) isolates.Methods: Fourteen clinical P. aeruginosa isolates obtained from urine samples and P. aeruginosa PA01 strain were included in the study. The antibacterial effects of phenolic compounds were screened by well diffusion assay. Pyocyanin and biofilm activity were measured from culture supernatants and the absorbance values were measured using a spectrophotometer. Swarming plates supplemented with phenolic acids were point inoculated with P. aeruginosa strains and the ability to swarm was determined by measuring the distance of swarming from the central inoculation site.Results: Tested phenolic compounds reduced the production of pyocyanin and biofilm formation without affecting growth compared to untreated cultures. Moreover, these compounds blocked about 50% of biofilm production and swarming motility in P. aeruginosa isolates.Conclusions: We may suggest that if swarming and consecutive biofilm formation could be inhibited by the natural products as shown in our study, the bacteria could not attach to the surfaces and produce chronic infections. Antimicrobials and natural products could be combined and the dosage of antimicrobials could be reduced to overcome antimicrobial resistance and drug side effects.

  8. 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

    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......, 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...

  9. 医院获得性肺炎铜绿假单胞菌生物被膜形成能力与基因型分析%Biofilm-forming ability and genotypes in Pseudomonas aeruginosa causing hospital-acquired pneumonia

    Institute of Scientific and Technical Information of China (English)

    陈列彬; 郭雪君; 陈峰

    2012-01-01

    OBJECTIVE To explore the biofilm-forming ability and genotypes of Pseudomonas aeruginosa causing nosocomial pneumonia. METHODS Forty-two P. aeruginosa strains isolated from nosocomial pneumonia were tested for the biofilm-forming ability using a modified microtiter test. Enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) was used to analyze their genotypes. RESULTS The best time to culture the biofilm of P. aeruginosa isolates in nosocomial pneumonia was 5~6 days. Absorbance value A57O of the biofilm of these strains ranged from —0. 114 to 2. 822 and film area ratio was from 0. 0449 to 0. 9065. Most of these isolates demonstrated different biofilm-forming ability by absorbance and film area ratio test. Among these isolates tested, thirteen clusters(A,B,C, D,E, F,G, II and so on) showed different genotypes in fingerprints by ERIC-PCR. CONCLUSION P. aeruginosa varies in the ability of forming biofilm and shows polymorphism in the genotype, those isolates with the genotypes of A, D, E ,F, or H have stronger ability in forming most of the biofilm.%目的 研究医院获得性肺炎铜绿假单胞菌的生物被膜形成能力及其基因型.方法 用微量平板法对临床分离42株医院获得性肺炎铜绿假单胞菌建立生物被膜模型并测定其形成能力,通过肠杆菌科细菌基因间重复一致序列-PCR(ERIC-PCR)方法进行基因分型.结果 医院获得性肺炎铜绿假单胞菌BF最适培养时间为5~6 d,吸光度值A570为-0.114~2.822,成膜面积比为0.0449~0.9065,绝大多数菌株具有生物被膜形成能力且形成能力不同,用ERIC-PCR法可分为A、B、C、D、E、F、G、H等13个基因型.结论 铜绿假单胞菌生物被膜形成能力不同,在基因型上表现出多态性,基因型为A、D、E、F、H型的菌株大多生物被膜形成能力较强.

  10. Beneficial biofilms in marine aquaculture? Linking points of biofilm formation mechanisms in Pseudomonas aeruginosa and Pseudoalteromonas species

    Directory of Open Access Journals (Sweden)

    Wiebke Wesseling

    2015-07-01

    Full Text Available For marine aquaculture it is suggested that a specific substrate coated with a beneficial biofilm could prevent fish egg clutches from pathogenic infestations and improve the water quality and health of adult fish while, at the same time, minimising the need for the application of antibiotics. In marine biotopes, the habitat of Pseudoalteromonas species (a strain with suggested beneficial properties, biofilms are mostly discussed in the context of fouling processes. Hence research focuses on unravelling the mechanisms of biofilm formation aiming to prevent formation or to destroy existing biofilms. Initially in this review, particular components of biofilm formation in Pseudomonas aeruginosa, a gram-negative model organism that is responsible for nosocomial infections and considered as a food spoiling agent, are described (extracellular appendages, role of matrix components, cell-cell signalling to get an advanced understanding of biofilm formation. The aim of this treatise is to seek linking points for biofilm formation of P. aeruginosa and Pseudoalteromonas sp., respectively. Furthermore, approaches are discussed for how biofilm formation can be realized to improve fish (larvae rearing by species of the genus Pseudoalteromonas.

  11. 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...... 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...

  12. 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

    ). Gfp-based reporter technology has been applied for non-destructive, single-cell-level detection of quorum sensing in laboratory-based P. aeruginosa biofilms. It is reported that a synthetic halogenated furanone compound, which is a derivative of the secondary metabolites produced by the Australian...... of important virulence factors, indicating a general effect on target genes of the las quorum sensing circuit. The furanone was applied to P. aeruginosa biofilms established in biofilm flow chambers. The Gfp-based analysis reveals that the compound penetrates microcolonies and blocks cell signalling and quorum...... sensing in most biofilm cells. The compound did not affect initial attachment to the abiotic substratum. It does, however, affect the architecture of the biofilm and enhances the process of bacterial detachment, leading to a loss of bacterial biomass from the substratum....

  13. Nanoparticle (star polymer) delivery of nitric oxide effectively negates Pseudomonas aeruginosa biofilm formation.

    Science.gov (United States)

    Duong, Hien T T; Jung, Kenward; Kutty, Samuel K; Agustina, Sri; Adnan, Nik Nik M; Basuki, Johan S; Kumar, Naresh; Davis, Thomas P; Barraud, Nicolas; Boyer, Cyrille

    2014-07-14

    Biofilms are increasingly recognized as playing a major role in human infectious diseases, as they can form on both living tissues and abiotic surfaces, with serious implications for applications that rely on prolonged exposure to the body such as implantable biomedical devices or catheters. Therefore, there is an urgent need to develop improved therapeutics to effectively eradicate unwanted biofilms. Recently, the biological signaling molecule nitric oxide (NO) was identified as a key regulator of dispersal events in biofilms. In this paper, we report a new class of core cross-linked star polymers designed to store and release nitric oxide, in a controlled way, for the dispersion of biofilms. First, core cross-linked star polymers were prepared by reversible addition-fragmentation chain transfer polymerization (RAFT) via an arm first approach. Poly(oligoethylene methoxy acrylate) chains were synthesized by RAFT polymerization, and then chain extended in the presence of 2-vinyl-4,4-dimethyl-5-oxazolone monomer (VDM) with N,N-methylenebis(acrylamide) employed as a cross-linker to yield functional core cross-linked star polymers. Spermine was successfully attached to the star core by reaction with VDM. Finally, the secondary amine groups were reacted with NO gas to yield NO-core cross-linked star polymers. The core cross-linked star polymers were found to release NO in a controlled, slow delivery in bacterial cultures showing great efficacy in preventing both cell attachment and biofilm formation in Pseudomonas aeruginosa over time via a nontoxic mechanism, confining bacterial growth to the suspended liquid.

  14. Extracellular DNA Acidifies Biofilms and Induces Aminoglycoside Resistance in Pseudomonas aeruginosa.

    Science.gov (United States)

    Wilton, Mike; Charron-Mazenod, Laetitia; Moore, Richard; Lewenza, Shawn

    2015-11-09

    Biofilms consist of surface-adhered bacterial communities encased in an extracellular matrix composed of DNA, exopolysaccharides, and proteins. Extracellular DNA (eDNA) has a structural role in the formation of biofilms, can bind and shield biofilms from aminoglycosides, and induces antimicrobial peptide resistance mechanisms. Here, we provide evidence that eDNA is responsible for the acidification of Pseudomonas aeruginosa planktonic cultures and biofilms. Further, we show that acidic pH and acidification via eDNA constitute a signal that is perceived by P. aeruginosa to induce the expression of genes regulated by the PhoPQ and PmrAB two-component regulatory systems. Planktonic P. aeruginosa cultured in exogenous 0.2% DNA or under acidic conditions demonstrates a 2- to 8-fold increase in aminoglycoside resistance. This resistance phenotype requires the aminoarabinose modification of lipid A and the production of spermidine on the bacterial outer membrane, which likely reduce the entry of aminoglycosides. Interestingly, the additions of the basic amino acid L-arginine and sodium bicarbonate neutralize the pH and restore P. aeruginosa susceptibility to aminoglycosides, even in the presence of eDNA. These data illustrate that the accumulation of eDNA in biofilms and infection sites can acidify the local environment and that acidic pH promotes the P. aeruginosa antibiotic resistance phenotype.

  15. Pseudomonas aeruginosa bacteriophage PA1Ø requires type IV pili for infection and shows broad bactericidal and biofilm removal activities.

    Science.gov (United States)

    Kim, Shukho; Rahman, Marzia; Seol, Sung Yong; Yoon, Sang Sun; Kim, Jungmin

    2012-09-01

    We isolated a new lytic Pseudomonas aeruginosa phage that requires type IV pili for infection. PA1Ø has a broad bactericidal spectrum, covering Gram-positive and Gram-negative bacteria, and can eradicate biofilm cells. PA1Ø may be developed as a therapeutic agent for biofilm-related mixed infections with P. aeruginosa and Staphylococcus aureus.

  16. Dynamics of Mutator and Antibiotic-Resistant Populations in a Pharmacokinetic/Pharmacodynamic Model of Pseudomonas aeruginosa Biofilm Treatment

    DEFF Research Database (Denmark)

    Macià, María D.; Pérez, José L.; Molin, Søren;

    2011-01-01

    Biofilm growth, antibiotic resistance, and mutator phenotypes are key components of chronic respiratory infections by Pseudomonas aeruginosa in cystic fibrosis patients. We examined the dynamics of mutator and antibiotic-resistant populations in P. aeruginosa flow-cell biofilms, using fluorescent...

  17. Nitroxides as anti-biofilm compounds for the treatment of Pseudomonas aeruginosa and mixed-culture biofilms.

    Science.gov (United States)

    Alexander, Stefanie-Ann; Kyi, Caroline; Schiesser, Carl H

    2015-04-28

    A series of 23 nitroxides () was tested for biofilm modulatory activity using a crystal violet staining technique. 3-(Dodecane-1-thiyl)-4-(hydroxymethyl)-2,2,5,5-tetramethyl-1-pyrrolinoxyl () was found to significantly suppress biofilm formation and elicit dispersal events in both Pseudomonas aeruginosa and mixed-culture biofilms. Twitching and swarming motilities were enhanced by nitroxide , leaving the planktonic-specific swimming motility unaffected and suggesting that the mechanism of -mediated biofilm modulation is linked to the hyperactivation of surface-associated cell motilities. Preliminary structure-activity relationship studies identify the dodecanethiyl chain, hydroxymethyl substituent and the free radical moiety to be structural features pertinent to the anti-biofilm activity of .

  18. 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.

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

    Science.gov (United States)

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

    2016-08-01

    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.

  20. Influence of clove oil on certain quorum-sensing-regulated functions and biofilm of Pseudomonas aeruginosa and Aeromonas hydrophila

    Indian Academy of Sciences (India)

    Fohad Mabood Husain; Iqbal Ahmad; Mohammad Asif; Qudsia Tahseen

    2013-12-01

    Quorum sensing (QS) plays an important role in virulence, biofilm formation and survival of many pathogenic bacteria including Pseudomonas aeruginosa. This signalling pathway is considered as novel and promising target for anti-infective agents. In the present investigation, effect of the Sub-MICs of clove oil on QS regulated virulence factors and biofilm formation was evaluated against P. aeruginosa PAO1 and Aeromonas hydrophila WAF-38 strain. Sub-inhibitory concentrations of the clove oil demonstrated statistically significant reduction of las- and rhl-regulated virulence factors such as LasB, total protease, chitinase and pyocyanin production, swimming motility and exopolysaccharide production. The biofilm forming capability of PAO1 and A. hydrophila WAF-38 was also reduced in a concentration-dependent manner at all tested sub-MIC values. Further, the PAO1-preinfected Caenorhabditis elegans displayed an enhanced survival when treated with 1.6% v/v of clove oil. The above findings highlight the promising anti-QS-dependent therapeutic function of clove oil against P. aeruginosa.

  1. Quantification of Pseudomonas aeruginosa in multispecies biofilms using PMA-qPCR

    Directory of Open Access Journals (Sweden)

    Sarah Tavernier

    2015-02-01

    Full Text Available Multispecies biofilms are an important healthcare problem and may lead to persistent infections. These infections are difficult to treat, as cells in a biofilm are highly resistant to antimicrobial agents. While increasingly being recognized as important, the properties of multispecies biofilms remain poorly studied. In order to do so, the quantification of the individual species is needed. The current cultivation-based approaches can lead to an underestimation of the actual cell number and are time-consuming. In the present study we set up a culture-independent approach based on propidium monoazide qPCR (PMA-qPCR to quantify Pseudomonas aeruginosa in a multispecies biofilm. As a proof of concept, we explored the influence of the combined presence of Staphylococcus aureus, Streptococcus anginosus and Burkholderia cenocepacia on the antimicrobial susceptibility of P. aeruginosa using this PMA-qPCR approach.

  2. 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.

  3. 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.

  4. 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.

  5. Plasma-mediated inactivation of Pseudomonas aeruginosa biofilms grown on borosilicate surfaces under continuous culture system.

    Science.gov (United States)

    Vandervoort, Kurt G; Brelles-Mariño, Graciela

    2014-01-01

    Biofilms are microbial communities attached to a surface and embedded in a matrix composed of exopolysaccharides and excreted nucleic acids. Bacterial biofilms are responsible for undesirable effects such as disease, prostheses colonization, biofouling, equipment damage, and pipe plugging. Biofilms are also more resilient than free-living cells to regular sterilization methods and therefore it is indispensable to develop better ways to control and remove them. The use of gas discharge plasmas is a good alternative since plasmas contain a mixture of reactive agents well-known for their decontamination potential against free microorganisms. We have previously reported that Pseudomonas aeruginosa biofilms were inactivated after a 1-min plasma exposure. We determined that the adhesiveness and the thickness of Pseudomonas biofilms grown on borosilicate were reduced. We also reported sequential morphological changes and loss of viability upon plasma treatment. However, the studies were carried out in batch cultures. The use of a continuous culture results in a more homogenous environment ensuring reproducible biofilm growth. The aim of this work was to study plasma-mediated inactivation of P. aeruginosa biofilms grown on borosilicate in a continuous culture system. In this paper we show that biofilms grown on glass under continuous culture can be inactivated by using gas discharge plasma. Both biofilm architecture and cell culturability are impacted by the plasma treatment. The inactivation kinetics is similar to previously described ones and cells go through sequential changes ranging from minimal modification without loss of viability at short plasma exposure times, to major structure and viability loss at longer exposure times. We report that changes in biofilm structure leading to the loss of culturability and viability are related to a decrease of the biofilm matrix adhesiveness. To our knowledge, there has been no attempt to evaluate the inactivation

  6. Plasma-mediated inactivation of Pseudomonas aeruginosa biofilms grown on borosilicate surfaces under continuous culture system.

    Directory of Open Access Journals (Sweden)

    Kurt G Vandervoort

    Full Text Available Biofilms are microbial communities attached to a surface and embedded in a matrix composed of exopolysaccharides and excreted nucleic acids. Bacterial biofilms are responsible for undesirable effects such as disease, prostheses colonization, biofouling, equipment damage, and pipe plugging. Biofilms are also more resilient than free-living cells to regular sterilization methods and therefore it is indispensable to develop better ways to control and remove them. The use of gas discharge plasmas is a good alternative since plasmas contain a mixture of reactive agents well-known for their decontamination potential against free microorganisms. We have previously reported that Pseudomonas aeruginosa biofilms were inactivated after a 1-min plasma exposure. We determined that the adhesiveness and the thickness of Pseudomonas biofilms grown on borosilicate were reduced. We also reported sequential morphological changes and loss of viability upon plasma treatment. However, the studies were carried out in batch cultures. The use of a continuous culture results in a more homogenous environment ensuring reproducible biofilm growth. The aim of this work was to study plasma-mediated inactivation of P. aeruginosa biofilms grown on borosilicate in a continuous culture system. In this paper we show that biofilms grown on glass under continuous culture can be inactivated by using gas discharge plasma. Both biofilm architecture and cell culturability are impacted by the plasma treatment. The inactivation kinetics is similar to previously described ones and cells go through sequential changes ranging from minimal modification without loss of viability at short plasma exposure times, to major structure and viability loss at longer exposure times. We report that changes in biofilm structure leading to the loss of culturability and viability are related to a decrease of the biofilm matrix adhesiveness. To our knowledge, there has been no attempt to evaluate the

  7. Pseudomonas aeruginosa Biofilm Response and Resistance to Cold Atmospheric Pressure Plasma Is Linked to the Redox-Active Molecule Phenazine.

    Science.gov (United States)

    Mai-Prochnow, Anne; Bradbury, Mark; Ostrikov, Kostya; Murphy, Anthony B

    2015-01-01

    Pseudomonas aeruginosa is an important opportunistic pathogen displaying high antibiotic resistance. Its resistance is in part due to its outstanding ability to form biofilms on a range of biotic and abiotic surfaces leading to difficult-to-treat, often long-term infections. Cold atmospheric plasma (CAP) is a new, promising antibacterial treatment to combat antibiotic-resistant bacteria. Plasma is ionized gas that has antibacterial properties through the generation of a mix of reactive oxygen and nitrogen species (RONS), excited molecules, charged particles and UV photons. Our results show the efficient removal of P. aeruginosa biofilms using a plasma jet (kINPen med), with no viable cells detected after 5 min treatment and no attached biofilm cells visible with confocal microscopy after 10 min plasma treatment. Because of its multi-factorial action, it is widely presumed that the development of bacterial resistance to plasma is unlikely. However, our results indicate that a short plasma treatment (3 min) may lead to the emergence of a small number of surviving cells exhibiting enhanced resistance to subsequent plasma exposure. Interestingly, these cells also exhibited a higher degree of resistance to hydrogen peroxide. Whole genome comparison between surviving cells and control cells revealed 10 distinct polymorphic regions, including four belonging to the redox active, antibiotic pigment phenazine. Subsequently, the interaction between phenazine production and CAP resistance was demonstrated in biofilms of transposon mutants disrupted in different phenazine pathway genes which exhibited significantly altered sensitivity to CAP.

  8. Pseudomonas aeruginosa Biofilm Response and Resistance to Cold Atmospheric Pressure Plasma Is Linked to the Redox-Active Molecule Phenazine.

    Directory of Open Access Journals (Sweden)

    Anne Mai-Prochnow

    Full Text Available Pseudomonas aeruginosa is an important opportunistic pathogen displaying high antibiotic resistance. Its resistance is in part due to its outstanding ability to form biofilms on a range of biotic and abiotic surfaces leading to difficult-to-treat, often long-term infections. Cold atmospheric plasma (CAP is a new, promising antibacterial treatment to combat antibiotic-resistant bacteria. Plasma is ionized gas that has antibacterial properties through the generation of a mix of reactive oxygen and nitrogen species (RONS, excited molecules, charged particles and UV photons. Our results show the efficient removal of P. aeruginosa biofilms using a plasma jet (kINPen med, with no viable cells detected after 5 min treatment and no attached biofilm cells visible with confocal microscopy after 10 min plasma treatment. Because of its multi-factorial action, it is widely presumed that the development of bacterial resistance to plasma is unlikely. However, our results indicate that a short plasma treatment (3 min may lead to the emergence of a small number of surviving cells exhibiting enhanced resistance to subsequent plasma exposure. Interestingly, these cells also exhibited a higher degree of resistance to hydrogen peroxide. Whole genome comparison between surviving cells and control cells revealed 10 distinct polymorphic regions, including four belonging to the redox active, antibiotic pigment phenazine. Subsequently, the interaction between phenazine production and CAP resistance was demonstrated in biofilms of transposon mutants disrupted in different phenazine pathway genes which exhibited significantly altered sensitivity to CAP.

  9. Contribution of stress responses to antibiotic tolerance in Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Stewart, Philip S; Franklin, Michael J; Williamson, Kerry S; Folsom, James P; Boegli, Laura; James, Garth A

    2015-07-01

    Enhanced tolerance of biofilm-associated bacteria to antibiotic treatments is likely due to a combination of factors, including changes in cell physiology as bacteria adapt to biofilm growth and the inherent physiological heterogeneity of biofilm bacteria. In this study, a transcriptomics approach was used to identify genes differentially expressed during biofilm growth of Pseudomonas aeruginosa. These genes were tested for statistically significant overlap, with independently compiled gene lists corresponding to stress responses and other putative antibiotic-protective mechanisms. Among the gene groups tested were those associated with biofilm response to tobramycin or ciprofloxacin, drug efflux pumps, acyl homoserine lactone quorum sensing, osmotic shock, heat shock, hypoxia stress, and stationary-phase growth. Regulons associated with Anr-mediated hypoxia stress, RpoS-regulated stationary-phase growth, and osmotic stress were significantly enriched in the set of genes induced in the biofilm. Mutant strains deficient in rpoS, relA and spoT, or anr were cultured in biofilms and challenged with ciprofloxacin and tobramycin. When challenged with ciprofloxacin, the mutant strain biofilms had 2.4- to 2.9-log reductions in viable cells compared to a 0.9-log reduction of the wild-type strain. Interestingly, none of the mutants exhibited a statistically significant alteration in tobramycin susceptibility compared to that with the wild-type biofilm. These results are consistent with a model in which multiple genes controlled by overlapping starvation or stress responses contribute to the protection of a P. aeruginosa biofilm from ciprofloxacin. A distinct and as yet undiscovered mechanism protects the biofilm bacteria from tobramycin.

  10. 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.

  11. 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

    Resistance to beta-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 beta-lactamase and biofilm formation. The purpose of this study was to investigate...... 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...... 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...

  12. Evaluation of Mannosidase and Trypsin Enzymes Effects on Biofilm Production of Pseudomonas aeruginosa Isolated from Burn Wound Infections

    Science.gov (United States)

    Banar, Maryam; Emaneini, Mohammad; Satarzadeh, Mhboubeh; Abdellahi, Nafiseh; Beigverdi, Reza; van Leeuwen, Willem B.; Jabalameli, Fereshteh

    2016-01-01

    Biofilm is an important virulence factor in Pseudomonas aeruginosa and has a substantial role in antibiotic resistance and chronic burn wound infections. New therapeutic agents against P. aeruginosa, degrading biofilms in burn wounds and improving the efficacy of current antimicrobial agents, are required. In this study, the effects of α-mannosidase, β-mannosidase and trypsin enzymes on the degradation of P. aeruginosa biofilms and on the reduction of ceftazidime minimum biofilm eliminating concentrations (MBEC) were evaluated. All tested enzymes, destroyed the biofilms and reduced the ceftazidime MBECs. However, only trypsin had no cytotoxic effect on A-431 human epidermoid carcinoma cell lines. In conclusion, since trypsin had better features than mannosidase enzymes, it can be a promising agent in combatting P. aeruginosa burn wound infections. PMID:27736961

  13. In vivo pharmacokinetics/pharmacodynamics of colistin and imipenem in Pseudomonas aeruginosa biofilm infection

    DEFF Research Database (Denmark)

    Hengzhuang, Wang; Wu, Hong; Ciofu, Oana;

    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...

  14. Discovery of an operon that participates in agmatine metabolism and regulates biofilm formation in Pseudomonas aeruginosa.

    Science.gov (United States)

    Williams, Bryan J; Du, Rui-Hong; Calcutt, M Wade; Abdolrasulnia, Rasul; Christman, Brian W; Blackwell, Timothy S

    2010-04-01

    Agmatine is the decarboxylation product of arginine and a number of bacteria have devoted enzymatic pathways for its metabolism. Pseudomonas aeruginosa harbours the aguBA operon that metabolizes agmatine to putrescine, which can be subsequently converted into other polyamines or shunted into the TCA cycle for energy production. We discovered an alternate agmatine operon in the P. aeruginosa strain PA14 named agu2ABCA' that contains two genes for agmatine deiminases (agu2A and agu2A'). This operon was found to be present in 25% of clinical P. aeruginosa isolates. Agu2A' contains a twin-arginine translocation signal at its N-terminus and site-directed mutagenesis and cell fractionation experiments confirmed this protein is secreted to the periplasm. Analysis of the agu2ABCA' promoter demonstrates that agmatine induces expression of the operon during the stationary phase of growth and during biofilm growth and agu2ABCA' provides only weak complementation of aguBA, which is induced during log phase. Biofilm assays of mutants of all three agmatine deiminase genes in PA14 revealed that deletion of agu2ABCA', specifically its secreted product Agu2A', reduces biofilm production of PA14 following addition of exogenous agmatine. Together, these findings reveal a novel role for the agu2ABCA' operon in the biofilm development of P. aeruginosa.

  15. 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.

  16. Optimal dosing regimen of nitric oxide donor compounds for the reduction of Pseudomonas aeruginosa biofilm and isolates from wastewater membranes.

    Science.gov (United States)

    Barnes, Robert J; Bandi, Ratnaharika R; Wong, Wee Seng; Barraud, Nicolas; McDougald, Diane; Fane, Anthony; Kjelleberg, Staffan; Rice, Scott A

    2013-01-01

    Membrane fouling by bacterial biofilms remains a key challenge for membrane-based water purification systems. Here, the optimal biofilm dispersal potential of three nitric oxide (NO) donor compounds, viz. sodium nitroprusside, 6-(2-hydroxy-1-methyl-2-nitrosohydrazino)-N-methyl-1-hexanamine (MAHMA NONOate) and 1-(hydroxy-NNO-azoxy)-L-proline, disodium salt, was investigated using Pseudomonas aeruginosa PAO1 as a model organism. Dispersal was quantitatively assessed by confocal microscopy [bacterial cells and the components of the extracellular polymeric substances (EPS) (polysaccharides and extracellular DNA)] and colony-forming unit counts. The three NO donor compounds had different optimal exposure times and concentrations, with MAHMA NONOate being the optimal NO donor compound. Biofilm dispersal correlated with a reduction in both bacterial cells and EPS. MAHMA NONOate also reduced single species biofilms formed by bacteria isolated from industrial membrane bioreactor and reverse osmosis membranes, as well as in isolates combined to generate mixed species biofilms. The data present strong evidence for the application of these NO donor compounds for prevention of biofouling in an industrial setting.

  17. 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

    Bacterial biofilms on long-term catheters are a major source of infection. Exposure to ultraviolet C (UVC - 265 nm) light was shown in an earlier study to reduce the number of bacteria substantially on ex vivo treated urinary patient catheters. Very large doses (long treatment times) should......, however, be applied to obtain 99.9% disinfection rates. The major reason was that besides cells the mature biofilm contained absorbing and scattering particulates, which made the biofilm opaque. The potential of UVC light emitting diodes (LED) for disinfection purposes in catheter-like tubes contaminated...... 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...

  18. 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

    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......Pseudomonas aeruginosa produces extracellular DNA which functions as a cell-to-cell interconnecting matrix component in biofilms. Comparison of extracellular DNA and chromosomal DNA by the use of polymerase chain reaction and Southern analysis suggested that the extracellular DNA is similar...... with the wild type and lasIrhlI, pqsA, pqsL and fliMpilA mutants indicated that the extracellular DNA is generated via a mechanism which is dependent on acyl homoserine lactone and Pseudomonas quinolone signalling, as well as on flagella and type IV pili. Microscopic investigation of flow chamber-grown wild...

  19. Pseudomonas aeruginosa lipopolysaccharide inhibits Candida albicans hyphae formation and alters gene expression during biofilm development.

    Science.gov (United States)

    Bandara, H M H N; K Cheung, B P; Watt, R M; Jin, L J; Samaranayake, L P

    2013-02-01

    Elucidation of bacterial and fungal interactions in multispecies biofilms will have major impacts on understanding the pathophysiology of infections. The objectives of this study were to (i) evaluate the effect of Pseudomonas aeruginosa lipopolysaccharide (LPS) on Candida albicans hyphal development and transcriptional regulation, (ii) investigate protein expression during biofilm formation, and (iii) propose likely molecular mechanisms for these interactions. The effect of LPS on C. albicans biofilms was assessed by XTT-reduction and growth curve assays, light microscopy, scanning electron microscopy (SEM), and confocal laser scanning microscopy (CLSM). Changes in candidal hypha-specific genes (HSGs) and transcription factor EFG1 expression were assessed by real-time polymerase chain reaction and two-dimensional gel electrophoresis, respectively. Proteome changes were examined by mass spectrometry. Both metabolic activities and growth rates of LPS-treated C. albicans biofilms were significantly lower (P GDH1), CaO19.11135(PGK1), CaO19.9877(HNT1) by P. aeruginosa LPS. Our data imply that bacterial LPS inhibit C. albicans biofilm formation and hyphal development. The P. aeruginosa LPS likely target glycolysis-associated mechanisms during candidal filamentation.

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

    Directory of Open Access Journals (Sweden)

    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.

  1. Control of Candida albicans metabolism and biofilm formation by Pseudomonas aeruginosa phenazines.

    Science.gov (United States)

    Morales, Diana K; Grahl, Nora; Okegbe, Chinweike; Dietrich, Lars E P; Jacobs, Nicholas J; Hogan, Deborah A

    2013-01-29

    Candida albicans has developmental programs that govern transitions between yeast and filamentous morphologies and between unattached and biofilm lifestyles. Here, we report that filamentation, intercellular adherence, and biofilm development were inhibited during interactions between Candida albicans and Pseudomonas aeruginosa through the action of P. aeruginosa-produced phenazines. While phenazines are toxic to C. albicans at millimolar concentrations, we found that lower concentrations of any of three different phenazines (pyocyanin, phenazine methosulfate, and phenazine-1-carboxylate) allowed growth but affected the development of C. albicans wrinkled colony biofilms and inhibited the fungal yeast-to-filament transition. Phenazines impaired C. albicans growth on nonfermentable carbon sources and led to increased production of fermentation products (ethanol, glycerol, and acetate) in glucose-containing medium, leading us to propose that phenazines specifically inhibited respiration. Methylene blue, another inhibitor of respiration, also prevented the formation of structured colony biofilms. The inhibition of filamentation and colony wrinkling was not solely due to lowered extracellular pH induced by fermentation. Compared to smooth, unstructured colonies, wrinkled colony biofilms had higher oxygen concentrations within the colony, and wrinkled regions of these colonies had higher levels of respiration. Together, our data suggest that the structure of the fungal biofilm promotes access to oxygen and enhances respiratory metabolism and that the perturbation of respiration by bacterial molecules such as phenazines or compounds with similar activities disrupts these pathways. These findings may suggest new ways to limit fungal biofilms in the context of disease. IMPORTANCE Many of the infections caused by Candida albicans, a major human opportunistic fungal pathogen, involve both morphological transitions and the formation of surface-associated biofilms. Through the

  2. Accelerated corrosion of 2205 duplex stainless steel caused by marine aerobic Pseudomonas aeruginosa biofilm.

    Science.gov (United States)

    Xu, Dake; Xia, Jin; Zhou, Enze; Zhang, Dawei; Li, Huabing; Yang, Chunguang; Li, Qi; Lin, Hai; Li, Xiaogang; Yang, Ke

    2017-02-01

    Microbiologically influenced corrosion (MIC) of 2205 duplex stainless steel (DSS) in the presence of Pseudomonas aeruginosa was investigated through electrochemical and surface analyses. The electrochemical results showed that P. aeruginosa significantly reduced the corrosion resistance of 2205 DSS. Confocal laser scanning microscopy (CLSM) images showed that the depths of the largest pits on 2205 DSS with and without P. aeruginosa were 14.0 and 4.9μm, respectively, indicating that the pitting corrosion was accelerated by P. aeruginosa. X-ray photoelectron spectroscopy (XPS) results revealed that CrO3 and CrN formed on the 2205 DSS surface in the presence of P. aeruginosa.

  3. Legionella pneumophila persists within biofilms formed by Klebsiella pneumoniae, Flavobacterium sp., and Pseudomonas fluorescens under dynamic flow conditions.

    Directory of Open Access Journals (Sweden)

    Catherine R Stewart

    Full Text Available Legionella pneumophila, the agent of Legionnaires' disease pneumonia, is transmitted to humans following the inhalation of contaminated water droplets. In aquatic systems, L. pneumophila survives much of time within multi-organismal biofilms. Therefore, we examined the ability of L. pneumophila (clinical isolate 130 b to persist within biofilms formed by various types of aquatic bacteria, using a bioreactor with flow, steel surfaces, and low-nutrient conditions. L. pneumophila was able to intercalate into and persist within a biofilm formed by Klebsiella pneumoniae, Flavobacterium sp. or Pseudomonas fluorescens. The levels of L. pneumophila within these biofilms were as much as 4 × 10(4 CFU per cm(2 of steel coupon and lasted for at least 12 days. These data document that K. pneumoniae, Flavobacterium sp., and P. fluorescens can promote the presence of L. pneumophila in dynamic biofilms. In contrast to these results, L. pneumophila 130 b did not persist within a biofilm formed by Pseudomonas aeruginosa, confirming that some bacteria are permissive for Legionella colonization whereas others are antagonistic. In addition to colonizing certain mono-species biofilms, L. pneumophila 130 b persisted within a two-species biofilm formed by K. pneumoniae and Flavobacterium sp. Interestingly, the legionellae were also able to colonize a two-species biofilm formed by K. pneumoniae and P. aeruginosa, demonstrating that a species that is permissive for L. pneumophila can override the inhibitory effect(s of a non-permissive species.

  4. Phage ΦPan70, a Putative Temperate Phage, Controls Pseudomonas aeruginosa in Planktonic, Biofilm and Burn Mouse Model Assays

    Science.gov (United States)

    Holguín, Angela V.; Rangel, Guillermo; Clavijo, Viviana; Prada, Catalina; Mantilla, Marcela; Gomez, María Catalina; Kutter, Elizabeth; Taylor, Corinda; Fineran, Peter C.; Barrios, Andrés Fernando González; Vives, Martha J.

    2015-01-01

    Pseudomonas aeruginosa is one of the Multi-Drug-Resistant organisms most frequently isolated worldwide and, because of a shortage of new antibiotics, bacteriophages are considered an alternative for its treatment. Previously, P. aeruginosa phages were isolated and best candidates were chosen based on their ability to form clear plaques and their host range. This work aimed to characterize one of those phages, ΦPan70, preliminarily identified as a good candidate for phage-therapy. We performed infection curves, biofilm removal assays, transmission-electron-microscopy, pulsed-field-gel-electrophoresis, and studied the in vivo ΦPan70 biological activity in the burned mouse model. ΦPan70 was classified as a member of the Myoviridae family and, in both planktonic cells and biofilms, was responsible for a significant reduction in the bacterial population. The burned mouse model showed an animal survival between 80% and 100%, significantly different from the control animals (0%). However, analysis of the ΦPan70 genome revealed that it was 64% identical to F10, a temperate P. aeruginosa phage. Gene annotation indicated ΦPan70 as a new, but possible temperate phage, therefore not ideal for phage-therapy. Based on this, we recommend genome sequence analysis as an early step to select candidate phages for potential application in phage-therapy, before entering into a more intensive characterization. PMID:26274971

  5. Physiology of Pseudomonas aeruginosa in biofilms as revealed by transcriptome analysis

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    Parker Albert

    2010-11-01

    Full Text Available Abstract Background Transcriptome analysis was applied to characterize the physiological activities of Pseudomonas aeruginosa grown for three days in drip-flow biofilm reactors. Conventional applications of transcriptional profiling often compare two paired data sets that differ in a single experimentally controlled variable. In contrast this study obtained the transcriptome of a single biofilm state, ranked transcript signals to make the priorities of the population manifest, and compared ranki ngs for a priori identified physiological marker genes between the biofilm and published data sets. Results Biofilms tolerated exposure to antibiotics, harbored steep oxygen concentration gradients, and exhibited stratified and heterogeneous spatial patterns of protein synthetic activity. Transcriptional profiling was performed and the signal intensity of each transcript was ranked to gain insight into the physiological state of the biofilm population. Similar rankings were obtained from data sets published in the GEO database http://www.ncbi.nlm.nih.gov/geo. By comparing the rank of genes selected as markers for particular physiological activities between the biofilm and comparator data sets, it was possible to infer qualitative features of the physiological state of the biofilm bacteria. These biofilms appeared, from their transcriptome, to be glucose nourished, iron replete, oxygen limited, and growing slowly or exhibiting stationary phase character. Genes associated with elaboration of type IV pili were strongly expressed in the biofilm. The biofilm population did not indicate oxidative stress, homoserine lactone mediated quorum sensing, or activation of efflux pumps. Using correlations with transcript ranks, the average specific growth rate of biofilm cells was estimated to be 0.08 h-1. Conclusions Collectively these data underscore the oxygen-limited, slow-growing nature of the biofilm population and are consistent with antimicrobial tolerance due

  6. Anti-Biofilm Activities from Marine Cold Adapted Bacteria Against Staphylococci and Pseudomonas aeruginosa.

    Science.gov (United States)

    Papa, Rosanna; Selan, Laura; Parrilli, Ermenegilda; Tilotta, Marco; Sannino, Filomena; Feller, Georges; Tutino, Maria L; Artini, Marco

    2015-01-01

    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 and to test their

  7. 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

  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. 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

    developments of the biofilms were quantified by the computer program COMSTAT (A. Heydorn, A. T. Nielsen, M. Hentzer, C. Sternberg, M. Givskov, B. K. Ersboll, and S. Molin, Microbiology 146:2395-2407, 2000). Two structural key variables, average thickness and roughness, formed the basis for an analysis......Four strains of Pseudomonas aeruginosa (wild type, DeltapilHIJK mutant, lasI mutant, and rpoS mutant) were genetically tagged with the green fluorescent protein, and the development of flow chamber-grown biofilms by each of them was investigated by confocal laser scanning microscopy. The structural...... 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...

  10. c-di-GMP and its Effects on Biofilm Formation and Dispersion: a Pseudomonas Aeruginosa Review.

    Science.gov (United States)

    Ha, Dae-Gon; O'Toole, George A

    2015-04-01

    Since its initial discovery as an allosteric factor regulating cellulose biosynthesis in Gluconacetobacter xylinus, the list of functional outputs regulated by c-di-GMP has grown. We have focused this article on one of these c-di-GMP-regulated processes, namely, biofilm formation in the organism Pseudomonas aeruginosa. The majority of diguanylate cyclases and phosphodiesterases encoded in the P. aeruginosa genome still remain uncharacterized; thus, there is still a great deal to be learned about the link between c-di-GMP and biofilm formation in this microbe. In particular, while a number of c-di-GMP metabolizing enzymes have been identified that participate in reversible and irreversible attachment and biofilm maturation, there is a still a significant knowledge gap regarding the c-di-GMP output systems in this organism. Even for the well-characterized Pel system, where c-di-GMP-mediated transcriptional regulation is now well documented, how binding of c-di-GMP by PelD stimulates Pel production is not understood in any detail. Similarly, c-di-GMP-mediated control of swimming, swarming and twitching also remains to be elucidated. Thus, despite terrific advances in our understanding of P. aeruginosa biofilm formation and the role of c-di-GMP in this process since the last version of this book (indeed there was no chapter on c-di-GMP!) there is still much to learn.

  11. Effect of bacteriophage infection in combination with tobramycin on the emergence of resistance in Escherichia coli and Pseudomonas aeruginosa biofilms.

    Science.gov (United States)

    Coulter, Lindsey B; McLean, Robert J C; Rohde, Rodney E; Aron, Gary M

    2014-10-03

    Bacteriophage infection and antibiotics used individually to reduce biofilm mass often result in the emergence of significant levels of phage and antibiotic resistant cells. In contrast, combination therapy in Escherichia coli biofilms employing T4 phage and tobramycin resulted in greater than 99% and 39% reduction in antibiotic and phage resistant cells, respectively. In P. aeruginosa biofilms, combination therapy resulted in a 60% and 99% reduction in antibiotic and PB-1 phage resistant cells, respectively. Although the combined treatment resulted in greater reduction of E. coli CFUs compared to the use of antibiotic alone, infection of P. aeruginosa biofilms with PB-1 in the presence of tobramycin was only as effective in the reduction of CFUs as the use of antibiotic alone. The study demonstrated phage infection in combination with tobramycin can significantly reduce the emergence of antibiotic and phage resistant cells in both E. coli and P. aeruginosa biofilms, however, a reduction in biomass was dependent on the phage-host system.

  12. Antimicrobial and anti-biofilm effect of a novel BODIPY photosensitizer against Pseudomonas aeruginosa PAO1

    DEFF Research Database (Denmark)

    Orlandi, Viviana Teresa; Rybtke, Morten; Caruso, Enrico;

    2014-01-01

    Photodynamic therapy (PDT) combines the use of organic dyes (photosensitizers, PSs) and visible light in order to elicit a photo-oxidative stress which causes bacterial death. GD11, a recently synthesized PS belonging to the boron-dipyrromethene (BODIPY) class, was demonstrated to be efficient...... against planktonic cultures of Pseudomonas aeruginosa, causing a 7 log unit reduction of viable cells when administered at 2.5 μM. The effectiveness of GD11 against P. aeruginosa biofilms grown in flow-cells and microtiter trays was also demonstrated. Confocal laser scanning microscopy of flow...

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

    DEFF Research Database (Denmark)

    Hentzer, Morten; Riedel, Kathrin; Rasmussen, Thomas B;

    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 by expression of an unstable version of the green-fluorescent protein (Gfp...... macroalga Delisea pulchra, is capable of interfering with AHL-mediated quorum sensing in P. aeruginosa. It is demonstrated that the furanone compound specifically represses expression of a PlasB-gfp reporter fusion without affecting growth or protein synthesis. In addition, it reduces the production...

  14. Synergistic effect of polyaniline coverage and surface microstructure on the inhibition of Pseudomonas aeruginosa biofilm formation.

    Science.gov (United States)

    Gallarato, L A; Mulko, L E; Dardanelli, M S; Barbero, C A; Acevedo, D F; Yslas, E I

    2017-02-01

    Biofilm Formation is a survival strategy for microorganisms to adapt to their environment. Microbial cells in biofilm become tolerant and resistant to antibiotics and immune responses, increasing the difficulties for the clinical treatment of microbial infections. The surface chemistry and the micro/nano-topography of solid interfaces play a major role in mediating microorganism activity and adhesion. The effect of the surface chemical composition and topography on the adhesion and viability of Pseudomonas aeruginosa was studied. Polymeric (polyethylene terephthalate) surfaces were covered with a conducting polymer (polyaniline, PANI) film by in-situ polymerization and microstructured by Direct Laser Interference Patterning (DLIP). The viability of Pseudomonas aeruginosa on the different surfaces was investigated. The physicochemical properties of the surfaces were characterized by water contact angle measurements, scanning electron microscopy and atomic force microscopy. Bacterial biofilms were imaged by atomic force and scanning electron microscopies. The bacterial viability decreased on PANI compared with the substrate (polyethylene terephthalate) and it decreased even more upon micro-structuring the PANI films. In addition, the biofilm reduction could be improved using polymers with different chemical composition and/or the same polymer with different topographies. Both methods presented diminish the bacterial attachment and biofilm formation. These findings present a high impact related to materials for biomedical engineer applications regarding medical devices, as prostheses or catheters.

  15. Characterization of membrane lipidome changes in Pseudomonas aeruginosa during biofilm growth on glass wool.

    Science.gov (United States)

    Benamara, Hayette; Rihouey, Christophe; Abbes, Imen; Ben Mlouka, Mohamed Amine; Hardouin, Julie; Jouenne, Thierry; Alexandre, Stéphane

    2014-01-01

    Bacteria cells within biofilms are physiologically distinct from their planktonic counterparts. In particular they are more resistant to detrimental environmental conditions. In this study, we monitored the evolution of the phospholipid composition of the inner and outer membranes of P. aeruginosa during the biofilm formation (i.e., from 1-, 2-, to 6-day-old biofilm). Lipidome analyses were performed by electrospray ionization mass spectrometry. In addition to the lipidomic analysis, the fatty acid composition was analysed by gas chromatography/mass spectrometry. We found that the lipidome alterations of the inner and the outer membranes varied with the biofilm age. These alterations in phospholipid compositions reflect a higher diversity in sessile organisms than in planktonic counterparts. The diversity is characterized by the presence of PE 30∶1, PE 31∶0 and PG 31∶0 for the lower masses as well as PE 38∶1, 38∶2, 39∶1, 39∶2 and PG 38∶0, 38∶1, 38∶2, 39∶1, 39∶2 for the higher masses. However, this lipidomic feature tends to disappear with the biofilm age, in particular the high mass phospholipids tend to disappear. The amount of branched chains phospholipids mainly located in the outer membrane decreased with the biofilm age, whereas the proportion of cyclopropylated phospholipids increased in both membranes. In bacteria present in oldest biofilms, i.e., 6-day-old, the phospholipid distribution moved closer to that of planktonic bacteria.

