Silver colloidal nanoparticles: effect on matrix composition and structure of Candida albicans and Candida glabrata biofilms.

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Monteiro, D R; Silva, S; Negri, M; Gorup, L F; de Camargo, E R; Oliveira, R; Barbosa, D B; Henriques, M

2013-04-01

The aim of this study was to assess the effect of different silver nanoparticles (SN) concentrations on the matrix composition and structure of Candida albicans and Candida glabrata biofilms. Candida biofilms were developed in 6-well microtiter plates during 48 h. After, these biofilms were exposed to 13.5 or 54 μg SN ml(-1) for 24 h. Then, extracellular matrices were extracted from biofilms and analysed chemically in terms of proteins, carbohydrates and DNA. To investigate the biofilm structure, scanning electron microscopy (SEM) and epifluorescence microscopy were used. SN interfered with the matrix composition of Candida biofilms tested in terms of protein, carbohydrate and DNA, except for the protein content of C. albicans biofilm. By SEM, Candida biofilms treated with SN revealed structural differences, when compared with the control groups. Further, SN showed a trend of agglomeration within the biofilms. Epifluorescence microscopy images suggest that SN induced damage on cell walls of the Candida isolates tested. In general, irrespective of concentration, SN affected the matrix composition and structure of Candida biofilms and these findings may be related to the mechanisms of biocide action of SN. This study reveals new insights about the behaviour of SN when in contact with Candida biofilms. SN may contribute to the development of therapies to prevent or control Candida infections. © 2012 The Society for Applied Microbiology.

Simvastatin inhibits Candida albicans biofilm in vitro.

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Liu, Geoffrey; Vellucci, Vincent F; Kyc, Stephanie; Hostetter, Margaret K

2009-12-01

By inhibiting the conversion of 3-hydroxy-3-methylglutaryl CoA (HMG-CoA) to mevalonate, statins impair cholesterol metabolism in humans. We reasoned that statins might similarly interfere with the biosynthesis of ergosterol, the major sterol of the yeast cell membrane. As assessed by spectrophotometric and microscopic analysis, significant inhibition of biofilm production was noted after 16-h incubation with 1, 2.5, and 5 muM simvastatin, concentrations that did not affect growth, adhesion, or hyphal formation by C. albicans in vitro. Higher concentrations (10, 20, and 25 muM simvastatin) inhibited biofilm by >90% but also impaired growth. Addition of exogenous ergosterol (90 muM) overcame the effects of 1 and 2.5 muM simvastatin, suggesting that at least one mechanism of inhibition is interference with ergosterol biosynthesis. Clinical isolates from blood, skin, and mucosal surfaces produced biofilms; biofilms from bloodstream isolates were similarly inhibited by simvastatin. In the absence of fungicidal activity, simvastatin's interruption of a critical step in an essential metabolic pathway, highly conserved from yeast to man, has unexpected effects on biofilm production by a eukaryotic pathogen.

Species-Specific and Drug-Specific Differences in Susceptibility of Candida Biofilms to Echinocandins: Characterization of Less Common Bloodstream Isolates

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Simitsopoulou, Maria; Peshkova, Pavla; Tasina, Efthymia; Katragkou, Aspasia; Kyrpitzi, Daniela; Velegraki, Aristea; Walsh, Thomas J.

2013-01-01

Candida species other than Candida albicans are increasingly recognized as causes of biofilm-associated infections. This is a comprehensive study that compared the in vitro activities of all three echinocandins against biofilms formed by different common and infrequently identified Candida isolates. We determined the activities of anidulafungin (ANID), caspofungin (CAS), and micafungin (MFG) against planktonic cells and biofilms of bloodstream isolates of C. albicans (15 strains), Candida parapsilosis (6 strains), Candida lusitaniae (16 strains), Candida guilliermondii (5 strains), and Candida krusei (12 strains) by XTT [2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide] assay. Planktonic and biofilm MICs were defined as ≥50% fungal damage. Planktonic cells of all Candida species were susceptible to the three echinocandins, with MICs of ≤1 mg/liter. By comparison, differences in the MIC profiles of biofilms in response to echinocandins existed among the Candida species. Thus, C. lusitaniae and C. guilliermondii biofilms were highly recalcitrant to all echinocandins, with MICs of ≥32 mg/liter. In contrast, the MICs of all three echinocandins for C. albicans and C. krusei biofilms were relatively low (MICs ≤ 1 mg/liter). While echinocandins exhibited generally high MICs against C. parapsilosis biofilms, MFG exhibited the lowest MICs against these isolates (4 mg/liter). A paradoxical growth effect was observed with CAS concentrations ranging from 8 to 64 mg/liter against C. albicans and C. parapsilosis biofilms but not against C. krusei, C. lusitaniae, or C. guilliermondii. While non-albicans Candida planktonic cells were susceptible to all echinocandins, there were drug- and species-specific differences in susceptibility among biofilms of the various Candida species, with C. lusitaniae and C. guilliermondii exhibiting profiles of high MICs of the three echinocandins. PMID:23529739

Antifungal Activity of 14-Helical β-Peptides against Planktonic Cells and Biofilms of Candida Species

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Namrata Raman

2015-08-01

Full Text Available Candida albicans is the most prevalent cause of fungal infections and treatment is further complicated by the formation of drug resistant biofilms, often on the surfaces of implanted medical devices. In recent years, the incidence of fungal infections by other pathogenic Candida species such as C. glabrata, C. parapsilosis and C. tropicalis has increased. Amphiphilic, helical β-peptide structural mimetics of natural antimicrobial α-peptides have been shown to exhibit specific planktonic antifungal and anti-biofilm formation activity against C. albicans in vitro. Here, we demonstrate that β-peptides are also active against clinically isolated and drug resistant strains of C. albicans and against other opportunistic Candida spp. Different Candida species were susceptible to β-peptides to varying degrees, with C. tropicalis being the most and C. glabrata being the least susceptible. β-peptide hydrophobicity directly correlated with antifungal activity against all the Candida clinical strains and species tested. While β-peptides were largely ineffective at disrupting existing Candida biofilms, hydrophobic β-peptides were able to prevent the formation of C. albicans, C. glabrata, C. parapsilosis and C. tropicalis biofilms. The broad-spectrum antifungal activity of β-peptides against planktonic cells and in preventing biofilm formation suggests the promise of this class of molecules as therapeutics.

Biosurfactants prevent in vitro Candida albicans biofilm formation on resins and silicon materials for prosthetic devices.

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Cochis, Andrea; Fracchia, Letizia; Martinotti, Maria Giovanna; Rimondini, Lia

2012-06-01

The aim of this study was to evaluate in vitro the preventive antiadhesion activity of biosurfactants against Candida albicans biofilm. Disks of silicon and acrylic resin for denture prostheses were precoated with increasing concentrations of biosurfactants obtained from endophyte biofilms selected from Robinia pseudoacacia and from Nerium oleander, and afterward infected with C. albicans cells. The number of biofilm cells were detected by colony-forming unit (CFU) counting, cell viability was established by the 2,3-bis(2-methoxy-4-nitro-5-sulphophenyl)-5-[(phenyl amino)carbonyl]-2H-tetrazolium hydroxide (XTT) assay, and biosurfactant cytotoxicity was evaluated by the [3-(4,5-dimethyliazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulphophenyl)-2H-tetrazolium] (MTT) assay. Chlorhexidine was used as control. Precoating with biosurfactants caused a greater reduction (P biosurfactants was observed at low concentrations (78.12 μg/mL and 156.12 μg/mL) which were noncytotoxic. This study demonstrated the preventive antiadhesion activity of biosurfactants against C. albicans biofilm. These agents are amphiphilic, interfere with microbial adhesion, and demonstrate cycompatibility with epithelial cells and fibroblasts. Copyright © 2012 Elsevier Inc. All rights reserved.

Candida Biofilms: Development, Architecture, and Resistance

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CHANDRA, JYOTSNA; MUKHERJEE, PRANAB K.

2015-01-01

Intravascular device–related infections are often associated with biofilms (microbial communities encased within a polysaccharide-rich extracellular matrix) formed by pathogens on the surfaces of these devices. Candida species are the most common fungi isolated from catheter-, denture-, and voice prosthesis–associated infections and also are commonly isolated from contact lens–related infections (e.g., fungal keratitis). These biofilms exhibit decreased susceptibility to most antimicrobial agents, which contributes to the persistence of infection. Recent technological advances have facilitated the development of novel approaches to investigate the formation of biofilms and identify specific markers for biofilms. These studies have provided extensive knowledge of the effect of different variables, including growth time, nutrients, and physiological conditions, on biofilm formation, morphology, and architecture. In this article, we will focus on fungal biofilms (mainly Candida biofilms) and provide an update on the development, architecture, and resistance mechanisms of biofilms. PMID:26350306

Lab-scale preparations of Candida albicans and dual Candida albicans-Candida glabrata biofilms on the surface of medical-grade polyvinyl chloride (PVC) perfusion tube using a modified gravity-supported free-flow biofilm incubator (GS-FFBI).

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Shao, Jing; Lu, KeQiao; Tian, Ge; Cui, YanYan; Yan, YuanYuan; Wang, TianMing; Zhang, XinLong; Wang, ChangZhong

2015-02-01

The assembly of a man-made gravity-supported free-flow biofilm incubator (GS-FFBI) was described, which was composed of a gas cushion injector and four incubators. The GS-FFBI had the characteristics of (i) a bottom-up flow direction, and (ii) lab-scale biofilm preparation without the use of a multichannel pump. Two opportunistic fungal strains, namely Candida albicans and Candida glabrata, were employed to incubate C. albicans and dual C. albicans-C. glabrata biofilms on the surface of medical-grade polyvinyl chloride perfusion tube. In terms of the results from {2, 3-bis (2-methoxy-4-nitro-5-sulfo-phenyl)-2H-tetrazolium-5-carboxanilide} (XTT) assay, dry weight measurement, colony-forming unit counting, susceptibility test, and scanning electron microscopy, it was demonstrated that GS-FFBI could form both stable single and dual Candida biofilms with no significant variations among the four incubators or the three daily incubations within 21h, and could operate for at least 96h smoothly with no contamination of stock medium. The results also indicated, for the first time, that C. albicans and C. glabrata might be co-existent competitively and symbiotically in the dual biofilms with flowing media. GS-FFBI would be a useful device to study in vitro morphological and physiological features of microbial biofilms in the medical settings. Copyright © 2014 Elsevier B.V. All rights reserved.

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

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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

Mechanisms of Candida biofilm drug resistance

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Taff, Heather T; Mitchell, Kaitlin F; Edward, Jessica A; Andes, David R

2013-01-01

Candida commonly adheres to implanted medical devices, growing as a resilient biofilm capable of withstanding extraordinarily high antifungal concentrations. As currently available antifungals have minimal activity against biofilms, new drugs to treat these recalcitrant infections are urgently needed. Recent investigations have begun to shed light on the mechanisms behind the profound resistance associated with the biofilm mode of growth. This resistance appears to be multifactorial, involving both mechanisms similar to conventional, planktonic antifungal resistance, such as increased efflux pump activity, as well as mechanisms specific to the biofilm lifestyle. A unique biofilm property is the production of an extracellular matrix. Two components of this material, β-glucan and extracellular DNA, promote biofilm resistance to multiple antifungals. Biofilm formation also engages several stress response pathways that impair the activity of azole drugs. Resistance within a biofilm is often heterogeneous, with the development of a subpopulation of resistant persister cells. In this article we review the molecular mechanisms underlying Candida biofilm antifungal resistance and their relative contributions during various growth phases. PMID:24059922

Gallic acid/hydroxypropyl-β-cyclodextrin complex: Improving solubility for application on in vitro/ in vivo Candida albicans biofilms.

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Guilherme Rodrigues Teodoro

Full Text Available The aim of this study was to increase the solubility of gallic acid (GA for the treatment of Candida albicans biofilm, which is very difficult to treat and requires high drug concentrations. Cyclodextrins (CDs were used for this purpose. Complexes were evaluated by phase-solubility studies, prepared by spray drying and characterized by drug loading, scanning electron microscopy (SEM and differential scanning calorimetry (DSC. The complexes were tested on C. albicans biofilm using in vitro and in vivo models. HPβCD formed soluble inclusion complexes with GA. The percentage of GA in GA/HPβCD was 10.8 ± 0.01%. The SEM and DSC analyses confirmed the formation of inclusion complexes. GA/HPβCD maintained the antimicrobial activity of the pure GA. GA/HPβCD was effective on C. albicans biofilms of 24 and 48h. The in vivo results showed an anti-inflammatory activity of GA/HPβCD with no difference in invading hypha counting among the groups. This study encourages the development of new antifungal agents.

In vitro activity of xanthorrhizol isolated from the rhizome of Javanese turmeric (Curcuma xanthorrhiza Roxb.) against Candida albicans biofilms.

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Rukayadi, Yaya; Hwang, Jae-Kwan

2013-07-01

The purpose of this study was to investigate the activity of xanthorrhizol isolated from Curcuma xanthorrhiza Roxb. on Candida albicans biofilms at adherent, intermediate, and mature phase of growth. C. albicans biofilms were formed in flat-bottom 96-well microtiter plates. The biofilms of C. albicans at different phases of development were exposed to xanthorrhizol at different concentrations (0.5 µg/mL-256 µg/mL) for 24 h. The metabolic activity of cells within the biofilms was quantified using the XTT reduction assay. Sessile minimum inhibitory concentrations (SMICs) were determined at 50% and 80% reduction in the biofilm OD₄₉₀ compared to the control wells. The SMIC₅₀ and SMIC₈₀ of xanthorrhizol against 18 C. albicans biofilms were 4--16 µg/mL and 8--32 µg/mL, respectively. The results demonstrated that the activity of xanthorrhizol in reducing C. albicans biofilms OD₄₉₀ was dependent on the concentration and the phase of growth of biofilm. Xanthorrhizol at concentration of 8 µg/mL completely reduced in biofilm referring to XTT-colorimetric readings at adherent phase, whereas 32 µg/mL of xanthorrhizol reduced 87.95% and 67.48 % of biofilm referring to XTT-colorimetric readings at intermediate and mature phases, respectively. Xanthorrhizol displayed potent activity against C. albicans biofilms in vitro and therefore might have potential therapeutic implication for biofilm-associated candidal infections. Copyright © 2012 John Wiley & Sons, Ltd.

Culture media profoundly affect Candida albicans and Candida tropicalis growth, adhesion and biofilm development.

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Weerasekera, Manjula M; Wijesinghe, Gayan K; Jayarathna, Thilini A; Gunasekara, Chinthika P; Fernando, Neluka; Kottegoda, Nilwala; Samaranayake, Lakshman P

2016-11-01

As there are sparse data on the impact of growth media on the phenomenon of biofilm development for Candida we evaluated the efficacy of three culture media on growth, adhesion and biofilm formation of two pathogenic yeasts, Candida albicans and Candida tropicalis. The planktonic phase yeast growth, either as monocultures or mixed cultures, in sabouraud dextrose broth (SDB), yeast nitrogen base (YNB), and RPMI 1640 was compared, and adhesion as well as biofilm formation were monitored using MTT and crystal violet (CV) assays and scanning electron microscopy. Planktonic cells of C. albicans, C. tropicalis and their 1:1 co-culture showed maximal growth in SDB. C. albicans/C. tropicalis adhesion was significantly facilitated in RPMI 1640 although the YNB elicited the maximum growth for C. tropicalis. Similarly, the biofilm growth was uniformly higher for both species in RPMI 1640, and C. tropicalis was the slower biofilm former in all three media. Scanning electron microscopy images tended to confirm the results of MTT and CV assay. Taken together, our data indicate that researchers should pay heed to the choice of laboratory culture media when comparing relative planktonic/biofilm growth of Candida. There is also a need for standardisation of biofilm development media so as to facilitate cross comparisons between laboratories.

Sexual Biofilm Formation in Candida tropicalis Opaque Cells

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Jones, Stephen K.; Hirakawa, Matthew P.; Bennett, Richard J.

2014-01-01

Summary Candida albicans and Candida tropicalis are opportunistic fungal pathogens that can transition between white and opaque phenotypic states. White and opaque cells differ both morphologically and in their responses to environmental signals. In C. albicans, opaque cells respond to sexual pheromones by undergoing conjugation, while white cells are induced by pheromones to form sexual biofilms. Here, we show that sexual biofilm formation also occurs in C. tropicalis but, unlike C. albicans, biofilms are formed exclusively by opaque cells. C. tropicalis biofilm formation was dependent on the pheromone receptors Ste2 and Ste3, confirming the role of pheromone signaling in sexual biofilm development. Structural analysis of C. tropicalis sexual biofilms revealed stratified communities consisting of a basal layer of yeast cells and an upper layer of filamentous cells, together with an extracellular matrix. Transcriptional profiling showed that genes involved in pheromone signaling and conjugation were upregulated in sexual biofilms. Furthermore, FGR23, which encodes an agglutinin-like protein, was found to enhance both mating and sexual biofilm formation. Together, these studies reveal that C. tropicalis opaque cells form sexual biofilms with a complex architecture, and suggest a conserved role for sexual agglutinins in mediating mating, cell cohesion and biofilm formation. PMID:24612417

Biofilm extracellular DNA enhances mixed species biofilms of Staphylococcus epidermidis and Candida albicans.

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Pammi, Mohan; Liang, Rong; Hicks, John; Mistretta, Toni-Ann; Versalovic, James

2013-11-14

Polymicrobial infections are responsible for significant mortality and morbidity in adults and children. Staphylococcus epidermidis and Candida albicans are the most frequent combination of organisms isolated from polymicrobial infections. Vascular indwelling catheters are sites for mixed species biofilm formation and pose a significant risk for polymicrobial infections. We hypothesized that enhancement of biofilms in a mixed species environment increases patient mortality and morbidity. Mixed species biofilms of S. epidermidis and C. albicans were evaluated in vitro and in a subcutaneous catheter infection model in vivo. Mixed species biofilms were enhanced compared to single species biofilms of either S. epidermidis or C. albicans. A mixed species environment increased catheter infection and increased dissemination of S. epidermidis in mice. Microarrays were used to explore differential gene expression of S. epidermidis in the mixed species biofilms. In mixed species biofilms, compared to single species S. epidermidis biofilms, 2.7% of S. epidermidis genes were upregulated and 6% were down regulated. Staphylococcal autolysis repressors lrgA and lrgB were down regulated 36-fold and 27-fold respectively. The role of biofilm extracellular DNA was investigated by quantitation and by evaluating the effects of DNAse in a concentration and time dependent manner. S. epidermidis specific eDNA was increased in mixed species biofilms and further confirmed by degradation with DNAse. Mixed-species biofilms are enhanced and associated with increased S. epidermidis-specific eDNA in vitro and greater systemic dissemination of S. epidermidis in vivo. Down regulation of the lrg operon, a repressor of autolysis, associated with increased eDNA suggests a possible role for bacterial autolysis in mixed species biofilms. Enhancement and systemic dissemination of S. epidermidis may explain adverse outcomes after clinical polymicrobial infections of S. epidermidis and C. albicans.

Biofilm extracellular DNA enhances mixed species biofilms of Staphylococcus epidermidis and Candida albicans

Science.gov (United States)

2013-01-01

Background Polymicrobial infections are responsible for significant mortality and morbidity in adults and children. Staphylococcus epidermidis and Candida albicans are the most frequent combination of organisms isolated from polymicrobial infections. Vascular indwelling catheters are sites for mixed species biofilm formation and pose a significant risk for polymicrobial infections. We hypothesized that enhancement of biofilms in a mixed species environment increases patient mortality and morbidity. Results Mixed species biofilms of S. epidermidis and C. albicans were evaluated in vitro and in a subcutaneous catheter infection model in vivo. Mixed species biofilms were enhanced compared to single species biofilms of either S. epidermidis or C. albicans. A mixed species environment increased catheter infection and increased dissemination of S. epidermidis in mice. Microarrays were used to explore differential gene expression of S. epidermidis in the mixed species biofilms. In mixed species biofilms, compared to single species S. epidermidis biofilms, 2.7% of S. epidermidis genes were upregulated and 6% were down regulated. Staphylococcal autolysis repressors lrgA and lrgB were down regulated 36-fold and 27-fold respectively. The role of biofilm extracellular DNA was investigated by quantitation and by evaluating the effects of DNAse in a concentration and time dependent manner. S. epidermidis specific eDNA was increased in mixed species biofilms and further confirmed by degradation with DNAse. Conclusions Mixed-species biofilms are enhanced and associated with increased S. epidermidis-specific eDNA in vitro and greater systemic dissemination of S. epidermidis in vivo. Down regulation of the lrg operon, a repressor of autolysis, associated with increased eDNA suggests a possible role for bacterial autolysis in mixed species biofilms. Enhancement and systemic dissemination of S. epidermidis may explain adverse outcomes after clinical polymicrobial infections of S

Inhibitory effect of farnesol on biofilm formation by Candida tropicalis

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E Zibafar

2009-03-01

Full Text Available ABSTRACT Background: Candidiasis associated with indwelling medical devices is especially problematic since they can act as substrates for biofilm growth which are highly resistant to antifungal drugs. Farnesol is a quorum-sensing molecule that inhibits filamentation and biofilm formation in Candida albicans. Since in recent years Candida tropicalis have been reported as an important and common non-albicans Candida species with high drug resistance pattern, the inhibitory effect of farnesol on biofilm formation by Candida tropicalis was evaluated. Methods: Five Candida tropicalis strains were treated with different concentration of farnesol (0, 30 and 300 µM after 0, 1 and 4 hrs of adherence and then they were maintained under biofilm formation condition in polystyrene, 96-well microtiter plates at 37°C for 48 hrs. Biofilm formation was measured by a semiquantitative colorimetric technique based on reduction assay of 2,3- bis  -2H-tetrazolium- 5- carboxanilide (XTT. Results: The results indicated that the initial adherence time had no effect on biofilm formation and low concentration of farnesol (30 µM could not inhibit biofilm formation. However the presence of non-adherent cells increased biofilm formation significantly and the high concentration of farnesol (300 µM could inhibit biofilm formation. Conclusion: Results of this study showed that the high concentration of farnesol could inhibit biofilm formation and may be used as an adjuvant in prevention and in therapeutic strategies with antifungal drugs.

Lactobacillus rhamnosus inhibits Candida albicans virulence factors in vitro and modulates immune system in Galleria mellonella.

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Ribeiro, F C; de Barros, P P; Rossoni, R D; Junqueira, J C; Jorge, A O C

2017-01-01

The aim of this study was to evaluate the potential anti-Candida effects of Lactobacillus rhamnosus ATCC 9595 on Candida albicans ATCC 18804 using in vitro and in vivo models. The in vitro analysis evaluated the effects of L. rhamnosus on C. albicans's biofilm formation by CFU count and metabolic activity, filamentation capacity, and adhesion (ALS3 and HWP1) and transcriptional regulatory gene (BCR1 and CPH1) expression. The in vitro results showed that both the L. rhamnosus cells and supernatant reduced C. albicans biofilm formation, filamentation and gene expression. In the in vivo study, the treatment with L. rhamnosus supernatant increased 80% the survival of Galleria mellonella larvae infected with C. albicans. Furthermore, the supernatant of L. rhamnosus recruited haemocytes into the haemolymph (2·1-fold increase). Lactobacillus rhamnosus reduced the biofilm formation and filamentation of C. albicans in vitro by negatively regulating all studied C. albicans genes. Lactobacillus rhamnosus protected G. mellonella against experimental candidiasis in vivo. This study is the first study to report the anti-Candida properties of L. rhamnosus ATCC 9595. The supernatant of this strain has immunomodulatory effects on the G. mellonella model and protects the larvae against pathogens. © 2016 The Society for Applied Microbiology.

Identification, antifungal resistance profile, in vitro biofilm formation and ultrastructural characteristics of Candida species isolated from diabetic foot patients in Northern India.

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Kumar, D; Banerjee, T; Chakravarty, J; Singh, S K; Dwivedi, A; Tilak, R

2016-01-01

Diabetic foot ulcers are a serious cause of diagnostic and therapeutic concern. The following study was undertaken to determine the fungal causes of diabetic foot ulcers, with their phenotypic and genotypic characterisation. A total of 155 diabetic foot ulcers were studied for 1 year. Deep tissue specimen was collected from the wounds, and crushed samples were plated on Sabouraud dextrose agar with chloramphenicol (0.05 g). Identification was done by growth on cornmeal agar, germ tube formation and urease test. For molecular identification, conserved portion of the 18S rDNA region, the adjacent internal transcribed spacer 1 (ITS1) and a portion of the 28S rDNA region were amplified, using the ITS1 and ITS2 primers. Antifungal susceptibility against voriconazole, fluconazole and amphotericin B was determined by standard broth microdilution method. Biofilm formation was studied in three steps. First, on the surface of wells of microtiter plates followed by quantification of growth by fungal metabolism measurement. Finally, biofilms were analysed by scanning electron microscopy (SEM). Fungal aetiology was found in 75 patients (48.38%). All were identified as Candida species (100%). The prevalence of different species was Candida tropicalis (34.6%), Candida albicans (29.3%), Candida krusei (16.0%), Candida parapsilosis (10.6%), Candida glabrata (9.33%). All were susceptible to amphotericin B (100%). On microtiter plate, all the isolates were viable within 48 h showing biofilms. The metabolic activity of cells in the biofilm increased with cellular mass, especially in the first 24 h. On SEM, majority showed budding yeast form. Non-albicans Candida spp. with potential biofilm forming ability are emerging as a predominant cause of diabetic foot ulcers.

The European Confederation of Medical Mycology (ECMM) survey of candidaemia in Italy: in vitro susceptibility of 375 Candida albicans isolates and biofilm production.

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Tortorano, Anna Maria; Prigitano, Anna; Biraghi, Emanuela; Viviani, Maria Anna

2005-10-01

To investigate the in vitro antifungal susceptibility pattern of 375 Candida albicans bloodstream isolates recovered during the European Confederation of Medical Mycology survey of candidaemia performed in Lombardia, Italy and to test the ability to form biofilm. In vitro susceptibility to flucytosine, fluconazole, itraconazole, posaconazole, voriconazole and caspofungin was performed by broth microdilution following the NCCLS guidelines. Biofilm production was measured using the XTT reduction assay in 59 isolates selected as representative of different patterns of susceptibility to flucytosine and azoles. MICs (mg/L) at which 90% of the strains were inhibited were < or =0.25 for flucytosine, 0.25 for caspofungin, 4 for fluconazole and 0.06 for itraconazole, voriconazole and posaconazole. Flucytosine resistance was detected in five isolates and was associated with serotype B in 2/29 and serotype A in 3/346. Resistance to fluconazole was detected in 10 isolates; nine of these exhibited reduced susceptibility to the other azoles. Among the 10 patients with fluconazole-resistant C. albicans bloodstream infection, only one, an AIDS patient, had been previously treated with fluconazole. Biofilm production was observed in 23 isolates (39%) and was significantly associated with serotype B. No relationship was detected with the pattern of antifungal susceptibility. Resistance is uncommon in C. albicans isolates recovered from blood cultures, while biofilm production is a relatively frequent event. Periodic surveillance is warranted to monitor the incidence of in vitro antifungal resistance as well as of biofilm production.

Candida glabrata Biofilms: How Far Have We Come?

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Célia F. Rodrigues

2017-03-01

Full Text Available Infections caused by Candida species have been increasing in the last decades and can result in local or systemic infections, with high morbidity and mortality. After Candida albicans, Candida glabrata is one of the most prevalent pathogenic fungi in humans. In addition to the high antifungal drugs resistance and inability to form hyphae or secret hydrolases, C. glabrata retain many virulence factors that contribute to its extreme aggressiveness and result in a low therapeutic response and serious recurrent candidiasis, particularly biofilm formation ability. For their extraordinary organization, especially regarding the complex structure of the matrix, biofilms are very resistant to antifungal treatments. Thus, new approaches to the treatment of C. glabrata’s biofilms are emerging. In this article, the knowledge available on C. glabrata’s resistance will be highlighted, with a special focus on biofilms, as well as new therapeutic alternatives to control them.

Candida glabrata Biofilms: How Far Have We Come?

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Rodrigues, Célia F.; Rodrigues, Maria Elisa; Silva, Sónia; Henriques, Mariana

2017-01-01

Infections caused by Candida species have been increasing in the last decades and can result in local or systemic infections, with high morbidity and mortality. After Candida albicans, Candida glabrata is one of the most prevalent pathogenic fungi in humans. In addition to the high antifungal drugs resistance and inability to form hyphae or secret hydrolases, C. glabrata retain many virulence factors that contribute to its extreme aggressiveness and result in a low therapeutic response and serious recurrent candidiasis, particularly biofilm formation ability. For their extraordinary organization, especially regarding the complex structure of the matrix, biofilms are very resistant to antifungal treatments. Thus, new approaches to the treatment of C. glabrata’s biofilms are emerging. In this article, the knowledge available on C. glabrata’s resistance will be highlighted, with a special focus on biofilms, as well as new therapeutic alternatives to control them. PMID:29371530

Distribution of Candida albicans and non-albicans Candida species in oral candidiasis patients: Correlation between cell surface hydrophobicity and biofilm forming activities.

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Muadcheingka, Thaniya; Tantivitayakul, Pornpen

2015-06-01

The purposes of this investigation were to study the prevalence of Candida albicans and non-albicans Candida (NAC) species from oral candidiasis patients and evaluate the cell surface hydrophobicity (CSH) and biofilm forming capacity of the clinical isolates Candida species from oral cavity. This study identified a total of 250 Candida strains isolated from 207 oral candidiasis patients with PCR-RFLP technique. CSH value, total biomass of biofilm and biofilm forming ability of 117 oral Candida isolates were evaluated. C. albicans (61.6%) was still the predominant species in oral candidiasis patients with and without denture wearer, respectively, followed by C. glabrata (15.2%), C. tropicalis (10.4%), C. parapsilosis (3.2%), C. kefyr (3.6%), C. dubliniensis (2%), C. lusitaniae (2%), C. krusei (1.6%), and C. guilliermondii (0.4%). The proportion of mixed colonization with more than one Candida species was 18% from total cases. The relative CSH value and biofilm biomass of NAC species were greater than C. albicans (poral isolates NAC species had biofilm forming ability, whereas 78% of C. albicans were biofilm formers. Furthermore, the significant difference of relative CSH values between biofilm formers and non-biofilm formers was observed in the NAC species (poral cavity was gradually increasing. The possible contributing factors might be high cell surface hydrophobicity and biofilm forming ability. The relative CSH value could be a putative factor for determining biofilm formation ability of the non-albicans Candida species. Copyright © 2015 Elsevier Ltd. All rights reserved.

Identification, antifungal resistance profile, in vitro biofilm formation and ultrastructural characteristics of Candida species isolated from diabetic foot patients in Northern India

Directory of Open Access Journals (Sweden)

D Kumar

2016-01-01

Full Text Available Purpose: Diabetic foot ulcers are a serious cause of diagnostic and therapeutic concern. The following study was undertaken to determine the fungal causes of diabetic foot ulcers, with their phenotypic and genotypic characterisation. Materials and Methods: A total of 155 diabetic foot ulcers were studied for 1 year. Deep tissue specimen was collected from the wounds, and crushed samples were plated on Sabouraud dextrose agar with chloramphenicol (0.05 g. Identification was done by growth on cornmeal agar, germ tube formation and urease test. For molecular identification, conserved portion of the 18S rDNA region, the adjacent internal transcribed spacer 1 (ITS1 and a portion of the 28S rDNA region were amplified, using the ITS1 and ITS2 primers. Antifungal susceptibility against voriconazole, fluconazole and amphotericin B was determined by standard broth microdilution method. Biofilm formation was studied in three steps. First, on the surface of wells of microtiter plates followed by quantification of growth by fungal metabolism measurement. Finally, biofilms were analysed by scanning electron microscopy (SEM. Results: Fungal aetiology was found in 75 patients (48.38%. All were identified as Candida species (100%. The prevalence of different species was Candida tropicalis (34.6%, Candida albicans (29.3%, Candida krusei (16.0%, Candida parapsilosis (10.6%, Candida glabrata (9.33%. All were susceptible to amphotericin B (100%. On microtiter plate, all the isolates were viable within 48 h showing biofilms. The metabolic activity of cells in the biofilm increased with cellular mass, especially in the first 24 h. On SEM, majority showed budding yeast form. Conclusion: Non-albicans Candida spp. with potential biofilm forming ability are emerging as a predominant cause of diabetic foot ulcers.

Symbiotic Relationship between Streptococcus mutans and Candida albicans Synergizes Virulence of Plaque Biofilms In Vivo

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Falsetta, Megan L.; Klein, Marlise I.; Colonne, Punsiri M.; Scott-Anne, Kathleen; Gregoire, Stacy; Pai, Chia-Hua; Gonzalez-Begne, Mireya; Watson, Gene; Krysan, Damian J.; Bowen, William H.

2014-01-01

Streptococcus mutans is often cited as the main bacterial pathogen in dental caries, particularly in early-childhood caries (ECC). S. mutans may not act alone; Candida albicans cells are frequently detected along with heavy infection by S. mutans in plaque biofilms from ECC-affected children. It remains to be elucidated whether this association is involved in the enhancement of biofilm virulence. We showed that the ability of these organisms together to form biofilms is enhanced in vitro and in vivo. The presence of C. albicans augments the production of exopolysaccharides (EPS), such that cospecies biofilms accrue more biomass and harbor more viable S. mutans cells than single-species biofilms. The resulting 3-dimensional biofilm architecture displays sizeable S. mutans microcolonies surrounded by fungal cells, which are enmeshed in a dense EPS-rich matrix. Using a rodent model, we explored the implications of this cross-kingdom interaction for the pathogenesis of dental caries. Coinfected animals displayed higher levels of infection and microbial carriage within plaque biofilms than animals infected with either species alone. Furthermore, coinfection synergistically enhanced biofilm virulence, leading to aggressive onset of the disease with rampant carious lesions. Our in vitro data also revealed that glucosyltransferase-derived EPS is a key mediator of cospecies biofilm development and that coexistence with C. albicans induces the expression of virulence genes in S. mutans (e.g., gtfB, fabM). We also found that Candida-derived β1,3-glucans contribute to the EPS matrix structure, while fungal mannan and β-glucan provide sites for GtfB binding and activity. Altogether, we demonstrate a novel mutualistic bacterium-fungus relationship that occurs at a clinically relevant site to amplify the severity of a ubiquitous infectious disease. PMID:24566629

Hibiscus sabdariffa extract inhibits in vitro biofilm formation capacity of Candida albicans isolated from recurrent urinary tract infections.

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Alshami, Issam; Alharbi, Ahmed E

2014-02-01

To explore the prevention of recurrent candiduria using natural based approaches and to study the antimicrobial effect of Hibiscus sabdariffa (H. sabdariffa) extract and the biofilm forming capacity of Candida albicans strains in the present of the H. sabdariffa extract. In this particular study, six strains of fluconazole resistant Candida albicans isolated from recurrent candiduria were used. The susceptibility of fungal isolates, time-kill curves and biofilm forming capacity in the present of the H. sabdariffa extract were determined. Various levels minimum inhibitory concentration of the extract were observed against all the isolates. Minimum inhibitory concentration values ranged from 0.5 to 2.0 mg/mL. Time-kill experiment demonstrated that the effect was fungistatic. The biofilm inhibition assay results showed that H. sabdariffa extract inhibited biofilm production of all the isolates. The results of the study support the potential effect of H. sabdariffa extract for preventing recurrent candiduria and emphasize the significance of the plant extract approach as a potential antifungal agent.

In vitro and in vivo activity of a novel antifungal small molecule against Candida infections.

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Sarah Sze Wah Wong

Full Text Available Candida is the most common fungal pathogen of humans worldwide and has become a major clinical problem because of the growing number of immunocompromised patients, who are susceptible to infection. Moreover, the number of available antifungals is limited, and antifungal-resistant Candida strains are emerging. New and effective antifungals are therefore urgently needed. Here, we discovered a small molecule with activity against Candida spp. both in vitro and in vivo. We screened a library of 50,240 small molecules for inhibitors of yeast-to-hypha transition, a major virulence attribute of Candida albicans. This screening identified 20 active compounds. Further examination of the in vitro antifungal and anti-biofilm properties of these compounds, using a range of Candida spp., led to the discovery of SM21, a highly potent antifungal molecule (minimum inhibitory concentration (MIC 0.2-1.6 µg/ml. In vitro, SM21 was toxic to fungi but not to various human cell lines or bacterial species and was active against Candida isolates that are resistant to existing antifungal agents. Moreover, SM21 was relatively more effective against biofilms of Candida spp. than the current antifungal agents. In vivo, SM21 prevented the death of mice in a systemic candidiasis model and was also more effective than the common antifungal nystatin at reducing the extent of tongue lesions in a mouse model of oral candidiasis. Propidium iodide uptake assay showed that SM21 affected the integrity of the cell membrane. Taken together, our results indicate that SM21 has the potential to be developed as a novel antifungal agent for clinical use.

Planktonic growth and biofilm formation profiles in Candida haemulonii species complex.

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Ramos, Lívia S; Oliveira, Simone S C; Souto, Xênia M; Branquinha, Marta H; Santos, André L S

2017-10-01

Candida haemulonii species complex have emerged as multidrug-resistant yeasts able to cause fungemia worldwide. However, very little is known regarding their physiology and virulence factors. In this context, planktonic growth and biofilm formation of Brazilian clinical isolates of Candida haemulonii (n = 5), Candida duobushaemulonii (n = 4), and Candida haemulonii var. vulnera (n = 3) were reported. Overall, the fungal planktonic growth curves in Sabouraud dextrose broth reached the exponential phase in 48 h at 37°C. All the clinical isolates formed biofilm on polystyrene in a time-dependent event, as judged by the parameters evaluated: biomass (crystal violet staining), metabolic activity (XTT reduction), and extracellular matrix (safranin incorporation). No statistically significant differences were observed when the average measurements among the three Candida species were compared regarding both planktonic and biofilm lifestyles; however, typical isolate-specific differences were clearly noticed in fungal growth kinetics. © The Author 2017. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Candida tropicalis from veterinary and human sources shows similar in vitro hemolytic activity, antifungal biofilm susceptibility and pathogenesis against Caenorhabditis elegans.

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Brilhante, Raimunda Sâmia Nogueira; Oliveira, Jonathas Sales de; Evangelista, Antônio José de Jesus; Serpa, Rosana; Silva, Aline Lobão da; Aguiar, Felipe Rodrigues Magalhães de; Pereira, Vandbergue Santos; Castelo-Branco, Débora de Souza Collares Maia; Pereira-Neto, Waldemiro Aquino; Cordeiro, Rossana de Aguiar; Sidrim, José Júlio Costa; Rocha, Marcos Fábio Gadelha

2016-08-30

The aim of this study was to evaluate the in vitro hemolytic activity and biofilm antifungal susceptibility of veterinary and human Candida tropicalis strains, as well as their pathogenesis against Caenorhabditis elegans. Twenty veterinary isolates and 20 human clinical isolates of C. tropicalis were used. The strains were evaluated for their hemolytic activity and biofilm production. Biofilm susceptibility to itraconazole, fluconazole, voriconazole, amphotericin B and caspofungin was assessed using broth microdilution assay. The in vivo evaluation of strain pathogenicity was investigated using the nematode C. elegans. Hemolytic factor was observed in 95% of the strains and 97.5% of the isolates showed ability to form biofilm. Caspofungin and amphotericin B showed better results than azole antifungals against mature biofilms. Paradoxical effect on mature biofilm metabolic activity was observed at elevated concentrations of caspofungin (8-64μg/mL). Azole antifungals were not able to inhibit mature C. tropicalis biofilms, even at the higher tested concentrations. High mortality rates of C. elegans were observed when the worms were exposed to with C. tropicalis strains, reaching up to 96%, 96h after exposure of the worms to C. tropicalis strains. These results reinforce the high pathogenicity of C. tropicalis from veterinary and human sources and show the effectiveness of caspofungin and amphotericin B against mature biofilms of this species. Copyright © 2016 Elsevier B.V. All rights reserved.

Biofilm antifungal susceptibility of Candida urine isolated from ambulatory patients

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Débora da Luz Becker

2016-07-01

Full Text Available Background and Objectives: the association between the biofilm formations an antifungal resistance has been suggested to be an important factor in the pathogenesis of several Candida species. Besides, studies have included invasive candidiasis from hospitalized patients; however there are few studies that evaluated the species distribution, antifungal susceptibility and biofilm formation of Candida species isolated from ambulatory patients. Thus, the aim of this study was to evaluate whether biofilm producing contributes to antifungal resistance in Candida isolates from urine sample obtained from ambulatory patients. Methods: During one year, 25 urine samples positive for yeast were collected, stored and plated on agar supplemented with chloramphenicol and Sabouread left at room temperature for 5 days for subsequent: 52% (13/25 were C. albicans, 36% (9/25 C. tropicalis, 8% (2/25 C. krusei and 4% (1/25 C. parapsilosis. Results: The ability to form biofilm was detected in 23 (92% of the yeast studied and 15.4% (2/13 of C. albicans were fluconazole (FLU and ketoconazole (KET resistant, while 11.1% (1/9 of C. tropicalis were ketoconazole resistant and were anidulafungin (ANI non-susceptible. Conclusion: our results showed the high capacity for biofilm formation among Candida isolates from ambulatory patients.

The expression of genes involved in the ergosterol biosynthesis pathway in Candida albicans and Candida dubliniensis biofilms exposed to fluconazole.

LENUS (Irish Health Repository)

2009-03-01

The expression of the ERG1, ERG3, ERG7, ERG9, ERG11 and ERG25 genes in response to incubation with fluconazole and biofilm formation was investigated using reverse-transcription PCR and real-time PCR in Candida albicans and Candida dubliniensis clinical isolates. The viability of biofilm was measured using an 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) reduction assay and confocal scanning laser microscopy (CSLM). Expression of the ERG11 gene was found to be low or moderate and it was regulated by fluconazole addition more so than by biofilm formation. Very low or non-detectable expression of ERG1, ERG7 and ERG25 genes was detected in C. albicans. The expression of the ERG9 increased in the presence of fluconazole in some isolates. Following incubation with fluconazole, formation of biofilm by C. dubliniensis was coupled with up-regulation of the ERG3 and ERG25 genes as have been observed previously in C. albicans. Planktonic cells of both Candida species released from biofilm displayed similar resistance mechanisms to fluconazole like attached cells. The XTT reduction assay and CSLM revealed that although incubation with fluconazole decreased the biofilm thickness, these were still comprised metabolically active cells able to disseminate and produce biofilm. Our data indicate that biofilm represents a highly adapted community reflecting the individuality of clinical isolates.

Candida biofilms: is adhesion sexy?

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Soll, David R

2008-08-26

The development of Candida albicans biofilms requires two types of adhesion molecule - the Als proteins and Hwp1. Mutational analyses have recently revealed that these molecules play complementary roles, and their characteristics suggest that they may have evolved from primitive mating agglutinins.

Influence of artificial saliva in biofilm formation of Candida albicans in vitro

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Michelle Peneluppi Silva

2012-02-01

Full Text Available Due to the increase in life expectancy, new treatments have emerged which, although palliative, provide individuals with a better quality of life. Artificial saliva is a solution that contains substances that moisten a dry mouth, thus mimicking the role of saliva in lubricating the oral cavity and controlling the existing normal oral microbiota. This study aimed to assess the influence of commercially available artificial saliva on biofilm formation by Candida albicans. Artificial saliva I consists of carboxymethylcellulose, while artificial saliva II is composed of glucose oxidase, lactoferrin, lysozyme and lactoperoxidase. A control group used sterile distilled water. Microorganisms from the oral cavity were transferred to Sabouraud Dextrose Agar and incubated at 37°C for 24 hours. Colonies of Candida albicans were suspended in a sterile solution of NaCl 0.9%, and standardisation of the suspension to 106 cells/mL was achieved. The acrylic discs, immersed in artificial saliva and sterile distilled water, were placed in a 24-well plate containing 2 mL of Sabouraud Dextrose Broth plus 5% sucrose and 0.1 mL aliquot of the Candida albicans suspension. The plates were incubated at 37°C for 5 days, the discs were washed in 2 mL of 0.9% NaCl and placed into a tube containing 10 mL of 0.9% NaCl. After decimal dilutions, aliquots of 0.1 mL were seeded on Sabouraud Dextrose Agar and incubated at 37°C for 48 hours. Counts were reported as CFU/mL (Log10. A statistically significant reduction of 29.89% (1.45 CFU/mL of Candida albicans was observed in saliva I when compared to saliva II (p = 0.002, considering p≤0.05.

Interactions between Candida albicans and Candida glabrata in biofilms: Influence of the strain type, culture medium and glucose supplementation.