  16. Escherichia coli BdcA controls biofilm dispersal in Pseudomonas aeruginosa and Rhizobium meliloti

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    Wood Thomas K

    2011-10-01

    Full Text Available Abstract Background Previously we showed that BdcA controls Escherichia coli biofilm dispersal by binding the ubiquitous bacterial signal cyclic diguanylate (c-di-GMP; upon reducing the concentration of c-di-GMP, the cell shifts to the planktonic state by increasing motility, decreasing aggregation, and decreasing production of biofilm adhesins. Findings Here we report that BdcA also increases biofilm dispersal in other Gram-negative bacteria including Pseudomonas aeruginosa, Pseudomonas fluorescens, and Rhizobium meliloti. BdcA binds c-di-GMP in these strains and thereby reduces the effective c-di-GMP concentrations as demonstrated by increases in swimming motility and swarming motility as well as by a reduction in extracellular polysaccharide production. We also develop a method to displace existing biofilms by adding BdcA via conjugation from E. coli in mixed-species biofilms. Conclusion Since BdcA shows the ability to control biofilm dispersal in diverse bacteria, BdcA has the potential to be used as a tool to disperse biofilms for engineering and medical applications.

  17. 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.

  18. 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.

  19. Eradication of Pseudomonas aeruginosa biofilms by atmospheric pressure non-thermal plasma.

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    Mahmoud Y Alkawareek

    Full Text Available Bacteria exist, in most environments, as complex, organised communities of sessile cells embedded within a matrix of self-produced, hydrated extracellular polymeric substances known as biofilms. Bacterial biofilms represent a ubiquitous and predominant cause of both chronic infections and infections associated with the use of indwelling medical devices such as catheters and prostheses. Such infections typically exhibit significantly enhanced tolerance to antimicrobial, biocidal and immunological challenge. This renders them difficult, sometimes impossible, to treat using conventional chemotherapeutic agents. Effective alternative approaches for prevention and eradication of biofilm associated chronic and device-associated infections are therefore urgently required. Atmospheric pressure non-thermal plasmas are gaining increasing attention as a potential approach for the eradication and control of bacterial infection and contamination. To date, however, the majority of studies have been conducted with reference to planktonic bacteria and rather less attention has been directed towards bacteria in the biofilm mode of growth. In this study, the activity of a kilohertz-driven atmospheric pressure non-thermal plasma jet, operated in a helium oxygen mixture, against Pseudomonas aeruginosa in vitro biofilms was evaluated. Pseudomonas aeruginosa biofilms exhibit marked susceptibility to exposure of the plasma jet effluent, following even relatively short (≈ 10's s exposure times. Manipulation of plasma operating conditions, for example, plasma operating frequency, had a significant effect on the bacterial inactivation rate. Survival curves exhibit a rapid decline in the number of surviving cells in the first 60 seconds followed by slower rate of cell number reduction. Excellent anti-biofilm activity of the plasma jet was also demonstrated by both confocal scanning laser microscopy and metabolism of the tetrazolium salt, XTT, a measure of bactericidal

  20. Association of biofilm production with multidrug resistance among clinical isolates of Acinetobacter baumannii and Pseudomonas aeruginosa from intensive care unit

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    Jeetendra Gurung

    2013-01-01

    Full Text Available Background and Aims: Given choice, bacteria prefer a community-based, surface-bound colony to an individual existence. The inclination for bacteria to become surface bound is so ubiquitous in diverse ecosystems that it suggests a strong survival strategy and selective advantage for surface dwellers over their free-ranging counterparts. Virtually any surface, biotic or abiotic (animal, mineral, or vegetable is suitable for bacterial colonization and biofilm formation. Thus, a biofilm is "a functional consortium of microorganisms organized within an extensive exopolymeric matrix." Materials and Methods: The present study was undertaken to detect biofilm production from the repertoire stocks of Acinetobacter baumannii (A. baumannii and Pseudomonas aeruginosa (P. aeruginosa obtained from clinical specimens. The tube method was performed to qualitatively detect biofilm production. Results: A total of 109 isolates of both organisms were included in the study, out of which 42% (46/109 isolates showed biofilm detection. Among the biofilm producers, 57% of P. aeruginosa and 73% of A. baumannii showed multidrug resistance (MDR pattern which was statistically significant in comparison to nonbiofilm producers (P < 0.001. Conclusion: To the best of our knowledge, this is the only study to have tested the biofilm production in both P. aeruginosa and A. baumannii in a single study. Biofilm production and MDR pattern were found to be significantly higher in A. baumannii than P. aeruginosa. Antibiotic resistance was significantly higher among biofilm producing P. aeruginosa than non producers. Similarly, antibiotic resistance was significantly higher among biofilm producing A. baumannii than non producers.

  1. 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.

  2. Effects of Combined Treatment with Sansanmycin and Macrolides on Pseudomonas aeruginosa and Formation of Biofilm

    Institute of Scientific and Technical Information of China (English)

    YUE LI; YUN-YING XIE; RU-XIAN CHEN; HONG-ZHANG XU; GUO-JI ZHANG; JIN-ZHE LI; XIAO-MIAN LI

    2009-01-01

    Objective To observe the effects of combined treatment with sansanmycin and macrolides on Pseudomonas aeruginosa and formation of biofilm. Methods Micro-dilution method was used to determine the minimal inhibitory concentrations (MICs) of sansanmycin, gentamycin, carbenicillin, polymyxin B, roxithromycin, piperacillin, and tazobactam. PA1 and PA27853 biofilms were observed under optical microscope after staining and under SEM after treatment with sansanmycin at different dosages and combined treatment with sansanmycin and roxithromycin. Viable bacteria in PA1 and PA27853 biofilms were counted after treatment with sansanmycin at different dosages or combined treatment with sansanmycin and roxithromycin. Results The MIC of sansanmycin was lower than that of gentamycin and polymyxin B, but was higher than that of carbenicillin. Roxithromycin enhanced the penetration of sansanmycin to PA1 and PA27853 strains through biofilms. PA1 and PA27853 biofilms were gradually cleared with the increased dosages of sansanmycin or with the combined sansanmycin and roxithromycin. Conclusion Sub-MIC levels of roxithromycin and sansanmycin substantially inhibit the generation of biofilms and proliferation of bacteria. Therefore, combined antibiotics can be used in treatment of intractable bacterial infection.

  3. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  5. Inhibition of quorum sensing-mediated biofilm formation in Pseudomonas aeruginosa by a locally isolated Bacillus cereus.

    Science.gov (United States)

    Wahman, Shaimaa; Emara, Mohamed; Shawky, Riham M; El-Domany, Ramadan A; Aboulwafa, Mohammad Mabrouk

    2015-12-01

    Quorum sensing has been shown to play a crucial role in Pseudomonas aeruginosa pathogenesis where it activates expression of myriad genes that regulate the production of important virulence factors such as biofilm formation. Antagonism of quorum sensing is an excellent target for antimicrobial therapy and represents a novel approach to combat drug resistance. In this study, Chromobacterium violaceum biosensor strain was employed as a fast, sensitive, reliable, and easy to use tool for rapid screening of soil samples for Quorum Sensing Inhibitors (QSI) and the optimal conditions for maximal QSI production were scrutinized. Screening of 127 soil isolates showed that 43 isolates were able to breakdown the HHL signal. Out of the 43 isolates, 38 isolates were able to inhibit the violet color of the biosensor and to form easily detectable zones of color inhibition around their growth. A confirmatory bioassay was carried out after concentrating the putative positive cell-free lysates. Three different isolates that belonged to Bacillus cereus group were shown to have QSI activities and their QSI activities were optimized by changing their culture conditions. Further experiments revealed that the cell-free lysates of these isolates were able to inhibit biofilm formation by P. aeruginosa clinical isolates.

  6. Chronic lung infection by Pseudomonas aeruginosa biofilm is cured by L-Methionine in combination with antibiotic therapy.

    Science.gov (United States)

    Gnanadhas, Divya Prakash; Elango, Monalisha; Datey, Akshay; Chakravortty, Dipshikha

    2015-11-02

    Bacterial biofilms are associated with 80-90% of infections. Within the biofilm, bacteria are refractile to antibiotics, requiring concentrations >1,000 times the minimum inhibitory concentration. Proteins, carbohydrates and DNA are the major components of biofilm matrix. Pseudomonas aeruginosa (PA) biofilms, which are majorly associated with chronic lung infection, contain extracellular DNA (eDNA) as a major component. Herein, we report for the first time that L-Methionine (L-Met) at 0.5 μM inhibits Pseudomonas aeruginosa (PA) biofilm formation and disassembles established PA biofilm by inducing DNase expression. Four DNase genes (sbcB, endA, eddB and recJ) were highly up-regulated upon L-Met treatment along with increased DNase activity in the culture supernatant. Since eDNA plays a major role in establishing and maintaining the PA biofilm, DNase activity is effective in disrupting the biofilm. Upon treatment with L-Met, the otherwise recalcitrant PA biofilm now shows susceptibility to ciprofloxacin. This was reflected in vivo, in the murine chronic PA lung infection model. Mice treated with L-Met responded better to antibiotic treatment, leading to enhanced survival as compared to mice treated with ciprofloxacin alone. These results clearly demonstrate that L-Met can be used along with antibiotic as an effective therapeutic against chronic PA biofilm infection.

  7. 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

    biofilms, and the development of tolerance to the antimicrobial agents was found to be affected as well. Mutations in genes interfering with lipopolysaccharide modification (pmr) eliminated the biofilm-associated colistin tolerance phenotype. Experiments with a PAO1 strain harboring a pmr-gfp fusion showed...... 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....

  8. 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.

  9. A quorum-sensing inhibitor blocks Pseudomonas aeruginosa virulence and biofilm formation.

    Science.gov (United States)

    O'Loughlin, Colleen T; Miller, Laura C; Siryaporn, Albert; Drescher, Knut; Semmelhack, Martin F; Bassler, Bonnie L

    2013-10-29

    Quorum sensing is a chemical communication process that bacteria use to regulate collective behaviors. Disabling quorum-sensing circuits with small molecules has been proposed as a potential strategy to prevent bacterial pathogenicity. The human pathogen Pseudomonas aeruginosa uses quorum sensing to control virulence and biofilm formation. Here, we analyze synthetic molecules for inhibition of the two P. aeruginosa quorum-sensing receptors, LasR and RhlR. Our most effective compound, meta-bromo-thiolactone (mBTL), inhibits both the production of the virulence factor pyocyanin and biofilm formation. mBTL also protects Caenorhabditis elegans and human lung epithelial cells from killing by P. aeruginosa. Both LasR and RhlR are partially inhibited by mBTL in vivo and in vitro; however, RhlR, not LasR, is the relevant in vivo target. More potent antagonists do not exhibit superior function in impeding virulence. Because LasR and RhlR reciprocally control crucial virulence factors, appropriately tuning rather than completely inhibiting their activities appears to hold the key to blocking pathogenesis in vivo.

  10. 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...... 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...

  11. Adsorption to metal oxides of the Pseudomonas aeruginosa siderophore pyoverdine and implications for bacterial biofilm formation on metals.

    Science.gov (United States)

    Upritchard, Hamish G; Yang, Jing; Bremer, Philip J; Lamont, Iain L; McQuillan, A James

    2007-06-19

    The initiation of biofilm formation is poorly understood, and in particular, the contribution of chemical bond formation between bacterial cells and metal surfaces has received little attention. We have previously used in situ infrared spectroscopy to show, during the initial stages of Pseudomonas aeruginosa biofilm formation, the formation of coordinate covalent bonds between titanium dioxide particle films and pyoverdine, a mixed catecholate and hydroxamate siderophore. Here we show using infrared spectroscopy that pyoverdine can also form covalent bonds with particle films of Fe2O3, CrOOH, and AlOOH. Adsorption to the metal oxides through the catechol-like 2,3-diamino-6,7-dihydroxyquinoline part of pyoverdine was most evident in the infrared spectrum of the adsorbed pyoverdine molecule. Weaker infrared absorption bands that are consistent with the hydroxamic acids of pyoverdine binding covalently to TiO2, Fe2O3, and AlOOH surfaces were also observed. The adsorption of pyoverdine to TiO2 and Fe2O3 surfaces showed a pH dependence that is indicative of the dominance of the catechol-like ligand of pyoverdine. Infrared absorption bands were also evident for pyoverdine associated with the cells of P. aeruginosa on TiO2 and Fe2O3 surfaces and were notably absent for genetically modified cells unable to synthesize or bind pyoverdine at the cell surface. These studies confirm the generality of pyoverdine-metal bond formation and suggest a wider involvement of siderophores in bacterial biofilm initiation on metals.

  12. Effects of quorum sensing autoinducer degradation gene on virulence and biofilm formation of Pseudomonas aeruginosa

    Institute of Scientific and Technical Information of China (English)

    WANG Yao; DAI Yue; ZHANG Yong; HU YangBo; YANG BaoYu; CHEN ShiYun

    2007-01-01

    The aiiA gene from Bacillus thuringiensis was cloned into the Pseudomonas/E. coli shuttle vector and transformed into Pseudomonas aeruginosa strain PAO1. Western blotting showed that the AiiA protein was expressed in PAO1. After induction by IPTG for 6 h and 18 h, expression of the aiiA gene in PAO1completely degraded the quorum sensing autoinducers N-acylhomoserine lactones (AHLs):N-oxododecanoyI-L-homoserine lactone (OdDHL) and N-butyryI-L-homoserine lactone (BHL). The reduced amount of AHLs in PAO1 was also correlated with decreased expression and production of several virulence factors such as elastase and pyocyanin. AiiA expression also influenced bacterial swarming motility. Most importantly, our studies indicated that aiiA played significant roles in P.aeruginosa biofilm formation and dispersion, as observed by the differences of the biofilm formation on liquid and solid surfaces, and biofilm structures under a scanning electron microscope.

  13. Pathogenic effects of biofilm with chronic pseudomonas aeruginosa lung infection in rats

    Institute of Scientific and Technical Information of China (English)

    Ping Yan; Yiqiang Chen; Zhijun Song; Hong Wu; Jinliang Kong; Xuejun Qin

    2008-01-01

    Objective: To establish an animal model of P.aeruginosa biofilm associated with chronic pulmonary infection and investigate the pathogenic effects of biofilm. Methods: Experiments in vitro, measuring the MICS, MBCS of ievofloxacin(LFX), ceftazidime(CAZ) in PAO579 in alginate beads and planktonic PAO579. Rats were challenged with 0.1 ml of PAO579(109CFU/ml) in alginate beads or 0.1 ml of planktonic PAO579(109CFU/ml), 3,7,14 days after challenging, bacteriological, pathological features were observed. Results: The MICS, MBCS of LFX, CAZ in PAO579 in alginate beads were higher than those in planktonic PAO579 in vitro. CFU/lung in alginate beads group was significantly higher than that in planktonic bacteria group(P = 0.002, P =0.004, P = 0.002, respectively); macroscopic lung pathology and the inflammation in alginate beads group were significantly more severe compared to those in planktonic bacteria group in vivo. Conclusion: P.aeruginosa biofilm protected bacterium from killing of antibiotics and might mediate the host immune damage in the lung tissue and made bacterium evade the host immune defense.

  14. Effects of quorum sensing autoinducer degradation gene on virulence and biofilm formation of Pseudomonas aeruginosa

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The aiiA gene from Bacillus thuringiensis was cloned into the Pseudomonas/E. coli shuttle vector and transformed into Pseudomonas aeruginosa strain PAO1. Western blotting showed that the AiiA protein was expressed in PAO1. After induction by IPTG for 6 h and 18 h, expression of the aiiA gene in PAO1 completely degraded the quorum sensing autoinducers N-acylhomoserine lactones (AHLs): N-oxododecanoyl-L-homoserine lactone (OdDHL) and N-butyryl-L-homoserine lactone (BHL). The re- duced amount of AHLs in PAO1 was also correlated with decreased expression and production of several virulence factors such as elastase and pyocyanin. AiiA expression also influenced bacterial swarming motility. Most importantly, our studies indicated that aiiA played significant roles in P. aeruginosa biofilm formation and dispersion, as observed by the differences of the biofilm formation on liquid and solid surfaces, and biofilm structures under a scanning electron microscope.

  15. 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...

  16. 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.

  17. Impact of Pseudomonas aeruginosa quorum sensing on biofilm persistence in an in vivo intraperitoneal foreign-body infection model

    DEFF Research Database (Denmark)

    Christensen, Louise D; Moser, Claus; Jensen, Peter Ø

    2007-01-01

    Pseudomonas aeruginosa is an opportunistic human pathogen that causes chronic biofilm-based infections in host organisms. P. aeruginosa employs quorum sensing (QS) to control expression of its virulence, and to establish and maintain chronic infections. Under such conditions, the biofilm mode....... 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...... to the placebo-treated group. These results were obtained with both an inbred (BALB/c) and an outbred (NMRI) mouse strain. The present results support a model by which functional QS systems play a pivotal role in the ability of bacteria to resist clearing by the innate immune system and strongly suggest...

  18. Evaluation and optimization of multiple fluorophore analysis of a Pseudomonas aeruginosa biofilm

    Science.gov (United States)

    Baird, Fiona J.; Wadsworth, Marilyn P.; Hill, Jane E.

    2015-01-01

    Conventional laser scanning microscopy for multiple fluorescent stains can be a useful tool if the problems of autofluorescence and cross-talk are eliminated. The technique of spectral imaging was employed to unmix five different fluorophores – ranging in emission from 435 to 665 nm – applied to a Pseudomonas aeruginosa biofilm with overlapping spectra and which was not possible using traditional channel mode operation. Using lambda scanning and linear unmixing, the five fluorophores could be distinguished with regions of differentiation apparent. PMID:22587931

  19. Microbial interaction between a CTXM-15 -producing Escherichia coli and a susceptible Pseudomonas aeruginosa isolated from bronchoalveolar lavage: influence of cefotaxime in the dual-species biofilm formation.

    Science.gov (United States)

    Bessa, Lucinda J; Mendes, Ângelo; Gomes, Rita; Curvelo, Sara; Cravo, Sara; Sousa, Emília; Vasconcelos, Vitor; Martins da Costa, Paulo

    2015-06-01

    Two isolates, Escherichia coli ella00 and Pseudomonas aeruginosa ella01, obtained from bronchoalveolar lavage, were found to be closely associated in clusters in agar medium. Escherichia coli ella00 was multidrug resistant and CTXM-15 extended-spectrum β-lactamase producer, while P. aeruginosa ella01 was susceptible to all antimicrobials tested. These observations impelled for further studies aimed to understand their microbial interaction. The P. aeruginosa ella01 biofilm-forming capacity was reduced and not affected when it was co-cultured with E. coli ella00 and E. coli ATCC 25922 respectively. Interestingly, the co-culture of ella isolates in the presence of high concentrations, such as 160 μg ml(-1) , of cefotaxime allowed the formation of more biofilm than in the absence of the antibiotic. As revealed by fluorescence in situ hybridization, in co-culture, P. aeruginosa ella01 survived and subsequently flourished when exposed to this third-generation cephalosporin at a concentration 10 × higher than its minimum inhibitory concentration (MIC), and this was mostly due to β-lactamases production by E. coli ella00. In fact, it was demonstrated by high-performance liquid chromatography that cefotaxime was absent for the culture medium 4 h after application. In conclusion, we demonstrate that bacterial species can interact differently depending on the surrounding conditions (favourable or stressing), and that those interactions can switch from unprofitable to beneficial.

  20. 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

    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.......Stress response plays an important role on microbial adaptation under hostile environmental conditions. It is generally unclear how the signaling transduction pathway mediates a stress response in planktonic and biofilm modes of microbial communities simultaneously. Here, we showed that metalloid...

  1. Evolution of Ecological Diversity in Biofilms of Pseudomonas aeruginosa by Altered Cyclic Diguanylate Signaling

    Science.gov (United States)

    Flynn, Kenneth M.; Dowell, Gabrielle; Johnson, Thomas M.; Koestler, Benjamin J.; Waters, Christopher M.

    2016-01-01

    ABSTRACT The ecological and evolutionary forces that promote and maintain diversity in biofilms are not well understood. To quantify these forces, three Pseudomonas aeruginosa populations were experimentally evolved from strain PA14 in a daily cycle of attachment, assembly, and dispersal for 600 generations. Each biofilm population evolved diverse colony morphologies and mutator genotypes defective in DNA mismatch repair. This diversity enhanced population fitness and biofilm output, owing partly to rare, early colonizing mutants that enhanced attachment of others. Evolved mutants exhibited various levels of the intracellular signal cyclic-di-GMP, which associated with their timing of adherence. Manipulating cyclic-di-GMP levels within individual mutants revealed a network of interactions in the population that depended on various attachment strategies related to this signal. Diversification in biofilms may therefore arise and be reinforced by initial colonists that enable community assembly. IMPORTANCE How biofilm diversity assembles, evolves, and contributes to community function is largely unknown. This presents a major challenge for understanding evolution during chronic infections and during the growth of all surface-associated microbes. We used experimental evolution to probe these dynamics and found that diversity, partly related to altered cyclic-di-GMP levels, arose and persisted due to the emergence of ecological interdependencies related to attachment patterns. Clonal isolates failed to capture population attributes, which points to the need to account for diversity in infections. More broadly, this study offers an experimental framework for linking phenotypic variation to distinct ecological strategies in biofilms and for studying eco-evolutionary interactions. PMID:27021563

  2. 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

  3. 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

    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 ... in the chronic wounds of BALB/c mice was observed at day 7 (p biofilm infection in mice. The results showed an aggravating impact of local inflammation induced by PA biofilms......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...

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

    Science.gov (United States)

    Souto, Renata; Silva-Boghossian, Carina M; Colombo, Ana Paula Vieira

    2014-01-01

    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 and > 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 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.

  5. 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.

  6. Effect of ceftazidime of formation of biofilm of Pseudomonas aeruginosa and mechanism%头孢他啶对铜绿假单胞菌生物膜形成的影响与机制的探讨

    Institute of Scientific and Technical Information of China (English)

    李琬琛; 宋林; 李立艳; 孙续国; 魏爱琳; 魏殿军

    2016-01-01

    目的 探讨头孢他啶在铜绿假单胞菌(PAE)生物膜形成过程中的抑制 、清除作用与机制,更好的指导临床用药.方法 通过96孔板结晶紫染色方法,定量分析头孢他啶对PAE标准菌株生物膜的抑制 、清除作用,通过real time-PCR方法对生物膜形成相关基因的表达进行相对定量分析,探讨头孢他啶对PAE生物膜的作用机制.结果 未形成成熟生物膜时,头孢他啶对PAE生长的抑制效果较好,而在有成熟生物膜形成时,头孢他啶不能有效抑制PAE的生长;头孢他啶与PAE生物膜形成相关基因的增多表达相关.结论 头孢他啶能够促进PAE生物膜形成相关基因的表达,临床对慢性PAE感染患者的治疗应避免使用头孢他啶.%OBJECTIVE To observe the effect of ceftazidime on inhibition of formation and clearance of Pseudomonas aeruginosa biofilm and analyze the mechanisms so as to provide guidance for clinical use of antibiotics .METHODS The 96 well plate crystal violet staining method was used to quantitatively analyze the effect of ceftazidime on inhi-bition and clearance of the P .aeruginosa biofilm ,and the real-time PCR method was employed to perform the rela-tive quantitative analysis of the expression of biofilm formation-related genes so as to observe the effect of ceftazi-dime on the formation of P .aeruginosa biofilm .RESULTS Ceftazidime had better effect on inhibition of growth of P .aeruginosa when the mature biofilms were not formed ,however ,ceftazidime could not effectively inhibit the growth of P .aeruginosa when the mature biofilms were formed .Ceftazidime was associated with the increased ex-pression of the biofilm formation-related genes of P .aeruginosa .CONCLUSION Ceftazidime can promote the ex-pression of the biofilm formation-related genes of P .aeruginosa .It is necessary for the hospital to avoid the use of ceftazidime for the treatment of the patients with chronic P .aeruginosa infection .

  7. 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

    Biofilms are reported to be inherently refractory toward antimicrobial attack and, therefore, cause problems in industrial and medical settings. Pseudomonas aeruginosa biofilms contain subpopulations that exhibit high metabolic activity and subpopulations that exhibit low metabolic activity. We......-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...

  8. In vitro characterization of biofilms formed by Kingella kingae.

    Science.gov (United States)

    Kaplan, J B; Sampathkumar, V; Bendaoud, M; Giannakakis, A K; Lally, E T; Balashova, N V

    2016-10-07

    The Gram-negative bacterium Kingella kingae is part of the normal oropharyngeal mucosal flora of children kingae can enter the submucosa and cause infections of the skeletal system in children, including septic arthritis and osteomyelitis. The organism is also associated with infective endocarditis in children and adults. Although biofilm formation has been coupled with pharyngeal colonization, osteoarticular infections, and infective endocarditis, no studies have investigated biofilm formation in K. kingae. In this study we measured biofilm formation by 79 K. kingae clinical isolates using a 96-well microtiter plate crystal violet binding assay. We found that 37 of 79 strains (47%) formed biofilms. All strains that formed biofilms produced corroding colonies on agar. Biofilm formation was inhibited by proteinase K and DNase I. DNase I also caused the detachment of pre-formed K. kingae biofilm colonies. A mutant strain carrying a deletion of the pilus gene cluster pilA1pilA2fimB did not produce corroding colonies on agar, autoaggregate in broth, or form biofilms. Biofilm forming strains have higher levels of pilA1 expression. The extracellular components of biofilms contained 490 μg cm(-2) of protein, 0.68 μg cm(-2) of DNA, and 0.4 μg cm(-2) of total carbohydrates. We concluded that biofilm formation is common among K. kingae clinical isolates, and that biofilm formation is dependent on the production of proteinaceous pili and extracellular DNA. Biofilm development may have relevance to the colonization, transmission, and pathogenesis of this bacterium. Extracellular DNA production by K. kingae may facilitate horizontal gene transfer within the oral microbial community.

  9. 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.

  10. Multivalency effects on Pseudomonas aeruginosa biofilm inhibition and dispersal by glycopeptide dendrimers targeting lectin LecA.

    Science.gov (United States)

    Bergmann, Myriam; Michaud, Gaëlle; Visini, Ricardo; Jin, Xian; Gillon, Emilie; Stocker, Achim; Imberty, Anne; Darbre, Tamis; Reymond, Jean-Louis

    2016-01-01

    The galactose specific lectin LecA partly mediates the formation of antibiotic resistant biofilms by Pseudomonas aeruginosa, an opportunistic pathogen causing lethal airways infections in immunocompromised and cystic fibrosis patients, suggesting that preventing LecA binding to natural saccharides might provide new opportunities for treatment. Here 8-fold (G3) and 16-fold (G4) galactosylated analogs of GalAG2, a tetravalent G2 glycopeptide dendrimer LecA ligand and P. aeruginosa biofilm inhibitor, were obtained by convergent chloroacetyl thioether (ClAc) ligation between 4-fold or 8-fold chloroacetylated dendrimer cores and digalactosylated dendritic arms. Hemagglutination inhibition, isothermal titration calorimetry and biofilm inhibition assays showed that G3 dendrimers bind LecA slightly better than their parent G2 dendrimers and induce complete biofilm inhibition and dispersal of P. aeruginosa biofilms, while G4 dendrimers show reduced binding and no biofilm inhibition. A binding model accounting for the observed saturation of glycopeptide dendrimer galactosyl groups and LecA binding sites is proposed based on the crystal structure of a G3 dendrimer LecA complex.

  11. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  13. Detection of bacteria bearing resistant biofilm forms, by using the universal and specific PCR is still unhelpful in the diagnosis of periprosthetic joint infections(PJI.

    Directory of Open Access Journals (Sweden)

    Batool eZegaer

    2014-09-01

    Full Text Available Intraoperative conventional bacteriological cultures were compared with different polymerase chain reaction (PCR methods in patients with total joint arthroplasties. The isolated bacteria were investigated for biofilm formation, and the biofilm forming strains, in their planktonic and biofilm forms, were further tested for their antimicrobial resistance against several clinically important antimicrobials. Forty four bone and joint samples were included and classified as infected or non-infected according to standard criteria for periprosthetic hip and knee infections. For the bacteriological diagnosis, conventional culture, two types of universal PCR and species specific PCR for three selected pathogens (S. aureus, S. epidermidis, P. aeruginosa were applied. Biofilm formation determination was performed by the tissue culture plate method. Antimicrobial susceptibility of the planktonic bacteria was performed by the minimal inhibitory concentration determination and, of the biofilm forms, by the minimal inhibitory concentration for bacterial regrowth from the biofilm. Twenty samples were culture positive, with S. epidermidis, S. aureus or P. aeruginosa. All PCR methods were very ineffective in detecting only one pathogen. All isolates were biofilm positive and their biofilm forms, were highly resistant. In this study, compared to PCR, culture remains the gold standard. The biofilm formation by the causative bacteria and the concomitant manifold increased antimicrobial resistance may explain the clinical failure of treatment in some cases and should be considered in the future for therapeutic planning.

  14. [Surviving Forms in Antibiotic-Treated Pseudomonas aeruginosa].

    Science.gov (United States)

    Mulyukin, A L; Kozlova, A N; Sorokin, V V; Suzina, N E; Cherdyntseva, T A; Kotova, I B; Gaponov, A M; Tutel'yan, A V; El'-Registan, G I

    2015-01-01

    Survival of bacterial populations treated with lethal doses of antibiotics is ensured by the presence of very small numbers of persister cells. Unlike antibiotic-resistant cells, antibiotic tolerance of persisters is not inheritable and reversible. The present work provides evidence supporting the hypothesis of transformation (maturation) of persisters of an opportunistic pathogen Pseudomonas aeruginosa revealed by ciprofloxacin (CF) treatment (25-100 μg/mL) into dormant cystlike cells (CLC) and non-culturable cells (NC), as was described previously for a number. of non-spore-forming bacteria. Subpopulations of type 1 and type 2 persisters, which survived antibiotic treatment and developed into dormant forms, were heterogeneous in their capacity to form colonies or microcolonies upon germination, in resistance to heating at 70 degrees C, and in cell morphology Type 1 persisters, which were formed after 1-month incubation in the stationary-phase cultures in the medium with decreased C and N concentrations, developed in several types of surviving cells, including those similar to CLC in cell morphology. In the course of 1-month incubation of type 2 persisters, which were formed in exponentially growing cultures, other types of surviving cells developed: immature CLC and L-forms. Unlike P. aeruginosa CLC formed in the control post-stationary phase cultures without antibiotic treatment, most of 1-month persisters, especially type 2 ones, were characterized by the loss of colony-forming capacity, probably due to transition into an uncultured state with relatively high numbers of live intact cells (Live/Dead test). Another survival strategy of P. aeruginosa populations was ensured by a minor subpopulation of CF-tolerant and CF-resistant cells able to grow in the form of microcolonies or regular colonies of decreased size in the presence of the antibiotic. The described P. aeruginosa dormant forms may be responsible for persistent forms in bacteria carriers and latent

  15. Pseudomonas aeruginosa biofilms exposed to imipenem exhibit changes in global gene expression and beta-lactamase and alginate production

    DEFF Research Database (Denmark)

    Bagge, Niels; Schuster, Martin; Hentzer, Morten

    2004-01-01

    . The treatment often includes beta-lactam antibiotics. How these antibiotics influence gene expression in the surviving biofilm population of P. aeruginosa is not clear. Thus, we used the microarray technology to study the effects of subinhibitory concentrations of a beta-lactam antibiotic, imipenem, on gene...

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

    DEFF Research Database (Denmark)

    Jensen, Peter Ø.; 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...

  17. 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

    2017-01-01

    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. DOI: http://dx.doi.org/10.7554/eLife.21855.001 PMID:28084994

  18. Effect of Silver Nanoparticles Against the Formation of Biofilm by Pseudomonas aeruginosa an In silico Approach.

    Science.gov (United States)

    Vyshnava, Satyanarayana Swamy; Kanderi, Dileep Kumar; Panjala, Shiva Prasad; Pandian, Kamesh; Bontha, Rajasekhar Reddy; Goukanapalle, Praveen Kumar Reddy; Banaganapalli, Babajan

    2016-10-01

    Studies were undertaken to examine the mechanism of mediation of silver nanoparticles in inhibiting biofilm formation by Pseudomonas aeruginosa through LuxI/LuxR system of signal transduction. This study includes the basic signaling transduction mechanism LasR, QscR, RhlR, and Vfr signaling model systems. The arbitrary homology models built with the I-TASSER server were evaluated and validated with the Qmean web server. Based on the Z-score and the relative square mean distance (RMSD) values, the structures were validated. The interaction results of the nanoparticle with the rigid docking proved the requirement of minimal energy for the inhibition of the protein active site by the silver nanoparticle. This principle docking experiment suggests that the biofilm formation in Gram-negative bacteria can be inhibited by the silver nanoparticles at the signal transduction level. Graphical abstract Systematic outline of present study; Stage one provides the data sampling and generation of pdb systems to conform the structure of bacterial signal sytems like LasR/LasI; RhlR/RhrI; QscR/QscI; VfrR/VfrI. Stage two involves docking of silver nanoparticles with Bacterial signal protein strucutres which are listed in Stage one. The Final Stage involves in understanding the development of appropriate mechanism behind the biofilm inhibition by silver nanoparticles.

  19. 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.

  20. Raffinose, a plant galactoside, inhibits Pseudomonas aeruginosa biofilm formation via binding to LecA and decreasing cellular cyclic diguanylate levels

    Science.gov (United States)

    Kim, Han-Shin; Cha, Eunji; Kim, Yunhye; Jeon, Young Ho; Olson, Betty H.; Byun, Youngjoo; Park, Hee-Deung

    2016-05-01

    Biofilm formation on biotic or abiotic surfaces has unwanted consequences in medical, clinical, and industrial settings. Treatments with antibiotics or biocides are often ineffective in eradicating biofilms. Promising alternatives to conventional agents are biofilm-inhibiting compounds regulating biofilm development without toxicity to growth. Here, we screened a biofilm inhibitor, raffinose, derived from ginger. Raffinose, a galactotrisaccharide, showed efficient biofilm inhibition of Pseudomonas aeruginosa without impairing its growth. Raffinose also affected various phenotypes such as colony morphology, matrix formation, and swarming motility. Binding of raffinose to a carbohydrate-binding protein called LecA was the cause of biofilm inhibition and altered phenotypes. Furthermore, raffinose reduced the concentration of the second messenger, cyclic diguanylate (c-di-GMP), by increased activity of a c-di-GMP specific phosphodiesterase. The ability of raffinose to inhibit P. aeruginosa biofilm formation and its molecular mechanism opens new possibilities for pharmacological and industrial applications.

  1. Effect of Cinnamon Oil on Quorum Sensing-Controlled Virulence Factors and Biofilm Formation in Pseudomonas aeruginosa.

    Science.gov (United States)

    Kalia, Manmohit; Yadav, Vivek Kumar; Singh, Pradeep Kumar; Sharma, Deepmala; Pandey, Himanshu; Narvi, Shahid Suhail; Agarwal, Vishnu

    2015-01-01

    Quorum sensing (QS) is a system of stimuli and responses in bacterial cells governed by their population density, through which they regulate genes that control virulence factors and biofilm formation. Despite considerable research on QS and the discovery of new antibiotics, QS-controlled biofilm formation by microorganisms in clinical settings has remained a problem because of nascent drug resistance, which requires screening of diverse compounds for anti-QS activities. Cinnamon is a dietary phytochemical that is traditionally used to remedy digestive problems and assorted contagions, which suggests that cinnamon might contain chemicals that can hinder the QS process. To test this hypothesis, the anti-QS activity of cinnamon oil against P. aeruginosa was tested, measured by the inhibition of biofilm formation and other QS-associated phenomena, including virulence factors such as pyocyanin, rhamnolipid, protease, alginate production, and swarming activity. To this end, multiple microscopy analyses, including light, scanning electron and confocal microscopy, revealed the ability of cinnamon oil to inhibit P. aeruginosa PAO1 biofilms and their accompanying extracellular polymeric substances. This work is the first to demonstrate that cinnamon oil can influence various QS-based phenomena in P. aeruginosa PAO1, including biofilm formation.

  2. Rhamnolipid but not motility is associated with the initiation of biofilm seeding dispersal of Pseudomonas aeruginosa strain PA17

    Indian Academy of Sciences (India)

    Jingjing Wang; Bing Yu; Deying Tian; Ming Ni

    2013-03-01

    Seeding dispersal is an active detachment exhibit in aging Pseudomonas aeruginosa biofilm. Yet, effect factors of this process in the biofilm of clinical isolated mucoid P. aeruginosa strain remain to be better characterized. In our previous work, one mucoid P. earuginosa strain PA17 was isolated from a patient with recurrent pulmonary infection. In this study, confocal scanning laser microscope combined with LIVE/DEAD viability staining revealed that PA17 biofilm exhibited earlier seeding dispersal than non-mucoid PAO1. We further compared the motility and the expression of motility-associated gene during biofilm development between PA17 and PAO1. PA17 was found to be impaired in all three kinds of motility compared to PAO1. Moreover, we investigated the expression of rhamnolipid-associated genes in PA17 and PAO1 biofilm. The expression of these genes was in accordance with the process of seeding dispersal. Our results indicated that rhamnolipid but not motility is associated with the initiation of seeding dispersal of PA17 biofilm.

  3. A multivariate approach to correlate bacterial surface properties to biofilm formation by lipopolysaccharide mutants of Pseudomonas aeruginosa.

    Science.gov (United States)

    Ruhal, Rohit; Antti, Henrik; Rzhepishevska, Olena; Boulanger, Nicolas; Barbero, David R; Wai, Sun Nyunt; Uhlin, Bernt Eric; Ramstedt, Madeleine

    2015-03-01

    Bacterial biofilms are involved in various medical infections and for this reason it is of great importance to better understand the process of biofilm formation in order to eradicate or mitigate it. It is a very complex process and a large range of variables have been suggested to influence biofilm formation. However, their internal importance is still not well understood. In the present study, a range of surface properties of Pseudomonas aeruginosa lipopolysaccharide mutants were studied in relation to biofilm formation measured in different kinds of multi-well plates and growth conditions in order to better understand the complexity of biofilm formation. Multivariate analysis was used to simultaneously evaluate the role of a range of physiochemical parameters under different conditions. Our results suggest the presence of serum inhibited biofilm formation due to changes in twitching motility. From the multivariate analysis it was observed that the most important parameters, positively correlated to biofilm formation on two types of plates, were high hydrophobicity, near neutral zeta potential and motility. Negative correlation was observed with cell aggregation, as well as formation of outer membrane vesicles and exopolysaccharides. This work shows that the complexity of biofilm formation can be better understood using a multivariate approach that can interpret and rank the importance of different factors being present simultaneously under several different environmental conditions, enabling a better understanding of this complex process.

  4. A Microfluidic Approach to Investigating a Synergistic Effect of Tobramycin and Sodium Dodecyl Sulfate on Pseudomonas aeruginosa Biofilms.

    Science.gov (United States)

    Shin, Soojeong; Ahmed, Ishtiaq; Hwang, Jangsun; Seo, Youngmin; Lee, Eunwon; Choi, Jonghoon; Moon, Sangjun; Hong, Jong Wook

    2016-01-01

    In recent years, a microfluidic technology has contributed a significant role in biological research, specifically for the study of biofilms. Bacterial biofilms are a source of infections and contamination in the environment due to an extra polymeric matrix. Inadequate uses of antibiotics make the bacterial biofilms antibiotic resistant. Therefore, it is important to determine the effective concentration of antibiotics in order to eliminate bacterial biofilms. The present microfluidic study was carried out to analyze the activities of tobramycin and sodium dodecyl sulfate (SDS) against Pseudomonas aeruginosa biofilms with a continuous flow in order to achieve a greater delivery of the agents. The results show that a co-treatment of tobramycin and SDS significantly reduced the biomass of biofilms (by more than 99%) after 24 h. Tobramycin and SDS killed and detached bacteria in the cores of biofilms. Evidently, our data suggest that a microchannel would be effective for both quantitative and qualitative evaluations in order to test combinatorial effect of drugs and chemicals on a complexed biological system including biofilm.

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

    Science.gov (United States)

    Chua, Song Lin; Sivakumar, Krishnakumar; Rybtke, Morten; Yuan, Mingjun; Andersen, Jens Bo; Nielsen, Thomas E; Givskov, Michael; Tolker-Nielsen, Tim; Cao, Bin; Kjelleberg, Staffan; Yang, Liang

    2015-01-01

    Stress response plays an important role on microbial adaptation under hostile environmental conditions. It is generally unclear how the signaling transduction pathway mediates a stress response in planktonic and biofilm modes of microbial communities simultaneously. Here, we showed that metalloid 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.

  6. Polymicrobial Ventilator-Associated Pneumonia: Fighting In Vitro Candida albicans-Pseudomonas aeruginosa Biofilms with Antifungal-Antibacterial Combination Therapy

    Science.gov (United States)

    Pereira, Cláudia R.; Azevedo, Nuno F.; Lourenço, Anália; Henriques, Mariana; Pereira, Maria O.

    2017-01-01

    The polymicrobial nature of ventilator-associated pneumonia (VAP) is now evident, with mixed bacterial-fungal biofilms colonizing the VAP endotracheal tube (ETT) surface. The microbial interplay within this infection may contribute for enhanced pathogenesis and exert impact towards antimicrobial therapy. Consequently, the high mortality/morbidity rates associated to VAP and the worldwide increase in antibiotic resistance has promoted the search for novel therapeutic strategies to fight VAP polymicrobial infections. Under this scope, this work aimed to assess the activity of mono- vs combinational antimicrobial therapy using one antibiotic (Polymyxin B; PolyB) and one antifungal (Amphotericin B; AmB) agent against polymicrobial biofilms of Pseudomonas aeruginosa and Candida albicans. The action of isolated antimicrobials was firstly evaluated in single- and polymicrobial cultures, with AmB being more effective against C. albicans and PolyB against P. aeruginosa. Mixed planktonic cultures required equal or higher antimicrobial concentrations. In biofilms, only PolyB at relatively high concentrations could reduce P. aeruginosa in both monospecies and polymicrobial populations, with C. albicans displaying only punctual disturbances. PolyB and AmB exhibited a synergistic effect against P. aeruginosa and C. albicans mixed planktonic cultures, but only high doses (256 mg L-1) of PolyB were able to eradicate polymicrobial biofilms, with P. aeruginosa showing loss of cultivability (but not viability) at 2 h post-treatment, whilst C. albicans only started to be inhibited after 14 h. In conclusion, combination therapy involving an antibiotic and an antifungal agent holds an attractive therapeutic option to treat severe bacterial-fungal polymicrobial infections. Nevertheless, optimization of antimicrobial doses and further clinical pharmacokinetics/pharmacodynamics and toxicodynamics studies underpinning the optimal use of these drugs are urgently required to improve therapy

  7. Detection and imaging of quorum sensing in Pseudomonas aeruginosa biofilm communities by surface-enhanced resonance Raman scattering

    Science.gov (United States)

    Bodelón, Gustavo; Montes-García, Verónica; López-Puente, Vanesa; Hill, Eric H.; Hamon, Cyrille; Sanz-Ortiz, Marta N.; Rodal-Cedeira, Sergio; Costas, Celina; Celiksoy, Sirin; Pérez-Juste, Ignacio; Scarabelli, Leonardo; La Porta, Andrea; Pérez-Juste, Jorge; Pastoriza-Santos, Isabel; Liz-Marzán, Luis M.

    2016-11-01

    Most bacteria in nature exist as biofilms, which support intercellular signalling processes such as quorum sensing (QS), a cell-to-cell communication mechanism that allows bacteria to monitor and respond to cell density and changes in the environment. As QS and biofilms are involved in the ability of bacteria to cause disease, there is a need for the development of methods for the non-invasive analysis of QS in natural bacterial populations. Here, by using surface-enhanced resonance Raman scattering spectroscopy, we report rationally designed nanostructured plasmonic substrates for the in situ, label-free detection of a QS signalling metabolite in growing Pseudomonas aeruginosa biofilms and microcolonies. The in situ, non-invasive plasmonic imaging of QS in biofilms provides a powerful analytical approach for studying intercellular communication on the basis of secreted molecules as signals.

  8. Biofilms and Cyclic di-GMP (c-di-GMP) Signaling: Lessons from Pseudomonas aeruginosa and Other Bacteria.

    Science.gov (United States)

    Valentini, Martina; Filloux, Alain

    2016-06-10

    The cyclic di-GMP (c-di-GMP) second messenger represents a signaling system that regulates many bacterial behaviors and is of key importance for driving the lifestyle switch between motile loner cells and biofilm formers. This review provides an up-to-date compendium of c-di-GMP pathways connected to biofilm formation, biofilm-associated motilities, and other functionalities in the ubiquitous and opportunistic human pathogen Pseudomonas aeruginosa This bacterium is frequently adopted as a model organism to study bacterial biofilm formation. Importantly, its versatility and adaptation capabilities are linked with a broad range of complex regulatory networks, including a large set of genes involved in c-di-GMP biosynthesis, degradation, and transmission.

  9. Interspecies signalling via the Stenotrophomonas maltophilia diffusible signal factor influences biofilm formation and polymyxin tolerance in Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Ryan, R.P.; Fouhy, Y.; Garcia, B.F.

    2008-01-01

    including the rhizosphere of plants and the cystic fibrosis lung. In mixed species biofilms, S. maltophilia substantially influenced the architecture of P. aeruginosa structures, which developed as extended filaments. This effect depended upon the synthesis of the diffusible signal factor (DSF) by S....... maltophilia and could be mimicked by the addition of synthetic DSF. This response of P. aeruginosa to DSF required PA1396, a sensor kinase with an input domain of related amino acid sequence to the sensory input domain of RpfC, which is responsible for DSF perception in xanthomonads. Mutation of PA1396...

  10. 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.

  11. Pseudomonas aeruginosa outcompetes other bacteria in the manifestation and maintenance of a biofilm in polyvinylchloride tubing as used in dental devices.

    Science.gov (United States)

    Ammann, Christoph Gert; Nagl, Markus; Nogler, Michael; Coraça-Huber, Débora Cristina

    2016-05-01

    In a PVC tube as a model system for dental devices, Pseudomonas aeruginosa outcompetes Staphylococcus aureus and Klebsiella pneumoniae for the biofilm formation. P. aeruginosa has advantage over the other strains due to higher tolerance for low-nutrient situations or direct killing by the production of soluble factors like pyocyanin.

  12. D-amino acids enhance the activity of antimicrobials against biofilms of clinical wound isolates of Staphylococcus aureus and Pseudomonas aeruginosa.

    Science.gov (United States)

    Sanchez, Carlos J; Akers, Kevin S; Romano, Desiree R; Woodbury, Ronald L; Hardy, Sharanda K; Murray, Clinton K; Wenke, Joseph C

    2014-08-01

    Within wounds, microorganisms predominantly exist as biofilms. Biofilms are associated with chronic infections and represent a tremendous clinical challenge. As antibiotics are often ineffective against biofilms, use of dispersal agents as adjunctive, topical therapies for the treatment of wound infections involving biofilms has gained interest. We evaluated in vitro the dispersive activity of D-amino acids (D-AAs) on biofilms from clinical wound isolates of Staphylococcus aureus and Pseudomonas aeruginosa; moreover, we determined whether combinations of D-AAs and antibiotics (clindamycin, cefazolin, oxacillin, rifampin, and vancomycin for S. aureus and amikacin, colistin, ciprofloxacin, imipenem, and ceftazidime for P. aeruginosa) enhance activity against biofilms. D-Met, D-Phe, and D-Trp at concentrations of ≥ 5 mM effectively dispersed preformed biofilms of S. aureus and P. aeruginosa clinical isolates, an effect that was enhanced when they were combined as an equimolar mixture (D-Met/D-Phe/D-Trp). When combined with D-AAs, the activity of rifampin was significantly enhanced against biofilms of clinical isolates of S. aureus, as indicated by a reduction in the minimum biofilm inhibitory concentration (MBIC) (from 32 to 8 μg/ml) and a >2-log reduction of viable biofilm bacteria compared to treatment with antibiotic alone. The addition of D-AAs was also observed to enhance the activity of colistin and ciprofloxacin against biofilms of P. aeruginosa, reducing the observed MBIC and the number of viable bacteria by >2 logs and 1 log at 64 and 32 μg/ml in contrast to antibiotics alone. These findings indicate that the biofilm dispersal activity of D-AAs may represent an effective strategy, in combination with antimicrobials, to release bacteria from biofilms, subsequently enhancing antimicrobial activity.

  13. Evaluation of antibiofilm effect of benzalkonium chloride, iodophore and sodium hypochlorite against biofilm of Pseudomonas aeruginosa of dairy origin.

    Science.gov (United States)

    Pagedar, Ankita; Singh, Jitender

    2015-08-01

    The present study was undertaken with objectives of; a) to investigate and compare Pseudomonas aeruginosa isolates from two dairies for biofilm formation potential and, b) to compares three common biocides for biofilm eradication efficiencies. Amongst the isolates from commercial dairy, 70 % were strong and/or moderate biofilm former in comparison to 40 % isolates from small scale dairy. All isolates, irrespective of source, exhibited higher susceptibility to biocides in planktonic stage than in biofilm. Antibiofilm efficiencies of three biocides i.e. benzalkonium chloride, sodium hypochlorite and iodophore were determined in terms of their microbial biofilms eradicating concentration (MBEC). Our findings show that the three biocides were ineffective against preformed biofilms at recommended in-use concentrations. Biofilms were the most resistant to benzalkonium chloride and least against iodophore. A trend of decreasing MBECs was observed with extended contact time. The findings of present study warrant for a systematic approach for selecting types and concentrations of biocide for application as antibiofilm agent in food industry.

  14. 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.

  15. Assessment of the working range and effect of sodium dichloroisocyanurate on Pseudomonas aeruginosa biofilms and planktonic cells.

    Science.gov (United States)

    Morgenthau, Ari; Nicolae, Alexandru M; Laursen, Andrew E; Foucher, Daniel A; Wolfaardt, Gideon M; Hausner, Martina

    2012-01-01

    Sodium dichloroisocyanurate (NaDCC) is a chemical agent that acts against microorganisms in a manner similar to that of sodium hypochlorite by releasing free available chlorine. NaDCC has been approved by the WHO for the emergency treatment of water and by the US EPA for routine treatment of water. Previous studies assessing the effectiveness of NaDCC for the treatment of water implied that NaDCC should have a wide array of disinfecting effects beyond the treatment of planktonic cells in potable water. In this study the biocidal effects of NaDCC against Pseudomonas aeruginosa cells in different growth modes including planktonic cells and biofilms were explored. The data showed that a 60% dilution of the standard NaDCC solution was effective in the treatment of both P. aeruginosa planktonic cells and biofilms.

  16. 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...... of the majority of the bacteria in in vitro-grown P. aeruginosa biofilms. Subsequently, we demonstrated that P. aeruginosa biofilms growing on silicone implants, located in the peritoneal cavity of mice, dispersed after induction of the YhjH protein. Bacteria accumulated temporarily in the spleen after induction...

  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. Characterization of biofilm formed on intrauterine devices

    OpenAIRE

    2003-01-01

    PURPOSE: Intrauterine device (IUD) is one of the most convenient contraceptive procedures used by women of Asian and African countries. Previous surveys have revealed that 75% of the IUDs recovered from patients suffering from reproductive tract infections (RTIs) were covered with a consortium of microbes. This study was designed to characterize these microbes and recommend remedial measures. METHODS: Quantitative measurement of biofilm formation was assessed by a microtitre plate assa...

  19. Interactions between the antimicrobial agent triclosan and the bloom-forming cyanobacteria Microcystis aeruginosa

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Xiaolong [State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072 (China); Wuhan Zhongke Hydrobiological Environment Engineering Co., Ltd, Wuhan 430071 (China); Tu, Yenan; Song, Chaofeng; Li, Tiancui; Lin, Juan [State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072 (China); Graduate University of the Chinese Academy of Sciences, Beijing 100049 (China); Wu, Yonghong [State Key Laboratory of Soil and Sustainable Agriculture, Institute of Soil Sciences, Chinese Academy of Sciences, Nanjing 210008 (China); Liu, Jiantong [State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072 (China); Wu, Chenxi, E-mail: chenxi.wu@ihb.ac.cn [State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072 (China)

    2016-03-15

    Highlights: • Triclosan inhibit the growth and photosynthesis of M. aeruginosa at environmental relevant level. • TEM imaging showed destruction of M. aeruginosa cell ultrastructure during triclosan exposure. • Triclosan can be biotransformed by M. aeruginosa with methylation as a major pathway. • Presence of M. aeruginosa enhanced the photodegradation of triclosan. - Abstract: Cyanobacteria can co-exist in eutrophic waters with chemicals or other substances derived from personal care products discharged in wastewater. In this work, we investigate the interactions between the antimicrobial agent triclosan (TCS) and the bloom-forming cyanobacteria Microcystis aeruginosa. M. aeruginosa was very sensitive to TCS with the 96 h lowest observed effect concentration of 1.0 and 10 μg/L for inhibition of growth and photosynthetic activity, respectively. Exposure to TCS at environmentally relevant levels (0.1–2.0 μg/L) also affected the activities of superoxide dismutase (SOD) and the generation of reduced glutathione (GSH), while microcystin production was not affected. Transmission electron microscope (TEM) examination showed the destruction of M. aeruginosa cell ultrastructure during TCS exposure. TCS however, can be biotransformed by M. aeruginosa with methylation as a major biotransformation pathway. Furthermore, the presence of M. aeruginosa in solution promoted the photodegradation of TCS. Overall, our results demonstrate that M. aeruginosa plays an important role in the dissipation of TCS in aquatic environments but high residual TCS can exert toxic effects on M. aeruginosa.

  20. 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.

  1. 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.

  2. The Study of Synergistic Effects of n.butanolic Cyclamen coum Extract and Ciprofloxacin on inhibition of Pseudomonas aeruginosa biofilm formation

    Directory of Open Access Journals (Sweden)

    ahya abdi ali

    2015-02-01

    Full Text Available   Introduction : Infections caused by Pseudomonas aeruginosa biofilm are the major causes of death in patients with cystic fibrosis (CF. Some studies revealed that biofilms are resistant to several antibiotics because of their impermeable structures. In order to re-sensitize bacteria to different antibiotics, biofilm formation should be inhibited. In this research, evaluation of antibiofilm activity of n-butanolic Cyclamen coum extract as a medici­nal plant from Myrsinaceae family, in combination with ciprofloxacin was carried out.   Materials and method s: The biofilm formation ability by P. aeruginosa PAO1 and one clinically isolated P. aeruginosa (PA214 was confirmed by microtiter plate method. Extraction of the tubers of Cyclamen coum was done by fractionation method . The antibiofilm and antibacterial properties of n-butanolic C. coum extract (which includes saponin compounds alone and in combination with ciprofloxacin by using microdilution and crystal violet methods were examined. The cytotoxicity effect of the n-butanolic extract on HT-29 cells was assayed by MTT (3- (4,5-dimethylthiazol-2-yl -2,5-diphenyl-tetrazolium bromide test.   Results : The biofilm formation ability by P. aeruginosa strains was quantitatively confirmed. Saponin content of the n-butanolic C.coum extract was 156 µg/mL. The extract revealed antibacterial activity against the growth of planktonic P. aeruginosa strains. The combination of n-butanolic C.coum extract and ciprofloxacin significantly inhibited P.aeruginosa biofilm formation (ΣFBIC = 0.5. The n-butanolic C.coum extract showed insignificant cytotoxic effect against HT-29 human cancer cell line after 48 hours and 72 hours incubation .   Discussion and conclusion : It can be concluded that n-butanolic C.coum extract in combination with ciprofloxacin significantly revealed antibiofilm activity against P. aeruginosa biofilm however, further clinical investigations are required.

  3. 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...

  4. Does Campylobacter jejuni form biofilms in food-related environments?

    Science.gov (United States)

    Teh, Amy Huei Teen; Lee, Sui Mae; Dykes, Gary A

    2014-09-01

    Campylobacter jejuni is one of the most frequent causes of bacterial gastrointestinal food-borne infection worldwide. This species is part of the normal flora of the gastrointestinal tracts of animals used for food production, including poultry, which is regarded as the primary source of human Campylobacter infections. The survival and persistence of C. jejuni in food processing environments, especially in poultry processing plants, represent significant risk factors that contribute to the spread of this pathogen through the food chain. Compared to other food-borne pathogens, C. jejuni is more fastidious in its growth requirements and is very susceptible to various environmental stressors. Biofilm formation is suggested to play a significant role in the survival of C. jejuni in the food production and processing environment. The aims of this minireview were (i) to examine the evidence that C. jejuni forms biofilms and (ii) to establish the extent to which reported and largely laboratory-based studies of C. jejuni biofilms provide evidence for biofilm formation by this pathogen in food processing environments. Overall existing studies do not provide strong evidence for biofilm formation (as usually defined) by most C. jejuni strains in food-related environments under the combined conditions of atmosphere, temperature, and shear that they are likely to encounter. Simple attachment to and survival on surfaces and in existing biofilms of other species are far more likely to contribute to C. jejuni survival in food-related environments based on our current understanding of this species.

  5. 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.

  6. In vitro and in vivo generation and characterization of Pseudomonas aeruginosa biofilm-dispersed cells via c-di-GMP manipulation.

    Science.gov (United States)

    Chua, Song Lin; Hultqvist, Louise D; Yuan, Mingjun; Rybtke, Morten; Nielsen, Thomas E; Givskov, Michael; Tolker-Nielsen, Tim; Yang, Liang

    2015-08-01

    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. However, freshly dispersed biofilm cells exhibit a physiology distinct from biofilm and planktonic cells, and they might have a clinically relevant role in infections. Here we present in vitro and in vivo protocols for the generation and characterization of dispersed cells from Pseudomonas aeruginosa 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 are identified and quantified, serving as potential targets for treating the dispersed cells. The in vitro protocol can be completed within 4 d, whereas the in vivo protocol requires 7 d.

  7. Effect of Cinnamomum burmannii Nees ex Bl. and Massoia aromatica Becc. Essential Oils on Planktonic Growth and Biofilm formation of Pseudomonas aeruginosa and Staphylococcus aureus In Vitro

    Directory of Open Access Journals (Sweden)

    Sylvia Utami Tunjung Pratiwi

    2015-03-01

    Full Text Available Summary. Biofilms are communities of microorganisms that can be found in almost every habitat. They can be attached to a surface and protected by an extracellular matrix of biomolecules that substantially protect microorganisms from environmental effects. The aim of this research is to explore the potency of essential oils from Cinnamomum burmannii Nees ex Bl. and Massoia aromatica Becc. against planktonic growth and biofilm formation of, two opportunistic pathogens, Pseudomonas aeruginosa PAO1 and Staphylococcus aureus Cowan I. Essential oil from C. burmannii  and M. aromatica showed a 50% inhibition of  P. aeruginosa and S. aureus planktonic growth (PMIC50 at concentration of 0.12 % v/v. Essential oil from C. burmannii and M.  aromatica showed capability to inhibit 50% (MBIC50 of P. aeruginosa and S. aureus biofilm formation at concentration of 0.03 % v/v, whereas higher concentration (0.12 % v/v was needed by C. burmannii and M. aromatica oil to disrupt 50% of P. aeruginosa and S. aureus established biofilm. The analysis by GC-MS showed cinnamic aldehyde (92.02 % to be the major component of C. burmannii essential oil, whereas Massoialactone (92.05 % was the main constituent of M. aromatica essential oil. The results obtained in this study have made the oil of C. burmannii and M. aromatica oil as an interesting source for antibiofilm agents in the development of new strategies to treat infections caused by P. aeruginosa and  S. aureus biofilm.Industrial Relevance. Instead of freely swimming in solution (planktonic, in nature microbial tends to adhere to surfaces, and develop microbial biofilms. Microbial biofilms are exhibits resistance to both antimicrobial drugs and the host defence systems, which often results in persistent and difficult-to-treat infections. This makes the discovery of anti-infective agents which are active against planktonic and biofilm microbial represents an important goal. Plant is an interesting source for finding

  8. 黄芩苷对铜绿假单胞菌生物膜的影响%Effect of Baicalin on Pseudomonas Aeruginosa Biofilms

    Institute of Scientific and Technical Information of China (English)

    王贵年; 范莹; 王龙梓; 吴娟

    2011-01-01

    Objective To investigate the effect of baicalin on Pseudomonas aeruginosa biofilms in vitro. Methods Clinical isolates of Pseudomonas aeruginosa were cuhured in Luria - Bertani medium - aspirate sputum pipe system to establish biofilm formation. Biofilm was observed in AgNO3 staining. Viable bacterial counts were determined by serial dilution. MICs were measured by doubling dilution.Results The biofilm was inhibited and destructed by 15. 65mg/L of baicalin. Conclusion Baicalin displayed potent activity against Pseudomonas aeruginosa biofilms.%目的 研究黄芩苷对体外铜绿假单胞菌(pseudomonas aeruginosa,Pa)生物膜(biofilm,BF)的影响.方法 选取临床分离呼吸道Pa,采用LB肉汤-吸痰管系统培养BF,建立体外BF模型,银染法观察BF变化.将黄芩苷作用于BF,连续稀释法进行活菌计数,微量稀释法测定最低抑菌浓度(MIC).结果 15.65mg/L的黄芩苷即可抑制和破坏BF.结论 黄芩苷对体外铜绿假单胞菌生物膜有较强的抑制作用.

  9. Evaluation of antibiotic efficacy against infections caused by planktonic or biofilm cultures of Pseudomonas aeruginosa and Klebsiella pneumoniae in Galleria mellonella.

    Science.gov (United States)

    Benthall, Gabriel; Touzel, Rebecca E; Hind, Charlotte K; Titball, Richard W; Sutton, J Mark; Thomas, Rachael J; Wand, Matthew E

    2015-11-01

    The lack of novel antibiotics for more than a decade has placed increased pressure on existing therapies to combat the emergence of multidrug-resistant (MDR) bacterial pathogens. This study evaluated the Galleria mellonella insect model in determining the efficacy of available antibiotics against planktonic and biofilm infections of MDR Pseudomonas aeruginosa and Klebsiella pneumoniae strains in comparison with in vitro minimum inhibitory concentration (MIC) determination. In general, in vitro analysis agreed with the G. mellonella studies, and susceptibility in Galleria identified different drug resistance mechanisms. However, the carbapenems tested appeared to perform better in vivo than in vitro, with meropenem and imipenem able to clear K. pneumoniae and P. aeruginosa infections with strains that had bla(NDM-1) and bla(VIM) carbapenemases. This study also established an implant model in G. mellonella to allow testing of antibiotic efficacy against biofilm-derived infections. A reduction in antibiotic efficacy of amikacin against K. pneumoniae and P. aeruginosa biofilms was observed compared with a planktonic infection. Ciprofloxacin was found to be less effective at clearing a P. aeruginosa biofilm infection compared with a planktonic infection, but no statistical difference was seen between K. pneumoniae biofilm and planktonic infections treated with this antibiotic (P>0.05). This study provides important information regarding the suitability of Galleria as a model for antibiotic efficacy testing both against planktonic and biofilm-derived MDR infections.

  10. Confocal laser scanning microscopy for assessment of pseudomonas aeruginosa biofilm formation%激光共聚焦显微镜观察铜绿假单胞菌生物膜的形成

    Institute of Scientific and Technical Information of China (English)

    王忠; 季文; 张瑞琴

    2013-01-01

    Objective To investigate the Pseudomonas aeruginosa biofilm by using confocal laser scanning microscopy (CLSM), thus revealing the formation of biofilm. Methods The cover slide biofilm culture approach was employed for induction of Pseudomonas aeruginosa biofilm formation. Following the culture for 2, 4, 8, 12, 16, 24, 48 and 72 hours, the cover slide was removed for subsequent staining with the fluoreseein isothiocyanate-labeled con-eanavalin A (FTTC-ConA) and iodidepyrtdine (PI)- This was followed by determination of the formation and characteristics of Pseudomonas aeruginosa biofilm by using CXSM. Results The CLSM images of biofilm formation at different time points were captured, suggesting that the biofilm formed at hour 24 and grew robustly at hour 72. Conclusion The double immunofluorescence staining and CLSM are effective means for assessment of Pseudomonas aeruginosa biofilm formation.%目的 采用激光共聚焦显微镜技术观察铜绿假单胞菌形成生物膜,进一步揭示了铜绿假单胞菌生物膜形成过程.方法 使用盖玻片生物膜培养法,培养铜绿假单胞菌的生物膜,在培养2、4、8、12、16、24、48和72 h后取出盖玻片,用异硫氰酸荧光素(FTTC)标记的刀豆蛋白A(FTTC-ConA)和碘化吡啶(PI)双重免疫荧光技术染色,用激光共聚焦显微镜(CLSM)观察铜绿假单胞菌生物膜形成过程与特点.结果 获得生物膜形成过程不同时间点的CLSM图像,观察到铜绿假单胞菌一般在24h后开始逐渐形成生物膜,在72 h形成稳定的生物膜.结论 双重免疫荧光染色技术和CLSM是观察细菌生物膜形成过程的有效手段.

  11. Mechanistic insights into c-di-GMP–dependent control of the biofilm regulator FleQ from Pseudomonas aeruginosa

    Energy Technology Data Exchange (ETDEWEB)

    Matsuyama, Bruno Y.; Krasteva, Petya V.; Baraquet, Claudine; Harwood, Caroline S.; Sondermann, Holger; Navarro, Marcos V.A. S. (UWASH); (U. Sao Paulo); (Cornell); (CNRS-UMR)

    2016-07-05

    Pseudomonas aeruginosa, an opportunistic pathogen that can cause fatal chronic infections, relies on the intracellular second-messenger c-di-GMP to form robust multicellular biofilms during host tissue colonization. c-di-GMP is sensed directly by the transcription regulator FleQ, which inversely regulates flagellar motility and exopolysaccharide secretion to secure a planktonic to sessile life-form transition. FleQ belongs to the diverse family of AAA+ ATPase enhancer-binding proteins, but how its noncanonical function on transcriptional regulation is controlled by c-di-GMP remains enigmatic. Here, we report structural and functional data that identify an unusual mode of c-di-GMP recognition accompanied by a major quaternary structure reorganization. Our analyses offer a consensus to previous studies and unique insights into the mechanism of action of FleQ and FleQ-like proteins.

  12. 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

    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...

  13. Pseudomonas aeruginosa tolerance to tobramycin, hydrogen peroxide and polymorphonuclear leukocytes is quorum-sensing dependent

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Jensen, Peter Østrup; Burmølle, Mette;

    2005-01-01

    The opportunistic human pathogen Pseudomonas aeruginosa is the predominant micro-organism of chronic lung infections in cystic fibrosis (CF) patients. P. aeruginosa colonizes the CF lungs by forming biofilm structures in the alveoli. In the biofilm mode of growth the bacteria are highly tolerant...

  14. Unsaturated fatty acid, cis-2-decenoic acid, in combination with disinfectants or antibiotics removes pre-established biofilms formed by food-related bacteria.

    Science.gov (United States)

    Sepehr, Shayesteh; Rahmani-Badi, Azadeh; Babaie-Naiej, Hamta; Soudi, Mohammad Reza

    2014-01-01

    Biofilm formation by food-related bacteria and food-related pathogenesis are significant problems in the food industry. Even though much disinfection and mechanical procedure exist for removal of biofilms, they may fail to eliminate pre-established biofilms. cis-2 decenoic acid (CDA), an unsaturated fatty acid messenger produced by Pseudomonas aeruginosa, is reportedly capable of inducing the dispersion of established biofilms by multiple types of microorganisms. However, whether CDA has potential to boost the actions of certain antimicrobials is unknown. Here, the activity of CDA as an inducer of pre-established biofilms dispersal, formed by four main food pathogens; Staphylococcus aureus, Bacillus cereus, Salmonella enterica and E. coli, was measured using both semi-batch and continuous cultures bioassays. To assess the ability of CDA combined biocides treatments to remove pre-established biofilms formed on stainless steel discs, CFU counts were performed for both treated and untreated cultures. Eradication of the biofilms by CDA combined antibiotics was evaluated using crystal violet staining. The effect of CDA combined treatments (antibiotics and disinfectants) on biofilm surface area and bacteria viability was evaluated using fluorescence microscopy, digital image analysis and LIVE/DEAD staining. MICs were also determined to assess the probable inhibitory effects of CDA combined treatments on the growth of tested microorganisms' planktonic cells. Treatment of pre-established biofilms with only 310 nM CDA resulted in at least two-fold increase in the number of planktonic cells in all cultures. While antibiotics or disinfectants alone exerted a trivial effect on CFU counts and percentage of surface area covered by the biofilms, combinational treatments with both 310 nM CDA and antibiotics or disinfectants led to approximate 80% reduction in biofilm biomass. These data suggests that combined treatments with CDA would pave the way toward developing new strategies

  15. Investigation of natural biofilms formed during the production of drinking water from surface water embankment filtration.

    Science.gov (United States)

    Emtiazi, Farahnaz; Schwartz, Thomas; Marten, Silke Mareike; Krolla-Sidenstein, Peter; Obst, Ursula

    2004-03-01

    Populations of bacteria in biofilms from different sites of a drinking water production system were analysed. Polymerase chain reaction (PCR) and denaturing gradient gel electrophoresis (DGGE) analyses revealed changing DNA band patterns, suggesting a population shift during bank filtration and processing at the waterworks. In addition, common DNA bands that were attributed to ubiquitous bacteria were found. Biofilms even developed directly after UV disinfection (1-2m distance). Their DNA band patterns only partly agreed with those of the biofilms from the downstream distribution system. Opportunistic pathogenic bacteria in biofilms were analysed using PCR and Southern blot hybridisation (SBH). Surface water appeared to have a direct influence on the composition of biofilms in the drinking water distribution system. In spite of preceding filtration and UV disinfection, opportunistic pathogens such as atypical mycobacteria and Legionella spp. were found in biofilms of drinking water, and Pseudomonas aeruginosa was detected sporadically. Enterococci were not found in any biofilm. Bacterial cell counts in the biofilms from surface water to drinking water dropped significantly, and esterase and alanine-aminopeptidase activity decreased. beta-glucosidase activity was not found in the biofilms. Contrary to the results for planktonic bacteria, inhibitory effects were not observed in biofilms. This suggested an increased tolerance of biofilm bacteria against toxic compounds.

  16. 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

    an efficient dosing regimen and to minimize the development of antimicrobial tolerance and resistance in biofilm infections. Unfortunately, most previous PK/PD studies of antibiotics have been done on planktonic cells, and extrapolation of the results on biofilms is problematic as bacterial biofilms differ...... certificated for clinical use or proved useful for guidance of antibiotic therapy....

  17. 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.

  18. Aeromonas spp.: evaluation of genomic diversity and biofilm forming ability

    OpenAIRE

    Craveiro, Sara Sofia Pereira, 1986-

    2013-01-01

    Tese de mestrado. Biologia (Microbiologia Aplicada). Universidade de Lisboa, Faculdade de Ciências, 2013 Aeromonas spp. are ubiquitous bacteria widely distributed among aquatic environments. Their persistence in water distribution systems is related to their ability to form biofilms, even in the presence of residual chlorine. Once in distribution water systems, aeromonads can contaminate drinking water, food processing facilities and food products. Moreover, members of this genus are known...

  19. 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

    The present study was undertaken to investigate the appearance and location of Pseudomonas aeruginosa in the cystic fibrosis (CF) lung and in sputum. Samples include preserved tissues of CF patients who died due to chronic P. aeruginosa lung infection prior to the advent of intensive antibiotic...

  20. Pseudomonas aeruginosa tolerance to tobramycin, hydrogen peroxide and polymorphonuclear leukocytes is quorum-sensing dependent

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Jensen, Peter Østrup; Burmølle, Mette;

    2005-01-01

    The opportunistic human pathogen Pseudomonas aeruginosa is the predominant micro-organism of chronic lung infections in cystic fibrosis (CF) patients. P. aeruginosa colonizes the CF lungs by forming biofilm structures in the alveoli. In the biofilm mode of growth the bacteria are highly tolerant...

  1. [The biological kinetics of biofilms of clinical strains of Staphylococcus aureus and Pseudomonas aeruginosa separated from patients with bronchopulmonary complications under traumatic disease of spinal cord].

    Science.gov (United States)

    Ul'ianov, V Iu; Opredelentseva, S V; Shvidenko, I G; Norkin, I A; Korshunov, G V; Gladkova, E V

    2014-08-01

    The capacity and intensity of formation of microbial biofilms was analyzed in 24 strains of Staphylococcus aureus and Pseudomonas aeruginosa in static conditions of cultivation during 24, 48, 72 and 96 yours. The microorganisms were separated from patients with bronchopulmonary infectious complications in acute and early periods of traumatic disease of spinal cord.

  2. 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 inv

  3. 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....

  4. 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.

  5. 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

  6. Pseudomonas aeruginosa exhibits deficient biofilm formation in the absence of class II and III ribonucleotide reductases due to hindered anaerobic growth.

    Directory of Open Access Journals (Sweden)

    Anna eCrespo

    2016-05-01

    Full Text Available 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

  7. 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

    , however, be applied to obtain 99.9% disinfection rates. The major reason was that besides cells the mature biofilm contained absorbing and scattering particulates, which made the biofilm opaque. The potential of UVC light emitting diodes ( LED) for disinfection purposes in catheter-like tubes contaminated...

  8. Biofilm-forming activity of bacteria isolated from toilet bowl biofilms and the bactericidal activity of disinfectants against the isolates.

    Science.gov (United States)

    Mori, Miho; Gomi, Mitsuhiro; Matsumune, Norihiko; Niizeki, Kazuma; Sakagami, Yoshikazu

    2013-01-01

    To evaluate the sanitary conditions of toilets, the bacterial counts of the toilet bowl biofilms in 5 Kansai area and 11 Kansai and Kanto area homes in Japan were measured in winter and summer seasons, respectively. Isolates (128 strains) were identified by analyzing 16S ribosomal RNA sequences. The number of colonies and bacterial species from biofilms sampled in winter tended to be higher and lower, respectively, than those in summer. Moreover, the composition of bacterial communities in summer and winter samples differed considerably. In summer samples, biofilms in Kansai and Kanto areas were dominated by Blastomonas sp. and Mycobacterium sp., respectively. Methylobacterium sp. was detected in all toilet bowl biofilms except for one sample. Methylobacterium sp. constituted the major presence in biofilms along with Brevundimonas sp., Sphingomonas sp., and/or Pseudomonas sp. The composition ratio of the sum of their genera was 88.0 from 42.9% of the total bacterial flora. The biofilm formation abilities of 128 isolates were investigated, and results suggested that Methylobacterium sp. and Sphingomonas sp. were involved in biofilm formation in toilet bowls. The biofilm formation of a mixed bacteria system that included bacteria with the highest biofilm-forming ability in a winter sample was greater than mixture without such bacteria. This result suggests that isolates possessing a high biofilm-forming activity are involved in the biofilm formation in the actual toilet bowl. A bactericidal test against 25 strains indicated that the bactericidal activities of didecyldimethylammonium chloride (DDAC) tended to be higher than those of polyhexamethylene biguanide (PHMB) and N-benzyl-N,N-dimethyldodecylammonium chloride (ADBAC). In particular, DDAC showed high bactericidal activity against approximately 90% of tested strains under the 5 h treatment.

  9. 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.

    Science.gov (United States)

    Pamp, Sünje Johanna; Gjermansen, Morten; Johansen, Helle Krogh; Tolker-Nielsen, Tim

    2008-04-01

    Bacteria living as biofilm are frequently reported to exhibit inherent tolerance to antimicrobial compounds, and might therefore contribute to the persistence of infections. Antimicrobial peptides are attracting increasing interest as new potential antimicrobial therapeutics; however, little is known about potential mechanisms, which might contribute to resistance or tolerance development towards these compounds in biofilms. Here we provide evidence that a spatially distinct subpopulation of metabolically active cells in Pseudomonas aeruginosa biofilms is able to develop tolerance to the antimicrobial peptide colistin. On the contrary, biofilm cells exhibiting low metabolic activity were killed by colistin. We demonstrate that the subpopulation of metabolically active cells is able to adapt to colistin by inducing a specific adaptation mechanism mediated by the pmr operon, as well as an unspecific adaptation mechanism mediated by the mexAB-oprM genes. Mutants defective in either pmr-mediated lipopolysaccharide modification or in mexAB-oprM-mediated antimicrobial efflux were not able to develop a tolerant subpopulation in biofilms. In contrast to the observed pattern of colistin-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.

  10. Visualization of Microbiological Processes Underlying Stress Relaxation in Pseudomonas aeruginosa Biofilms

    NARCIS (Netherlands)

    Peterson, Brandon W.; Busscher, Henk J.; Sharma, Prashant K.; van der Mei, Henny C.

    2014-01-01

    Bacterial biofilms relieve themselves from external stresses through internal rearrangement, as mathematically modeled in many studies, but never microscopically visualized for their underlying microbiological processes. The aim of this study was to visualize rearrangement processes occurring in mec

  11. Next Generation Biofilm Inhibitors for Pseudomonas aeruginosa: Synthesis and Rational Design Approaches

    Digital Repository Service at National Institute of Oceanography (India)

    Majik, M.S.; Parvatkar, P.T.

    The bacterial biofilms and the emergence of multiple drug resistance have become a major threat for current medical treatment of nosocomial infections. It has been estimated that about 65-80% of microbial infections in the developed countries...

  12. Noncontact, Low Frequency Ultrasound as an Effective Therapy against Pseudomonas aeruginosa-infected Biofilm Wounds

    Science.gov (United States)

    2013-03-01

    effective. Previous studies have shown physical effects on cells and their surrounding matrix due to ultrasound energy, termed cavitation and...Noncontact, low-frequency ultrasound as an effective therapy against Pseudomonas aeruginosa–infected biofilm wounds Akhil K. Seth, MD1; Khang T...devices may potentially improve healing, but with no evidence of efficacy against biofilms. This study evaluates noncontact, low-frequency ultrasound

  13. 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.

  14. Pseudomonas aeruginosa PA1006, which plays a role in molybdenum homeostasis, is required for nitrate utilization, biofilm formation, and virulence.

    Directory of Open Access Journals (Sweden)

    Melanie J Filiatrault

    Full Text Available Pseudomonas aeruginosa (Pae is a clinically important opportunistic pathogen. Herein, we demonstrate that the PA1006 protein is critical for all nitrate reductase activities, growth as a biofilm in a continuous flow system, as well as virulence in mouse burn and rat lung model systems. Microarray analysis revealed that ΔPA1006 cells displayed extensive alterations in gene expression including nitrate-responsive, quorum sensing (including PQS production, and iron-regulated genes, as well as molybdenum cofactor and Fe-S cluster biosynthesis factors, members of the TCA cycle, and Type VI Secretion System components. Phenotype Microarray™ profiles of ΔPA1006 aerobic cultures using Biolog plates also revealed a reduced ability to utilize a number of TCA cycle intermediates as well as a failure to utilize xanthine as a sole source of nitrogen. As a whole, these data indicate that the loss of PA1006 confers extensive changes in Pae metabolism. Based upon homology of PA1006 to the E. coli YhhP protein and data from the accompanying study, loss of PA1006 persulfuration and/or molybdenum homeostasis are likely the cause of extensive metabolic alterations that impact biofilm development and virulence in the ΔPA1006 mutant.

  15. 铜绿假单胞菌细菌生物膜形成及耐药性分析%Formation of pseudomonas aeruginosa biofilm and antibiotic resistance

    Institute of Scientific and Technical Information of China (English)

    袁晨燕; 韩勍; 陈建明; 芦慧霞

    2011-01-01

    目的 建立铜绿假单胞菌细菌生物膜模型,比较浮游状态和生物膜中铜绿假单胞菌对临床常用抗菌药物的耐药性.方法 用临床分离的铜绿假单胞菌孵育医用硅胶材料(硅胶导尿管)建立细菌生物膜,经银染法和扫描电镜观察生物膜的形成情况,测定 6种抗菌药物对分离菌在浮游状态的最低抑菌浓度(MIC)、最小杀菌浓度(MBC)及生物膜形成以后最小生物膜清除浓度(MBEc).结果 经银染法和扫描电镜观察,导尿管表面形成了细菌生物膜;抗菌药物对生物膜的MBEC为浮游状态下MIC和MBC的100~1000倍.结论 体外建立细菌生物膜,并经银染法观察生物膜的形成是方便可行的;铜绿假单胞菌形成细菌生物膜以后对常用抗菌药物的耐药性远远高于浮游菌.%OBJECTIVE To establish the models of Pseudomonas aeruginosa, to compare the antibiotic susceptibility of P. aeruginosa in biofilms versus its counterparts in planctonic culture. METHODS Incubated isolates from clinic with the silicon catheters and observed the formation of biofilm on catheters by silver staining and scanning electron microscopy (SEM). Assay of determination the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of the planctonic cells and the minimal biofilm eradication concentration (MBEC) of the cells in biofilm were undertaken. RESULTS P. aeruginosa biofilms were observed on the surface of catheters. The MBEC was 100 - 1000 times than MIC and MBC. CONCLUSION The assay of establishing P. aeruginosa biofilm and observed by silver staining is convenient and feasible. P. aeruginosa in biofilm exhibits far more resistance to antimicrobial than its planctonic counterparts does.

  16. Characterization of biofilms formed by Candida parapsilosis, C. metapsilosis, and C. orthopsilosis.

    Science.gov (United States)

    Lattif, Ali Abdul; Mukherjee, Pranab K; Chandra, Jyotsna; Swindell, Kim; Lockhart, Shawn R; Diekema, Daniel J; Pfaller, Michael A; Ghannoum, Mahmoud A

    2010-04-01

    Infections due to Candida parapsilosis have been associated with the ability of this fungus to form biofilms on indwelling medical devices. Recently, C. parapsilosis isolates were reclassified into 3 genetically non-identical classes: C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Little information is available regarding the ability of these newly reclassified species to form biofilms on biomedical substrates. In this study, we characterized biofilm formation by 10 clinical isolates each of C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Biofilms were allowed to form on silicone elastomer discs to early (6h) or mature (48 h) phases and quantified by tetrazolium (XTT) and dry weight assays. Surface topography and three-dimensional architecture of the biofilms were visualized using scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM), respectively. Metabolic activity assay revealed strain-dependent biofilm forming ability of the 3 species tested, while biomass determination revealed that all 3 species formed equivalent biofilms (P>0.05 for all comparisons). SEM analyses of representative isolates of these species showed biofilms with clusters of yeast cells adherent to the catheter surface. Additionally, confocal microscopy analyses showed the presence of cells embedded in biofilms ranging in thickness between 62 and 85 microm. These results demonstrate that similar to C. parapsilosis, the 2 newly identified Candida species (C. orthopsilosis and C. metapsilosis) were able to form biofilms.

  17. Alternative to antibiotics against Pseudomonas aeruginosa: Effects of Glycyrrhiza glabra on membrane permeability and inhibition of efflux activity and biofilm formation in Pseudomonas aeruginosa and its in vitro time-kill activity.

    Science.gov (United States)

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

    2016-09-01

    The multi-drug resistance offered by Pseudomonas aeruginosa to antibiotics can be attributed towards its propensity to develop biofilm, modification in cell membrane and to efflux antibacterial drugs. The present study explored the activity of Glycyrrhiza glabra and one of its pure compounds, glycyrrhizic acid against P. aeruginosa and their mechanism of action in terms of the effect on membrane permeability, efflux activity, and biofilm formation were determined. Minimum inhibitory concentrations were determined by using broth dilution technique. The minimum bactericidal concentrations were assessed on agar plate. The MIC of the extract and glycyrrhizic acid was found to be 200 and 100 μg ml(-1), respectively. The MBC was found to be 800 and 400 μg ml(-1) in the case of extract and glycyrrhizic acid, respectively. Time -dependent killing efficacy was also estimated. Flowcytometric analysis with staining methods was used to determine the effect of extract and glycyrrhizic acid at 2 × MIC on different physiological parameters and compared it with the standard (antibiotic). The growth of P. aeruginosa was significantly inhibited by extract and the pure compound. The herbal extract and the glycyrrhic acid were also found to effective in targeting the physiological parameters of the bacteria that involve cell membrane permeabilization, efflux activity, and biofilm formation. This study reports the antipseudomonal action of Glycyrrhiza glabra and one of its compound and provides insight into their mode of action.

  18. Studies on formation, control and application of biofilm formed by food related microorganisms.

    Science.gov (United States)

    Furukawa, Soichi

    2015-01-01

    Biofilms are sessile microbial aggregates on the interfaces, and they were usually considered as microbial contamination sources in medical care and various industries. We studied the control and application of biofilms formed by food-related microorganisms, and mechanism of the biofilm formation was also investigated. We studied the biofilm formation in mixed cultures using various combinations of two strains of food-related microorganisms. There were various microorganisms that showed decreased or increased biofilm formation in the mixed culture in comparison with that in a single culture. Biofilm formed by lactic acid bacteria and yeast isolated from traditional fermented food, Fukuyama pot vinegar, exhibited unique feature in that structure and formation mechanism, and expected to be used as an immobilized microorganism in fermentation production. Here our studies on the control and application of biofilms and the mechanisms of its formation were described.

  19. Correlation study between c-di-GMP and biofilm formation by Pseudomonas aeruginosa%环二鸟苷酸与铜绿假单胞菌形成生物膜的相关性研究

    Institute of Scientific and Technical Information of China (English)

    王岚; 刘新; 吴怡; 方芳

    2012-01-01

    Objective To investigate the correlation between c-di-GMP and biofilm formation by Pseudomonas aeruginosa through constructing strains with phospho-diesterase gene mutation. Methods The plasmid for genetic mutation was constructed with gene engineering technique and transformed into wild Pseudomonas aeruginosa by homologous recombination. The genetic mutation strains were screened by resistance and sucrose. Results Compared with wild strains, the genetic mutation strains were more prone to form microclone and biofilm on catheter material. Conclusion The increase of c-di-GMP can promotes biofilm formation. C-di-GMP has closely correlation with nosocomial infection and drug resistance caused by Pseudomonas aeruginosa.%目的:通过构建环二鸟苷酸降解酶磷脂酶D基因突变株,探讨环二鸟苷酸与铜绿假单胞菌形成生物膜的相关性.方法:采用基因工程技术构建磷脂酶D基因突变质粒,通过同源重组转化至野生型铜绿假单胞菌PA01,抗性筛选获得基因突变株bf-.扫描电子显微镜下观察基因突变株与野生菌株生物膜表型的差异.结果:与野生菌株相比,基因突变株bf-细菌微克隆的形成增加,有利于细菌在导管材料表面形成生物膜.结论:环二鸟苷酸的增加可促进细菌生物膜的形成,与铜绿假单胞菌引起的院内感染和耐药性密切相关.

  20. Epigallocatechin Gallate Remodels overexpressed Functional Amyloids in Pseudomonas aeruginosa and Increases Biofilm Susceptibility to Antibiotic Treatment

    DEFF Research Database (Denmark)

    Stenvang, Marcel; Dueholm, Morten S; Vad, Brian S;

    2016-01-01

    Epigallocatechin-3-gallate (EGCG) is the major polyphenol in green tea. It has anti-microbial properties and disrupts the ordered structure of amyloid fibrils involved in human disease. The anti-microbial effect of EGCG against the opportunistic pathogen Pseudomonas aeruginosa has been shown...

  1. Cell surface physico chemistry alters biofilm development of Pseudomonas aeruginosa lipopolysaccharide mutants

    NARCIS (Netherlands)

    Flemming, CA; Palmer, RJ; Arrage, AA; Van der Mei, HC; 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 (A(+)B(-

  2. In situ growth rates and biofilm development of Pseudomonas aeruginosa populations in chronic lung infections

    DEFF Research Database (Denmark)

    Yang, L.; Haagensen, J.A.; Jelsbak, L.

    2008-01-01

    The growth dynamics of bacterial pathogens within infected hosts are a fundamental but poorly understood feature of most infections. We have focused on the in situ distribution and growth characteristics of two prevailing and transmissible Pseudomonas aeruginosa clones that have caused chronic lu...

  3. Delayed wound healing in diabetic (db/db) mice with Pseudomonas aeruginosa biofilm challenge: a model for the study of chronic wounds.

    Science.gov (United States)

    Zhao, Ge; Hochwalt, Phillip C; Usui, Marcia L; Underwood, Robert A; Singh, Pradeep K; James, Garth A; Stewart, Philip S; Fleckman, Philip; Olerud, John E

    2010-01-01

    Chronic wounds are a major clinical problem that lead to considerable morbidity and mortality. We hypothesized that an important factor in the failure of chronic wounds to heal was the presence of microbial biofilm resistant to antibiotics and protected from host defenses. A major difficulty in studying chronic wounds is the absence of suitable animal models. The goal of this study was to create a reproducible chronic wound model in diabetic mice by the application of bacterial biofilm. Six-millimeter punch biopsy wounds were created on the dorsal surface of diabetic (db/db) mice, subsequently challenged with Pseudomonas aeruginosa (PAO1) biofilms 2 days postwounding, and covered with semiocclusive dressings for 2 weeks. Most of the control wounds were epithelialized by 28 days postwounding. In contrast, none of biofilm-challenged wounds were closed. Histological analysis showed extensive inflammatory cell infiltration, tissue necrosis, and epidermal hyperplasia adjacent to challenged wounds-all indicators of an inflammatory nonhealing wound. Quantitative cultures and transmission electron microscopy demonstrated that the majority of bacteria were in the scab above the wound bed rather than in the wound tissue. The model was reproducible, allowed localized cutaneous wound infections without high mortality, and demonstrated delayed wound healing following a biofilm challenge. This model may provide an approach to study the role of microbial biofilms in chronic wounds as well as the effect of specific biofilm therapy on wound healing.