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Hosida, Thayse Yumi; Cavazana, Thamires Priscila; Henriques, Mariana; Pessan, Juliano Pelim; Delbem, Alberto Carlos Botazzo; Monteiro, Douglas Roberto

2018-04-01

The relationship among Candida species may be influenced by several factors. Thus, this study evaluated the interactions between Candida albicans and Candida glabrata in biofilms, varying the strain type, culture medium and glucose supplementation. Biofilms were formed for 48 hours in Sabouraud dextrose broth (SDB) or RPMI 1640, supplemented with 0%, 1% or 5% glucose. Each strain of C. albicans was combined with two strains of C. glabrata, generating four biofilm associations, which were quantified by colony-forming units (CFUs), total biomass and metabolic activity. Data were analysed by ANOVA and Tukey's HSD test (α = 0.05). For CFUs, all associations were classified as indifferent for biofilms formed in RPMI 1640, while for SDB the interactions were antagonistic for C. albicans and indifferent for C. glabrata. The association of reference strains resulted in a dual-species biofilm with biomass significantly higher than that observed for each single biofilm developed in SDB. The metabolic activity of dual-species biofilms did not significantly differ from that found for single ones, except for co-culture of the reference strains. Glucose supplementation and culture media had a significant influence on all parameters. In conclusion, the strain type, culture medium and glucose supplementation influenced the interactions between C. albicans and C. glabrata. © 2017 Blackwell Verlag GmbH.

Effect of Candida albicans and Candida dubliniensis planktonic/biofilm quorum sensing molecules on yeast morphogenesis

OpenAIRE

Henriques, Mariana; Martins, Margarida Isabel Barros Coelho; Azeredo, Joana; Oliveira, Rosário

2006-01-01

One of the aims of this work was to study the effect of farnesol, a quorum sensing molecule for Candida albicans, on morphologic inhibition of Candida dubliniensis. The second goal of this work was to confirm if Candida dubliniensis also excreted quorum sensing molecules, on both planktonic and biofilm forms. The results clearly demonstrate that Candida dubliniensis undergoes morphological alterations triggered by farnesol. It was also found that supernatants of Candida dubliniensis and Ca...

Candida species isolated from different body sites and their antifungal susceptibility pattern: Cross-analysis of Candida albicans and Candida glabrata biofilms.

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Cataldi, Valentina; Di Campli, Emanuela; Fazii, Paolo; Traini, Tonino; Cellini, Luigina; Di Giulio, Mara

2017-08-01

Candida species are regular commensal in humans, but-especially in immunocompromised patients-they represent opportunistic pathogens giving rise to systemic infection. The aim of the present work was to isolate and characterize for their antifungal profile Candida species from different body sites and to analyze the biofilms produced by C. albicans and C. glabrata isolates. Eighty-one strains of Candida species from 77 patients were identified. Epidemiological study showed that the most isolated species were C. albicans (44), C. glabrata (13) and C. parapsilosis (13) mainly from Hematology, Infectious Diseases, Medicine, Neonatology and Oncology Divisions, the majority of the biological samples were swabs (44) and blood cultures (16). The analysis of the biofilm formation was performed at 24 and 48-hours comparing resistant and susceptible strains of C. albicans to resistant and susceptible strains of C. glabrata. Candida albicans has a greater ability to form biofilm compared to C. glabrata, both in the susceptible and resistant strains reaching maturity after 24 hours with a complex structure composed of blastospores, pseudohyphae, and hyphae embedded in a matrix. On the contrary, C. glabrata biofilm was composed exclusively of blastospores that in the resistant strain, after 24 hours, were organized in a compact multilayer different to the discontinuous structure observed in the susceptible analyzed strains. In conclusion, the increasing of the incidence of Candida species infection together with their emerging drug resistance also related to the biofilm forming capability underline the need to monitor their distribution and susceptibility patterns for improving the surveillance and for a correct management of the infection. © The Author 2016. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Comparison of the Candida albicans and biofilm formation amount on natural tooth,

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Serdar Polat

2012-03-01

Full Text Available Objective: This study compared the retention of the Candidaalbicans and biofilm formation on natural teeth, porcelainand acrylic resin.Materials and methods: Samples are taken with thesterile ecuvion sticks from the buccal embrasures of thenatural tooth, porcelain and acrylic. The biofilm productionof candida reproducing strains was determined withmicroplate method. Samples are settled in 0.5 ml sterilephosphate buffered saline. Reproduction detected coloniesdefined to species in accordance with their macroscopicand microscopic features and germ tube test inmicrobiology laboratory.Results: There was significant differences for retention ofCandida albicans and biofilm formation on the surface oftooth, porcelain and acrylic (p<0.05.Conclusion: Adherence of Candida albicans and biofilmformation on the porcelain significantly less than naturaltooth and acrylic, and retention and biofilm formation onthe tooth less than acrylic.

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

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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. © The Author 2014. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Silver colloidal nanoparticles: antifungal effect against adhered cells and biofilms of Candida albicans and Candida glabrata.

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Monteiro, D R; Gorup, L F; Silva, S; Negri, M; de Camargo, E R; Oliveira, R; Barbosa, D B; Henriques, M

2011-08-01

The aim of this study was to evaluate the effect of silver nanoparticles (SN) against Candida albicans and Candida glabrata adhered cells and biofilms. SN (average diameter 5 nm) were synthesized by silver nitrate reduction with sodium citrate and stabilized with ammonia. Minimal inhibitory concentration (MIC) tests were performed for C. albicans (n = 2) and C. glabrata (n = 2) grown in suspension following the Clinical Laboratory Standards Institute microbroth dilution method. SN were applied to adhered cells (2 h) or biofilms (48 h) and after 24 h of contact their effect was assessed by enumeration of colony forming units (CFUs) and quantification of total biomass (by crystal violet staining). The MIC results showed that SN were fungicidal against all strains tested at very low concentrations (0.4-3.3 μg ml(-1)). Furthermore, SN were more effective in reducing biofilm biomass when applied to adhered cells (2 h) than to pre-formed biofilms (48 h), with the exception of C. glabrata ATCC, which in both cases showed a reduction ∼90%. Regarding cell viability, SN were highly effective on adhered C. glabrata and respective biofilms. On C. albicans the effect was not so evident but there was also a reduction in the number of viable biofilm cells. In summary, SN may have the potential to be an effective alternative to conventional antifungal agents for future therapies in Candida-associated denture stomatitis.

[Fungal infectivities of implanted catheters due to Candida sp. Biofilms formation and resistance].

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Seddiki, S M L; Boucherit-Otmani, Z; Boucherit, K; Kunkel, D

2015-06-01

Candidemia are the most common fungal infections in hospitals. However, the catheters are subject to be altered by Candida biofilms which increase the risk of invasive nosocomial infections due to the high resistance to antifungal agents. Therefore, the minimum inhibitory concentrations of planktonic (MIC) and sessile cells (CIMS) were evaluated. To review the in vivo biofilms structures of Candida sp. formed on the inner and/or external surfaces of collected catheters, we used scanning electron microscopy (SEM). The level of biofilm resistance was assessed against two conventional antifungal agents: amphotericin B (AmB), which belongs to the class of polyenes, and fluconazole (FLZ) which is an azole. The SEM observation of biofilms of Candida sp. reveals complex structures. Compared to MICs, the calculation of CIMS showed an increase of 32 times with AmB and of 128 times with FLZ. Catheters offer an ideal surface to Candida sp. to form biofilms. This complex structure induces the increase of the resistance of sessile cells against two antifungal agents, AmB and FLZ. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Conservation and Divergence in the Candida Species Biofilm Matrix Mannan-Glucan Complex Structure, Function, and Genetic Control.

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Dominguez, Eddie; Zarnowski, Robert; Sanchez, Hiram; Covelli, Antonio S; Westler, William M; Azadi, Parastoo; Nett, Jeniel; Mitchell, Aaron P; Andes, David R

2018-04-03

Candida biofilms resist the effects of available antifungal therapies. Prior studies with Candida albicans biofilms show that an extracellular matrix mannan-glucan complex (MGCx) contributes to antifungal sequestration, leading to drug resistance. Here we implement biochemical, pharmacological, and genetic approaches to explore a similar mechanism of resistance for the three most common clinically encountered non- albicans Candida species (NAC). Our findings reveal that each Candida species biofilm synthesizes a mannan-glucan complex and that the antifungal-protective function of this complex is conserved. Structural similarities extended primarily to the polysaccharide backbone (α-1,6-mannan and β-1,6-glucan). Surprisingly, biochemical analysis uncovered stark differences in the branching side chains of the MGCx among the species. Consistent with the structural analysis, similarities in the genetic control of MGCx production for each Candida species also appeared limited to the synthesis of the polysaccharide backbone. Each species appears to employ a unique subset of modification enzymes for MGCx synthesis, likely accounting for the observed side chain diversity. Our results argue for the conservation of matrix function among Candida spp. While biogenesis is preserved at the level of the mannan-glucan complex backbone, divergence emerges for construction of branching side chains. Thus, the MGCx backbone represents an ideal drug target for effective pan- Candida species biofilm therapy. IMPORTANCE Candida species, the most common fungal pathogens, frequently grow as a biofilm. These adherent communities tolerate extremely high concentrations of antifungal agents, due in large part, to a protective extracellular matrix. The present studies define the structural, functional, and genetic similarities and differences in the biofilm matrix from the four most common Candida species. Each species synthesizes an extracellular mannan-glucan complex (MGCx) which

Effect of ferrocene-substituted porphyrin RL-91 on Candida albicans biofilm formation.

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Lippert, Rainer; Vojnovic, Sandra; Mitrovic, Aleksandra; Jux, Norbert; Ivanović-Burmazović, Ivana; Vasiljevic, Branka; Stankovic, Nada

2014-08-01

Ferrocene-substituted porphyrin RL-91 exhibits antifungal activity against opportune human pathogen Candida albicans. RL-91 efficiently inhibits growth of both planktonic C. albicans cells and cells within biofilms without photoactivation. The minimal inhibitory concentration for plankton form (PMIC) was established to be 100 μg/mL and the same concentration killed 80% of sessile cells in the mature biofilm (SMIC80). Furthermore PMIC of RL-91 efficiently prevents C. albicans biofilm formation. RL-91 is cytotoxic for human fibroblasts in vitro in concentration of 10 μg/mL, however it does not cause hemolysis in concentrations of up to 50 μg/mL. These findings open possibility for application of RL-91 as an antifungal agent for external antibiofilm treatment of medical devices as well as a scaffold for further development of porphyrin based systemic antifungals. Copyright © 2014 Elsevier Ltd. All rights reserved.

An In Vitro Model for Candida albicans–Streptococcus gordonii Biofilms on Titanium Surfaces

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Daniel Montelongo-Jauregui

2018-06-01

Full Text Available The oral cavity serves as a nutrient-rich haven for over 600 species of microorganisms. Although many are essential to maintaining the oral microbiota, some can cause oral infections such as caries, periodontitis, mucositis, and endodontic infections, and this is further exacerbated with dental implants. Most of these infections are mixed species in nature and associated with a biofilm mode of growth. Here, after optimization of different parameters including cell density, growth media, and incubation conditions, we have developed an in vitro model of C. albicans–S. gordonii mixed-species biofilms on titanium discs that is relevant to infections of peri-implant diseases. Our results indicate a synergistic effect for the development of biofilms when both microorganisms were seeded together, confirming the existence of beneficial, mutualistic cross-kingdom interactions for biofilm formation. The morphological and architectural features of these dual-species biofilms formed on titanium were determined using scanning electron microscopy (SEM and confocal laser scanning microscopy (CLSM. Mixed biofilms formed on titanium discs showed a high level of resistance to combination therapy with antifungal and antibacterial drugs. This model can serve as a platform for further analyses of complex fungal/bacterial biofilms and can also be applied to screening of new drug candidates against mixed-species biofilms.

The Candida albicans Biofilm Matrix: Composition, Structure and Function.

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Pierce, Christopher G; Vila, Taissa; Romo, Jesus A; Montelongo-Jauregui, Daniel; Wall, Gina; Ramasubramanian, Anand; Lopez-Ribot, Jose L

2017-03-01

A majority of infections caused by Candida albicans -the most frequent fungal pathogen-are associated with biofilm formation. A salient feature of C. albicans biofilms is the presence of the biofilm matrix. This matrix is composed of exopolymeric materials secreted by sessile cells within the biofilm, in which all classes of macromolecules are represented, and provides protection against environmental challenges. In this review, we summarize the knowledge accumulated during the last two decades on the composition, structure, and function of the C. albicans biofilm matrix. Knowledge of the matrix components, its structure, and function will help pave the way to novel strategies to combat C. albicans biofilm infections.

Inhibitory effects of Lactobacillus rhamnosus and Lactobacillus casei on Candida biofilm of denture surface.

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Song, Young-Gyun; Lee, Sung-Hoon

2017-04-01

Candida albicans biofilm is associated with denture-related stomatitis and oral candidiasis of elderly. Probiotics are beneficial bacteria and have antibacterial activity against pathogenic bacteria. The purpose of this study was to investigate the antifungal activity of various probiotics against C. albicans and the inhibitory effects of probiotics on Candida biofilm on the denture surface. The spent culture media of various probiotics were investigated the antifungal efficacy against C. albicans. Candida biofilm was formed on a denture base resin and was then treated with Lactobacillus rhamnosus and Lactobacillus casei. Also, the biofilms of L. rhamnosus and L. casei were formed and were sequentially treated with C. albicans. Colony-forming units of C. albicans on the denture surface were counted after spreading on agar plate. The denture base resin was treated with the spent culture media for 30days, after which the denture surface roughness was analyzed with an atomic force microscope. L. rhamnosus and L. casei exhibited stronger antifungal activity than other probiotics. The spent culture medium of L. rhamnosus and L. casei exhibited the antifungal activity against blastoconidia and biofilm of C. albicans. L. rhamnosus and L. casei showed the antifungal activity against Candida biofilm, and the biofilm of L. rhamnosus and L. casei inhibited formation of Candida biofilm on denture surface. Neither of the probiotics affected the surface roughness of the denture base resin. L. rhamnosus and L. casei may be the ideal probiotics for the prevention and treatment of denture-related stomatitis. Copyright © 2017 Elsevier Ltd. All rights reserved.

Candida species from oral cavity of HIV-infected children exhibit reduced virulence factors in the HAART era.

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Portela, Maristela Barbosa; Lima de Amorim, Elaine; Santos, Adrielle Mangabeira; Alexandre da Rocha Curvelo, José; de Oliveira Martins, Karol; Capillé, Cauli Lima; Maria de Araújo Soares, Rosangela; Barbosa de Araújo Castro, Gloria Fernanda

2017-01-01

This study aimed to assess, in vitro, the biofilm viability and the phospholipase and protease production of Candida spp. from the saliva of HIV infected children and healthy controls, and to correlate the results with the use of medical data. A total of 79 isolates were analyzed: 48 Candida albicans isolates (33/15) and 20 Candida parapsilosis sensu lato complex isolates (12/8) (from HIV/control patients, respectively), and 8 Candida krusei, 1 Candida tropicalis, 1 Candida dubliniensis and 1 Candida guilliermondii from HIV patients. The XTT (2, 3-bis (2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-Carboxanilide) reduction assay analyzed the biofilm viability. Phospholipase and protease assays were performed using the egg yolk and Bovine Serum Albumin agar plate methods, respectively. All isolates were able to form biofilm with cell viability. Quantitatively, Candida isolates from both groups presented a similar ability to form biofilm (p > 0.05). The biofilm viability activity was higher in C. albicans isolates than in non-albicans Candida isolates (p Candida spp. isolates from HIV-positive children presented higher phospholipase production, in vitro they exhibited reduced virulence factors compared to isolates from healthy individuals. This finding may enlighten the role played by immunosuppression in the modulation of Candida virulence attributes. Copyright © 2016 Elsevier Ltd. All rights reserved.

Evaluation of CAMP-Like Effect, Biofilm Formation, and Discrimination of Candida africana from Vaginal Candida albicans Species

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Keyvan Pakshir

2017-01-01

Full Text Available Candida africana as a species recovered from female genital specimens is highly close to C. albicans. The present study was conducted to discriminate C. africana from presumptive vaginal C. albicans strains by molecular assay and evaluate their hemolysin activity, biofilm formation, and cohemolytic effect (CAMP with vaginal bacterial flora. A total of 110 stock vaginal C. albicans isolates were examined by HWP1 gene amplification. Hemolysin activity and the ability of biofilm formation were evaluated by blood plate assay and visual detection methods, respectively. Staphylococcus aureus, Staphylococcus epidermidis, and Streptococcus agalactiae were used to evaluate the CAMP-like effects in Sabouraud blood agar media. Based on the size of the amplicons (941 bp, all isolates were identified as C. albicans. All samples were able to produce beta-hemolysin. Moreover, 69 out of 110 of the isolates (62.7% were biofilm-positive, 54 out of 110 Candida isolates (49% demonstrated cohemolytic effects with S. agalactiae, and 48 out of 110 showed this effect with S. aureus (43.6%. All isolates were CAMP-negative with S. epidermidis. We detected all isolates as Candida albicans and almost half of the isolates were CAMP-positive with S. aureus and S. agalactiae, suggesting that these bacteria increase the pathogenicity of Candida in vaginal candidiasis.

Evaluation of CAMP-Like Effect, Biofilm Formation, and Discrimination of Candida africana from Vaginal Candida albicans Species

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Bordbar, Mahboubeh; Nouraei, Hasti; Khodadadi, Hossein

2017-01-01

Candida africana as a species recovered from female genital specimens is highly close to C. albicans. The present study was conducted to discriminate C. africana from presumptive vaginal C. albicans strains by molecular assay and evaluate their hemolysin activity, biofilm formation, and cohemolytic effect (CAMP) with vaginal bacterial flora. A total of 110 stock vaginal C. albicans isolates were examined by HWP1 gene amplification. Hemolysin activity and the ability of biofilm formation were evaluated by blood plate assay and visual detection methods, respectively. Staphylococcus aureus, Staphylococcus epidermidis, and Streptococcus agalactiae were used to evaluate the CAMP-like effects in Sabouraud blood agar media. Based on the size of the amplicons (941 bp), all isolates were identified as C. albicans. All samples were able to produce beta-hemolysin. Moreover, 69 out of 110 of the isolates (62.7%) were biofilm-positive, 54 out of 110 Candida isolates (49%) demonstrated cohemolytic effects with S. agalactiae, and 48 out of 110 showed this effect with S. aureus (43.6%). All isolates were CAMP-negative with S. epidermidis. We detected all isolates as Candida albicans and almost half of the isolates were CAMP-positive with S. aureus and S. agalactiae, suggesting that these bacteria increase the pathogenicity of Candida in vaginal candidiasis. PMID:29318048

Anti-Candida activity assessment of Pelargonium graveolens oil free and nanoemulsion in biofilm formation in hospital medical supplies.

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Giongo, Janice Luehring; de Almeida Vaucher, Rodrigo; Fausto, Viviane Pedroso; Quatrin, Priscilla Maciel; Lopes, Leonardo Quintana Soares; Santos, Roberto Christ Vianna; Gündel, André; Gomes, Patrícia; Steppe, Martin

2016-11-01

Infections due to microbial biofilm formation on the surface of catheters and other medical devices are constantly reported as a major cause of morbidity and mortality in patients admitted to hospitals. Furthermore, sessile cells are more resistant to phagocytosis and most antimicrobial, which complicates the treatment of such infections. Researches aimed at new antimicrobial originating mainly from plants have increased in recent years and the development of new strategies for their release is critical in combating the formation of biofilms. Geranium oil (GO) has proven antimicrobial activity. Because of this, the aim of this study was to develop nanoemulsions containing this oil (NEG) and evaluate its activity after the biofilm formation of Candida albicans, Candida tropicalis, Candida glabrata, and Candida krusei in hospital medical supplies. For quantification of the biofilm, crystal violet, total protein, and ATP-bioluminescence assays were used. The results revealed that GO and NEG showed lower MIC for C. albicans and C. tropicalis. The biofilms formed by different species of Candida on the surfaces of polyethylene and polyurethane were quantified. GO and NEG significantly inhibited the formation of biofilms in all species tested on the surfaces of polyethylene. However, NEG antibiofilm has had better activity than GO for C. albicans, C. tropicalis and C. glabrata, according to the surface potential analysis by atomic force microscopy (AFM). The analysis of the biofilm formation on the polyethylene surface by ATP-bioluminescence and CFU showed similar results. In both methods the formation of biofilm in the catheter occurred in greater quantity for C. albicans and C. tropicalis. GO did not significantly inhibit the formation of biofilms only in C. krusei, although NEG significantly increased this activity GO in all species tested when compared to the control training biofilm. The following study shows that the development of NEG may become an effective

Minocycline Inhibits Candida albicans Budded-to-Hyphal-Form Transition and Biofilm Formation.

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Kurakado, Sanae; Takatori, Kazuhiko; Sugita, Takashi

2017-09-25

Candida albicans frequently causes bloodstream infections; its budded-to-hyphalform transition (BHT) and biofilm formation are major contributors to virulence. During an analysis of antibacterial compounds that inhibit C. albicans BHT, we found that the tetracycline derivative minocycline inhibited BHT and subsequent biofilm formation. Minocycline decreased expression of hypha-specific genes HWP1 and ECE1, and adhesion factor gene ALS3 of C. albicans. In addition, minocycline decreased cell surface hydrophobicity and the extracellular β-glucan level in biofilms. Minocycline has been widely used for catheter antibiotic lock therapy to prevent bacterial infection; this compound may also be prophylactically effective against Candida infection.

Lipidomics of Candida albicans biofilms reveals phase-dependent production of phospholipid molecular classes and role for lipid rafts in biofilm formation.

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Lattif, Ali Abdul; Mukherjee, Pranab K; Chandra, Jyotsna; Roth, Mary R; Welti, Ruth; Rouabhia, Mahmoud; Ghannoum, Mahmoud A

2011-11-01

Candida albicans-associated bloodstream infections are linked to the ability of this yeast to form biofilms. In this study, we used lipidomics to compare the lipid profiles of C. albicans biofilms and planktonic cells, in early and mature developmental phases. Our results showed that significant differences exist in lipid composition in both developmental phases. Biofilms contained higher levels of phospholipid and sphingolipids than planktonic cells (nmol per g biomass, Pbiofilms compared to planktonic cells (P≤0.05). The ratio of phosphatidylcholine to phosphatidylethanolamine was lower in biofilms compared to planktonic cells in both early (1.17 vs 2.52, P≤0.001) and late (2.34 vs 3.81, P≤0.001) developmental phases. The unsaturation index of phospholipids decreased with time, with this effect being particularly strong for biofilms. Inhibition of the biosynthetic pathway for sphingolipid [mannosyl diinositolphosphoryl ceramide, M(IP)₂C] by myriocin or aureobasidin A, and disruption of the gene encoding inositolphosphotransferase (Ipt1p), abrogated the ability of C. albicans to form biofilms. The differences in lipid profiles between biofilms and planktonic Candida cells may have important implications for the biology and antifungal resistance of biofilms.

Biofilm development by blastospores and hyphae of Candida albicans on abraded denture acrylic resin surfaces.

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Jackson, Sarah; Coulthwaite, Lisa; Loewy, Zvi; Scallan, Anthony; Verran, Joanna

2014-10-01

Candida albicans is a known etiologic agent of denture stomatitis. Candida hyphae exhibit the ability to respond directionally to environmental stimuli. This characteristic is thought to be important in the penetration of substrata such as resilient denture liners and host epithelium. It has been suggested that hyphal production also enhances adhesion and survival of Candida on host and denture surfaces. Surface roughness, in addition, can enhance adhesion where stronger interactions occur between cells and surface features of similar dimensions. The purpose of this study was to assess the development of hyphal and blastospore biofilms on abraded denture acrylic resin specimens and measure the ease of removal of these biofilms. Biofilms were grown for 48 hours on abraded 1-cm² denture acrylic resin specimens from adhered hyphal phase C albicans or from adhered blastospores. Subsequently, all specimens were stained with Calcofluor White and examined with confocal scanning laser microscopy. Biofilms were removed by vortex mixing in sterile phosphate buffered saline solution. Removed cells were filtered (0.2-μm pore size). Filters were dried at 37°C for 24 hours for dry weight measurements. Any cells that remained on the acrylic resin specimens were stained with 0.03% acridine orange and examined with epifluorescence microscopy. Biofilms grown from both cell types contained all morphologic forms of C albicans. Although the underlying surface topography did not affect the amount of biofilm produced, biofilms grown from hyphal phase Candida were visibly thicker and had greater biomass (Phyphae in early Candida biofilms increased biofilm mass and resistance to removal. Increased surface roughness enhances retention of hyphae and yeast cells, and, therefore, will facilitate plaque regrowth. Therefore, minimization of denture abrasion during cleaning is desirable. Copyright © 2014 Editorial Council for the Journal of Prosthetic Dentistry. Published by Elsevier Inc. All

Long-term efficacy of denture cleansers in preventing Candida spp. biofilm recolonization on liner surface

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Ana Paula Coelho Vieira

2010-09-01

Full Text Available This study evaluated the long-term efficacy of denture cleansers against Candida spp. biofilm recolonization on liner surface. Specimens were fabricated of a poly(methyl methacrylate-based denture liner and had their surface roughness evaluated at baseline and after cleansing treatments. C. albicans or C. glabrata biofilms were formed on liner surface for 48 h, and then the specimens were randomly assigned to one of cleaning treatments: two alkaline peroxides (soaking for 3 or 15 min, 0.5% sodium hypochlorite (10 min or distilled water (control; 15 min. After the treatments, the specimens were sonicated to disrupt the biofilm, and residual cells were counted (cell/mL. Long-term effectiveness of the cleaning processes was determined by submitting a set of cleaned specimens to biofilm growth conditions for 48 h followed by estimation of cell counts. The topography of specimens after cleaning treatments was analyzed by SEM. Data were analyzed by ANOVA and Tukey's test (α; = 0.05. Results of cell count estimation showed significant differences in cleanliness among the treatments (p 0.05 was observed among the Candida species regarding the recolonization condition. Alkaline denture cleansers showed similar cleaning performance and both differed from the control (p < 0.001. Sodium hypochlorite was the only treatment that removed biofilm efficiently, since no viable cells were found after its use. In conclusion, alkaline peroxide denture cleansers were not effective in removing Candida spp. biofilm from denture liner surfaces and preventing biofilm recolonization.

Streptococcus mutans Can Modulate Biofilm Formation and Attenuate the Virulence of Candida albicans

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Barbosa, Júnia Oliveira; Rossoni, Rodnei Dennis; Vilela, Simone Furgeri Godinho; de Alvarenga, Janaína Araújo; Velloso, Marisol dos Santos; Prata, Márcia Cristina de Azevedo; Jorge, Antonio Olavo Cardoso; Junqueira, Juliana Campos

2016-01-01

Streptococcus mutans and Candida albicans are found together in the oral biofilms on dental surfaces, but little is known about the ecological interactions between these species. Here, we studied the effects of S. mutans UA159 on the growth and pathogencity of C. albicans. Initially, the effects of S. mutans on the biofilm formation and morphogenesis of C. albicans were tested in vitro. Next, we investigate the influence of S. mutans on pathogenicity of C. albicans using in vivo host models, in which the experimental candidiasis was induced in G. mellonella larvae and analyzed by survival curves, C. albicans count in hemolymph, and quantification of hyphae in the host tissues. In all the tests, we evaluated the direct effects of S. mutans cells, as well as the indirect effects of the subproducts secreted by this microorganism using a bacterial culture filtrate. The in vitro analysis showed that S. mutans cells favored biofilm formation by C. albicans. However, a reduction in biofilm viable cells and inhibition of hyphal growth was observed when C. albicans was in contact with the S. mutans culture filtrate. In the in vivo study, injection of S. mutans cells or S. mutans culture filtrate into G. mellonella larvae infected with C. albicans increased the survival of these animals. Furthermore, a reduction in hyphal formation was observed in larval tissues when C. albicans was associated with S. mutans culture filtrate. These findings suggest that S. mutans can secrete subproducts capable to inhibit the biofilm formation, morphogenesis and pathogenicity of C. albicans, attenuating the experimental candidiasis in G. mellonella model. PMID:26934196

RNA-Seq Reveals Enhanced Sugar Metabolism in Streptococcus mutans Co-cultured with Candida albicans within Mixed-Species Biofilms

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He, Jinzhi; Kim, Dongyeop; Zhou, Xuedong; Ahn, Sang-Joon; Burne, Robert A.; Richards, Vincent P.; Koo, Hyun

2017-01-01

Early childhood caries (ECC), which can lead to rampant tooth-decay that is painful and costly to treat, is one of the most prevalent infectious diseases affecting children worldwide. Previous studies support that interactions between Streptococcus mutans and Candida albicans are associated with the pathogenesis of ECC. The presence of Candida enhances S. mutans growth, fitness and accumulation within biofilms in vitro, although the molecular basis for these behaviors is undefined. Using an established co-cultivation biofilm model and RNA-Seq, we investigated how C. albicans influences the transcriptome of S. mutans. The presence of C. albicans dramatically altered gene expression in S. mutans in the dual-species biofilm, resulting in 393 genes differentially expressed, compared to mono-species biofilms of S. mutans. By Gene Ontology analysis, the majority of up-regulated genes were related to carbohydrate transport and metabolic/catabolic processes. KEGG pathway impact analysis showed elevated pyruvate and galactose metabolism, suggesting that co-cultivation with C. albicans influences carbohydrate utilization by S. mutans. Analysis of metabolites confirmed the increases in carbohydrate metabolism, with elevated amounts of formate in the culture medium of co-cultured biofilms. Moreover, co-cultivation with C. albicans altered transcription of S. mutans signal transduction (comC and ciaRH) genes associated with fitness and virulence. Interestingly, the expression of genes for mutacins (bacteriocins) and CRISPR were down-regulated. Collectively, the data provide a comprehensive insight into S. mutans transcriptomic changes induced by C. albicans, and offer novel insights into how bacterial–fungal interactions may enhance the severity of dental caries. PMID:28642749

RNA-Seq Reveals Enhanced Sugar Metabolism in Streptococcus mutans Co-cultured with Candida albicans within Mixed-Species Biofilms

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Jinzhi He

2017-06-01

Full Text Available Early childhood caries (ECC, which can lead to rampant tooth-decay that is painful and costly to treat, is one of the most prevalent infectious diseases affecting children worldwide. Previous studies support that interactions between Streptococcus mutans and Candida albicans are associated with the pathogenesis of ECC. The presence of Candida enhances S. mutans growth, fitness and accumulation within biofilms in vitro, although the molecular basis for these behaviors is undefined. Using an established co-cultivation biofilm model and RNA-Seq, we investigated how C. albicans influences the transcriptome of S. mutans. The presence of C. albicans dramatically altered gene expression in S. mutans in the dual-species biofilm, resulting in 393 genes differentially expressed, compared to mono-species biofilms of S. mutans. By Gene Ontology analysis, the majority of up-regulated genes were related to carbohydrate transport and metabolic/catabolic processes. KEGG pathway impact analysis showed elevated pyruvate and galactose metabolism, suggesting that co-cultivation with C. albicans influences carbohydrate utilization by S. mutans. Analysis of metabolites confirmed the increases in carbohydrate metabolism, with elevated amounts of formate in the culture medium of co-cultured biofilms. Moreover, co-cultivation with C. albicans altered transcription of S. mutans signal transduction (comC and ciaRH genes associated with fitness and virulence. Interestingly, the expression of genes for mutacins (bacteriocins and CRISPR were down-regulated. Collectively, the data provide a comprehensive insight into S. mutans transcriptomic changes induced by C. albicans, and offer novel insights into how bacterial–fungal interactions may enhance the severity of dental caries.

The differences in the isoelectric points of biofilm-positive and biofilm-negative Candida parapsilosis strains.

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Ruzicka, Filip; Horka, Marie; Hola, Veronika; Kubesova, Anna; Pavlik, Tomas; Votava, Miroslav

2010-03-01

The isoelectric points of 39 Candida parapsilosis strains were determined by means of capillary isoelectric focusing. The value of the isoelectric point corresponded well with cell surface hydrophobicity, as well as with the ability to form biofilm in these yeasts. Copyright 2010 Elsevier B.V. All rights reserved.

Molecular characterization of Candida in the oral cavity and factors involved in biofilm formation and virulence

NARCIS (Netherlands)

Kraneveld, E.A.

2014-01-01

The research described in this thesis addresses current issues related to oral Candida infections. Interactions of Candida with the oral microbiome were characterized and factors involved in biofilm formation and virulence were studied. All in all, the work described in this thesis contributes

Inactivation of Candida biofilms by non-thermal plasma and its enhancement for fungistatic effect of antifungal drugs.

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Yi Sun

Full Text Available We investigated the antifungal effect of non-thermal plasma, as well as its combination with common antifungal drugs, against Candida biofilms. A direct current atmospheric pressure He/O(2 (2% plasma microjet (PMJ was used to treat Candida biofilms in a 96-well plate. Inactivation efficacies of the biofilms were evaluated by XTT assay and counting colony forming units (CFUs. Morphological properties of the biofilms were evaluated by Scanning Electron Microscope (SEM. The sessile minimal inhibitory concentrations (SMICs of fluconazole, amphotericin B, and caspofungin for the biofilms were also tested. Electron Spin Resonance (ESR spectroscopy was used to detect the reactive oxygen species (ROS generated directly and indirectly by PMJ. The Candida biofilms were completely inactivated after 1 min PMJ treatment, where severely deformed fungal elements were observed in SEM images. The SMICs of the tested antifungal drugs for the plasma-treated biofilms were decreased by 2-6 folds of dilution, compared to those of the untreated controls. ROS such as hydroxyl radical ((•OH, superoxide anion radical ((•O(2 (- and singlet molecular oxygen ((1O(2 were detected by ESR. We hence conclude that He/O(2 (2% plasma alone, as well as in combination with common antifungal drugs, is able to inactivate Candida biofilms rapidly. The generation of ROS is believed to be one of the underlying mechanisms for the fungicidal activity of plasma.

Candida parapsilosis Biofilm Identification by Raman Spectroscopy

Czech Academy of Sciences Publication Activity Database

Samek, Ota; Mlynariková, K.; Bernatová, Silvie; Ježek, Jan; Krzyžánek, Vladislav; Šiler, Martin; Zemánek, Pavel; Růžička, F.; Holá, Miroslava; Mahelová, M.

2014-01-01

Roč. 15, č. 12 (2014), s. 23924-23935 E-ISSN 1422-0067 R&D Projects: GA MŠk(CZ) LO1212; GA MŠk ED0017/01/01; GA ČR GAP205/11/1687 Institutional support: RVO:68081731 Keywords : Raman spectroscopy * Candida parapsilosis * biofilm Subject RIV: BH - Optics, Masers, Lasers Impact factor: 2.862, year: 2014

Levorotatory carbohydrates and xylitol subdue Streptococcus mutans and Candida albicans adhesion and biofilm formation.

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Brambilla, Eugenio; Ionescu, Andrei C; Cazzaniga, Gloria; Ottobelli, Marco; Samaranayake, Lakshman P

2016-05-01

Dietary carbohydrates and polyols affect the microbial colonization of oral surfaces by modulating adhesion and biofilm formation. The aim of this study was to evaluate the influence of a select group of l-carbohydrates and polyols on either Streptococcus mutans or Candida albicans adhesion and biofilm formation in vitro. S. mutans or C. albicans suspensions were inoculated on polystyrene substrata in the presence of Tryptic soy broth containing 5% of the following compounds: d-glucose, d-mannose, l-glucose, l-mannose, d- and l-glucose (raceme), d- and l-mannose (raceme), l-glucose and l-mannose, sorbitol, mannitol, and xylitol. Microbial adhesion (2 h) and biofilm formation (24 h) were evaluated using MTT-test and Scanning Electron Microscopy (SEM). Xylitol and l-carbohydrates induced the lowest adhesion and biofilm formation in both the tested species, while sorbitol and mannitol did not promote C. albicans biofilm formation. Higher adhesion and biofilm formation was noted in both organisms in the presence of d-carbohydrates relative to their l-carbohydrate counterparts. These results elucidate, hitherto undescribed, interactions of the individually tested strains with l- and d-carbohydrates, and how they impact fungal and bacterial colonization. In translational terms, our data raise the possibility of using l-form of carbohydrates and xylitol for dietary control of oral plaque biofilms. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Candida albicans Biofilms Do Not Trigger Reactive Oxygen Species and Evade Neutrophil Killing

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Xie, Zhihong; Thompson, Angela; Sobue, Takanori; Kashleva, Helena; Xu, Hongbin; Vasilakos, John; Dongari-Bagtzoglou, Anna

2012-01-01

Neutrophils are found within Candida albicans biofilms in vivo and could play a crucial role in clearing the pathogen from biofilms forming on catheters and mucosal surfaces. Our goal was to compare the antimicrobial activity of neutrophils against developing and mature C. albicans biofilms and identify biofilm-specific properties mediating resistance to immune cells. Antibiofilm activity was measured with the 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)2H-tetrazolium-5-carboxanilide assay and a molecular Candida viability assay. Reactive oxygen species generation was assessed by measuring fluorescence of 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate, acetyl ester in preloaded neutrophils. We found that mature biofilms were resistant to leukocytic killing and did not trigger reactive oxygen species, even though neutrophils retained their viability and functional activation potential. Beta-glucans found in the extracellular matrix negatively affected antibiofilm activities. We conclude that these polymers act as a decoy mechanism to prevent neutrophil activation and that this represents an important innate immune evasion mechanism of C. albicans biofilms. PMID:23033146

In Vitro Study on the Adhesion and Colonization of Candida Albicans on Metal and Acrylic Piercings

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Stamenov N.

2016-03-01

Full Text Available Oral/perioral piercing may provide an ideal environment for adhesion and colonization of microorganisms. The aim of this study is to perform an “in vitro” research on the capabilities of adhesion of Candida albicans on oral piercings made of plastic and metal. Acrylic and metal piercings were incubated with Candida albicans and then were observed using scanning electron microscopy under different magnifications. A lot of irregularities and roughness were observed on the surface of the plastic piercing unlike the surface of the metal one, which is not so rough. Nevertheless, the number of Candida albicans colonies was considerably larger on the scanned metal surface in comparison to the plastic surface. In vitro the metal surface of the piercing creates better environment for the adhesion and colonization of microorganisms than the acrylic. This could be attributed to the electrostatic forces that most likely attract Candida albicans to the metal piercing in the early stages of biofilm formation.

[FUNCTION OF INTERCELLULAR ADHESION A, FIBRINOGEN BINDING PROTEIN, AND ACCUMULATION-ASSOCIATED PROTEIN GENES IN FORMATION OF STAPHYLOCOCCUS EPIDERMIDIS-CANDIDA ALBICANS MIXED SPECIES BIOFILMS].

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Wang, Xiaoyan; Chen, Ying; Huang, Yunchao; Zhou, Youquan; Zhao, Guangqiang; Ye, Lianhua; Lei, Yujie; Tang, Qi

2015-01-01

To explore the function of intercellular adhesion A (icaA), fibrinogen binding protein (fbe), and accumulation-associated protein (aap) genes in formation of Staphylococcus epidermidis-Candida albicans mixed species biofilms. The experiment was divided into 3 groups: single culture of Staphylococcus epidermidis ATCC35984 (S. epidermidis group) or Candida albicans ATCC10231 (C. albicans group), and co-culture of two strains (mixed group) to build in vitro biofilm model. Biofilm mass was detected by crystal violet semi-quantitative adherence assay at 2, 4, 6, 8, 12, 24, 48, and 72 hours after incubation. XTT assay was performed to determine the growth kinetics in the same time. Scanning electron microscopy (SEM) was used to observe the ultrastructure of the biofilms after 24 and 72 hours of incubation. The expressions of icaA, fbe, and aap genes were analyzed by real-time fluorescent quantitative PCR. Crystal violet semi-quantitative adherence assay showed that the biofilms thickened at 12 hours in the S. epidermidis and mixed groups; after co-cultured for 72 hours the thickness of biofilm in mixed group was more than that in the S. epidermidis group, and there was significant difference between 2 groups at the other time (P 0.05). In C. albicans group, the biofilm started to grow at 12 hours of cultivation, but the thickness of the biofilm was significantly lower than that in the mixed group in all the time points (P 0.05) except at 12 hours (P 0.05); the A value of mixed group was significantly higher than that of the C. albicans group after 6 hours (P biofilms with complex structure formed in all groups. The real-time fluorescent quantitative PCR showed the expressions of fbe, icaA, and aap genes in mixed group increased 1.93, 1.52, and 1.46 times respectively at 72 hours compared with the S. epidermidis group (P biofilms have more complex structure and are thicker than single species biofilms of Staphylococcus epidermidis or Candida albicans, which is related to

Anti-Candida albicans biofilm effect of novel heterocyclic compounds.

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Kagan, Sarah; Jabbour, Adel; Sionov, Edward; Alquntar, Abed A; Steinberg, Doron; Srebnik, Morris; Nir-Paz, Ran; Weiss, Aryeh; Polacheck, Itzhack

2014-02-01

The aims of this study were to develop new anti-biofilm drugs, examine their activity against Candida albicans biofilm and investigate their structure-activity relationship and mechanism of action. A series of thiazolidinedione and succinimide derivatives were synthesized and their ability to inhibit C. albicans biofilm formation and destroy pre-formed biofilm was tested. The biofilms' structure, metabolic activity and viability were determined by XTT assay and propidium iodide and SYTO 9 live/dead stains combined with confocal microscopic analysis. The effect of the most active compounds on cell morphology, sterol distribution and cell wall morphology and composition was then determined by specific fluorescent stains and transmission electron microscopy. Most of the compounds were active at sub-MICs. Elongation of the aliphatic side chain resulted in reduced anti-biofilm activity and the sulphur atom contributed to biofilm killing, indicating a structure-activity relationship. The compounds differed in their effects on biofilm viability, yeast-to-hyphal form transition, hyphal morphology, cell wall morphology and composition, and sterol distribution. The most effective anti-biofilm compounds were the thiazolidinedione S8H and the succinimide NA8. We developed novel anti-biofilm agents that both inhibited and destroyed C. albicans biofilm. With some further development, these agents might be suitable for therapeutic purposes.

Candida albicans mannans mediate Streptococcus mutans exoenzyme GtfB binding to modulate cross-kingdom biofilm development in vivo.

Science.gov (United States)

Hwang, Geelsu; Liu, Yuan; Kim, Dongyeop; Li, Yong; Krysan, Damian J; Koo, Hyun

2017-06-01

Candida albicans is frequently detected with heavy infection by Streptococcus mutans in plaque-biofilms from children with early-childhood caries (ECC). This cross-kingdom biofilm contains an extensive matrix of extracellular α-glucans that is produced by an exoenzyme (GtfB) secreted by S. mutans. Here, we report that mannans located on the outer surface of C. albicans cell-wall mediates GtfB binding, enhancing glucan-matrix production and modulating bacterial-fungal association within biofilms formed in vivo. Using single-molecule atomic force microscopy, we determined that GtfB binds with remarkable affinity to mannans and to the C. albicans surface, forming a highly stable and strong bond (1-2 nN). However, GtfB binding properties to C. albicans was compromised in strains defective in O-mannan (pmt4ΔΔ) or N-mannan outer chain (och1ΔΔ). In particular, the binding strength of GtfB on och1ΔΔ strain was severely disrupted (>3-fold reduction vs. parental strain). In turn, the GtfB amount on the fungal surface was significantly reduced, and the ability of C. albicans mutant strains to develop mixed-species biofilms with S. mutans was impaired. This phenotype was independent of hyphae or established fungal-biofilm regulators (EFG1, BCR1). Notably, the mechanical stability of the defective biofilms was weakened, resulting in near complete biomass removal by shear forces. In addition, these in vitro findings were confirmed in vivo using a rodent biofilm model. Specifically, we observed that C. albicans och1ΔΔ was unable to form cross-kingdom biofilms on the tooth surface of rats co-infected with S. mutans. Likewise, co-infection with S. mutans defective in GtfB was also incapable of forming mixed-species biofilms. Taken together, the data support a mechanism whereby S. mutans-secreted GtfB binds to the mannan layer of C. albicans to promote extracellular matrix formation and their co-existence within biofilms. Enhanced understanding of GtfB-Candida interactions

Persea americana Glycolic Extract: In Vitro Study of Antimicrobial Activity against Candida albicans Biofilm and Cytotoxicity Evaluation

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D. Jesus

2015-01-01

Full Text Available This study evaluated the antifungal activity of Persea americana extract on Candida albicans biofilm and its cytotoxicity in macrophage culture (RAW 264.7. To determine the minimum inhibitory concentration (MIC, microdilution in broth (CLSI M27-S4 protocol was performed. Thereafter, the concentrations of 12.5, 25, 50, 100, and 200 mg/mL (n=10 with 5 min exposure were analyzed on mature biofilm in microplate wells for 48 h. Saline was used as control (n=10. After treatment, biofilm cells were scraped off and dilutions were plated on Sabouraud dextrose agar. After incubation (37°C/48 h, the values of colony forming units per milliliter (CFU/mL were converted to log10 and analyzed (ANOVA and Tukey test, 5%. The cytotoxicity of the P. americana extract was evaluated on macrophages by MTT assay. The MIC of the extract was 6.25 mg/mL and with 12.5 mg/mL there was elimination of 100% of planktonic cultures. Regarding the biofilms, a significant reduction (P<0.001 of the biofilm at concentrations of 50 (0.580±0.209 log10, 100 (0.998±0.508 log10, and 200 mg/mL (1.093±0.462 log10 was observed. The concentrations of 200 and 100 mg/mL were cytotoxic for macrophages, while the concentrations of 50, 25, and 12.5 mg/mL showed viability higher than 55%.