  4. Inhibition of Aspergillus fumigatus and Its Biofilm by Pseudomonas aeruginosa Is Dependent on the Source, Phenotype and Growth Conditions of the Bacterium.

    Science.gov (United States)

    Ferreira, Jose A G; Penner, John C; Moss, Richard B; Haagensen, Janus A J; Clemons, Karl V; Spormann, Alfred M; Nazik, Hasan; Cohen, Kevin; Banaei, Niaz; Carolino, Elisabete; Stevens, David A

    2015-01-01

    Aspergillus fumigatus (Af) and Pseudomonas aeruginosa (Pa) are leading fungal and bacterial pathogens, respectively, in many clinical situations. Relevant to this, their interface and co-existence has been studied. In some experiments in vitro, Pa products have been defined that are inhibitory to Af. In some clinical situations, both can be biofilm producers, and biofilm could alter their physiology and affect their interaction. That may be most relevant to airways in cystic fibrosis (CF), where both are often prominent residents. We have studied clinical Pa isolates from several sources for their effects on Af, including testing involving their biofilms. We show that the described inhibition of Af is related to the source and phenotype of the Pa isolate. Pa cells inhibited the growth and formation of Af biofilm from conidia, with CF isolates more inhibitory than non-CF isolates, and non-mucoid CF isolates most inhibitory. Inhibition did not require live Pa contact, as culture filtrates were also inhibitory, and again non-mucoid>mucoid CF>non-CF. Preformed Af biofilm was more resistant to Pa, and inhibition that occurred could be reproduced with filtrates. Inhibition of Af biofilm appears also dependent on bacterial growth conditions; filtrates from Pa grown as biofilm were more inhibitory than from Pa grown planktonically. The differences in Pa shown from these different sources are consistent with the extensive evolutionary Pa changes that have been described in association with chronic residence in CF airways, and may reflect adaptive changes to life in a polymicrobial environment.

  5. Biofilm-Forming Staphylococcus epidermidis Expressing Vancomycin Resistance Early after Adhesion to a Metal Surface

    Directory of Open Access Journals (Sweden)

    Toshiyuki Sakimura

    2015-01-01

    Full Text Available We investigated biofilm formation and time of vancomycin (VCM resistance expression after adhesion to a metal surface in Staphylococcus epidermidis. Biofilm-forming Staphylococcus epidermidis with a VCM MIC of 1 μg/mL was used. The bacteria were made to adhere to a stainless steel washer and treated with VCM at different times and concentrations. VCM was administered 0, 2, 4, and 8 hours after adhesion. The amount of biofilm formed was evaluated based on the biofilm coverage rates (BCRs before and after VCM administration, bacterial viability in biofilm was visually observed using the fluorescence staining method, and the viable bacterial count in biofilm was measured. The VCM concentration required to decrease BCR significantly compared with that of VCM-untreated bacteria was 4 μg/mL, even in the 0 hr group. In the 4 and 8 hr groups, VCM could not inhibit biofilm growth even at 1,024 μg/mL. In the 8 hr group, viable bacteria remained in biofilm at a count of 104 CFU even at a high VCM concentration (1,024 μg/mL. It was suggested that biofilm-forming Staphylococcus epidermidis expresses resistance to VCM early after adhesion to a metal surface. Resistance increased over time after adhesion as the biofilm formed, and strong resistance was expressed 4–8 hours after adhesion.

  6. Effect of pH on biologic degradation of Microcystis aeruginosa by alga-lysing bacteria in sequencing batch biofilm reactors

    Institute of Scientific and Technical Information of China (English)

    Hongjing LI; Mengli HAO; Jingxian LIU; Chen CHEN1; Zhengqiu FAN; Xiangrong WANG

    2012-01-01

    In this paper, the effect of pH on biological degradation of Microcystis aeruginosa by alga-lysing bacteria in laboratory-scale sequencing batch biofilm reactors (SBBRs) was investigated. After 10 d filming with waste activated sludge, the biological film could be formed, and the bioreactors in which laid polyolefin resin filler were used to treat algal culture. By comparing the removal efficiency of chlorophyll a at different aerobic time, the optimum time was determined as 5 h. Under pH 6.5, 7.5, and 8.5 conditions, the removal rates of Microcystis aeruginosa were respectively 75.9%, 83.6%, and 78.3% (in term of chlorophyll a), and that of Chemical Oxygen Demand (CODMn) were 30.6%, 35.8%, and 33.5%. While the removal efficiencies of ammonia nitrogen (NH+ -N) were all 100%. It was observed that the sequence of the removal efficiencies of algae, NH+ -N and organic matter were pH 7.5 〉 pH 8.5 〉 pH 6.5. The results showed that the dominant alga-lysing bacteria in the SBBRs was strain HM-01, which was identified as Bacillus sp. by Polymerase Chain Reaction (PCR) amplification of the 16S rRNA gene, Basic Local Alignment Search Tool (BLAST) analysis, and compar- ison with sequences in the GenBank nucleotide database. The algicidal activated substance which HM-01 strain excreted could withstand high temperature and pressure, also had better hydrophily and stronger polarity.

  7. 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

    Impact of a novel, antimicrobial dressing on in vivo, Pseudomonas aeruginosa wound biofilm: Quantitative comparative analysis using a rabbit ear...for the dressing in the clinical setting. METHODS Animals Under an approved protocol by the Animal Care and Use Committee at the Northwestern... animals were housed in individual cages under constant temperature and humidity with a 12-hour light-dark cycle. A total of 12 animals were used for this

  8. 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...

  9. 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

    We exploited the ability of photocatalytic titanium dioxide (TiO(2) ) as an agent for the biofilm control. Two photocatalytic systems were investigated: a 3g/l suspension of TiO(2) nanopowder in demineralised water and glass slides coated with a TiO(2) thin film, achieved by sol-gel deposition...

  10. 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 co...

  11. In situ growth rates and biofilm development of Pseudomonas aeruginosa populations in chronic lung infections

    DEFF Research Database (Denmark)

    Yang, Lei; Haagensen, Janus Anders Juul; Jelsbak, Lars;

    2008-01-01

    The growth dynamics of bacterial pathogens within infected hosts are a fundamental but poorly understood feature of most infections. We have focused on the in situ distribution and growth characteristics of two prevailing and transmissible Pseudomonas aeruginosa clones that have caused chronic lung...... matrix, whereas nonmucoid variants were present mainly as dispersed cells. To obtain estimates of the growth rates of P. aeruginosa in CF lungs, we used quantitative FISH to indirectly measure growth rates of bacteria in sputum samples (reflecting the in vivo lung conditions). The concentration of r......RNA in bacteria isolated from sputa was measured and correlated with the rRNA contents of the same bacteria growing in vitro at defined rates. The results showed that most cells were actively growing with doubling times of between 100 and 200 min, with some growing even faster. Only a small stationary...

  12. Antimicrobial activities against biofilm formed by Proteus mirabilis isolates from wound and urinary tract infections

    Directory of Open Access Journals (Sweden)

    R Wasfi

    2012-01-01

    Full Text Available Background: Bacterial species are capable of living as biofilm and/or planktonic forms. There is increasing evidence for the role of bacterial biofilm in various wound and urinary tract infections (UTIs. The aim of the present study was to evaluate the ability of the bacteria, isolated from urinary tract infections (UTIs and wound infections, to form biofilm and correlate the role of biofilm with their antimicrobial resistance. Materials and Methods: All the isolated bacteria were screened for their ability to form biofilm using the microtitre plate method. Results: Wound isolates of Staphylococcus aureus and Enterobacter sp. had more biofilm forming capacity than the UTI isolates. Proteus mirabilis isolates were among the strongest biofilm forming bacteria and were chosen for antimicrobial study. In sub-MIC concentrations of antimicrobial agents used, ciprofloxacin was found to be the most effective in decreasing biofilm formation. On the other hand, ceftriaxone and ciprofloxacin were effective in partial removal of preformed biofilm biomass. Conclusion: Ciprofloxacin was more effective in killing bacterial cells especially at high antimicrobial concentrations that could be reached in urine levels and can be used in impregenating catheters.

  13. Rifampicin fails to eradicate mature biofilm formed by methicillin-resistant Staphylococcus aureus

    Directory of Open Access Journals (Sweden)

    Keli Cristine Reiter

    2012-08-01

    Full Text Available INTRODUCTION: Antimicrobial activity on biofilms depends on their molecular size, positive charges, permeability coefficient, and bactericidal activity. Vancomycin is the primary choice for methicillin-resistant Staphylococcus aureus (MRSA infection treatment; rifampicin has interesting antibiofilm properties, but its effectivity remains poorly defined. METHODS: Rifampicin activity alone and in combination with vancomycin against biofilm-forming MRSA was investigated, using a twofold serial broth microtiter method, biofilm challenge, and bacterial count recovery. RESULTS: Minimal inhibitory concentration (MIC and minimal bactericidal concentration for vancomycin and rifampicin ranged from 0.5 to 1mg/l and 0.008 to 4mg/l, and from 1 to 4mg/l and 0.06 to 32mg/l, respectively. Mature biofilms were submitted to rifampicin and vancomycin exposure, and minimum biofilm eradication concentration ranged from 64 to 32,000 folds and from 32 to 512 folds higher than those for planktonic cells, respectively. Vancomycin (15mg/l in combination with rifampicin at 6 dilutions higher each isolate MIC did not reach in vitro biofilm eradication but showed biofilm inhibitory capacity (1.43 and 0.56log10 CFU/ml reduction for weak and strong biofilm producers, respectively; p<0.05. CONCLUSIONS: In our setting, rifampicin alone failed to effectively kill biofilm-forming MRSA, demonstrating stronger inability to eradicate mature biofilm compared with vancomycin.

  14. Characterisation of biofilms formed by Lactobacillus plantarum WCFS1 and food spoilage isolates.

    Science.gov (United States)

    Fernández Ramírez, Mónica D; Smid, Eddy J; Abee, Tjakko; Nierop Groot, Masja N

    2015-08-17

    Lactobacillus plantarum has been associated with food spoilage in a wide range of products and the biofilm growth mode has been implicated as a possible source of contamination. In this study we analysed the biofilm forming capacity of L. plantarum WCFS1 and six food spoilage isolates. Biofilm formation as quantified by crystal violet staining and colony forming units was largely affected by the medium composition, growth temperature and maturation time and by strain specific features. All strains showed highest biofilm formation in Brain Heart Infusion medium supplemented with manganese and glucose. For L. plantarum biofilms the crystal violet (CV) assay, that is routinely used to quantify total biofilm formation, correlates poorly with the number of culturable cells in the biofilm. This can in part be explained by cell death and lysis resulting in CV stainable material, conceivably extracellular DNA (eDNA), contributing to the extracellular matrix. The strain to strain variation may in part be explained by differences in levels of eDNA, likely as result of differences in lysis behaviour. In line with this, biofilms of all strains tested, except for one spoilage isolate, were sensitive to DNase treatment. In addition, biofilms were highly sensitive to treatment with Proteinase K suggesting a role for proteins and/or proteinaceous material in surface colonisation. This study shows the impact of a range of environmental factors and enzyme treatments on biofilm formation capacity for selected L. plantarum isolates associated with food spoilage, and may provide clues for disinfection strategies in food industry.

  15. Expression of Fap amyloids in Pseudomonas aeruginosa, P. fluorescens, and P. putida results in aggregation and increased biofilm formation

    DEFF Research Database (Denmark)

    Dueholm, Morten S; Søndergaard, Mads; Nilsson, Martin

    2013-01-01

    resulted in highly aggregative phenotypes and in increased biofilm formation. Detailed biophysical investigations of purified fibrils confirmed FapC as the main fibril monomer and supported the role of FapB as a minor, nucleating constituent as also indicated by bioinformatic analysis. Bioinformatics......The fap operon, encoding functional amyloids in Pseudomonas (Fap), is present in most pseudomonads, but so far the expression and importance for biofilm formation has only been investigated for P. fluorescens strain UK4. In this study, we demonstrate the capacity of P. aeruginosa PAO1, P....... fluorescens Pf-5, and P. putida F1 to express Fap fibrils, and investigated the effect of Fap expression on aggregation and biofilm formation. The fap operon in all three Pseudomonas species conferred the ability to express Fap fibrils as shown using a recombinant approach. This Fap overexpression consistently...

  16. Mechanistic insights into c-di-GMP-dependent control of the biofilm regulator FleQ from Pseudomonas aeruginosa.

    Science.gov (United States)

    Matsuyama, Bruno Y; Krasteva, Petya V; Baraquet, Claudine; Harwood, Caroline S; Sondermann, Holger; Navarro, Marcos V A S

    2016-01-12

    Bacterial biofilm formation during chronic infections confers increased fitness, antibiotic tolerance, and cytotoxicity. In many pathogens, the transition from a planktonic lifestyle to collaborative, sessile biofilms represents a regulated process orchestrated by the intracellular second-messenger c-di-GMP. A main effector for c-di-GMP signaling in the opportunistic pathogen Pseudomonas aeruginosa is the transcription regulator FleQ. FleQ is a bacterial enhancer-binding protein (bEBP) with a central AAA+ ATPase σ(54)-interaction domain, flanked by a C-terminal helix-turn-helix DNA-binding motif and a divergent N-terminal receiver domain. Together with a second ATPase, FleN, FleQ regulates the expression of flagellar and exopolysaccharide biosynthesis genes in response to cellular c-di-GMP. Here we report structural and functional data that reveal an unexpected mode of c-di-GMP recognition that is associated with major conformational rearrangements in FleQ. Crystal structures of FleQ's AAA+ ATPase domain in its apo-state or bound to ADP or ATP-γ-S show conformations reminiscent of the activated ring-shaped assemblies of other bEBPs. As revealed by the structure of c-di-GMP-complexed FleQ, the second messenger interacts with the AAA+ ATPase domain at a site distinct from the ATP binding pocket. c-di-GMP interaction leads to active site obstruction, hexameric ring destabilization, and discrete quaternary structure transitions. Solution and cell-based studies confirm coupling of the ATPase active site and c-di-GMP binding, as well as the functional significance of crystallographic interprotomer interfaces. Taken together, our data offer unprecedented insight into conserved regulatory mechanisms of gene expression under direct c-di-GMP control via FleQ and FleQ-like bEBPs.

  17. Regulation of biofilm formation in Pseudomonas aeruginosa by quorum sensing%群体感应对铜绿假单胞菌生物被膜形成的调控

    Institute of Scientific and Technical Information of China (English)

    黄媛媛; 宋水山

    2011-01-01

    生物被膜是一种与浮游细胞相对应的生长方式,由细菌和自身分泌的包外基质组成.铜绿假单胞菌是研究这一生长方式的模式生物.在过去十年,对铜绿假单胞菌生物被膜的研究已取得显著进展.群体感应(QS)的细胞沟通机制在铜绿假单胞菌生物被膜形成中发挥着重要作用.介绍生物被膜的特点,并重点讨论了QS和生物被膜之间的关系.%Compared with planktonic cells, bacterial biofilm is a kind of particular colonial life style which consists of bacteria and their extracellular matrix. Pseudomonas aeruginosa has become a model organism for studying biofilm formation. Over the past decade, significant strides have been made towards understanding biofilm development in Pseudomonas aeruginosa. Quorum sensing (QS)has been found to play a role in Pseudomonas aeruginosa biofilm formation. This paper introduced the biofilm characteristics,focusing on the relationship between QS and biofilm formation in Pseudomonas aeruginosa biofilm.

  18. A tribute to disorder in the genome of the bloom-forming freshwater cyanobacterium Microcystis aeruginosa.

    Directory of Open Access Journals (Sweden)

    Jean-François Humbert

    Full Text Available Microcystis aeruginosa is one of the most common bloom-forming cyanobacteria in freshwater ecosystems worldwide. This species produces numerous secondary metabolites, including microcystins, which are harmful to human health. We sequenced the genomes of ten strains of M. aeruginosa in order to explore the genomic basis of their ability to occupy varied environments and proliferate. Our findings show that M. aeruginosa genomes are characterized by having a large open pangenome, and that each genome contains similar proportions of core and flexible genes. By comparing the GC content of each gene to the mean value of the whole genome, we estimated that in each genome, around 11% of the genes seem to result from recent horizontal gene transfer events. Moreover, several large gene clusters resulting from HGT (up to 19 kb have been found, illustrating the ability of this species to integrate such large DNA molecules. It appeared also that all M. aeruginosa displays a large genomic plasticity, which is characterized by a high proportion of repeat sequences and by low synteny values between the strains. Finally, we identified 13 secondary metabolite gene clusters, including three new putative clusters. When comparing the genomes of Microcystis and Prochlorococcus, one of the dominant picocyanobacteria living in marine ecosystems, our findings show that they are characterized by having almost opposite evolutionary strategies, both of which have led to ecological success in their respective environments.

  19. In vitro biofilm forming potential of Streptococcus suis isolated from human and swine in China

    Directory of Open Access Journals (Sweden)

    Guo Dawei

    2012-09-01

    Full Text Available Streptococcus suis is a swine pathogen and also a zoonotic agent. The formation of biofilms allows S. suis to become persistent colonizers and resist clearance by the host immune system and antibiotics. In this study, biofilm forming potentials of various S. suis strains were characterized by confocal laser scanning microscopy (CLSM, scanning electron microscopy (SEM and tissue culture plates stained with crystal violet. In addition, the effects of five antimicrobial agents on biofilm formation were assayed in this study. S. suis produced biofilms on smooth and rough surface. The nutritional contents including glucose and NaCl in the growth medium modulated biofilm formation. There was a significant difference in their biofilm-forming ability among all 46 S. suis strains. The biofilm-forming potential of S. suis serotype 9 was stronger than type 2 and all other types. However, biofilm formation was inhibited by five commonly used antimicrobial agents, penicillin, erythromycin, azithromycin, ciprofloxacin, and ofloxacin at subinhibitory concentrations, among which inhibition of ciprofloxacin and ofloxacin was stronger than that of other three antimicrobial agents.Our study provides a detailed analysis of biofilm formation potential in S. suis, which is a step towards understanding its role in pathogenesis, and eventually lead to a better understanding of how to eradicate S. suis growing as biofilms with antibiotic therapy.

  20. Mixed biofilms formed by C. albicans and non-albicans species: a study of microbial interactions.

    Science.gov (United States)

    Santos, Jéssica Diane dos; Piva, Elisabete; Vilela, Simone Furgeri Godinho; Jorge, Antonio Olavo Cardoso; Junqueira, Juliana Campos

    2016-01-01

    Most Candida infections are related to microbial biofilms often formed by the association of different species. The objective of this study was to evaluate the interactions between Candida albicans and non-albicans species in biofilms formed in vitro. The non-albicans species studied were:Candida tropicalis, Candida glabrata and Candida krusei. Single and mixed biofilms (formed by clinical isolates of C. albicans and non-albicans species) were developed from standardized suspensions of each strain (10(7) cells/mL), on flat-bottom 96-well microtiter plates for 48 hour. These biofilms were analyzed by counting colony-forming units (CFU/mL) in Candida HiChrome agar and by determining cell viability, using the XTT 2,3-bis (2-methoxy-4-nitro-5-sulphophenyl)-5-[(phenylamino) carbonyl]-2H-tetrazolium hydroxide colorimetric assay. The results for both the CFU/mL count and the XTT colorimetric assay showed that all the species studied were capable of forming high levels of in vitro biofilm. The number of CFU/mL and the metabolic activity of C. albicans were reduced in mixed biofilms with non-albicans species, as compared with a single C. albicans biofilm. Among the species tested, C. krusei exerted the highest inhibitory action against C. albicans. In conclusion, C. albicans established antagonistic interactions with non-albicans Candida species in mixed biofilms.

  1. 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-wise pro...

  2. Inhibition of Staphylococcus aureus by antimicrobial biofilms formed by competitive exclusion microorganisms on stainless steel.

    Science.gov (United States)

    Son, Hyeri; Park, Sunhyung; Beuchat, Larry R; Kim, Hoikyung; Ryu, Jee-Hoon

    2016-12-05

    The goal of this study was to develop a desiccation resistant antimicrobial surface using biofilm of competitive exclusion (CE) microorganism inhibitory to Staphylococcus aureus. We isolated 161 microorganisms from soils, foods, and food-contact surfaces that are inhibitory to S. aureus. Among them, three CE microorganisms (Streptomyces spororaveus strain Gaeunsan-18, Bacillus safensis strain Chamnamu-sup 5-25, and Pseudomonas azotoformans strain Lettuce-9) exhibiting strong antibacterial activity and high growth rates were selected for evaluation. These isolates formed biofilms within 24h on stainless steel coupons (SSCs) immersed in Bennet's broth and tryptic soy broth at 25°C. Cells in these biofilms showed significantly (P≤0.05) enhanced resistance to a desiccation (43% relative humidity [RH]) compared to those attached to SSCs but not in biofilms. The antimicrobial activities of biofilms formed by these isolates on SSCs against S. aureus at 25°C and 43% RH were determined. Compared to SSCs lacking biofilms formed by CE microorganisms, populations of S. aureus on SSCs harboring CE biofilms were significantly lower (P≤0.05). Results indicate that persistent antimicrobial activity against S. aureus on stainless steel surfaces can be achieved by the presence of biofilms of CE microorganisms. This information will be useful when developing strategies to improve the microbiological safety of foods during storage, processing, and distribution by facilitating the development of effective antimicrobial food-contact surfaces.

  3. Biofilm-forming bacteria with varying tolerance to peracetic acid from a paper machine.

    Science.gov (United States)

    Rasimus, Stiina; Kolari, Marko; Rita, Hannu; Hoornstra, Douwe; Salkinoja-Salonen, Mirja

    2011-09-01

    Biofilms cause runnability problems in paper machines and are therefore controlled with biocides. Peracetic acid is usually effective in preventing bulky biofilms. This study investigated the microbiological status of a paper machine where low concentrations (≤ 15 ppm active ingredient) of peracetic acid had been used for several years. The paper machine contained a low amount of biofilms. Biofilm-forming bacteria from this environment were isolated and characterized by 16S rRNA gene sequencing, whole-cell fatty acid analysis, biochemical tests, and DNA fingerprinting. Seventy-five percent of the isolates were identified as members of the subclades Sphingomonas trueperi and S. aquatilis, and the others as species of the genera Burkholderia (B. cepacia complex), Methylobacterium, and Rhizobium. Although the isolation media were suitable for the common paper machine biofoulers Deinococcus, Meiothermus, and Pseudoxanthomonas, none of these were found, indicating that peracetic acid had prevented their growth. Spontaneous, irreversible loss of the ability to form biofilm was observed during subculturing of certain isolates of the subclade S. trueperi. The Sphingomonas isolates formed monoculture biofilms that tolerated peracetic acid at concentrations (10 ppm active ingredient) used for antifouling in paper machines. High pH and low conductivity of the process waters favored the peracetic acid tolerance of Sphingomonas sp. biofilms. This appears to be the first report on sphingomonads as biofilm formers in warm water using industries.

  4. Helicobacter pylori-coccoid forms and biofilm formation

    DEFF Research Database (Denmark)

    Andersen, Leif Percival; Rasmussen, Lone

    2009-01-01

    be detected by PCR in water supplies. There is no substantial evidence for viable H. pylori persisting in water supplies. Epidemiological studies suggest that environmental water is a risk factor for H. pylori infection when compared with tap water, and formation of H. pylori biofilm cannot be excluded....... Helicobacter pylori does not seem to take part in biofilm formation in the oral cavity even though the bacterium may be detected....

  5. Biofilm-Forming Abilities of Listeria monocytogenes Serotypes Isolated from Different Sources.

    Directory of Open Access Journals (Sweden)

    Swapnil P Doijad

    Full Text Available A total of 98 previously characterized and serotyped L. monocytogenes strains, comprising 32 of 1/2a; 20 of 1/2b and 46 of 4b serotype, from clinical and food sources were studied for their capability to form a biofilm. The microtiter plate assay revealed 62 (63.26% strains as weak, 27 (27.55% strains as moderate, and 9 (9.18% strains as strong biofilm formers. Among the strong biofilm formers, 6 strains were of serotype 1/2a and 3 strains were of serotype 1/2b. None of the strain from 4b serotype exhibited strong biofilm formation. No firm correlation (p = 0.015 was noticed between any serotype and respective biofilm formation ability. Electron microscopic studies showed that strong biofilm forming isolates could synthesize a biofilm within 24 h on surfaces important in food industries such as stainless steel, ceramic tiles, high-density polyethylene plastics, polyvinyl chloride pipes, and glass. Cell enumeration of strong, moderate, and weak biofilm was performed to determine if the number of cells correlated with the biofilm-forming capabilities of the isolates. Strong, moderate, and weak biofilm showed 570±127× 103 cells/cm2, 33±26× 103 cells/cm2, 5±3× 103 cells/cm2, respectively, indicating that the number of cells was directly proportional to the strength of the biofilm. The hydrophobicity index (HI analysis revealed higher hydrophobicity with an increased biofilm formation. Fatty acid methyl esterase analysis revealed the amount of certain fatty acids such as iso-C15:0, anteiso-C15:0, and anteiso-C17:0 fatty acids correlated with the biofilm-forming capability of L. monocytogenes. This study showed that different strains of L. monocytogenes form biofilm of different intensities which did not completely correlate with their serotype; however, it correlated with the number of cells, hydrophobicity, and amount of certain fatty acids.

  6. Inhibition of Aspergillus fumigatus and Its Biofilm by Pseudomonas aeruginosa Is Dependent on the Source, Phenotype and Growth Conditions of the Bacterium.

    Directory of Open Access Journals (Sweden)

    Jose A G Ferreira

    Full Text Available Aspergillus fumigatus (Af and Pseudomonas aeruginosa (Pa are leading fungal and bacterial pathogens, respectively, in many clinical situations. Relevant to this, their interface and co-existence has been studied. In some experiments in vitro, Pa products have been defined that are inhibitory to Af. In some clinical situations, both can be biofilm producers, and biofilm could alter their physiology and affect their interaction. That may be most relevant to airways in cystic fibrosis (CF, where both are often prominent residents. We have studied clinical Pa isolates from several sources for their effects on Af, including testing involving their biofilms. We show that the described inhibition of Af is related to the source and phenotype of the Pa isolate. Pa cells inhibited the growth and formation of Af biofilm from conidia, with CF isolates more inhibitory than non-CF isolates, and non-mucoid CF isolates most inhibitory. Inhibition did not require live Pa contact, as culture filtrates were also inhibitory, and again non-mucoid>mucoid CF>non-CF. Preformed Af biofilm was more resistant to Pa, and inhibition that occurred could be reproduced with filtrates. Inhibition of Af biofilm appears also dependent on bacterial growth conditions; filtrates from Pa grown as biofilm were more inhibitory than from Pa grown planktonically. The differences in Pa shown from these different sources are consistent with the extensive evolutionary Pa changes that have been described in association with chronic residence in CF airways, and may reflect adaptive changes to life in a polymicrobial environment.

  7. 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...

  8. In vitro biofilm forming capacity on abiotic contact surfaces by outbreak-associated Vibrio harveyi strains

    Institute of Scientific and Technical Information of China (English)

    Pallaval Veera Bramha Chari; Kuchipudi Viswadeepika; Bottu Anand Kumar

    2014-01-01

    Objective:To evaluate the in vitro biofilm forming capacity on abiotic food contact surfaces by Vibrio harveyi (V. harveyi) strains. Methods:Thirty six Gram-negative V. harveyi strains were isolated from various street vended seafood outlets in a food processing line and evaluated for their ability to produce mucoid biofilms on food contact surfaces using a microplate assay. Phenotypic characterization of mucoid biofilm producing V. harveyi strains were screened on Congo red agar, thiosulfate-citrate-bile salts-sucrose agar and tryptic soy agar, respectively. Results: Only five V. harveyi strains (14%) were mucoid biofilm producers characterized by formation of black colonies, whereas the remaining 31 strains (86%) were not capable of producing biofilm characterized by formation of red colonies or pinkish-red colonies with darkening at the centre. The morphological, physiological and biochemical characteristics of these isolates were studied using standard protocols. Strain identification was confirmed by polymerase chain reaction targeted to species-specific polymerase chain reaction primers VH-1 and VH-2 corresponding to variable regions of V. harveyi 16S rRNA sequence. All the biofilm-forming strains showed resistance to at least three antimicrobial compounds tested. V. harveyi strains isolated from various seafood were able to form biofilms of different capacity, and the strains VB267, VB238 and VB166 isolated from cat fish, shrimp and eel fish exhibited significantly greater biofilm forming ability compared to other isolates. Conclusions: It can be concluded from the present study that the strain VB166 was able to better attach and form subsequent biofilms on glass and stainless steel compared to high density polyethylene. These properties allow these bacteria to survive, proliferate and persist in street vended seafood outlets.

  9. In vitro biofilm forming capacity on abiotic contact surfaces by outbreak-associated Vibrio harveyi strains

    Directory of Open Access Journals (Sweden)

    Pallaval Veera Bramha Chari

    2014-02-01

    Full Text Available Objective: To evaluate the in vitro biofilm forming capacity on abiotic food contact surfaces by Vibrio harveyi (V. harveyi strains. Methods: Thirty six Gram-negative V. harveyi strains were isolated from various street vended seafood outlets in a food processing line and evaluated for their ability to produce mucoid biofilms on food contact surfaces using a microplate assay. Phenotypic characterization of mucoid biofilm producing V. harveyi strains were screened on Congo red agar, thiosulfate-citrate-bile salts-sucrose agar and tryptic soy agar, respectively. Results: Only five V. harveyi strains (14% were mucoid biofilm producers characterized by formation of black colonies, whereas the remaining 31 strains (86% were not capable of producing biofilm characterized by formation of red colonies or pinkish-red colonies with darkening at the centre. The morphological, physiological and biochemical characteristics of these isolates were studied using standard protocols. Strain identification was confirmed by polymerase chain reaction targeted to species-specific polymerase chain reaction primers VH-1 and VH-2 corresponding to variable regions of V. harveyi 16S rRNA sequence. All the biofilm-forming strains showed resistance to at least three antimicrobial compounds tested. V. harveyi strains isolated from various seafood were able to form biofilms of different capacity, and the strains VB267, VB238 and VB166 isolated from cat fish, shrimp and eel fish exhibited significantly greater biofilm forming ability compared to other isolates. Conclusions: It can be concluded from the present study that the strain VB166 was able to better attach and form subsequent biofilms on glass and stainless steel compared to high density polyethylene. These properties allow these bacteria to survive, proliferate and persist in street vended seafood outlets.

  10. Genotypic diversity of S. mutans in dental biofilm formed in situ under sugar stress exposure.

    Science.gov (United States)

    Arthur, Rodrigo Alex; Tabchoury, Cínthia Pereira Machado; Mattos-Graner, Renata de Oliveira; Del Bel Cury, Altair A; Paes Leme, Adriana Franco; Vale, Gláuber Campos; Cury, Jaime Aparecido

    2007-01-01

    In situ dental biofilm composition under sugar exposure is well known, but sugar effect on the genotypic diversity of S. mutans in dental biofilm has not been explored. This study evaluated S. mutans genotypic diversity in dental biofilm formed in situ under frequent exposure to sucrose and its monosaccharide constituents (glucose and fructose). Saliva of 7 volunteers was collected for isolation of S. mutans and the same volunteers wore intraoral palatal appliances, containing enamel slabs, which were submitted to the following treatments: distilled and deionized water (negative control), 10% glucose + 10% fructose (fermentable carbohydrates) solution or 20% sucrose (fermentable and EPS inductor) solution, 8x/day. After 3, 7 and 14 days, the biofilms were collected and S. mutans colonies were isolated. Arbitrarily primed polymerase chain reaction (AP-PCR) of S. mutans showed that salivary genotypes were also detected in almost all biofilm samples, independently of the treatment, and seemed to reflect those genotypes present at higher proportion in biofilms. In addition to the salivary genotypes, others were found in biofilms but in lower proportions and were distinct among treatment. The data suggest that the in situ model seems to be useful to evaluate genotypic diversity of S. mutans, but, under the tested conditions, it was not possible to clearly show that specific genotypes were selected in the biofilm due to the stress induced by sucrose metabolism or simple fermentation of its monosaccharides.

  11. The influence of oral Veillonella species on biofilms formed by Streptococcus species.

    Science.gov (United States)

    Mashima, Izumi; Nakazawa, Futoshi

    2014-08-01

    Oral Veillonella, Veillonella atypica, Veillonella denticariosi, Veillonella dispar, Veillonella parvula, Veillonella rogosae, and Veillonella tobetsuensis are known as early colonizers in oral biofilm formation. To investigate the role of oral Veillonella, biofilms formed by the co-culture of Streptococcus gordonii, Streptococcus mutans, Streptococcus salivarius, or Streptococcus sanguinis, with oral Veillonella were examined at the species level. The amount of biofilm formed by S. mutans, S. gordonii, and S. salivarius in the presence of the six Veillonella species was greater than that formed in the control experiments, with the exception of S. mutans with V. dispar. In contrast, in the case of biofilm formation by S. sanguinis, the presence of Veillonella species reduced the amount of the biofilm, with the exception of V. parvula and V. dispar. The time-dependent changes in the amount of biofilm and the number of planktonic cells were grouped into four patterns over the 24 combinations. Only that of S. gordonii with V. tobetsuensis showed a unique pattern. These results indicate that the mode of action of this combination differed from that of the other combinations with respect to biofilm formation. It is possible that there may be several factors involved in the interaction between Streptococcus and Veillonella species.

  12. Efficacy of a marine bacterial nuclease against biofilm forming microorganisms isolated from chronic rhinosinusitis.

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    Robert C Shields

    Full Text Available BACKGROUND: The persistent colonization of paranasal sinus mucosa by microbial biofilms is a major factor in the pathogenesis of chronic rhinosinusitis (CRS. Control of microorganisms within biofilms is hampered by the presence of viscous extracellular polymers of host or microbial origin, including nucleic acids. The aim of this study was to investigate the role of extracellular DNA in biofilm formation by bacteria associated with CRS. METHODS/PRINCIPAL FINDINGS: Obstructive mucin was collected from patients during functional endoscopic sinus surgery. Examination of the mucous by transmission electron microscopy revealed an acellular matrix punctuated occasionally with host cells in varying states of degradation. Bacteria were observed in biofilms on mucosal biopsies, and between two and six different species were isolated from each of 20 different patient samples. In total, 16 different bacterial genera were isolated, of which the most commonly identified organisms were coagulase-negative staphylococci, Staphylococcus aureus and α-haemolytic streptococci. Twenty-four fresh clinical isolates were selected for investigation of biofilm formation in vitro using a microplate model system. Biofilms formed by 14 strains, including all 9 extracellular nuclease-producing bacteria, were significantly disrupted by treatment with a novel bacterial deoxyribonuclease, NucB, isolated from a marine strain of Bacillus licheniformis. Extracellular biofilm matrix was observed in untreated samples but not in those treated with NucB and extracellular DNA was purified from in vitro biofilms. CONCLUSION/SIGNIFICANCE: Our data demonstrate that bacteria associated with CRS form robust biofilms which can be reduced by treatment with matrix-degrading enzymes such as NucB. The dispersal of bacterial biofilms with NucB may offer an additional therapeutic target for CRS sufferers.

  13. DKP对3株病原菌生物膜的抑制作用研究%Study on the Inhibition Effect of DKP on the Biofilms Formed by Three Pathogens

    Institute of Scientific and Technical Information of China (English)

    王建华; 权春善; 赵朋超; 范圣第

    2011-01-01

    The biofilms make it more difficult to treat the diseases caused by pathogens with antibiotics. A kind of diketopiperazine ( DKP )-cyclo ( Pro-Phe ) were found can inhibit the biofilms formation of Staphylococcus aureus,Pseudomonas aeruginosa and Candida albicans. The results of crystal violet staining,CFU (colony forming unit) analysis, and the structure analysis by optical microscope and atomic force microscope indicated that the biofilms of S. aureus and P. aeruginosa were almost disappeared with 10 mg/ml DKP,and the biofilms of C. albicans was significantly inhibited by 12 mg/ml DKP. This brings hope to the research work of novel biofilm inhibitors and cure of the biofilms-associated infections.%生物膜的存在使一些由病原菌引发的疾病变得更加难以治疗.经研究发现一种环二肽物质DKP-cyclo(Pro-Phe)能够抑制这3株病原菌(Staphylococcus aureus,Pseudomonas aeruginosa,Candida albicans)生物膜的形成.通过对不同浓度DKP作用下所形成的生物膜进行结晶紫定量、菌落计数分析和结构显微分析表明:在DKP的浓度达到10 mg/ml时,S.aureus和P.aeruginosa的生物膜几乎消失;在DKP的浓度达到12 mg/ml时,C.albicans的生物膜被显著抑制.这一发现为寻找新型的生物膜抑制剂治愈顽固疾病带来了新的希望.

  14. Forming of a functional biofilm on wood surfaces

    NARCIS (Netherlands)

    Sailer, M.F.; Nieuwenhuijzen, E.J. van; Knol, W.

    2010-01-01

    The protecting and staining properties of biofilms grown on oil-treated surfaces of Pinus sylvestris L. sapwood were investigated with respect to their potential to create homogeneous coloured surfaces. Linseed oil pressure-treated blocks of P. sylvestris L. were evaluated after 36 months of outdoor

  15. Hydrophobicity of biofilm coatings influences the transport dynamics of polystyrene nanoparticles in biofilm-coated sand.

    Science.gov (United States)

    Mitzel, Michael R; Sand, Stefanie; Whalen, Joann K; Tufenkji, Nathalie

    2016-04-01

    Engineered nanoparticles (ENPs) are used in the manufacture of over 2000 industrial and consumer products to enhance their material properties and functions or to enable new nanoparticle-dependent functions. The widespread use of ENPs will result in their release to the subsurface and aquatic environments, where they will interact with indigenous biota. Laboratory column experiments were designed to understand the influence of two different Pseudomonas aeruginosa biofilms on the mobility of polystyrene latex nanoparticles in granular porous media representative of groundwater aquifers or riverbank filtration settings. The transport behavior of 20 nm carboxylate-modified (CLPs) and sulfate (SLPs) polystyrene latex ENPs suspended in NaCl or CaCl2 (1 and 10 mM ionic strength, pH 7) was studied in columns packed with quartz sand coated with biofilms formed by two P. aeruginosa strains that differed in cell surface hydrophobicity (P. aeruginosa 9027™, relatively hydrophilic and P. aeruginosa PAO1, relatively hydrophobic). Biofilm-coated quartz sand retained more of the electrostatically-stabilized latex ENPs than clean, uncoated sand, regardless of the serotype. As IS increased, clear differences in the shape of the ENP breakthrough curves were observed for each type of biofilm coating. ENP breakthrough in the P. aeruginosa PAO1 biofilm-coated sand was generally constant with time whereby breakthrough in the P. aeruginosa 9027 biofilm-coated sand showed dynamic behavior. This indicates a fundamental difference in the mechanisms of ENP deposition onto hydrophilic or hydrophobic biofilm coatings due to the hydration properties of these biofilms. The results of this study demonstrate the importance of considering the surface properties of aquifer grain coatings when evaluating ENP fate in natural subsurface environments.

  16. Pseudomonas aeruginosa Pore-Forming Exolysin and Type IV Pili Cooperate To Induce Host Cell Lysis

    Science.gov (United States)

    Basso, Pauline; Ragno, Michel; Elsen, Sylvie; Reboud, Emeline; Golovkine, Guillaume; Bouillot, Stephanie; Huber, Philippe; Lory, Stephen; Faudry, Eric

    2017-01-01

    ABSTRACT   Clinical strains of Pseudomonas aeruginosa lacking the type III secretion system genes employ a toxin, exolysin (ExlA), for host cell membrane disruption. Here, we demonstrated that ExlA export requires a predicted outer membrane protein, ExlB, showing that ExlA and ExlB define a new active two-partner secretion (TPS) system of P. aeruginosa. In addition to the TPS signals, ExlA harbors several distinct domains, which include one hemagglutinin domain, five arginine-glycine-aspartic acid (RGD) motifs, and a C-terminal region lacking any identifiable sequence motifs. However, this C-terminal region is important for the toxic activity, since its deletion abolishes host cell lysis. Using lipid vesicles and eukaryotic cells, including red blood cells, we demonstrated that ExlA has a pore-forming activity which precedes cell membrane disruption of nucleated cells. Finally, we developed a high-throughput cell-based live-dead assay and used it to screen a transposon mutant library of an ExlA-producing P. aeruginosa clinical strain for bacterial factors required for ExlA-mediated toxicity. The screen resulted in the identification of proteins involved in the formation of type IV pili as being required for ExlA to exert its cytotoxic activity by promoting close contact between bacteria and the host cell. These findings represent the first example of cooperation between a pore-forming toxin of the TPS family and surface appendages in host cell intoxication. PMID:28119472

  17. The Hybrid Histidine Kinase LadS Forms a Multicomponent Signal Transduction System with the GacS/GacA Two-Component System in Pseudomonas aeruginosa.

    Directory of Open Access Journals (Sweden)

    Gaël Chambonnier

    2016-05-01

    Full Text Available In response to environmental changes, Pseudomonas aeruginosa is able to switch from a planktonic (free swimming to a sessile (biofilm lifestyle. The two-component system (TCS GacS/GacA activates the production of two small non-coding RNAs, RsmY and RsmZ, but four histidine kinases (HKs, RetS, GacS, LadS and PA1611, are instrumental in this process. RetS hybrid HK blocks GacS unorthodox HK autophosphorylation through the formation of a heterodimer. PA1611 hybrid HK, which is structurally related to GacS, interacts with RetS in P. aeruginosa in a very similar manner to GacS. LadS hybrid HK phenotypically antagonizes the function of RetS by a mechanism that has never been investigated. The four sensors are found in most Pseudomonas species but their characteristics and mode of signaling may differ from one species to another. Here, we demonstrated in P. aeruginosa that LadS controls both rsmY and rsmZ gene expression and that this regulation occurs through the GacS/GacA TCS. We additionally evidenced that in contrast to RetS, LadS signals through GacS/GacA without forming heterodimers, either with GacS or with RetS. Instead, we demonstrated that LadS is involved in a genuine phosphorelay, which requires both transmitter and receiver LadS domains. LadS signaling ultimately requires the alternative histidine-phosphotransfer domain of GacS, which is here used as an Hpt relay by the hybrid kinase. LadS HK thus forms, with the GacS/GacA TCS, a multicomponent signal transduction system with an original phosphorelay cascade, i.e. H1LadS→D1LadS→H2GacS→D2GacA. This highlights an original strategy in which a unique output, i.e. the modulation of sRNA levels, is controlled by a complex multi-sensing network to fine-tune an adapted biofilm and virulence response.

  18. The Hybrid Histidine Kinase LadS Forms a Multicomponent Signal Transduction System with the GacS/GacA Two-Component System in Pseudomonas aeruginosa

    Science.gov (United States)

    Redelberger, David; Fadel, Firas; Filloux, Alain; Sivaneson, Melissa; de Bentzmann, Sophie; Bordi, Christophe

    2016-01-01

    In response to environmental changes, Pseudomonas aeruginosa is able to switch from a planktonic (free swimming) to a sessile (biofilm) lifestyle. The two-component system (TCS) GacS/GacA activates the production of two small non-coding RNAs, RsmY and RsmZ, but four histidine kinases (HKs), RetS, GacS, LadS and PA1611, are instrumental in this process. RetS hybrid HK blocks GacS unorthodox HK autophosphorylation through the formation of a heterodimer. PA1611 hybrid HK, which is structurally related to GacS, interacts with RetS in P. aeruginosa in a very similar manner to GacS. LadS hybrid HK phenotypically antagonizes the function of RetS by a mechanism that has never been investigated. The four sensors are found in most Pseudomonas species but their characteristics and mode of signaling may differ from one species to another. Here, we demonstrated in P. aeruginosa that LadS controls both rsmY and rsmZ gene expression and that this regulation occurs through the GacS/GacA TCS. We additionally evidenced that in contrast to RetS, LadS signals through GacS/GacA without forming heterodimers, either with GacS or with RetS. Instead, we demonstrated that LadS is involved in a genuine phosphorelay, which requires both transmitter and receiver LadS domains. LadS signaling ultimately requires the alternative histidine-phosphotransfer domain of GacS, which is here used as an Hpt relay by the hybrid kinase. LadS HK thus forms, with the GacS/GacA TCS, a multicomponent signal transduction system with an original phosphorelay cascade, i.e. H1LadS→D1LadS→H2GacS→D2GacA. This highlights an original strategy in which a unique output, i.e. the modulation of sRNA levels, is controlled by a complex multi-sensing network to fine-tune an adapted biofilm and virulence response. PMID:27176226

  19. The role of biofilms in the sedimentology of actively forming gypsum deposits at Guerrero Negro, Mexico.

    Science.gov (United States)

    Vogel, Marilyn B; Des Marais, David J; Turk, Kendra A; Parenteau, Mary N; Jahnke, Linda L; Kubo, Michael D Y

    2009-11-01

    Actively forming gypsum deposits at the Guerrero Negro sabkha and saltern system provided habitats for stratified, pigmented microbial communities that exhibited significant morphological and phylogenetic diversity. These deposits ranged from meter-thick gypsum crusts forming in saltern seawater concentration ponds to columnar microbial mats with internally crystallized gypsum granules developing in natural anchialine pools. Gypsum-depositing environments were categorized as forming precipitation surfaces, biofilm-supported surfaces, and clastic surfaces. Each surface type was described in terms of depositional environment, microbial diversity, mineralogy, and sedimentary fabrics. Precipitation surfaces developed in high-salinity subaqueous environments where rates of precipitation outpaced the accumulation of clastic, organic, and/or biofilm layers. These surfaces hosted endolithic biofilms comprised predominantly of oxygenic and anoxygenic phototrophs, sulfate-reducing bacteria, and bacteria from the phylum Bacteroidetes. Biofilm-supported deposits developed in lower-salinity subaqueous environments where light and low water-column turbulence supported dense benthic microbial communities comprised mainly of oxygenic phototrophs. In these settings, gypsum granules precipitated in the extracellular polymeric substance (EPS) matrix as individual granules exhibiting distinctive morphologies. Clastic surfaces developed in sabkha mudflats that included gypsum, carbonate, and siliclastic particles with thin gypsum/biofilm components. Clastic surfaces were influenced by subsurface brine sheets and capillary evaporation and precipitated subsedimentary gypsum discs in deeper regions. Biofilms appeared to influence both chemical and physical sedimentary processes in the various subaqueous and subaerially exposed environments studied. Biofilm interaction with chemical sedimentary processes included dissolution and granularization of precipitation surfaces, formation of

  20. 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; McDevitt, Christopher A

    2014-11-01

    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 (MoO4 (2-)). 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.

  1. The isolation and characterization of lipopolysaccharides from Microcystis aeruginosa, a prominent toxic water bloom forming cyanobacteria.

    Science.gov (United States)

    Bláhová, Lucie; Adamovský, Ondřej; Kubala, Lukáš; Švihálková Šindlerová, Lenka; Zounková, Radka; Bláha, Luděk

    2013-12-15

    Massive toxic blooms of cyanobacteria represent a major threat to water supplies worldwide, yet serious gaps exist in understanding their complex toxic effects, including the role of lipopolysaccharides (LPS). The present comparative study focused on the levels and biological activities of LPS isolated from Microcystis aeruginosa, which is one of the most globally distributed toxic species. Using hot phenol extraction, LPS was isolated from 3 laboratory cultures and 11 natural water blooms. It formed 0.2-0.7% of the original dry biomass of the cyanobacteria, based on gravimetry. Additional analyses by commercial anti-LPS ELISA were correlated with gravimetry but showed concentrations that were about 7-times lower, which indicated either impurities in isolated LPS or the poor cross-reactivity of the antibodies used. LPS isolates from M. aeruginosa were potent pyrogens in the traditional Limulus amebocyte lysate (LAL)-test, but comparison with the PyroGene test demonstrated the limited selectivity of LAL with several interferences. The determined pyrogenicity (endotoxin units, EU) ranged from very low values in laboratory cultures (less than 0.003 up to 0.008-EU per 100 pg LPS) to higher values in complex bloom samples (0.01-0.078 EU per 100 pg of LPS), which suggested the role of bloom-associated bacteria in the overall effects. Potent pro-inflammatory effects of the studied LPS from both cultures and bloom samples were observed in a highly-relevant ex vivo human blood model by studying reactive oxygen species production in phagocytes as well as increased productions of interleukin 8, IL-8, and tumor necrosis factor α, TNF-α. LPS from M. aeruginosa seem to modulate several pathways involved in the regulation of both innate immunity and specific responses. In comparison to the standard pathogenic bacterial LPS (World Health Organization Escherichia coli O113:10 endotoxin; activity 1 EU per 100 pg), the studied cyanobacterial samples had pyrogenicity potencies

  2. Biofilm forming and leaching mechanism during bioleaching chalcopyrite by Thiobacillus ferrooxidans

    Institute of Scientific and Technical Information of China (English)

    傅建华; 胡岳华; 邱冠周; 柳建设; 徐竞

    2004-01-01

    The mechanism of attachment and leaching of thiobacillus ferrooxcidans (T. f. ) on chalcopyrite were studied. The shaking flasks with bacteria were observed by SEM. The process of T. f attached to the surface of the mineral sample and the biofilm forming were described. The promoting role of the biofilm for bioleaching was discussed. The existence of Fe2+ in the exopolysaccharide layer of T. f was demonstrated by EM(electronic microscope)cell-chemistry analysis. These results show that under the proper growth condition of bacteria, bioleaching of chalcopyrite results in the formation of complete biofilm after 2 - 3 weeks. There are iron ions in the outer layer polymer of T. f. , which provides the micro-environment for themselves, and can guaruntee the energy needed for the bacteria growth in the biofilm. At the same time, Fe3+ ions produced oxidize sulfide which brings about the increase of both growth rate of the bacterial and leaching rate of sulfide minerals.

  3. Functional analysis of stress protein data in a flor yeast subjected to a biofilm forming condition

    Science.gov (United States)

    Moreno-García, Jaime; Mauricio, Juan Carlos; Moreno, Juan; García-Martínez, Teresa

    2016-01-01

    In this data article, an OFFGEL fractionator coupled to LTQ Orbitrap XL MS equipment and a SGD filtering were used to detect in a biofilm-forming flor yeast strain, the maximum possible number of stress proteins under the first stage of a biofilm formation conditions (BFC) and under an initial stage of fermentation used as reference, so-called non-biofilm formation condition (NBFC). Protein functional analysis – based on cellular components and biological process GO terms – was performed for these proteins through the SGD Gene Ontology Slim Mapper tool. A detailed analysis and interpretation of the data can be found in “Stress responsive proteins of a flor yeast strain during the early stages of biofilm formation” [1]. PMID:27104213

  4. Functional analysis of stress protein data in a flor yeast subjected to a biofilm forming condition

    Directory of Open Access Journals (Sweden)

    Jaime Moreno-García

    2016-06-01

    Full Text Available In this data article, an OFFGEL fractionator coupled to LTQ Orbitrap XL MS equipment and a SGD filtering were used to detect in a biofilm-forming flor yeast strain, the maximum possible number of stress proteins under the first stage of a biofilm formation conditions (BFC and under an initial stage of fermentation used as reference, so-called non-biofilm formation condition (NBFC. Protein functional analysis – based on cellular components and biological process GO terms – was performed for these proteins through the SGD Gene Ontology Slim Mapper tool. A detailed analysis and interpretation of the data can be found in “Stress responsive proteins of a flor yeast strain during the early stages of biofilm formation” [1].

  5. Electron transfer mechanism in Shewanella loihica PV-4 biofilms formed at graphite electrode.

    Science.gov (United States)

    Jain, Anand; Zhang, Xiaoming; Pastorella, Gabriele; Connolly, Jack O; Barry, Niamh; Woolley, Robert; Krishnamurthy, Satheesh; Marsili, Enrico

    2012-10-01

    Electron transfer mechanisms in Shewanella loihica PV-4 viable biofilms formed at graphite electrodes were investigated in potentiostat-controlled electrochemical cells poised at oxidative potentials (0.2V vs. Ag/AgCl). Chronoamperometry (CA) showed a repeatable biofilm growth of S. loihica PV-4 on graphite electrode. CA, cyclic voltammetry (CV) and its first derivative shows that both direct electron transfer (DET) mediated electron transfer (MET) mechanism contributes to the overall anodic (oxidation) current. The maximum anodic current density recorded on graphite was 90 μA cm(-2). Fluorescence emission spectra shows increased concentration of quinone derivatives and riboflavin in the cell-free supernatant as the biofilm grows. Differential pulse voltammetry (DPV) show accumulation of riboflavin at the graphite interface, with the increase in incubation period. This is the first study to observe a gradual shift from DET to MET mechanism in viable S. loihica PV-4 biofilms.

  6. Biofilm-forming capacity in biogenic amine-producing bacteria isolated from dairy products.

    Directory of Open Access Journals (Sweden)

    Maria eDiaz

    2016-05-01

    Full Text Available Biofilms on the surface of food industry equipment are reservoirs of potentially food-contaminating bacteria - both spoilage and pathogenic. However, the capacity of biogenic amine (BA-producers to form biofilms has remained largely unexamined. BAs are low molecular weight, biologically active compounds that in food can reach concentrations high enough to be a toxicological hazard. Fermented foods, especially some types of cheese, accumulate the highest BA concentrations of all. The present work examines the biofilm-forming capacity of 56 BA-producing strains belonging to three genera and 10 species (12 Enterococcus faecalis, 6 Enterococcus faecium, 6 Enterococcus durans, 1 Enterococcus hirae, 12 Lactococcus lactis, 7 Lactobacillus vaginalis, 2 Lactobacillus curvatus, 2 Lactobacillus brevis, 1 Lactobacillus reuteri and 7 Lactobacillus parabuchneri, all isolated from dairy products. Strains of all the tested species - except for L. vaginalis - were able to produce biofilms on polystyrene and adhered to stainless steel. However, the biomass produced in biofilms was strain-dependent. These results suggest that biofilms may provide a route via which fermented foods can become contaminated by BA-producing microorganisms.

  7. Potentially pathogenic Escherichia coli can form a biofilm under conditions relevant to the food production chain.

    Science.gov (United States)

    Nesse, Live L; Sekse, Camilla; Berg, Kristin; Johannesen, Karianne C S; Solheim, Heidi; Vestby, Lene K; Urdahl, Anne Margrete

    2014-04-01

    The biofilm-producing abilities of potentially human-pathogenic serotypes of Escherichia coli from the ovine reservoir were studied at different temperatures and on different surfaces. A possible influence of the hydrophobicity of the bacterial cells, as well as the presence of two virulence factors, the Shiga toxin-encoding (Stx) bacteriophage and the eae gene, was also studied. A total of 99 E. coli isolates of serotypes O26:H11, O103:H2, and O103:H25 isolated from sheep feces were included. The results show that isolates of all three E. coli serotypes investigated can produce biofilm on stainless steel, glass, and polystyrene at 12, 20, and 37°C. There was a good general correlation between the results obtained on the different surfaces. E. coli O103:H2 isolates produced much more biofilm than those of the other two serotypes at all three temperatures. In addition, isolates of serotype O26:H11 produced more biofilm than those of O103:H25 at 37°C. The hydrophobicity of the isolates varied between serotypes and was also influenced by temperature. The results strongly indicated that hydrophobicity influenced the attachment of the bacteria rather than their ability to form biofilm once attached. Isolates with the eae gene produced less biofilm at 37°C than isolates without this gene. The presence of a Stx bacteriophage did not influence biofilm production. In conclusion, our results show that potentially human-pathogenic E. coli from the ovine reservoir can form biofilm on various surfaces and at several temperatures relevant for food production and handling.

  8. Potential novel therapeutic strategies in cystic fibrosis: antimicrobial and anti-biofilm activity of natural and designed α-helical peptides against Staphylococcus aureus, Pseudomonas aeruginosa, and Stenotrophomonas maltophilia

    Directory of Open Access Journals (Sweden)

    Pompilio Arianna

    2012-07-01

    Full Text Available Abstract Background Treatment of cystic fibrosis-associated lung infections is hampered by the presence of multi-drug resistant pathogens, many of which are also strong biofilm producers. Antimicrobial peptides, essential components of innate immunity in humans and animals, exhibit relevant in vitro antimicrobial activity although they tend not to select for resistant strains. Results Three α-helical antimicrobial peptides, BMAP-27 and BMAP-28 of bovine origin, and the artificial P19(9/B peptide were tested, comparatively to Tobramycin, for their in vitro antibacterial and anti-biofilm activity against 15 Staphylococcus aureus, 25 Pseudomonas aeruginosa, and 27 Stenotrophomonas maltophilia strains from cystic fibrosis patients. All assays were carried out in physical-chemical experimental conditions simulating a cystic fibrosis lung. All peptides showed a potent and rapid bactericidal activity against most P. aeruginosa, S. maltophilia and S. aureus strains tested, at levels generally higher than those exhibited by Tobramycin and significantly reduced biofilm formation of all the bacterial species tested, although less effectively than Tobramycin did. On the contrary, the viability-reducing activity of antimicrobial peptides against preformed P. aeruginosa biofilms was comparable to and, in some cases, higher than that showed by Tobramycin. Conclusions The activity shown by α-helical peptides against planktonic and biofilm cells makes them promising “lead compounds” for future development of novel drugs for therapeutic treatment of cystic fibrosis lung disease.

  9. The effect of biyuanshu oral liquid on the formation of pseudomonas aeruginosa biofilms in vitro%鼻渊舒口服液对铜绿假单胞菌生物膜体外形成的抑制作用

    Institute of Scientific and Technical Information of China (English)

    刘向; 陈海红; 汪审清

    2012-01-01

    目的:观察鼻渊舒口服液对铜绿假单胞菌生物膜体外形成的抑制作用.方法:平板法建立铜绿假单胞菌细菌生物膜体外模型,银染法及扫描电子显微镜鉴定.不同浓度的鼻渊舒口服液及红霉素作用于成熟前阶段的及已形成的铜绿假单胞菌生物膜,银染法及连续稀释法菌落计数观察其对生物膜的抑制作用.结果:扫描电镜观察铜绿假单胞菌在硅胶片上7d形成生物膜,与银染结果一致.红霉素及鼻渊舒体外能抑制铜绿假单胞菌生物膜的形成,且抑制作用随药物浓度的增加而加强,但对已形成的细菌生物膜清除作用不明显.连续稀释法菌落计数结果表明,不同浓度红霉素及鼻渊舒能抑制成熟前的生物膜膜内细菌生长,与对照组比较差异有统计学意义(P<0.05).结论:鼻渊舒口服液及红霉素体外对铜绿假单胞菌生物膜的形成有抑制作用.%Objective:To observe the effect of biyuanshu oral liquid on the formation of pseudomonas aeruginosa biofilms in vitro. Method: Pseudomonas aeruginosa biofilm was established by plate culture and detected by Scanning electron microscopy and AgNO3 stainning . After treated with different dosages of biyuanshu oral liquid and erythromycin, the pseudomonas aeruginosa biofilms were observed by AgNO3 stainning and the number of viable bacteria were measured by serial dilution. Result; The pseudomonas aeruginosa biofilms could be detected by SEM at the seventh culture day and it was consistent with the detetion of AgNO3 stainning. 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<0. 05). Conclusion

  10. Influence of Amphibian Antimicrobial Peptides and Short Lipopeptides on Bacterial Biofilms Formed on Contact Lenses

    Directory of Open Access Journals (Sweden)

    Magdalena Maciejewska

    2016-10-01

    Full Text Available The widespread use of contact lenses is associated with several complications, including ocular biofilm-related infections. They are very difficult to manage with standard antimicrobial therapies, because bacterial growth in a biofilm is associated with an increased antibiotic resistance. The principal aim of this study was to evaluate the efficacy of antimicrobial peptides (AMPs in eradication of bacterial biofilms formed on commercially available contact lenses. AMPs were synthesized according to Fmoc/tBu chemistry using the solid-phase method. Minimum inhibitory concentration (MIC and minimum biofilm eradication concentration (MBEC of the compounds were determined. Anti-biofilm activity of the antimicrobial peptides determined at different temperatures (25 °C and 37 °C were compared with the effectiveness of commercially available contact lens solutions. All of the tested compounds exhibited stronger anti-biofilm properties as compared to those of the tested lens solutions. The strongest activity of AMPs was noticed against Gram-positive strains at a temperature of 25 °C. Conclusions: The results of our experiments encourage us toward further studies on AMPs and their potential application in the prophylaxis of contact lens-related eye infections.

  11. The efficacy of immediate versus delayed antibiotic administration on bacterial growth and biofilm production of selected strains of uropathogenic Escherichia coli and Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Leah Gandee

    2015-02-01

    Full Text Available Purpose The treatment of urinary tract infections (UTI with antibiotics is commonly used, but recurrence and antibiotic resistance have been growing and concerning clinicians. We studied whether the rapid onset of a protective biofilm may be responsible for the lack of effectiveness of antibiotics against selected bacteria. Materials and Methods Two established uropathogenic Escherichia coli strains, UTI89 and CFT073, and two Pseudomonas aeruginosa strains, PA01 and Boston-41501, were studied to establish a reliable biofilm formation process. Bacterial growth (BG was determined by optical density at 600 nm (OD 600 using a spectrophotometer, while biofilm formation (BF using crystal violet staining was measured at OD 550. Next, these bacterial strains were treated with clinically relevant antibiotics, ciprofloxacin HCl (200 ng/mL and 2 μg/mL, nitrofurantoin (20 μg/mL and 40 μg/mL and ampicillin (50 μg/mL at time points of 0 (T0 or after 6 hours of culture (T6. All measurements, including controls (bacteria -1% DMSO, were done in triplicates and repeated three times for consistency. Results The tested antibiotics effectively inhibited both BG and BF when administered at T0 for UPEC strains, but not when the antibiotic administration started 6 hours later. For Pseudomonas strains, only Ciprofloxacin was able to significantly inhibit bacterial growth at T0 but only at the higher concentration of 2 μg/mL for T6. Conclusion When established UPEC and Pseudomonas bacteria were allowed to culture for 6 hours before initialization of treatment, the therapeutic effect of selected antibiotics was greatly suppressed when compared to immediate treatment, probably as a result of the protective nature of the biofilm.

  12. Repeated applications of photodynamic therapy on Candida glabrata biofilms formed in acrylic resin polymerized.

    Science.gov (United States)

    de Figueiredo Freitas, Lírian Silva; Rossoni, Rodnei Dennis; Jorge, Antonio Olavo Cardoso; Junqueira, Juliana Campos

    2017-04-01

    Previous studies have been suggested that photodynamic therapy (PDT) can be used as an adjuvant treatment for denture stomatitis. In this study, we evaluated the effects of multiple sessions of PDT on Candida glabrata biofilms in specimens of polymerized acrylic resin formed after 5 days. Subsequently, four applications of PDT were performed on biofilms in 24-h intervals (days 6-9). Also, we evaluated two types of PDT, including application of laser and methylene blue or light-emitting diode (LED) and erythrosine. The control groups were treated with physiological solution. The effects of PDT on biofilm were evaluated after the first and fourth application of PDT. The biofilm analysis was performed by counting the colony-forming units. The results showed that between the days 6 and 9, the biofilms not treated by PDT had an increase of 5.53 to 6.05 log (p = 0.0271). Regarding the treatments, after one application of PDT, the biofilms decreased from 5.53 to 0.89 log. When it was done four applications, the microbial reduction ranged from 6.05 log to 0.11 log. We observed that one application of PDT with laser or LED caused a reduction of 3.36 and 4.64 compared to the control groups, respectively (p = 0.1708). When it was done four applications of PDT, the reductions achieved were 1.57 for laser and 5.94 for LED (p = 0.0001). It was concluded that repeated applications of PDT on C. glabrata biofilms showed higher antimicrobial activity compared to single application. PDT mediated by LED and erythrosine was more efficient than the PDT mediated by laser and methylene blue.

  13. Comparative genomics reveals diversified CRISPR-Cas systems of globally distributed Microcystis aeruginosa, a freshwater bloom-forming cyanobacterium

    Directory of Open Access Journals (Sweden)

    Chen eYang

    2015-05-01

    Full Text Available Microcystis aeruginosa is one of the most common and dominant bloom-forming cyanobacteria in freshwater lakes around the world. Microcystis cells can produce toxic secondary metabolites, such as microcystins, which are harmful to human health. Two M. aeruginosa strains were isolated from two highly eutrophic lakes in China and their genomes were sequenced. Comparative genomic analysis was performed with the 12 other available M. aeruginosa genomes and closely related unicellular cyanobacterium. Each genome of M. aeruginosa containing at least one clustered regularly interspaced short palindromic repeat (CRISPR locus and total 71 loci were identified, suggesting it is ubiquitous in M. aeruginosa genomes. In addition to the previously reported subtype I-D cas gene sets, three CAS subtypes I-A, III-A and III-B were identified and characterized in this study. Seven types of CRISPR direct repeat have close association with CAS subtype, confirming that different and specific secondary structures of CRISPR repeats are important for the recognition, binding and process of corresponding cas gene sets. Homology search of the CRISPR spacer sequences provides a history of not only resistance to bacteriophages and plasmids known to be associated with M. aeruginosa, but also the ability to target much more exogenous genetic material in the natural environment. These adaptive and heritable defense mechanisms play a vital role in keeping genomic stability and self-maintenance by restriction of horizontal gene transfer. Maintaining genomic stability and modulating genomic plasticity are both important evolutionary strategies for M. aeruginosa in adaptation and survival in various habitats.

  14. Comparative genomics reveals diversified CRISPR-Cas systems of globally distributed Microcystis aeruginosa, a freshwater bloom-forming cyanobacterium.

    Science.gov (United States)

    Yang, Chen; Lin, Feibi; Li, Qi; Li, Tao; Zhao, Jindong

    2015-01-01

    Microcystis aeruginosa is one of the most common and dominant bloom-forming cyanobacteria in freshwater lakes around the world. Microcystis cells can produce toxic secondary metabolites, such as microcystins, which are harmful to human health. Two M. aeruginosa strains were isolated from two highly eutrophic lakes in China and their genomes were sequenced. Comparative genomic analysis was performed with the 12 other available M. aeruginosa genomes and closely related unicellular cyanobacterium. Each genome of M. aeruginosa containing at least one clustered regularly interspaced short palindromic repeat (CRISPR) locus and total 71 loci were identified, suggesting it is ubiquitous in M. aeruginosa genomes. In addition to the previously reported subtype I-D cas gene sets, three CAS subtypes I-A, III-A and III-B were identified and characterized in this study. Seven types of CRISPR direct repeat have close association with CAS subtype, confirming that different and specific secondary structures of CRISPR repeats are important for the recognition, binding and process of corresponding cas gene sets. Homology search of the CRISPR spacer sequences provides a history of not only resistance to bacteriophages and plasmids known to be associated with M. aeruginosa, but also the ability to target much more exogenous genetic material in the natural environment. These adaptive and heritable defense mechanisms play a vital role in keeping genomic stability and self-maintenance by restriction of horizontal gene transfer. Maintaining genomic stability and modulating genomic plasticity are both important evolutionary strategies for M. aeruginosa in adaptation and survival in various habitats.

  15. The absence of the Pseudomonas aeruginosa OprF protein leads to increased biofilm formation through variation in c-di-GMP level.

    Science.gov (United States)

    Bouffartigues, Emeline; Moscoso, Joana A; Duchesne, Rachel; Rosay, Thibaut; Fito-Boncompte, Laurène; Gicquel, Gwendoline; Maillot, Olivier; Bénard, Magalie; Bazire, Alexis; Brenner-Weiss, Gerald; Lesouhaitier, Olivier; Lerouge, Patrice; Dufour, Alain; Orange, Nicole; Feuilloley, Marc G J; Overhage, Joerg; Filloux, Alain; Chevalier, Sylvie

    2015-01-01

    OprF is the major outer membrane porin in bacteria belonging to the Pseudomonas genus. In previous studies, we have shown that OprF is required for full virulence expression of the opportunistic pathogen Pseudomonas aeruginosa. Here, we describe molecular insights on the nature of this relationship and report that the absence of OprF leads to increased biofilm formation and production of the Pel exopolysaccharide. Accordingly, the level of c-di-GMP, a key second messenger in biofilm control, is elevated in an oprF mutant. By decreasing c-di-GMP levels in this mutant, both biofilm formation and pel gene expression phenotypes were restored to wild-type levels. We further investigated the impact on two small RNAs, which are associated with the biofilm lifestyle, and found that expression of rsmZ but not of rsmY was increased in the oprF mutant and this occurs in a c-di-GMP-dependent manner. Finally, the extracytoplasmic function (ECF) sigma factors AlgU and SigX displayed higher activity levels in the oprF mutant. Two genes of the SigX regulon involved in c-di-GMP metabolism, PA1181 and adcA (PA4843), were up-regulated in the oprF mutant, partly explaining the increased c-di-GMP level. We hypothesized that the absence of OprF leads to a cell envelope stress that activates SigX and results in a c-di-GMP elevated level due to higher expression of adcA and PA1181. The c-di-GMP level can in turn stimulate Pel synthesis via increased rsmZ sRNA levels and pel mRNA, thus affecting Pel-dependent phenotypes such as cell aggregation and biofilm formation. This work highlights the connection between OprF and c-di-GMP regulatory networks, likely via SigX (ECF), on the regulation of biofilm phenotypes.

  16. Silver colloidal nanoparticle stability: influence on Candida biofilms formed on denture acrylic.

    Science.gov (United States)

    Monteiro, Douglas Roberto; Takamiya, Aline Satie; Feresin, Leonardo Perina; Gorup, Luiz Fernando; de Camargo, Emerson Rodrigues; Delbem, Alberto Carlos Botazzo; Henriques, Mariana; Barbosa, Debora Barros

    2014-08-01

    Our aim in this study was to evaluate how the chemical stability of silver nanoparticles (SNs) influences their efficacy against Candida albicans and C. glabrata biofilms. Several parameters of SN stability were tested, namely, temperature (50ºC, 70ºC, and 100ºC), pH (5.0 and 9.0), and time of contact (5 h and 24 h) with biofilms. The control was defined as SNs without temperature treatment, pH 7, and 24 h of contact. These colloidal suspensions at 54 mg/L were used to treat mature Candida biofilms (48 h) formed on acrylic. Their efficacy was determined by total biomass and colony-forming unit quantification. Data were analyzed using analysis of variance and the Bonferroni post hoc test (α = 0.05). The temperature and pH variations of SNs did not affect their efficacy against the viable cells of Candida biofilms (P > 0.05). Moreover, the treatment periods were not decisive in terms of the susceptibility of Candida biofilms to SNs. These findings provide an important advantage of SNs that may be useful in the treatment of Candida-associated denture stomatitis.

  17. Frequency of biofilm formation in toothbrushes and wash basin junks

    Directory of Open Access Journals (Sweden)

    Abdulazeez A Abubakar

    2013-01-01

    Full Text Available Background: Biofilms are known to be resistant to several antibiotics once they are allowed to form on any surface. Aim: To investigate the biofilm forming ability of some bacterial isolates in toothbrushes and wash basin junks. Materials and Methods: A total of 606 students of Federal University of Technology, Yola were provided with new toothbrushes, which were collected after 1 month of usage and screened for biofilm formation. Another 620 swabs were collected from the wash basins of Federal Medical Centre, Specialist Hospital, Federal University of Technology, and students′ hostels in Yola and from some residence in Jimeta, Yola Metropolis; they were all screened for biofilm formation. Results: A total of 38.3% biofilm formation rate was recorded. Three types of bacterial isolates were identified in the biofilms of toothbrushes and wash basin junks, namely Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa at the prevalence rate of 48.0%, 29.1%, and 22.6%, respectively. Overall, 83.3% of the toothbrush biofilm were identified from female students, while 16.7% were from their male counterparts. Statistically, the frequency of biofilm formation showed a significant difference by gender (X 2 = 10.242, P 0.05. Conclusion: This study identified three microorganisms namely S. aureus, E. coli, and P. aeruginosa that were involved in wash basin junk biofilm formation. The findings also showed that occurrence of biofilm in females′ toothbrushes were significantly higher than in males′ (X 2 = 10.242, P < 0.05.

  18. Dynamic remodeling of microbial biofilms by functionally distinct exopolysaccharides.

    Science.gov (United States)

    Chew, Su Chuen; Kundukad, Binu; Seviour, Thomas; van der Maarel, Johan R C; Yang, Liang; Rice, Scott A; Doyle, Patrick; Kjelleberg, Staffan

    2014-08-05

    Biofilms are densely populated communities of microbial cells protected and held together by a matrix of extracellular polymeric substances. The structure and rheological properties of the matrix at the microscale influence the retention and transport of molecules and cells in the biofilm, thereby dictating population and community behavior. Despite its importance, quantitative descriptions of the matrix microstructure and microrheology are limited. Here, particle-tracking microrheology in combination with genetic approaches was used to spatially and temporally study the rheological contributions of the major exopolysaccharides Pel and Psl in Pseudomonas aeruginosa biofilms. Psl increased the elasticity and effective cross-linking within the matrix, which strengthened its scaffold and appeared to facilitate the formation of microcolonies. Conversely, Pel reduced effective cross-linking within the matrix. Without Psl, the matrix becomes more viscous, which facilitates biofilm spreading. The wild-type biofilm decreased in effective cross-linking over time, which would be advantageous for the spreading and colonization of new surfaces. This suggests that there are regulatory mechanisms to control production of the exopolysaccharides that serve to remodel the matrix of developing biofilms. The exopolysaccharides were also found to have profound effects on the spatial organization and integration of P. aeruginosa in a mixed-species biofilm model of P. aeruginosa-Staphylococcus aureus. Pel was required for close association of the two species in mixed-species microcolonies. In contrast, Psl was important for P. aeruginosa to form single-species biofilms on top of S. aureus biofilms. Our results demonstrate that Pel and Psl have distinct physical properties and functional roles during biofilm formation. Importance: Most bacteria grow as biofilms in the environment or in association with eukaryotic hosts. Removal of biofilms that form on surfaces is a challenge in clinical

  19. Antibiofilm activity of α-amylase from Bacillus subtilis S8-18 against biofilm forming human bacterial pathogens.

    Science.gov (United States)

    Kalpana, Balu Jancy; Aarthy, Subramonian; Pandian, Shunmugiah Karutha

    2012-07-01

    The extracellular α-amylase enzyme from Bacillus subtilis S8-18 of marine origin was proved as an antibiofilm agent against methicillin-resistant Staphylococcus aureus (MRSA), a clinical strain isolated from pharyngitis patient, Vibrio cholerae also a clinical isolate from cholera patient and Pseudomonas aeruginosa ATCC10145. The spectrophotometric and microscopic investigations revealed the potential of α-amylase to inhibit biofilm formation in these pathogens. At its BIC level, the crude enzyme caused 51.81-73.07% of biofilm inhibition. Beyond the inhibition, the enzyme was also effective in degradation of preformed mature biofilm by disrupting the exopolysaccharide (EPS), an essential component in biofilm architecture. Furthermore, the enzyme purified to its homogeneity by chromatographic techniques was also effective in biofilm inhibition (43.83-61.68%) as well as in degradation of EPS. A commercial α-amylase enzyme from B. subtilis was also used for comparative purpose. Besides, the effect of various enzymes and temperature on the antibiofilm activity of amylase enzymes was also investigated. This study, for the first time, proved that α-amylase enzyme alone can be used to inhibit/disrupt the biofilms of V. cholerae and MRSA strains and beholds much promise in clinical applications.

  20. Coliform retention and release in biofilms formed on new and weathered irrigation pipes

    Science.gov (United States)

    Irrigation waters have come under increasing scrutiny as a source of pathogenic microorganisms contaminating fresh produce. It is generally assumed that the microbial concentrations entering and leaving irrigation pipe networks are identical. However, this may not be true if biofilms form on the i...

  1. Biofilm-Forming Capability of Highly Virulent, Multidrug-Resistant Candida auris

    Science.gov (United States)

    Sherry, Leighann; Ramage, Gordon; Kean, Ryan; Borman, Andrew; Johnson, Elizabeth M.; Richardson, Malcolm D.

    2017-01-01

    The emerging multidrug-resistant yeast pathogen Candida auris has attracted considerable attention as a source of healthcare–associated infections. We report that this highly virulent yeast has the capacity to form antifungal resistant biofilms sensitive to the disinfectant chlorhexidine in vitro. PMID:28098553

  2. In vitro study of biofilm growth on biologic prosthetics.

    Science.gov (United States)

    Bellows, Charles; Smith, Alison

    2014-01-01

    Biologic prosthetics are increasingly used for the repair of abdominal wall hernia defects but can become infected as a result of peri- or early post-operative bacterial contamination. Data evaluating biofilm formation on biologic prosthetics is lacking. The aim of this study was to investigate the influence of different biologic prosthetics on the growth behavior of two different bacterial species and their ability to form biofilms. Methicillin resistant Staphylococcus aureus (MRSA) or Pseudomrnonas aeruginosa were incubated on disks of two biologic prosthetics-human acellular dermis (ADM), and porcine small intestinal submucosa (SIS). The bacteria were allowed to attach to the prosthetics and propagate into mature biofilms for 24 hours at 370C. Images of biofilms were obtained using confocal microscopy and scanning electron microscopy (SEM). The number of viable cells and the biofilm biomass were quantified by colony forming units (CFUs) and crystal violet staining respectively. Analysis of variance was performed to compare the mean values for the different prosthetics. Each biologic matrix had a distinct surface characteristic. SEM visualized mature biofilms characterized by highly organized multi-cellular structures on surface of both biologic prosthetics. Quantification of bacterial growth over time showed that ADM had the lowest CFUs and biofilm biomass at 24 hours post-inoculation compared to SIS for both bacterial strains. MRSA and P. aeruginosa can form mature biofilms on biologic prosthetics but the relative abundance of the biofilm varies on different prosthetic constructs. Biologic material composition and manufacturing methods may influence bacterial adherence.

  3. 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.

  4. Role of biofilm in protection of the replicative form of Legionella pneumophila.

    Science.gov (United States)

    Andreozzi, Elisa; Di Cesare, Andrea; Sabatini, Luigia; Chessa, Elisa; Sisti, Davide; Rocchi, Marco; Citterio, Barbara

    2014-12-01

    The dual nature of Legionella pneumophila enables its survival in free and intracellular environments and underpins its infection and spread mechanisms. Experiments using bacterial cultures and improved RTqPCR protocols were devised to gain fresh insights into the role of biofilm in protecting the replicative form of L. pneumophila. mip gene expression was used as a marker of virulence in sessile (biofilm-bound) and planktonic (free-floating) cells of L. pneumophila serotype 1 ATCC 33152. The ratio of mip gene expression to transcriptionally active Legionella cells increased both in sessile and free-floating cells demonstrating an up-regulation of mip gene under nutrient depletion. However, a different trend was observed between the two forms, in planktonic cells the mip gene expression/transcriptionally active Legionella cells increased until the end of the experiment, while in the biofilm such increase was observed at the end of the experiment. These findings suggest a possible association between the switch to the transmissive phase of Legionella and a mip up-regulation and a role for biofilm in preserving Legionella cells in replicative form. Moreover, it has been shown that improved RTqPCR protocols are valuable tools to explore bacterial virulence.

  5. Mucoid Pseudomonas aeruginosa isolates maintain the biofilm formation capacity and the gene expression profiles during the chronic lung infection of CF patients

    DEFF Research Database (Denmark)

    Lee, Bao le ri; Schjerling, Charlotte K.; Kirkby, Nikolai;

    2011-01-01

    Danish CF patients were investigated. The in vitro biofilm formation capacity was studied under static and flow through conditions and the global gene expression profiles were investigated by Affymetrix GeneChip. Regulatory genes of alginate production and quorum sensing (QS) system were sequenced...... and measurements of the alginate production and the detection of the QS signal molecules were performed. Comparisons of mucoid and non-mucoid isolates from early and late stages of the infection showed that the mucoid phenotype maintained over a decade the capacity to form in vitro biofilm and showed an unaltered...

  6. Studies of protein adsorption on implant materials in relation to biofilm formation I. Activity of Pseudomonas aeruginosa on Polypropylene and High density Polyethylene in presence of serum albumin

    CERN Document Server

    Sinha, S Dutta; Maity, P K; Tarafdar, S; Moulik, S P

    2014-01-01

    The surface of biomaterials used as implants are highly susceptible to bacterial colonization and subsequent infection. The amount of protein adsorption on biomaterials, among other factors, can affect the nature and quality of biofilms formed on them. The variation in the adsorption time of the protein on the biomaterial surface produces a phenotypic change in the bacteria by alteration of the production of EPS (exoplysaccharide) matrix. Knowledge of the effects of protein adsorption on implant infection will be very useful in understanding the chemistry of the biomaterial surfaces, which can deter the formation of biofilms. It is observed that the adsorption of BSA on the biomaterial surfaces increases with time and concentration, irrespective of their type and the nature of the EPS matrix of the bacterial biofilm is dependent on the amount of protein adsorbed on the biomaterial surface. The adsorption of protein (BSA) on the biomaterials, polypropylene (PP) and high density polyethylene (HDPE) has been stu...

  7. Pseudomonas aeruginosa biofilms exposed to imipenem exhibit changes in global gene expression and beta-lactamase and alginate production

    DEFF Research Database (Denmark)

    Bagge, N.; Schuster, M.; Hentzer, Morten

    2004-01-01

    . As expected, the most strongly induced gene was ampC, which codes for chromosomal beta-lactamase. We also found that genes coding for alginate biosynthesis were induced by exposure to imipenem. Alginate production is correlated to the development of impaired lung function, and P. aeruginosa strains isolated...

  8. 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 Ø;

    2007-01-01

    of growth contributes significantly to P. aeruginosa tolerance to the action of the innate and adaptive defence system and numerous antibiotics. In the present study, an in vivo foreign-body infection model was established in the peritoneal cavity of mice. Experimental data showed that QS-deficient P...

  9. Long-term succession of structure and diversity of a biofilm formed in a model drinking water distribution system

    DEFF Research Database (Denmark)

    Martiny, A.C.; Jørgensen, T.M.; Albrechtsen, Hans-Jørgen

    2003-01-01

    In this study, we examined the long-term development of the overall structural morphology and community composition of a biofilm formed in a model drinking water distribution system with biofilms from 1 day to 3 years old. Visualization and subsequent quantification showed how the biofilm developed...... length polymorphisms showed a correlation between the population profile and the age of the sample, separating the samples into young (1 to 94 days) and old (571 to 1,093 days) biofilms, whereas a limited spatial variation in the biofilm was observed. A more detailed analysis with cloning and sequencing......% of the community by day 256, and which resulted in a reduction in the overall richness. After 500 days, the biofilm entered a stable population state, which was characterized by a greater richness of bacteria, including Nitrospira, Planctomyces, Acidobacterium, and Pseudomonas. The combination of different...

  10. Application of alkaliphilic biofilm-forming bacteria to improve compressive strength of cement-sand mortar.

    Science.gov (United States)

    Park, Sung-Jin; Chun, Woo-Young; Kim, Wha-Jung; Ghim, Sa-Youl

    2012-03-01

    The application of microorganisms in the field of construction material is rapidly increasing worldwide; however, almost all studies that were investigated were bacterial sources with mineral-producing activity and not with organic substances. The difference in the efficiency of using bacteria as an organic agent is that it could improve the durability of cement material. This study aimed to assess the use of biofilm-forming microorganisms as binding agents to increase the compressive strength of cement-sand material. We isolated 13 alkaliphilic biofilmforming bacteria (ABB) from a cement tetrapod block in the West Sea, Korea. Using 16S RNA sequence analysis, the ABB were partially identified as Bacillus algicola KNUC501 and Exiguobacterium marinum KNUC513. KNUC513 was selected for further study following analysis of pH and biofilm formation. Cement-sand mortar cubes containing KNUC513 exhibited greater compressive strength than mineral-forming bacteria (Sporosarcina pasteurii and Arthrobacter crystallopoietes KNUC403). To determine the biofilm effect, Dnase I was used to suppress the biofilm formation of KNUC513. Field emission scanning electron microscopy image revealed the direct involvement of organic-inorganic substance in cement-sand mortar.

  11. Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) isolates of swine origin form robust biofilms.

    Science.gov (United States)

    Nicholson, Tracy L; Shore, Sarah M; Smith, Tara C; Frana, Timothy S; Fraena, Timothy S

    2013-01-01

    Methicillin-resistant Staphylococcus aureus (MRSA) colonization of livestock animals is common and prevalence rates for pigs have been reported to be as high as 49%. Mechanisms contributing to the persistent carriage and high prevalence rates of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) strains in swine herds and production facilities have not been investigated. One explanation for the high prevalence of MRSA in swine herds is the ability of these organisms to exist as biofilms. In this report, the ability of swine LA-MRSA strains, including ST398, ST9, and ST5, to form biofilms was quantified and compared to several swine and human isolates. The contribution of known biofilm matrix components, polysaccharides, proteins and extracellular DNA (eDNA), was tested in all strains as well. All MRSA swine isolates formed robust biofilms similar to human clinical isolates. The addition of Dispersin B had no inhibitory effect on swine MRSA isolates when added at the initiation of biofilm growth or after pre-established mature biofilms formed. In contrast, the addition of proteinase K inhibited biofilm formation in all strains when added at the initiation of biofilm growth and was able to disperse pre-established mature biofilms. Of the LA-MRSA strains tested, we found ST398 strains to be the most sensitive to both inhibition of biofilm formation and dispersal of pre-formed biofilms by DNaseI. Collectively, these findings provide a critical first step in designing strategies to control or eliminate MRSA in swine herds.

  12. Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA isolates of swine origin form robust biofilms.

    Directory of Open Access Journals (Sweden)

    Tracy L Nicholson

    Full Text Available Methicillin-resistant Staphylococcus aureus (MRSA colonization of livestock animals is common and prevalence rates for pigs have been reported to be as high as 49%. Mechanisms contributing to the persistent carriage and high prevalence rates of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA strains in swine herds and production facilities have not been investigated. One explanation for the high prevalence of MRSA in swine herds is the ability of these organisms to exist as biofilms. In this report, the ability of swine LA-MRSA strains, including ST398, ST9, and ST5, to form biofilms was quantified and compared to several swine and human isolates. The contribution of known biofilm matrix components, polysaccharides, proteins and extracellular DNA (eDNA, was tested in all strains as well. All MRSA swine isolates formed robust biofilms similar to human clinical isolates. The addition of Dispersin B had no inhibitory effect on swine MRSA isolates when added at the initiation of biofilm growth or after pre-established mature biofilms formed. In contrast, the addition of proteinase K inhibited biofilm formation in all strains when added at the initiation of biofilm growth and was able to disperse pre-established mature biofilms. Of the LA-MRSA strains tested, we found ST398 strains to be the most sensitive to both inhibition of biofilm formation and dispersal of pre-formed biofilms by DNaseI. Collectively, these findings provide a critical first step in designing strategies to control or eliminate MRSA in swine herds.

  13. Characterization of typical taste and odor compounds formed by Microcystis aeruginosa

    Institute of Scientific and Technical Information of China (English)

    Kejia Zhang; Tsair Fuh Lin; Tuqiao Zhang; Cong Li; Naiyun Gao

    2013-01-01

    Production and characteristics of typical taste and odor (T&O) compounds by Microcystis aeruginosa were investigated.A few terpenoid chemicals,including 2-MIB,β-cyclocitral,and β-ionone,and a few sulfur compounds,such as dimethyl sulfide and dimethyl trisulfide,were detected.β-Cyclocitral and β-carotene concentrations were observed to be relevant to the growth phases of Microcystis.During the stable growth phase,41-865 fg/cell of β-cyclocitral were found in the laboratory culture.β-Cyclocitral concentrations correlated closely with β-carotene concentrations,with the correlation coefficient R2 =0.96,as it is formed from the cleavage reaction of β-carotene.For dead cell cases,a high concentration of dimethyl trisulfide was detected at 3.48-6.37 fg/cell.Four T&O compounds,including β-cyclocitral,β-ionone,heptanal and dimethyl trisulfide,were tested and found to be able to inhibit and damage Microcystis cells to varying degrees.Among these chemicals,β-cyclocitral has the strongest ability to quickly rupture cells.

  14. Azithromycin-Ciprofloxacin-Impregnated Urinary Catheters Avert Bacterial Colonization, Biofilm Formation, and Inflammation in a Murine Model of Foreign-Body-Associated Urinary Tract Infections Caused by Pseudomonas aeruginosa.

    Science.gov (United States)

    Saini, Hina; Vadekeetil, Anitha; Chhibber, Sanjay; Harjai, Kusum

    2017-03-01

    Pseudomonas aeruginosa is a multifaceted pathogen causing a variety of biofilm-mediated infections, including catheter-associated urinary tract infections (CAUTIs). The high prevalence of CAUTIs in hospitals, their clinical manifestations, such as urethritis, cystitis, pyelonephritis, meningitis, urosepsis, and death, and the associated economic challenges underscore the need for management of these infections. Biomaterial modification of urinary catheters with two drugs seems an interesting approach to combat CAUTIs by inhibiting biofilm. Previously, we demonstrated the in vitro efficacy of urinary catheters impregnated with azithromycin (AZM) and ciprofloxacin (CIP) against P. aeruginosa Here, we report how these coated catheters impact the course of CAUTI induced by P. aeruginosa in a murine model. CAUTI was established in female LACA mice with uncoated or AZM-CIP-coated silicone implants in the bladder, followed by transurethral inoculation of 10(8) CFU/ml of biofilm cells of P. aeruginosa PAO1. AZM-CIP-coated implants (i) prevented biofilm formation on the implant's surface (P ≤ 0.01), (ii) restricted bacterial colonization in the bladder and kidney (P < 0.0001), (iii) averted bacteriuria (P < 0.0001), and (iv) exhibited no major histopathological changes for 28 days in comparison to uncoated implants, which showed persistent CAUTI. Antibiotic implants also overcame implant-mediated inflammation, as characterized by trivial levels of inflammatory markers such as malondialdehyde (P < 0.001), myeloperoxidase (P < 0.05), reactive oxygen species (P ≤ 0.001), and reactive nitrogen intermediates (P < 0.01) in comparison to those in uncoated implants. Further, AZM-CIP-coated implants showed immunomodulation by manipulating the release of inflammatory cytokines interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-α), and IL-10 to the benefit of the host. Overall, the study demonstrates long-term in vivo effectiveness of AZM-CIP-impregnated catheters, which may

  15. 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

    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...