The Extracellular Matrix of Candida albicans Biofilms Impairs Formation of Neutrophil Extracellular Traps.

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Johnson, Chad J; Cabezas-Olcoz, Jonathan; Kernien, John F; Wang, Steven X; Beebe, David J; Huttenlocher, Anna; Ansari, Hamayail; Nett, Jeniel E

2016-09-01

Neutrophils release extracellular traps (NETs) in response to planktonic C. albicans. These complexes composed of DNA, histones, and proteins inhibit Candida growth and dissemination. Considering the resilience of Candida biofilms to host defenses, we examined the neutrophil response to C. albicans during biofilm growth. In contrast to planktonic C. albicans, biofilms triggered negligible release of NETs. Time lapse imaging confirmed the impairment in NET release and revealed neutrophils adhering to hyphae and migrating on the biofilm. NET inhibition depended on an intact extracellular biofilm matrix as physical or genetic disruption of this component resulted in NET release. Biofilm inhibition of NETosis could not be overcome by protein kinase C activation via phorbol myristate acetate (PMA) and was associated with suppression of neutrophil reactive oxygen species (ROS) production. The degree of impaired NET release correlated with resistance to neutrophil attack. The clinical relevance of the role for extracellular matrix in diminishing NET production was corroborated in vivo using a rat catheter model. The C. albicans pmr1Δ/Δ, defective in production of matrix mannan, appeared to elicit a greater abundance of NETs by scanning electron microscopy imaging, which correlated with a decreased fungal burden. Together, these findings show that C. albicans biofilms impair neutrophil response through an inhibitory pathway induced by the extracellular matrix.

Evaluation of Candida albicans biofilm formation on various dental ...

African Journals Online (AJOL)

Evaluation of Candida albicans biofilm formation on various dental restorative material surfaces. ... Nigerian Journal of Clinical Practice ... was significantly lower on the resin-modified glass ionomer and glass-ionomer cement samples. ... Conclusion: This finding emphasizes the use of glass ionomer restorative cements and ...

Activities of Fluconazole, Caspofungin, Anidulafungin, and Amphotericin B on Planktonic and Biofilm Candida Species Determined by Microcalorimetry

Science.gov (United States)

Maiolo, Elena Maryka; Furustrand Tafin, Ulrika; Borens, Olivier

2014-01-01

We investigated the activities of fluconazole, caspofungin, anidulafungin, and amphotericin B against Candida species in planktonic form and biofilms using a highly sensitive assay measuring growth-related heat production (microcalorimetry). C. albicans, C. glabrata, C. krusei, and C. parapsilosis were tested, and MICs were determined by the broth microdilution method. The antifungal activities were determined by isothermal microcalorimetry at 37°C in RPMI 1640. For planktonic Candida, heat flow was measured in the presence of antifungal dilutions for 24 h. Candida biofilm was formed on porous glass beads for 24 h and exposed to serial dilutions of antifungals for 24 h, and heat flow was measured for 48 h. The minimum heat inhibitory concentration (MHIC) was defined as the lowest antifungal concentration reducing the heat flow peak by ≥50% (≥90% for amphotericin B) at 24 h for planktonic Candida and at 48 h for Candida biofilms (measured also at 24 h). Fluconazole (planktonic MHICs, 0.25 to >512 μg/ml) and amphotericin B (planktonic MHICs, 0.25 to 1 μg/ml) showed higher MHICs than anidulafungin (planktonic MHICs, 0.015 to 0.5 μg/ml) and caspofungin (planktonic MHICs, 0.125 to 0.5 μg/ml). Against Candida species in biofilms, fluconazole's activity was reduced by >1,000-fold compared to its activity against the planktonic counterparts, whereas echinocandins and amphotericin B mainly preserved their activities. Fluconazole induced growth of planktonic C. krusei at sub-MICs. At high concentrations of caspofungin (>4 μg/ml), paradoxical growth of planktonic C. albicans and C. glabrata was observed. Microcalorimetry enabled real-time evaluation of antifungal activities against planktonic and biofilm Candida organisms. It can be used in the future to evaluate new antifungals and antifungal combinations and to study resistant strains. PMID:24566186

In vitro modification of Candida albicans invasiveness.

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Fontenla de Petrino, S E; de Jorrat, M E; Sirena, A; Valdez, J C; Mesón, O

1986-05-01

Candida albicans produces germ-tubes (GT) when it is incubated in animal or human serum. This dimorphism is responsible for its invasive ability. The purpose of the present paper is (1) to evaluate the ability of rat peritoneal macrophages to inhibit GT production of ingested Candida albicans, obtained from immunized rats and then activated in vitro with Candida-induced lymphokines; (2) to determinate any possible alteration of phagocytic and candidacidal activities. The phagocytes were obtained from rats immunized with viable C. albicans. Some of them were exposed to Candida-induced lymphokines in order to activate the macrophages in vitro. The monolayers of activated, immune and normal macrophages were infected with a C. albicans suspension during 4 hr. Activated macrophages presented not only the highest phagocytic and candidacidal activities but a noticeable inhibition of GT formation and incremented candidacidal activity.

Atividade antimicrobiana, antiaderente e antifúngica in vitro de plantas medicinais brasileiras sobre microrganismos do biofilme dental e cepas do gênero Candida In vitro antimicrobial, antiadherent and antifungal activity of Brazilian medicinal plants on oral biofilm microorganisms and strains of the genus Candida

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Pollianna Muniz Alves

2009-04-01

Full Text Available Avaliou-se in vitro a atividade antimicrobiana, antifúngica e antiaderente da aroeira-do-sertão, malva e goiabeira sobre microrganismos do biofilme dental e candidose oral. Os extratos mostraram-se eficazes, inibindo o crescimento das bactérias do biofilme dental e fungos da candidose oral, sugerindo a utilização dessas plantas como meio alternativo na terapêutica odontológica.The antimicrobial, antifungal and antiadherent activity of aroeira-do-sertão, mallow and guava tree on oral biofilm microorganisms and oral candidiasis was evaluated in vitro. The extracts were shown to be effective in inhibiting the growth of bacteria of the oral biofilm and fungi of oral candidiasis, thus suggesting that these extracts can be used as alternative means of dental therapy.

Anaerobic bacteria grow within Candida albicans biofilms and induce biofilm formation in suspension cultures.

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Fox, Emily P; Cowley, Elise S; Nobile, Clarissa J; Hartooni, Nairi; Newman, Dianne K; Johnson, Alexander D

2014-10-20

The human microbiome contains diverse microorganisms, which share and compete for the same environmental niches. A major microbial growth form in the human body is the biofilm state, where tightly packed bacterial, archaeal, and fungal cells must cooperate and/or compete for resources in order to survive. We examined mixed biofilms composed of the major fungal species of the gut microbiome, Candida albicans, and each of five prevalent bacterial gastrointestinal inhabitants: Bacteroides fragilis, Clostridium perfringens, Escherichia coli, Klebsiella pneumoniae, and Enterococcus faecalis. We observed that biofilms formed by C. albicans provide a hypoxic microenvironment that supports the growth of two anaerobic bacteria, even when cultured in ambient oxic conditions that are normally toxic to the bacteria. We also found that coculture with bacteria in biofilms induces massive gene expression changes in C. albicans, including upregulation of WOR1, which encodes a transcription regulator that controls a phenotypic switch in C. albicans, from the "white" cell type to the "opaque" cell type. Finally, we observed that in suspension cultures, C. perfringens induces aggregation of C. albicans into "mini-biofilms," which allow C. perfringens cells to survive in a normally toxic environment. This work indicates that bacteria and C. albicans interactions modulate the local chemistry of their environment in multiple ways to create niches favorable to their growth and survival. Copyright © 2014 Elsevier Ltd. All rights reserved.

Influence of Culture Media on Biofilm Formation by Candida Species and Response of Sessile Cells to Antifungals and Oxidative Stress

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Isela Serrano-Fujarte

2015-01-01

Full Text Available The aims of the study were to evaluate the influence of culture media on biofilm formation by C. albicans, C. glabrata, C. krusei, and C. parapsilosis and to investigate the responses of sessile cells to antifungals and reactive oxygen species (ROS as compared to planktonic cells. For biofilm formation, the Candida species were grown at different periods of time in YP or YNB media supplemented or not with 0.2 or 2% glucose. Sessile and planktonic cells were exposed to increasing concentrations of antifungals, H2O2, menadione or silver nanoparticles (AgNPs. Biofilms were observed by scanning electron microscopy (SEM and quantified by the XTT assay. C. albicans formed biofilms preferentially in YPD containing 2% glucose (YPD/2%, C. glabrata in glucose-free YNB or supplemented with 0.2% glucose (YNB/0.2%, while C. krusei and C. parapsilosis preferred YP, YPD/0.2%, and YPD/2%. Interestingly, only C. albicans produced an exopolymeric matrix. This is the first report dealing with the in vitro effect of the culture medium and glucose on the formation of biofilms in four Candida species as well as the resistance of sessile cells to antifungals, AgNPs, and ROS. Our results suggest that candidiasis in vivo is a multifactorial and complex process where the nutritional conditions, the human immune system, and the adaptability of the pathogen should be considered altogether to provide an effective treatment of the patient.

Influence of culture media on biofilm formation by Candida species and response of sessile cells to antifungals and oxidative stress.

Science.gov (United States)

Serrano-Fujarte, Isela; López-Romero, Everardo; Reyna-López, Georgina Elena; Martínez-Gámez, Ma Alejandrina; Vega-González, Arturo; Cuéllar-Cruz, Mayra

2015-01-01

The aims of the study were to evaluate the influence of culture media on biofilm formation by C. albicans, C. glabrata, C. krusei, and C. parapsilosis and to investigate the responses of sessile cells to antifungals and reactive oxygen species (ROS) as compared to planktonic cells. For biofilm formation, the Candida species were grown at different periods of time in YP or YNB media supplemented or not with 0.2 or 2% glucose. Sessile and planktonic cells were exposed to increasing concentrations of antifungals, H2O2, menadione or silver nanoparticles (AgNPs). Biofilms were observed by scanning electron microscopy (SEM) and quantified by the XTT assay. C. albicans formed biofilms preferentially in YPD containing 2% glucose (YPD/2%), C. glabrata in glucose-free YNB or supplemented with 0.2% glucose (YNB/0.2%), while C. krusei and C. parapsilosis preferred YP, YPD/0.2%, and YPD/2%. Interestingly, only C. albicans produced an exopolymeric matrix. This is the first report dealing with the in vitro effect of the culture medium and glucose on the formation of biofilms in four Candida species as well as the resistance of sessile cells to antifungals, AgNPs, and ROS. Our results suggest that candidiasis in vivo is a multifactorial and complex process where the nutritional conditions, the human immune system, and the adaptability of the pathogen should be considered altogether to provide an effective treatment of the patient.

Influence of Culture Media on Biofilm Formation by Candida Species and Response of Sessile Cells to Antifungals and Oxidative Stress

Science.gov (United States)

Serrano-Fujarte, Isela; Reyna-López, Georgina Elena; Martínez-Gámez, Ma. Alejandrina; Vega-González, Arturo; Cuéllar-Cruz, Mayra

2015-01-01

The aims of the study were to evaluate the influence of culture media on biofilm formation by C. albicans, C. glabrata, C. krusei, and C. parapsilosis and to investigate the responses of sessile cells to antifungals and reactive oxygen species (ROS) as compared to planktonic cells. For biofilm formation, the Candida species were grown at different periods of time in YP or YNB media supplemented or not with 0.2 or 2% glucose. Sessile and planktonic cells were exposed to increasing concentrations of antifungals, H2O2, menadione or silver nanoparticles (AgNPs). Biofilms were observed by scanning electron microscopy (SEM) and quantified by the XTT assay. C. albicans formed biofilms preferentially in YPD containing 2% glucose (YPD/2%), C. glabrata in glucose-free YNB or supplemented with 0.2% glucose (YNB/0.2%), while C. krusei and C. parapsilosis preferred YP, YPD/0.2%, and YPD/2%. Interestingly, only C. albicans produced an exopolymeric matrix. This is the first report dealing with the in vitro effect of the culture medium and glucose on the formation of biofilms in four Candida species as well as the resistance of sessile cells to antifungals, AgNPs, and ROS. Our results suggest that candidiasis in vivo is a multifactorial and complex process where the nutritional conditions, the human immune system, and the adaptability of the pathogen should be considered altogether to provide an effective treatment of the patient. PMID:25705688

Modulation of Candida albicans virulence by bacterial biofilms on titanium surfaces.

Science.gov (United States)

Cavalcanti, Yuri Wanderley; Wilson, Melanie; Lewis, Michael; Del-Bel-Cury, Altair Antoninha; da Silva, Wander José; Williams, David W

2016-01-01

Whilst Candida albicans occurs in peri-implant biofilms, its role in peri-implantitis remains unclear. This study therefore examined the virulence of C. albicans in mixed-species biofilms on titanium surfaces. Biofilms of C. albicans (Ca), C. albicans with streptococci (Streptococcus sanguinis, S. mutans) (Ca-Ss-Sm) and those incorporating Porphyromonas gingivalis (Ca-Pg and Ca-Ss-Sm-Pg) were developed. Expression of C. albicans genes associated with adhesion (ALS1, ALS3, HWP1) and hydrolytic enzymes (SAP2, SAP4, SAP6, PLD1) was measured and hyphal production by C. albicans quantified. Compared with Ca biofilms, significant (pbiofilms containing streptococci (Ca-Ss-Sm). In Ca-Pg biofilms, down-regulation of HWP1 and SAP4 expression, with reduced hyphal production occurred. Ca-Ss-Sm-Pg biofilms had increased hyphal proportions and up-regulation of ALS3, SAP2 and SAP6. In conclusion, C. albicans expressed virulence factors in biofilms that could contribute to peri-implantitis, but this was dependent on associated bacterial species.

Daya hambat xylitol dan nistation terhadap pertumbuhan Candida albicans (in vitro (Inhibition effect of xylitol and nistatin combination on Candida albicans growth (in vitro

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Sarah Kartimah Djajusman

2014-09-01

Full Text Available Background: The growth of Candida albicans can be controlled by using antifungal such as nystatin. These days we found that using antifungal is not enough to control Candida albicans, we also have to control the intake of sugar by using xylitol. Purpose: Purpose of the study was to determine the optimal inhibitory concentration of xylitol-nystatin in the Candida albicans growth. Methods: This was an in-vitro study using an antimicrobial test of serial dilution with xylitol-nystatin and sucrose–nystatin consentration of 1%, 3%, 5%, 7%, 9%, and 10%.Growth inhibition of C. albicans was determined by the inhibition zone of xylitol + nystatin on C. albicans culture media (in vitro Results: The result of study was the inhibitory consentration of xylitol-nystatin to inhibit Candida albicans growth was 3%-10%. Conclusion: The study showed that combination of xylitol and nystation could inhibit the growth of Candida albicans.Latar belakang: Pertumbuhan Candida albicans dapat dikontrol dengan menggunakan antijamur seperti nistatin. Penggunakan antijamur saja tidak cukup untuk mengontrol Candida albicans, namun perlu pula mengontrol asupan gula dengan menggunakan xylitol. Tujuan: Tujuan dari penelitian ini adalah untuk menentukan konsentrasi hambat optimal xylitol-nistatin dalam pertumbuhan Candida albicans. Metode: Penelitian ini merupakan penelitian in vitro menggunakan uji antimikroba pengenceran serial dengan xylitol-nistatin dan nystatin-sukrosa konsentrasi 1%, 3 %, 5 %, 7%, 9%, dan 10%. Daya hambat pertumbuhan C. albicans diukur dari zona hambat xylitol + nistatin pada media kultur C. albicans (in vitro Hasil: Konsentrasi penghambatan xylitol-nistatin untuk menghambat pertumbuhan Candida albicans adalah 3-10%. Simpulan: Hasil penelitian menunjukkan bahwa kombinasi xylitol dan nystation bisa menghambat pertumbuhan Candida albicans.

Voice Prosthetic Biofilm Formation and Candida Morphogenic Conversions in Absence and Presence of Different Bacterial Strains and Species on Silicone-Rubber

NARCIS (Netherlands)

van der Mei, Henny C.; Buijssen, Kevin J. D. A.; van der Laan, Bernard F. A. M.; Ovchinnikova, Ekatarina; Geertsema-Doornbusch, Gesinda I.; Atema-Smit, Jelly; van de Belt-Gritter, Betsy; Busscher, Henk J.

2014-01-01

Morphogenic conversion of Candida from a yeast to hyphal morphology plays a pivotal role in the pathogenicity of Candida species. Both Candida albicans and Candida tropicalis, in combination with a variety of different bacterial strains and species, appear in biofilms on silicone-rubber voice

The N-Terminus of Human Lactoferrin Displays Anti-biofilm Activity on Candida parapsilosis in Lumen Catheters.

Science.gov (United States)

Fais, Roberta; Di Luca, Mariagrazia; Rizzato, Cosmeri; Morici, Paola; Bottai, Daria; Tavanti, Arianna; Lupetti, Antonella

2017-01-01

Candida parapsilosis is a major cause of hospital-acquired infection, often related to parenteral nutrition administered via catheters and hand colonization of health care workers, and its peculiar biofilm formation ability on plastic surfaces. The mortality rate of 30% points to the pressing need for new antifungal drugs. The present study aimed at analyzing the inhibitory activity of the N-terminal lactoferrin-derived peptide, further referred to as hLF 1-11, against biofilms produced by clinical isolates of C. parapsilosis characterized for their biofilm forming ability and fluconazole susceptibility. hLF 1-11 anti-biofilm activity was assessed in terms of reduction of biofilm biomass, metabolic activity, and observation of sessile cell morphology on polystyrene microtiter plates and using an in vitro model of catheter-associated C. parapsilosis biofilm production. Moreover, fluctuation in transcription levels of genes related to cell adhesion, hyphal development and extracellular matrix production upon peptide exposure were evaluated by quantitative real time RT-PCR. The results revealed that hLF 1-11 exhibits an inhibitory effect on biofilm formation by all the C. parapsilosis isolates tested, in a dose-dependent manner, regardless of their fluconazole susceptibility. In addition, hLF 1-11 induced a statistically significant dose-dependent reduction of preformed-biofilm cellular density and metabolic activity at high peptide concentrations only. Interestingly, when assessed in a catheter lumen, hLF 1-11 was able to induce a 2-log reduction of sessile cell viability at both the peptide concentrations used in RPMI diluted in NaPB. A more pronounced anti-biofilm effect was observed (3.5-log reduction) when a 10% glucose solution was used as experimental condition on both early and preformed C. parapsilosis biofilm. Quantitative real time RT-PCR experiments confirmed that hLF 1-11 down-regulates key biofilm related genes. The overall findings suggest hLF 1-11 as a

Treatment of Candida albicans biofilms with low-temperature plasma induced by dielectric barrier discharge and atmospheric pressure plasma jet

International Nuclear Information System (INIS)

Koban, Ina; Welk, Alexander; Meisel, Peter; Holtfreter, Birte; Kocher, Thomas; Matthes, Rutger; Huebner, Nils-Olaf; Kramer, Axel; Sietmann, Rabea; Kindel, Eckhard; Weltmann, Klaus-Dieter

2010-01-01

Because of some disadvantages of chemical disinfection in dental practice (especially denture cleaning), we investigated the effects of physical methods on Candida albicans biofilms. For this purpose, the antifungal efficacy of three different low-temperature plasma devices (an atmospheric pressure plasma jet and two different dielectric barrier discharges (DBDs)) on Candida albicans biofilms grown on titanium discs in vitro was investigated. As positive treatment controls, we used 0.1% chlorhexidine digluconate (CHX) and 0.6% sodium hypochlorite (NaOCl). The corresponding gas streams without plasma ignition served as negative treatment controls. The efficacy of the plasma treatment was determined evaluating the number of colony-forming units (CFU) recovered from titanium discs. The plasma treatment reduced the CFU significantly compared to chemical disinfectants. While 10 min CHX or NaOCl exposure led to a CFU log 10 reduction factor of 1.5, the log 10 reduction factor of DBD plasma was up to 5. In conclusion, the use of low-temperature plasma is a promising physical alternative to chemical antiseptics for dental practice.

Treatment of Candida albicans biofilms with low-temperature plasma induced by dielectric barrier discharge and atmospheric pressure plasma jet

Energy Technology Data Exchange (ETDEWEB)

Koban, Ina; Welk, Alexander; Meisel, Peter; Holtfreter, Birte; Kocher, Thomas [Unit of Periodontology, Dental School, University of Greifswald, Rotgerberstr. 8, 17475 Greifswald (Germany); Matthes, Rutger; Huebner, Nils-Olaf; Kramer, Axel [Institute for Hygiene and Environmental Medicine, University of Greifswald, Walther-Rathenau-Str. 49 a, 17487 Greifswald (Germany); Sietmann, Rabea [Institute of Microbiology, University of Greifswald, Friedrich-Ludwig-Jahn-Str. 15, 17487 Greifswald (Germany); Kindel, Eckhard; Weltmann, Klaus-Dieter, E-mail: ina.koban@uni-greifswald.d [Leibniz Institute for Plasma Science and Technology (INP), Felix-Hausdorff-Str. 2, 17489 Greifswald (Germany)

2010-07-15

Because of some disadvantages of chemical disinfection in dental practice (especially denture cleaning), we investigated the effects of physical methods on Candida albicans biofilms. For this purpose, the antifungal efficacy of three different low-temperature plasma devices (an atmospheric pressure plasma jet and two different dielectric barrier discharges (DBDs)) on Candida albicans biofilms grown on titanium discs in vitro was investigated. As positive treatment controls, we used 0.1% chlorhexidine digluconate (CHX) and 0.6% sodium hypochlorite (NaOCl). The corresponding gas streams without plasma ignition served as negative treatment controls. The efficacy of the plasma treatment was determined evaluating the number of colony-forming units (CFU) recovered from titanium discs. The plasma treatment reduced the CFU significantly compared to chemical disinfectants. While 10 min CHX or NaOCl exposure led to a CFU log{sub 10} reduction factor of 1.5, the log{sub 10} reduction factor of DBD plasma was up to 5. In conclusion, the use of low-temperature plasma is a promising physical alternative to chemical antiseptics for dental practice.

Effect of alcohols on filamentation, growth, viability and biofilm development in Candida albicans.

Science.gov (United States)

Chauhan, Nitin M; Shinde, Ravikumar B; Karuppayil, S Mohan

2013-12-01

In this study we report the potential of alcohols as morphogenetic regulators in Candida albicans. All the alcohols tested influenced various modes of growth like planktonic as well as biofilm forms. Viability was affected at high concentrations. Among the alcohols, the response of C. albicans to amyl alcohol (pentanol) was noteworthy. Amyl alcohol at a concentration 0.5% which was not inhibitory to growth and viability specifically inhibited morphogenetic switching from yeast to hyphal forms. It also inhibited normal biofilm development favoring yeast dominated biofilms. Based on this study we hypothesize that alcohols produced under anaerobic conditions may not favor biofilm development and support dissemination of yeast cells. Since anaerobic conditions are not found to favor production of quorum sensing molecules like farnesol, the alcohols may play a role in morphogenetic regulation.

An in vitro Model for Oral Mixed Biofilms of Candida albicans and Streptococcus gordonii in Synthetic Saliva

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Daniel eMontelongo-Jauregui

2016-05-01

Full Text Available As a member of the normal human oral microbiota, the fungus C. albicans is often found in association with Streptococcus gordonii, a member of dental plaque forming bacteria. Evidence suggests that S. gordonii serves as a facilitator of C. albicans adherence to dental tissues, which represents a clinically relevant problem, particularly for immunocompromised individuals that could subsequently develop fungal infections. In this study we describe the development of a relatively simple and economical in vitro model that allows for the growth of mixed bacterial/fungal biofilms in 96-well microtiter plates. We have applied this method to test and compare the growth characteristics of single and dual species biofilms in traditional microbiological media versus a synthetic saliva medium (basal medium mucin, BMM that more closely resembles physiological conditions within the oral cavity. Results indicated a synergistic effect for the formation of biofilms when both microorganisms were seeded together under all conditions tested. The structural and architectural features of the resulting biofilms were further characterized using scanning electron microscopy (SEM and confocal scanning laser microscopy (CSLM. We also performed drug susceptibility assays against single and mixed species biofilms using commonly used antifungals and antibacterial antibiotics, both in monotherapy and in combination therapy, for a direct comparison of resistance against antimicrobial treatment. As expected, mixed species biofilms displayed higher levels of resistance to antimicrobial treatment at every dose tested in both traditional media and BMM synthetic saliva, as compared to single-species biofilms.

Development of a high-throughput Candida albicans biofilm chip.

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Anand Srinivasan

2011-04-01

Full Text Available We have developed a high-density microarray platform consisting of nano-biofilms of Candida albicans. A robotic microarrayer was used to print yeast cells of C. albicans encapsulated in a collagen matrix at a volume as low as 50 nL onto surface-modified microscope slides. Upon incubation, the cells grow into fully formed "nano-biofilms". The morphological and architectural complexity of these biofilms were evaluated by scanning electron and confocal scanning laser microscopy. The extent of biofilm formation was determined using a microarray scanner from changes in fluorescence intensities due to FUN 1 metabolic processing. This staining technique was also adapted for antifungal susceptibility testing, which demonstrated that, similar to regular biofilms, cells within the on-chip biofilms displayed elevated levels of resistance against antifungal agents (fluconazole and amphotericin B. Thus, results from structural analyses and antifungal susceptibility testing indicated that despite miniaturization, these biofilms display the typical phenotypic properties associated with the biofilm mode of growth. In its final format, the C. albicans biofilm chip (CaBChip is composed of 768 equivalent and spatially distinct nano-biofilms on a single slide; multiple chips can be printed and processed simultaneously. Compared to current methods for the formation of microbial biofilms, namely the 96-well microtiter plate model, this fungal biofilm chip has advantages in terms of miniaturization and automation, which combine to cut reagent use and analysis time, minimize labor intensive steps, and dramatically reduce assay costs. Such a chip should accelerate the antifungal drug discovery process by enabling rapid, convenient and inexpensive screening of hundreds-to-thousands of compounds simultaneously.

Long-term efficacy of denture cleansers in preventing Candida spp. biofilm recolonization on liner surface.

Science.gov (United States)

Vieira, Ana Paula Coelho; Senna, Plínio Mendes; Silva, Wander José da; Del Bel Cury, Altair Antoninha

2010-01-01

This study evaluated the long-term efficacy of denture cleansers against Candida spp. biofilm recolonization on liner surface. Specimens were fabricated of a poly(methyl methacrylate)-based denture liner and had their surface roughness evaluated at baseline and after cleansing treatments. C. albicans or C. glabrata biofilms were formed on liner surface for 48 h, and then the specimens were randomly assigned to one of cleaning treatments: two alkaline peroxides (soaking for 3 or 15 min), 0.5% sodium hypochlorite (10 min) or distilled water (control; 15 min). After the treatments, the specimens were sonicated to disrupt the biofilm, and residual cells were counted (cell/mL). Long-term effectiveness of the cleaning processes was determined by submitting a set of cleaned specimens to biofilm growth conditions for 48 h followed by estimation of cell counts. The topography of specimens after cleaning treatments was analyzed by SEM. Data were analyzed by ANOVA and Tukey's test (α; = 0.05). Results of cell count estimation showed significant differences in cleanliness among the treatments (p 0.05) was observed among the Candida species regarding the recolonization condition. Alkaline denture cleansers showed similar cleaning performance and both differed from the control (p recolonization.

Efficacy of citric acid denture cleanser on the Candida albicans biofilm formed on poly(methyl methacrylate): effects on residual biofilm and recolonization process.

Science.gov (United States)

Faot, Fernanda; Cavalcanti, Yuri Wanderley; Mendonça e Bertolini, Martinna de; Pinto, Luciana de Rezende; da Silva, Wander José; Cury, Altair Antoninha Del Bel

2014-06-23

It is well known that the use of denture cleansers can reduce Candida albicans biofilm accumulation; however, the efficacy of citric acid denture cleansers is uncertain. In addition, the long-term efficacy of this denture cleanser is not well established, and their effect on residual biofilms is unknown. This in vitro study evaluated the efficacy of citric acid denture cleanser treatment on C. albicans biofilm recolonization on poly(methyl methacrylate) (PMMA) surface. C. albicans biofilms were developed for 72 h on PMMA resin specimens (n = 168), which were randomly assigned to 1 of 3 cleansing treatments (CTs) overnight (8 h). CTs included purified water as a control (CTC) and two experimental groups that used either a 1:5 dilution of citric acid denture cleanser (CT5) or a 1:8 dilution of citric acid denture cleanser (CT8). Residual biofilms adhering to the specimens were collected and quantified at two time points: immediately after CTs (ICT) and after cleaning and residual biofilm recolonization (RT). Residual biofilms were analyzed by quantifying the viable cells (CFU/mL), and biofilm architecture was evaluated by confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM). Denture cleanser treatments and evaluation periods were considered study factors. Data were analyzed using two-way ANOVA and Tukey's Honestly Significant Difference (HSD) test (α = 0.05). Immediately after treatments, citric acid denture cleansing solutions (CT5 and CT8) reduced the number of viable cells as compared with the control (p recolonization (p recolonization was also detected by CLSM and SEM analysis, which revealed a higher biomass and average biofilm thickness for the CT8 group (p recolonization.

Candida Species Biofilms’ Antifungal Resistance

Science.gov (United States)

Silva, Sónia; Rodrigues, Célia F.; Araújo, Daniela; Rodrigues, Maria Elisa; Henriques, Mariana

2017-01-01

Candida infections (candidiasis) are the most prevalent opportunistic fungal infection on humans and, as such, a major public health problem. In recent decades, candidiasis has been associated to Candida species other than Candida albicans. Moreover, biofilms have been considered the most prevalent growth form of Candida cells and a strong causative agent of the intensification of antifungal resistance. As yet, no specific resistance factor has been identified as the sole responsible for the increased recalcitrance to antifungal agents exhibited by biofilms. Instead, biofilm antifungal resistance is a complex multifactorial phenomenon, which still remains to be fully elucidated and understood. The different mechanisms, which may be responsible for the intrinsic resistance of Candida species biofilms, include the high density of cells within the biofilm, the growth and nutrient limitation, the effects of the biofilm matrix, the presence of persister cells, the antifungal resistance gene expression and the increase of sterols on the membrane of biofilm cells. Thus, this review intends to provide information on the recent advances about Candida species biofilm antifungal resistance and its implication on intensification of the candidiasis. PMID:29371527

The in vitro and in vivo efficacy of fluconazole in combination with farnesol against Candida albicans isolates using a murine vulvovaginitis model.

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Bozó, Aliz; Domán, Marianna; Majoros, László; Kardos, Gábor; Varga, István; Kovács, Renátó

2016-11-01

Farnesol is a quorum-sensing molecule that inhibits biofilm formation in Candida albicans. Previous in vitro data suggest that, in combination with certain antifungals, farnesol may have an adjuvant anti-biofilm agent. However, the in vivo efficacy of farnesol is very questionable. Therefore, the in vitro and in vivo activity of fluconazole combined with farnesol was evaluated against C. albicans biofilms using fractional inhibitory concentration index (FICI) determination, time-kill experiments and a murine vulvovaginitis model. The median biofilm MICs of fluconazole-sensitive C. albicans isolates ranged between 4 -> 512 mg/L and 150-300 μM for fluconazole and farnesol, respectively. These values were 512 -> 512 mg/L and > 300 μM for fluconazole-resistant clinical isolates. Farnesol decreased the median MICs of fluconazole by 2-64-fold for biofilms. Based on FICI, synergistic interaction was observed only in the case of the sessile SC5314 reference strain (FICIs: 0.16-0.27). In time-kill studies, only the 512 mg/L fluconazole and 512 mg/L fluconazole + 75 μM farnesol reduced biofilm mass significantly at each time point in the case of all isolates. The combination reduced the metabolic activity of biofilms for all isolates in a concentration- and time-dependent manner. Our findings revealed that farnesol alone was not protective in a murine vulvovaginitis model. Farnesol was not beneficial in combination with fluconazole for fluconazole-susceptible isolates, but partially increased fluconazole activity against one fluconazole-resistant isolate, but not the other one.

Screening of pharmacologically active small molecule compounds identifies antifungal agents against Candida biofilms

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Takao eWatamoto

2015-12-01

Full Text Available Candida species have emerged as important and common opportunistic human pathogens, particularly in immunocompromised individuals. The current antifungal therapies either have toxic side effects or are insufficiently effect. The aim of this study is develop new small-molecule antifungal compounds by library screening methods using C. albicans, and to evaluate their antifungal effects on Candida biofilms and cytotoxic effects on human cells. Wild-type C. albicans strain SC5314 was used in library screening. To identify antifungal compounds, we screened a small-molecule library of 1,280 pharmacologically active compounds (LOPAC1280TM using an antifungal susceptibility test (AST. To investigate the antifungal effects of the hit compounds, ASTs were conducted using Candida strains in various growth modes, including biofilms. We tested the cytotoxicity of the hit compounds using human gingival fibroblast (hGF cells to evaluate their clinical safety. Only 35 compounds were identified by screening, which inhibited the metabolic activity of C. albicans by >50%. Of these, 26 compounds had fungistatic effects and 9 compounds had fungicidal effects on C. albicans. Five compounds, BAY11-7082, BAY11-7085, sanguinarine chloride hydrate, ellipticine and CV-3988, had strong fungicidal effects and could inhibit the metabolic activity of Candida biofilms. However, BAY11-7082, BAY11-7085, sanguinarine chloride hydrate and ellipticine were cytotoxic to hGF cells at low concentrations. CV-3988 showed no cytotoxicity at a fungicidal concentration.Four of the compounds identified, BAY11-7082, BAY11-7085, sanguinarine chloride hydrate and ellipticine, had toxic effects on Candida strains and hGF cells. In contrast, CV-3988 had fungicidal effects on Candida strains, but low cytotoxic effects on hGF cells. Therefore, this screening reveals agent, CV-3988 that was previously unknown to be antifungal agent, which could be a novel therapies for superficial mucosal

Genetic Control of Conventional and Pheromone-Stimulated Biofilm Formation in Candida albicans

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Lin, Ching-Hsuan; Kabrawala, Shail; Fox, Emily P.; Nobile, Clarissa J.; Johnson, Alexander D.; Bennett, Richard J.

2013-01-01

Candida albicans can stochastically switch between two phenotypes, white and opaque. Opaque cells are the sexually competent form of C. albicans and therefore undergo efficient polarized growth and mating in the presence of pheromone. In contrast, white cells cannot mate, but are induced – under a specialized set of conditions – to form biofilms in response to pheromone. In this work, we compare the genetic regulation of such “pheromone-stimulated” biofilms with that of “conventional” C. albicans biofilms. In particular, we examined a network of six transcriptional regulators (Bcr1, Brg1, Efg1, Tec1, Ndt80, and Rob1) that mediate conventional biofilm formation for their potential roles in pheromone-stimulated biofilm formation. We show that four of the six transcription factors (Bcr1, Brg1, Rob1, and Tec1) promote formation of both conventional and pheromone-stimulated biofilms, indicating they play general roles in cell cohesion and biofilm development. In addition, we identify the master transcriptional regulator of pheromone-stimulated biofilms as C. albicans Cph1, ortholog of Saccharomyces cerevisiae Ste12. Cph1 regulates mating in C. albicans opaque cells, and here we show that Cph1 is also essential for pheromone-stimulated biofilm formation in white cells. In contrast, Cph1 is dispensable for the formation of conventional biofilms. The regulation of pheromone- stimulated biofilm formation was further investigated by transcriptional profiling and genetic analyses. These studies identified 196 genes that are induced by pheromone signaling during biofilm formation. One of these genes, HGC1, is shown to be required for both conventional and pheromone-stimulated biofilm formation. Taken together, these observations compare and contrast the regulation of conventional and pheromone-stimulated biofilm formation in C. albicans, and demonstrate that Cph1 is required for the latter, but not the former. PMID:23637598

Quercetin Assists Fluconazole to Inhibit Biofilm Formations of Fluconazole-Resistant Candida Albicans in In Vitro and In Vivo Antifungal Managements of Vulvovaginal Candidiasis

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Mei Gao

2016-11-01

Full Text Available Background: Vulvovaginal candidiasis (VVC is a common gynecological disease. Candida albicans is believed to be mainly implicated in VVC occurrence, the biofilm of which is one of the virulence factors responsible for resistance to traditional antifungal agents especially to fluconazole (FCZ. Quercetin (QCT is a dietary flavonoid and has been demonstrated to be antifungal against C. albicans biofilm. Methods: 17 C. albicans isolates including 15 clinical ones isolated from VVC patients were employed to investigate the effects of QCT and/or FCZ on the inhibition of C. albicans biofilm. Results: We observed that 64 µg/mL QCT and/or 128 µg/mL FCZ could (i be synergistic against 10 FCZ-resistant planktonic and 17 biofilm cells of C. albicans, (ii inhibit fungal adherence, cell surface hydrophobicity (CSH, flocculation, yeast-to-hypha transition, metabolism, thickness and dispersion of biofilms; (iii down-regulate the expressions of ALS1, ALS3, HWP1, SUN41, UME6 and ECE1 and up-regulate the expressions of PDE2, NRG1 and HSP90, and we also found that (iv the fungal burden was reduced in vaginal mucosa and the symptoms were alleviated in a murine VVC model after the treatments of 5 mg/kg QCT and/or 20 mg/kg FCZ. Conclusion: Together with these results, it could be demonstrated that QCT could be a favorable antifungal agent and a promising synergist with FCZ in the clinical management of VVC caused by C. albicans biofilm.

Inhibition of Candida albicans Biofilm Formation by the Synthetic Lactoferricin Derived Peptide hLF1-11.

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Morici, Paola; Fais, Roberta; Rizzato, Cosmeri; Tavanti, Arianna; Lupetti, Antonella

2016-01-01

The aim of this study was to evaluate the in vitro activity of the synthetic peptide hLF1-11 against biofilm produced by clinical isolates of Candida albicans with different fluconazole susceptibility. The antibiofilm activity of the peptide hLF1-11 was assessed in terms of reduction of biofilm cellular density, metabolic activity and sessile cell viability. The extent of morphogenesis in hLF1-11 treated and untreated biofilms was also investigated microscopically. Transcription levels of genes related to cell adhesion, hyphal development and extracellular matrix production were analysed by qRT-PCR in hLF1-11 treated and untreated biofilms. Exogenous dibutyryl-cAMP (db-cAMP) was used to rescue morphogenesis in cells exposed to the peptide. The results revealed that hLF1-11 exhibited an inhibitory effect on biofilm formation by all C. albicans isolates tested in a dose-dependent manner, regardless of their fluconazole susceptibility. Visual inspection of treated or untreated biofilm cells with an inverted microscope revealed a significant reduction in hyphal formation by hLF1-11 treated cells, as early as 3 hours of incubation. Moreover, hLF1-11 showed a reduced activity on preadherent cells. hLF1-11 induced the down-regulation of biofilm and hyphal-associated genes, which were predominantly regulated via the Ras1-cAMP-Efg1 pathway. Indeed, exogenous db-cAMP restored morphogenesis in hLF1-11 treated cells. The hLF1-11 peptide significantly inhibited biofilm formation by C. albicans mainly at early stages, interfering with biofilm cellular density and metabolic activity, and affected morphogenesis through the Ras1-cAMP-Efg1 pathway. Our findings provide the first evidence that hLF1-11 could represent a potential candidate for the prevention of biofilm formation by C. albicans.

Inhibition of Candida albicans Biofilm Formation by the Synthetic Lactoferricin Derived Peptide hLF1-11

Science.gov (United States)

Morici, Paola; Fais, Roberta; Rizzato, Cosmeri

2016-01-01

The aim of this study was to evaluate the in vitro activity of the synthetic peptide hLF1-11 against biofilm produced by clinical isolates of Candida albicans with different fluconazole susceptibility. The antibiofilm activity of the peptide hLF1-11 was assessed in terms of reduction of biofilm cellular density, metabolic activity and sessile cell viability. The extent of morphogenesis in hLF1-11 treated and untreated biofilms was also investigated microscopically. Transcription levels of genes related to cell adhesion, hyphal development and extracellular matrix production were analysed by qRT-PCR in hLF1-11 treated and untreated biofilms. Exogenous dibutyryl-cAMP (db-cAMP) was used to rescue morphogenesis in cells exposed to the peptide. The results revealed that hLF1-11 exhibited an inhibitory effect on biofilm formation by all C. albicans isolates tested in a dose-dependent manner, regardless of their fluconazole susceptibility. Visual inspection of treated or untreated biofilm cells with an inverted microscope revealed a significant reduction in hyphal formation by hLF1-11 treated cells, as early as 3 hours of incubation. Moreover, hLF1-11 showed a reduced activity on preadherent cells. hLF1-11 induced the down-regulation of biofilm and hyphal-associated genes, which were predominantly regulated via the Ras1-cAMP-Efg1 pathway. Indeed, exogenous db-cAMP restored morphogenesis in hLF1-11 treated cells. The hLF1-11 peptide significantly inhibited biofilm formation by C. albicans mainly at early stages, interfering with biofilm cellular density and metabolic activity, and affected morphogenesis through the Ras1-cAMP-Efg1 pathway. Our findings provide the first evidence that hLF1-11 could represent a potential candidate for the prevention of biofilm formation by C. albicans. PMID:27902776

Inhibition of Candida albicans Biofilm Formation by the Synthetic Lactoferricin Derived Peptide hLF1-11.

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Paola Morici

Full Text Available The aim of this study was to evaluate the in vitro activity of the synthetic peptide hLF1-11 against biofilm produced by clinical isolates of Candida albicans with different fluconazole susceptibility. The antibiofilm activity of the peptide hLF1-11 was assessed in terms of reduction of biofilm cellular density, metabolic activity and sessile cell viability. The extent of morphogenesis in hLF1-11 treated and untreated biofilms was also investigated microscopically. Transcription levels of genes related to cell adhesion, hyphal development and extracellular matrix production were analysed by qRT-PCR in hLF1-11 treated and untreated biofilms. Exogenous dibutyryl-cAMP (db-cAMP was used to rescue morphogenesis in cells exposed to the peptide. The results revealed that hLF1-11 exhibited an inhibitory effect on biofilm formation by all C. albicans isolates tested in a dose-dependent manner, regardless of their fluconazole susceptibility. Visual inspection of treated or untreated biofilm cells with an inverted microscope revealed a significant reduction in hyphal formation by hLF1-11 treated cells, as early as 3 hours of incubation. Moreover, hLF1-11 showed a reduced activity on preadherent cells. hLF1-11 induced the down-regulation of biofilm and hyphal-associated genes, which were predominantly regulated via the Ras1-cAMP-Efg1 pathway. Indeed, exogenous db-cAMP restored morphogenesis in hLF1-11 treated cells. The hLF1-11 peptide significantly inhibited biofilm formation by C. albicans mainly at early stages, interfering with biofilm cellular density and metabolic activity, and affected morphogenesis through the Ras1-cAMP-Efg1 pathway. Our findings provide the first evidence that hLF1-11 could represent a potential candidate for the prevention of biofilm formation by C. albicans.

Action of Coriandrum sativum L. Essential Oil upon Oral Candida albicans Biofilm Formation.