  16. Growth and virulence properties of biofilm-forming Salmonella enterica serovar typhimurium under different acidic conditions.

    Science.gov (United States)

    Xu, Hua; Lee, Hyeon-Yong; Ahn, Juhee

    2010-12-01

    This study was designed to characterize the viability and potential virulence of bofilm-forming Salmonella enterica serovar Typhimurium under different pH levels, ranging from 5 to 7. The plate count method and real-time reverse transcription-PCR (RT-PCR) were used to evaluate the survival of S. Typhimurium grown in Trypticase soy broth (TSB) adjusted to pH 5, 6, and 7 (TSB-5, TSB-6, and TSB-7, respectively) at 37°C for 10 days. In TSB-5 and TSB-6, the numbers of viable cells estimated by using the real-time RT-PCR were greater than the culturable counts enumerated by the plate count method. Reflectance micro-Fourier transform infrared (micro-FTIR) spectroscopy was used to evaluate the biochemical changes in biofilm cells. Considerable changes in chemical components were observed in the biofilm cells grown in TSB-5 and TSB-6 when compared to the cells grown in TSB-7. The enterotoxin production and invasive ability of planktonic and biofilm S. Typhimurium cells were inferred by the relative levels of expression of stn and invA. The levels of expression of stn and invA were significantly increased in biofilm S. Typhimurium cells grown in TSB-5 (1.9-fold and 3.2-fold) and TSB-6 (2.1-fold and 22.3-fold) after 10 days of incubation. These results suggest that the biofilm-forming S. Typhimurium under different pH levels might change the virulence production and stress response mechanisms.

  17. [Investigation of the correlation between biofilm forming ability of urinary Candida isolates with the use of urinary catheters and change of antifungal susceptibility in the presence of biofilm].

    Science.gov (United States)

    Aslan, Hacer; Gülmez, Dolunay

    2016-04-01

    Frequency of Candida species causing urinary tract infections is increasing, and this increase is outstanding in nosocomial urinary tract infections especially in intensive care units. The ability of biofilm formation that is contributed to the virulence of the yeast, plays a role in the pathogenesis of biomaterial-related infections and also constitutes a risk for treatment failure. The aims of this study were to compare biofilm forming abilities of Candida strains isolated from urine cultures of patients with and without urinary catheters, and to investigate the change of antifungal susceptibility in the presence of biofilm. A total of 50 Candida strains isolated from urine cultures of 25 patients with urinary catheters (10 C.tropicalis, 6 C.glabrata, 4 C.albicans, 4 C.parapsilosis, 1 C.krusei) and 25 without urinary catheters (8 C.tropicalis, 6 C.albicans, 4 C.krusei, 3 C.parapsilosis, 2 C.kefyr, 1 C.glabrata, 1 C.lusitaniae) were included in the study. Biofilm forming ability was tested by Congo red agar (CRA) and microplate XTT [2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide] reduction methods. Fluconazole (FLU) and amphotericin B (AMP-B) susceptibilities of the isolates were determined by reference microdilution method recommended by Clinical and Laboratory Standards Institute for planktonic cells and by XTT reduction assay in case of biofilm presence. Biofilm formation was detected in 12 (24%) by CRA and 50 (100%) of the isolates by XTT reduction method. None of the C.albicans (n= 10) and C.tropicalis (n= 18) strains were detected as biofilm positive by CRA, however, these strains were strongly positive by XTT reduction method. No statistically significant correlation was detected between the presence of urinary catheter and biofilm forming ability of the isolate (p> 0.05). This might be caused by the advantage of biofilm forming strains in adhesion to bladder mucosa at the initial stages of infection. For all of the isolates in

  18. Pore-forming pyocin S5 utilizes the FptA ferripyochelin receptor to kill Pseudomonas aeruginosa.

    Science.gov (United States)

    Elfarash, Ameer; Dingemans, Jozef; Ye, Lumeng; Hassan, Ahmed Amir; Craggs, Michael; Reimmann, Cornelia; Thomas, Mark S; Cornelis, Pierre

    2014-02-01

    Pyocins are toxic proteins produced by some strains of Pseudomonas aeruginosa that are lethal for related strains of the same species. Some soluble pyocins (S2, S3 and S4) were previously shown to use the pyoverdine siderophore receptors to enter the cell. The P. aeruginosa PAO1 pore-forming pyocin S5 encoding gene (PAO985) was cloned into the expression vector pET15b, and the affinity-purified protein product tested for its killing activity against different P. aeruginosa strains. The results, however, did not show any correlation with a specific ferripyoverdine receptor. To further identify the S5 receptor, transposon mutants were generated. Pooled mutants were exposed to pyocin S5 and the resistant colonies growing in the killing zone were selected. The majority of S5-resistant mutants had an insertion in the fptA gene encoding the receptor for the siderophore pyochelin. Complementation of an fptA transposon mutant with the P. aeruginosa fptA gene in trans restored the sensitivity to S5. In order to define the receptor-binding domain of pyocin S5, two hybrid pyocins were constructed containing different regions from pyocin S5 fused to the C-terminal translocation and DNase killing domains of pyocin S2. Only the protein containing amino acid residues 151 to 300 from S5 showed toxicity, indicating that the pyocin S5 receptor-binding domain is not at the N-terminus of the protein as in other S-type pyocins. Pyocin S5 was, however, unable to kill Burkholderia cenocepacia strains producing a ferripyochelin FptA receptor, nor was the B. cenocepacia fptA gene able to restore the sensitivity of the resistant fptA mutant P. aeruginosa strain.

  19. 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.

  20. Application of bacteriophages to reduce biofilms formed by hydrogen sulfide producing bacteria on surfaces in a rendering plant.

    Science.gov (United States)

    Gong, Chao; Jiang, Xiuping

    2015-08-01

    Hydrogen sulfide producing bacteria (SPB) in raw animal by-products are likely to grow and form biofilms in the rendering processing environments, resulting in the release of harmful hydrogen sulfide (H2S) gas. The objective of this study was to reduce SPB biofilms formed on different surfaces typically found in rendering plants by applying a bacteriophage cocktail. Using a 96-well microplate method, we determined that 3 SPB strains of Citrobacter freundii and Hafnia alvei are strong biofilm formers. Application of 9 bacteriophages (10(7) PFU/mL) from families of Siphoviridae and Myoviridae resulted in a 33%-70% reduction of biofilm formation by each SPB strain. On stainless steel and plastic templates, phage treatment (10(8) PFU/mL) reduced the attached cells of a mixed SPB culture (no biofilm) by 2.3 and 2.7 log CFU/cm(2) within 6 h at 30 °C, respectively, as compared with 2 and 1.5 log CFU/cm(2) reductions of SPB biofilms within 6 h at 30 °C. Phage treatment was also applied to indigenous SPB biofilms formed on the environmental surface, stainless steel, high-density polyethylene plastic, and rubber templates in a rendering plant. With phage treatment (10(9) PFU/mL), SPB biofilms were reduced by 0.7-1.4, 0.3-0.6, and 0.2-0.6 log CFU/cm(2) in spring, summer, and fall trials, respectively. Our study demonstrated that bacteriophages could effectively reduce the selected SPB strains either attached to or in formed biofilms on various surfaces and could to some extent reduce the indigenous SPB biofilms on the surfaces in the rendering environment.

  1. The inhibitory effect of Thymus vulgaris extracts on the planktonic form and biofilm structures of six human pathogenic bacteria

    Directory of Open Access Journals (Sweden)

    Zinab Mohsenipour

    2015-06-01

    Full Text Available Objective: Microorganisms are responsible for many problems in industry and medicine because of biofilm formation. Therefore, this study was aimed to examine the effect of Thymus vulgaris (T. vulgaris extracts on the planktonic form and biofilm structures of six pathogenic bacteria. Materials and methods: Antimicrobial activities of the plant extracts against the planktonic form of the bacteria were determined using the disc diffusion method. MIC and MBC values were evaluated using macrobroth dilution technique. Anti-biofilm effects were assessed by microtiter plate method. Results: According to disc diffusion test (MIC and MBC, the ability of Thymus vulgaris (T. vulgaris extracts for inhibition of bacteria in planktonic form was confirmed. In dealing with biofilm structures, the inhibitory effect of the extracts was directly correlated to their concentration. Except for the inhibition of biofilm formation, efficacy of each extract was independent from type of solvent. Conclusion: According to the potential of Thymus vulgaris (T. vulgaris extracts to inhibit the test bacteria in planktonic and biofilm form, it can be suggested that Thymus vulgaris(T. vulgaris extracts can be applied as antimicrobial agents against the pathogenic bacteria particularly in biofilm forms.

  2. Effects of environmental parameters on the dual-species biofilms formed by Escherichia coli O157:H7 and Ralstonia insidiosa, a strong biofilm producer isolated from a fresh-cut processing plant

    Science.gov (United States)

    Biofilm forming bacteria resident to food processing facilities are a food safety concern due to the potential of biofilms to harbor foodborne bacterial pathogens. When cultured together, Ralstonia insidiosa, a strong biofilm former frequently isolated from produce processing environments, has been ...

  3. Genotypic and phenotypic analysis of S. mutans isolated from dental biofilms formed in vivo under high cariogenic conditions.

    Science.gov (United States)

    Arthur, Rodrigo Alex; Cury, Altair Antoninha Del Bel; Graner, Renata Oliveira Mattos; Rosalen, Pedro Luiz; Vale, Gláuber Campos; Paes Leme, Adriana Franco; Cury, Jaime Aparecido; Tabchoury, Cínthia Pereira Machado

    2011-01-01

    The oral cavity harbors several Streptococcus mutans genotypes, which could present distinct virulence properties. However, little is known about the diversity and virulence traits of S. mutans genotypes isolated in vivo under controlled conditions of high cariogenic challenge. This study evaluated the genotypic diversity of S. mutans isolated from dental biofilms formed in vivo under sucrose exposure, as well as their acidogenicity and aciduricity. To form biofilms, subjects rinsed their mouths with distilled water or sucrose solution 8 times/day for 3 days. S. mutans collected from saliva and biofilms were genotyped by arbitrarily-primed PCR. Genotypes identified in the biofilms were evaluated regarding their ability to lower the suspension pH through glycolysis and their acid susceptibility and F-ATPase activity. Most subjects harbored only one genotype in saliva, which was detected in almost all biofilm samples at high proportions. Genotypes isolated only in the presence of sucrose had higher acidogenicity than those isolated only in the presence of water. Genotypes from biofilms formed with sucrose were more aciduric after 30 and 60 min of incubation at pH 2.8 and 5.0, respectively. The present results suggest that biofilms formed under high cariogenic conditions may harbor more aciduric and acidogenic S. mutans genotypes.

  4. Comparative pyrosequencing analysis of bacterial community change in biofilm formed on seawater reverse osmosis membrane.

    Science.gov (United States)

    Kim, In S; Lee, Jinwook; Kima, Sung-Jo; Yu, Hye-Weon; Jang, Am

    2014-01-01

    The change in bacterial community structure induced by bacterial competition and succession was investigated during seawater reverse osmosis (SWRO) in order to elucidate a possible link between the bacterial consortium on SWRO membranes and biofouling. To date, there has been no definitive characterization of the microbial diversity in SWRO in terms of distinguishing time-dependent changes in the richness or abundance of bacterial species. For bacterial succession within biofilms on the membrane surface, SWRO using a cross-flow filtration membrane test unit was operated for 5 and 100h, respectively. As results of the pyrosequencing analysis, bacterial communities differed considerably among seawater and the 5 and 100 h samples. From a total of 33,876 pyrosequences (using a 95% sequence similarity), there were less than 1% of shared species, confirming the influence of the operational time factor and lack of similarity of these communities. During SWRO operation, the abundance of Pseudomonas stutzeri BBSPN3 (GU594474) belonging to gamma-Proteobacteria suggest that biofouling of SWRO membrane might be driven by the dominant influence of a specific species. In addition, among the bacterial competition of five bacterial species (Pseudomonas aeruginosa, Bacillus sp., Rhodobacter sp., Flavobacterium sp., and Mycobacterium sp.) competing for bacterial colonization on the SWRO membrane surfaces, it was exhibited that Bacillus sp. was the most dominant. The dominant influences ofPseudomonas sp. and Bacillus sp. on biofouling during actual SWRO is decisive depending on higher removal efficiency of the seawater pretreatment.

  5. The inhibitory effect of Thymus vulgaris extracts on the planktonic form and biofilm structures of six human pathogenic bacteria

    OpenAIRE

    Zinab Mohsenipour; Mehdi Hassanshahian

    2015-01-01

    Objective: Microorganisms are responsible for many problems in industry and medicine because of biofilm formation. Therefore, this study was aimed to examine the effect of Thymus vulgaris (T. vulgaris) extracts on the planktonic form and biofilm structures of six pathogenic bacteria. Materials and methods: Antimicrobial activities of the plant extracts against the planktonic form of the bacteria were determined using the disc diffusion method. MIC and MBC values were evaluated using macrobrot...

  6. Growth inhibition of bloom forming cyanobacterium Microcystis aeruginosa by green route fabricated copper oxide nanoparticles.

    Science.gov (United States)

    Sankar, Renu; Prasath, Barathan Balaji; Nandakumar, Ravichandran; Santhanam, Perumal; Shivashangari, Kanchi Subramanian; Ravikumar, Vilwanathan

    2014-12-01

    The cyanobacterium Microcystis aeruginosa can potentially proliferate in a wide range of freshwater bionetworks and create extensive secondary metabolites which are harmful to human and animal health. The M. aeruginosa release toxic microcystins that can create a wide range of health-related issues to aquatic animals and humans. It is essential to eliminate them from the ecosystem with convenient method. It has been reported that engineered metal nanoparticles are potentially toxic to pathogenic organisms. In the present study, we examined the growth inhibition effect of green synthesized copper oxide nanoparticles against M. aeruginosa. The green synthesized copper oxide nanoparticles exhibit an excitation of surface plasmon resonance (SPR) at 270 nm confirmed using UV-visible spectrophotometer. The dynamic light scattering (DLS) analysis revealed that synthesized nanoparticles are colloidal in nature and having a particle size of 551 nm with high stability at -26.6 mV. The scanning electron microscopy (SEM) analysis shows that copper oxide nanoparticles are spherical, rod and irregular in shape, and consistently distributed throughout the solution. The elemental copper and oxide peak were confirmed using energy dispersive x-ray analysis (EDAX). Fourier-transform infrared (FT-IR) spectroscopy indicates the presence of functional groups which is mandatory for the reduction of copper ions. Besides, green synthesized copper oxide nanoparticles shows growth inhibition against M. aeruginosa. The inhibition efficiency was 31.8 % at lower concentration and 89.7 % at higher concentration of copper oxide nanoparticles, respectively. The chlorophyll (a and b) and carotenoid content of M. aeruginosa declined in dose-dependent manner with respect to induction of copper oxide nanoparticles. Furthermore, we analyzed the mechanism behind the cytotoxicity of M. aeruginosa induced by copper oxide nanoparticles through evaluating membrane integrity, reactive oxygen species (ROS

  7. The Activity of Cotinus coggygria Scop. Leaves on Staphylococcus aureus Strains in Planktonic and Biofilm Growth Forms

    Directory of Open Access Journals (Sweden)

    Katarína Rendeková

    2015-12-01

    Full Text Available The purpose of this study was to detect the effectiveness of Cotinus coggygria Scop. leaves methanol extract against planktonic and biofilm growth forms of Staphylococcus aureus. The antimicrobial activity was determined by the broth microdilution test. Minimal inhibitory concentrations and minimal bactericidal concentrations were detected against two collection and ten clinical S. aureus strains. Anti-biofilm activity of the tested extract was detected using 24 h bacterial biofilm on the surface of microtiter plate wells. The biofilm inhibitory activity was evaluated visually after 24 h interaction of extract with biofilm, and the eradicating activity by a regrowth method. The tested extract showed bactericidal activity against all S. aureus strains (methicillin susceptible or methicillin resistant in concentrations ranging from 0.313 to 0.625 mg·mL−1. Biofilm inhibitory concentrations were 10-times higher and biofilm eradicating concentrations 100-times higher (8 and 32 mg·mL−1, respectively. The phytochemical analysis of C. coggygria leaves 60% methanol extract performed by LC-DAD-MS/MS revealed quercetin rhamnoside, methyl gallate, and methyl trigallate as main constituents. Results of our study indicate that C. coggygria, rich in tannins and flavonoids, seems to be a prospective topical antibacterial agent with anti-biofilm activity.

  8. Biofilm-Forming Methicillin-Resistant Staphylococcus aureus Survive in Kupffer Cells and Exhibit High Virulence in Mice

    Directory of Open Access Journals (Sweden)

    Takuto Oyama

    2016-06-01

    Full Text Available Although Staphylococcus aureus is part of the normal body flora, heavy usage of antibiotics has resulted in the emergence of methicillin-resistant strains (MRSA. MRSA can form biofilms and cause indwelling foreign body infections, bacteremia, soft tissue infections, endocarditis, and osteomyelitis. Using an in vitro assay, we screened 173 clinical blood isolates of MRSA and selected 20 high-biofilm formers (H-BF and low-biofilm formers (L-BF. These were intravenously administered to mice and the general condition of mice, the distribution of bacteria, and biofilm in the liver, lung, spleen, and kidney were investigated. MRSA count was the highest in the liver, especially within Kupffer cells, which were positive for acid polysaccharides that are associated with intracellular biofilm. After 24 h, the general condition of the mice worsened significantly in the H-BF group. In the liver, bacterial deposition and aggregation and the biofilm-forming spot number were all significantly greater for H-BF group than for L-BF. CFU analysis revealed that bacteria in the H-BF group survived for long periods in the liver. These results indicate that the biofilm-forming ability of MRSA is a crucial factor for intracellular persistence, which could lead to chronic infections.

  9. Experimental study on effect of mesna on Pseudomonas aeruginosa biofilm%巯乙磺酸钠对铜绿假单胞菌生物被膜作用的实验研究

    Institute of Scientific and Technical Information of China (English)

    陈盛; 余加林; 罗则佳; 何念海; 孙凤军

    2013-01-01

    OBJECTIVE To investigate the effect of mesna on the formation of Pseudomonas aeruginosa biofilm, and study the effect of mesna on P. aeruginosa biofilm. METHODS The broth microdilution method was performed to determine the minimal inhibitory concentration of mesna to PAO1, then a biofilm model of Pseudo-monas aeruginosa in vitro was established , the appearance of biofilm was detected by scanning electron microscope (SEM ) to assess the effect of mesna on the formation of P. aeruginosa biofilm; the bacteria colony counts in biofilm was measured by agar plate after the biofilm was treated by mesna, biofilm structure was observed under confocal laser scanning microscope (CLSM), and the parameters of biofilm structure were analyzed through pictures from CLSM with image structure analyzer (ISA) software. RESULTS The MIC value against PAO1 was 10mg/mL for mesna. In the process of Pseudomonas aeruginosa biofilm formation, scanning electron microscope showed that the mucoid materials among bacteria was significantly reduced and the thickness of biofilm was decreased in mesna group. In comparison with normal saline group, viable counts in biofilms in the mesna treatment group were less than those in the saline group, and the high-dose group (4. 06 ± 0. 12) had less positive effect than did the low-dose group(5. 84 ± 0. 24)(P<0. 05). Confocal laser scanning microscope showed that the biofilm was thinner and more scattered than the saline control group. The results of ISA showed that with the treatment of mesna, biofilm was decreased in thickness, average diffusion distance (ADD) and textual entropy (TE) in comparison with the saline control group(P<0. 05),however areal porosity(AP) was increased (P< 0. 05) , and the high-doses group was more significant than the low-doses group (P<0. 05). CONCLUSION Mesna can inhibit the formation of P. aeruginosa biofilm and disrupt the structure of P. aeruginosa biofilm.%目的 研究巯乙磺酸钠(Mesna)对铜绿假单胞菌生物被

  10. Biofilm formation of Achromobacter xylosoxidans on contact lens.

    Science.gov (United States)

    Konstantinović, Neda; Ćirković, Ivana; Đukić, Slobodanka; Marić, Vesna; Božić, Dragana D

    2017-02-20

    Achromobacter spp. may contaminate lenses, lens cases, and contact lens solutions and cause ocular infections. The aim of this study was to investigate the possibility of isolated strain of Achromobacter xylosoxidans to form biofilm on the surface of soft contact lenses (CL), to quantify the production of the formed biofilm, and compare it with the reference strains (Pseudomonas aeruginosa, Staphylococcus aureus, and Haemophilus influenzae). Bacterial strain isolated from one contact lens case was identified as A. xylosoxidans using Vitek2 Automated System. Biofilm forming capacity of isolated strain of A. xylosoxidans and reference strains of P. aeruginosa, S. aureus, and H. influenzae on soft CL were analyzed by commonly used microtitre plate method. Our results showed that isolated strain of A. xylosoxidans was capable to form biofilm on the surface of soft contact lens. A. xylosoxidans was strong biofilm producer while all examined reference strains were moderate biofilm producers. A. xylosoxidans appears to be superior biofilm producer on soft CL compared to reference strains.

  11. 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.

  12. Clinical isolates of Acinetobacter baumannii from a Portuguese hospital: PFGE characterization, antibiotic susceptibility and biofilm-forming ability.

    Science.gov (United States)

    Duarte, Andreia; Ferreira, Susana; Almeida, Sofia; Domingues, Fernanda C

    2016-04-01

    Acinetobacter baumannii is an emerging pathogen associated with nosocomial infections that in addition has shown an increasing resistance to antibiotics. In this work the genetic diversity of A. baumannii isolates from a Portuguese hospital, their antibiotic resistance profiles and ability to form biofilms was studied. Seventy-nine clinical A. baumannii isolates were characterized by pulsed-field gel electrophoresis (PFGE) with 9 different PFGE profiles being obtained. Concerning the antimicrobial susceptibility, all A. baumannii isolates were resistant to 12 of the 17 tested antibiotics and classified as multidrug-resistant (MDR). In addition, 74.7% of the isolates showed biofilm formation ability, however no statistical significance with antibiotic resistance was observed. In contrast, urine samples isolates were more likely to form biofilms than strains isolated from other sources. Our findings highlight the high number of MDR A. baumannii isolates and the importance of the formation of biofilms as a potential virulence factor.

  13. Igneous phosphate rock solubilization by biofilm-forming mycorrhizobacteria and hyphobacteria associated with Rhizoglomus irregulare DAOM 197198.

    Science.gov (United States)

    Taktek, Salma; St-Arnaud, Marc; Piché, Yves; Fortin, J André; Antoun, Hani

    2017-01-01

    Biofilm formation on abiotic and biotic surfaces was studied with two hyphobacteria, strongly attached to the surface of the arbuscular mycorrhizal fungus (AMF) Rhizoglomus irregulare (Ri) DAOM 197198 and two mycorrhizobacteria, loosely attached to the roots of different mycorrhizal plants. When the sparingly soluble igneous phosphate rock (PR) from Quebec, or when the chemical hydroxyapatite were used as sole phosphorus (P) source, hyphobacteria Rhizobium miluonense Rm3 and Burkholderia anthina Ba8 produced significantly more biofilms than mycorrhizobacteria Rahnella sp. Rs11 and Burkholderia phenazinium Bph12, as indicated by the crystal violet assay or by quantifying biofilm exopolysaccharides. As previously observed with planktonic bacteria, biofilms mobilized P by lowering the pH and releasing gluconic acid. The high efficiency of P mobilization by the hyphobacteria Ba8 was linked to the presence of more viable cells in its biofilm as revealed by the hydrolysis of fluorescein diacetate. Scanning electron microscopy micrographs showed a high adherence of the best P-solubilizer hyphobacteria Ba8 on the surface of Quebec PR. Hydroxyapatite porous structure did not allow a good adherence of Ba8. Ba8 formed an important biofilm on the hyphae of Ri DAOM 197198 with low reactive Quebec PR while no biofilm was observed with the high reactive hydroxyapatite. Results confirm the possible presence of specificity between the Ri DAOM 197198 and the hyphobacteria and suggest that the interaction would be regulated by the availability of P.

  14. Synergistic effect of amphotericin B and tyrosol on biofilm formed by Candida krusei and Candida tropicalis from intrauterine device users.

    Science.gov (United States)

    Shanmughapriya, Santhanam; Sornakumari, Haridevvenkatesan; Lency, Arumugam; Kavitha, Senthil; Natarajaseenivasan, Kalimuthusamy

    2014-11-01

    The presence of intrauterine contraceptive devices (IUDs) provides a solid surface for attachment of microorganisms and an ideal niche for the biofilm to form and flourish. Vaginal candidiasis is often associated with the use of IUDs. Treatment of vaginal candidiasis that develops in connection with IUD use requires their immediate removal. Here, we present in vitro evidence to support the use of combination therapy to inhibit Candida biofilm. Twenty-three clinical Candida isolates (10 C. krusei and 13 C. tropicalis) recovered from endocervical swabs obtained from IUD and non-IUD users were assessed for biofilm-formation ability. The rate of isolation of Candida did not differ significantly among IUD and non-IUD users (P = 0.183), but the biofilm-formation ability of isolates differed significantly (P = 0.02). An in vitro biofilm model with the obtained isolates was subjected to treatment with amphotericin B, tyrosol, and a combination of amphotericin B and tyrosol. Inhibition of biofilm by amphotericin B or tyrosol was found to be concentration dependent, with 50% reduction (P tyrosol and amphotericin B was studied. Interestingly, approximately 90% reduction in biofilm was observed with use of 80 μM tyrosol combined with 4 mg/l amphotericin B (P < 0.001). This represents a first step in establishing an appropriate antibiofilm therapy when yeasts are present.

  15. Enzymatic removal and disinfection of bacterial biofilms

    DEFF Research Database (Denmark)

    Johansen, Charlotte; Falholt, Per; Gram, Lone

    1997-01-01

    Model biofilms of Staphylococcus aureus, Staphylococcus epidermidis, Pseudomonas fluorescens, and Pseudomonas aeruginosa were made on steel and polypropylene substrata. Plaque-resembling biofilms of Streptococcus mutans, Actinomyces, viscosus, and Fusobacterium nucleatum were made on saliva...

  16. Zingerone silences quorum sensing and attenuates virulence of Pseudomonas aeruginosa.

    Science.gov (United States)

    Kumar, Lokender; Chhibber, Sanjay; Kumar, Rajnish; Kumar, Manoj; Harjai, Kusum

    2015-04-01

    Quorum sensing in Pseudomonas aeruginosa plays an imperative role in virulence factor, biofilm formation and antimicrobial resistance. Blocking quorum sensing pathways are viewed as viable anti-virulent therapy in association with traditional antimicrobial therapy. Anti-quorum sensing dietary phytochemicals with may prove to be a safe and viable choice as anti-virulent drug candidates. Previously, our lab proved zingerone as potent anti-biofilm agent hence; further its anti-virulent and anti-quorum activities were evaluated. Zingerone, besides decreasing swimming, swarming and twitching phenotypes of P. aeruginosa PAO1, reduced biofilm forming capacity and production of virulence factors including rhamnolipid, elastase, protease, pyocyanin, cell free and cell bound hemolysin (pquorum sensing signal molecules by clinical isolates of P. aeruginosa but also showed significant interference with the activation of QS reporter strains. To study the mechanism of blocking quorum sensing cascade, in silico analysis was carried out. Anti-QS activity was attributed to interference with the ligand receptor interaction of zingerone with QS receptors (TraR, LasR, RhlR and PqsR). Zingerone showed a good comparative docking score to respective autoinducer molecules which was even higher than that of vanillin, a proven anti-quorum sensing phytochemical. The results of the present study revealed the anti-quorum sensing activity of zingerone targeting ligand-receptor interaction, hence proposing zingerone as a suitable anti-virulent drug candidate against P. aeruginosa infections.

  17. Diarrhea-associated biofilm formed by enteroaggregative Escherichia coli and aggregative Citrobacter freundii: a consortium mediated by putative F pili

    Directory of Open Access Journals (Sweden)

    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.

  18. Caracterización del mutante nfxB de Pseudomonas aeruginosa: papel en la resistencia antibiótica de los biofilms e interacción con los mecanismos de resistencia intrínsecos.

    OpenAIRE

    Mulet Aguiló, Francisco Javier

    2015-01-01

    La azitromicina ha mostrado resultados prometedores en el tratamiento de la infección respiratoria crónica por P. aeruginosa. No presenta actividad bactericida frente a este microorganismo en cultivo planctónico, pero se sabe que este macrólido inhibe la formación de biofilms gracias a su capacidad para interferir en el sistema quorum sensing bacteriano. En las infecciones crónicas frecuentemente se seleccionan mutantes que presentan un alto grado de adaptación al nicho ecológico y a los anti...

  19. Cultivation and characterization of the MaMV-DC cyanophage that infects bloom-forming cyanobacterium Microcystis aeruginosa.

    Science.gov (United States)

    Ou, Tong; Li, Sanhua; Liao, Xiangyong; Zhang, Qiya

    2013-10-01

    The MaMV-DC cyanophage, which infects the bloom-forming cyanobacterium Microcystis aeruginosa, was isolated from Lake Dianchi, Kunming, China. Twenty-one cyanobacterial strains were used to detect the host range of MaMV-DC. Microcystic aeruginosa FACHB-524 and plaque purification were used to isolate individual cyanophages, and culturing MaMV-DC with cyanobacteria allowed us to prepare purified cyanophages for further analysis. Electron microscopy demonstrated that the negatively stained viral particles are tadpole-shaped with an icosahedral head approximately 70 nm in diameter and a contractile tail approximately 160 nm in length. Using one-step growth experiments, the latent period and burst size of MaMV-DC were estimated to be 24-48 hours and approximately 80 infectious units per cell, respectively. Restriction endonuclease digestion and agarose gel electrophoresis were performed using purified MaMV-DC genomic DNA, and the genome size was estimated to be approximately 160 kb. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis revealed four major structural proteins. These results support the growing interest in using freshwater cyanophages to control bloom-forming cyanobacterium.

  20. Paradoxical antifungal activity and structural observations in biofilms formed by echinocandin-resistant Candida albicans clinical isolates.

    Science.gov (United States)

    Walraven, Carla J; Bernardo, Stella M; Wiederhold, Nathan P; Lee, Samuel A

    2014-02-01

    Echinocandin-resistant clinical isolates of Candida albicans have been reported, and key-hot spot mutations in the FKS1 gene, which encodes a major glucan synthase subunit, have been identified in these (caspofungin-resistant [CAS-R]) strains. Although these mutations result in phenotypic resistance to echinocandins in planktonic cells, there is little data on antifungal susceptibilities of CAS-R C. albicans strains within biofilms. Thus, we analyzed biofilms formed by 12 C. albicans CAS-R clinical strains in which we previously identified FKS1 hot-spot mutations and compared the sessile antifungal and paradoxical activity of anidulafungin (ANID), caspofungin (CAS), and micafungin (MICA). Biofilms were formed in a 96-well static microplate model and assayed using both tetrazolium-salt reduction and crystal violet assays, as well as examination by scanning electron microscopy. We first sought to assess biofilm formation and structure in these fks1 mutants and found that the biofilm mass and metabolic activities were reduced in most of the fks1 mutants as compared with reference strain SC5314. Structural analyses revealed that the fks1 mutant biofilms were generally less dense and had a clear predominance of yeast and pseudohyphae, with unusual "pit"-like cell surface structures. We also noted that sessile minimum inhibitory concentrations (MICs) to ANID, CAS, and MICA were higher than planktonic MICs of all but one strain. The majority of strains demonstrated a paradoxical effect (PE) to particular echinocandins, in either planktonic or sessile forms. Overall, biofilms formed by echinocandin-resistant clinical isolates demonstrated varied PEs to echinocandins and were structurally characterized by a preponderance of yeast, pseudohyphae, and pit-like structures.

  1. The effect of Streptococcus mutans and Candida glabrata on Candida albicans biofilms formed on different surfaces

    NARCIS (Netherlands)

    Pereira-Cenci, T.; Deng, D.M.; Kraneveld, E.A.; Manders, E.M.M.; Del Bel Cury, A.A.; ten Cate, J.M.; Crielaard, W.

    2008-01-01

    Although Candida containing biofilms contribute to the development of oral candidosis, the characteristics of multi-species Candida biofilms and how oral bacteria modulate these biofilms is poorly understood. The aim of this study was to investigate interactions between Candida albicans and either C

  2. Control of the Biofilms Formed by Curli- and Cellulose-Expressing Shiga Toxin-Producing Escherichia coli Using Treatments with Organic Acids and Commercial Sanitizers.

    Science.gov (United States)

    Park, Yoen Ju; Chen, Jinru

    2015-05-01

    Biofilms are a mixture of bacteria and extracellular products secreted by bacterial cells and are of great concern to the food industry because they offer physical, mechanical, and biological protection to bacterial cells. This study was conducted to quantify biofilms formed by different Shiga toxin-producing Escherichia coli (STEC) strains on polystyrene and stainless steel surfaces and to determine the effectiveness of sanitizing treatments in control of these biofilms. STEC producing various amounts of cellulose (n = 6) or curli (n = 6) were allowed to develop biofilms on polystyrene and stainless steel surfaces at 28°C for 7 days. The biofilms were treated with 2% acetic or lactic acid and manufacturer-recommended concentrations of acidic or alkaline sanitizers, and residual biofilms were quantified. Treatments with the acidic and alkaline sanitizers were more effective than those with the organic acids for removing the biofilms. Compared with their counterparts, cells expressing a greater amount of cellulose or curli formed more biofilm mass and had greater residual mass after sanitizing treatments on polystyrene than on stainless steel. Research suggests that the organic acids and sanitizers used in the present study differed in their ability to control biofilms. Bacterial surface components and cell contact surfaces can influence both biofilm formation and the efficacy of sanitizing treatments. These results provide additional information on control of biofilms formed by STEC.

  3. Atypical Enteropathogenic Escherichia coli Strains form Biofilm on Abiotic Surfaces Regardless of Their Adherence Pattern on Cultured Epithelial Cells

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    Hebert F. Culler

    2014-01-01

    Full Text Available The aim of this study was to determine the capacity of biofilm formation of atypical enteropathogenic Escherichia coli (aEPEC strains on abiotic and biotic surfaces. Ninety-one aEPEC strains, isolated from feces of children with diarrhea, were analyzed by the crystal violet (CV assay on an abiotic surface after 24 h of incubation. aEPEC strains representing each HEp-2 cell type of adherence were analyzed after 24 h and 6, 12, and 18 days of incubation at 37°C on abiotic and cell surfaces by CFU/cm2 counting and confocal laser scanning microscopy (CLSM. Biofilm formation on abiotic surfaces occurred in 55 (60.4% of the aEPEC strains. There was no significant difference in biofilm biomass formation on an abiotic versus prefixed cell surface. The biofilms could be visualized by CLSM at various developmental stages. aEPEC strains are able to form biofilm on an abiotic surface with no association with their adherence pattern on HEp-2 cells with the exception of the strains expressing UND (undetermined adherence. This study revealed the capacity of adhesion and biofilm formation by aEPEC strains on abiotic and biotic surfaces, possibly playing a role in pathogenesis, mainly in cases of persistent diarrhea.

  4. 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.

  5. Intricate interactions between the bloom-forming cyanobacterium Microcystis aeruginosa and foreign genetic elements, revealed by diversified clustered regularly interspaced short palindromic repeat (CRISPR) signatures.

    Science.gov (United States)

    Kuno, Sotaro; Yoshida, Takashi; Kaneko, Takakazu; Sako, Yoshihiko

    2012-08-01

    Clustered regularly interspaced short palindromic repeats (CRISPR) confer sequence-dependent, adaptive resistance in prokaryotes against viruses and plasmids via incorporation of short sequences, called spacers, derived from foreign genetic elements. CRISPR loci are thus considered to provide records of past infections. To describe the host-parasite (i.e., cyanophages and plasmids) interactions involving the bloom-forming freshwater cyanobacterium Microcystis aeruginosa, we investigated CRISPR in four M. aeruginosa strains and in two previously sequenced genomes. The number of spacers in each locus was larger than the average among prokaryotes. All spacers were strain specific, except for a string of 11 spacers shared in two closely related strains, suggesting diversification of the loci. Using CRISPR repeat-based PCR, 24 CRISPR genotypes were identified in a natural cyanobacterial community. Among 995 unique spacers obtained, only 10 sequences showed similarity to M. aeruginosa phage Ma-LMM01. Of these, six spacers showed only silent or conservative nucleotide mutations compared to Ma-LMM01 sequences, suggesting a strategy by the cyanophage to avert CRISPR immunity dependent on nucleotide identity. These results imply that host-phage interactions can be divided into M. aeruginosa-cyanophage combinations rather than pandemics of population-wide infectious cyanophages. Spacer similarity also showed frequent exposure of M. aeruginosa to small cryptic plasmids that were observed only in a few strains. Thus, the diversification of CRISPR implies that M. aeruginosa has been challenged by diverse communities (almost entirely uncharacterized) of cyanophages and plasmids.

  6. Histamine-producing Lactobacillus parabuchneri strains isolated from grated cheese can form biofilms on stainless steel.

    Science.gov (United States)

    Diaz, Maria; Del Rio, Beatriz; Sanchez-Llana, Esther; Ladero, Victor; Redruello, Begoña; Fernández, María; Martin, M Cruz; Alvarez, Miguel A

    2016-10-01

    The consumption of food containing large amounts of histamine can lead to histamine poisoning. Cheese is one of the most frequently involved foods. Histamine, one of the biogenic amines (BAs) exhibiting the highest safety risk, accumulates in food contaminated by microorganisms with histidine decarboxylase activity. The origin of these microorganisms may be very diverse with contamination likely occurring during post-ripening processing, but the microorganisms involved during this manufacturing step have never been identified. The present work reports the isolation of 21 histamine-producing Lactobacillus parabuchneri strains from a histamine-containing grated cheese. PCR revealed that every isolate carried the histidine decarboxylase gene (hdcA). Eight lineages were identified based on the results of genome PFGE restriction analysis plus endonuclease restriction profile analysis of the carried plasmids. Members of all lineages were able to form biofilms on polystyrene and stainless steel surfaces. L. parabuchneri is therefore an undesirable species in the dairy industry; the biofilms it can produce on food processing equipment represent a reservoir of histamine-producing bacteria and thus a source of contamination of post-ripening-processed cheeses.

  7. THE COMPLEX EXTRACELLULAR POLYSACCHARIDES OF MAINLY CHAIN-FORMING FRESHWATER DIATOM SPECIES FROM EPILITHIC BIOFILMS(1).

    Science.gov (United States)

    Bahulikar, Rahul A; Kroth, Peter G

    2008-12-01

    Diatoms are dominant organisms in phototrophic biofilms in aquatic habitats. They produce copious amounts of extracellular polymeric substances (EPS), which mainly consist of carbohydrates and traces of proteins and glycoproteins. This study focuses on the characterization of EPS from a total of 14 diatoms belonging to the six genera Achnanthes, Cymbella, Fragilaria, Punctastriata, Staurosira, and Pseudostaurosira, all of which were isolated from epilithic biofilms of the littoral zone of Lake Constance. EPS from all isolates were extracted by a sequential extraction procedure resulting in five different fractions. The monosaccharide composition of each fraction was analyzed by HPLC equipped with a pulse amperiometric detector, yielding results similar to those obtained by probing the EPS structures with monomer-specific fluorophore-linked lectins. Significant differences in carbohydrate composition occurred in the different fractions of single isolates. Most of the diatom isolates in our study form chain-like colonies in which the cells are attached to each other by intercellular pads. Here we demonstrate that these pads can be dissolved in hot bicarbonate and that they show a heterogeneous composition of monosaccharides in contrast to other fractions, which mostly were dominated by one or two monosaccharides. Principal component analysis indicates a correlation between carbohydrate composition of EPS fractions and the phylogenetic relationship of the respective species, indicating that EPS analyses under defined culture conditions may support taxonomic analyses.

  8. Zinc-dependent mechanical properties of Staphylococcus aureus biofilm-forming surface protein SasG.

    Science.gov (United States)

    Formosa-Dague, Cécile; Speziale, Pietro; Foster, Timothy J; Geoghegan, Joan A; Dufrêne, Yves F

    2016-01-12

    Staphylococcus aureus surface protein SasG promotes cell-cell adhesion during the accumulation phase of biofilm formation, but the molecular basis of this interaction remains poorly understood. Here, we unravel the mechanical properties of SasG on the surface of living bacteria, that is, in its native cellular environment. Nanoscale multiparametric imaging of living bacteria reveals that Zn(2+) strongly increases cell wall rigidity and activates the adhesive function of SasG. Single-cell force measurements show that SasG mediates cell-cell adhesion via specific Zn(2+)-dependent homophilic bonds between β-sheet-rich G5-E domains on neighboring cells. The force required to unfold individual domains is remarkably strong, up to ∼500 pN, thus explaining how SasG can withstand physiological shear forces. We also observe that SasG forms homophilic bonds with the structurally related accumulation-associated protein of Staphylococcus epidermidis, suggesting the possibility of multispecies biofilms during host colonization and infection. Collectively, our findings support a model in which zinc plays a dual role in activating cell-cell adhesion: adsorption of zinc ions to the bacterial cell surface increases cell wall cohesion and favors the projection of elongated SasG proteins away from the cell surface, thereby enabling zinc-dependent homophilic bonds between opposing cells. This work demonstrates an unexpected relationship between mechanics and adhesion in a staphylococcal surface protein, which may represent a general mechanism among bacterial pathogens for activating cell association.

  9. Relationship between clinical site of isolation and ability to form biofilms in vitro in nontypeable Haemophilus influenzae.

    Science.gov (United States)

    Obaid, Najla A; Jacobson, Glenn A; Tristram, Stephen

    2015-03-01

    Nontypeable Haemophilus influenzae (NTHi) is an opportunistic pathogen associated with a range of infections, including various lower respiratory infections, otitis media, and conjunctivitis. There is some debate as to whether or not NTHi produces biofilms and, if so, whether or not this is relevant to pathogenesis. Although many studies have examined the association between in vitro biofilm formation and isolates from a specific infection type, few have made comparisons from isolates from a broad range of isolates grouped by clinical source. In our study 50 NTHi from different clinical sources, otitis media, conjunctivitis, lower respiratory tract infections in both cystic fibrosis and non-cystic fibrosis patients, and nasopharyngeal carriage, plus 10 nasopharyngeal isolates of the commensal Haemophilus haemolyticus were tested for the ability to form biofilm by using a static microtitre plate crystal violet assay. A high degree of variation in biofilm forming ability was observed across all isolates, with no statistically significant differences observed between the groups, with the exception of the isolates from conjunctivitis. These isolates had uniformly lower biofilm forming ability compared with isolates from the other groups (p < 0.005).

  10. Synthesis of Bioinspired Carbohydrate Amphiphiles that Promote and Inhibit Biofilms.

    Science.gov (United States)

    Dane, Eric L; Ballok, Alicia E; O'Toole, George A; Grinstaff, Mark W

    2014-02-01

    The synthesis and characterization of a new class of bioinspired carbohydrate amphiphiles that modulate Pseudomonas aeruginosa biofilm formation are reported. The carbohydrate head is an enantiopure poly-amido-saccharide (PAS) prepared by a controlled anionic polymerization of β-lactam monomers derived from either glucose or galactose. The supramolecular assemblies formed by PAS amphiphiles are investigated in solution using fluorescence assays and dynamic light scattering. Dried samples are investigated using X-ray, infrared spectroscopy, and transmission electron microscopy. Additionally, the amphiphiles are evaluated for their ability to modulate biofilm formation by the Gram-negative bacterium Pseudomonas aeruginosa. Remarkably, from a library of eight amphiphiles, we identify a structure that promotes biofilm formation and two structures that inhibit biofilm formation. Using biological assays and electron microscopy, we relate the chemical structure of the amphiphiles to the observed activity. Materials that modulate the formation of biofilms by bacteria are important both as research tools for microbiologists to study the process of biofilm formation and for their potential to provide new drug candidates for treating biofilm-associated infections.

  11. Effects of environmental parameters on the dual-species biofilms formed by Escherichia coli O157:H7 and Ralstonia insidiosa, a strong biofilm producer isolated from a fresh-cut produce processing plant.

    Science.gov (United States)

    Liu, Nancy T; Nou, Xiangwu; Bauchan, Gary R; Murphy, Charles; Lefcourt, Alan M; Shelton, Daniel R; Lo, Y Martin

    2015-01-01

    Biofilm-forming bacteria resident to food processing facilities are a food safety concern due to the potential of biofilms to harbor foodborne bacterial pathogens. When cultured together, Ralstonia insidiosa, a strong biofilm former frequently isolated from produce processing environments, has been shown to promote the incorporation of Escherichia coli O157:H7 into dual-species biofilms. In this study, interactions between E. coli O157:H7 and R. insidiosa were examined under different incubating conditions. Under static culture conditions, the incorporation of E. coli O157:H7 into biofilms with R. insidiosa was not significantly affected by either low incubating temperature (10°C) or by limited nutrient availability. Greater enhancement of E. coli O157:H7 incorporation in dual-species biofilms was observed by using a continuous culture system with limited nutrient availability. Under the continuous culture conditions used in this study, E coli O157:H7 cells showed a strong tendency of colocalizing with R. insidiosa on a glass surface at the early stage of biofilm formation. As the biofilms matured, E coli O157:H7 cells were mostly found at the bottom layer of the dual-species biofilms, suggesting an effective protection by R. insidiosa in the mature biofilms.

  12. Raman spectroscopic differentiation of planktonic bacteria and biofilms.

    Science.gov (United States)

    Kusić, Dragana; Kampe, Bernd; Ramoji, Anuradha; Neugebauer, Ute; Rösch, Petra; Popp, Jürgen

    2015-09-01

    Both biofilm formations as well as planktonic cells of water bacteria such as diverse species of the Legionella genus as well as Pseudomonas aeruginosa, Klebsiella pneumoniae, and Escherichia coli were examined in detail by Raman microspectroscopy. Production of various molecules involved in biofilm formation of tested species in nutrient-deficient media such as tap water was observed and was particularly evident in the biofilms formed by six Legionella species. Biofilms of selected species of the Legionella genus differ significantly from the planktonic cells of the same organisms in their lipid amount. Also, all Legionella species have formed biofilms that differ significantly from the biofilms of the other tested genera in the amount of lipids they produced. We believe that the significant increase in the synthesis of this molecular species may be associated with the ability of Legionella species to form biofilms. In addition, a combination of Raman microspectroscopy with chemometric approaches can distinguish between both planktonic form and biofilms of diverse bacteria and could be used to identify samples which were unknown to the identification model. Our results provide valuable data for the development of fast and reliable analytic methods based on Raman microspectroscopy, which can be applied to the analysis of tap water-adapted microorganisms without any cultivation step.

  13. Assessment of Changes in Biodiversity when a Community of Ultramicrobacteria Isolated from Groundwater Is Stimulated to Form a Biofilm.

    Science.gov (United States)

    Ross, N.; Villemur, R.; Marcandella, E.; Deschênes, L.

    2001-07-01

    The stimulation of groundwater bacteria to form biofilms, for the remediation of polluted aquifers, is subjected to environmental regulations that include measurement of effects on microbial biodiversity. Groundwater microorganisms contain a proportion of unidentified and uncharacterized ultramicrobacteria (UMB) that might play a major role in the bioclogging of geological materials. This study aimed to assess the changes in genetic and metabolic biodiversity when a community of UMB, isolated from groundwater, is stimulated to form biofilms on a ceramic surface. UMB were stimulated with aerobic conditions and injection of molasses, in reactors reproducing groundwater composition and temperature. Concentration of planktonic viable UMB, secretion of extracellular polymeric substances (EPS), and biofilm thickness were monitored. The assessment of changes in biodiversity was achieved by comparing the initial UMB community to the biofilm community, using the single strand conformational polymorphism (SSCP) method, the cloning and sequencing of 16S rRNA gene (16S rDNA) sequences, and the Biolog microplate system. The hypothesis stating that indigenous UMB would play a significant role of in the biofilm development was corroborated. Within 13 days of stimulation, the UMB produced 700 mg L?1 of planktonic EPS and formed a biofilm up to a thickness of 1100 mm. This stimulation led to a decrease in genetic diversity and an increase in metabolic diversity. The decrease in genetic diversity was shown by a reduced number of single strand DNA fragments in the SSCP profiles. As such, 16S rDNA sequences from the biofilm revealed the predominance of four bacterial groups: Zoogloea, Bacillus/Paenibacillus, Enterobacteriaceae, and Pseudomonads. A significant increase in metabolic diversity was shown by a highest substrate richness profile and a lower substrate evenness profile of the biofilm bacterial population (p = 0.0 and p = 0.09, respectively). This higher metabolic diversity

  14. Enzymatic quorum quenching increases antibiotic susceptibility of multidrug resistant Pseudomonas aeruginosa

    Science.gov (United States)

    Kiran, S; Sharma, P; Harjai, K; Capalash, N

    2011-01-01

    Background and Objectives There is increasing emergence of multidrug resistant Pseudomonas aeruginosa (MDRPA) strains and drug resistance is positively-correlated with biofilm-forming ability. Since about 10% of P. aeruginosa genome is controlled by quorum sensing (QS), alteration in its antibiotic susceptibility by targeting QS was the focus of the present study. Materials and Methods One day biofilms of PAO1 and three urinary tract infection MDRPA isolates (PA2, PA8 and PA18) were formed in 96-well microtiter plate. Biofilms were exposed to concentration gradient of ciprofloxacin and gentamicin to obtain Minimum Biofilm Eradication Concentration (MBEC) by direct enumeration method. Susceptibility of 24 h biofilms was evaluated by treatment with ciprofloxacin and gentamicin per se and in combination with lactonase. The effect was also examined on 72 h biofilms by Scanning Electron Microscopy. Results Lactonase treatment did not have any effect on growth of the selected strains but 73.42, 69.1, 77.34 and 72.5% reduction of biofilm was observed after lactonase (1 unit) treatment, respectively. Antibiotics in combination with lactonase (0.3 units) resulted in an increased susceptibility of the biofilm forms by>3.3, 4, 5 and 1.5 folds of MBEC, for ciprofloxacin and>6.67, 12.5, 6 and>2.5 folds, for gentamicin respectively, which could be due to the disruption of biofilm by lactonase treatment as shown by scanning electron microscopy. Also there was significant reduction (p<0.001) in virulence factor production by the strains. Conclusion Lactonase treatment increased antibiotic susceptibility of the biofilms of MDRPA isolates underscoring the potential of quorum quenching in antimicrobial therapeutics. PMID:22347576

  15. Co-occurence of filamentation defects and impaired biofilms in Candida albicans protein kinase mutants.

    Science.gov (United States)

    Konstantinidou, Nina; Morrissey, John Patrick

    2015-12-01

    Pathogenicity of Candida albicans is linked with its developmental stages, notably the capacity switch from yeast-like to hyphal growth, and to form biofilms on surfaces. To better understand the cellular processes involved in C. albicans development, a collection of 63 C. albicans protein kinase mutants was screened for biofilm formation in a microtitre plate assay. Thirty-eight mutants displayed some degree of biofilm impairment, with 20 categorised as poor biofilm formers. All the poor biofilm formers were also defective in the switch from yeast to hyphae, establishing it as a primary defect. Five genes, VPS15, IME2, PKH3, PGA43 and CEX1, encode proteins not previously reported to influence hyphal development or biofilm formation. Network analysis established that individual components of some processes, most interestingly MAP kinase pathways, are not required for biofilm formation, most likely indicating functional redundancy. Mutants were also screened for their response to bacterial supernatants and it was found that Pseudomonas aeruginosa supernatants inhibited biofilm formation in all mutants, regardless of the presence of homoserine lactones (HSLs). In contrast, Candida morphology was only affected by supernatant containing HSLs. This confirms the distinct HSL-dependent inhibition of filamentation and the HSL-independent impairment of biofilm development by P. aeruginosa.

  16. Effect of peracetic acid on biofilms formed by Staphylococcus aureus and Listeria monocytogenes isolated from dairy plants.

    Science.gov (United States)

    Lee, S H I; Cappato, L P; Corassin, C H; Cruz, A G; Oliveira, C A F

    2016-03-01

    This research investigated the removal of adherent cells of 4 strains of Staphylococcus aureus and 1 Listeria monocytogenes strain (previously isolated from dairy plants) from polystyrene microtiter plates using peracetic acid (PAA, 0.5%) for 15, 30, 60, and 120 s, and the inactivation of biofilms formed by those strains on stainless steel coupons using the same treatment times. In the microtiter plates, PAA removed all S. aureus at 15 s compared with control (no PAA treatment). However, L. monocytogenes biofilm was not affected by any PAA treatment. On the stainless steel surface, epifluorescence microscopy using LIVE/DEAD staining (BacLight, Molecular Probes/Thermo Fisher Scientific, Eugene, OR) showed that all strains were damaged within 15 s, with almost 100% of cells inactivated after 30 s. Results of this trial indicate that, although PAA was able to inactivate both S. aureus and L. monocytogenes monospecies biofilms on stainless steel, it was only able to remove adherent cells of S. aureus from polystyrene microplates. The correct use of PAA is critical for eliminating biofilms formed by S. aureus strains found in dairy plants, although further studies are necessary to determine the optimal PAA treatment for removing biofilms of L. monocytogenes.

  17. Investigation of the antibiotic resistance and biofilm-forming ability of Staphylococcus aureus from subclinical bovine mastitis cases.

    Science.gov (United States)

    Aslantaş, Özkan; Demir, Cemil

    2016-11-01

    A total of 112 Staphylococcus aureus isolates obtained from subclinical bovine mastitis cases were examined for antibiotic susceptibility and biofilm-forming ability as well as genes responsible for antibiotic resistance, biofilm-forming ability, and adhesin. Antimicrobial susceptibility of the isolates were determined by disk diffusion method. Biofilm forming ability of the isolates were investigated by Congo red agar method, standard tube method, and microplate method. The genes responsible for antibiotic resistance, biofilm-forming ability, and adhesion were examined by PCR. Five isolates (4.5%) were identified as methicillin-resistant Staph. aureus by antibiotic susceptibility testing and confirmed by mecA detection. The resistance rates to penicillin, ampicillin, tetracycline, erythromycin, trimethoprim-sulfamethoxazole, enrofloxacin, and amoxicillin-clavulanic acid were 45.5, 39.3, 33, 26.8, 5.4, 0.9, and 0.9%, respectively. All isolates were susceptible against vancomycin and gentamicin. The blaZ (100%), tetK (67.6%), and ermA (70%) genes were the most common antibiotic-resistance genes. Using Congo red agar, microplate, and standard tube methods, 70.5, 67, and 62.5% of the isolates were found to be biofilm producers, respectively. The percentage rate of icaA, icaD, and bap genes in Staph. aureus isolates were 86.6, 86.6, and 13.4%, respectively. The adhesion molecules fnbA, can, and clfA were detected in 87 (77.7%), 98 (87.5%), and 75 (70%) isolates, respectively. The results indicated that Staph. aureus from sublinical bovine mastitis cases were mainly resistant to β-lactams and, to a lesser extent, to tetracycline and erythromycin. Also, biofilm- and adhesion-related genes, which are increasingly accepted as an important virulence factor in the pathogenesis of Staph. aureus infections, were detected at a high rate.

  18. Cyanophage infection in the bloom-forming cyanobacteria Microcystis aeruginosa in surface freshwater.

    Science.gov (United States)

    Yoshida-Takashima, Yukari; Yoshida, Mitsuhiro; Ogata, Hiroyuki; Nagasaki, Keizo; Hiroishi, Shingo; Yoshida, Takashi

    2012-01-01

    Host-like genes are often found in viral genomes. To date, multiple host-like genes involved in photosynthesis and the pentose phosphate pathway have been found in phages of marine cyanobacteria Synechococcus and Prochlorococcus. These gene products are predicted to redirect host metabolism to deoxynucleotide biosynthesis for phage replication while maintaining photosynthesis. A cyanophage, Ma-LMM01, infecting the toxic cyanobacterium Microcystis aeruginosa, was isolated from a eutrophic freshwater lake and assigned as a member of a new lineage of the Myoviridae family. The genome encodes a host-like NblA. Cyanobacterial NblA is known to be involved in the degradation of the major light harvesting complex, the phycobilisomes. Ma-LMM01 nblA gene showed an early expression pattern and was highly transcribed during phage infection. We speculate that the co-option of nblA into Microcystis phages provides a significant fitness advantage to phages by preventing photoinhibition during infection and possibly represents an important part of the co-evolutionary interactions between cyanobacteria and their phages.

  19. Characterization of structures in biofilms formed by a Pseudomonas fluorescens isolated from soil

    Directory of Open Access Journals (Sweden)

    Wu Siva

    2009-05-01

    Full Text Available Abstract Background Microbial biofilms represent an incompletely understood, but fundamental mode of bacterial growth. These sessile communities typically consist of stratified, morphologically-distinct layers of extracellular material, where numerous metabolic processes occur simultaneously in close proximity. Limited reports on environmental isolates have revealed highly ordered, three-dimensional organization of the extracellular matrix, which may hold important implications for biofilm physiology in vivo. Results A Pseudomonas spp. isolated from a natural soil environment produced flocculent, nonmucoidal biofilms in vitro with unique structural features. These mature biofilms were made up of numerous viable bacteria, even after extended culture, and contained up to 50% of proteins and accumulated 3% (by dry weight calcium, suggesting an important role for the divalent metal in biofilm formation. Ultrastructurally, the mature biofilms contained structural motifs consisting of dense, fibrillary clusters, nanofibers, and ordered, honeycomb-like chambers enveloped in thin sheets. Conclusion Mature biofilms contained living bacteria and were structurally, chemically, and physiologically heterogeneous. The principal architectural elements observed by electron microscopy may represent useful morphological clues for identifying bacterial biofilms in vivo. The complexity and reproducibility of the structural motifs observed in bacterial biofilms appear to be the result of organized assembly, suggesting that this environmental isolate may possess ecological advantages in its natural habitat.

  20. Particulate Bioglass reduces the viability of bacterial biofilms formed on its surface in an in vitro model.

    Science.gov (United States)

    Allan, Iain; Newman, Hubert; Wilson, Michael

    2002-02-01

    45S5 Bioglass is a bioactive implant material which, in its particulate form, is used in the repair of periodontal defects. The surface reactions undergone by this material in an aqueous environment may exert an antibacterial effect that would be beneficial to periodontal surgical treatment. Biofilms of Streptococcus sanguis, an early plaque former, and mixed species biofilms from a salivary inoculum grown under conditions similar to those associated with periodontal implants, were grown on particulate Bioglass in a constant depth film fermenter (CDFF). Control biofilms were grown on inert glass particulates. At sample times of 3, 24 and 48 hours the viability of biofilms of S. sanguis grown on Bioglass was significantly lower than for those grown on inert glass. In the experiments with subgingivally-modelled mixed species biofilms, the total anaerobic counts were significantly lower on Bioglass after 24 and 48 hours, but not 96 or 168 hours, compared to inert glass. Thus, particulate Bioglass has the potential to reduce bacterial colonisation of its surface in vivo, a feature relevant to post-surgical periodontal wound healing.

  1. Staphylococcus aureus Clinical Isolates: Antibiotic Susceptibility, Molecular Characteristics, and Ability to Form Biofilm

    Directory of Open Access Journals (Sweden)

    N. Indrawattana

    2013-01-01

    Full Text Available Periodic monitoring of Staphylococcus aureus characteristics in a locality is imperative as their drug-resistant variants cause treatment problem. In this study, antibiograms, prevalence of toxin genes (sea-see, seg-ser, seu, tsst-1, eta, etb, and etd, PFGE types, accessory gene regulator (agr groups, and ability to form biofilm of 92 S. aureus Thailand clinical isolates were investigated. They were classified into 10 drug groups: groups 1–7 (56 isolates were methicillin resistant (MRSA and 8–10 (36 isolates were methicillin sensitive (MSSA. One isolate did not have any toxin gene, 4 isolates carried one toxin gene (seq, and 87 isolates had two or more toxin genes. No isolate had see, etb, or tsst-1; six isolates had eta or etd. Combined seg-sei-sem-sen-seo of the highly prevalent egc locus was 26.1%. The seb, sec, sel, seu, and eta associated significantly with MSSA; sek was more in MRSA. The sek-seq association was 52.17% while combined sed-sej was not found. Twenty-three PFGE types were revealed, no association of toxin genes with PFGE types. All four agr groups were present; agr group 1 was predominant (58.70% but agr group 2 strains carried more toxin genes and were more frequent toxin producers. Biofilm formation was found in 72.83% of the isolates but there was no association with antibiograms. This study provides insight information on molecular and phenotypic markers of Thailand S. aureus clinical isolates which should be useful for future active surveillance that aimed to control a spread of existing antimicrobial resistant bacteria and early recognition of a newly emerged variant.

  2. D-Amino Acids Enhance the Activity of Antimicrobials against Biofilms of Clinical Wound Isolates of Staphylococcus aureus and Pseudomonas aeruginosa

    Science.gov (United States)

    2014-05-19

    biofilm eradication concentration high-throughput (MBEC- HTP ) assay plates (Innovotech, Canada) for biofilm antimicrobial susceptibility test - ing (38, 39...susceptibility testing . M100-S22. CLSI, Wayne, PA. 38. Coraca-Huber DC, Fille M, Hausdorfer J, Pfaller K, Nogler M. 2012. Evaluation of MBEC- HTP biofilm model...Standards Institute (37). Test performance for antimicrobial agents was monitored using P. aerugi- nosa ATCC 27853 and S. aureus ATCC 29213 as control

  3. Antibiofilm and anti-infection of a marine bacterial exopolysaccharide against Pseudomonas aeruginosa.

    Directory of Open Access Journals (Sweden)

    Shimei eWu

    2016-02-01

    Full Text Available Pseudomonas aeruginosa is a well-known pathogenic bacterium that forms biofilms and produces virulence factors, thus leading to major problems in many fields, such as clinical infection, food contamination and marine biofouling. In this study, we report the purification and characterization of an exopolysaccharide EPS273 from the culture supernatant of marine bacterium Pseudomonas stutzeri 273. The exopolysaccharide EPS273 not only effectively inhibits biofilm formation but also disperses preformed biofilm of Pseudomonas aeruginosa PAO1. High performance liquid chromatography traces of the hydrolyzed polysaccharides shows that EPS273 primarily consists of glucosamine, rhamnose, glucose and mannose. Further investigation demonstrates that EPS273 reduces the production of the virulence factors pyocyanin, exoprotease and rhamnolipid, and the virulence of P. aeruginosa PAO1 to human lung cells A549 and zebrafish embryos is also obviously attenuated by EPS273. In addition, EPS273 also greatly reduces the production of hydrogen peroxide (H2O2 and extracellular DNA (eDNA, which are important factors for biofilm formation. Furthermore, EPS273 exhibits strong antioxidant potential by quenching hydroxyl and superoxide anion radicals. Notably, the antibiofouling activity of EPS273 is observed in the marine environment up to two weeks according to the amounts of bacteria and diatoms in the glass slides submerged in the ocean. Taken together, the properties of EPS273 indicate that it has a promising prospect in combating bacterial biofilm-associated infection, food-processing contamination and marine biofouling.

  4. Multi-channel microfluidic biosensor platform applied for online monitoring and screening of biofilm formation and activity.

    Science.gov (United States)

    Bruchmann, Julia; Sachsenheimer, Kai; Rapp, Bastian E; Schwartz, Thomas

    2015-01-01

    Bacterial colonization of surfaces and interfaces has a major impact on various areas including biotechnology, medicine, food industries, and water technologies. In most of these areas biofilm development has a strong impact on hygiene situations, product quality, and process efficacies. In consequence, biofilm manipulation and prevention is a fundamental issue to avoid adverse impacts. For such scenario online, non-destructive biofilm monitoring systems become important in many technical and industrial applications. This study reports such a system in form of a microfluidic sensor platform based on the combination of electrical impedance spectroscopy and amperometric current measurement, which allows sensitive online measurement of biofilm formation and activity. A total number of 12 parallel fluidic channels enable real-time online screening of various biofilms formed by different Pseudomonas aeruginosa and Stenotrophomonas maltophilia strains and complex mixed population biofilms. Experiments using disinfectant and antibiofilm reagents demonstrate that the biofilm sensor is able to discriminate between inactivation/killing of bacteria and destabilization of biofilm structures. The impedance and amperometric sensor data demonstrated the high dynamics of biofilms as a consequence of distinct responses to chemical treatment strategies. Gene expression of flagellar and fimbrial genes of biofilms grown inside the microfluidic system supported the detected biofilm growth kinetics. Thus, the presented biosensor platform is a qualified tool for assessing biofilm formation in specific environments and for evaluating the effectiveness of antibiofilm treatment strategies.

  5. An in vitro urinary tract catheter system to investigate biofilm development in catheter-associated urinary tract infections.

    Science.gov (United States)

    Dohnt, Katrin; Sauer, Marie; Müller, Maren; Atallah, Karin; Weidemann, Marina; Gronemeyer, Petra; Rasch, Detlev; Tielen, Petra; Krull, Rainer

    2011-12-01

    Biofilm development in urinary tract catheters is an often underestimated problem. However, this form of infection leads to high mortality rates and causes significant costs in health care. Therefore, it is important to analyze these biofilms and establish avoiding strategies. In this study a continuous flow-through system for the cultivation of biofilms under catheter-associated urinary tract infection conditions was established and validated. The in vitro urinary tract catheter system implies the composition of urine (artificial urine medium), the mean volume of urine of adults (1 mL min(-1)), the frequently used silicone catheter (foley silicon catheter) as well as the infection with uropathogenic microorganisms like Pseudomonas aeruginosa. Three clinical isolates from urine of catheterized patients were chosen due to their ability to form biofilms, their mobility and their cell surface hydrophobicity. As reference strain P. aeruginosa PA14 has been used. Characteristic parameters as biofilm thickness, specific biofilm growth rate and substrate consumption were observed. Biofilm thicknesses varied from 105±16 μm up to 246±67 μm for the different isolates. The specific biofilm growth rate could be determined with a non invasive optical biomass sensor. This sensor allows online monitoring of the biofilm growth in the progress of the cultivation.

  6. Bacteriophages as an alternative strategy for fighting biofilm development.

    Science.gov (United States)

    Parasion, Sylwia; Kwiatek, Magdalena; Gryko, Romuald; Mizak, Lidia; Malm, Anna

    2014-01-01

    The ability of microbes to form biofilms is an important element of their pathogenicity, and biofilm formation is a serious challenge for today's medicine. Fighting the clinical complications associated with biofilm formation is very difficult and linked to a high risk of failure, especially in a time of increasing bacterial resistance to antibiotics. Bacterial species most commonly isolated from biofilms include coagulase-negative staphylococci, Staphylococcus aureus, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, Proteus mirabilis, Klebsiella pneumoniae, Pseudomonas aeruginosa and Acinetobacter spp. The frequent failure of antibiotic therapy led researchers to look for alternative methods and experiment with the use of antibacterial factors with a mechanism of action different from that of antibiotics. Experimental studies with bacteriophages and mixtures thereof, expressing lytic properties against numerous biofilm-forming bacterial species showed that bacteriophages may both prevent biofilm formation and contribute to eradication of biofilm bacteria. A specific role is played here by phage depolymerases, which facilitate the degradation of extracellular polymeric substances (EPS) and thus the permeation of bacteriophages into deeper biofilm layers and lysis of the susceptible bacterial cells. Much hope is placed in genetic modifications of bacteriophages that would allow the equipping bacteriophages with the function of depolymerase synthesis. The use of phage cocktails prevents the development of phage-resistant bacteria.

  7. A semi-quantitative approach to assess biofilm formation using wrinkled colony development.

    Science.gov (United States)

    Ray, Valerie A; Morris, Andrew R; Visick, Karen L

    2012-06-07

    Biofilms, or surface-attached communities of cells encapsulated in an extracellular matrix, represent a common lifestyle for many bacteria. Within a biofilm, bacterial cells often exhibit altered physiology, including enhanced resistance to antibiotics and other environmental stresses. Additionally, biofilms can play important roles in host-microbe interactions. Biofilms develop when bacteria transition from individual, planktonic cells to form complex, multi-cellular communities. In the laboratory, biofilms are studied by assessing the development of specific biofilm phenotypes. A common biofilm phenotype involves the formation of wrinkled or rugose bacterial colonies on solid agar media. Wrinkled colony formation provides a particularly simple and useful means to identify and characterize bacterial strains exhibiting altered biofilm phenotypes, and to investigate environmental conditions that impact biofilm formation. Wrinkled colony formation serves as an indicator of biofilm formation in a variety of bacteria, including both Gram-positive bacteria, such as Bacillus subtilis, and Gram-negative bacteria, such as Vibrio cholerae, Vibrio parahaemolyticus, Pseudomonas aeruginosa, and Vibrio fischeri. The marine bacterium V. fischeri has become a model for biofilm formation due to the critical role of biofilms during host colonization: biofilms produced by V. fischeri promote its colonization of the Hawaiian bobtail squid Euprymna scolopes. Importantly, biofilm phenotypes observed in vitro correlate with the ability of V. fischeri cells to effectively colonize host animals: strains impaired for biofilm formation in vitro possess a colonization defect, while strains exhibiting increased biofilm phenotypes are enhanced for colonization. V. fischeri therefore provides a simple model system to assess the mechanisms by which bacteria regulate biofilm formation and how biofilms impact host colonization. In this report, we describe a semi-quantitative method to assess

  8. Electrocatalytic Study of Pseudomonas Aeruginosa BTE-1 Strain%绿脓杆菌Pseudomonas aeruginosa BTE-1直接电催化特征

    Institute of Scientific and Technical Information of China (English)

    周萍; 张恩仁; 周立; 刁国旺; 牛俊乐

    2012-01-01

    , the temperatures higher than 60 ℃ reduced the electrocatalytic activity of the biofilms quickly. Cyclic voltammetry analysis indicated that P. aeruginosa BTE-1 biofilms formed under anaerobic conditions exhibited an electrochemical catalytic wave in the potential range of -0.4 V to -0.2 V ( vs. SCE), similar to that observed with a typical electricity-generating strain, C, eobacter sulfurreducens. Organic substrates, such as compounds in yeast and glucose, could be oxidized through the catalysis of P. aeruginosa BTE-1 biofilms,whereas acetate could not be catalyzed to oxidize.

  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. Biological features of biofilm-forming ability of Acinetobacter baumannii strains derived from 121 elderly patients with hospital-acquired pneumonia.

    Science.gov (United States)

    Zhang, Duchao; Xia, Jingjing; Xu, Yaping; Gong, Meiliang; Zhou, Yu; Xie, Lixin; Fang, Xiangqun

    2016-02-01

    This study is to investigate a biological activity of Acinetobacter baumannii isolates from sputum specimens of 121 elderly patients with hospital-acquired pneumonia. The ability of the isolates to form biofilms was quantitatively assessed by crystal violet staining, and adhesive property was examined using Giemsa staining. Biofilm-forming ability by the isolates was employed to test antimicrobial resistance and examine sources and clinical manifestations. The isolates grew as biofilm on abiotic surface at the indicated temperatures after a 48 h of incubation. 27.3 % of the isolates were strongly biofilm-positive in the samples, and 84.8 % displayed high adhesion ability (P < 0.05). All of the isolates showed antibiotic resistance at different levels, and the isolates produced strong biofilm exhibited low-level resistance to gentamicin, minocycline and ceftazidime (P < 0.05). The patients' experience in ICU, use of antibiotics and estimation of APACHE II (<17) were related to incidence of strong biofilm formation with no clinical manifestations found in the study. All clinical isolates are able to form biofilms which refer to adhesive efficiency and antibiotic resistance. Patient experiences in ICU surveillance, use of antibiotics and APACHE II scores are involved in biofilm-forming ability by the nosocomial pathogen derived from the hospitalized patients.

  11. Sanitizer resistance of biofilm-forming Salmonella isolated from meat products

    OpenAIRE

    Milan, C.; Agostinetto,A.; R.C.S. Conceição; H.L. Gonzalez; Timm, C. D.

    2015-01-01

    Este estudo avaliou a capacidade de Salmonella enterica subsp. enterica isolada de produtos cárneos formar biofilme e testou sua resistência a diferentes sanitizantes. Vinte cepas foram avaliadas quanto à capacidade de formar biofilme em placas de microtitulação. As cepas formadoras de biofilme foram testadas em superfícies de polietileno de alta densidade, aço inoxidável e vidro e tiveram a sensibilidade ao hipoclorito de sódio e ao iodo avaliada. Duas cepas de Salmonella Enteritidis isolada...

  12. 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

  13. Interactions between laponite and microbial biofilms in porous media: implications for colloid transport and biofilm stability.

    Science.gov (United States)

    Leon-Morales, C Felipe; Leis, Andrew P; Strathmann, Martin; Flemming, Hans-Curt

    2004-09-01

    Quartz sand columns and sand-filled microscope flow cells were used to investigate the transport characteristics of the clay colloid laponite, and a biofilm-forming bacterium, Pseudomonas aeruginosa SG81. Separate experiments were performed with each particle to determine their individual transport characteristics in clean sand columns. In a second set of experiments, bacterial biofilms were formed prior to introduction of the clay colloids. In the independent transport experiments, bacteria and laponite each conformed to known physicochemical principles. A sodium chloride concentration of 7 x 10(-2) M caused complete retention of the laponite within the sand columns. P. aeruginosa SG81 was generally less influenced by ionic strength effects; it showed relatively low mobility at all ionic strengths tested and some (albeit reduced) mobility when introduced to the columns in 1M NaCl, the highest concentration tested, but nevertheless showed reproducible trends. Under conditions favourable to laponite retention and biofilm stability (7 x 10(-2) MNaCl), laponite suspensions were able to remobilise a portion of the attached bacterial biomass. At low ionic strength, the profile of laponite elution was also altered in the presence of a P. aeruginosa biofilm. These observations suggest that while a reduction in ionic strength has a dominant influence on the mobilisation of biological and inorganic colloids, the presence of laponite and biomass can have a distinct influence on the mobility of both types of colloids. Since these events are likely to occur in subsurface environments, our results suggest that colloid-biofilm interactions will have implications for colloid-bound contaminant transport and the remobilisation of pathogens.

  14. 铜绿假单胞菌生物膜与亚抑菌浓度抗菌药物的相关研究%Effect of subinhibitory concentrations on formation of biofilm of Pseudomonas aeruginosa

    Institute of Scientific and Technical Information of China (English)

    李俊娟; 王强; 孙开宇; 蒋捍东

    2012-01-01

    OBJECTIVE To study the effects of the subinhibitory concentrations on the biofilm formation of Pseudomonas aeruginosa (PAE).- METHODS We first determined the minimal inhibitory concentration of azithromycin, amikacin, ciprofloxacin and ceftazidime of the PAE cultured in MHB and LB. We made the static biofilm model of P. aeruginosa in 96-well microtiter plates with the four antibiotics and medical silica gel with the azithromycin concentration, the OD value was determined with crystal violet staining microplate reader, and the biofilm was observed with silver staining microscope. RESULTS The MIC of azithromycin of PAE in MHB and LB were 512 mg/L and 16 mg/L, respectively; the MIC of amikacin, ciprofloxacin and ceftazidime of the PAE were 4 mg/L, 0. 125 mg/L and 4 mg/L, respectively; the results of the static-made biofilm were as follows: in the MHB and LB media, azithromycin could induce the biofilm formation of PAE at the subinhibitory concentration. Ciprofloxacin and ceftazidime could inhibit the biofilm formation at these concentrations. Amikacin could inhibit the biofilm formation at the concentration of 0. 25 to 4 mg/L (1/16MIC-MIC) , but could induce biofilm formation at the concentration of 0. 125 mg/L (1/16 MIC)in MHB; and when PAE was cultured in LB, amikacin could inhibit the biofilm formation at the concentration of 1 to 4 mg/L (1/4MIC-MIC) , and induce the biofilm formation at 0.25 mg/L(l/16MIC). All the results above had statistical differences compared with the control group. Silverstaining results: when the concentration of azithromycin was 8mg/L,the formation of the biofilm was the most, the second was the blank group and it was the least as the concentration was 256 mg/L. CONCLUSION The sensitivity of PAE planktonic bacteria to azithromycin varies in different culture condition; and the azithromycin may induce the biofilm formation at the subinhibitory concentration; ciprofloxacin and ceftazidime can inhibit the biofilm formation; the inhibitory of

  15. Comparison of quantification methods illustrates reduced Pseudomonas aeruginosa activity on nanorough polyvinyl chloride.

    Science.gov (United States)

    Seil, Justin T; Rubien, Nathan M; Webster, Thomas J; Tarquinio, Keiko M

    2011-07-01

    Patients on mechanical ventilators for extended periods of time are faced with a high probability of developing ventilator associated pneumonia. Although this has been mostly addressed through the re-engineering of endotracheal tubes (ETTs) with antimicrobial materials, such material coatings may easily delaminate during use. However, the potential exists to apply nanotechnology to the ETT to avoid delamination but implement antibacterial properties. Selecting a protocol to evaluate in vitro material for anti-infection is difficult, partially due to the existence of conflicting reported methods of analysis. In this study, the susceptibility of conventional and nanorough polymeric materials to bacterial biofilm growth were evaluated. After creating nanorough polyvinyl chloride (PVC) ETTs, Pseudomonas aeruginosa biofilms were then grown on sample surfaces during a 24-h culture. Biofilms were then removed and assayed from sample surfaces using a variety of techniques. Comparisons between the different techniques used for biofilm removal indicated that vortexing provided adequate removal of the biofilm from sample surfaces. Most importantly, a protocol following the vortexing method of biofilm and bacteria removal provided an ∼40% lower yield of colony forming units from nanorough PVC compared to conventional PVC. This suggests that Pseudomonas aeruginosa are less adherent on nanorough PVC than conventional PVC.

  16. Mammary Gland Pathology Subsequent to Acute Infection with Strong versus Weak Biofilm Forming Staphylococcus aureus Bovine Mastitis Isolates: A Pilot Study Using Non-Invasive Mouse Mastitis Model

    Science.gov (United States)

    Gogoi-Tiwari, Jully; Williams, Vincent; Waryah, Charlene Babra; Costantino, Paul; Al-Salami, Hani; Mathavan, Sangeetha; Wells, Kelsi; Tiwari, Harish Kumar; Hegde, Nagendra; Isloor, Shrikrishna; Al-Sallami, Hesham; Mukkur, Trilochan

    2017-01-01

    Background Biofilm formation by Staphylococcus aureus is an important virulence attribute because of its potential to induce persistent antibiotic resistance, retard phagocytosis and either attenuate or promote inflammation, depending upon the disease syndrome, in vivo. This study was undertaken to evaluate the potential significance of strength of biofilm formation by clinical bovine mastitis-associated S. aureus in mammary tissue damage by using a mouse mastitis model. Methods Two S. aureus strains of the same capsular phenotype with different biofilm forming strengths were used to non-invasively infect mammary glands of lactating mice. Biofilm forming potential of these strains were determined by tissue culture plate method, ica typing and virulence gene profile per detection by PCR. Delivery of the infectious dose of S. aureus was directly through the teat lactiferous duct without invasive scraping of the teat surface. Both bacteriological and histological methods were used for analysis of mammary gland pathology of mice post-infection. Results Histopathological analysis of the infected mammary glands revealed that mice inoculated with the strong biofilm forming S. aureus strain produced marked acute mastitic lesions, showing profuse infiltration predominantly with neutrophils, with evidence of necrosis in the affected mammary glands. In contrast, the damage was significantly less severe in mammary glands of mice infected with the weak biofilm-forming S. aureus strain. Although both IL-1β and TNF-α inflammatory biomarkers were produced in infected mice, level of TNF-α produced was significantly higher (p<0.05) in mice inoculated with strong biofilm forming S. aureus than the weak biofilm forming strain. Conclusion This finding suggests an important role of TNF-α in mammary gland pathology post-infection with strong biofilm-forming S. aureus in the acute mouse mastitis model, and offers an opportunity for the development of novel strategies for reduction of

  17. [Genetic identification and study of the ability of lactobacilli isolated from the oral cavity of healthy individuals to form biofilms].

    Science.gov (United States)

    Chervinets, Iu V; Botina, S G; Glazova, A A; Koroban, N V; Chervinets, V M; Samoukina, A M; Gavrilova, O A; Lebedev, D V; Mironov, A Iu

    2011-02-01

    The highly antagonistic lactobacillus strains isolated from the oral cavity of human individuals were genetically passported as L. fermentum 39, L. rhamnosus 50, and L. rhamnosus 24, by applying the RAPD-PCR technique with two types of primers (M13, MSP). These lactobacillus strains showed high degrees of autoaggregation, surface hydrophobicity, coaggregation, and adhesion. These characteristics determine the obvious capacity of lactobacilli to form biofilms, which may be used to design new probiotic agents.

  18. Agaricus Blazei Hot Water Extract Shows Anti Quorum Sensing Activity in the Nosocomial Human Pathogen Pseudomonas Aeruginosa

    Directory of Open Access Journals (Sweden)

    Marina Soković

    2014-04-01

    Full Text Available The edible mushroom Agaricus blazei Murill is known to induce protective immunomodulatory action against a variety of infectious diseases. In the present study we report potential anti-quorum sensing properties of A. blazei hot water extract. Quorum sensing (QS plays an important role in virulence, biofilm formation and survival of many pathogenic bacteria, including the Gram negative Pseudomonas aeruginosa, and is considered as a novel and promising target for anti-infectious agents. In this study, the effect of the sub-MICs of Agaricus blazei water extract on QS regulated virulence factors and biofilm formation was evaluated against P. aeruginosa PAO1. Sub-MIC concentrations of the extract which did not kill P. aeruginosa nor inhibited its growth, demonstrated a statistically significant reduction of virulence factors of P. aeruginosa, such as pyocyanin production, twitching and swimming motility. The biofilm forming capability of P. aeruginosa was also reduced in a concentration-dependent manner at sub-MIC values. Water extract of A. blazei is a promising source of antiquorum sensing and antibacterial compounds.

  19. Type III Secretion System Translocon Component EseB Forms Filaments on and Mediates Autoaggregation of and Biofilm Formation by Edwardsiella tarda.

    Science.gov (United States)

    Gao, Zhi Peng; Nie, Pin; Lu, Jin Fang; Liu, Lu Yi; Xiao, Tiao Yi; Liu, Wei; Liu, Jia Shou; Xie, Hai Xia

    2015-09-01

    The type III secretion system (T3SS) of Edwardsiella tarda plays an important role in infection by translocating effector proteins into host cells. EseB, a component required for effector translocation, is reported to mediate autoaggregation of E. tarda. In this study, we demonstrate that EseB forms filamentous appendages on the surface of E. tarda and is required for biofilm formation by E. tarda in Dulbecco's modified Eagle's medium (DMEM). Biofilm formation by E. tarda in DMEM does not require FlhB, an essential component for assembling flagella. Dynamic analysis of EseB filament formation, autoaggregation, and biofilm formation shows that the formation of EseB filaments occurs prior to autoaggregation and biofilm formation. The addition of an EseB antibody to E. tarda cultures before bacterial autoaggregation prevents autoaggregation and biofilm formation in a dose-dependent manner, whereas the addition of the EseB antibody to E. tarda cultures in which biofilm is already formed does not destroy the biofilm. Therefore, EseB filament-mediated bacterial cell-cell interaction is a prerequisite for autoaggregation and biofilm formation.

  20. Destruction of oral biofilms formed in situ on machined titanium (Ti) surfaces by cold atmospheric plasma.

    Science.gov (United States)

    Idlibi, Ahmad Nour; Al-Marrawi, Fuad; Hannig, Matthias; Lehmann, Antje; Rueppell, Andre; Schindler, Axel; Jentsch, Holger; Rupf, Stefan

    2013-01-01

    The decontamination of implant surfaces represents the basic procedure in the management of peri-implant diseases, but it is still a challenge. The study aimed to evaluate the degradation of oral biofilms grown in situ on machined titanium (Ti) discs by cold atmospheric plasma (CAP). ~200 Ti discs were exposed to the oral cavities of five healthy human volunteers for 72 h. The resulting biofilms were divided randomly between the following treatments: CAP (which varied in mean power, treatment duration, and/or the gas mixture), and untreated and treated controls (diode laser, air-abrasion, chlorhexidine). The viability, quantity, and morphology of the biofilms were determined by live/dead staining, inoculation onto blood agar, quantification of the total protein content, and scanning electron microscopy. Exposure to CAP significantly reduced the viability and quantity of biofilms compared with the positive control treatments. The efficacy of treatment with CAP correlated with the treatment duration and plasma power. No single method achieved complete biofilm removal; however, CAP may provide an effective support to established decontamination techniques for treatment of peri-implant diseases.

  1. A combination of cis-2-decenoic acid and antibiotics eradicates pre-established catheter-associated biofilms.

    Science.gov (United States)

    Rahmani-Badi, Azadeh; Sepehr, Shayesteh; Mohammadi, Parisa; Soudi, Mohammad Reza; Babaie-Naiej, Hamta; Fallahi, Hossein

    2014-11-01

    The catheterized urinary tract provides ideal conditions for the development of biofilm populations. Catheter-associated urinary tract infections (CAUTIs) are recalcitrant to existing antimicrobial treatments; therefore, established biofilms are not eradicated completely after treatment and surviving biofilm cells will carry on the infection. Cis-2-decenoic acid (CDA), an unsaturated fatty acid, is capable of inhibiting biofilm formation by Pseudomonas aeruginosa and of inducing the dispersion of established biofilms by multiple types of micro-organisms. Here, the ability of CDA to induce dispersal in pre-established single- and dual-species biofilms formed by Escherichia coli and Klebsiella pneumoniae was measured by using both semi-batch and continuous cultures bioassays. Removal of the biofilms by combined CDA and antibiotics (ciprofloxacin or ampicillin) was evaluated using microtitre plate assays (crystal violet staining). The c.f.u. counts were determined to assess the potential of combined CDA treatments to kill and eradicate pre-established biofilms formed on catheters. The effects of combined CDA treatments on biofilm surface area and bacteria viability were evaluated using fluorescence microscopy, digital image analysis and live/dead staining. To investigate the ability of CDA to prevent biofilm formation, single and mixed cultures were grown in the presence and absence of CDA. Treatment of pre-established biofilms with only 310 nM CDA resulted in at least threefold increase in the number of planktonic cells in all cultures tested. Whilst none of the antibiotics alone exerted a significant effect on c.f.u. counts and percentage of surface area covered by the biofilms, combined CDA treatments led to at least a 78% reduction in biofilm biomass in all cases. Moreover, most of the biofilm cells remaining on the surface were killed by antibiotics. The addition of 310 nM CDA significantly prevented biofilm formation by the tested micro-organisms, even within

  2. 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.

  3. 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

  4. Efficacy of Various Chemical Disinfectants on Biofilms Formed in Spacecraft Potable Water System Component

    Science.gov (United States)

    Wong, Willy; Garcia, Veronica; Castro, Victoria; Ott, Mark; Duane

    2009-01-01

    As the provision of potable water is critical for successful habitation of the International Space Station (ISS), life support systems were installed in December 2008 to recycle both humidity from the atmosphere and urine to conserve available water in the vehicle. Pre-consumption testing from the dispensing needle at the Potable Water Dispenser (PWD) indicated that bacterial concentrations exceeded the current ISS specifications of 50 colony forming units (CFU) per ml. Subsequent investigations revealed that a corrugated stainless steel flex hose upstream of the dispensing needle in the PWD was filled with non-sterile water and left at room temperature for over one month before launch. To simulate biofilm formation that was suspected in the flight system, sterile flex hoses were seeded with a consortium of bacterial isolates previously recovered from other ISS water systems, which included Ralstonia pickettii, Burkholderia multivorans, Caulobacter vibrioides., and Cupriavidus pauculus. After 5 days of incubation, these hoses were challenged with various chemical disinfectants including hydrogen peroxide, colloidal silver, and buffered pH solutions to determine the ability of the disinfectants to decrease and maintain bacterial concentrations below ISS specifications. Disinfection efficacy over time was measured by collecting daily heterotrophic plate counts following exposure to the disinfectants. A single flush with either 6% hydrogen peroxide solution or a mixture of 3% hydrogen peroxide and 400 ppb colloidal silver effectively reduced the bacterial concentrations to less than 1 CFU/ml for a period of up to 2 months. Testing results indicated that hydrogen peroxide and mixtures of hydrogen peroxide and colloidal silver have tremendous potential as alternative disinfectants for ISS water systems.