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Furletti, V F; Teixeira, I P; Obando-Pereda, G; Mardegan, R C; Sartoratto, A; Figueira, G M; Duarte, R M T; Rehder, V L G; Duarte, M C T; Höfling, J F

2011-01-01

The efficacy of extracts and essential oils from Allium tuberosum, Coriandrum sativum, Cymbopogon martini, Cymbopogon winterianus, and Santolina chamaecyparissus was evaluated against Candida spp. isolates from the oral cavity of patients with periodontal disease. The most active oil was fractionated and tested against C. albicans biofilm formation. The oils were obtained by water-distillation and the extracts were prepared with macerated dried plant material. The Minimal Inhibitory Concentration-MIC was determined by the microdilution method. Chemical characterization of oil constituents was performed using Gas Chromatography and Mass Spectrometry (GC-MS). C. sativum activity oil upon cell and biofilm morphology was evaluated by Scanning Electron Microscopy (SEM). The best activities against planktonic Candida spp. were observed for the essential oil and the grouped F(8-10) fractions from C. sativum. The crude oil also affected the biofilm formation in C. albicans causing a decrease in the biofilm growth. Chemical analysis of the F(8-10) fractions detected as major active compounds, 2-hexen-1-ol, 3-hexen-1-ol and cyclodecane. Standards of these compounds tested grouped provided a stronger activity than the oil suggesting a synergistic action from the major oil constituents. The activity of C. sativum oil demonstrates its potential for a new natural antifungal formulation.

Action of Coriandrum sativum L. Essential Oil upon Oral Candida albicans Biofilm Formation

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V. F. Furletti

2011-01-01

Full Text Available The efficacy of extracts and essential oils from Allium tuberosum, Coriandrum sativum, Cymbopogon martini, Cymbopogon winterianus, and Santolina chamaecyparissus was evaluated against Candida spp. isolates from the oral cavity of patients with periodontal disease. The most active oil was fractionated and tested against C. albicans biofilm formation. The oils were obtained by water-distillation and the extracts were prepared with macerated dried plant material. The Minimal Inhibitory Concentration—MIC was determined by the microdilution method. Chemical characterization of oil constituents was performed using Gas Chromatography and Mass Spectrometry (GC-MS. C. sativum activity oil upon cell and biofilm morphology was evaluated by Scanning Electron Microscopy (SEM. The best activities against planktonic Candida spp. were observed for the essential oil and the grouped F8–10 fractions from C. sativum. The crude oil also affected the biofilm formation in C. albicans causing a decrease in the biofilm growth. Chemical analysis of the F8–10 fractions detected as major active compounds, 2-hexen-1-ol, 3-hexen-1-ol and cyclodecane. Standards of these compounds tested grouped provided a stronger activity than the oil suggesting a synergistic action from the major oil constituents. The activity of C. sativum oil demonstrates its potential for a new natural antifungal formulation.

Promising results of cranberry in the prevention of oral Candida biofilms.

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Girardot, Marion; Guerineau, Amandine; Boudesocque, Leslie; Costa, Damien; Bazinet, Laurent; Enguehard-Gueiffier, Cécile; Imbert, Christine

2014-04-01

In the context of dental caries prevention by natural foodstuff sources, antifungal and antibiofilm activities of dry commercial extracts of cranberry fruit (Vaccinium macrocarpon Aiton) and two other red fruits (Vaccinium myrtillus L. and Malpighia punicifolia L.) were assessed on Candida albicans and Candida glabrata yeasts. When added to the culture medium, the cranberry extract displayed a significant anti-adhesion activity against Candida spp. when used at low concentrations. In addition, the pretreatment of surfaces with this extract induced an anti-adhesion activity mainly against C. glabrata yeasts and an antibiofilm activity against C. albicans. This activity was dependent on concentration, species, and strain. A phytochemical investigation bioguided by anti-adhesion tests against the two Candida species was carried out on crude cranberry juice to determine the active fractions. Three subfractions enriched in proanthocyanidins showed an anti-adhesion activity at low concentrations. This study investigated for the first time the interest of crude extracts of cranberry and cranberry juice fractions to prevent biofilms of C. glabrata. It highlighted the potency of consuming this fruit and using it as a source of anti-adhesion agents. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

The Role of Isocitrate Lyase (ICL1) in the Metabolic Adaptation of Candida albicans Biofilms

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Ishola, Oluwaseun Ayodeji; Ting, Seng Yeat; Tabana, Yasser M; Ahmed, Mowaffaq Adam; Yunus, Muhammad Amir; Mohamed, Rafeezul; Lung Than, Leslie Thian; Sandai, Doblin

2016-01-01

Background A major characteristic of Candida biofilm cells that differentiates them from free-floating cells is their high tolerance to antifungal drugs. This high resistance is attributed to particular biofilm properties, including the accumulation of extrapolymeric substances, morphogenetic switching, and metabolic flexibility. Objectives This study evaluated the roles of metabolic processes (in particular the glyoxylate cycle) on biofilm formation, antifungal drug resistance, morphology, and cell wall components. Methods Growth, adhesion, biofilm formation, and cell wall carbohydrate composition were quantified for isogenic Candida albicans ICL1/ICL1, ICL1/icl1, and icl1/icl1 strains. The morphology and topography of these strains were compared by light microscopy and scanning electron microscopy. FKS1 (glucan synthase), ERG11 (14-α-demethylase), and CDR2 (efflux pump) mRNA levels were quantified using qRT-PCR. Results The ICL1/icl1 and icl1/icl1 strains formed similar biofilms and exhibited analogous drug-tolerance levels to the control ICL1/ICL1 strains. Furthermore, the drug sequestration ability of β-1, 3-glucan, a major carbohydrate component of the extracellular matrix, was not impaired. However, the inactivation of ICL1 did impair morphogenesis. ICL1 deletion also had a considerable effect on the expression of the FKS1, ERG11, and CDR2 genes. FKS1 and ERG11 were upregulated in ICL1/icl1 and icl1/icl1 cells throughout the biofilm developmental stages, and CDR2 was upregulated at the early phase. However, their expression was downregulated compared to the control ICL1/ICL1 strain. Conclusions We conclude that the glyoxylate cycle is not a specific determinant of biofilm drug resistance. PMID:27800147

How can in vitro models best reflect in vivo Staphylococcus biofilms?

DEFF Research Database (Denmark)

Meyer, Rikke Louise

In vitro biofilm models are the basis for most studies of biofilm biology because they enable high-throughput analyses without the expenditure of animals. But how do we ensure that what we learn from in vitro studies is relevant in vivo? Biofilms grown in standard laboratory media do not interact...... with host factors and are thus profoundly different from in vivo biofilms. We therefore need in vitro models that are as in vivo-like as possible. We investigated how the addition of divalent cations and human plasma to brain heart infusion broth affected biofilm formation by Staphylococcus aureus...

Candida albicans biofilms and MMA surface treatment influence the adhesion of soft denture liners to PMMA resin

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Martinna de Mendonça e Bertolini

2014-01-01

Full Text Available The effect of Candida albicans biofilms and methyl methacrylate (MMA pretreatment on the bond strength between soft denture liners and polymethyl methacrylate (PMMA resin was analyzed. Specimens were prepared and randomly divided with respect to PMMA pretreatment, soft liner type (silicone-based or PMMA-based, and presence or absence of a C. albicans biofilm. Samples were composed of a soft denture liner bonded between two PMMA bars. Specimens (n = 10 were incubated to produce a C. albicans biofilm or stored in sterile PBS for 12 days. The tensile bond strength test was performed and failure type was determined using a stereomicroscope. Surface roughness (SR and scanning electron microscopy (SEM analysis were performed on denture liners (n = 8. Highest bond strength was observed in samples containing a silicone-based soft liner and stored in PBS, regardless of pretreatment (p < 0.01. Silicone-based specimens mostly underwent adhesive failures, while samples containing PMMA-based liners predominantly underwent cohesive failures. The silicone-based specimens SR decreased after 12 days of biofilm accumulation or PBS storage, while the SR of PMMA-based soft liners increased (p < 0.01. The PMMA-based soft liners surfaces presented sharp valleys and depressions, while silicone-based specimens surfaces exhibited more gentle features. In vitro exposure to C. albicans biofilms reduced the adhesion of denture liners to PMMA resin, and MMA pretreatment is recommended during relining procedures.

Flexible camphor diamond-like carbon coating on polyurethane to prevent Candida albicans biofilm growth.

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Santos, Thaisa B; Vieira, Angela A; Paula, Luciana O; Santos, Everton D; Radi, Polyana A; Khouri, Sônia; Maciel, Homero S; Pessoa, Rodrigo S; Vieira, Lucia

2017-04-01

Camphor was incorporated in diamond-like carbon (DLC) films to prevent the Candida albicans yeasts fouling on polyurethane substrates, which is a material commonly used for catheter manufacturing. The camphor:DLC and DLC film for this investigation was produced by plasma enhanced chemical vapor deposition (PECVD), using an apparatus based on the flash evaporation of organic liquid (hexane) containing diluted camphor for camphor:DLC and hexane/methane, mixture for DLC films. The film was deposited at a low temperature of less than 25°C. We obtained very adherent camphor:DLC and DLC films that accompanied the substrate flexibility without delamination. The adherence of camphor:DLC and DLC films on polyurethane segments were evaluated by scratching test and bending polyurethane segments at 180°. The polyurethane samples, with and without camphor:DLC and DLC films were characterized by Raman spectroscopy, scanning electron microscopy, atomic force microscopy, and optical profilometry. Candida albicans biofilm formation on polyurethane, with and without camphor:DLC and DLC, was assessed. The camphor:DLC and DLC films reduced the biofilm growth by 99.0% and 91.0% of Candida albicans, respectively, compared to bare polyurethane. These results open the doors to studies of functionalized DLC coatings with biofilm inhibition properties used in the production of catheters or other biomedical applications. Copyright © 2017 Elsevier Ltd. All rights reserved.

Formation of In Vitro Mixed-Species Biofilms by Lactobacillus pentosus and Yeasts Isolated from Spanish-Style Green Table Olive Fermentations.

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León-Romero, Ángela; Domínguez-Manzano, Jesús; Garrido-Fernández, Antonio; Arroyo-López, Francisco Noé; Jiménez-Díaz, Rufino

2016-01-15

The present work details the in vitro interactions between Lactobacillus pentosus and yeast strains isolated from table olive processing to form mixed biofilms. Among the different pairs assayed, the strongest biofilms were obtained from L. pentosus and Candida boidinii strain cocultures. However, biofilm formation was inhibited in the presence of d-(+)-mannose. In addition, biofilm formation by C. boidinii monoculture was stimulated in the absence of cell-cell contact with L. pentosus. Scanning electron microscopy revealed that a sort of "sticky" material formed by the yeasts contributed to substrate adherence. Hence, the data obtained in this work suggest that yeast-lactobacilli biofilms may be favored by the presence of a specific mate of yeast and L. pentosus, and that more than one mechanism might be implicated in the biofilm formation. This knowledge will help in the design of appropriate mixed starter cultures of L. pentosus-yeast species pairs that are able to improve the quality and safety of Spanish-style green table olive processing. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Adhesion, biofilm formation, cell surface hydrophobicity, and antifungal planktonic susceptibility: relationship among Candida spp.

OpenAIRE

Silva-Dias, Ana; Miranda, Isabel M.; Branco, Joana; Monteiro-Soares, Matilde; Pina-Vaz, Cid?lia; Rodrigues, Ac?cio G.

2015-01-01

We have performed the characterization of the adhesion profile, biofilm formation, cell surface hydrophobicity (CSH) and antifungal susceptibility of 184 Candida clinical isolates obtained from different human reservoirs. Adhesion was quantified using a flow cytometric assay and biofilm formation was evaluated using two methodologies: XTT and crystal violet assay. CSH was quantified with the microbial adhesion to hydrocarbons test while planktonic susceptibility was assessed accordingly the C...

Anti-Biofilm Efficacy of Nitric Oxide-Releasing Silica Nanoparticles

OpenAIRE

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

2009-01-01

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

The essential oil of Allium sativum as an alternative agent against Candida isolated from dental prostheses.

Science.gov (United States)

Mendoza-Juache, Alejandro; Aranda-Romo, Saray; Bermeo-Escalona, Josué R; Gómez-Hernández, Araceli; Pozos-Guillén, Amaury; Sánchez-Vargas, Luis Octavio

The colonization of the surfaces of dental prostheses by Candida albicans is associated with the development of denture stomatitis. In this context, the use of fluconazole has been proposed, but its disadvantage is microbial resistance. Meanwhile, the oil of Allium sativum has shown an effect in controlling biofilm formation by C. albicans. The objective of this study was to determine the antifungal activities of the essential oil of A. sativum and fluconazole against clinical isolates of Candida species obtained from rigid, acrylic-based partial or total dentures and to compare these agents' effects on both biofilm and planktonic cells. A total of 48 clinical isolates obtained from the acrylic surface of partial or complete dentures were examined, and the following species were identified: C. albicans, Candida glabrata, Candida tropicalis, and Candida krusei. For each isolate, the antifungal activities of the essential oil of A. sativum and fluconazole against both biofilm and planktonic cells were evaluated using the Clinical & Laboratory Standards Institute (CLSI) M27-A3 method. The isolates were also evaluated by semiquantitative XTT reduction. All planktonic Candida isolates were susceptible to the essential oil of A. sativum, whereas 4.2% were resistant to fluconazole. Regarding susceptibilities in biofilms, 43.8% of biofilms were resistant to A. sativum oil, and 91.7% were resistant to fluconazole. All planktonic cells of the different Candida species tested are susceptible to sativum oil, and the majority are susceptible to fluconazole. Susceptibility decreases in biofilm cells, with increased resistance to fluconazole compared with A. sativum oil. The essential oil of A. sativum is thus active against clinical isolates of Candida species obtained from dentures, with effects on both biofilm and planktonic cells in vitro. Copyright © 2017 Asociación Española de Micología. Publicado por Elsevier España, S.L.U. All rights reserved.

The Impact of Biofilm Formation on the Persistence of Candidemia

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Wei-Sin Li

2018-06-01

Full Text Available This study aimed to determine the predictors of persistent candidemia and examine the impact of biofilm formation by Candida isolates in adult patients with candidemia. Of the adult patients with candidemia in Kaohsiung Chang Gung Memorial Hospital between January 2007 and December 2012, 68 case patients with persistent candidemia (repeated candidemia after a 3-day systemic antifungal therapy and 68 control patients with non-persistent candidemia (Candida clearance from the bloodstream after a 3-day systemic antifungal therapy were included based on propensity score matching and matching for the Candida species isolated. Biofilm formation by the Candida species was assessed in vitro using standard biomass assays. Presence of central venous catheters (CVCs at diagnosis (adjusted odd ratio [AOR], 3.77; 95% confidence interval [CI], 1.09–13.00, p = 0.04, infection with higher biofilm forming strains of Candida species (AOR, 8.03; 95% CI, 2.50–25.81; p < 0.01, and receipt of suboptimal fluconazole doses as initial therapy (AOR, 5.54; 95% CI, 1.53–20.10; p < 0.01 were independently associated with persistent candidemia. Biofilm formation by Candida albicans, C. tropicalis, and C. glabrata strains was significantly higher in the case patients than in the controls. There were no significant differences in the overall mortality and duration of hospitalization between the two groups. Our data suggest that, other than presence of retained CVCs and use of suboptimal doses of fluconazole, biofilm formation was highly associated with development of persistent candidemia.

Candida albicans biofilm on titanium: effect of peroxidase precoating

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Mohamed Ahariz

2010-08-01

Full Text Available Mohamed Ahariz1, Philippe Courtois1,21Laboratory of Experimental Hormonology, Université Libre de Bruxelles, Brussels, 2UER de Biologie Médicale, Haute Ecole Francisco Ferrer, Brussels, BelgiumAbstract: The present study aimed to document Candida albicans biofilm development on titanium and its modulation by a peroxidase-precoated material which can generate antimicrobials, such as hypoiodite or hypothiocyanite, from hydrogen peroxide, iodide, or thiocyanate. For this purpose, titanium (powder or foil was suspended in Sabouraud liquid medium inoculated with C. albicans ATCC10231. After continuous stirring for 2–21 days at room temperature, the supernatant was monitored by turbidimetry at 600 nm and titanium washed three times in sterile Sabouraud broth. Using the tetrazolium salt MTT-formazan assay, the titanium-adherent fungal biomass was measured as 7.50 ± 0.60 × 106 blastoconidia per gram of titanium powder (n = 30 and 0.50 ± 0.04 × 106 blastoconidia per cm² of titanium foil (n = 12. The presence of yeast on the surface of titanium was confirmed by microscopy both on fresh preparations and after calcofluor white staining. However, in the presence of peroxidase systems (lactoperoxidase with substrates such as hydrogen peroxide donor, iodide, or thiocyanate, Candida growth in both planktonic and attached phases appeared to be inhibited. Moreover, this study demonstrates the possible partition of peroxidase systems between titanium material (peroxidase-precoated and liquid environment (containing peroxidase substrates to limit C. albicans biofilm formation.Keywords: adhesion, material, oral, yeast

Identification, Typing, Antifungal Resistance Profile, and Biofilm Formation of Candida albicans Isolates from Lebanese Hospital Patients

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Ibrahim Bitar

2014-01-01

Full Text Available As leading opportunistic fungal pathogens identification and subtyping of Candida species are crucial in recognizing outbreaks of infection, recognizing particularly virulent strains, and detecting the emergence of drug resistant strains. In this study our objective was to compare identification of Candida albicans by the hospitals through the use of conventional versus identification based on the ITS (Internal Transcribed Spacer and to assess biofilm forming capabilities, drug resistance patterns and correlate these with MLST typing. ITS typing revealed a 21.2% hospital misidentification rate. Multidrug resistance to three drugs out of four tested was detected within 25% of the isolates raising concerns about the followed treatment regimens. Drug resistant strains as well as biofilm formers were phylogenetically related, with some isolates with significant biofilm forming capabilities being correlated to those that were multidrug resistant. Such isolates were grouped closely together in a neighbor-joining tree generated by MLST typing indicating phylogenetic relatedness, microevolution, or recurrent infection. In conclusion, this pilot study gives much needed insight concerning C. albicans isolates circulating in Lebanese hospitals and is the first study of its kind correlating biofilm formation, antifungal resistance, and evolutionary relatedness.

AI-2 of Aggregatibacter actinomycetemcomitans Inhibits Candida albicans Biofilm Formation

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Endang W. Bachtiar

2014-07-01

Full Text Available Aggregatibacter actinomycetemcomitans, a Gram-negative bacterium, and Candida albicans, a polymorphic fungus, are both commensals of the oral cavity but both are opportunistic pathogens that can cause oral diseases. A. actinomycetemcomitans produces a quorum-sensing molecule called autoinducer-2 (AI-2, synthesized by LuxS, that plays an important role in expression of virulence factors, in intra- but also in interspecies communication. The aim of this study was to investigate the role of AI-2 based signaling in the interactions between C. albicans and A. actinomycetemcomitans. A. actinomycetemcomitans adhered to C. albicans and inhibited biofilm formation by means of a molecule that was secreted during growth. C. albicans biofilm formation increased significantly when co-cultured with A. actinomycetemcomitans luxS, lacking AI-2 production. Addition of wild-type-derived spent medium or synthetic AI-2 to spent medium of the luxS strain, restored inhibition of C. albicans biofilm formation to wild-type levels. Addition of synthetic AI-2 significantly inhibited hypha formation of C. albicans possibly explaining the inhibition of biofilm formation. AI-2 of A. actinomycetemcomitans is synthesized by LuxS, accumulates during growth and inhibits C. albicans hypha- and biofilm formation. Identifying the molecular mechanisms underlying the interaction between bacteria and fungi may provide important insight into the balance within complex oral microbial communities.

Synergistic antifungal effect of chitosan-stabilized selenium nanoparticles synthesized by pulsed laser ablation in liquids against Candida albicans biofilms.

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Lara, Humberto H; Guisbiers, Gregory; Mendoza, Jonathan; Mimun, Lawrence C; Vincent, Brandy A; Lopez-Ribot, Jose L; Nash, Kelly L

2018-01-01

Candida albicans is a major opportunistic fungal pathogen. One of the most important virulence factors that contribute to the pathogenesis of candidiasis is its ability to form biofilms. A key characteristic of Candida biofilms is their resistance to antifungal agents. Due to significant morbidity and mortality rates related to biofilm-associated drug resistance, there is an urgency to develop novel nanotechnology-based approaches preventing biofilm-related infections. In this study, we report, for the first time, the synthesis of selenium nanoparticles by irradiating selenium pellets by nanosecond pulsed laser ablation in liquid chitosan as a capping agent. Synergy of the fungicidal effect of selenium nanoparticles and chitosan was quantified by the combination index theorem of Chou-Talalay. This drug combination resulted in a potent fungicidal effect against a preformed C. albicans biofilm in a dose-response manner. By advanced electron microscopy techniques, we documented the adhesive and permeabilizing properties of chitosan, therefore allowing selenium nanoparticles to enter as the cell wall of the yeast became disrupted and distorted. Most importantly, we demonstrated a potent quantitative synergistic effect when compounds such as selenium and chitosan are combined. These chitosan-stabilized selenium nanoparticles could be used for ex vivo applications such as sterilizers for surfaces and biomedical devices.

Real-time evaluation of two light delivery systems for photodynamic disinfection of Candida albicans biofilm in curved root canals.

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Sabino, C P; Garcez, A S; Núñez, S C; Ribeiro, M S; Hamblin, M R

2015-08-01

Antimicrobial photodynamic therapy (APDT) combined with endodontic treatment has been recognized as an alternative approach to complement conventional root canal disinfection methods on bacterial biofilms. We developed an in  vitro model of bioluminescent Candida albicans biofilm inside curved dental root canals and investigated the microbial reduction produced when different light delivery methods are employed. Each light delivery method was evaluated in respect to the light distribution provided inside curved root canals. After conventional endodontic preparation, teeth were sterilized before canals were contaminated by a bioluminescent strain of C. albicans (CEC789). Methylene blue (90 μM) was introduced into the canals and then irradiated (λ = 660 nm, P = 100 mW, beam diameter = 2 mm) with laser tip either in contact with pulp chamber or within the canal using an optical diffuser fiber. Light distribution was evaluated by CCD camera, and microbial reduction was monitored through bioluminescence imaging. Our findings demonstrated that the bioluminescent C. albicans biofilm model had good reproducibility and uniformity. Light distribution in dental tissue was markedly dependent on the light delivery system, and this strategy was directly related to microbial destruction. Both light delivery systems performed significant fungal inactivation. However, when irradiation was performed with optical diffuser fiber, microbial burden reduction was nearly 100 times more effective. Bioluminescence is an interesting real-time analysis to endodontic C. albicans biofilm inactivation. APDT showed to be an effective way to inactivate C. albicans biofilms. Diffuser fibers provided optimized light distribution inside curved root canals and significantly increased APDT efficiency.

Produção de fatores de virulência in vitro por espécies patogênicas do gênero Candida Production of virulence factors in vitro by pathogenic species of the genus Candida

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Kelly Cristina Ortolan Rörig

2009-04-01

Full Text Available Avaliou-se, in vitro, a capacidade de crescimento em 39ºC e 42ºC, a produção de enzimas hidrolíticas e a atividade hemolítica de 21 cepas clínicas e de referência de sete espécies de Candida spp, Candida dubliniensis e Candida krusei demonstraram menor potencial de virulência e Candida albicans maior.The growth capacity at 39ºC and 42ºC, production of hydrolytic enzymes and hemolytic activity of 21 clinical and reference strains of seven species of Candida spp were evaluated in vitro.Candida dubliniensis and Candida krusei demonstrated lower virulence potential and Candida albicans higher potential.

In vitro phenotypic differentiation towards commensal and pathogenic oral biofilms

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Janus, M.M.; Keijser, B.J.F.; Bikker, F.J.; Exterkate, R.A.M.; Crielaard, W.; Krom, B.P.

2015-01-01

Commensal oral biofilms, defined by the absence of pathology-related phenotypes, are ubiquitously present. In contrast to pathological biofilms commensal biofilms are rarely studied. Here, the effect of the initial inoculum and subsequent growth conditions on in vitro oral biofilms was studied.

Biophysical Effects of a Polymeric Biosurfactant in Candida krusei and Candida albicans Cells.

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Ferreira, Gabriella Freitas; Dos Santos Pinto, Bruna Lorrana; Souza, Eliene Batista; Viana, José Lima; Zagmignan, Adrielle; Dos Santos, Julliana Ribeiro Alves; Santos, Áquila Rodrigues Costa; Tavares, Priscila Batista; Denadai, Ângelo Márcio Leite; Monteiro, Andrea Souza

2016-12-01

This study evaluated the effects of a polymeric biosurfactant produced by Trichosporon montevideense CLOA72 in the adhesion of Candida albicans and Candida krusei cells to human buccal epithelial cells and its interference in biofilm formation by these strains. The biofilm inhibition by biosurfactant (25 mg/mL) in C. krusei and C. albicans in polystyrene was reduced up to 79.5 and 85 %, respectively. In addition, the zeta potential and hydrodynamic diameter of the yeasts altered as a function of the biosurfactant concentration added to the cell suspension. The changes in the cell surface characteristics and the interface modification can contribute to the inhibition of the initial adherence of yeasts cells to the surface. In addition, the analyses of the biofilm matrix and planktonic cell surfaces demonstrated differences in carbohydrate and protein concentrations for the two studied strains, which may contribute to the modulation of cell adhesion or consolidation of biofilms, especially in C. krusei. This study suggests a possible application of the of CLOA72 biosurfactant in inhibiting the adhesion and formation of biofilms on biological surfaces by yeasts of the Candida genus.

Proteus vulgaris and Proteus mirabilis Decrease Candida albicans Biofilm Formation by Suppressing Morphological Transition to Its Hyphal Form.

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Lee, Kyoung Ho; Park, Su Jung; Choi, Sun Ju; Park, Joo Young

2017-11-01

Candida albicans (C. albicans) and Proteus species are causative agents in a variety of opportunistic nosocomial infections, and their ability to form biofilms is known to be a virulence factor. In this study, the influence of co-cultivation with Proteus vulgaris (P. vulgaris) and Proteus mirabilis (P. mirabilis) on C. albicans biofilm formation and its underlying mechanisms were examined. XTT reduction assays were adopted to measure biofilm formation, and viable colony counts were performed to quantify yeast growth. Real-time reverse transcriptase polymerase chain reaction was used to evaluate the expression of yeast-specific genes (rhd1 and rbe1), filament formation inhibiting genes (tup1 and nrg1), and hyphae-related genes (als3, ece1, hwp1, and sap5). Candida biofilm formation was markedly inhibited by treatment with either living or heat-killed P. vulgaris and P. mirabilis. Proteus-cultured supernatant also inhibited Candida biofilm formation. Likewise, treatment with live P. vulgaris or P. mirabilis or with Proteus-cultured supernatant decreased expression of hyphae-related C. albicans genes, while the expression of yeast-specific genes and the filament formation inhibiting genes of C. albicans were increased. Heat-killed P. vulgaris and P. mirabilis treatment, however, did not affect the expression of C. albicans morphology-related genes. These results suggest that secretory products from P. vulgaris and P. mirabilis regulate the expression of genes related to morphologic changes in C. albicans such that transition from the yeast form to the hyphal form can be inhibited. © Copyright: Yonsei University College of Medicine 2017

Características da formação do biofilme de Candida tropicalis eresistência a antifúngicos

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Fernando César Bizerra

2006-01-01

Candida tropicalis é um dos principais agentes de candidíase, destacando-se em casos de infecções da corrente sanguínea e urinária em pacientes hospitalizados. Grande parte das infecções por Candida spp. estão associadas com a formação de biofilme na superfície de materiais médicos ou epitélio do hospedeiro. As células sésseis que constituem o biofilme apresentam fenótipos drasticamente diferentes das células planctônicas, tais como resistência aos agentes antimicrobianos e aos mecanismos de ...

Adhesion, biofilm formation, cell surface hydrophobicity and antifungal planktonic susceptibility: relationship among Candida spp.

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Ana Isabel Silva-Dias

2015-03-01

Full Text Available We have performed the characterization of the adhesion profile, biofilm formation, cell surface hydrophobicity (CSH and antifungal susceptibility of 184 Candida clinical isolates obtained from different human reservoirs. Adhesion was quantified using a flow cytometric assay and biofilm formation was evaluated using two methodologies: XTT and crystal violet assay. CSH was quantified with the microbial adhesion to hydrocarbons test while planktonic susceptibility was assessed accordingly the CLSI protocol for yeast M27-A3 S4.Yeast cells of non-albicans species exhibit increased ability to adhere and form biofilm. However the correlation between adhesion and biofilm formation varied according to species and also with the methodology used for biofilm assessment. No association was found between strain´s site of isolation or planktonic antifungal susceptibility and adhesion or biofilm formation. Finally CSH seemed to be a good predictor for biofilm formation but not for adhesion.Despite the marked variability registered intra and inter species, C. tropicalis and C. parapsilosis were the species exhibiting high adhesion profile. C. tropicalis, C. guilliermondii and C. krusei revealed higher biofilm formation values in terms of biomass. C. parapsilosis was the species with lower biofilm metabolic activity.

Adhesion, biofilm formation, cell surface hydrophobicity, and antifungal planktonic susceptibility: relationship among Candida spp.

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Silva-Dias, Ana; Miranda, Isabel M; Branco, Joana; Monteiro-Soares, Matilde; Pina-Vaz, Cidália; Rodrigues, Acácio G

2015-01-01

We have performed the characterization of the adhesion profile, biofilm formation, cell surface hydrophobicity (CSH) and antifungal susceptibility of 184 Candida clinical isolates obtained from different human reservoirs. Adhesion was quantified using a flow cytometric assay and biofilm formation was evaluated using two methodologies: XTT and crystal violet assay. CSH was quantified with the microbial adhesion to hydrocarbons test while planktonic susceptibility was assessed accordingly the CLSI protocol for yeast M27-A3 S4. Yeast cells of non-albicans species exhibit increased ability to adhere and form biofilm. However, the correlation between adhesion and biofilm formation varied according to species and also with the methodology used for biofilm assessment. No association was found between strain's site of isolation or planktonic antifungal susceptibility and adhesion or biofilm formation. Finally CSH seemed to be a good predictor for biofilm formation but not for adhesion. Despite the marked variability registered intra and inter species, C. tropicalis and C. parapsilosis were the species exhibiting high adhesion profile. C. tropicalis, C. guilliermondii, and C. krusei revealed higher biofilm formation values in terms of biomass. C. parapsilosis was the species with lower biofilm metabolic activity.

Antifungal effects of undecylenic acid on the biofilm formation of Candida albicans.

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Shi, Dongmei; Zhao, Yaxin; Yan, Hongxia; Fu, Hongjun; Shen, Yongnian; Lu, Guixia; Mei, Huan; Qiu, Ying; Li, Dongmei; Liu, Weida

2016-05-01

Undecylenic acid can effectively control skin fungal infection, but the mechanism of its fungal inhibition is unclear. Hyphal growth of Candida albicans (C. albicans) and biofilm formation have been well recognized as important virulence factors for the initiation of skin infection and late development of disseminated infection. In this study, we seek to investigate antifungal mechanisms of undecylenic acid by evaluating the virulence factors of C. albicans during biofilm formation. We found that undecylenic acid inhibits biofilm formation of C. albicans effectively with optimal concentration above 3 mM. In the presence of this compound, the morphological transition from yeast to filamentous phase is abolished ultimately when the concentration of undecylenic acid is above 4 mM. Meanwhile, the cell surface is crumpled, and cells display an atrophic appearance under scanning electron microscopy even with low concentration of drug treatment. On the other hand, the drug treatment decreases the transcriptions of hydrolytic enzymes such as secreted aspartic protease, lipase, and phospholipase. Hyphal formation related genes, like HWP1, are significantly reduced in transcriptional level in drug-treated biofilm condition as well. The down-regulated profile of these genes leads to a poorly organized biofilm in undecylenic acid treated environment.

Real-time PCR expression profiling of genes encoding potential virulence factors in Candida albicans biofilms: identification of model-dependent and -independent gene expression

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Řičicová Markéta

2010-04-01

Full Text Available Abstract Background Candida albicans infections are often associated with biofilm formation. Previous work demonstrated that the expression of HWP1 (hyphal wall protein and of genes belonging to the ALS (agglutinin-like sequence, SAP (secreted aspartyl protease, PLB (phospholipase B and LIP (lipase gene families is associated with biofilm growth on mucosal surfaces. We investigated using real-time PCR whether genes encoding potential virulence factors are also highly expressed in biofilms associated with abiotic surfaces. For this, C. albicans biofilms were grown on silicone in microtiter plates (MTP or in the Centres for Disease Control (CDC reactor, on polyurethane in an in vivo subcutaneous catheter rat (SCR model, and on mucosal surfaces in the reconstituted human epithelium (RHE model. Results HWP1 and genes belonging to the ALS, SAP, PLB and LIP gene families were constitutively expressed in C. albicans biofilms. ALS1-5 were upregulated in all model systems, while ALS9 was mostly downregulated. ALS6 and HWP1 were overexpressed in all models except in the RHE and MTP, respectively. The expression levels of SAP1 were more pronounced in both in vitro models, while those of SAP2, SAP4 and SAP6 were higher in the in vivo model. Furthermore, SAP5 was highly upregulated in the in vivo and RHE models. For SAP9 and SAP10 similar gene expression levels were observed in all model systems. PLB genes were not considerably upregulated in biofilms, while LIP1-3, LIP5-7 and LIP9-10 were highly overexpressed in both in vitro models. Furthermore, an elevated lipase activity was detected in supernatans of biofilms grown in the MTP and RHE model. Conclusions Our findings show that HWP1 and most of the genes belonging to the ALS, SAP and LIP gene families are upregulated in C. albicans biofilms. Comparison of the fold expression between the various model systems revealed similar expression levels for some genes, while for others model-dependent expression

Biofilms of vaginal Lactobacillus in vitro test.

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Wei, Xiao-Yu; Zhang, Rui; Xiao, Bing-Bing; Liao, Qin-Ping

2017-01-01

This paper focuses on biofilms of Lactobacillus spp. - a type of normal flora isolated from healthy human vaginas of women of childbearing age; thereupon, it broadens the research scope of investigation of vaginal normal flora. The static slide culture method was adopted to foster biofilms, marked by specific fluorescence staining. Laser scanning confocal and scanning electron microscopy were used to observe the microstructure of the biofilms. Photographs taken from the microstructure were analysed to calculate the density of the biofilms. The body of Lactobacillus spp., though red, turned yellow when interacting with the green extracellular polysaccharides. The structure of the biofilm and aquaporin within the biofilm were imaged. Lactobacillus density increases over time. This study provides convincing evidence that Lactobacillus can form biofilms and grow over time in vitro. This finding establishes an important and necessary condition for selecting proper strains for the pharmaceutics of vaginal ecology.

A Candida biofilm-induced pathway for matrix glucan delivery: implications for drug resistance.

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Heather T Taff

Full Text Available Extracellular polysaccharides are key constituents of the biofilm matrix of many microorganisms. One critical carbohydrate component of Candida albicans biofilms, β-1,3 glucan, has been linked to biofilm protection from antifungal agents. In this study, we identify three glucan modification enzymes that function to deliver glucan from the cell to the extracellular matrix. These enzymes include two predicted glucan transferases and an exo-glucanase, encoded by BGL2, PHR1, and XOG1, respectively. We show that the enzymes are crucial for both delivery of β-1,3 glucan to the biofilm matrix and for accumulation of mature matrix biomass. The enzymes do not appear to impact cell wall glucan content of biofilm cells, nor are they necessary for filamentation or biofilm formation. We demonstrate that mutants lacking these genes exhibit enhanced susceptibility to the commonly used antifungal, fluconazole, during biofilm growth only. Transcriptional analysis and biofilm phenotypes of strains with multiple mutations suggest that these enzymes act in a complementary fashion to distribute matrix downstream of the primary β-1,3 glucan synthase encoded by FKS1. Furthermore, our observations suggest that this matrix delivery pathway works independently from the C. albicans ZAP1 matrix formation regulatory pathway. These glucan modification enzymes appear to play a biofilm-specific role in mediating the delivery and organization of mature biofilm matrix. We propose that the discovery of inhibitors for these enzymes would provide promising anti-biofilm therapeutics.

Antifungal and Anti-Biofilm Activities of Acetone Lichen Extracts against Candida albicans

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Marion Millot

2017-04-01

Full Text Available Candida albicans is a commensal coloniser of the human gastrointestinal tract and an opportunistic pathogen, especially thanks to its capacity to form biofilms. This lifestyle is frequently involved in infections and increases the yeast resistance to antimicrobials and immune defenses. In this context, 38 lichen acetone extracts have been prepared and evaluated for their activity against C. albicans planktonic and sessile cells. Minimum inhibitory concentrations of extracts (MICs were determined using the broth microdilution method. Anti-biofilm activity was evaluated using tetrazolium salt (XTT assay as the ability to inhibit the maturation phase (anti-maturation or to eradicate a preformed 24 h old biofilm (anti-biofilm. While none of the extracts were active against planktonic cells, biofilm maturation was limited by 11 of the tested extracts. Seven extracts displayed both anti-maturation and anti-biofilm activities (half maximal inhibitory concentrations IC50_mat and IC50_biof ≤ 100 µg/mL; Evernia prunastri and Ramalina fastigiata were the most promising lichens (IC50_mat < 4 µg/mL and IC50_biof < 10 µg/mL. Chemical profiles of the active extracts performed by thin layer chromatography (TLC and high performance liquid chromatography (HPLC have been analyzed. Depsides, which were present in large amounts in the most active extracts, could be involved in anti-biofilm activities. This work confirmed that lichens represent a reservoir of compounds with anti-biofilm potential.

In vitro phagocytosis of several Candida berkhout species by murine leukocytes.

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Fontenla de Petrino, S E; Bibas Bonet de Jorrat, M E; Sirena, A

1985-03-01

In vitro phagocytosis of thirteen Candida berkhout species by rat leukocytes was studied to assess a possible correlation between pathogenicity and phagocytosis Yeast-leukocyte suspensions were mixed up for 3 h and phagocytic index, germ-tube formation and leukocyte candidacidal activity were evaluated. Highest values for phagocytosis were reached in all cases at the end of the first hour. Leukocyte candidacidal activity was absent. Only C. albicans produced germ-tubes. The various phagocytosis indices were determined depending on the Candida species assayed. Under these conditions, the more pathogenic species presented the lower indices of phagocytosis. It is determined that the in vitro phagocytic index may bear a close relationship with the pathogenicity of the Candida berkhout.

Influência de Lactobacillus rhamnosus na patogenicidade e na expressão de genes de virulência de Candida albicans: estudo in vitro e in vivo

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Ribeiro, Felipe de Camargo [UNESP

2015-01-01

The high incidence of candidiasis caused by Candida albicans and the adaptability of this species, as well as resistance to antifungal drive the development of research on alternative therapies to control this infection. The aim of this study was to evaluate the influence of Lactobacillus rhamnosus and products of their metabolism against C. albicans, evaluating the pathogenicity and the expression of genes that regulate the formation of C. albicans biofilms in vitro and in vivo in invertebra...

In Vitro Antifungal Susceptibility of Oral Candida Isolates from Patients Suffering from Caries and Chronic Periodontitis.

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De-la-Torre, Janire; Ortiz-Samperio, María Esther; Marcos-Arias, Cristina; Marichalar-Mendia, Xabier; Eraso, Elena; Echebarria-Goicouria, María Ángeles; Aguirre-Urizar, José Manuel; Quindós, Guillermo

2017-06-01

Caries and chronic periodontitis are common oral diseases where a higher Candida colonization is reported. Antifungal agents could be adjuvant drugs for the therapy of both clinical conditions. The aim of the current study has been to evaluate the in vitro activities of conventional and new antifungal drugs against oral Candida isolates from patients suffering from caries and/or chronic periodontitis. In vitro activities of amphotericin B, fluconazole, itraconazole, miconazole, nystatin, posaconazole and voriconazole against 126 oral Candida isolates (75 Candida albicans, 18 Candida parapsilosis, 11 Candida dubliniensis, six Candida guilliermondii, five Candida lipolytica, five Candida glabrata, four Candida tropicalis and two Candida krusei) from 61 patients were tested by the CLSI M27-A3 method. Most antifungal drugs were highly active, and resistance was observed in less than 5% of tested isolates. Miconazole was the most active antifungal drug, being more than 98% of isolates susceptible. Fluconazole, itraconazole, and the new triazoles, posaconazole and voriconazole, were also very active. Miconazole, fluconazole and voriconazole have excellent in vitro activities against all Candida isolates and could represent suitable treatment for a hypothetically adjunctive therapy of caries and chronic periodontitis.

Commensal Protection of Staphylococcus aureus against Antimicrobials by Candida albicans Biofilm Matrix

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Kong, Eric F.; Tsui, Christina; Kucharíková, Sona; Andes, David

2016-01-01

ABSTRACT Biofilm-associated polymicrobial infections, particularly those involving fungi and bacteria, are responsible for significant morbidity and mortality and tend to be challenging to treat. Candida albicans and Staphylococcus aureus specifically are considered leading opportunistic fungal and bacterial pathogens, respectively, mainly due to their ability to form biofilms on catheters and indwelling medical devices. However, the impact of mixed-species biofilm growth on therapy remains largely understudied. In this study, we investigated the influence of C. albicans secreted cell wall polysaccharides on the response of S. aureus to antibacterial agents in biofilm. Results demonstrated significantly enhanced tolerance for S. aureus to drugs in the presence of C. albicans or its secreted cell wall polysaccharide material. Fluorescence confocal time-lapse microscopy revealed impairment of drug diffusion through the mixed biofilm matrix. Using C. albicans mutant strains with modulated cell wall polysaccharide expression, exogenous supplementation, and enzymatic degradation, the C. albicans-secreted β-1,3-glucan cell wall component was identified as the key matrix constituent providing the bacteria with enhanced drug tolerance. Further, antibody labeling demonstrated rapid coating of the bacteria by the C. albicans matrix material. Importantly, via its effect on the fungal biofilm matrix, the antifungal caspofungin sensitized the bacteria to the drugs. Understanding such symbiotic interactions with clinical relevance between microbial species in biofilms will greatly aid in overcoming the limitations of current therapies and in defining potential new targets for treating polymicrobial infections. PMID:27729510

The anthraquinones rubiadin and its 1-methyl ether isolated from Heterophyllaea pustulata reduces Candida tropicalis biofilms formation.

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Marioni, Juliana; da Silva, María Angel; Cabrera, José Luis; Montoya, Susana C Núñez; Paraje, María Gabriela

2016-11-15

Candida tropicalis is increasingly becoming among the most commonly isolated pathogens causing fungal infections with an important biofilm-forming capacity. This study addresses the antifungal effect of rubiadin (AQ1) and rubiadin 1-methyl ether (AQ2), two photosensitizing anthraquinones (AQs) isolated from Heterophyllaea pustulata, against C. tropicalis biofilms, by studying the cellular stress and antioxidant response in two experimental conditions: darkness and irradiation. The combination with Amphotericin B (AmB) was assayed to evaluate the synergic effect. Biofilms of clinical isolates and reference strain of Candida tropicalis were treated with AQs (AQ1 or AQ2) and/or AmB, and the biofilms depletion was studied by crystal violet and confocal scanning laser microscopy (CSLM). The oxidant metabolites production and the response of antioxidant defense system were also evaluated under dark and irradiation conditions, being the light a trigger for photo-activation of the AQs. The Reactive Oxygen Species (ROS) were detected by the reduction of Nitro Blue Tetrazolium test, and Reactive Nitrogen Intermediates (RNI) by the Griess assay. ROS accumulation was also detected inside biofilms by using 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) probe, which was visualized by CSLM. Superoxide dismutase (SOD) activity and the total antioxidant capacity of biofilms were measured by spectrophotometric methods. The minimun inhibitory concentration for sessile cells (SMIC) was determined for each AQs and AmB. The fractional inhibitory concentration index (FICI) was calculated for the combinations of each AQ with AmB by the checkerboard microdilution method. Biofilm reduction of both strains was more effective with AQ1 than with AQ2. The antifungal effect was mediated by an oxidative and nitrosative stress under irradiation, with a significant accumulation of endogenous ROS detected by CSLM and an increase in the SOD activity. Thus, the prooxidant-antioxidant balance was

Psd1 Effects on Candida albicans Planktonic Cells and Biofilms

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Sónia Gonçalves

2017-06-01

Full Text Available Candida albicans is an important human pathogen, causing opportunistic infections. The adhesion of planktonic cells to a substrate is the first step for biofilm development. The antimicrobial peptide (AMP Psd1 is a defensin isolated from Pisum sativum seeds. We tested the effects of this AMP on C. albicans biofilms and planktonic cells, comparing its activity with amphotericin B and fluconazole. Three C. albicans variants were studied, one of them a mutant deficient in glucosylceramide synthase, conferring resistance to Psd1 antifungal action. Atomic force microscopy (AFM was used to assess morphological and biomechanical changes on fungal cells. Surface alterations, with membrane disruption and leakage of cellular contents, were observed. Cytometry assays and confocal microscopy imaging showed that Psd1 causes cell death, in a time and concentration-dependent manner. These results demonstrate Psd1 pleiotropic action against a relevant fungal human pathogen, suggesting its use as natural antimycotic agent.