  5. Synthesis of 2(5H)-pyrrinones and the inhibitory effects on the Biofilms of P.aeruginosa in vitro%2(5H)-吡咯酮衍生物的合成及对铜绿假单胞菌生物膜的体外作用

    Institute of Scientific and Technical Information of China (English)

    郭嘉亮; 刘兆祥; 巢伟; 孙平华

    2013-01-01

    以乙酰乙酸乙酯为原料,设计合成2(5H)-吡咯酮及其衍生物,所得化合物经1H NMR、13C NMR、MS和元素分析等确证.通过扫描电镜观察2(5H)-吡咯酮对黏液型铜绿假单胞菌P.aeruginosa(PA)体外生物被膜(biofilms,BF)的影响,可见BF被破坏,基质样物变稀疏,细菌群聚大为降低.活性实验结果表明,2(5H)-吡咯酮类化合物具有一定的抑制细菌群体感应的能力.%A series of pyrroles and 2 (5H) -pyrrinones were successfully synthesized from ethyl aceto-acetate, and characterized by 'H NMR, 13C NMR, MS spectra and elemental analysis. Scanning electron microscope (SEM) was used to evaluate the effects of the 2 (5H) -pyrrolones on the quorum sensing (QS) system and the bacterial biofilm (BF) formation biofilm formation. The biofilms of P. aeruginosa were inhibited, the base became sparseness, and the swarming was decreased. The results of biological activity tests indicated that 2 (5H) -pyrrinones showed potent ability to inhibit the formation of bacterial biofilms.

  6. Investigations of Biofilm-Forming Bacterial Cells by Atomic Force Microscopy Prior to and Following Treatment from Gas Discharge Plasmas

    Science.gov (United States)

    Vandervoort, K. G.; Joaquin, J. C.; Kwan, C.; Bray, J. D.; Torrico, R.; Abramzon, N.; Brelles-Marino, G.

    2007-03-01

    We present investigations of biofilm-forming bacteria before and after treatment from gas discharge plasmas. Gas discharge plasmas represent a way to inactivate bacteria under conditions where conventional disinfection methods are often ineffective. These conditions involve bacteria in biofilm communities, where cooperative interactions between cells make organisms less susceptible to standard killing methods. Rhizobium gallicum and Chromobacterium violaceum were imaged before and after plasma treatment using an atomic force microscope (AFM). In addition, cell wall elasticity was studied by measuring force distance curves as the AFM tip was pressed into the cell surface. Results for cell surface morphology and micromechanical properties for plasma treatments lasting from 5 to 60 minutes were obtained and will be presented.

  7. Analysis of hyper-baric biofilms on engineering surfaces formed in the Deep Sea

    Science.gov (United States)

    Meier, A.; Tsaloglou, N. M.; Connelly, D.; Keevil, B.; Mowlem, M.

    2012-04-01

    Long-term monitoring of the environment is essential to our understanding of global processes, such as global warming, and their impact. As biofilm formation occurs after only short deployment periods in the marine environment, it is a major problem in long-term operation of environmental sensors. This makes the development of anti-fouling strategies for in situ sensors critical to their function. The effects on sensors can range from measurement drift, which can be compensated, to blockage of channels and material degradation, rendering them inoperative. In general, the longer the deployment period the more severe the effects of the biofouling become. Until now, biofilm research has focused mainly on the eutrophic and euphotic zones of the oceans. Hyper-baric biofilms are poorly understood due to difficulties in experimental setup and the assumption that biofouling in these oligotrophic regions could be regarded as insignificant. Our study shows significant biofilm formation occurs in the deep sea. We deployed a variety of materials, typically used in engineering structures, on a 4500 metre deep mooring during a cruise to the Cayman Trough, for 10 days. The materials were clear plain glass, poly-methyl methacrylate (PMMA), Delrin™, and copper, a known antifouling agent. The biofilms were studied by fluorescence microscopy and molecular analysis. For microscopy the nucleic acid stain, SYTO©9, was used and surface coverage was quantified by using a custom MATLAB™ program. Further molecular analyses, including UV Vis spectrometric quantification of DNA, nucleic acid amplification using Polymerase Chain Reaction (PCR), and Denaturing Gradient Gel Electrophoresis (DGGE), were utilised for the analysis of the microbial community composition of these biofilms. Six 16S/18S universal primer sets representative for the three kingdoms, Archea, Bacteria, and Eukarya were used for the PCR and DGGE. Preliminary results from fluorescence microscopy showed that the biofilm

  8. The in vivo biofilm

    DEFF Research Database (Denmark)

    Bjarnsholt, Thomas; Alhede, Maria; Alhede, Morten

    2013-01-01

    Bacteria can grow and proliferate either as single, independent cells or organized in aggregates commonly referred to as biofilms. When bacteria succeed in forming a biofilm within the human host, the infection often becomes very resistant to treatment and can develop into a chronic state. Biofilms...... have been studied for decades using various in vitro models, but it remains debatable whether such in vitro biofilms actually resemble in vivo biofilms in chronic infections. In vivo biofilms share several structural characteristics that differ from most in vitro biofilms. Additionally, the in vivo...... experimental time span and presence of host defenses differ from chronic infections and the chemical microenvironment of both in vivo and in vitro biofilms is seldom taken into account. In this review, we discuss why the current in vitro models of biofilms might be limited for describing infectious biofilms...

  9. Periplasmic Domains of Pseudomonas aeruginosa PilN and PilO Form a Stable Heterodimeric Complex

    Energy Technology Data Exchange (ETDEWEB)

    Sampaleanu, L.M.; Bonanno, J.B.; Ayers, M.; Koo, J.; Tammam, S.; Burley, S.K.; Almo, S.C.; Burrows, L.L.; Howell, P.L.; (HSC); (Einstein); (McMaster U.); (Lilly)

    2010-01-12

    Type IV pili (T4P) are bacterial virulence factors responsible for attachment to surfaces and for twitching motility, a motion that involves a succession of pilus extension and retraction cycles. In the opportunistic pathogen Pseudomonas aeruginosa, the PilM/N/O/P proteins are essential for T4P biogenesis, and genetic and biochemical analyses strongly suggest that they form an inner-membrane complex. Here, we show through co-expression and biochemical analysis that the periplasmic domains of PilN and PilO interact to form a heterodimer. The structure of residues 69-201 of the periplasmic domain of PilO was determined to 2.2 {angstrom} resolution and reveals the presence of a homodimer in the asymmetric unit. Each monomer consists of two N-terminal coiled coils and a C-terminal ferredoxin-like domain. This structure was used to generate homology models of PilN and the PilN/O heterodimer. Our structural analysis suggests that in vivo PilN/O heterodimerization would require changes in the orientation of the first N-terminal coiled coil, which leads to two alternative models for the role of the transmembrane domains in the PilN/O interaction. Analysis of PilN/O orthologues in the type II secretion system EpsL/M revealed significant similarities in their secondary structures and the tertiary structures of PilO and EpsM, although the way these proteins interact to form inner-membrane complexes appears to be different in T4P and type II secretion. Our analysis suggests that PilN interacts directly, via its N-terminal tail, with the cytoplasmic protein PilM. This work shows a direct interaction between the periplasmic domains of PilN and PilO, with PilO playing a key role in the proper folding of PilN. Our results suggest that PilN/O heterodimers form the foundation of the inner-membrane PilM/N/O/P complex, which is critical for the assembly of a functional T4P complex.

  10. 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

    to the antimicrobial peptide colistin. On the contrary, biofilm cells exhibiting low metabolic activity were killed by colistin. We demonstrate that the subpopulation of metabolically active cells is able to adapt to colistin by inducing a specific adaptation mechanism mediated by the pmr operon, as well...... as an unspecific adaptation mechanism mediated by the mexAB-oprM genes. Mutants defective in either pmr-mediated lipopolysaccharide modification or in mexAB-oprM-mediated antimicrobial efflux were not able to develop a tolerant subpopulation in biofilms. In contrast to the observed pattern of colistin...

  11. Study on resistance of pseudomonas aeruginosa biofilm to different disinfectants%铜绿假单胞菌生物膜对不同消毒因子的抗力研究

    Institute of Scientific and Technical Information of China (English)

    陆龙喜; 陆烨; 林军明; 许激; 朱一凡

    2011-01-01

    OBJECTIVE To study the resistance of Psedomonas aeruginosa biofilm to different disinfectants.METHODS The resistance of artificially cultured biofilm to different disinfectants were tested by improved Brown plate method, continuous perfusion culture, quantitative germicidal test and scanning electron microscopy.RESULTS After cultured for 5 ~ 7 days, PAE biofilm adhered to teflon tube-wall stably and the bacterial concentration was up to 105~106 CFU/cm2 approximately. This biofilm could not be washed by water easily. The endure time of PAE biofilm to 0. 55% ortho-phthalaldehyde was 3min, and its killing time was 5min, The endure time of PAE biofilm to 200mg/L available chlorine was 5min, and its killing time was 7min, The endure time of PAE biofilm to 500mg/L available iodine was 10min, and its killing time was 20min, There were quite amount PAE alive after 30min contact with 1000mg/L PHMB. CONCLUSION The PAE lm to 0.55% orthophthalaldehyde, 200mg/L available chlorine, 500mg/L available iodine, 1000mg/L PHMB all show different degree resistance, It should be attached importance in hospital disinfection.%目的 研究铜绿假单胞菌生物膜对不同消毒因子的抗力.方法采用Brown平板改良法、持续灌流培养技术、载体定量杀菌试验以及扫描电镜技术进行人工培养生物膜对不同消毒因子的抗力研究.结果体外连续培养5~7 d的聚四氟乙烯管内壁的铜绿假单胞菌生物膜形成稳定,不易冲洗脱落,膜中细菌浓度达105~106 CFU/cm2,生物膜对0.55%邻苯二甲醛(OPA)耐受时间为3 min,作用5 min可达到完全杀灭;对200 mg/L有效氯耐受时间为5 min,作用7 min可达到完全杀灭;500 mg/L有效碘耐受时间为10 min,作用20 min可达到完全杀灭;对1000 mg/L聚六亚甲基双胍盐酸盐(PHMB)作用30 min仍然有大量细菌存活.结论体外培养生物膜对0.55%邻苯二甲醛、200 mg/L有效氯、500 mg/L有效碘、1000 mg/L PHMB均有不同程度的抵抗力,

  12. Influence of medium composition on the characteristics of a denitrifying biofilm formed by Alcaligenes denitrificans in a fluidised bed reactor

    OpenAIRE

    Alves, C. F.; Melo, L. F.; Vieira, M. J.

    2002-01-01

    The influence of the ratio carbon/nitrogen and phosphorus concentration on the performance of a biofilm fluidised bed reactor used for denitrification and on the properties of the biofilm was studied. Although the removal efficiencies of C and N reached steady-state values, the thickness of the biofilm steadily increased. The dry density of the biofilm did not seem to be dependent on the loading conditions, although a denser biofilm was obtained when there was no nutrient limitation ...

  13. A novel strategy for acetonitrile wastewater treatment by using a recombinant bacterium with biofilm-forming and nitrile-degrading capability.

    Science.gov (United States)

    Li, Chunyan; Yue, Zhenlei; Feng, Fengzhao; Xi, Chuanwu; Zang, Hailian; An, Xuejiao; Liu, Keran

    2016-10-01

    There is a great need for efficient acetonitrile removal technology in wastewater treatment to reduce the discharge of this pollutant in untreated wastewater. In this study, a nitrilase gene (nit) isolated from a nitrile-degrading bacterium (Rhodococcus rhodochrous BX2) was cloned and transformed into a biofilm-forming bacterium (Bacillus subtilis N4) that expressed the recombinant protein upon isopropylthio-β-galactoside (IPTG) induction. The recombinant bacterium (B. subtilis N4-pHT01-nit) formed strong biofilms and had nitrile-degrading capability. Further testing demonstrated that biofilms formed by B. subtilis N4-pHT01-nit were highly resistant to loading shock from acetonitrile and almost completely degraded the initial concentration of acetonitrile (800 mg L(-1)) within 24 h in a moving bed biofilm reactor (MBBR) after operation for 35 d. The bacterial composition of the biofilm, identified by high-throughput sequencing, in a reactor in which the B. subtilis N4-pHT01-nit bacterium was introduced indicated that the engineered bacterium was successfully immobilized in the reactor and became dominant genus. This work demonstrates that an engineered bacterium with nitrile-degrading and biofilm-forming capacity can improve the degradation of contaminants in wastewater. This approach offers a novel strategy for enhancing the biological oxidation of toxic pollutants in wastewater.

  14. 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.

  15. Biocidal effect of cathodic protection on bacterial viability in biofilm attached to carbon steel.

    Science.gov (United States)

    Miyanaga, Kazuhiko; Terashi, Ryosuke; Kawai, Hirofumi; Unno, Hajime; Tanji, Yasunori

    2007-07-01

    Biofilm formed on carbon steel by various species of bacterial cells causes serious problems such as corrosion of steel, choking of flow in the pipe, deterioration of the heat-transfer efficiency, and so on. Cathodic protection is known to be a reliable method for protecting carbon steel from corrosion. However, the initial attachment of bacteria to the surface and the effects of cathodic protection on bacterial viability in the biofilm have not been clarified. In this study, cathodic protection was applied to an artificial biofilm containing Pseudomonas aeruginosa (PAO1), a biofilm constituent, on carbon steel. The aims of this study were to evaluate the inhibition effect of cathodic protection on biofilm formation and to reveal the inhibition mechanisms. The viability of PAO1 in artificial biofilm of 5 mm thickness on cathodically protected steel decreased to 1% of the initial cell concentration. Analysis of pH distribution in the artificial biofilm by pH microelectrode revealed that pH in proximity to carbon steel increased to approximately 11 after cathodic protection for 5 h. Moreover, 99% of region in the artificial biofilm was under the pH conditions of over nine. A simulation of pH profile was shown to correspond to experimental values. These results indicate cells in the artificial biofilm were killed or damaged by cathodic protection due to pH increase.

  16. A novel Pseudomonas aeruginosa bacteriophage, Ab31, a chimera formed from temperate phage PAJU2 and P. putida lytic phage AF: characteristics and mechanism of bacterial resistance.

    Directory of Open Access Journals (Sweden)

    Libera Latino

    Full Text Available A novel temperate bacteriophage of Pseudomonas aeruginosa, phage vB_PaeP_Tr60_Ab31 (alias Ab31 is described. Its genome is composed of structural genes related to those of lytic P. putida phage AF, and regulatory genes similar to those of temperate phage PAJU2. The virion structure resembles that of phage AF and other lytic Podoviridae (S. enterica Epsilon 15 and E. coli phiv10 with similar tail spikes. Ab31 was able to infect P. aeruginosa strain PA14 and two genetically related strains called Tr60 and Tr162, out of 35 diverse strains from cystic fibrosis patients. Analysis of resistant host variants revealed different phenotypes, including induction of pigment and alginate overproduction. Whole genome sequencing of resistant variants highlighted the existence of a large deletion of 234 kbp in two strains, encompassing a cluster of genes required for the production of CupA fimbriae. Stable lysogens formed by Ab31 in strain Tr60, permitted the identification of the insertion site. During colonization of the lung in cystic fibrosis patients, P. aeruginosa adapts by modifying its genome. We suggest that bacteriophages such as Ab31 may play an important role in this adaptation by selecting for bacterial characteristics that favor persistence of bacteria in the lung.

  17. Synergistic algicidal effect and mechanism of two diketopiperazines produced by Chryseobacterium sp. strain GLY-1106 on the harmful bloom-forming Microcystis aeruginosa.

    Science.gov (United States)

    Guo, Xingliang; Liu, Xianglong; Pan, Jianliang; Yang, Hong

    2015-10-01

    A potent algicidal bacterium isolated from Lake Taihu, Chryseobacterium sp. strain GLY-1106, produces two algicidal compounds: 1106-A (cyclo(4-OH-Pro-Leu)) and 1106-B (cyclo(Pro-Leu)). Both diketopiperazines showed strong algicidal activities against Microcystis aeruginosa, the dominant bloom-forming cyanobacterium in Lake Taihu. Interestingly, these two algicidal compounds functioned synergistically. Compared with individual treatment, combined treatment with cyclo(4-OH-Pro-Leu) and cyclo(Pro-Leu) significantly enhanced algicidal activity, accelerated the increase in intracellular reactive oxygen species (ROS) levels in M. aeruginosa, and further decreased the activities of antioxidases, effective quantum yield and maximal electron transport rate of M. aeruginosa. The results also showed that the algicidal characteristics of cyclo(4-OH-Pro-Leu) are distinct from those of cyclo(Pro-Leu). Cyclo(4-OH-Pro-Leu) mainly interrupted the flux of electron transport in the cyanobacterial photosynthetic system, whereas cyclo(Pro-Leu) mainly inhibited the activity of cyanobacterial intracellular antioxidases. A possible algicidal mechanism for the synergism between cyclo(4-OH-Pro-Leu) and cyclo(Pro-Leu) is proposed, which is in accordance with their distinct algicidal characteristics in individual and combined treatment. These findings suggest that synergism between algicidal compounds might be used as an effective strategy for the future control of Microcystis blooms.

  18. Community Composition of Bacterial Biofilms Formed on Simple Soil Based Bioelectrochemical Cell Anodes and Cathodes

    Science.gov (United States)

    2012-04-01

    3 Table 2. Relative molar percentages and absolute abundances of prokaryotic and eukaryotic fatty acid (FA...density for the three soils varied from 5 to 8 × 108 cells per gram of soil. 3.1 Anode biofilms Although eukaryotic biomarkers were detected on the...percentages and absolute abundances of prokaryotic and eukaryotic fatty acid (FA) biomarkers detected in the soils, top middle and bottom fractions, on the

  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. Pseudomonas aeruginosa dose response and bathing water infection.

    Science.gov (United States)

    Roser, D J; van den Akker, B; Boase, S; Haas, C N; Ashbolt, N J; Rice, S A

    2014-03-01

    Pseudomonas aeruginosa is the opportunistic pathogen mostly implicated in folliculitis and acute otitis externa in pools and hot tubs. Nevertheless, infection risks remain poorly quantified. This paper reviews disease aetiologies and bacterial skin colonization science to advance dose-response theory development. Three model forms are identified for predicting disease likelihood from pathogen density. Two are based on Furumoto & Mickey's exponential 'single-hit' model and predict infection likelihood and severity (lesions/m2), respectively. 'Third-generation', mechanistic, dose-response algorithm development is additionally scoped. The proposed formulation integrates dispersion, epidermal interaction, and follicle invasion. The review also details uncertainties needing consideration which pertain to water quality, outbreaks, exposure time, infection sites, biofilms, cerumen, environmental factors (e.g. skin saturation, hydrodynamics), and whether P. aeruginosa is endogenous or exogenous. The review's findings are used to propose a conceptual infection model and identify research priorities including pool dose-response modelling, epidermis ecology and infection likelihood-based hygiene management.

  1. Selective reactivity of monochloramine with extracellular matrix components affects the disinfection of biofilm and detached clusters.

    Science.gov (United States)

    Xue, Zheng; Lee, Woo Hyoung; Coburn, Kimberly M; Seo, Youngwoo

    2014-04-01

    The efficiency of monochloramine disinfection was dependent on the quantity and composition of extracellular polymeric substances (EPS) in biofilms, as monochloramine has a selective reactivity with proteins over polysaccharides. Biofilms with protein-based (Pseudomonas putida) and polysaccharide based EPS (Pseudomonas aeruginosa), as well as biofilms with varied amount of polysaccharide EPS (wild-type and mutant P. aeruginosa), were compared. The different reactivity of EPS components with monochloramine influenced disinfectant penetration, biofilm inactivation, as well as the viability of detached clusters. Monochloramine transport profiling measured by a chloramine-sensitive microelectrode revealed a broader diffusion boundary layer between bulk and biofilm surface in the P. putida biofilm compared to those of P. aeruginosa biofilms. The reaction with proteins in P. putida EPS multiplied both the time and the monochloramine mass required to achieve a full biofilm penetration. Cell viability in biofilms was also spatially influenced by monochloramine diffusion and reaction within biofilms, showing a lower survival in the surface section and a higher persistence in the middle section of the P. putida biofilm compared to the P. aeruginosa biofilms. While polysaccharide EPS promoted biofilm cell viability by obstructing monochloramine reactive sites on bacterial cells, protein EPS hindered monochloramine penetration by reacting with monochloramine and reduced its concentration within biofilms. Furthermore, the persistence of bacterial cells detached from biofilm (over 70% for P. putida and ∼40% for polysaccharide producing P. aeruginosa) suggested that currently recommended monochloramine residual levels may underestimate the risk of water quality deterioration caused by biofilm detachment.

  2. The induced expression of AmpC gene in pseudomonas aeruginosa biofilms%生物膜铜绿假单胞菌AmpC基因诱导表达的研究

    Institute of Scientific and Technical Information of China (English)

    赵京明; 成炜; 蒋捍东

    2009-01-01

    目的 研究铜绿假单胞菌的产酶基因AmpC在浮游和生物膜状态下的表达差异.方法 改良的平板法建立铜绿假单胞菌生物膜模型,抗生素诱导浮游菌和生物膜菌AmpC基因表达,实时荧光定量聚合酶链反应测定PAOI的AmpC基因表达水平.结果 抗生素诱导前,PAO1浮游菌和生物膜菌的AmpC基因表达均较低,诱导后AmpC基因表达均明显上调.运用产最大AmpC酶活性浓度的抗生素诱导后,亚胺培南和头孢他啶诱导的生物膜菌的AmpC基因表达量高于其诱导的浮游菌.在浮游和生物膜状态下,亚胺培南诱导PAO1的AmpC基因表达量均高于头孢他啶.结论 生物膜PA较浮游菌更易被诱导产生AmpC酶,亚胺培南的诱导能力高于头孢他啶.%Objective To study the expression of AmpC gene in Pseudomonas aeruginosa induced by antibiotics at plank tonic and biofilms phases. Methods An in vitro model of PAO1 biofilms was established with modified flat-board method. Being induced by antibiotics at plank tonic and biofilms phases, the expression of AmpC gene in PAO1 was quantified by real-time quantitative PCR. Results Without the effect of antibiotics, the expres-sion of AmpC gene in PAO1 was low at both plank tonic and biofiims phases. Being induced by imipenem and eeftazidime that can induce the maximal AmpC β-lactamase activities, the expression of AmpC gene in biofilms PAO1 was higher than that in plank tonic PAO1. During both plank tonic and biofilms phases, the expression of AmpC gene in PAO1 induced by imipenem was higher than that induced by ceftazidime. Conclusion Induced by antibiotics,biofilms PAO1 showes stronger ability for the expression of AmpC gene than plank tonic PAO1. The ex-pression of AmpC gene induced by imipenem is higher than that induced by ceftazidime.

  3. A novel model of chronic wounds: importance of redox imbalance and biofilm-forming bacteria for establishment of chronicity.

    Directory of Open Access Journals (Sweden)

    Sandeep Dhall

    Full Text Available Chronic wounds have a large impact on health, affecting ∼6.5 M people and costing ∼$25B/year in the US alone. We previously discovered that a genetically modified mouse model displays impaired healing similar to problematic wounds in humans and that sometimes the wounds become chronic. Here we show how and why these impaired wounds become chronic, describe a way whereby we can drive impaired wounds to chronicity at will and propose that the same processes are involved in chronic wound development in humans. We hypothesize that exacerbated levels of oxidative stress are critical for initiation of chronicity. We show that, very early after injury, wounds with impaired healing contain elevated levels of reactive oxygen and nitrogen species and, much like in humans, these levels increase with age. Moreover, the activity of anti-oxidant enzymes is not elevated, leading to buildup of oxidative stress in the wound environment. To induce chronicity, we exacerbated the redox imbalance by further inhibiting the antioxidant enzymes and by infecting the wounds with biofilm-forming bacteria isolated from the chronic wounds that developed naturally in these mice. These wounds do not re-epithelialize, the granulation tissue lacks vascularization and interstitial collagen fibers, they contain an antibiotic-resistant mixed bioflora with biofilm-forming capacity, and they stay open for several weeks. These findings are highly significant because they show for the first time that chronic wounds can be generated in an animal model effectively and consistently. The availability of such a model will significantly propel the field forward because it can be used to develop strategies to regain redox balance that may result in inhibition of biofilm formation and result in restoration of healthy wound tissue. Furthermore, the model can lead to the understanding of other fundamental mechanisms of chronic wound development that can potentially lead to novel therapies.

  4. Treatment of Pseudomonas aeruginosa Biofilm Infected Wounds with Clinical Wound Care Strategies: A Quantitative Study Using an In Vivo Rabbit Ear Model

    Science.gov (United States)

    2012-02-01

    Microbiology Branch, U.S. Army Dental and Trauma Research Detach- ment, Institute of Surgical Research. Received for publication April 22, 2011...polysaccharide matrix, biofilm is the predominant state of bacteria23 found throughout the body (e.g., gastrointestinal tract, dental enamel) and in association...and approximately 0.1 cc of Silvadene Cream 1% (Monarch Pharmaceuticals, Bristol, Tenn.) was applied topically to any wounds in a treatment group

  5. Chemically Specific Cellular Imaging of Biofilm Formation

    Energy Technology Data Exchange (ETDEWEB)

    Herberg, J L; Schaldach, C; Horn, J; Gjersing, E; Maxwell, R

    2006-02-09

    This document and the accompanying manuscripts summarize the technical accomplishments for our one-year LDRD-ER effort. Biofilm forming microbes have existed on this planet for billions of years and make up 60% of the biological mass on earth. Such microbes exhibit unique biochemical pathways during biofilm formation and play important roles in human health and the environment. Microbial biofilms have been directly implicated in, for example, product contamination, energy losses, and medical infection that cost the loss of human lives and billions of dollars. In no small part due to the lack of detailed understanding, biofilms unfortunately are resistant to control, inhibition, and destruction, either through treatment with antimicrobials or immunological defense mechanisms of the body. Current biofilm research has concentrated on the study of biofilms in the bulk. This is primarily due to the lack of analytical and physical tools to study biofilms non-destructively, in three dimensions, and on the micron or sub-micron scale. This has hindered the development of a clear understanding of either the early stage mechanisms of biofilm growth or the interactions of biofilms with their environment. Enzymatic studies have deduced a biochemical reaction that results in the oxidation of reduced sulfur species with the concomitant reduction of nitrate, a common groundwater pollutant, to dinitrogen gas by the bacterium, Thiobacillus denitrificans (TD). Because of its unique involvement in biologically relevant environmental pathways, TD is scheduled for genome sequencing in the near future by the DOE's Joint Genome Institute and is of interest to DOE's Genomes to Life Program. As our ecosystem is exposed to more and more nitrate contamination large scale livestock and agricultural practices, a further understanding of biofilm formation by organisms that could alleviate these problems is necessary in order to protect out biosphere. However, in order to study this

  6. 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.

  7. Prevalence and biofilm-forming ability of Listeria monocytogenes in New Zealand mussel (Perna canaliculus) processing plants.

    Science.gov (United States)

    Cruz, Cristina D; Fletcher, Graham C

    2011-10-01

    Greenshell™ mussels are New Zealand's largest seafood export species. Some export markets require compliance with 'zero' tolerance legislation for Listeria monocytogenes in 25 g of product. Even though individually quick frozen (IQF) mussel products are labeled 'to be cooked', and are not classified as ready-to-eat, some markets still require them to comply with the strict policy. Three mussel processing plants were assessed for the pattern of L. monocytogenes contamination on raw material, environment, food contact surfaces, and in the final product. Cultures (n = 101) obtained from an industrial Listeria monitoring program from August 2007 to June 2009 were characterized by serotyping and pulsed field gel electrophoresis. Using the crystal violet method, isolates were assessed for their ability to form biofilms. This work confirmed the presence of L. monocytogenes in raw and processed product, and the importance of cross-contamination from external and internal environments. Processing plants had L. monocytogenes pulsotypes that were detected more than once over 6 months. No correlation was found between biofilm-forming ability and persistent isolates. Two pulsotypes (including a persistent one), were previously isolated in human cases of listeriosis in New Zealand, but none of the pulsotypes matched those involved in international outbreaks.

  8. Biofilm inhibition activity of compounds isolated from two Eunicea species collected at the Caribbean Sea

    Directory of Open Access Journals (Sweden)

    Yenny Martínez Díaz

    2015-12-01

    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.

  9. Chemical, physical and morphological properties of bacterial biofilms affect survival of encased Campylobacter jejuni F38011 under aerobic stress.

    Science.gov (United States)

    Feng, Jinsong; Lamour, Guillaume; Xue, Rui; Mirvakliki, Mehr Negar; Hatzikiriakos, Savvas G; Xu, Jie; Li, Hongbin; Wang, Shuo; Lu, Xiaonan

    2016-12-05

    Campylobacter jejuni is a microaerophilic pathogen and leading cause of human gastroenteritis. The presence of C. jejuni encased in biofilms found in meat and poultry processing facilities may be the major strategy for its survival and dissemination in aerobic environment. In this study, Staphylococcus aureus, Salmonella enterica, or Pseudomonas aeruginosa was mixed with C. jejuni F38011 as a culture to form dual-species biofilms. After 4days' exposure to aerobic stress, no viable C. jejuni cells could be detected from mono-species C. jejuni biofilm. In contrast, at least 4.7logCFU/cm(2) of viable C. jejuni cells existed in some dual-species biofilms. To elucidate the mechanism of protection mode, chemical, physical and morphological features of biofilms were characterized. Dual-species biofilms contained a higher level of extracellular polymeric substances with a more diversified chemical composition, especially for polysaccharides and proteins, than mono-species C. jejuni biofilm. Structure of dual-species biofilms was more compact and their surface was >8 times smoother than mono-species C. jejuni biofilm, as indicated by atomic force microscopy. Under desiccation stress, water content of dual-species biofilms decreased slowly and remained at higher levels for a longer time than mono-species C. jejuni biofilm. The surface of all biofilms was hydrophilic, but total surface energy of dual-species biofilms (ranging from 52.5 to 56.2mJ/m(2)) was lower than that of mono-species C. jejuni biofilm, leading to more resistance to wetting by polar liquids. This knowledge can aid in developing intervention strategies to decrease the survival and dispersal of C. jejuni into foods or environment.

  10. The Biofilm Challenge

    DEFF Research Database (Denmark)

    Alhede, Maria; Alhede, Morten

    2014-01-01

    The concept of biofilms has emerged in the clinical setting during the last decade. Infections involving biofilms have been documented in all parts of the human body, and it is currently believed that the presence of biofilm-forming bacteria is equivalent to chronic infection. A quick Pubmed search...

  11. 茶树精油对铜绿假单胞杆菌生物膜的抑制作用%Restraining effects of tea tree oil to the Pseudomonas aeruginosa biofilm

    Institute of Scientific and Technical Information of China (English)

    黄晓敏; 李海妙; 罗宝正; 管文华; 林少芸; 王明明

    2012-01-01

    目的 观察茶树精油对铜绿假单胞杆菌生物膜(BF)的抑制作用.方法 采用二倍稀释法测定茶树精油对铜绿假单胞杆菌的最小抑菌浓度(MIC),以一次性输液管和微孔滤膜作为生物膜吸附载体,用改良后的Brown平板连续培养法制备铜绿假单胞杆菌生物膜体外模型,采用光学高倍显微镜连续观察生物膜的形成,采用扫描电子显微镜进行生物膜结构观察和鉴定,以琼脂平板菌落计数法测定茶树精油对受试菌生物膜的抑制效果.结果 茶树精油对铜绿假单胞杆菌的MIC(孟乙烯4-ol浓度含量)为9.6μg/ml; 0.5倍MIC (4.8 μg/ml)、1倍MIC(9.6 μg/ml)虽能使成熟铜绿假单胞杆菌生物膜菌落数呈梯度性减少,但不能完全清除生物膜细菌,而2倍MIC(19.2 μg/ml)对细菌生物膜作用20 h后可完全清除生物膜细菌,3倍MIC(28.8μg/ml)则可在4h内完全清除生物膜.结论 茶树精油对铜绿假单胞菌生物膜有清除效应,可抑制生物膜形成,但需进一步研究验证.%OBJECTIVE To find out and study the restraining effect of tea tree oil to Pseudomonas aeruginos biofilm. METHODS PVC biomedical material and millipore was used as adsorption carrier of biofilm and double-dilution method was used to detect the MIC of tea tree oil on Pseudomonas aeruginos, the in vitro model of Pseudomonas aeruginos biofilm was built by continuous development method on Brown' s plate, and scanning with electron microcopy (SEM) was used to evaluate the formation of biofilm. The viable bacterial colony was counted on the plate of agar so as to evaluate the restraining effect of tea tree oil for the tested bacterial biofilm. RESULTS The MIC of tea tree oil on Pseudomonas aeruginos (the concentration of Terpinen 4-ol) was 9.6 (μg/ml. The concentration of tea tree oil of half of MIC (4.8 μg/ml), and one of MIC (9.6 μg/ml) can lower the number of the viable bacterial of the formed Pseudomonas aeruginos biofilm. For example, the

  12. Microbiota formed on attached stainless steel coupons correlates with the natural biofilm of the sink surface in domestic kitchens.

    Science.gov (United States)

    Moen, Birgitte; Røssvoll, Elin; Måge, Ingrid; Møretrø, Trond; Langsrud, Solveig

    2016-02-01

    Stainless steel coupons are frequently used in biofilm studies in the laboratory, as this material is commonly used in the food industry. The coupons are attached to different surfaces to create a "natural" biofilm to be studied further in laboratory trials. However, little has been done to investigate how well the microbiota on such coupons represents the surrounding environment. The microbiota on sink wall surfaces and on new stainless steel coupons attached to the sink wall for 3 months in 8 domestic kitchen sinks was investigated by next-generation sequencing (MiSeq) of the 16S rRNA gene derived from DNA and RNA (cDNA), and by plating and identification of colonies. The mean number of colony-forming units was about 10-fold higher for coupons than sink surfaces, and more variation in bacterial counts between kitchens was seen on sink surfaces than coupons. The microbiota in the majority of biofilms was dominated by Moraxellaceae (genus Moraxella/Enhydrobacter) and Micrococcaceae (genus Kocuria). The results demonstrated that the variation in the microbiota was mainly due to differences between kitchens (38.2%), followed by the different nucleic acid template (DNA vs RNA) (10.8%), and that only 5.1% of the variation was a result of differences between coupons and sink surfaces. The microbiota variation between sink surfaces and coupons was smaller for samples based on their RNA than on their DNA. Overall, our results suggest that new stainless steel coupons are suited to model the dominating part of the natural microbiota of the surrounding environment and, furthermore, are suitable for different downstream studies.

  13. Molecular Characterisation and Co-cultivation of Bacterial Biofilm Communities Associated with the Mat-Forming Diatom Didymosphenia geminata.

    Science.gov (United States)

    Brandes, Josephin; Kuhajek, Jeanne M; Goodwin, Eric; Wood, Susanna A

    2016-10-01

    Didymosphenia geminata (Lyngbye) M. Schmidt is a stalked freshwater diatom that is expanding its range globally. In some rivers, D. geminata forms thick and expansive polysaccharide-dominated mats. Like other stalked diatoms, D. geminata cells attach to the substratum with a pad of adhesive extracellular polymeric substance. Research on D. geminata and other diatoms suggests that bacterial biofilm composition may contribute to successful attachment. The aim of this study was to investigate the composition and role of bacterial biofilm communities in D. geminata attachment and survival. Bacterial biofilms were collected at four sites in the main stem of a river (containing D. geminata) and in four tributaries (free of D. geminata). Samples were characterised using automated rRNA intergenic spacer analysis and high-throughput sequencing (HTS). Mat-associated bacteria were isolated and their effect on the early establishment of D. geminata cells assessed using co-culturing experiments. ARISA and HTS data showed differences in bacterial communities between samples with and without D. geminata at two of the four sites. Samples with D. geminata had a higher relative abundance of Sphingobacteria (p < 0.01) and variability in community composition was reduced. Analysis of the 76 bacteria isolated from the mat revealed 12 different strains representing 8 genera. Co-culturing of a Carnobacterium sp. with D. geminata reduced survival (p < 0.001) and attachment (p < 0.001) of D. geminata. Attachment was enhanced by Micrococcus sp. and Pseudomonas sp. (p < 0.001 and p < 0.01, respectively). These data provide evidence that bacteria play a role in the initial attachment and on-going survival of D. geminata, and may partly explain observed distribution patterns.

  14. Application of a biofilm formed by a mixture of yeasts isolated in Vietnam to degrade aromatic hydrocarbon polluted wastewater collected from petroleum storage.

    Science.gov (United States)

    Nhi Cong, Le Thi; Ngoc Mai, Cung Thi; Thanh, Vu Thi; Nga, Le Phi; Minh, Nghiem Ngoc

    2014-01-01

    In this study, three good biofilm-forming yeast strains, including Candida viswanathii TH1, Candida tropicalis TH4 and Trichosporon asahii B1, were isolated from oil-contaminated water and sediment samples collected in coastal zones of Vietnam. These strains were registered in the GenBank database with the accession numbers JX129175, JX129176 and KC139404 for strain TH1, TH4 and B1, respectively. The biofilm formed by a mixture of these organisms degraded 90, 85, 82 and 67% of phenol, naphthalene, anthracene and pyrene, respectively, after a 7-day incubation period using an initial concentration of 600 ppm phenol and 200 ppm of each of the other compounds. In addition, this biofilm completely degraded these aromatic compounds, which were from wastewater collected from petroleum tanks in Do Xa, Hanoi after 14 days of incubation based on gas chromatography mass spectrometry analysis. To the best of our knowledge, reports on polycyclic aromatic hydrocarbon and phenol degradation by biofilm-forming yeasts are limited. The results obtained indicate that the biofilm formed by multiple yeast strains may considerably increase the degradation efficiency of aromatic hydrocarbon compounds, and may lead to a new approach for eliminating petroleum oil-contaminated water in Vietnam.

  15. Lysophosphatidic acid inhibition of the accumulation of Pseudomonas aeruginosa PAO1 alginate, pyoverdin, elastase and LasA

    DEFF Research Database (Denmark)

    Laux, D.C.; Corson, J.M.; Givskov, Michael Christian;

    2002-01-01

    The pathogenesis of Pseudomonas aeruginosa is at least partially attributable to its ability to synthesize and secrete the siderophore pyoverdin and the two zinc metal loproteases elastase and LasA, and its ability to form biofilms in which bacterial cells are embedded in an alginate matrix...... pyoverdin. MPPA also inhibited biofilm formation. The inhibitory effects of MPPA occur independently of rpoS expression and without affecting the accumulation of the autoinducers N-(3-oxododecanoyl) homoserine lactone and N-butyryl-(L)-homoserine lactone, and may be due, at least in part, to the ability...

  16. Utility of in vivo transcription profiling for identifying Pseudomonas aeruginosa genes needed for gastrointestinal colonization and dissemination

    DEFF Research Database (Denmark)

    Koh, Andrew Y; Mikkelsen, Per J; Smith, Roger S;

    2010-01-01

    Microarray analysis of Pseudomonas aeruginosa mRNA transcripts expressed in vivo during animal infection has not been previously used to investigate potential virulence factors needed in this setting. We compared mRNA expression in bacterial cells recovered from the gastrointestinal (GI) tracts...... to test mutants in genes identified as having increased transcription during in vivo colonization. All of the Tn-library mutants in biofilm-associated genes had an attenuated ability to form biofilms in vitro, but there were no significant differences in GI colonization and dissemination between...

  17. Marine Sponge-Derived Streptomyces sp. SBT343 Extract Inhibits Staphylococcal Biofilm Formation

    Science.gov (United States)

    Balasubramanian, Srikkanth; Othman, Eman M.; Kampik, Daniel; Stopper, Helga; Hentschel, Ute; Ziebuhr, Wilma; Oelschlaeger, Tobias A.; Abdelmohsen, Usama R.

    2017-01-01

    Staphylococcus epidermidis and Staphylococcus aureus are opportunistic pathogens that cause nosocomial and chronic biofilm-associated infections. Indwelling medical devices and contact lenses are ideal ecological niches for formation of staphylococcal biofilms. Bacteria within biofilms are known to display reduced susceptibilities to antimicrobials and are protected from the host immune system. High rates of acquired antibiotic resistances in staphylococci and other biofilm-forming bacteria further hamper treatment options and highlight the need for new anti-biofilm strategies. Here, we aimed to evaluate the potential of marine sponge-derived actinomycetes in inhibiting biofilm formation of several strains of S. epidermidis, S. aureus, and Pseudomonas aeruginosa. Results from in vitro biofilm-formation assays, as well as scanning electron and confocal microscopy, revealed that an organic extract derived from the marine sponge-associated bacterium Streptomyces sp. SBT343 significantly inhibited staphylococcal biofilm formation on polystyrene, glass and contact lens surfaces, without affecting bacterial growth. The extract also displayed similar antagonistic effects towards the biofilm formation of other S. epidermidis and S. aureus strains tested but had no inhibitory effects towards Pseudomonas biofilms. Interestingly the extract, at lower effective concentrations, did not exhibit cytotoxic effects on mouse fibroblast, macrophage and human corneal epithelial cell lines. Chemical analysis by High Resolution Fourier Transform Mass Spectrometry (HRMS) of the Streptomyces sp. SBT343 extract proportion revealed its chemical richness and complexity. Preliminary physico-chemical characterization of the extract highlighted the heat-stable and non-proteinaceous nature of the active component(s). The combined data suggest that the Streptomyces sp. SBT343 extract selectively inhibits staphylococcal biofilm formation without interfering with bacterial cell viability. Due to

  18. 铜绿假单胞菌生物膜形成机制及耐药相关性%A study on the mechanism of biofilm formation of Pseudomonas aeruginosa and its relationship with drug resistance

    Institute of Scientific and Technical Information of China (English)

    沈健; 冯晨; 林允照; 郑文华; 顾华; 蒋锦琴; 冯磊

    2015-01-01

    目的:探究铜绿假单胞菌在生物膜状态与浮游状态时对常见抗菌药物的耐药性差异,为临床合理用药提供依据。方法在体外培养基中对铜绿假单胞菌进行孵育处理,并分为浮游液对照培养组(A组)、放置生物膜培养后的培养液组(B组)和生物膜培养硅胶片粘附组(C组)。取浮游培养(培养液)、生物膜培养(培养液)以及生物膜培养(硅胶片粘附)铜绿假单胞菌进行银染色、胞外糖染色并且扫描电镜鉴定。向3组培养基中加入不同抗菌药物,利用K-B方法观察细菌耐药性情况。结果 B组与A组药物耐药率和最低抑菌浓度(MIC)值差异均无统计学意义(p>0.05);C组菌株与A组比较,对美洛培南(50.00%vs 26.47%)、头孢吡肟(53.52%vs 25.00%)、头孢他啶(60.94% vs 20.59%)、头孢曲松(70.31% vs 61.76%)、左氧氟沙星(39.84%vs 2.94%)及哌拉西林(48.05%vs 8.82%)的耐药性差异均有统计学意义( p0. 01). Compared with group A, group C showed that their drug resistance on meropenem,cefepime,ceftazidime,ceftriaxone,amoxicillin and levofloxacin were much higher(p<0. 01). The minimal biofilm eradication concentration(MBEC)of group C was about 100 times than that of group A and group B. Conclusion The assay of establishment of pseudomonas aeruginosa biofilm and the methods observed by silver stainingin vitro is convenient and feasible. At the same time,pseudomonas aeruginosa in biofilm exhibits far more resistance to antimicrobial than its planctonic counterparts does.

  19. Eradication of Bacterial Biofilms Using Atmospheric Pressure Non-Thermal Plasmas

    Science.gov (United States)

    Alkawareek, Mahmoud; Gilmore, Brendan; Gorman, Sean; Algwari, Qais; Graham, William; O'Connell, Deborah

    2011-10-01

    Bacterial biofilms are ubiquitous in natural and clinical settings and form a major health risk. Biofilms are recognised to be the predominant mode of bacterial growth, and are an immunological challenge compared to planktonic bacteria of the same species. Eradication of biofilms with atmospheric pressure plasma jets is investigated. Cold non-equilibrium plasmas, operated at ambient atmospheric pressure and temperature, are efficient sources for controlled energy transport through highly reactive neutrals (e.g. ROS, RNS), charged particles (ions and electrons), UV radiation, and electro-magnetic fields. A focused panel of clinically significant biofilms, including Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, and Bacillus cereus, are exposed to various plasma jet configurations operated in helium and oxygen mixtures. Viability of surviving cells was determined using both standard plate counting method and XTT viability assay. These are correlated with measurements and simulations of relevant reactive plasma species.

  20. Biofilm and siderophore effects on secondary waste water disinfection.

    Science.gov (United States)

    Saidi, N; Kouki, S; Mehri, I; Ben Rejeb, A; Belila, A; Hassen, A; Ouzari, H

    2011-10-01

    The efficiency of ultraviolet (UV) light disinfection of wastewater effluent using a large-scale pilot system was studied. The relationship between biofilm and siderophore production and UV doses received by Pseudomonas aeruginosa strain ATCC 15442 was determined. UV decreased pyoverdine production and enhanced biofilm production. Consequently external factors conditioned by both pyoverdine and biofilm may affect the UV effect on bacterial disinfection.