Fungal Biofilms: In Vivo Models for Discovery of Anti-Biofilm Drugs.

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Nett, Jeniel E; Andes, David R

2015-06-01

During infection, fungi frequently transition to a biofilm lifestyle, proliferating as communities of surface-adherent aggregates of cells. Phenotypically, cells in a biofilm are distinct from free-floating cells. Their high tolerance of antifungals and ability to withstand host defenses are two characteristics that foster resilience. Biofilm infections are particularly difficult to eradicate, and most available antifungals have minimal activity. Therefore, the discovery of novel compounds and innovative strategies to treat fungal biofilms is of great interest. Although many fungi have been observed to form biofilms, the most well-studied is Candida albicans. Animal models have been developed to simulate common Candida device-associated infections, including those involving vascular catheters, dentures, urinary catheters, and subcutaneous implants. Models have also reproduced the most common mucosal biofilm infections: oropharyngeal and vaginal candidiasis. These models incorporate the anatomical site, immune components, and fluid dynamics of clinical niches and have been instrumental in the study of drug resistance and investigation of novel therapies. This chapter describes the significance of fungal biofilm infections, the animal models developed for biofilm study, and how these models have contributed to the development of new strategies for the eradication of fungal biofilm infections.

Impact of Environmental Conditions on the Form and Function of Candida albicans Biofilms

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Daniels, Karla J.; Park, Yang-Nim; Srikantha, Thyagarajan; Pujol, Claude

2013-01-01

Candida albicans, like other pathogens, can form complex biofilms on a variety of substrates. However, as the number of studies of gene regulation, architecture, and pathogenic traits of C. albicans biofilms has increased, so have differences in results. This suggests that depending upon the conditions employed, biofilms may vary widely, thus hampering attempts at a uniform description. Gene expression studies suggest that this may be the case. To explore this hypothesis further, we compared the architectures and traits of biofilms formed in RPMI 1640 and Spider media at 37°C in air. Biofilms formed by a/α cells in the two media differed to various degrees in cellular architecture, matrix deposition, penetrability by leukocytes, fluconazole susceptibility, and the facilitation of mating. Similar comparisons of a/a cells in the two media, however, were made difficult given that in air, although a/a cells form traditional biofilms in RPMI medium, they form polylayers composed primarily of yeast cells in Spider medium. These polylayers lack an upper hyphal/matrix region, are readily penetrated by leukocytes, are highly fluconazole susceptible, and do not facilitate mating. If, however, air is replaced with 20% CO2, a/a cells make a biofilm in Spider medium similar architecturally to that of a/α cells, which facilitates mating. A second, more cursory comparison is made between the disparate cellular architectures of a/a biofilms formed in air in RPMI and Lee's media. The results demonstrate that C. albicans forms very different types of biofilms depending upon the composition of the medium, level of CO2 in the atmosphere, and configuration of the MTL locus. PMID:23954841

Effects of nanosecond pulsed electric fields (nsPEFs) on the human fungal pathogen Candida albicans: an in vitro study

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Guo, Jinsong; Dang, Jie; Wang, Kaile; Zhang, Jue; Fang, Jing

2018-05-01

Candida albicans is the leading human fungal pathogen that causes many life-threatening infections. Notably, the current clinical trial data indicate that Candida species shows the emerging resistance to anti-fungal drugs. The aim of this study was to evaluate the antifungal effects of nanosecond pulsed electric fields (nsPEFs) as a novel drug-free strategy in vitro. In this study, we investigated the inactivation and permeabilization effects of C. albicans under different nsPEFs exposure conditions (100 pulses, 100 ns in duration, intensities of 20, 40 kV cm‑1). Cell death was studied by annexin-V and propidium iodide staining. The changes of intracellular Ca2+ concentration after nsPEFs treatment were observed using Fluo-4 AM. Results show that C. albicans cells and biofilms were both obviously inhibited and destroyed after nsPEFs treatment. Furthermore, C. albicans cells were significantly permeabilized after nsPEFs treatment. Additionally, nsPEFs exposure led to a large amount of DNA and protein leakage. Importantly, nsPEFs induced a field strength-dependent apoptosis in C. albicans cells. Further experiments revealed that Ca2+ involved in nsPEFs induced C. albicans apoptosis. In conclusion, this proof-of-concept study provides a potential alternative drug-free strategy for killing pathogenic Candida species.

Evaluation of Antimicrobial and Antifungal efficacy of Chitosan as endodontic irrigant against Enterococcus Faecalis and Candida Albicans Biofilm formed on tooth substrate.

Science.gov (United States)

Yadav, Pankaj; Chaudhary, Sarika; Saxena, Rajendra K; Talwar, Sangeeta; Yadav, Sudha

2017-03-01

Bacterial biofilms formed on the root canal wall are often difficult to remove. This study aimed to evaluate the cytotoxic effect and antibacterial efficacy of chitosan when used as root canal irrigant against E. Faecalis and Candida albicans biofilm formed on tooth substrate. The present study evaluated antibacterial effect of 0.25% Chitosan, 0.5% Chitosan, 2% chlorhexidine and 3% sodium hypochlorite against Enterococcus faecalis and Candida Albicans . Agar-well diffusion methods, minimal inhibitory concentration tests and biofilm susceptibility assays were used to determine antibacterial activity. Teeth specimens were sectioned to obtain a standardized tooth length of 12mm. Specimens were inoculated with 10 mL of the freshly prepared E. Faecalis suspension and Candida albicans for 4 weeks. The specimens were then instrumented with ProTaper rotary files F3 size. After irrigation with test solution, three sterile paper points were placed into one canal, left for 60 s and transferred to a test tube containing 1 mL of reduced transport fluid. The number of CFU in 1 mL was determined. 3-week biofilm qualitative assay showed complete inhibition of bacterial growth with 3% Sodium hypochlorite, 2% Chlorhexidine and Chitosan except saline, which showed presence of bacterial growth. Significant reduction of colony forming units (CFU)/mL was observed for the chitosan groups and the antibacterial activity of the chitosan groups was at par with 3% NaOCl and 2% Chlorhexidine. It was observed that the chitosan showed no cytotoxicity at 3mg/ml and 10% cytotoxicity at 6mg/ml. The use of chitosan as a root canal irrigant might be an alternative considering the various undesirable properties of NaOCl and chlorhexidine. Key words: Biofilm, Candida albicans, Chitosan, Cytotoxicity, Enterococcus faecalis.

Culture Supernatants of Lactobacillus gasseri and L. crispatus Inhibit Candida albicans Biofilm Formation and Adhesion to HeLa Cells.

Science.gov (United States)

Matsuda, Yuko; Cho, Otomi; Sugita, Takashi; Ogishima, Daiki; Takeda, Satoru

2018-03-30

Vulvovaginal candidiasis (VVC) is a common superficial infection of the vaginal mucous membranes caused by the fungus Candida albicans. The aim of this study was to assess the mechanisms underlying the inhibitory effects of the culture supernatants of Lactobacillus gasseri and L. crispatus, the predominant microbiota in Asian healthy women, on C. albicans biofilm formation. The inhibition of C. albicans adhesion to HeLa cells by Lactobacillus culture supernatant was also investigated. Candida albicans biofilm was formed on polystyrene flat-bottomed 96-well plates, and the inhibitory effects on the initial colonization and maturation phases were determined using the XTT reduction assay. The expression levels of biofilm formation-associated genes (HWP1, ECE1, ALS3, BCR1, EFG1, TEC1, and CPH1) were determined by reverse transcription quantitative polymerase chain reaction. The inhibition of C. albicans adhesion to HeLa cells by Lactobacillus culture supernatant was evaluated by enumerating viable C. albicans cells. The culture supernatants of both Lactobacillus species inhibited the initial colonization and maturation of C. albicans biofilm. The expression levels of all biofilm formation-related genes were downregulated in the presence of Lactobacillus culture supernatant. The culture supernatant also inhibited C. albicans adhesion to HeLa cells. The culture supernatants of L. gasseri and L. crispatus inhibited C. albicans biofilm formation by downregulating biofilm formation-related genes and C. albicans adhesion to HeLa cells. These findings support the notion that Lactobacillus metabolites may be useful alternatives to antifungal drugs for the management of VVC.

Differential effects of antifungal agents on expression of genes related to formation of Candida albicans biofilms.

Science.gov (United States)

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

2016-01-01

The purpose of this study was to analyse specific molecular mechanisms involved in the intrinsic resistance of C. albicans biofilms to antifungals. We investigated the transcriptional profile of three genes (BGL2, SUN41, ECE1) involved in Candida cell wall formation in response to voriconazole or anidulafungin after the production of intermediate and mature biofilms. C. albicans M61, a well-documented biofilm producer strain, was used for the development of intermediate (12 h and 18 h) and completely mature biofilms (48 h). After exposure of cells from each biofilm growth mode to voriconazole (128 and 512 mg l(-1)) or anidulafungin (0.25 and 1 mg l(-1)) for 12-24 h, total RNA samples extracted from biofilm cells were analysed by RT-PCR. The voriconazole and anidulafungin biofilm MIC was 512 and 0.5 mg l(-1) respectively. Anidulafungin caused significant up-regulation of SUN41 (3.7-9.3-fold) and BGL2 (2.2-2.8 fold) in intermediately mature biofilms; whereas, voriconazole increased gene expression in completely mature biofilms (SUN41 2.3-fold, BGL2 2.1-fold). Gene expression was primarily down-regulated by voriconazole in intermediately, but not completely mature biofilms. Both antifungals caused down-regulation of ECE1 in intermediately mature biofilms. © 2015 Blackwell Verlag GmbH.

Mimicking disinfection and drying of biofilms in contaminated endoscopes

NARCIS (Netherlands)

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

2010-01-01

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

Penicillenols from a deep-sea fungus Aspergillus restrictus inhibit Candida albicans biofilm formation and hyphal growth.

Science.gov (United States)

Wang, Jie; Yao, Qi-Feng; Amin, Muhammad; Nong, Xu-Hua; Zhang, Xiao-Yong; Qi, Shu-Hua

2017-06-01

Penicillenols (A1, A2, B1, B2, C1 and C2) were isolated from Aspergillus restrictus DFFSCS006, and could differentially inhibit biofilm formation and eradicate pre-developed biofilms of Candida albicans. Their structure-bioactivity relationships suggested that the saturation of hydrocarbon chain at C-8, R-configuration of C-5 and trans-configuration of the double bond between C-5 and C-6 of pyrrolidine-2,4-dione unit were important for their anti-biofilm activities. Penicillenols A2 and B1 slowed the hyphal growth and suppressed the transcripts of hypha specific genes HWP1, ALS1, ALS3, ECE1 and SAP4. Moreover, penicillenols A2 and B1 were found to act synergistically with amphotericin B against C. albicans biofilm formation.

[Activity of amphotericin B and anidulafungin, alone and combined, against Candida tropicalis biofilms developed on Teflon® and titanium].

Science.gov (United States)

Fernández-Rivero, Marcelo Ernesto; Del Pozo, José L; Valentín, Amparo; Fornes, Victoria; Molina de Diego, Araceli; Pemán, Javier; Cantón, Emilia

Current therapeutic strategies have a limited efficacy against Candida biofilms that form on the surfaces of biomedical devices. Few studies have evaluated the activity of antifungal agents against Candida tropicalis biofilms. To evaluate the activity of amphotericin B (AMB) and anidulafungin (AND), alone and in combination, against C. tropicalis biofilms developed on polytetrafluoroethylene (teflon -PTFE) and titanium surfaces using time-kill assays. Assays were performed using the CDC Biofilm Reactor equipped with PTFE and titanium disks with C. tropicalis biofilms after 24h of maturation. The concentrations assayed were 40mg/l for AMB and 8mg/l for AND, both alone and combined. After 24, 48 and 72h of exposure to the antifungals, the cfu/cm 2 was determined by a vortexing-sonication procedure. AMB reduced biofilm viable cells attached to PTFE and titanium by ≥99% and AND by 89.3% on PTFE and 96.8% on titanium. The AMB+AND combination was less active than AMB alone, both on PTFE (decrease of cfu/cm 2 3.09 Log 10 vs. 1.08 when combined) and titanium (4.51 vs. 1.53 when combined), being the interaction irrelevant on both surfaces. AMB is more active than AND against C. tropicalis biofilms. Yeast killing rates are higher on titanium than on PTFE surfaces. The combination of AMB plus AND is less effective than AMB alone on both surfaces. Copyright © 2017 Asociación Española de Micología. Publicado por Elsevier España, S.L.U. All rights reserved.

The Limitations of In Vitro Experimentation in Understanding Biofilms and Chronic Infection

DEFF Research Database (Denmark)

Roberts, Aled E. L.; Kragh, Kasper N.; Bjarnsholt, Thomas

2015-01-01

a systematic review of the most widely used in vitro biofilm systems, and we discuss why they are not always representative of the in vivo biofilms found in chronic infections. We present examples of methods that will help us to bridge the gap between in vitro and in vivo biofilm work so that we can ultimately......We have become increasingly aware that, during infection, pathogenic bacteria often grow in multicellular biofilms that are often highly resistant to antibacterial strategies. In order to understand how biofilms form and contribute to infection, many research groups around the world have heavily...

[The evaluation of relationship between the origin of Candida sp. and the ability of biofilm formation on surface of different biomaterials].

Science.gov (United States)

Ciok-Pater, Emilia; Gospodarek, Eugenia; Prazyńska, Małgorzata; Bogiel, Tomasz

2009-01-01

The increase of fungal infections in recent years is connected with the progress in medicine. The vast usage of biomaterials is an inseparable element of contemporary medicine but it also leads to development of infections. The ability to produce biofilm by those yeasts plays an important role in the pathogenesis of candidiasis. Candida biofilm can form on the surface of plastic materials (silicon, polychloride vinyl, polymethacrylate methyl) used to catheters, drains and dentures production that is why it is a serious problem in case of fungal infections in patients who during the diagnosis and treatment have contact with biomaterials. The aim of the study was the assessment of ability to form biofilm on the surface of different biomaterials (latex silicon, polychloride vinyl, polystyrene, nylon and polymethacrylate methyl). 150 strains of Candida sp. were examined: 85 (56.7%) C. albicans and 65 (43.3%) C. non-albicans. The examined yeasts produced biofilm on the surface of polymethacrylate methyl in 39.3%, latex silicone in 38.7%, polychloride vinyl in 38.0%, polystyrene in 35.3% and nylon in 30.7%. Biofilm was most frequently produced by the strains of C. albicans, C. tropicalis, C. glabrata, C. parapsilosis, C. krusei and C. lusitaniae species.

Cross-feeding and interkingdom communication in dual-species biofilms of Streptococcus mutans and Candida albicans

Science.gov (United States)

Sztajer, Helena; Szafranski, Szymon P; Tomasch, Jürgen; Reck, Michael; Nimtz, Manfred; Rohde, Manfred; Wagner-Döbler, Irene

2014-01-01

Polymicrobial biofilms are of large medical importance, but relatively little is known about the role of interspecies interactions for their physiology and virulence. Here, we studied two human pathogens co-occuring in the oral cavity, the opportunistic fungus Candida albicans and the caries-promoting bacterium Streptococcus mutans. Dual-species biofilms reached higher biomass and cell numbers than mono-species biofilms, and the production of extracellular polymeric substances (EPSs) by S. mutans was strongly suppressed, which was confirmed by scanning electron microscopy, gas chromatography–mass spectrometry and transcriptome analysis. To detect interkingdom communication, C. albicans was co-cultivated with a strain of S. mutans carrying a transcriptional fusion between a green fluorescent protein-encoding gene and the promoter for sigX, the alternative sigma factor of S. mutans, which is induced by quorum sensing signals. Strong induction of sigX was observed in dual-species biofilms, but not in single-species biofilms. Conditioned media from mixed biofilms but not from C. albicans or S. mutans cultivated alone activated sigX in the reporter strain. Deletion of comS encoding the synthesis of the sigX-inducing peptide precursor abolished this activity, whereas deletion of comC encoding the competence-stimulating peptide precursor had no effect. Transcriptome analysis of S. mutans confirmed induction of comS, sigX, bacteriocins and the downstream late competence genes, including fratricins, in dual-species biofilms. We show here for the first time the stimulation of the complete quorum sensing system of S. mutans by a species from another kingdom, namely the fungus C. albicans, resulting in fundamentally changed virulence properties of the caries pathogen. PMID:24824668

In situ rheology of yeast biofilms.

Science.gov (United States)

Brugnoni, Lorena I; Tarifa, María C; Lozano, Jorge E; Genovese, Diego

2014-01-01

The aim of the present work was to investigate the in situ rheological behavior of yeast biofilms growing on stainless steel under static and turbulent flow. The species used (Rhodototula mucilaginosa, Candida krusei, Candida kefyr and Candida tropicalis) were isolated from a clarified apple juice industry. The flow conditions impacted biofilm composition over time, with a predominance of C. krusei under static and turbulent flow. Likewise, structural variations occurred, with a tighter appearance under dynamic flow. Under turbulent flow there was an increase of 112 μm in biofilm thickness at 11 weeks (p < 0.001) and cell morphology was governed by hyphal structures and rounded cells. Using the in situ growth method introduced here, yeast biofilms were determined to be viscoelastic materials with a predominantly solid-like behavior, and neither this nor the G'0 values were significantly affected by the flow conditions or the growth time, and at large deformations their weak structure collapsed beyond a critical strain of about 1.5-5%. The present work could represent a starting point for developing in situ measurements of yeast rheology and contribute to a thin body of knowledge about fungal biofilm formation.

Antifungal activity, mode of action and anti-biofilm effects of Laurus nobilis Linnaeus essential oil against Candida spp.

Science.gov (United States)

Peixoto, Larissa Rangel; Rosalen, Pedro Luiz; Ferreira, Gabriela Lacet Silva; Freires, Irlan Almeida; de Carvalho, Fabíola Galbiatti; Castellano, Lúcio Roberto; de Castro, Ricardo Dias

2017-01-01

The present study demonstrated the antifungal potential of the chemically characterized essential oil (EO) of Laurus nobilis L. (bay laurel) against Candida spp. biofilm adhesion and formation, and further established its mode of action on C. albicans. L. nobilis EO was obtained and tested for its minimum inhibitory and fungicidal concentrations (MIC/MFC) against Candida spp., as well as for interaction with cell wall biosynthesis and membrane ionic permeability. Then we evaluated its effects on the adhesion, formation, and reduction of 48hC. albicans biofilms. The EO phytochemical profile was determined by gas chromatography coupled to mass spectrometry (GC/MS). The MIC and MFC values of the EO ranged from (250 to 500) μg/mL. The MIC values increased in the presence of sorbitol (osmotic protector) and ergosterol, which indicates that the EO may affect cell wall biosynthesis and membrane ionic permeability, respectively. At 2 MIC the EO disrupted initial adhesion of C. albicans biofilms (p0.05). When applied for 1min, every 8h, for 24h and 48h, the EO reduced the amount of C. albicans mature biofilm with no difference in relation to nystatin (p>0.05). The phytochemical analysis identified isoeugenol as the major compound (53.49%) in the sample. L. nobilis EO has antifungal activity probably due to monoterpenes and sesquiterpenes in its composition. This EO may affect cell wall biosynthesis and membrane permeability, and showed deleterious effects against C. albicans biofilms. Copyright © 2016. Published by Elsevier Ltd.

Standardization and classification of In vitro biofilm formation by clinical isolates of Staphylococcus aureus

Directory of Open Access Journals (Sweden)

Ashish Kumar Singh

2017-01-01

Full Text Available Background: Staphylococcus aureus is Gram-positive bacterium commonly associated with nosocomial infections. The development of biofilm exhibiting drug resistance especially in foreign body associated infections has enabled the bacterium to draw considerable attention. However, till date, consensus guidelines for in vitro biofilm quantitation and categorization criterion for the bacterial isolates based on biofilm-forming capacity are lacking. Therefore, it was intended to standardize in vitro biofilm formation by clinical isolates of S. aureus and then to classify them on the basis of their biofilm-forming capacity. Materials and Methods: A study was conducted for biofilm quantitation by tissue culture plate (TCP assay employing 61 strains of S. aureus isolated from clinical samples during May 2015– December 2015 wherein several factors influencing the biofilm formation were optimized. Therefore, it was intended to propose a biofilm classification criteria based on the standard deviation multiples of the control differentiating them into non, low, medium, and high biofilm formers. Results: Brain-heart infusion broth was found to be more effective in biofilm formation compared to trypticase soy broth. Heat fixation was more effective than chemical fixation. Although, individually, glucose, sucrose, and sodium chloride (NaCl had no significant effect on biofilm formation, a statistically significant increase in absorbance was observed after using the supplement mix consisting of 222.2 mM glucose, 116.9 mM sucrose, and 1000 mM NaCl (P = 0.037. Conclusions: The present study puts forth a standardized in vitro TCP assay for biofilm biomass quantitation and categorization criteria for clinical isolates of S. aureus based on their biofilm-forming capacity. The proposed in vitro technique may be further evaluated for its usefulness in the management of persistent infections caused by the bacterium.

Propolis Is an Efficient Fungicide and Inhibitor of Biofilm Production by Vaginal Candida albicans

Directory of Open Access Journals (Sweden)

Isis Regina Grenier Capoci

2015-01-01

Full Text Available Vulvovaginal candidiasis (VVC is one of the most common genital infections in women. The therapeutic arsenal remains restricted, and some alternatives to VVC treatment are being studied. The present study evaluated the influence of a propolis extractive solution (PES on biofilm production by Candida albicans isolated from patients with VVC. Susceptibility testing was used to verify the minimum inhibitory concentration (MIC of PES, with fluconazole and nystatin as controls. The biofilm formation of 29 vaginal isolates of C. albicans and a reference strain that were exposed to PES was evaluated using crystal violet staining. Colony-forming units were evaluated, proteins and carbohydrates of the matrix biofilm were quantified, and scanning electron microscopy was performed. The MIC of PES ranged from 68.35 to 546.87 μg/mL of total phenol content in gallic acid. A concentration of 546.87 μg/mL was able to cause the death of 75.8% of the isolates. PES inhibited biofilm formation by C. albicans from VVC. Besides antifungal activity, PES appears to present important antibiofilm activity on abiotic surfaces, indicating that it may have an additional beneficial effect in the treatment of VVC.

The novel oral glucan synthase inhibitor SCY-078 shows in vitro activity against sessile and planktonic Candida spp.

Science.gov (United States)

Marcos-Zambrano, Laura Judith; Gómez-Perosanz, Marta; Escribano, Pilar; Bouza, Emilio; Guinea, Jesús

2017-07-01

We studied the antifungal activity of SCY-078 (an orally bioavailable 1,3-β -d- glucan synthesis inhibitor), micafungin and fluconazole against the planktonic and sessile forms of 178 Candida and non- Candida isolates causing fungaemia in patients recently admitted to a large European hospital. The in vitro activity of SCY-078, micafungin and fluconazole against the planktonic form of the isolates was assessed using EUCAST EDef 7.3 and CLSI M27-A3. Antibiofilm activity was assessed using the XTT reduction assay. SCY-078 and micafungin showed potent in vitro activity against Candida and non- Candida isolates. The in vitro activity of both drugs was similar, but SYC-078 displayed significantly lower MIC values than micafungin against Candida parapsilosis and non- Candida isolates, whereas micafungin displayed significantly lower MIC values for the remaining species ( P  Candida glabrata , in which the micafungin sessile MIC values were significantly lower ( P  Candida isolates in both sessile and planktonic forms is comparable to that of micafungin. © The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

The Small Molecule DAM Inhibitor, Pyrimidinedione, Disrupts Streptococcus pneumoniae Biofilm Growth In Vitro.

Directory of Open Access Journals (Sweden)

Mukesh Kumar Yadav

Full Text Available Streptococcus pneumoniae persist in the human nasopharynx within organized biofilms. However, expansion to other tissues may cause severe infections such as pneumonia, otitis media, bacteremia, and meningitis, especially in children and the elderly. Bacteria within biofilms possess increased tolerance to antibiotics and are able to resist host defense systems. Bacteria within biofilms exhibit different physiology, metabolism, and gene expression profiles than planktonic cells. These differences underscore the need to identify alternative therapeutic targets and novel antimicrobial compounds that are effective against pneumococcal biofilms. In bacteria, DNA adenine methyltransferase (Dam alters pathogenic gene expression and catalyzes the methylation of adenine in the DNA duplex and of macromolecules during the activated methyl cycle (AMC. In pneumococci, AMC is involved in the biosynthesis of quorum sensing molecules that regulate competence and biofilm formation. In this study, we examine the effect of a small molecule Dam inhibitor, pyrimidinedione, on Streptococcus pneumoniae biofilm formation and evaluate the changes in global gene expression within biofilms via microarray analysis. The effects of pyrimidinedione on in vitro biofilms were studied using a static microtiter plate assay, and the architecture of the biofilms was viewed using confocal and scanning electron microscopy. The cytotoxicity of pyrimidinedione was tested on a human middle ear epithelium cell line by CCK-8. In situ oligonucleotide microarray was used to compare the global gene expression of Streptococcus pneumoniae D39 within biofilms grown in the presence and absence of pyrimidinedione. Real-time RT-PCR was used to study gene expression. Pyrimidinedione inhibits pneumococcal biofilm growth in vitro in a concentration-dependent manner, but it does not inhibit planktonic cell growth. Confocal microscopy analysis revealed the absence of organized biofilms, where cell

Physical disruption of oral biofilms by sodium bicarbonate: an in vitro study.

Science.gov (United States)

Pratten, J; Wiecek, J; Mordan, N; Lomax, A; Patel, N; Spratt, D; Middleton, A M

2016-08-01

Sodium bicarbonate has been shown clinically to be efficacious at removing dental plaque; however, its effect of mechanism against biofilms has not been evaluated in vitro. Here, we used a well-established in vitro plaque biofilm model to investigate the disruption of dental plaque biofilms. Biofilms were grown in a constant depth film fermentor for up to 14 days. The fermentor was inoculated with pooled human saliva and growth maintained with artificial saliva. After various time points, replicate biofilms were removed and subjected to treatment at varying concentrations of sodium bicarbonate. Disruption of the plaque was assessed by viable counts and microscopy. The viable count results showed that younger biofilms were less susceptible to the action of sodium bicarbonate; however, biofilms of 7 days and older were increasingly susceptible to the material with the oldest biofilms being the most susceptible. Sixty-seven percentage of sodium bicarbonate slurry was able to reduce the number of organisms present by approx. 3 log10 . These quantitative data were corroborated qualitatively with both confocal and electron microscopy, which both showed substantial qualitative removal of mature biofilms. The results from this study have shown that sodium bicarbonate is able to disrupt mature dental plaque grown in vitro and that its reported efficacy in maintaining oral hygiene may be related to this key factor. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Candida virulence and ethanol-derived acetaldehyde production in oral cancer and non-cancer subjects.

Science.gov (United States)

Alnuaimi, A D; Ramdzan, A N; Wiesenfeld, D; O'Brien-Simpson, N M; Kolev, S D; Reynolds, E C; McCullough, M J

2016-11-01

To compare biofilm-forming ability, hydrolytic enzymes and ethanol-derived acetaldehyde production of oral Candida isolated from the patients with oral cancer and matched non-oral cancer. Fungal biofilms were grown in RPMI-1640 medium, and biofilm mass and biofilm activity were assessed using crystal violet staining and XTT salt reduction assays, respectively. Phospholipase, proteinase, and esterase production were measured using agar plate method, while fungal acetaldehyde production was assessed via gas chromatography. Candida isolated from patients with oral cancer demonstrated significantly higher biofilm mass (P = 0.031), biofilm metabolic activity (P oral cancer. High ethanol-derived acetaldehyde-producing Candida were more prevalent in patients with oral cancer than non-oral cancer (P = 0.01). In univariate regression analysis, high biofilm mass (P = 0.03) and biofilm metabolic activity (P production ability (0.01) were significant risk factors for oral cancer; while in the multivariate regression analysis, high biofilm activity (0.01) and phospholipase (P = 0.01) were significantly positive influencing factors on oral cancer. These data suggest a significant positive association between the ability of Candida isolates to form biofilms, to produce hydrolytic enzymes, and to metabolize alcohol to acetaldehyde with their ability to promote oral cancer development. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Combination of micropreparative solution isoelectric focusing and high-performance liquid chromatography for differentiation of biofilm-positive and biofilm-negative Candida parapsilosis group from vascular catheter

Czech Academy of Sciences Publication Activity Database

Vykydalová, Marie; Horká, Marie; Růžička, F.; Duša, Filip; Moravcová, Dana; Kahle, Vladislav; Šlais, Karel

2014-01-01

Roč. 812, Feb (2014), s. 243-249 ISSN 0003-2670 R&D Projects: GA MV VG20112015021 Institutional support: RVO:68081715 Keywords : Candida parapsilosis group * biofilm * sonicate from vascular cathether * sIEF * HPLC Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.513, year: 2014 http://hdl.handle.net/11104/0230766

In vitro Effects of Lemongrass Extract on Candida albicans Biofilms, Human Cells Viability, and Denture Surface.

Science.gov (United States)

Madeira, Petrus L B; Carvalho, Letícia T; Paschoal, Marco A B; de Sousa, Eduardo M; Moffa, Eduardo B; da Silva, Marcos A Dos Santos; Tavarez, Rudys de Jesus Rodolfo; Gonçalves, Letícia M

2016-01-01

The purpose of this study was to investigate whether immersion of a denture surface in lemongrass extract (LGE) has effects on C. albicans biofilms, human cell viability and denture surface. Minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC) were performed for LGE against C. albicans. For biofilm analysis, discs were fabricated using a denture acrylic resin with surface roughness standardization. C. albicans biofilms were developed on saliva-coated discs, and the effects of LGE at MIC, 5XMIC, and 10XMIC were investigated during biofilm formation and after biofilm maturation. Biofilms were investigated for cell counting, metabolic activity, and microscopic analysis. The cytotoxicity of different concentrations of LGE to peripheral blood mononuclear cells (PBMC) was analyzed using MTT. The effects of LGE on acrylic resin were verified by measuring changes in roughness, color and flexural strength after 28 days of immersion. Data were analyzed by ANOVA, followed by a Tukey test at a 5% significance level. The minimal concentration of LGE required to inhibit C. albicans growth was 0.625 mg/mL, while MFC was 2.5 mg/mL. The presence of LGE during biofilm development resulted in a reduction of cell counting (p 0.05). There were no verified differences in color perception, roughness, or flexural strength after immersion in LGE at MIC compared to the control (p > 0.05). It could be concluded that immersion of the denture surface in LGE was effective in reducing C. albicans biofilms with no deleterious effects on acrylic properties at MIC. MIC was also an effective and safe concentration for use.

The in vitro effect of xylitol on chronic rhinosinusitis biofilms.

Science.gov (United States)

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

2016-12-01

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

[Infectious risk related to the formation of multi-species biofilms (Candida - bacteria) on peripheral vascular catheters].

Science.gov (United States)

Seghir, A; Boucherit-Otmani, Z; Sari-Belkharroubi, L; Boucherit, K

2017-03-01

The Candida yeasts are the fourth leading cause of death from systemic infections, the risk may increase when the infection also involves bacteria. Yeasts and bacteria can adhere to medical implants, such as peripheral vascular catheters, and form a multicellular structures called "mixed biofilms" more resistant to antimicrobials agents. However, the formation of mixed biofilms on implants leads to long-term persistent infections because they can act as reservoirs of pathogens that have poorly understood interactions. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

In Vitro Activities of Terbinafine against Cutaneous Isolates of Candida albicans and Other Pathogenic Yeasts

Science.gov (United States)

Ryder, Neil S.; Wagner, Sonja; Leitner, Ingrid

1998-01-01

Terbinafine is active in vitro against a wide range of pathogenic fungi, including dermatophytes, molds, dimorphic fungi, and some yeasts, but earlier studies indicated that the drug had little activity against Candida albicans. In contrast, clinical studies have shown topical and oral terbinafine to be active in cutaneous candidiasis and Candida nail infections. In order to define the anti-Candida activity of terbinafine, we tested the drug against 350 fresh clinical isolates and additional strains by using a broth dilution assay standardized according to the guidelines of the National Committee for Clinical Laboratory Standards (NCCLS) M27-A assay. Terbinafine was found to have an MIC of 1 μg/ml for reference C. albicans strains. For 259 clinical isolates, the MIC at which 50% of the isolates are inhibited (MIC50) of terbinafine was 1 μg/ml (fluconazole, 0.5 μg/ml), and the MIC90 was 4 μg/ml (fluconazole, 1 μg/ml). Terbinafine was highly active against Candida parapsilosis (MIC90, 0.125 μg/ml) and showed potentially interesting activity against isolates of Candida dubliniensis, Candida guilliermondii, Candida humicola, and Candida lusitaniae. It was not active against the Candida glabrata, Candida krusei, and Candida tropicalis isolates in this assay. Cryptococcus laurentii and Cryptococcus neoformans were highly susceptible to terbinafine, with MICs of 0.06 to 0.25 μg/ml. The NCCLS macrodilution assay provides reproducible in vitro data for terbinafine against Candida and other yeasts. The MICs for C. albicans and C. parapsilosis are compatible with the known clinical efficacy of terbinafine in cutaneous infections, while the clinical relevance of its activities against the other species has yet to be determined. PMID:9593126

Multitask Imidazolium Salt Additives for Innovative Poly(l-lactide) Biomaterials: Morphology Control, Candida spp. Biofilm Inhibition, Human Mesenchymal Stem Cell Biocompatibility, and Skin Tolerance.

Science.gov (United States)

Schrekker, Clarissa M L; Sokolovicz, Yuri C A; Raucci, Maria G; Selukar, Balaji S; Klitzke, Joice S; Lopes, William; Leal, Claudio A M; de Souza, Igor O P; Galland, Griselda B; Dos Santos, João Henrique Z; Mauler, Raquel S; Kol, Moshe; Dagorne, Samuel; Ambrosio, Luigi; Teixeira, Mário L; Morais, Jonder; Landers, Richard; Fuentefria, Alexandre M; Schrekker, Henri S

2016-08-24

Candida species have great ability to colonize and form biofilms on medical devices, causing infections in human hosts. In this study, poly(l-lactide) films with different imidazolium salt (1-n-hexadecyl-3-methylimidazolium chloride (C16MImCl) and 1-n-hexadecyl-3-methylimidazolium methanesulfonate (C16MImMeS)) contents were prepared, using the solvent casting process. Poly(l-lactide)-imidazolium salt films were obtained with different surface morphologies (spherical and directional), and the presence of the imidazolium salt in the surface was confirmed. These films with different concentrations of the imidazolium salts C16MImCl and C16MImMeS presented antibiofilm activity against isolates of Candida tropicalis, Candida parapsilosis, and Candida albicans. The minor antibiofilm concentration assay enabled one to determine that an increasing imidazolium salt content promoted, in general, an increase in the inhibition percentage of biofilm formation. Scanning electron microscopy micrographs confirmed the effective prevention of biofilm formation on the imidazolium salt containing biomaterials. Lower concentrations of the imidazolium salts showed no cytotoxicity, and the poly(l-lactide)-imidazolium salt films presented good cell adhesion and proliferation percentages with human mesenchymal stem cells. Furthermore, no acute microscopic lesions were identified in the histopathological evaluation after contact between the films and pig ear skin. In combination with the good morphological, physicochemical, and mechanical properties, these poly(l-lactide)-based materials with imidazolium salt additives can be considered as promising biomaterials for use in the manufacturing of medical devices.

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...

The influence of photodynamic therapy parameters on the inactivation of Candida spp: in vitro and in vivo studies

International Nuclear Information System (INIS)

Alves, F; Mima, E G; Jorge, J H; Pavarina, A C; Dovigo, L N; Bagnato, V S; De Souza Costa, C A

2014-01-01

The influence of parameters of photodynamic therapy (PDT), such as pre-irradiation time (PIT), on the inactivation of Candida spp. was assessed in vitro and in vivo. Suspensions of Candida albicans, Candida tropicalis, Candida krusei and Candida glabrata were treated with Photogem ® , incubated for 5, 10 or 15 min and illuminated with a blue LED light. Colonies were cultivated and log values of CFU ml −1 were analyzed by ANOVA and Kruskall–Wallis test. For in vivo evaluation, immunosuppressed mice were inoculated with C. albicans. PDT was performed on the dorsum of the tongue by topical administration of Photogem ®  and illumination after 5, 10 or 15 min. C. albicans was recovered from the tongue and the number of CFU ml −1 was analyzed by ANOVA and Tukey test. Animals were killed and the tongues were surgically removed for histological analysis. Susceptibility of Candida spp. suspensions to PDT was in decreasing order: C. albicans  = C. tropicalis  < C. krusei  < C. glabrata. No significant difference was observed among the different PIT (p > 0.05), both in vivo and in vitro. A significant reduction (p < 0.05) of log(CFU ml −1 ) of C. albicans from tongues of mice was observed with no adverse effects in the tissue. PDT was effective to inactivate in vitroCandida spp. and for reduction of C. albicans in vivo, independently of the PIT used. (paper)

In Vitro Effect of Local Anesthetics on Candida albicans Germ Tube Formation

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Acácio Rodrigues

1994-01-01

Full Text Available Objective: This study was planned to clarify the in vitro effect of lidocaine and bupivacaine on germ tube formation by Candida albicans isolates from cases of clinical vaginal candidiasis.

The MP65 gene is required for cell wall integrity, adherence to epithelial cells and biofilm formation in Candida albicans

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Girolamo Antonietta

2011-05-01

Full Text Available Abstract Background The MP65 gene of Candida albicans (orf19.1779 encodes a putative β-glucanase mannoprotein of 65 kDa, which plays a main role in a host-fungus relationship, morphogenesis and pathogenicity. In this study, we performed an extensive analysis of a mp65Δ mutant to assess the role of this protein in cell wall integrity, adherence to epithelial cells and biofilm formation. Results The mp65Δ mutant showed a high sensitivity to a range of cell wall-perturbing and degrading agents, especially Congo red, which induced morphological changes such as swelling, clumping and formation of hyphae. The mp65Δ mutant showed an activation of two MAPKs (Mkc1p and Cek1p, a high level of expression of two stress-related genes (DDR48 and SOD5, and a modulated expression of β-glucan epitopes, but no gross changes in cell wall polysaccharide composition. Interestingly, the mp65Δ mutant displayed a marked reduction in adhesion to BEC and Caco-2 cells and severe defects in biofilm formation when compared to the wild type. All of the mentioned properties were totally or partially recovered in a revertant strain, demonstrating the specificity of gene deletion. Conclusions We demonstrate that the MP65 gene of Candida albicans plays a significant role in maintaining cell wall integrity, as well as in adherence to epithelia and biofilm formation, which are major virulence attributes of this fungus.

The Effect of Essential Oils and Bioactive Fractions on Streptococcus mutans and Candida albicans Biofilms: A Confocal Analysis

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Irlan Almeida Freires

2015-01-01

Full Text Available The essential oils (EO and bioactive fractions (BF from Aloysia gratissima, Baccharis dracunculifolia, Coriandrum sativum, Cyperus articulatus, and Lippia sidoides were proven to have strong antimicrobial activity on planktonic microorganisms; however, little is known about their effects on the morphology or viability of oral biofilms. Previously, we determined the EO/fractions with the best antimicrobial activity against Streptococcus mutans and Candida spp. In this report, we used a confocal analysis to investigate the effect of these EO and BF on the morphology of S. mutans biofilms (thickness, biovolume, and architecture and on the metabolic viability of C. albicans biofilms. The analysis of intact treated S. mutans biofilms showed no statistical difference for thickness in all groups compared to the control. However, a significant reduction in the biovolume of extracellular polysaccharides and bacteria was observed for A. gratissima and L. sidoides groups, indicating that these BF disrupt biofilm integrity and may have created porosity in the biofilm. This phenomenon could potentially result in a weakened structure and affect biofilm dynamics. Finally, C. sativum EO drastically affected C. albicans viability when compared to the control. These results highlight the promising antimicrobial activity of these plant species and support future translational research on the treatment of dental caries and oral candidiasis.

In vitro activity of 23 tea extractions and epigallocatechin gallate against Candida species

DEFF Research Database (Denmark)

Chen, Ming; Zhai, Lin; Arendrup, Maiken Cavling

2015-01-01

In this study, we investigate the susceptibility of Candida albicans, Candida glabrata, Candida krusei, Candida parapsilosis, Candida tropicalis, and Aspergillus fumigatus using the EUCAST microdilution minimum inhibitory concentration (MIC) method (final tea supernatant concentration range 5.......0-0.005 mg/ml) to 23 different teas and tea catechins including epigallocatechin gallate (EGCG) isolated from green tea. All teas exhibited potent in vitro antifungal activity against C. glabrata. Six out of nine green teas and three of eight black teas had an MIC of 0.078 mg/ml, one white tea had an MIC...... of 0.156 mg/ml, and finally three of five oolong teas had an MIC of 0.156 mg/ml. Three teas exhibited activity against C. albicans (MIC 1.25 mg/ml), one green tea was active against C. parapsilosis (MIC 1.25 mg/ml), but none were effective against C. krusei, C. tropicalis or A. fumigatus...

The Anti-Adhesive Effect of Curcumin on Candida albicans Biofilms on Denture Materials

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Gordon Ramage

2017-04-01

Full Text Available The use of natural compounds as an alternative source of antimicrobials has become a necessity given the growing concern over global antimicrobial resistance. Polyphenols, found in various edible plants, offers one potential solution to this. We aimed to investigate the possibility of using curcumin within the context of oral health as a way of inhibiting and preventing the harmful development of Candida albicans biofilms. We undertook a series of adsorption experiments with varying concentrations of curcumin, showing that 50 μg/ml could prevent adhesion. This effect could be further synergized by the curcumin pre-treatment of yeast cells to obtain significantly greater inhibition (>90%, p < 0.001. Investigation of the biological impact of curcumin showed that it preferentially affected immature morphological forms (yeast and germlings, and actively promoted aggregation of the cells. Transcriptional analyses showed that key adhesins were down-regulated (ALS1 and ALS3, whereas aggregation related genes (ALS5 and AAF1 were up-regulated. Collectively, these data demonstrated that curcumin elicits anti-adhesive effects and that induces transcription of genes integrally involved in the processes related to biofilm formation. Curcumin and associated polyphenols therefore have the capacity to be developed for use in oral healthcare to augment existing preventative strategies for candidal biofilms on the denture surface.

Characterization of biofilm formation by Borrelia burgdorferi in vitro.

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Eva Sapi

Full Text Available Borrelia burgdorferi, the causative agent of Lyme disease, has long been known to be capable of forming aggregates and colonies. It was recently demonstrated that Borrelia burgdorferi aggregate formation dramatically changes the in vitro response to hostile environments by this pathogen. In this study, we investigated the hypothesis that these aggregates are indeed biofilms, structures whose resistance to unfavorable conditions are well documented. We studied Borrelia burgdorferi for several known hallmark features of biofilm, including structural rearrangements in the aggregates, variations in development on various substrate matrices and secretion of a protective extracellular polymeric substance (EPS matrix using several modes of microscopic, cell and molecular biology techniques. The atomic force microscopic results provided evidence that multilevel rearrangements take place at different stages of aggregate development, producing a complex, continuously rearranging structure. Our results also demonstrated that Borrelia burgdorferi is capable of developing aggregates on different abiotic and biotic substrates, and is also capable of forming floating aggregates. Analyzing the extracellular substance of the aggregates for potential exopolysaccharides revealed the existence of both sulfated and non-sulfated/carboxylated substrates, predominately composed of an alginate with calcium and extracellular DNA present. In summary, we have found substantial evidence that Borrelia burgdorferi is capable of forming biofilm in vitro. Biofilm formation by Borrelia species might play an important role in their survival in diverse environmental conditions by providing refuge to individual cells.

Acute and substantive action of antimicrobial toothpastes and mouthrinses on oral biofilm in vitro

NARCIS (Netherlands)

Otten, Marieke P. T.; Busscher, Henk J.; van der Mei, Henny C.; van Hoogmoed, Chris G.; Abbas, Frank; Hoogmoed, G.G. van

The aim of this study was to compare acute action by killing or disrupting oral biofilms through the use of antimicrobial toothpastes and mouthrinses in vitro and to investigate substantive action by absorption of antimicrobials in a biofilm. Biofilms from freshly collected human saliva were grown

Candida and Porphyromonas gingivalis: the effect on wound closure in vitro

NARCIS (Netherlands)

Haverman, Thijs M.; Laheij, Alexa M. G. A.; de Soet, Johannes J.; de Lange, Jan; Rozema, Frederik R.

2017-01-01

Microorganisms play a role in oral mucositis after cancer therapy. The current study explored the hypothesis that Candida spp. alone and together with Porphyromonas gingivalis cause delayed healing of oral ulcerations due to the inhibition of wound closure. An in vitro scratch assay model was used

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

DEFF Research Database (Denmark)

Roizman, Dan; Vidaillac, Celine; Givskov, Michael

2017-01-01

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

Candida krusei and Candida glabrata reduce the filamentation of Candida albicans by downregulating expression of HWP1 gene.

Science.gov (United States)

de Barros, Patrícia Pimentel; Freire, Fernanda; Rossoni, Rodnei Dennis; Junqueira, Juliana Campos; Jorge, Antonio Olavo Cardoso

2017-07-01

Pathogenicity of Candida albicans is associated with its capacity switch from yeast-like to hyphal growth. The hyphal form is capable to penetrate the epithelial surfaces and to damage the host tissues. Therefore, many investigations have focused on mechanisms that control the morphological transitions of C. albicans. Recently, certain studies have showed that non-albicans Candida species can reduce the capacity of C. albicans to form biofilms and to develop candidiasis in animal models. Then, the objective of this study was to evaluate the effects of Candida krusei and Candida glabrata on the morphogenesis of C. albicans. Firstly, the capacity of reference and clinical strains of C. albicans in forming hyphae was tested in vitro. After that, the expression of HWP1 (hyphal wall protein 1) gene was determined by quantitative real-time PCR (polymerase chain reaction) assay. For both reference and clinical strains, a significant inhibition of the hyphae formation was observed when C. albicans was incubated in the presence of C. krusei or C. glabrata compared to the control group composed only by C. albicans. In addition, the culture mixed of C. albicans-C. krusei or C. albicans-C. glabrata reduced significantly the expression of HWP1 gene of C. albicans in relation to single cultures of this specie. In both filamentation and gene expression assays, C. krusei showed the higher inhibitory activity on the morphogenesis of C. albicans compared to C. glabrata. C. krusei and C. glabrata are capable to reduce the filamentation of C. albicans and consequently decrease the expression of the HWP1 gene.

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

Science.gov (United States)

Liu, Xiang; Chen, Haihong; Wang, Shengqing

2012-07-01

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

[Antimycotic activity in vitro and in vivo of 5-fluorocytosine on pathogenic strains of Candida albicans and Cryptococcus neoformans].

Science.gov (United States)

Costa, A L; Valenti, A; Costa, G; Calogero, F

1976-01-01

The authors have analyzed the 5 Fluoro Cytosine (5FC) activity on strains of Candida albicans and Criptococcus neoformans, both in vitro and in vivo. In vitro the minimal inhibitory concentration (MIC) was determined; in vivo tests of pathogenicity on rabbit and mouse have been executed. The various findings obtained have shown a strong activity of the 5FC on strains of Candida and Criptococcus.

Effects of fluconazole treatment of mice infected with fluconazole-susceptible and -resistant Candida tropicalis on fungal cell surface hydrophobicity, adhesion and biofilm formation

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R L Kanoshiki

2015-01-01

Full Text Available Background : The incidence of Candida tropicalis less susceptible to fluconazole (FLC has been reported in many parts of the world. Objectives : The aim of this study was to examine the changes of putative virulence attributes of Candida tropicalis accompanying the development of resistance to FLC in vitro and in vivo. Materials and Methods : A FLC-resistant strain (FLC-R was obtained after sequential exposure of a clinical isolate FLC-sensitive (FLC-S to increasing concentrations of the antifungal. The course of infection by both strains was analyzed in BALB/c mice. Analyses of gene expression were performed by real-time polymerase chain reaction PCR. The cell surface hydrophobicity, adhesion and biofilm formation were also determined. Results : Development of resistance to FLC could be observed after 15 days of subculture in azole-containing medium. Overexpression of MDR1 and ERG11 genes were observed in FLC-R, and this strain exhibited enhanced virulence in mice, as assessed by the mortality rate. All mice challenged with the FLC-R died and FLC-treatment caused earlier death in mice infected with this strain. All animals challenged with FLC-S survived the experiment, regardless of FLC-treatment. Overall, FLC-R derivatives strains were significantly more hydrophobic than FLC-S strains and showed greater adherence and higher capacity to form biofilm on polystyrene surface. Conclusions : The expression of virulence factors was higher in FLC-R-C. tropicalis and it was enhanced after FLC-exposure. These data alert us to the importance of identifying microorganisms that show resistance to the antifungals to establish an appropriate management of candidiasis therapy.

From in vitro to in vivo Models of Bacterial Biofilm-Related Infections

Science.gov (United States)

Lebeaux, David; Chauhan, Ashwini; Rendueles, Olaya; Beloin, Christophe

2013-01-01

The influence of microorganisms growing as sessile communities in a large number of human infections has been extensively studied and recognized for 30–40 years, therefore warranting intense scientific and medical research. Nonetheless, mimicking the biofilm-life style of bacteria and biofilm-related infections has been an arduous task. Models used to study biofilms range from simple in vitro to complex in vivo models of tissues or device-related infections. These different models have progressively contributed to the current knowledge of biofilm physiology within the host context. While far from a complete understanding of the multiple elements controlling the dynamic interactions between the host and biofilms, we are nowadays witnessing the emergence of promising preventive or curative strategies to fight biofilm-related infections. This review undertakes a comprehensive analysis of the literature from a historic perspective commenting on the contribution of the different models and discussing future venues and new approaches that can be merged with more traditional techniques in order to model biofilm-infections and efficiently fight them. PMID:25437038

[THE CHARACTERISTICS OF MORPHOLOGY OF BIOFILM OF PERIODONTIUM UNDER INFLAMMATORY DISEASES OF GUMS (CHRONIC CATARRHAL GINGIVITIS, CHRONIC PERIODONTITIS, CANDIDA-ASSOCIATED PERIODONTITIS) ACCORDING RESULTS OF ELECTRONIC MICROSCOPY].

Science.gov (United States)

Ippolitov, E V; Didenko, L V; Tzarev, V N

2015-12-01

The study was carried out to analyze morphology of biofilm of periodontium and to develop electronic microscopic criteria of differentiated diagnostic of inflammatory diseases of gums. The scanning electronic microscopy was applied to analyze samples of bioflm of periodont from 70 patients. Including ten patients with every nosologic form of groups with chronic catarrhal periodontitis. of light, mean and severe degree, chronic catarrhal gingivitis, Candida-associated paroperiodontitis and 20 healthy persons with intact periodontium. The analysis was implemented using dual-beam scanning electronic microscope Quanta 200 3D (FEI company, USA) and walk-through electronic micJEM 100B (JEOL, Japan). To detect marker DNA of periodont pathogenic bacteria in analyzed samples the kit of reagentsfor polymerase chain reaction "MultiDent-5" ("GenLab", Russia). The scanning electronic microscopy in combination with transmission electronic microscopy and polymerase chain reaction permits analyzing structure, composition and degree of development of biofilm of periodontium and to apply differentiated diagnostic of different nosologic forms of inflammatory diseases of periodontium, including light form of chronic periodontitis and gingivitis. The electronic microscopical indications of diseases ofperiodontium of inflammatory character are established: catarrhal gingivitis, (coccal morphological alternate), chronic periodontitis (bacillary morphological alternate), Candida-associated periodontitis (Candida morphological alternate of biofilm ofperiodontium).

Lactobacillus fermentum ATCC 23271 Displays In vitro Inhibitory Activities against Candida spp.

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do Carmo, Monique S.; Noronha, Francisca M. F.; Arruda, Mariana O.; Costa, Ênnio P. da Silva; Bomfim, Maria R. Q.; Monteiro, Andrea S.; Ferro, Thiago A. F.; Fernandes, Elizabeth S.; Girón, Jorge A.; Monteiro-Neto, Valério

2016-01-01

Lactobacilli are involved in the microbial homeostasis in the female genital tract. Due to the high prevalence of many bacterial diseases of the female genital tract and the resistance of microorganisms to various antimicrobial agents, alternative means to control these infections are necessary. Thus, this study aimed to evaluate the probiotic properties of well-characterized Lactobacillus species, including L. acidophilus (ATCC 4356), L. brevis (ATCC 367), L. delbrueckii ssp. delbrueckii (ATCC 9645), L. fermentum (ATCC 23271), L. paracasei (ATCC 335), L. plantarum (ATCC 8014), and L. rhamnosus (ATCC 9595), against Candida albicans (ATCC 18804), Neisseria gonorrhoeae (ATCC 9826), and Streptococcus agalactiae (ATCC 13813). The probiotic potential was investigated by using the following criteria: (i) adhesion to host epithelial cells and mucus, (ii) biofilm formation, (iii) co-aggregation with bacterial pathogens, (iv) inhibition of pathogen adhesion to mucus and HeLa cells, and (v) antimicrobial activity. Tested lactobacilli adhered to mucin, co-aggregated with all genital microorganisms, and displayed antimicrobial activity. With the exception of L. acidophilus and L. paracasei, they adhered to HeLa cells. However, only L. fermentum produced a moderate biofilm and a higher level of co-aggregation and mucin binding. The displacement assay demonstrated that all Lactobacillus strains inhibit C. albicans binding to mucin (p < 0.001), likely due to the production of substances with antimicrobial activity. Clinical isolates belonging to the most common Candida species associated to vaginal candidiasis were inhibited by L. fermentum. Collectively, our data suggest that L. fermentum ATCC 23271 is a potential probiotic candidate, particularly to complement candidiasis treatment, since presented with the best probiotic profile in comparison with the other tested lactobacilli strains. PMID:27833605

Lactobacillus fermentum ATCC 23271 displays in vitro inhibitory activities against Candida spp.

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Monique Santos Carmo

2016-10-01

Full Text Available Lactobacilli are involved in the microbial homeostasis in the female genital tract. Due to the high prevalence of many bacterial diseases of the female genital tract and the resistance of microorganisms to various antimicrobial agents, alternative means to control these infections are necessary. Thus, this study aimed to evaluate the probiotic properties of well-characterized Lactobacillus species, including L. acidophilus (ATCC 4356, L. brevis (ATCC 367, L. delbrueckii ssp. delbrueckii (ATCC 9645, L. fermentum (ATCC 23271, L. paracasei (ATCC 335, L. plantarum (ATCC 8014, and L. rhamnosus (ATCC 9595, against Candida albicans (ATCC 18804, Neisseria gonorrhoeae (ATCC 9826, and Streptococcus agalactiae (ATCC 13813. The probiotic potential was investigated by using the following criteria: i adhesion to host epithelial cells and mucus, ii biofilm formation, iii co-aggregation with bacterial pathogens, iv inhibition of pathogen adhesion to mucus and HeLa cells, and v antimicrobial activity. Tested lactobacilli adhered to mucin, co-aggregated with all genital microorganisms, and displayed antimicrobial activity. With the exception of L. acidophilus and L. paracasei, they adhered to HeLa cells. However, only L. fermentum produced a moderate biofilm and a higher level of co-aggregation and mucin binding. The displacement assay demonstrated that all Lactobacillus strains inhibit C. albicans binding to mucin (p < 0.001, likely due to the production of substances with antimicrobial activity. Clinical isolates belonging to the most common Candida species associated to vaginal candidosis were inhibited by L. fermentum. Collectively, our data suggest that L. fermentum ATCC 23271 is a potential probiotic candidate, particularly to complement candidosis treatment, since presented with the best probiotic profile in comparison with the other tested lactobacilli strains.

Induction of suppressor cells in vitro by Candida albicans.

Science.gov (United States)

Cuff, C F; Rogers, C M; Lamb, B J; Rogers, T J

1986-06-01

Normal splenocytes cultured with Formalin-killed Candida albicans were shown to acquire significant suppressor cell activity in a period of 3 days. These cells were found to suppress both the phytohemagglutinin-induced mitogen response as well as the anti-sheep erythrocyte antibody response. Experiments were carried out to determine the nature of the suppressor cell population. Results showed that these cells were not susceptible to treatment with anti-Thy 1 antibody and complement. Panning experiments showed that the suppressor cells were not plastic-adherent or Mac-1 antigen-positive. The suppressor cells were, however, adherent to anti-mouse immunoglobulin (F(ab')2-fragment)-coated dishes. Additional experiments showed that the suppressor cell activity was susceptible to treatment with monoclonal anti-Lyb 2.1 antibody and complement. These results suggest that the suppressor cell induced in vitro by Candida is a member of the B-lymphocyte lineage.

Streptococcus uberis: In vitro biofilm production in response to carbohydrates and skim milk.

Science.gov (United States)

Dieser, Silvana A; Fessia, Aluminé S; Ferrari, Miriam P; Raspanti, Claudia G; Odierno, Liliana M

Streptococcus uberis has become one of the most important environmental pathogens associated with clinical and subclinical bovine mastitis. Biofilm confers to bacteria more resistance to physical and chemical agents as well as to different mechanisms of the innate immune system. The aim of this work was to evaluate the ability of in vitro biofilm production in 32 S. uberis isolates from bovine mastitis and identified by biochemical tests and subsequently confirmed by the amplification of the pauA gene. The isolates were cultivated in TMP broth and TMP broth with the addition of 0.5% glucose, 1% sucrose, 1% lactose or 0.5% skim milk in microtiter plates stained with crystal violet. We demonstrated that S. uberis isolated from bovine mastitis are able to produce biofilms in TMP broth and, also that biofilm formation by S. uberis can be significantly enhanced by the addition of 0.5% glucose or 1% sucrose to TMP broth. This may suggest that the carbohydrates in milk or within the ruminant gut might affect the growth mode of S. uberis. In addition, our results showed that in vitro biofilm production under different conditions of supplementation displays variation among the isolates and that each isolate shows a particular profile of biofilm production. This phenotypic heterogeneity in biofilm production exhibited by S. uberis could at least partly explain why this bacterium has the ability to adapt to different niches facilitating survival to diverse and stressful conditions. Copyright © 2017 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

Soluble factors from biofilm of Candida albicans and Staphylococcus aureus promote cell death and inflammatory response.

Science.gov (United States)

de Carvalho Dias, Kassia; Barbugli, Paula Aboud; de Patto, Fernanda; Lordello, Virginia Barreto; de Aquino Penteado, Letícia; Medeiros, Alexandra Ivo; Vergani, Carlos Eduardo

2017-06-30

The objective of this study was to better understand the effects of soluble factors from biofilm of single- and mixed-species Candida albicans (C. albicans) and methicillin-sensitive Staphylococcus aureus (MSSA) cultures after 36 h in culture on keratinocytes (NOK-si and HaCaT) and macrophages (J774A.1). Soluble factors from biofilms of C. albicans and MSSA were collected and incubated with keratinocytes and macrophages, which were subsequently evaluated by cell viability assays (MTT). Lactate dehydrogenase (LDH) enzyme release was measured to assess cell membrane damage to keratinocytes. Cells were analysed by brightfield microscopy after 2 and 24 h of exposure to the soluble factors from biofilm. Cell death was detected by labelling apoptotic cells with annexin V and necrotic cells with propidium iodide (PI) and was visualized via fluorescence microscopy. Soluble factors from biofilm were incubated with J774A.1 cells for 24 h; the subsequent production of NO and the cytokines IL-6 and TNF-α was measured by ELISA. The cell viability assays showed that the soluble factors of single-species C. albicans cultures were as toxic as the soluble factors from biofilm of mixed cultures, whereas the soluble factors of MSSA cultures were less toxic than those of C. albicans or mixed cultures. The soluble factors from biofilm of mixed cultures were the most toxic to the NOK-si and HaCaT cells, as confirmed by analyses of PI labelling and cell morphology. Soluble factors from biofilm of single-species MSSA and mixed-species cultures induced the production of IL-6, NO and TNF-α by J744A.1 macrophages. The production of IL-6 and NO induced by the soluble factors from biofilm of mixed cultures was lower than that induced by the soluble factors from biofilm of single-species MSSA cultures, whereas the soluble factors from biofilm of C. albicans cultures induced only low levels of NO. Soluble factors from 36-h-old biofilm of C. albicans and MSSA cultures promoted cell death and

In vitro activity of Origanum vulgare essential oil against Candida species

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Marlete Brum Cleff

2010-03-01

Full Text Available The aim of this study was to evaluate the in vitro activity of the essential oil extracted from Origanum vulgare against sixteen Candida species isolates. Standard strains tested comprised C. albicans (ATCC strains 44858, 4053, 18804 and 3691, C. parapsilosis (ATCC 22019, C. krusei (ATCC 34135, C. lusitaniae (ATCC 34449 and C. dubliniensis (ATCC MY646. Six Candida albicans isolates from the vaginal mucous membrane of female dogs, one isolate from the cutaneous tegument of a dog and one isolate of a capuchin monkey were tested in parallel. A broth microdilution technique (CLSI was used, and the inoculum concentration was adjusted to 5 x 10(6 CFU mL-1. The essential oil was obtained by hydrodistillation in a Clevenger apparatus and analyzed by gas chromatography. Susceptibility was expressed as Minimal Inhibitory Concentration (MIC and Minimal Fungicidal Concentration (MFC. All isolates tested in vitro were sensitive to O. vulgare essential oil. The chromatographic analysis revealed that the main compounds present in the essential oil were 4-terpineol (47.95%, carvacrol (9.42%, thymol (8.42% and □-terpineol (7.57%. C. albicans isolates obtained from animal mucous membranes exhibited MIC and MFC values of 2.72 µL mL-1 and 5 µL mL-1, respectively. MIC and MFC values for C. albicans standard strains were 2.97 µL mL-1 and 3.54 µL mL-1, respectively. The MIC and MFC for non-albicans species were 2.10 µL mL-1 and 2.97 µL mL-1, respectively. The antifungal activity of O. vulgare essential oil against Candida spp. observed in vitro suggests its administration may represent an alternative treatment for candidiasis.

Biofilm formation on stainless steel and gold wires for bonded retainers in vitro and in vivo and their susceptibility to oral antimicrobials.

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Jongsma, Marije A; Pelser, Floris D H; van der Mei, Henny C; Atema-Smit, Jelly; van de Belt-Gritter, Betsy; Busscher, Henk J; Ren, Yijin

2013-05-01

Bonded retainers are used in orthodontics to maintain treatment result. Retention wires are prone to biofilm formation and cause gingival recession, bleeding on probing and increased pocket depths near bonded retainers. In this study, we compare in vitro and in vivo biofilm formation on different wires used for bonded retainers and the susceptibility of in vitro biofilms to oral antimicrobials. Orthodontic wires were exposed to saliva, and in vitro biofilm formation was evaluated using plate counting and live/dead staining, together with effects of exposure to toothpaste slurry alone or followed by antimicrobial mouthrinse application. Wires were also placed intra-orally for 72 h in human volunteers and undisturbed biofilm formation was compared by plate counting and live/dead staining, as well as by denaturing gradient gel electrophoresis for compositional differences in biofilms. Single-strand wires attracted only slightly less biofilm in vitro than multi-strand wires. Biofilms on stainless steel single-strand wires however, were much more susceptible to antimicrobials from toothpaste slurries and mouthrinses than on single-strand gold wires and biofilms on multi-strand wires. Also, in vivo significantly less biofilm was found on single-strand than on multi-strand wires. Microbial composition of biofilms was more dependent on the volunteer involved than on wire type. Biofilms on single-strand stainless steel wires attract less biofilm in vitro and are more susceptible to antimicrobials than on multi-strand wires. Also in vivo, single-strand wires attract less biofilm than multi-strand ones. Use of single-strand wires is preferred over multi-strand wires, not because they attract less biofilm, but because biofilms on single-strand wires are not protected against antimicrobials as in crevices and niches as on multi-strand wires.

AÇÃO ANTIFÚNGICA DE UM JATO DE PLASMA NÃO-TERMICO DE HÉLIO/AR COMPRIMIDO SOBRE BIOFILMES DE CANDIDA ALBICANS

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Fernanda Ramos Figueira

2017-01-01

Full Text Available Os biofilmes fúngicos são uma predominante causa de infecções crônicas associadas à utilização de cateteres e próteses, conferindo resistência aos antibióticos e fatores imunológicos do hospedeiro, sendo as leveduras, do gênero Candida spp, as mais frequentemente isoladas. O plasma não-térmico, operado à pressão atmosférica, vem ganhando destaque como uma nova estratégia antimicrobiana, inclusive para erradicação de biofilmes. Este trabalho teve como objetivo avaliar a eficácia da inativação de biofilmes de Candida albicans ATCC sobre substrato de poliuretano, utilizando jatos de plasma atmosférico de 6L/min de hélio e 4L/min de ar comprimido, alternando entre sistema contínuo e pulsado, com frequência de 60Hz e distâncias de 10 a 30mm entre o bocal e substrato. Após tratamento, realizou-se a contagem das unidades formadoras de colônia e a análise morfológica da superfície do biofilme por Microscopia Eletrônica de Varredura. O melhor grupo foi o plasma de modo pulsado com distância de 30mm com redução de 92% das unidades formadoras de colônia, demonstrando ser uma tecnologia promissora para o controle de biofilmes de C. albicans.

Antimicrobial activity of denture adhesive associated with Equisetum giganteum- and Punica granatum-enriched fractions against Candida albicans biofilms on acrylic resin surfaces.

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Almeida, Nara Ligia Martins; Saldanha, Luiz Leonardo; da Silva, Rafaela Alves; Pinke, Karen Henriette; da Costa, Eliane Ferraz; Porto, Vinicius Carvalho; Dokkedal, Anne Lígia; Lara, Vanessa Soares

2018-01-01

Candida biofilms adhere to the internal surface of removable dentures, which is an etiological factor in the pathogenesis of denture stomatitis (DS). Adhesive materials are used at the base of maxillary complete dentures to improve their retention and chewing qualities. This article reports the antimicrobial activity of the enriched fractions of Equisetum giganteum and Punica granatum incorporated into a denture adhesive against C. albicans biofilm. The biofilms were induced on the surface of heat-cured acrylic resin specimens that were previously treated with a mixture of adhesive/herb extracts. The antimicrobial activity was evaluated by CFU counts, XTT reduction, and SEM and CLSM analysis. Both herb extracts amplified the anti-biofilm action of the adhesive on the acrylic resin by up to 12 h. Therefore, when these extracts were combined with COREGA®, they played a collaborative and innovative role in biofilm control and can be considered alternatives for temporary use in the treatment and/or prevention of DS.

The Effectiveness of Voriconazole in Therapy of Candida glabrata's Biofilms Oral Infections and Its Influence on the Matrix Composition and Gene Expression.

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Rodrigues, Célia F; Gonçalves, Bruna; Rodrigues, Maria Elisa; Silva, Sónia; Azeredo, Joana; Henriques, Mariana

2017-08-01

Candida glabrata is one of most prevalent yeast in fungal infections, especially in immunocompromised patients. Its azole resistance results in a low therapeutic response, particularly when associated with biofilms. The main goal of this work was to study the effectiveness of voriconazole (Vcz) against C. glabrata biofilms oral pathologies, as esophageal or oropharyngeal candidiasis. Antifungal susceptibilities were determined in pre-formed 24-h-biofilms and ERG genes expression was determined by qRT-PCR. Protein quantification was performed using BCA ® Kit, carbohydrate was estimated according to the Dubois assay and β-1,3 glucans concentration were determined using Glucatell ® kit. Finally, ergosterol, Vcz, and fluconazole (Flu) concentrations within the biofilm matrices were determined by RP-HPLC. Results showed that C. glabrata biofilms were more susceptible to Vcz than to Flu and that ERG genes expression evidenced an overexpression of the three ERG genes in the presence of both azoles. The matrix content presented a remarked decrease in proteins and an increase in carbohydrates, namely β-1,3 glucans. Ergosterol was successfully detected and quantified in the biofilm matrices, with no differences in all the considered conditions. Vcz demonstrated better diffusion through the biofilms and better cell penetration capacities, than Flu, indicating that the structure of the drug molecule fully influences its dissemination through the biofilm matrices. This work showed that Vcz is notably more effective than Flu for the treatment of resistant C. glabrata oral biofilms, which demonstrates a clinical relevance in its future use for the treatment of oropharyngeal/esophageal candidiasis caused by this species.

Effects of lactoferricin B against keratitis-associated fungal biofilms.

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Sengupta, Jayangshu; Saha, Suman; Khetan, Archana; Sarkar, Sujoy K; Mandal, Santi M

2012-10-01

Biofilms are considered as the most important developmental characteristics in ocular infections. Biofilm eradication is a major challenge today to overcome the incidence of drug resistance. This report demonstrates the in vitro ability of biofilm formation on contact lens by three common keratitis-associated fungal pathogens, namely, Aspergillus fumigatus, Fusarium solani, and Candida albicans. Antifungal sensitivity testing performed for both planktonic cells and biofilm revealed the sessile phenotype to be resistant at MIC levels for the planktonic cells and also at higher concentrations. A prototype lens care solution was also found to be partially effective in eradication of the mature biofilm from contact lenses. Lactoferricin B (Lacf, 64 μg/ml), an antimicrobial peptide, exhibited almost no effect on the sessile phenotype. However, the combinatory effect of Lacf with antifungals against planktonic cells and biofilms of three fungal strains that were isolated from keratitis patients exhibited a reduction of antifungal dose more than eightfold. Furthermore, the effect of Lacf in lens care solution against biofilms in which those strains formed was eradicated successfully. These results suggest that lactoferricin B could be a promising candidate for clinical use in improving biofilm susceptibility to antifungals and also as an antibiofilm-antifungal additive in lens care solution.

Cytotoxicity, interaction with dentine and efficacy on multispecies biofilms of a modified salt solution intended for endodontic disinfection in a new in vitro biofilm model.

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van der Waal, S V; Scheres, N; de Soet, J J; Wesselink, P R; Crielaard, W

2015-02-01

To investigate the cytotoxicity of a modified salt solution (MSS) and evaluate the antimicrobial properties of MSS on in vitro biofilm models. In a metabolic assay, fibroblasts derived from periodontal ligaments (PDL) of human extracted teeth were cultured and challenged with MSS or controls. Then, in active attachment biofilm models, the efficacy of MSS in the presence of dentine powder and in eliminating mature biofilms was investigated. In the dentine assay, a biofilm of Enterococcus faecalis was employed. For the final assay, microorganisms were retrieved from infected root canals and cultured to produce biofilms. After the treatments with MSS or the controls, the biofilms were collected, serially diluted and plated. The colony-forming units were counted. One-way anova was used to analyse the differences between the groups. A P 0.05). In endodontic biofilms, the culturable bacteria were equally reduced by MSS, 2% chlorhexidine (CHX) or 2% sodium hypochlorite (NaOCl) (P > 0.05). Modified salt solution is noncytotoxic in vitro and has good antimicrobial properties equal to CHX and NaOCl. Although the results are promising, ex vivo and in vivo studies are needed before its use as an interappointment root canal dressing can be considered. © 2014 International Endodontic Journal. Published by John Wiley & Sons Ltd.

From Mouth to Model: Combining in vivo and in vitro oral biofilm growth

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Barbara Klug

2016-09-01

Full Text Available Oral biofilm studies based on simplified experimental setups are difficult to interpret. Models are limited mostly by the number of bacterial species observed and the insufficiency of artificial media. Few studies have attempted to overcome these limitations and to cultivate native oral biofilm. This study aimed to grow oral biofilm in vivo before transfer to a biofilm reactor for ex-situ incubation. The in-vitro survival of this oral biofilm and the changes in bacterial composition over time were observed. Six human enamel-dentin slabs embedded buccally in dental splints were used as biofilm carriers. Fitted individually to the upper jaw of 25 non-smoking male volunteers, the splints were worn continuously for 48 hours. During this time, tooth-brushing and alcohol-consumption were not permitted. The biofilm was then transferred on slabs into a biofilm reactor and incubated there for 48 hours while being nourished in BHI medium. Live/dead staining and confocal laser scanning microscopy were used to observe bacterial survival over four points in time: directly after removal (T0 and after 1h (T1, 24h (T2 and 48h (T3 of incubation. Bacterial diversity at T0 and T3 was compared with 454-pyrosequencing. Fluorescence in situ hybridization was performed to show specific taxa. Survival curves were calculated with a specially designed MATLAB script. Acacia and QIIME 1.9.1 were used to process pyrosequencing data. SPSS 21.0 and R 3.3.1 were used for statistical analysis.After initial fluctuations at T1, survival curves mostly showed approximation of the bacterial numbers to the initial level at T3. Pyrosequencing analysis resulted in 117 OTUs common to all samples. The genera Streptococcus and Veillonella (both Firmicutes dominated at T0 and T3. They make up two thirds of the biofilm. Genera with lower relative abundance had grown significantly at T3. FISH analysis confirmed the pyrosequencing results, i.e. the predominant staining of Firmicutes. We

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.

A novel in vitro wound biofilm model used to evaluate low-frequency ultrasonic-assisted wound debridement

DEFF Research Database (Denmark)

Crone, S.; Garde, Christian; Bjarnsholt, T.

2015-01-01

Objective: Bacterial biofilms remain difficult to treat. The biofilm mode of growth enables bacteria to survive antibiotic treatment and the inflammatory reaction. Low-frequency ultrasound has recently been shown to improve healing in a variety of settings. It is hypothesised that ultrasound...... disrupts the biofilm leaving bacteria more vulnerable to antiseptic or antibiotic treatment. The objective of this study is to develop a realistic model to elucidate the effect of ultrasound on biofilms. Method: A novel in vitro wound biofilm model was developed. Biofilms of Staphylococcus aureus were...

Assessing the potential of four cathelicidins for the management of mouse candidiasis and Candida albicans biofilms.

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Yu, Haining; Liu, Xuelian; Wang, Chen; Qiao, Xue; Wu, Sijin; Wang, Hui; Feng, Lan; Wang, Yipeng

2016-02-01

As the most common fungal pathogen of humans, severe drug resistance has emerged in the clinically isolated Candida albicans, which lead to the urgency to develop novel antifungal agents. Here, four our previously characterized cathelicidins (cathelicidin-BF, Pc-CATH1, Cc-CATH2, Cc-CATH3) were selected and their antifungal activities against C. albicans were evaluated in vitro and in vivo using amphotericin B and LL-37 as control. Results showed that all four cathelicidins could eradicate standard and clinically isolated C. albicans strains with most MIC values ranging from 1 to 16 μg/ml, in less than 0.5 h revealed by time-kill kinetic assay. Four peptides only exhibited slight hemolytic activity with most HC50 > 200 μg/ml, and retained potent anti-C. albicans activity at salt concentrations below and beyond physiological level. In animal experiment, 50 mg/kg administration of the four cathelicidins could significantly reduce the fungal counts in a murine oral candidiasis model induced by clinically isolated C. albicans. The antibiofilm activity of cathelicidin-BF, the most potent among the five peptides was evaluated, and result showed that cathelicidin-BF strongly inhibited C. albicans biofilm formation at 20 μg/ml. Furthermore, cathelicidin-BF also exhibited potent anti-C. albicans activity in established biofilms as measured by metabolic and fluorescent viability assays. Structure-function analyses suggest that they mainly adopt an α-helical conformations, which enable them to act as a membrane-active molecule. Altogether, the four cathelicidins display great potential for antifungal agent development against candidiasis. Copyright © 2015 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

In vitro evaluation of a multispecies oral biofilm on different implant surfaces

International Nuclear Information System (INIS)

Violant, Deborah; Galofré, Marta; Nart, José; Teles, Ricardo Patricio

2014-01-01

Biofilm accumulation on implant surfaces is one of the most important factors for early and late implant failure. Because of the related clinical implications, the aim of this in vitro study was to compare the bacterial cell attachment of a four-species oral biofilm on titanium discs of purity grade 2 and 4, with machined surfaces and etched-thermochemically modified with Avantblast®. The in vitro biofilm model was composed of early (Actinomyces naeslundii, Streptococcus gordonii), secondary (Veillonella parvula), and intermediate (Fusobacterium nucleatum ssp. polymorphum) colonizers of tooth surfaces. A total of 36 discs were divided into four groups: Tigr2-c (titanium grade 2, machined surface), Tigr2-t (titanium grade 2, modified surface with Avantblast®), Tigr4-c (titanium grade 4, machined surface), Tigr4-t (titanium grade 4, modified surface with Avantblast®). The experiment was repeated three times. Biofilm viability was tested with 1% 2, 3, 5-triphenyltetrazolium chloride solution and bacterial cell quantification by checkerboard DNA–DNA hybridization. Descriptive analysis was performed to evaluate biofilm composition and differences between groups were checked with the Mann–Whitney test (p < 0.05). After one week, multispecies biofilms showed a similar pattern of bacterial composition on all analyzed implant surfaces. The most prevalent bacterium was V. parvula (∼50% of the total biomass), followed by S. gordonii (∼30%), F. nucleatum ssp. polymorphum (∼10%) and A. naeslundii (<5%). Total bacterial biomass was significantly higher in both grade-4-titanium surfaces (p < 0.05). The results demonstrated that not only implant surface treatment, but also titanium purity, influence early bacterial colonization. (paper)

Evaluation of the Enterococcus faecalis Biofilm-Associated Virulence Factors AhrC and Eep in Rat Foreign Body Osteomyelitis and In Vitro Biofilm-Associated Antimicrobial Resistance.

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Kristi L Frank

Full Text Available Enterococcus faecalis can cause healthcare-associated biofilm infections, including those of orthopedic devices. Treatment of enterococcal prosthetic joint infection is difficult, in part, due to biofilm-associated antimicrobial resistance. We previously showed that the E. faecalis OG1RF genes ahrC and eep are in vitro biofilm determinants and virulence factors in animal models of endocarditis and catheter-associated urinary tract infection. In this study, we evaluated the role of these genes in a rat acute foreign body osteomyelitis model and in in vitro biofilm-associated antimicrobial resistance. Osteomyelitis was established for one week following the implantation of stainless steel orthopedic wires inoculated with E. faecalis strains OG1RF, ΩahrC, and ∆eep into the proximal tibiae of rats. The median bacterial loads recovered from bones and wires did not differ significantly between the strains at multiple inoculum concentrations. We hypothesize that factors present at the infection site that affect biofilm formation, such as the presence or absence of shear force, may account for the differences in attenuation in the various animal models we have used to study the ΩahrC and ∆eep strains. No differences among the three strains were observed in the planktonic and biofilm antimicrobial susceptibilities to ampicillin, vancomycin, daptomycin, linezolid, and tetracycline. These findings suggest that neither ahrC nor eep directly contribute to E. faecalis biofilm-associated antimicrobial resistance. Notably, the experimental evidence that the biofilm attachment mutant ΩahrC displays biofilm-associated antimicrobial resistance suggests that surface colonization alone is sufficient for E. faecalis cells to acquire the biofilm antimicrobial resistance phenotype.

Differential control of Yersinia pestis biofilm formation in vitro and in the flea vector by two c-di-GMP diguanylate cyclases.

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Yi-Cheng Sun

2011-04-01

Full Text Available Yersinia pestis forms a biofilm in the foregut of its flea vector that promotes transmission by flea bite. As in many bacteria, biofilm formation in Y. pestis is controlled by intracellular levels of the bacterial second messenger c-di-GMP. Two Y. pestis diguanylate cyclase (DGC enzymes, encoded by hmsT and y3730, and one phosphodiesterase (PDE, encoded by hmsP, have been shown to control biofilm production in vitro via their opposing c-di-GMP synthesis and degradation activities, respectively. In this study, we provide further evidence that hmsT, hmsP, and y3730 are the only three genes involved in c-di-GMP metabolism in Y. pestis and evaluated the two DGCs for their comparative roles in biofilm formation in vitro and in the flea vector. As with HmsT, the DGC activity of Y3730 depended on a catalytic GGDEF domain, but the relative contribution of the two enzymes to the biofilm phenotype was influenced strongly by the environmental niche. Deletion of y3730 had a very minor effect on in vitro biofilm formation, but resulted in greatly reduced biofilm formation in the flea. In contrast, the predominant effect of hmsT was on in vitro biofilm formation. DGC activity was also required for the Hms-independent autoaggregation phenotype of Y. pestis, but was not required for virulence in a mouse model of bubonic plague. Our results confirm that only one PDE (HmsP and two DGCs (HmsT and Y3730 control c-di-GMP levels in Y. pestis, indicate that hmsT and y3730 are regulated post-transcriptionally to differentially control biofilm formation in vitro and in the flea vector, and identify a second c-di-GMP-regulated phenotype in Y. pestis.

Response to antiseptic agents of periodontal pathogens in in vitro biofilms on titanium and zirconium surfaces.

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Sánchez, M C; Fernández, E; Llama-Palacios, A; Figuero, E; Herrera, D; Sanz, M

2017-04-01

The aim of this study was to develop in vitro biofilms on SLA titanium (Ti-SLA) and zirconium oxide (ZrO 2 ) surfaces and to evaluate the effect of antiseptic agents on the number of putative periodontal pathogenic species. An in vitro biofilm model was developed on sterile discs of Ti-SLA and ZrO 2 . Three antiseptic agents [chlorhexidine and cetyl-pyridinium-chloride (CHX/CPC), essential oils (EEOOs) and cetyl-peridinium-chloride (CPC)] were applied to 72-h biofilms, immersing discs during 1min in the antiseptic solution, either with or without mechanical disruption. Viable bacteria [colony forming units (CFU/mL)] were measured by quantitative polymerase chain reaction (qPCR) combined with propidium monoazide. A generalized lineal model was constructed to determine the effect of the agents on the viable bacterial counts of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis and Fusobacterium nucleatum on each surface. The exposure to each antiseptic solution resulted in a statistically significant reductions in the number of viable target species included in the in vitro multi-species biofilm, on both Ti-SLA and ZrO 2 (pzirconium surfaces, in spite of the described structural differences between these bacterial communities. Copyright © 2017 The Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

EFEITO DOS COMPOSTOS ORAGANOCALCOGÊNEOS (PhSe2 E (p-Cl-PhSe2 SOBRE A VIABILIDADE DO BIOFILME FORMADO POR Candida albicans

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Marília Toledo Braga

2017-05-01

Full Text Available Os fungos do gênero Candida são capazes de produzir infecções superficiais ou sistêmicas em indivíduos com baixa resposta imunológica. Estes fungos promovem infecções a partir da sua capacidade de formar biofilmes. Os biofilmes formados por C. albicans são mais resistentes aos tratamentos antifúngicos convencionais. Portanto o objetivo deste trabalho foi avaliar e comparar o potencial antifúngico dos compostos organocalcogêneos (PhSe2 e (p­-Cl-PhSe2 sobre os biofilmes de C. albicans. Suspensões de C. albicans foram incubadas para a formação do biofilme e estes foram tratados com os compostos organocalcogêneos por 24 horas. Os resultados obtidos demonstraram o potencial antifúngico de ambos compostos sobre o biofilme formado por C. albicans, inibindo a viabilidade do biofilme e reduzindo a quantidade de células planctônicas. Esses compostos apresentaram uma resposta terapêutica promissora, uma vez que as opções para tratamento de infecções fúngicas é limitado.

Yeast casein kinase 2 governs morphology, biofilm formation, cell wall integrity, and host cell damage of Candida albicans.

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Jung, Sook-In; Rodriguez, Natalie; Irrizary, Jihyun; Liboro, Karl; Bogarin, Thania; Macias, Marlene; Eivers, Edward; Porter, Edith; Filler, Scott G; Park, Hyunsook

2017-01-01

The regulatory networks governing morphogenesis of a pleomorphic fungus, Candida albicans are extremely complex and remain to be completely elucidated. This study investigated the function of C. albicans yeast casein kinase 2 (CaYck2p). The yck2Δ/yck2Δ strain displayed constitutive pseudohyphae in both yeast and hyphal growth conditions, and formed enhanced biofilm under non-biofilm inducing condition. This finding was further supported by gene expression analysis of the yck2Δ/yck2Δ strain which showed significant upregulation of UME6, a key transcriptional regulator of hyphal transition and biofilm formation, and cell wall protein genes ALS3, HWP1, and SUN41, all of which are associated with morphogenesis and biofilm architecture. The yck2Δ/yck2Δ strain was hypersensitive to cell wall damaging agents and had increased compensatory chitin deposition in the cell wall accompanied by an upregulation of the expression of the chitin synthase genes, CHS2, CHS3, and CHS8. Absence of CaYck2p also affected fungal-host interaction; the yck2Δ/yck2Δ strain had significantly reduced ability to damage host cells. However, the yck2Δ/yck2Δ strain had wild-type susceptibility to cyclosporine and FK506, suggesting that CaYck2p functions independently from the Ca+/calcineurin pathway. Thus, in C. albicans, Yck2p is a multifunctional kinase that governs morphogenesis, biofilm formation, cell wall integrity, and host cell interactions.

In vitro sensitivity of Trichomonas vaginalis and Candida albicans to chemotherapeutic agents.

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Lövgren, T; Salmela, I

1978-06-01

Strains of fresh clinical isolates of Trichomonas vaginalis and Candida albicans have been tested in vitro for their sensitivity to eight drugs used in the therapy of monilial and trichomonal vaginitis. Three of the chemotherapeutic agents, chlorchinaldol, clotrimazole and broxyquinoline were effective against both organisms. Tinidazole and metronidazole were active against T. vaginalis. The strains of C. albicans were also sensitive to trichomycin, natamycin and nystatin. Tinidazole was the most effective trichomonacide, clotrimazole and chlorchinaldol were most effective against C. albicans, while chlorchinaldol had the best in vitro effect against both organisms. The ranges of the MICs are compared to values previously reported.

Silver-Containing Hydroxyapatite Coating Reduces Biofilm Formation by Methicillin-Resistant Staphylococcus aureus In Vitro and In Vivo

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Masaya Ueno

2016-01-01

Full Text Available Biofilm-producing bacteria are the principal causes of infections associated with orthopaedic implants. We previously reported that silver-containing hydroxyapatite (Ag-HA coatings exhibit high antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA. In the present study, we evaluated the effects of Ag-HA coating of implant surfaces on biofilm formation. Titanium disks (14-mm diameter, 1-mm thickness, one surface of which was coated with HA or 0.5%–3.0% Ag-HA with a thermal spraying technique, were used. In vitro, the disks were inoculated with an MRSA suspension containing 4×105 CFU and incubated for 1-2 weeks. In vivo, MRSA-inoculated HA and 3% Ag-HA disks (8.8–10.0 × 108 CFU were implanted subcutaneously on the back of rats for 1–7 days. All disks were subsequently stained with a biofilm dye and observed under a fluorescence microscope, and biofilm coverage rates (BCRs were calculated. The BCRs on the Ag-HA coating were significantly lower than those on the HA coating at all time points in vitro (p<0.05. Similar results were observed in vivo (p<0.001 without argyria. Ag-HA coating reduced biofilm formation by MRSA in vitro and in vivo; therefore, Ag-HA coating might be effective for reducing implant-associated infections.

In vitro susceptibility of Candida spp. to fluconazole, itraconazole and voriconazole and the correlation between triazoles susceptibility: Results from a five-year study.

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Lei, J; Xu, J; Wang, T

2018-06-01

Candida spp. is a common cause of invasive fungal disease. The aim of this study was to examine the susceptibility of Candida spp. to fluconazole, itraconazole and voriconazole and explore the correlation between triazoles susceptibility. The antifungal susceptibility in the present study was measured by ATB Fungus 3 method, and the potential relationship was examined by obtaining the correlation of measured minimal inhibitory concentrations (MICs) of Candida spp. isolates. A total of 2099 clinical isolates of Candida spp. from 1441 patients were analyzed. The organisms included 1435 isolates of Candida albicans, 207 isolates of Candida glabrata, 65 isolates of Candida parapsilosis, 31 isolates of Candida krusei, 268 isolates of Candida tropicalis. Voriconazole and itraconazole were more active than fluconazole and against Candida spp. in vitro. The fluconazole, itraconazole and voriconazole MIC 90 (MIC for 90% of the isolates) for all Candida spp. isolates was 4mg/L, 1mg/L and 0.25mg/L, respectively. There was a moderate correlation between the fluconazole MIC s for Candida spp. isolates and this for voriconazole (R 2 =0.475; P<0.01) and itraconazole (R 2 =0.431; P<0.01). Voriconazole MICs for the Candida spp. isolates also correlated with those for itraconazole (R 2 =0.401; P<0.01). These observations suggest that the in vitro susceptibility of Candida spp. to fluconazole, itraconazole and voriconazole exhibits a moderate correlation. Published by Elsevier Masson SAS.

Candida albicans importance to denture wearers. A literature review.

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Gleiznys, Alvydas; Zdanavičienė, Eglė; Žilinskas, Juozas

2015-01-01

Opportunistic oral fungal infections have spred, especially in denture wearers. Denture stomatitis is a common inflammatory reaction, multifactorial etiology, which is usually associated with Candida species, particularly Candida albicans, due to its high virulence, ability to adhere and form biofilms on oral cavity tissues and denture surfaces. This article highlights the pathogenesis, clinical presentation, and management strategies of Candida-associated denture stomatitis commonly encountered in dental practice.

Effect of a Lactobacillus Salivarius Probiotic on a Double-Species Streptococcus Mutans and Candida Albicans Caries Biofilm.

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Krzyściak, Wirginia; Kościelniak, Dorota; Papież, Monika; Vyhouskaya, Palina; Zagórska-Świeży, Katarzyna; Kołodziej, Iwona; Bystrowska, Beata; Jurczak, Anna

2017-11-14

The aim of the study was to evaluate the anti-cariogenic effects of Lactobacillus salivarius by reducing pathogenic species and biofilm mass in a double-species biofilm model. Coexistence of S. mutans with C. albicans can cause dental caries progression or recurrence of the disease in the future. Fifty-nine children with diagnosed early childhood caries (ECC) were recruited onto the study. The condition of the children's dentition was defined according to the World Health Organization guidelines. The participants were divided into children with initial enamel demineralization and children showing dentin damage. The study was performed on the S. mutans and C. albicans clinical strains, isolated from dental plaque of patients with ECC. The effect of a probiotic containing Lactobacillus salivarius on the ability of S. mutans and C. albicans to produce a double-species biofilm was investigated in an in vitro model. The biomass of the formed/non-degraded biofilm was analyzed on the basis of its crystal violet staining. The number of colonies of S. mutans and C. albicans (CFU/mL, colony forming units/mL) forming the biofilm was determined. Microorganism morphology in the biofilm was evaluated using a scanning electron microscope (SEM). In vitro analysis demonstrated that the presence of S. mutans increased the number of C. albicans colonies (CFU/mL); the double-species biofilm mass and hyphal forms produced in it by the yeast. L. salivarius inhibited the cariogenic biofilm formation of C. albicans and S. mutans . Under the influence of the probiotic; the biofilm mass and the number of S. mutans ; C. albicans and S. mutans with C. albicans colonies in the biofilm was decreased. Moreover; it can be noted that after the addition of the probiotic; fungi did not form hyphae or germ tubes of pathogenic potential. These results suggest that L. salivarius can secrete intermediates capable of inhibiting the formation of cariogenic S. mutans and C. albicans biofilm; and may

Effect of a Lactobacillus Salivarius Probiotic on a Double-Species Streptococcus Mutans and Candida Albicans Caries Biofilm

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Wirginia Krzyściak

2017-11-01

Full Text Available The aim of the study was to evaluate the anti-cariogenic effects of Lactobacillus salivarius by reducing pathogenic species and biofilm mass in a double-species biofilm model. Coexistence of S. mutans with C. albicans can cause dental caries progression or recurrence of the disease in the future. Fifty-nine children with diagnosed early childhood caries (ECC were recruited onto the study. The condition of the children’s dentition was defined according to the World Health Organization guidelines. The participants were divided into children with initial enamel demineralization and children showing dentin damage. The study was performed on the S. mutans and C. albicans clinical strains, isolated from dental plaque of patients with ECC. The effect of a probiotic containing Lactobacillus salivarius on the ability of S. mutans and C. albicans to produce a double-species biofilm was investigated in an in vitro model. The biomass of the formed/non-degraded biofilm was analyzed on the basis of its crystal violet staining. The number of colonies of S. mutans and C. albicans (CFU/mL, colony forming units/mL forming the biofilm was determined. Microorganism morphology in the biofilm was evaluated using a scanning electron microscope (SEM. In vitro analysis demonstrated that the presence of S. mutans increased the number of C. albicans colonies (CFU/mL; the double-species biofilm mass and hyphal forms produced in it by the yeast. L. salivarius inhibited the cariogenic biofilm formation of C. albicans and S. mutans. Under the influence of the probiotic; the biofilm mass and the number of S. mutans; C. albicans and S. mutans with C. albicans colonies in the biofilm was decreased. Moreover; it can be noted that after the addition of the probiotic; fungi did not form hyphae or germ tubes of pathogenic potential. These results suggest that L. salivarius can secrete intermediates capable of inhibiting the formation of cariogenic S. mutans and C. albicans biofilm

In vitro antimicrobial activity of Caesalpinia ferrea Martius fruits against oral pathogens.

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Sampaio, Fábio C; Pereira, Maria do Socorro V; Dias, Celidarque S; Costa, Vicente Carlos O; Conde, Nikeila C O; Buzalaf, Marília A R

2009-07-15

In the Amazon region of Brazil, the fruits of Caesalpinia ferrea Martius (Brazilian ironwood) are widely used as an antimicrobial and healing medicine in many situations including oral infections. This study aimed to evaluate the antimicrobial activity of Caesalpinia ferrea Martius fruit extract against oral pathogens. Polyphenols estimation and spectral analysis ((1)H NMR) of the methanol extract were carried out. The microorganisms Candida albicans, Streptococcus mutans, Streptococcus salivarius, Streptococcus oralis and Lactobacillus casei were tested using the microdilution method for planktonic cells (MIC) and a multispecies biofilm model. Chlorhexidine was used as positive control. Polyphenols in the extract were estimated at 7.3% and (1)H NMR analysis revealed hydroxy phenols and methoxilated compounds. MIC values for Candida albicans, Streptococcus mutans, Streptococcus salivarius, Streptococcus oralis and Lactobacillus casei were 25.0, 40.0, 66.0, 100.0, 66.0 microg/mL, respectively. For the biofilm assay, chlorhexidine and plant extract showed no growth at 10(-4) and 10(-5) microbial dilution, respectively. At 10(-4) and 10(-5) the growth values (mean+/-SD) of the negative controls (DMSO and saline solution) for Streptococcus mutans, Streptococcus sp. and Candida albicans were 8.1+/-0.7, 7.0+/-0.6 and 5.9+/-0.9 x 10(6)CFU, respectively. Caesalpinia ferrea fruit extract can inhibit in vitro growth of oral pathogens in planktonic and biofilm models supporting its use for oral infections.

Competitive Interactions between C. albicans, C. glabrata and C. krusei during Biofilm Formation and Development of Experimental Candidiasis

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Rossoni, Rodnei Dennis; Barbosa, Júnia Oliveira; Vilela, Simone Furgeri Godinho; dos Santos, Jéssica Diane; de Barros, Patrícia Pimentel; Prata, Márcia Cristina de Azevedo; Anbinder, Ana Lia; Fuchs, Beth Burgwyn; Jorge, Antonio Olavo Cardoso; Mylonakis, Eleftherios; Junqueira, Juliana Campos

2015-01-01

In this study, we evaluated the interactions between Candida albicans, Candida krusei and Candida glabrata in mixed infections. Initially, these interactions were studied in biofilms formed in vitro. CFU/mL values of C. albicans were lower in mixed biofilms when compared to the single biofilms, verifying 77% and 89% of C. albicans reduction when this species was associated with C. glabrata and C. krusei, respectively. After that, we expanded this study for in vivo host models of experimental candidiasis. G. mellonella larvae were inoculated with monotypic and heterotypic Candida suspensions for analysis of survival rate and quantification of fungal cells in the haemolymph. In the groups with single infections, 100% of the larvae died within 18 h after infection with C. albicans. However, interaction groups achieved 100% mortality after 72 h of infection by C. albicans-C. glabrata and 96 h of infection by C. albicans-C. krusei. C. albicans CFU/mL values from larvae hemolymph were lower in the interacting groups compared with the monoespecies group after 12 h of infection. In addition, immunosuppressed mice were also inoculated with monotypic and heterotypic microbial suspensions to induce oral candidiasis. C. albicans CFU/mL values recovered from oral cavity of mice were higher in the group with single infection by C. albicans than the groups with mixed infections by C. albicans-C. glabrata and C. albicans-C. krusei. Moreover, the group with single infection by C. albicans had a higher degree of hyphae and epithelial changes in the tongue dorsum than the groups with mixed infections. We concluded that single infections by C. albicans were more harmful for animal models than mixed infections with non-albicans species, suggesting that C. albicans establish competitive interactions with C. krusei and C. glabrata during biofilm formation and development of experimental candidiasis. PMID:26146832

Competitive Interactions between C. albicans, C. glabrata and C. krusei during Biofilm Formation and Development of Experimental Candidiasis.

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Rossoni, Rodnei Dennis; Barbosa, Júnia Oliveira; Vilela, Simone Furgeri Godinho; dos Santos, Jéssica Diane; de Barros, Patrícia Pimentel; Prata, Márcia Cristina de Azevedo; Anbinder, Ana Lia; Fuchs, Beth Burgwyn; Jorge, Antonio Olavo Cardoso; Mylonakis, Eleftherios; Junqueira, Juliana Campos

2015-01-01

In this study, we evaluated the interactions between Candida albicans, Candida krusei and Candida glabrata in mixed infections. Initially, these interactions were studied in biofilms formed in vitro. CFU/mL values of C. albicans were lower in mixed biofilms when compared to the single biofilms, verifying 77% and 89% of C. albicans reduction when this species was associated with C. glabrata and C. krusei, respectively. After that, we expanded this study for in vivo host models of experimental candidiasis. G. mellonella larvae were inoculated with monotypic and heterotypic Candida suspensions for analysis of survival rate and quantification of fungal cells in the haemolymph. In the groups with single infections, 100% of the larvae died within 18 h after infection with C. albicans. However, interaction groups achieved 100% mortality after 72 h of infection by C. albicans-C. glabrata and 96 h of infection by C. albicans-C. krusei. C. albicans CFU/mL values from larvae hemolymph were lower in the interacting groups compared with the monoespecies group after 12 h of infection. In addition, immunosuppressed mice were also inoculated with monotypic and heterotypic microbial suspensions to induce oral candidiasis. C. albicans CFU/mL values recovered from oral cavity of mice were higher in the group with single infection by C. albicans than the groups with mixed infections by C. albicans-C. glabrata and C. albicans-C. krusei. Moreover, the group with single infection by C. albicans had a higher degree of hyphae and epithelial changes in the tongue dorsum than the groups with mixed infections. We concluded that single infections by C. albicans were more harmful for animal models than mixed infections with non-albicans species, suggesting that C. albicans establish competitive interactions with C. krusei and C. glabrata during biofilm formation and development of experimental candidiasis.

Development of an in vitro Assay, based on the BioFilm Ring Test®, for Rapid Profiling of Biofilm-Growing Bacteria

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Enea Gino Di Domenico

2016-09-01

Full Text Available Microbial biofilm represents a major virulence factor associated with chronic and recurrent infections. Pathogenic bacteria embedded in biofilms are highly resistant to environmental and chemical agents, including antibiotics and therefore difficult to eradicate. Thus, reliable tests to assess biofilm formation by bacterial strains as well as the impact of chemicals or antibiotics on biofilm formation represent desirable tools for a most effective therapeutic management and microbiological risk control. Current methods to evaluate biofilm formation are usually time-consuming, costly, and hardly applicable in the clinical setting.The aim of the present study was to develop and assess a simple and reliable in vitro procedure for the characterization of biofilm-producing bacterial strains for future clinical applications based on the BioFilm Ring Test® (BRT technology. The procedure developed for clinical testing (cBRT can provide an accurate and timely (5 hours measurement of biofilm formation for the most common pathogenic bacteria seen in clinical practice. The results gathered by the cBRT assay were in agreement with the traditional crystal violet (CV staining test, according to the kappa coefficient test (kappa = 0.623. However, the cBRT assay showed higher levels of specificity (92.2% and accuracy (88.1% as compared to CV. The results indicate that this procedure offers an easy, rapid and robust assay to test microbial biofilm and a promising tool for clinical microbiology.

Species distribution and in vitro fluconazole susceptibility of clinical Candida isolates in a Brazilian tertiary-care hospital over a 3-year period

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Márcia Cristina Furlaneto

2011-10-01

Full Text Available INTRODUCTION: In this study, we aimed at identifying Candida isolates obtained from blood, urine, tracheal secretion, and nail/skin lesions from cases attended at the Hospital Universitário de Londrina over a 3-year period and at evaluating fluconazole susceptibilities of the isolates. METHODS: Candida isolates were identified by polymerase chain reaction (PCR using species-specific forward primers. The in vitro fluconazole susceptibility test was performed according to EUCAST-AFST reference procedure. RESULTS: Isolates were obtained from urine (53.4%, blood cultures (19.2%, tracheal secretion (17.8%, and nail/skin lesions (9.6%. When urine samples were considered, prevalence was similar in women (45.5% and in men (54.5% and was high in the age group >61 years than that in younger ones. For blood samples, prevalence was high in neonates (35% and advanced ages (22.5%. For nail and skin samples, prevalence was higher in women (71.4% than in men (28.6%. Candida albicans was the most frequently isolated in the hospital, but Candida species other than C. albicans accounted for 64% of isolates, including predominantly Candida tropicalis (33.2% and Candida parapsilosis (19.2%. The trend for non-albicans Candida as the predominant species was noted from all clinical specimens, except from urine samples. All Candida isolates were considered susceptible in vitro to fluconazole with the exception of isolates belonging to the intrinsically less-susceptible species C. glabrata. CONCLUSIONS: Non-albicans Candida species were more frequently isolated in the hospital. Fluconazole resistance was a rare finding in our study.

Identification of proteins involved in the adhesionof Candida species to different medical devices.

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Núñez-Beltrán, Arianna; López-Romero, Everardo; Cuéllar-Cruz, Mayra

2017-06-01

Adhesion is the first step for Candida species to form biofilms on medical devices implanted in the human host. Both the physicochemical nature of the biomaterial and cell wall proteins (CWP) of the pathogen play a determinant role in the process. While it is true that some CWP have been identified in vitro, little is known about the CWP of pathogenic species of Candida involved in adhesion. On this background, we considered it important to investigate the potential role of CWP of C. albicans, C. glabrata, C. krusei and C. parapsilosis in adhesion to different medical devices. Our results indicate that the four species strongly adher to polyvinyl chloride (PVC) devices, followed by polyurethane and finally by silicone. It was interesting to identify fructose-bisphosphate aldolase (Fba1) and enolase 1 (Eno1) as the CWP involved in adhesion of C. albicans, C. glabrata and C. krusei to PVC devices whereas phosphoglycerate kinase (Pgk) and Eno1 allow C. parapsilosis to adher to silicone-made implants. Results presented here suggest that these CWP participate in the initial event of adhesion and are probably followed by other proteins that covalently bind to the biomaterial thus providing conditions for biofilm formation and eventually the onset of infection. Copyright © 2017 Elsevier Ltd. All rights reserved.

Targeted changes of the cell wall proteome influence Candida albicans ability to form single- and multi-strain biofilms.

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Vitor Cabral

2014-12-01

Full Text Available Biofilm formation is an important virulence trait of the pathogenic yeast Candida albicans. We have combined gene overexpression, strain barcoding and microarray profiling to screen a library of 531 C. albicans conditional overexpression strains (∼10% of the genome for genes affecting biofilm development in mixed-population experiments. The overexpression of 16 genes increased strain occupancy within a multi-strain biofilm, whereas overexpression of 4 genes decreased it. The set of 16 genes was significantly enriched for those encoding predicted glycosylphosphatidylinositol (GPI-modified proteins, namely Ihd1/Pga36, Phr2, Pga15, Pga19, Pga22, Pga32, Pga37, Pga42 and Pga59; eight of which have been classified as pathogen-specific. Validation experiments using either individually- or competitively-grown overexpression strains revealed that the contribution of these genes to biofilm formation was variable and stage-specific. Deeper functional analysis of PGA59 and PGA22 at a single-cell resolution using atomic force microscopy showed that overexpression of either gene increased C. albicans ability to adhere to an abiotic substrate. However, unlike PGA59, PGA22 overexpression led to cell cluster formation that resulted in increased sensitivity to shear forces and decreased ability to form a single-strain biofilm. Within the multi-strain environment provided by the PGA22-non overexpressing cells, PGA22-overexpressing cells were protected from shear forces and fitter for biofilm development. Ultrastructural analysis, genome-wide transcript profiling and phenotypic analyses in a heterologous context suggested that PGA22 affects cell adherence through alteration of cell wall structure and/or function. Taken together, our findings reveal that several novel predicted GPI-modified proteins contribute to the cooperative behaviour between biofilm cells and are important participants during C. albicans biofilm formation. Moreover, they illustrate the power

Comparative Phenotypic Analysis of the Major Fungal Pathogens Candida parapsilosis and Candida albicans

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Holland, Linda M.; Schröder, Markus S.; Turner, Siobhán A.; Taff, Heather; Andes, David; Grózer, Zsuzsanna; Gácser, Attila; Ames, Lauren; Haynes, Ken; Higgins, Desmond G.; Butler, Geraldine

2014-01-01

Candida parapsilosis and Candida albicans are human fungal pathogens that belong to the CTG clade in the Saccharomycotina. In contrast to C. albicans, relatively little is known about the virulence properties of C. parapsilosis, a pathogen particularly associated with infections of premature neonates. We describe here the construction of C. parapsilosis strains carrying double allele deletions of 100 transcription factors, protein kinases and species-specific genes. Two independent deletions were constructed for each target gene. Growth in >40 conditions was tested, including carbon source, temperature, and the presence of antifungal drugs. The phenotypes were compared to C. albicans strains with deletions of orthologous transcription factors. We found that many phenotypes are shared between the two species, such as the role of Upc2 as a regulator of azole resistance, and of CAP1 in the oxidative stress response. Others are unique to one species. For example, Cph2 plays a role in the hypoxic response in C. parapsilosis but not in C. albicans. We found extensive divergence between the biofilm regulators of the two species. We identified seven transcription factors and one protein kinase that are required for biofilm development in C. parapsilosis. Only three (Efg1, Bcr1 and Ace2) have similar effects on C. albicans biofilms, whereas Cph2, Czf1, Gzf3 and Ume6 have major roles in C. parapsilosis only. Two transcription factors (Brg1 and Tec1) with well-characterized roles in biofilm formation in C. albicans do not have the same function in C. parapsilosis. We also compared the transcription profile of C. parapsilosis and C. albicans biofilms. Our analysis suggests the processes shared between the two species are predominantly metabolic, and that Cph2 and Bcr1 are major biofilm regulators in C. parapsilosis. PMID:25233198

The antagonistic effect of Saccharomyces boulardii on Candida albicans filamentation, adhesion and biofilm formation.

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Krasowska, Anna; Murzyn, Anna; Dyjankiewicz, Agnieszka; Łukaszewicz, Marcin; Dziadkowiec, Dorota

2009-12-01

The dimorphic fungus Candida albicans is a member of the normal flora residing in the intestinal tract of humans. In spite of this, under certain conditions it can induce both superficial and serious systemic diseases, as well as be the cause of gastrointestinal infections. Saccharomyces boulardii is a yeast strain that has been shown to have applications in the prevention and treatment of intestinal infections caused by bacterial pathogens. The purpose of this study was to determine whether S. boulardii affects the virulence factors of C. albicans. We demonstrate the inhibitory effect of live S. boulardii cells on the filamentation (hyphae and pseudohyphae formation) of C. albicans SC5314 strain proportional to the amount of S. boulardii added. An extract from S. boulardii culture has a similar effect. Live S. boulardii and the extract from S. boulardii culture filtrate diminish C. albicans adhesion to and subsequent biofilm formation on polystyrene surfaces under both aerobic and microaerophilic conditions. This effect is very strong and requires lower doses of S. boulardii cells or concentrations of the extract than serum-induced filamentation tests. Saccharomyces boulardii has a strong negative effect on very important virulence factors of C. albicans, i.e. the ability to form filaments and to adhere and form biofilms on plastic surfaces.

Early staphylococcal biofilm formation on solid orthopaedic implant materials: in vitro study.

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Hironobu Koseki

Full Text Available Biofilms forming on the surface of biomaterials can cause intractable implant-related infections. Bacterial adherence and early biofilm formation are influenced by the type of biomaterial used and the physical characteristics of implant surface. In this in vitro research, we evaluated the ability of Staphylococcus epidermidis, the main pathogen in implant-related infections, to form biofilms on the surface of the solid orthopaedic biomaterials, oxidized zirconium-niobium alloy, cobalt-chromium-molybdenum alloy (Co-Cr-Mo, titanium alloy (Ti-6Al-4V, commercially pure titanium (cp-Ti and stainless steel. A bacterial suspension of Staphylococcus epidermidis strain RP62A (ATCC35984 was added to the surface of specimens and incubated. The stained biofilms were imaged with a digital optical microscope and the biofilm coverage rate (BCR was calculated. The total amount of biofilm was determined with the crystal violet assay and the number of viable cells in the biofilm was counted using the plate count method. The BCR of all the biomaterials rose in proportion to culture duration. After culturing for 2-4 hours, the BCR was similar for all materials. However, after culturing for 6 hours, the BCR for Co-Cr-Mo alloy was significantly lower than for Ti-6Al-4V, cp-Ti and stainless steel (P0.05. These results suggest that surface properties, such as hydrophobicity or the low surface free energy of Co-Cr-Mo, may have some influence in inhibiting or delaying the two-dimensional expansion of biofilm on surfaces with a similar degree of smoothness.

The Emerging Pathogen Candida auris: Growth Phenotype, Virulence Factors, Activity of Antifungals, and Effect of SCY-078, a Novel Glucan Synthesis Inhibitor, on Growth Morphology and Biofilm Formation.

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Larkin, Emily; Hager, Christopher; Chandra, Jyotsna; Mukherjee, Pranab K; Retuerto, Mauricio; Salem, Iman; Long, Lisa; Isham, Nancy; Kovanda, Laura; Borroto-Esoda, Katyna; Wring, Steve; Angulo, David; Ghannoum, Mahmoud

2017-05-01

Candida auris , a new multidrug-resistant Candida spp. which is associated with invasive infection and high rates of mortality, has recently emerged. Here, we determined the virulence factors (germination, adherence, biofilm formation, phospholipase and proteinase production) of 16 C. auris isolates and their susceptibilities to 11 drugs belonging to different antifungal classes, including a novel orally bioavailable 1,3-β-d-glucan synthesis inhibitor (SCY-078). We also examined the effect of SCY-078 on the growth, ultrastructure, and biofilm-forming abilities of C. auris Our data showed that while the tested strains did not germinate, they did produce phospholipase and proteinase in a strain-dependent manner and had a significantly reduced ability to adhere and form biofilms compared to that of Candida albicans ( P = 0.01). C. auris isolates demonstrated reduced susceptibility to fluconazole and amphotericin B, while, in general, they were susceptible to the remaining drugs tested. SCY-078 had an MIC 90 of 1 mg/liter against C. auris and caused complete inhibition of the growth of C. auris and C. albicans Scanning electron microscopy analysis showed that SCY-078 interrupted C. auris cell division, with the organism forming abnormal fused fungal cells. Additionally, SCY-078 possessed potent antibiofilm activity, wherein treated biofilms demonstrated significantly reduced metabolic activity and a significantly reduced thickness compared to the untreated control ( P < 0.05 for both comparisons). Our study shows that C. auris expresses several virulence determinants (albeit to a lesser extent than C. albicans ) and is resistant to fluconazole and amphotericin B. SCY-078, the new orally bioavailable antifungal, had potent antifungal/antibiofilm activity against C. auris , indicating that further evaluation of this antifungal is warranted. Copyright © 2017 Larkin et al.

Rapid in Vitro Quantification of S. aureus Biofilms on Vascular Graft Surfaces

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Monika Herten

2017-12-01

Full Text Available Objectives: Increasing resistance of microorganisms and particularly tolerance of bacterial biofilms against antibiotics require the need for alternative antimicrobial substances. S. aureus is the most frequent pathogen causing vascular graft infections. In order to evaluate the antimicrobial efficacy, quantification of the bacterial biofilms is necessary. Aim of the present study was the validation of an in vitro model for quantification of bacterial biofilm on vascular graft surfaces using three different assays.Methods: Standardized discs of vascular graft material (Dacron or PTFE or polystyrene (PS as control surface with 0.25 cm2 surface area were inoculated with 10−3 diluted overnight culture of three biofilm-producing S. aureus isolates (BEB-029, BEB-295, SH1000 in 96-well PS culture plates. After incubation for 4 and 18 h, the biofilm was determined by three different methods: (a mitochondrial ATP concentration as measure of bacterial viability (ATP, (b crystal violet staining (Cry, and (c vital cell count by calculation of colony-forming units (CFU. The experiments were performed three times. Quadruplicates were used for each isolate, time point, and method. In parallel, bacterial biofilms were documented via scanning electron microscopy.Results: All three methods could quantify biofilms on the PS control. Time needed was 0:40, 13:10, and 14:30 h for ATP, Cry, and CFU, respectively. The Cry assay could not be used for vascular graft surfaces due to high unspecific background staining. However, ATP assay and CFU count showed comparable results on vascular graft material and control. The correlations between ATP and CFU assay differed according to the surface and incubation time and were significant only after 4 h on Dacron (BEB-029, p = 0.013 and on PS (BEB-029, p < 0.001. Between ATP and Cry assay on PS, a significant correlation could be detected after 4 h (BEB-295, p = 0.027 and after 18 h (all three strains, p < 0.026. The

Aspectos micológicos e suscetibilidade in vitro de leveduras do gênero Candida em pacientes HIV-positivos provenientes do Estado de Mato Grosso Mycological aspects and susceptibility in vitro the yeast of the genus Candida from HIV-positive patients in the State of Mato Grosso

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Olivia Cometti Favalessa

2010-12-01

Full Text Available INTRODUÇÃO: A candidíase é uma das infecções fúngicas mais frequentes entre os pacientes infectados pelo vírus da imunodeficiência humana. O presente estudo objetivou a caracterização das leveduras do gênero Candida de distintas amostras clínicas, provenientes de pacientes HIV - positivos, assim como a determinação do perfil de suscetibilidade in vitro a cinco drogas antifúngicas. MÉTODOS: A caracterização dos isolados de Candida sp foi realizada através da metodologia clássica, testes bioquímicos (zimograma e auxanograma e morfológicos (prova do tubo germinativo e microcultivo em lâmina. Também, foram realizadas a técnica genotípica (PCR e identificação pelo método comercial API 20C AUX (BioMeriéux. Para a determinação do perfil de suscetibilidade in vitro, foram utilizadas cinco drogas antifúngicas (cetoconazol, fluconazol, itraconazol, voriconazol e anfotericina B, através do método comercialmente disponível - Etest. RESULTADOS: Foram identificados 105 isolados de leveduras do gênero Candida provenientes de 102 pacientes infectados pelo vírus HIV. Destes, foram caracterizadas 82 (78,1% Candida albicans, 8 (7,6% Candida parapsilosis, 8 (7,6% Candida tropicalis, 4 (3,8% Candida krusei, 2 (1,9% Candida glabrata e 1 (1% Candida guilliermondii. CONCLUSÕES: Considerando o perfil geral de sensibilidade, 60% dos isolados foram suscetíveis a todos os antifúngicos testados, porém as espécies C. tropicalis e C. krusei demonstraram uma tendência a valores mais elevados de CIMs para os azóis do que os encontrados paraC. albicans, sugerindo resistência.INTRODUCTION: Candidiasis is one of the most common fungal infections among patients infected by human immunodeficiency virus. The present study aimed to characterize yeasts of the genus Candida from distinct clinical samples from HIV-positive patients and determine the in vitro susceptibility profile to five antifungal drugs. METHODS: Characterization of

Streptococcus mutans competence-stimulating peptide inhibits Candida albicans hypha formation

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Jarosz, L.M.; Deng, D.M.; van der Mei, H.C.; Crielaard, W.; Krom, B.P.

2009-01-01

The oral cavity is colonized by microorganisms growing in biofilms in which interspecies interactions take place. Streptococcus mutans grows in biofilms on enamel surfaces and is considered one of the main etiological agents of human dental caries. Candida albicans is also commonly found in the

Streptococcus mutans Competence-Stimulating Peptide Inhibits Candida albicans Hypha Formation

NARCIS (Netherlands)

Jarosz, Lucja M.; Deng, Dong Mei; van der Mei, Henny C.; Crielaard, Wim; Krom, Bastiaan P.

2009-01-01

The oral cavity is colonized by microorganisms growing in biofilms in which interspecies interactions take place. Streptococcus mutans grows in biofilms on enamel surfaces and is considered one of the main etiological agents of human dental caries. Candida albicans is also commonly found in the

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

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

2018-04-01

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

Effects of commonly used food preservatives on biofilm formation of Streptococcus mutans in vitro.

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Al-Ahmad, Ali; Wiedmann-Al-Ahmad, Margit; Auschill, Thorsten Mathias; Follo, Marie; Braun, Gabriele; Hellwig, Elmar; Arweiler, Nicole Birgit

2008-08-01

Sodium benzoate (SB), potassium sorbate (PS) and sodium nitrite (SN) are commonly used food preservatives. In this in vitro study, the effects of these substances on biofilm formation of Streptococcus mutans were analysed. In addition to the microtiter plate test (MPT), a biofilm reactor containing bovine enamel slabs (BES) was used to study the influence of food preservatives on biofilm formation in 5 independent periods of 4 days each. These included one period with chlorhexidine digluconate (CHX) as a positive control as well as a period with growth medium alone as a negative control. The vitality of the biofilm on BES was detected using live/dead staining and confocal laser scanning microscopy. Additionally, the number of colony forming units (CFU) was determined. In MPT 0.12% SN significantly reduced the biofilm formation. PS at a concentration of 0.4% tended to inhibit biofilm formation, whereas the inhibition for 0.8% PS was significant. Less inhibition was caused by 0.8% SB. In the biofilm reactor 0.06% of SN, 0.1% of SB and 0.1% PS significantly reduced the covering grade as well as the CFU of the biofilm. Biofilm vitality was reduced significantly by CHX to a level of 32.5% compared to the control. Only SB reduced the vitality to a level of 19.1%. SN and PS showed no influence on biofilm vitality. This study indicates the potential of food preservatives as inhibitory agents in S. mutans biofilm formation, which should be kept in mind when studying the effects of conserved food on dental plaque biofilm in situ.

Alternative Mating Type Configurations (a/α versus a/a or α/α) of Candida albicans Result in Alternative Biofilms Regulated by Different Pathways

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Srikantha, Thyagarajan; Huang, Guanghua; Garnaas, Adam M.; Soll, David R.

2011-01-01

Similar multicellular structures can evolve within the same organism that may have different evolutionary histories, be controlled by different regulatory pathways, and play similar but nonidentical roles. In the human fungal pathogen Candida albicans, a quite extraordinary example of this has occurred. Depending upon the configuration of the mating type locus (a/α versus a/a or α/α), C. albicans forms alternative biofilms that appear similar morphologically, but exhibit dramatically different characteristics and are regulated by distinctly different signal transduction pathways. Biofilms formed by a/α cells are impermeable to molecules in the size range of 300 Da to 140 kDa, are poorly penetrated by human polymorphonuclear leukocytes (PMNs), and are resistant to antifungals. In contrast, a/a or α/α biofilms are permeable to molecules in this size range, are readily penetrated by PMNs, and are susceptible to antifungals. By mutational analyses, a/α biofilms are demonstrated to be regulated by the Ras1/cAMP pathway that includes Ras1→Cdc35→cAMP(Pde2—|)→Tpk2(Tpk1)→Efg1→Tec1→Bcr1, and a/a biofilms by the MAP kinase pathway that includes Mfα→Ste2→ (Ste4, Ste18, Cag1)→Ste11→Hst7→Cek2(Cek1)→Tec1. These observations suggest the hypothesis that while the upstream portion of the newly evolved pathway regulating a/a and α/α cell biofilms was derived intact from the upstream portion of the conserved pheromone-regulated pathway for mating, the downstream portion was derived through modification of the downstream portion of the conserved pathway for a/α biofilm formation. C. albicans therefore forms two alternative biofilms depending upon mating configuration. PMID:21829325

Antimicrobial Efficacy of Salvadora persica Extracts on a Monospecies Biofilm on Orthodontic Brackets In Vitro.

Science.gov (United States)

Halawany, Hassan S; Abraham, Nimmi B; Siddiqui, Yunus M; Balto, Hanan A; Jacob, Vimal

2016-01-01

The oral cavity is a rich ecosystem with a plethora of microorganisms, and different components of fixed orthodontic appliances may contribute to a shift in the balance of oral ecology. The purpose of this study was to investigate the antimicrobial potential of hexane and ethanol extracts of Salvadora persica on a monospecies biofilm model established on orthodontic brackets in vitro. Streptococcus mutans biofilm was formed on mini diamond orthodontic brackets following three days of anaerobic incubation at 37˚C. The bacterial cell viability of this biofilm was measured after their exposure to saline, hexane extract of S. persica, ethanol extract of S. persica and 0.2% chlorhexidine using 3-(4, 5-dimethylthiazol- 2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulphophenyl)-2H-tetrazolium (MTS) assay. On half of the brackets, the colony forming units (CFU) were counted. Both experiments were performed in triplicate. The absorbance values obtained from the MTS reduction assay after exposure to the different test agents showed a decline in the bacterial cell viability of the S. mutans biofilm as follows: chlorhexidine (+)0.05). The CFU counts of S. mutans obtained from chlorhexidine exposure were lower than from hexane and ethanol extracts. S. persica extracts were found to have antimicrobial effects on S. mutans biofilm established in vitro on orthodontic brackets suggestive of its potential use as an oral antimicrobial agent for orthodontic patients.

Strawberry Extract’s Effects on Enterococcus faecalis and Porphyromonas gingivalis Biofilms in vitro

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Armelia Sari Widyarman

2017-09-01

Full Text Available Background: Enterococcus faecalis (E. faecalis and Porphyromonas gingivalis (P. gingivalis are oral bacteria related to root canal infection and periodontal disease pathogenesis. Strawberries (Fragaria x ananassa fruit are rich in vitamins and minerals, have antibacterial and antioxidant effects. Objective: This study investigated the inhibition effect of strawberry extract on monospecies and multispecies E. faecalis and P. gingivalis bacteria grown as biofilms in vitro. Methods: This study used E. faecalis ATCC 29212 and P. gingivalis ATCC 33277. It analyzed the effect of strawberry extract on bacteria biofilm formation using a biofilm assay on microplate wells. Five concentrations of strawberry extracts were used (100%, 50%, 25%, 12.5%, and 6.25%, and the inhibition effect was observed after a 1h, 3h, 6h, and 24h incubation period. Biofilms without the strawberry extract were used as the negative controls, and crystal violet and safranin (0.5%w/v were used to count the biofilm mass. The biofilms grown on microplates were counted using an ELISA reader at 450 nm after 200 mL of 90% ethanol was added to attract the absorbed stain. The strawberry extract inhibition effectiveness on the biofilm formation of each bacterium tested was analyzed using one-way Anova, where p<0.05 was defined as a significant difference. Result: The strawberry extract inhibited the tested monospecies and multispecies bacteria biofilm formation. The optimal strawberry extract concentration for the inhibition of either monospecies biofilms was 100%. However, the optimal incubation time for the strawberry extract to inhibit the multispecies biofilm formation was 24h, which was the study’s biofilm maturity phase. Conclusions: The 100% strawberry extract concentration inhibited the formation of both the monospecies and multispecies E. faecalis and P. gingivalis biofilms. Future studies are needed to evaluate the potential of strawberry extract as an alternative dental

Methods for dynamic investigations of surface-attached in vitro bacterial and fungal biofilms

DEFF Research Database (Denmark)

Sternberg, Claus; Bjarnsholt, Thomas; Shirtliff, Mark

2014-01-01

Three dynamic models for the investigation of in vitro biofilm formation are described in this chapter. In the 6-well plate assay presented here, the placing of the plate on a rotating platform provides shear, thereby making the system dynamic with respect to the static microtiter assay.The second...... reported model, especially suitable for harvesting high amounts of cells for transcriptomic or proteomic investigations, is based on numerous glass beads placed in a flask incubated with shaking on a rotating platform, thus increasing the surface area for biofilm formation. Finally, the flow-cell system...

In vitro synergy of natamycin and voriconazole against clinical isolates of Fusarium, Candida, Aspergillus and Curvularia spp.

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Sradhanjali, Swatishree; Yein, Bandana; Sharma, Savitri; Das, Sujata

2018-01-01

To determine the minimum inhibitory concentrations (MICs) of voriconazole and natamycin, alone and in combination, against the clinical isolates of Fungus and to evaluate the synergy between the drugs in an experimental in vitro study. In an experimental in vitro study, clinical isolates of Fusarium , Aspergillus , Candida and Curvularia spp were maintained on Sabouraud Dextrose Agar and used for the study. The MICs of natamycin and voriconazole, used alone and in combination, were evaluated by checkerboard microdilution technique based on the standard protocol proposed by the Clinical Laboratory Standards Institute. The interactions were assessed using the fractional inhibitory concentration (FIC) Index model. Tested with all the clinical isolates, the MICs ranged between 0.125 and 8 µg/mL both for natamycin and voriconazole. In descending order, maximum synergism (FIC ≤0.5) was observed in Candida spp (33.3%) followed by Curvularia spp and Fusarium spp (23.1%). Synergism was least for Aspergillus spp (22.2%). However, at 61.5% (8/13), maximum additive effect (>0.5-1) was observed in Aspergillus spp and minimum (33.3%, 2/6) in Candida spp. Indifference (FIC value >1 and≤4) was observed in 22.2% (2/9) of Aspergillus spp, 15.4% (2/13) of Fusarium spp, 33.3% (2/6) of Candida spp and 23.1% (3/13) of Curvularia spp. No cases of antagonism (FIC >4) were observed. Natamycin and voriconazole in combination demonstrated more effective antifungal activity than single-use in vitro treatment in all species tested, which implies that these combinations may be helpful in treating fungal keratitis. There was no antagonism between these two drugs. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Inhibitory Effect of Curcumin on Candida-albicans compared with Nystatin: an in-vitro Study

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Neda Babaii

2016-10-01

Full Text Available Introduction: Curcumin is the active ingredient in the traditional herbal remedy and dietary spice turmeric (Curcuma longa. Curcumin has a surprisingly wide range of beneficial properties, including anti-inflammatory, antioxidant, chemopreventive and chemotherapeutic activity. on basis of recent studies; it has antifungal and antibacterial effects. The aim of this study was in-vitro evaluation of antifungal effect of curcumin on candida albicans and comparing it with nystatin. Methods: after preparing curcumin powder, 3 laboratory methods were used to evaluate antifungal effect. The first method was cell count technique, used to evaluate the amount of candida albicans after time, in different concentrations of curcumin in Dimethyl sulfoxide (DMSO. The second was cup bioassay, in which inhibitory a zone of curcumin in DMSO was evaluated in sabouraud culture plates; and in third method, inhibitory zones of dried disks; which contained curcumin in DMSO were evaluated. Results: the result of all three methods showed that curcumin has antifungal effect and this effect increases in more concentrations. Conclusion: curcumin has apparent and dose dependent antifungal effect on candida albicans.

Disruption of the ECM33 Gene in Candida albicans Prevents Biofilm Formation, Engineered Human Oral Mucosa Tissue Damage and Gingival Cell Necrosis/Apoptosis

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Mahmoud Rouabhia

2012-01-01

Full Text Available In this study we demonstrated that ΔCaecm33 double mutant showed reduced biofilm formation and causes less damage to gingival mucosa tissues. This was confirmed by the reduced level of necrotic cells and Bax/Bcl2 gene expression as apoptotic markers. In contrast, parental and Caecm33 mutant strains decreased basement membrane protein production (laminin 5 and type IV collagen. We thus propose that ECM33 gene/protein represents a novel target for the prevention and treatment of infections caused by Candida.

Biofilm inhibitory and eradicating activity of wound care products against Staphylococcus aureus and Staphylococcus epidermidis biofilms in an in vitro chronic wound model.

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Brackman, G; De Meyer, L; Nelis, H J; Coenye, T

2013-06-01

Although several factors contribute to wound healing, bacterial infections and the presence of biofilm can significantly affect healing. Despite that this clearly indicates that therapies should address biofilm in wounds, only few wound care products have been evaluated for their antibiofilm effect. For this reason, we developed a rapid quantification approach to investigate the efficacy of wound care products on wounds infected with Staphylococcus spp. An in vitro chronic wound infection model was used in which a fluorescent Staph. aureus strain was used to allow the rapid quantification of the bacterial burden after treatment. A good correlation was observed between the fluorescence signal and the bacterial counts. When evaluated in this model, several commonly used wound dressings and wound care products inhibited biofilm formation resulting in a decrease between one and seven log CFU per biofilm compared with biofilm formed in the absence of products. In contrast, most dressings only moderately affected mature biofilms. Our model allowed the rapid quantification of the bacterial burden after treatment. However, the efficacy of treatment varied between the different types of dressings and/or wound care products. Our model can be used to compare the efficacy of wound care products to inhibit biofilm formation and/or eradicate mature biofilms. In addition, the results indicate that treatment of infected wounds should be started as soon as possible and that novel products with more potent antibiofilm activity are needed. © 2013 The Society for Applied Microbiology.

AzaSite® inhibits Staphylococcus aureus and coagulase-negative Staphylococcus biofilm formation in vitro.

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Wu, Eric C; Kowalski, Regis P; Romanowski, Eric G; Mah, Francis S; Gordon, Y Jerold; Shanks, Robert M Q

2010-12-01

The aim of this study was to analyze the effect of azithromycin (AZM) 1% ophthalmic solution in DuraSite® (AzaSite®) on biofilm formation by Staphylococcus aureus and coagulase-negative staphylococci in vitro. Susceptible and resistant clinical strains (n = 8) of S. aureus and coagulase-negative staphylococci were challenged with serial dilutions of AzaSite® and its components: AZM, benzalkonium chloride (BAK), and the DuraSite drug delivery vehicle. After 20 h of incubation, bacterial growth was quantified using a spectrophotometer (A = 600 nm). Plates were stained with crystal violet and biofilm formation was quantified spectrophotometrically at A = 590 nm. AzaSite® and AZM inhibited bacterial growth (P reduction in biofilm formation (P reduction in biofilm formation at concentrations from 1.25 to 10 mg/mL in most strains. DuraSite® inhibited biofilm formation at concentrations between 10% and 2.5% in all studied strains (P < 0.05), without affecting bacterial growth. BAK inhibited bacterial growth and biofilm formation in all strains between concentrations of 0.042 and 0.375 mg/mL (P < 0.05). AzaSite®, AZM, or BAK prevented biofilm formation by inhibiting growth of AZM-susceptible strains. AzaSite®, AZM, and DuraSite® also reduced biofilm formation at subinhibitory concentrations for growth. Our data indicate that AZM has a moderate inhibitory effect on biofilm formation, whereas DuraSite® appears to play a greater role in the inhibition of staphylococcal biofilm formation by AzaSite®.

Evaluation of Candida Albicans Biofilm Formation on Various Dental ...

African Journals Online (AJOL)

2016-06-24

Jun 24, 2016 ... This study compared the susceptibility of six dental restorative materials to Candida albicans adhesion. ... found for the composite and the compomer samples. ..... Candida colonization on acrylic resins and denture liners:.

Enzymatic degradation of in vitro Staphylococcus aureus biofilms supplemented with human plasma

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Watters CM

2016-04-01

Full Text Available Chase M Watters,1,2 Tarea Burton,1 Dickson K Kirui,1 Nancy J Millenbaugh1 1Maxillofacial Injury and Disease Department, Naval Medical Research Unit San Antonio, Joint Base San Antonio-Fort Sam Houston, TX, USA; 2Wound Infections Department, Naval Medical Research Center, Silver Spring, MD, USA Abstract: Enzymatic debridement is a therapeutic strategy used clinically to remove necrotic tissue from wounds. Some of the enzymes utilized for debridement have been tested against bacterial pathogens, but the effectiveness of these agents in dispersing clinically relevant biofilms has not been fully characterized. Here, we developed an in vitro Staphylococcus aureus biofilm model that mimics wound-like conditions and employed this model to investigate the antibiofilm activity of four enzymatic compounds. Human plasma at concentrations of 0%–50% was supplemented into growth media and used to evaluate biofilm biomass accumulation over 24 hours and 48 hours in one methicillin-sensitive and five methicillin-resistant strains of S. aureus. Supplementation of media with 10% human plasma resulted in the most robust biofilms in all six strains. The enzymes α-amylase, bromelain, lysostaphin, and papain were then tested against S. aureus biofilms cultured in 10% human plasma. Quantification of biofilms after 2 hours and 24 hours of treatment using the crystal violet assay revealed that lysostaphin decreased biomass by up to 76%, whereas a-amylase, bromelain, and papain reduced biomass by up to 97%, 98%, and 98%, respectively. Scanning electron microscopy confirmed that the dispersal agents detached the biofilm exopolysaccharide matrix and bacteria from the growth surface. Lysostaphin caused less visible dispersal of the biofilms, but unlike the other enzymes, induced morphological changes indicative of bacterial cell damage. Overall, our results indicate that use of enzymes may be an effective means of eradicating biofilms and a promising strategy to improve

Histone deacetylases and their inhibition in Candida species

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Cecile Garnaud

2016-08-01

Full Text Available Fungi are generally benign members of the human mucosal flora or live as saprophytes in the environment. However, they can become pathogenic, leading to invasive and life threatening infections in vulnerable patients. These invasive fungal infections are regarded as a major public health problem on a similar scale to tuberculosis or malaria. Current treatment for these infections is based on only four available drug classes. This limited therapeutic arsenal and the emergence of drug-resistant strains are a matter of concern due to the growing number of patients to be treated, and new therapeutic strategies are urgently needed. Adaptation of fungi to drug pressure involves transcriptional regulation, in which chromatin dynamics and histone modifications play a major role. Histone deacetylases (HDACs remove acetyl groups from histones and actively participate in controlling stress responses. HDAC inhibition has been shown to limit fungal development, virulence, biofilm formation and dissemination in the infected host, while also improving the efficacy of existing antifungal drugs towards Candida spp. In this article, we review the functional roles of HDACs and the biological effects of HDAC inhibitors on Candida spp., highlighting the correlations between their pathogenic effects in vitro and in vivo. We focus on how HDAC inhibitors could be used to treat invasive candidiasis while also reviewing recent developments in their clinical evaluation.

In Vitro Effects of Sports and Energy Drinks on Streptococcus mutans Biofilm Formation and Metabolic Activity.

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Vinson, LaQuia A; Goodlett, Amy K; Huang, Ruijie; Eckert, George J; Gregory, Richard L

2017-09-15

Sports and energy drinks are being increasingly consumed and contain large amounts of sugars, which are known to increase Streptococcus mutans biofilm formation and metabolic activity. The purpose of this in vitro study was to investigate the effects of sports and energy drinks on S. mutans biofilm formation and metabolic activity. S. mutans UA159 was cultured with and without a dilution (1:3 ratio) of a variety of sports and energy drinks in bacterial media for 24 hours. The biofilm was washed, fixed, and stained. Biofilm growth was evaluated by reading absorbance of the crystal violet. Biofilm metabolic activity was measured by the biofilm-reducing XTT to a water-soluble orange compound. Gatorade Protein Recovery Shake and Starbucks Doubleshot Espresso Energy were found to significantly increase biofilm (30-fold and 22-fold, respectively) and metabolic activity (2-fold and 3-fold, respectively). However, most of the remaining drinks significantly inhibited biofilm growth and metabolic activity. Several sports and energy drinks, with sugars or sugar substitutes as their main ingredients inhibited S. mutans biofilm formation. Among the drinks evaluated, Gatorade Protein Recovery Chocolate Shake and Starbucks Doubleshot Energy appear to have cariogenic potential since they increased the biofilm formation and metabolic activity of S. mutans.

In vitro Susceptibility of Fluconazole, Clotrimazole and Toucrium Polium Smoke Product on Candida Isolates of Vaginal Candidiasis

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B Bonyadpour

2009-07-01

Full Text Available ABSTRACT: Introduction & Objective: It has been estimated that up to 75% of women in their child-bearing age have been affected by vulvovaginal candidiasis at least once in their life time. Almost 45% of women experience this infection two or more times. The antifungal azole group, in topical and oral forms, is the common way of therapy. Herbal products are often used for vulvovaginal therapy. Nowadays, Toucrium polium (TP products are being used as traditional medicine to reduce signs of Candida vaginitis. There is no study regarding to antifungal activity of TP smoke product in Iran. The aim of this study was to evaluate the In vitro activity of TP smoke product against Candida, isolated from women with Candida vaginitis, compared with antifungal drugs which are ordinary used to cure Candida vaginitis. Materials & Methods: The present study was conducted at the University of Medical Sciences of Shiraz in 1387 (2008. During seven months, samples were taken from 450 patients suffering from urogenital infections and 105 Candida vaginitis were detected. Germ tube test was used for identification of fungal species. TP smoke product was prepared in suitable potency. Antifungal activity of fluconazole, clotrimazole and TP product were evaluated by disk diffusion method. Sterile blank disks were loaded by TP smoke product in potency of 10-240 microliter/disk. Inhibition zone around the disks were measured and compared with each other. Results: 105 Candida species were isolated from the patients. Candida species were identified by germ tube test as Candida albicans 74 (70.5% and Candida non-albicans 31(29.5%.The mean of inhibition zone around the clotrimazole disks was 22±5.39 along with one case of resistance. Forty seven species had resistance to fluconazole while 94% and 55.2.% of all samples were sensitive to clotrimazole and fluconazole respectively. All of the clinical isolates and standard Candida species were sensitive to TP smoke product

Determination of virulence factors and biofilm formation among isolates of vulvovaginal candidiasis

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Tapan Majumdar

2016-01-01

Full Text Available Context: Under morphogenesis-inducing conditions, Candida spp. begins to undergo yeast-to-hypha switch. This shift from commensal to pathogenic state is dependent on several virulence factors. Aim: To find out whether the isolated Candida spp. were pathogens causing vulvovaginal candidiasis or mere bystanders. Settings and Design: Cross-sectional observational study conducted on 275 symptomatic hospital patients in Tripura between August 2012 and April 2015. Subjects and Methods: Discharge was collected from patients and identified by Grams staining and wet mount test. Culturing was done in Sabouraud dextrose agar followed by speciation. To test for virulence factors, assays for adherence, plasma coagulase, phospholipase, lipase, protease, hemolysin, and biofilm formation were carried out. Statistical Analysis Used: Significance between two groups was compared using one-way analysis of variance along with Tukey test, and Chi-square 2 × 2 contingency table at 95% confidence interval. Results: Fifty-six Candida spp. could be isolated in the study which was used for further virulence tests. One hundred percent of isolates expressed adherence. Among other virulence factors, maximum virulence 25 (45% was shown through protease production. Hemolysin production and biofilm formation were the second most 22 (39% expressed virulence factors. In a comparison of virulence factors between biofilm-forming isolates and planktonic cells, significant difference was seen for plasma coagulase and hemolysin production. Conclusions: All the isolates expressed one or more virulence factors. Adherence was expressed in all isolates but highest number was observed for Candida albicans. Furthermore, C. albicans strain number was highest for protease, hemolysin and coagulase expression and biofilm formation. Candida krusei isolates were the least in number for expressing any of the virulence factors. Significantly higher number of biofilm forming isolates produced

Limonene inhibits Candida albicans growth by inducing apoptosis.

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Thakre, Archana; Zore, Gajanan; Kodgire, Santosh; Kazi, Rubina; Mulange, Shradha; Patil, Rajendra; Shelar, Amruta; Santhakumari, Bayitigeri; Kulkarni, Mahesh; Kharat, Kiran; Karuppayil, Sankunny Mohan

2018-07-01

Anti-Candida potential of limonene was evaluated against planktonic growth, biofilm (adhesion, development and maturation) and morphogenesis of Candida albicans in this study. Limonene is a major constituent of citrus oil and most frequently used terpene in food and beverage industry due to its pleasant fragrance, nontoxic, and is generally recognized as safe (GRAS) flavoring agent as well as treatment option in many gastrointestinal diseases.Limonene exhibited excellent anti-Candida activity and was equally effective against planktonic growth of C. albicans isolates differentially susceptible to FLC (N = 35). Limonene inhibited morphogenesis significantly at low concentration. However, it showed stage dependent activity against biofilm formation, that is, it was more effective against adhesion followed by development and maturation. Limonene also exhibited excellent synergy with FLC against planktonic and biofilm growth. SWATH-MS analysis led to identification of limonene responsive proteins that provided molecular insight of its anti-Candida activity. Proteomic analysis revealed upregulation of proteins involved in cell wall glucan synthesis (Kre6); oxidative stress (Rhr2, Adh7 and Ebp1); DNA damage stress (Mbf1 and Npl3); nucleolar stress (Rpl11, Rpl7, Rpl29, Rpl15) and down regulation of cytoskeleton organization (Crn1, Pin3, Cct8, Rbl2), and so forth, in response to limonene. Limonene mediated down regulation of Tps3 indicates activation of caspase (CaMca1) and induction of apoptosis in C. albicans. These results suggest that limonene inhibits C. albicans growth by cell wall/membrane damage induced oxidative stress that leads to DNA damage resulting into modulation of cell cycle and induction of apoptosis through nucleolar stress and metacaspase dependent pathway.

Plants’ Natural Products as Alternative Promising Anti-Candida Drugs

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Soliman, Sameh; Alnajdy, Dina; El-Keblawy, Ali A.; Mosa, Kareem A.; Khoder, Ghalia; Noreddin, Ayman M.

2017-01-01

Candida is a serious life-threatening pathogen, particularly with immunocompromised patients. Candida infections are considered as a major cause of morbidity and mortality in a broad range of immunocompromised patients. Candida infections are common in hospitalized patients and elderly people. The difficulty to eradicate Candida infections is owing to its unique switch between yeast and hyphae forms and more likely to biofilm formations that render resistance to antifungal therapy. Plants are known sources of natural medicines. Several plants show significant anti-Candida activities and some of them have lower minimum inhibitory concentration, making them promising candidates for anti-Candida therapy. However, none of these plant products is marketed for anti-Candida therapy because of lack of sufficient information about their efficacy, toxicity, and kinetics. This review revises major plants that have been tested for anti-Candida activities with recommendations for further use of some of these plants for more investigation and in vivo testing including the use of nanostructure lipid system. PMID:28989245

In Vitro Activity of Tea Tree Oil Vaginal Suppositories against Candida spp. and Probiotic Vaginal Microbiota.

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Di Vito, Maura; Mattarelli, Paola; Modesto, Monica; Girolamo, Antonietta; Ballardini, Milva; Tamburro, Annunziata; Meledandri, Marcello; Mondello, Francesca

2015-10-01

The aim of this work is to evaluate the in vitro microbicidal activity of vaginal suppositories (VS) containing tea tree oil (TTO-VS) towards Candida spp. and vaginal probiotics. A total of 20 Candida spp. strains, taken from patients with vaginitis and from an established type collection, including reference strains, were analysed by using the CLSI microdilution method. To study the action of VS towards the beneficial vaginal microbiota, the sensitivity of Bifidobacterium animalis subsp. lactis (DSM 10140) and Lactobacillus spp. (Lactobacillus casei R-215 and Lactobacillus acidophilus R-52) was tested. Both TTO-VS and TTO showed fungicidal activity against all strains of Candida spp. whereas placebo-VS or the Aloe gel used as controls were ineffective. The study of fractional fungicidal concentrations (FFC) showed synergistic interaction with the association between Amphotericin B and TTO (0.25 to 0.08 µg/ml, respectively) against Candida albicans. Instead, the probiotics were only affected by TTO concentration ≥ 4% v/v, while, at concentrations vaginal microbiota. In vivo studies are needed to confirm the efficacy to prevent acute or recurrent vaginal candidiasis. Copyright © 2015 John Wiley & Sons, Ltd.

Moonlight-like proteins of the cell wall protect sessile cells of Candida from oxidative stress.

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Serrano-Fujarte, Isela; López-Romero, Everardo; Cuéllar-Cruz, Mayra

2016-01-01

Biofilms of Candida species are associated with high morbidity and hospital mortality. Candida forms biofilms by adhering to human host epithelium through cell wall proteins (CWP) and simultaneously neutralizing the reactive oxygen species (ROS) produced during the respiratory burst by phagocytic cells. The purpose of this paper is to identify the CWP of Candida albicans, Candida glabrata, Candida krusei and Candida parapsilosis expressed after exposure to different concentrations of H2O2 using a proteomic approach. CWP obtained from sessile cells, both treated and untreated with the oxidizing agent, were resolved by one and two-dimensional (2D-PAGE) gels and identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. Some of these proteins were identified and found to correspond to moonlighting CWP such as: (i) glycolytic enzymes, (ii) heat shock, (iii) OSR proteins, (iv) general metabolic enzymes and (v) highly conserved proteins, which are up- or down-regulated in the presence or absence of ROS. We also found that the expression of these CWP is different for each Candida species. Moreover, RT-PCR assays allowed us to demonstrate that transcription of the gene coding for Eno1, one of the moonlight-like CWP identified in response to the oxidant agent, is differentially regulated. To our knowledge this is the first demonstration that, in response to oxidative stress, each species of Candida, differentially regulates the expression of moonlighting CWP, which may protect the organism from the ROS generated during phagocytosis. Presumptively, these proteins allow the pathogen to adhere and form a biofilm, and eventually cause invasive candidiasis in the human host. We propose that, in addition to the antioxidant mechanisms present in Candida, the moonlighting CWP also confer protection to these pathogens from oxidative stress. Copyright © 2015 Elsevier Ltd. All rights reserved.

Comparison of the adhesion ability of Candida albicans strains to ...

African Journals Online (AJOL)

The purpose of the present study is to investigate the ability of oral Candida albicans strains to adhere to Caco-2 and Hep-2 epithelial cells, to produce slime using Congo red and Safranin methods and to form a biofilm on polymethylmethacrylate. A total of 20 C. albicans strains were tested in the present work. The biofilm ...

Haemophilus parainfluenzae Strain ATCC 33392 Forms Biofilms In Vitro and during Experimental Otitis Media Infections.

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Pang, Bing; Swords, W Edward

2017-09-01

Haemophilus parainfluenzae is a nutritionally fastidious, Gram-negative bacterium with an oropharyngeal/nasopharyngeal carriage niche that is associated with a range of opportunistic infections, including infectious endocarditis and otitis media (OM). These infections are often chronic/recurrent in nature and typically involve bacterial persistence within biofilm communities that are highly resistant to host clearance. This study addresses the primary hypothesis that H. parainfluenzae forms biofilm communities that are important determinants of persistence in vivo The results from in vitro biofilm studies confirmed that H. parainfluenzae formed biofilm communities within which the polymeric matrix was mainly composed of extracellular DNA and proteins. Using a chinchilla OM infection model, we demonstrated that H. parainfluenzae formed surface-associated biofilm communities containing bacterial and host components that included neutrophil extracellular trap (NET) structures and that the bacteria mainly persisted in these biofilm communities. We also used this model to examine the possible interaction between H. parainfluenzae and its close relative Haemophilus influenzae , which is also commonly carried within the same host environments and can cause OM. The results showed that coinfection with H. influenzae promoted clearance of H. parainfluenzae from biofilm communities during OM infection. The underlying mechanisms for bacterial persistence and biofilm formation by H. parainfluenzae and knowledge about the survival defects of H. parainfluenzae during coinfection with H. influenzae are topics for future work. Copyright © 2017 American Society for Microbiology.

RT-PCR detection of Candida albicans ALS gene expression in the reconstituted human epithelium (RHE) model of oral candidiasis and in model biofilms.

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Green, Clayton B; Cheng, Georgina; Chandra, Jyotsna; Mukherjee, Pranab; Ghannoum, Mahmoud A; Hoyer, Lois L

2004-02-01

An RT-PCR assay was developed to analyse expression patterns of genes in the Candida albicans ALS (agglutinin-like sequence) family. Inoculation of a reconstituted human buccal epithelium (RHE) model of mucocutaneous candidiasis with strain SC5314 showed destruction of the epithelial layer by C. albicans and also formation of an upper fungal layer that had characteristics similar to a biofilm. RT-PCR analysis of total RNA samples extracted from C. albicans-inoculated buccal RHE showed that ALS1, ALS2, ALS3, ALS4, ALS5 and ALS9 were consistently detected over time as destruction of the RHE progressed. Detection of transcripts from ALS7, and particularly from ALS6, was more sporadic, but not associated with a strictly temporal pattern. The expression pattern of ALS genes in C. albicans cultures used to inoculate the RHE was similar to that observed in the RHE model, suggesting that contact of C. albicans with buccal RHE does little to alter ALS gene expression. RT-PCR analysis of RNA samples extracted from model denture and catheter biofilms showed similar gene expression patterns to the buccal RHE specimens. Results from the RT-PCR analysis of biofilm RNA specimens were consistent between various C. albicans strains during biofilm development and were comparable to gene expression patterns in planktonic cells. The RT-PCR assay described here will be useful for analysis of human clinical specimens and samples from other disease models. The method will provide further insight into the role of ALS genes and their encoded proteins in the diverse interactions between C. albicans and its host.

Evaluation of efficacy of commercial denture cleansing agents to reduce the fungal biofilm activity from heat polymerized denture acrylic resin: An in vitro study

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Mithilesh M Dhamande

2012-01-01

Full Text Available Aims: To compare and evaluate Candida removing effects of three most commonly available varieties of commercial denture cleansers from heat polymerized acrylic resins. To compare and evaluate Candida lytic effects of denture cleansers. To assess the effect of time on ability of denture cleansers in reducing Candidal biofilm. Materials and Methods: A specially designed metal mold was fabricated to obtain wax plates of uniform dimensions which were used to fabricate heat cure acrylic resin plates. A square-shaped window of dimension 15 mm and thickness of 1.5 mm was provided in metal mould to simulate thickness of denture base. All samples used in this study were prepared using this mould. Candida albicans colonies were then cultured on this acrylic resin plates by colonization assay. Yeast removing test for samples was performed using microscope and yeast lytic test was performed using photo colorimeter. Results: Denture cleanser D2 showed the highest Candida removing activity when compared with cleansers D1, D3, and control solution. Denture cleansers D2 showed increased yeast lytic ability when compared with denture cleansers D1, D3, and control solution. More time span shared a definite influence on yeast lytic ability of denture cleansers. Conclusions: The effect of cleansing agents on removal of colonized yeasts particularly fungal biofilm from acrylic resins was assessed for clinical implications. The observation indicated superior performance of cleanser D2 when compared with D1 and D3 even though they all belong to same chemical group of alkaline peroxide. The increased effectiveness may be due to presence of sodium lauryl sulphate in formula of D2.

Effects of Low-Level Laser Irradiation on the Pathogenicity of Candida albicans: In Vitro and in Vivo Study

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Seyedmousavi Tasieh, S.; Hashemi, S.J.; Rezaie, S.; Fateh, M.; Djavid, G.E.; Zibafar, E.; Morsali, F.; Zand, N.; Alinaghizadeh, M.; Ataie-Fashtami, L.

2014-01-01

Abstract Objective: The purpose of this study was to evaluate the effects of low-level laser irradiation (LLLI) on the in vitro growth characteristics and in vivo pathogenicity of Candida albicans in a murine model in the absence of a photosensitizer. Background data: C. albicans is an opportunistic

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...

Identification of Genes Upregulated by the Transcription Factor Bcr1 That Are Involved in Impermeability, Impenetrability, and Drug Resistance of Candida albicans a/α Biofilms

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Srikantha, Thyagarajan; Daniels, Karla J.; Pujol, Claude; Kim, Elena

2013-01-01

Candida albicans forms two types of biofilm, depending upon the configuration of the mating type locus. Although architecturally similar, a/α biofilms are impermeable, impenetrable, and drug resistant, whereas a/a and α/α biofilms lack these traits. The difference appears to be the result of an alternative matrix. Overexpression in a/a cells of BCR1, a master regulator of the a/α matrix, conferred impermeability, impenetrability, and drug resistance to a/a biofilms. Deletion of BCR1 in a/α cells resulted in the loss of these a/α-specific biofilm traits. Using BCR1 overexpression in a/a cells, we screened 107 genes of interest and identified 8 that were upregulated by Bcr1. When each was overexpressed in a/a biofilms, the three a/α traits were partially conferred, and when each was deleted in a/α cells, the traits were partially lost. Five of the eight genes have been implicated in iron homeostasis, and six encode proteins that are either in the wall or plasma membrane or secreted. All six possess sites for O-linked and N-linked glycosylation that, like glycosylphosphatidylinositol (GPI) anchors, can cross-link to the wall and matrix, suggesting that they may exert a structural role in conferring impermeability, impenetrability, and drug resistance, in addition to their physiological functions. The fact that in a screen of 107 genes, all 8 of the Bcr1-upregulated genes identified play a role in impermeability, impenetrability, and drug resistance suggests that the formation of the a/α matrix is highly complex and involves a larger number of genes than the initial ones identified here. PMID:23563485

Evaluation of the in vitro activity of isavuconazole and comparator voriconazole against 2635 contemporary clinical Candida and Aspergillus isolates

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Astvad, K. M.T.; Hare, R. K.; Arendrup, M. C.

2017-01-01

Objective The in vitro activity of isavuconazole was determined for 1677 Candida and 958 Aspergillus isolates from 2012 to 2014 with voriconazole as comparator. Methods Aspergillus isolates were screened for resistance using azole-agar. Aspergillus isolates that screened positive and all Candida...

In vitro differential activity of phospholipases and acid proteinases of clinical isolates of Candida

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Aurean D'Eça Júnior

2011-06-01

Full Text Available INTRODUCTION: Candida yeasts are commensals; however, if the balance of normal flora is disrupted or the immune defenses are compromised, Candida species can cause disease manifestations. Several attributes contribute to the virulence and pathogenicity of Candida, including the production of extracellular hydrolytic enzymes, particularly phospholipase and proteinase. This study aimed to investigate the in vitro activity of phospholipases and acid proteinases in clinical isolates of Candida spp. METHODS: Eighty-two isolates from hospitalized patients collected from various sites of origin were analyzed. Phospholipase production was performed in egg yolk medium and the production of proteinase was verified in a medium containing bovine serum albumin. The study was performed in triplicate. RESULTS: Fifty-six (68.3% of isolates tested were phospholipase positive and 16 (44.4% were positive for proteinase activity. C. tropicalis was the species with the highest number of positive isolates for phospholipase (91.7%. Statistically significant differences were observed in relation to production of phospholipases among species (p<0,0001 and among the strains from different sites of origin (p=0.014. Regarding the production of acid protease, the isolates of C. parapsilosis tested presented a larger number of producers (69.2%. Among the species analyzed, the percentage of protease producing isolates did not differ statistically (χ2=1.9 p=0.5901 (χ2=1.9 p=0.5901. CONCLUSIONS: The majority of C. non-albicans and all C. albicans isolates were great producers of hydrolytic enzymes and, consequently, might be able to cause infection under favorable conditions.

In Vitro Antimicrobial Susceptibility of Pathogenic Leptospira Biofilm.

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Vinod Kumar, Kirubakaran; Lall, Chandan; Raj, Ratchagadasse Vimal; Vedhagiri, Kumaresan; Sunish, Ittoop Pulikkottil; Vijayachari, Paluru

2016-10-01

Pathogenic Leptospira spp. are the causative agent of leptospirosis. Biofilm formation in leptospires is a new area of study, and its role in pathogenesis is not fully explored. As in other biofilm-forming bacteria, Leptospira biofilm may play a significant role in antibiotic resistance. In this study, the antimicrobial susceptibility of Leptospira biofilm was investigated by 96-well plate assay using Alamar Blue. Leptospira biofilm showed five to sixfold increase in resistance in all the strains used. The range of minimal bactericidal concentrations for penicillin G, ampicillin, tetracycline, and doxycycline was 1,600 U/ml, 800-1,600 μg/ml, 800-1,600 μg/ml, and 800-1,600 μg/ml, respectively. In agar substrate, the biofilm showed six- to sevenfold increase in resistance to antibiotics compared to planktonic cell. The present study emphasizes the importance of biofilm formation and its antibiotic susceptibility patterns. This could pave the way for devising appropriate strategy to prevent the occurrence of potential chronic leptospirosis in endemic areas and also during an outbreak situation.

Cariogenicity of soluble starch in oral in vitro biofilm and experimental rat caries studies: a comparison.

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Thurnheer, T; Giertsen, E; Gmür, R; Guggenheim, B

2008-09-01

Common belief suggests that starch is less cariogenic than sugar; however, the related literature is quite controversial. We aimed to compare cariogenic and microbiological effects of soluble starch in both a standard animal model and an oral biofilm system, and to assess the possible substitution of the animal model. Six-species biofilms were grown anaerobically on enamel discs in saliva and medium with glucose/sucrose, starch (average molecular weight of 5000, average polymerization grade of 31), or mixtures thereof. After 64.5 h of biofilm formation, the microbiota were quantitated by cultivation and demineralization was measured by quantitative light-induced fluorescence. To assess caries incidence in rats, the same microbiota as in the biofilm experiments were applied. The animals were fed diets containing either glucose, glucose/sucrose, glucose/sucrose/starch or starch alone. Results with both models show that demineralization was significantly smaller with starch than sucrose. The data demonstrate that soluble starch is substantially less cariogenic than glucose/sucrose. By leading to the same scientific evidence as its in vivo counterpart, the described in vitro biofilm system provides an interesting and valuable tool in the quest to reduce experimentation with animals.

Biofilm formation on stainless steel and gold wires for bonded retainers in vitro and in vivo and their susceptibility to oral antimicrobials

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Jongsma, Marije A.; Pelser, Floris D. H.; van der Mei, Henny C.; Atema-Smit, Jelly; van de Belt-Gritter, Betsy; Busscher, Henk J.; Ren, Yijin

OBJECTIVE: Bonded retainers are used in orthodontics to maintain treatment result. Retention wires are prone to biofilm formation and cause gingival recession, bleeding on probing and increased pocket depths near bonded retainers. In this study, we compare in vitro and in vivo biofilm formation on

Photodynamic therapy with water-soluble phtalocyanines against bacterial biofilms in teeth root canals

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Gergova, Raina; Georgieva, Tzvetelina; Angelov, Ivan; Mantareva, Vanya; Valkanov, Serjoga; Mitov, Ivan; Dimitrov, Slavcho

2012-06-01

The study presents the PDT with metal phthalocyanines on biofilms grown in root canals of ten representatives of the Gram-positive and the Gram-negative bacterial species and a fungus Candida albicans which cause aqute teeth infections in root canals.. The extracted human single-root teeth infected for 48 h with microorganisms in conditions to form biofilms of the above pathogens were PDT treated. The stage of biofilm formation and PDT effect of the samples of the teeth were determined by the scaning electron microscopy and with standard microbial tests. The PDT treating procedure included 10 min incubation with the respected phthalocyanine and irradiated with 660 nm Diode laser for 10 min. The most strongly antibacterial activity was achieved with zinc(II) phthalocyanine (ZnPc) against Enterococcus faecalis, Staphylococcus aureus and Moraxella catarrhalis. The other Gram-negative bacteria and Candida albicans were 10-100 times more resistant than the Gram-positive species. The Gram-negative Moraxella catarrhalis and Acinetobacter baumannii were more sensitive than the enterobacteria, but eradication of Pseudomonas aeruginosa in biofilm was insignificant. The influence of the stage of biofilm formation and the initial conditions (bacterial density, photosensitizer concentration and energy fluence of radiation) to the obtained level of inactivation of biofilms was investigated. The PDT with ZnPc photosensitizers show a powerful antimicrobial activity against the most frequent pathogens in endodontic infections and this method for inactivation of pathogens may be used with sucsses for treatment of the bacterial biofilms in the root canals.

Structural Properties and Antifungal Activity against Candida albicans Biofilm of Different Composite Layers Based on Ag/Zn Doped Hydroxyapatite-Polydimethylsiloxanes

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Andreea Groza

2016-04-01

Full Text Available Modern medicine is still struggling to find new and more effective methods for fighting off viruses, bacteria and fungi. Among the most dangerous and at times life-threatening fungi is Candida albicans. Our work is focused on surface and structural characterization of hydroxyapatite, silver doped hydroxyapatite and zinc doped hydroxyapatite deposited on a titanium substrate previously coated with polydimethylsiloxane (HAp-PDMS, Ag:HAp-PDMS, Zn:HAp-PDMS by different techniques: Scanning Electron Microscopy (SEM, Glow Discharge Optical Emission Spectroscopy (GDOES and Fourier Transform Infrared Spectroscopy (FTIR. The morphological studies revealed that the use of the PDMS polymer as an interlayer improves the quality of the coatings. The structural characterizations of the thin films revealed the basic constituents of both apatitic and PDMS structure. In addition, the GD depth profiles indicated the formation of a composite material as well as the successful embedding of the HAp, Zn:HAp and Ag:HAp into the polymer. On the other hand, in vitro evaluation of the antifungal properties of Ag:HAp-PDMS and Zn:HAp-PDMS demonstrated the fungicidal effects of Ag:HAp-PDMS and the potential antifungal effect of Zn:HAp-PDMS composite layers against C. albicans biofilm. The results acquired in this research complete previous research on the potential use of new complex materials produced by nanotechnology in biomedicine.

Molecular identification and in-vitro antifungal susceptibility testing of Candida species isolated from patients with onychomycosis

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Keyvan Pakshir

2015-03-01

Full Text Available Background and Purpose: Candida species are the most opportunistic fungi affecting the nails and resulting in onychomycosis. In this study, we identified and evaluated in-vitro susceptibility of the recovered isolates against fluconazole (FLC, voriconazole (VRC, and clotrimazole (CLT using the Clinical and Laboratory Standards Institute (CLSI M27-A3 document. Materials and Methods: From patients with either clinically or mycologically proven onychomycosis, 97 isolates comprising of seven Candida species were isolated, which were identified by both conventional and molecular techniques such as polymerase chain reaction-restriction fragment length polymorphism. In addition, Candida dubliniensis was confirmed by restriction endonuclease analysis. Antifungal susceptibility of each isolate against the three azoles applied in this study was determined using the CLSI microdilution reference method M27-A3. Results: Candida parapsilosis (C. parapsilosis was the most frequently isolated species (n=44, followed by C. albicans (n=23, C. tropicalis (n=13, C. glabrata (n=7, C. krusei (n=6, C. guilliermondii (n=3, and C. dubliniensis (n=1. All the isolates were susceptible to CLT. VRC had lower minimum inhibitory concentration (MIC values for the isolates compared to FLC. Geometric mean MIC values of VRC, FLC, and CLT for C. parapsilosis isolates were 0.07 µg/ml, 0.8 µg/ml, and 0.35 µg/ml, respectively. Collectively, all species exhibited greater susceptibility to VRC in comparison to C. albicans (P≤0.001. Conclusion: This study showed that non-albicans Candida species were the most common etiologic agents of non-dermatophyte onychomycosis. The major antifungal agents used in clinics to empirically treat yeast onychomycosis are FLC and CLT. Our data suggested that CLT is a better choice for the treatment of Candida onychomycosis, especially in drug resistant cases.

Special Issue: Candida and Candidiasis

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Jeniel E. Nett

2018-06-01

Full Text Available This special issue highlights emerging topics related to Candida, the most prevalent fungal pathogen in the hospital setting. The advantages and limitations of new, non-culture based diagnostic techniques are discussed. The issue reviews mammalian and non-mammalian infection models. The manuscripts present updates on several molecular mechanisms of pathogenicity, including filamentation, biofilm formation, and phospholipid production.

Antibiofilm activity of Streptomyces toxytricini Fz94 against Candida albicans ATCC 10231

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Sheir DH

2017-06-01

Full Text Available Candida albicans is a significant cause of morbidity and mortality in immunocompromised patients worldwide. Biofilm formation by Candida species is a significant virulence factor for disease pathogenesis. Keeping in view the importance of Streptomyces' metabolites, the present study was initiated during the bioprospecting programme of Egyptian Streptomyces carried by the authors since 2013. Native Streptomyces isolates were recovered from soil samples collected from different governorates. Antifungal activity of forty isolates of Streptomyces were performed against planktonic (free cells of C. albicans ATCC 10231 and resistant clinical Candida isolates. Streptomyces isolates showed high inhibition activity against free cells of Candida were further assayed against biofilm of C. albicans reference strain. The most active Streptomyces sp. (no.6 was identified phenotypically, biochemically and by using 16S rRNA. The 16S rRNA sequences obtained were compared with those deposited in the GenBank Database and registered with accession number KM052378 as S. toxytricini Fz94. Screening of S. toxytricini Fz94 extract capability in prevention and destruction of C. albicans reference strain biolfilm was assessed by resazurin dye adopted technique. In the pre-exposure scheme, the lowest concentration of 5 gL-1 showed biofilm viability inhibition of 92% after 120 min, while Ketoconazole® gave 90 % inhibition at concentration of 2 gL-1. In post exposure, the concentration of S. toxytricini Fz94 extract 7gL-1 caused 82 % inhibition of biofilms viability after 120 min, while Ketoconazole did not show any destruction capability. The cytotoxicity of S. toxytricini Fz94 crude extract results showed that it was nontoxic at 10 gL-1. S. toxytricini Fz94 is maintained in the Fungarium of Arab Society for Fungal Conservation (ASFC with accession number FSCU-2017-1110.

In vitro characterization of biofilms formed by Kingella kingae.

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

2017-08-01

The Gram-negative bacterium Kingella kingae is part of the normal oropharyngeal mucosal flora of children 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. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Myroides odoratimimus Forms Structurally Complex and Inherently Antibiotic-Resistant Biofilm in a Wound-Like in vitro Model

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Arianna Pompilio

2017-12-01

Full Text Available Myroides odoratimimus is an aerobic, non-fermenting Gram-negative multidrug-resistant bacterium widely distributed in nature that rarely causes infections in immunocompromised patients. We recently described in a diabetic patient a case of recurrent calcaneal ulcer infection caused by a M. odoratimimus strain showing potential for biofilm formation. For the first time, we therefore evaluated the ability of M. odoratimimus to form biofilm under different pH values and glucose concentrations using an in vitro “skin-like” model, and its susceptibility to levofloxacin, meropenem, and tigecycline. The expression of some antibiotic-resistance related genes was also monitored by RT-PCR during planktonic-to-biofilm transition. Our results indicated that M. odoratimimus can produce relevant amounts of biofilm biomass, in a time-dependent manner, especially at acidic pH and regardless of glucose concentration tested. The comparative analysis of MIC and MBC values between planktonic and sessile cells showed that resistance to antibiotics increased during the planktonic-to-biofilm transition. Viable cell count indicated that none of the tested antibiotics were able to completely eradicate preformed biofilms, although meropenem and levofloxacin were the most active causing a significant, dose-independent, reduction of biofilm's viability, as also confirmed by microscopic analysis. RT-PCR showed that antibiotic-resistance related gyrA and acrB genes are over-expressed during the transition from planktonic to sessile (biofilm lifestyle. Overall, our findings showed that M. odoratimimus can form relevant amounts of inherently antibiotic-resistant biofilm under conditions relevant to wound site, therefore suggesting a role in the pathogenesis of chronic ulcer infections.

Use of chlorhexidine gel (0.2%) to control gingivitis and candida species colonization in human immunodeficiency virus-infected children: a pilot study.

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Machado, Fernanda Campos; de Souza, Ivete Pomarico Ribeiro; Portela, Maristela Barbosa; de Araújo Soares, Rosangela Maria; Freitas-Fernandes, Liana Bastos; Castro, Gloria Fernanda

2011-01-01

The purpose of this study was to evaluate chlorhexidine to control gingivitis and Candida species (spp.) in children infected with the human immunodeficiency virus (HIV) and their acceptance of the therapy. Twenty-six HIV+ children were selected, and oral exam-established biofilm, gingival indexes, and stimulated saliva were collected for Candida ssp. identification. The children brushed their teeth for 21 days with chlorhexidine gel (0.2%). Salivary samples, biofilm, and gingival indexes were collected after 21-days and again 35 days after ceasing gel use. The children answered a questionnaire about the therapy. All children tested positive for Candida and gingivitis. After 21 days, Candida counts and gingivitis decreased in 25 and 26 children, respectively. Mean reduction was approximately 68% for Candida spp. and 74% for gingivitis. Thirty-five days after discontinuing gel use, gingivitis and Candida spp. increased in 13 and 16 patients, respectively. Considering the Candida spp., the heavy growth was lower in the first re-evaluation. Candida albicans was the most frequent species. Approximately 85% did not experience inconvenience with the gel, and approximately 48% thought it was good for tooth-brushing. Chlorhexidine therapy may be an option to treat and pre- vent gingivitis and reduce yeast counts in children infected with HIV.

In vitro antifungal activity against Candida species of Sri Lankan orthodox black tea (Camellia sinensis L. belonging to different agro-climatic elevations

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Wanigasekara Daya Ratnasooriya

2017-02-01

Full Text Available Objective: To investigate the antifungal potential of different grades of Sri Lankan orthodox black tea [orange pekoe, broken orange pekoe fannings (BOPF and Dust No. 1] belonging to the three agro-climatic elevations (low, mid and high. Methods: Antifungal activity was assessed in vitro using methanolic extracts (300 µg/disc and agar disc diffusion bioassay technique against three Candida species, Candida albicans (C. albicans, Candida glabrata (C. glabrata, and Candida tropicalis. ketoconazole and itraconazole mixture was used as positive control (10 µg/disc and methanol was used as the negative control. The minimum inhibitory concentrations were also determined using standard protocols. Results: None of the extracts were effective against Candida tropicalis. Furthermore, orange pekoe grade tea belonging to all agro-climatic elevations did not induce any antifungal activity against C. albicans and C. glabrata as well. Conversely, Dust No. 1 belonging to all three agro-climatic elevations and low-grown BOPF showed moderate antifungal activity against C. albicans and C. glabrata. Interestingly, the severity of the antifungal effect varied with agroclimatic elevations. The minimum inhibitory concentrations ranged from 64.00–128.00 µg/mL against C. glabrata and 128.00-256.00 µg/mL against C. albicans. Conclusions: Sri Lankan Dust No. 1 and BOPF have marked antifungal activity in vitro and offer promise to be used as a supplementary beverage in prophylaxis and during drug treatment in candidiasis.

The in vitro effect of fluoridated milk in a bacterial biofilm--enamel model.

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Arnold, Wolfgang H; Forer, Stefan; Heesen, Joerg; Yudovich, Keren; Steinberg, Doron; Gaengler, Peter

2006-07-01

The purpose of this study was to investigate the effect of milk and fluoridated milk on bacterially induced caries-like lesions. Extracted impacted human molars were cut in half and covered with a varnish leaving a 4*4 mm window. The samples were coated with biofilm of S. sobrinus and were further divided into three experimental groups of S. sobrinus, S. sobrinus and milk and S. sobrinus and fluoridated milk. As negative controls served teeth incubated in saline. Of twenty tooth halves serial ground sections were cut through the lesions and investigated with polarization light microscopy (PLM) and scanning electron microscopy (SEM) and EDX element analysis. The PLM photographs were used for 3D reconstruction, volumetric assessment and determination of the extension of the lesion zones. Of eight tooth halves the biofilm on the enamel surface was studied with SEM and EDX element analysis. Volumetric assessment showed a statistically significant difference in the volume of the body of the lesion and the translucent zone between the milk group and fluoridated milk group. Quantitative element analysis demonstrated significant differences between sound enamel and the superficial layer in the fluoridated milk group. The biofilm on the enamel surface showed an increased Ca content in the milk group and fluoridated milk group. Milk as a common nutrient seems to play a complex role in in-vitro biofilm--enamel interactions stimulating bacterial demineralization on one hand, and, as effective fluoride carrier, inhibits caries-like demineralization.

The effect of denture adhesives on Candida albicans growth in vitro.

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Sampaio-Maia, Benedita; Figueiral, Maria Helena; Sousa-Rodrigues, Patricia; Fernandes, Maria Helena; Scully, Crispian

2012-06-01

Denture-wearing favours the growth of Candida. In view of the fact that many denture wearers regularly use adhesives to enhance denture retention, stability and function, the aim of this work was to study the effect of denture adhesives on Candida albicans growth in vitro. The denture adhesives tested were Corega(®) cream, Kukident(®) cream, Novafix(®) cream, Polident(®) cream, Protefix(®) cream, Steradent(®) cream, Aderyn(®) powder, Corega(®) ultra powder, Protefix(®) powder and Corega(®) strip. C. albicans growth curves were obtained in the presence or absence of a 1% solution of the denture adhesive diluted in Sabouraud broth. Macro- and microscopic morphological changes in C. albicans were analysed, as was microbial contamination of the denture adhesive. Most of the denture adhesives studied induced morphological changes in C. albicans cells and colonies, but only two had any significant inhibitory effect on yeast growth. Kukident(®) cream markedly inhibited C. albicans growth in a concentration-dependent way, reducing the growth rate by 95%, whereas Corega(®) cream also inhibited C. albicans growth but in a non-concentration-dependent way, reducing the growth rate by 37%. In addition, denture adhesives available as powders had detectable microbial contamination. Some commercially available denture adhesives showed microbial contamination and some had significant inhibitory effect on C. albicans growth. © 2011 The Gerodontology Society and John Wiley & Sons A/S.

In vitro synergism of simvastatin and fluconazole against Candida species

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Everardo Albuquerque Menezes

2012-08-01

Full Text Available Systemic fungal infections are responsible for high mortality rates. Several species of fungi may be involved, but Candida spp. is the most prevalent. Simvastatin is used to lower cholesterol and also exhibits antifungal action. The aim of this study was to evaluate the synergistic action of simvastatin with fluconazole against strains of Candida spp. Susceptibility testing was performed according to protocol M27-A3, by broth microdilution method and the synergistic effect of simvastatin and fluconazole was calculated based on FICI (Fractional Inhibitory Concentration Index. Eleven strains were evaluated, and simvastatin showed a synergistic effect with fluconazole against 10 (91% of the Candida spp. strains tested. Simvastatin may be a valuable drug in the treatment of systemic infections caused by Candida spp.

Biofilm Formation by Pseudomonas Species Onto Graphene Oxide-TiO2 Nanocomposite-Coated Catheters: In vitro Analysis

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Deb, Ananya; Vimala, R.

The present study focuses on the development of an in vitro model system for biofilm growth by Pseudomonas aerouginosa onto small discs of foley catheter. Catheter disc used for the study was coated with graphene oxide-titanium oxide composite (GO-TiO2) and titanium oxide (TiO2) and characterized through XRD, UV-visible spectroscopy. Morphological analysis was done by scanning electron microscopy (SEM). The biofilm formed on the catheter surface was quantified by crystal violet (CV) staining method and a colorimetric assay (MTT assay) which involves the reduction of tetrazolium salt. The catheter coated with GO-TiO2 showed reduced biofilm growth in comparison to the TiO2-coated and uncoated catheter, thus indicating that it could be successfully used in coating biomedical devices to prevent biofilm formation which is a major cause of nosocomial infection.

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

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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.

Microcalorimetric determination of the effects of amoxicillin, metronidazole, and their combination on in vitro biofilm.

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Astasov-Frauenhoffer, Monika; Braissant, Olivier; Hauser-Gerspach, Irmgard; Weiger, Roland; Walter, Clemens; Zitzmann, Nicola U; Waltimo, Tuomas

2014-02-01

The mechanism of action of adjuvant antibiotic therapy in the treatment of peri-implantitis is not well understood. The aim of this study is to investigate antibiotic susceptibility of an in vitro biofilm by isothermal microcalorimetry (IMC). Titanium disks containing a 72-hour three-species biofilm (Streptococcus sanguinis DSM20068, Fusobacterium nucleatum ATCC10953, and Porphyromonas gingivalis DSM20709) were placed in a series of IMC ampoules with nutrient agar supplemented with increasing concentrations of amoxicillin, metronidazole, or their combination and incubated anaerobically for 10 days. Lag time and maximum growth rate were determined from continuous heat-flow recordings of metabolic activity. To validate the IMC biofilm results, adherent S. sanguinis and P. gingivalis were incubated anaerobically in medium supplemented with antibiotics at 37°C for 24 hours, and their vitality was determined by live/dead staining, conventional culturing, and IMC. In all biofilm samples incubated with antibiotics, a prolonged lag phase was observed compared with controls (P live/dead staining and conventional culturing. IMC gives new evidence about antibiotic effects on oral biofilms and is more informative than conventional culture and live/dead assays. The combination of antibiotics was found to be more efficient than metronidazole alone; however, only minor differences in growth inhibition were detected compared with amoxicillin alone.

The Interface between Fungal Biofilms and Innate Immunity

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John F. Kernien

2018-01-01

Full Text Available Fungal biofilms are communities of adherent cells surrounded by an extracellular matrix. These biofilms are commonly found during infection caused by a variety of fungal pathogens. Clinically, biofilm infections can be extremely difficult to eradicate due to their resistance to antifungals and host defenses. Biofilm formation can protect fungal pathogens from many aspects of the innate immune system, including killing by neutrophils and monocytes. Altered immune recognition during this phase of growth is also evident by changes in the cytokine profiles of monocytes and macrophages exposed to biofilm. In this manuscript, we review the host response to fungal biofilms, focusing on how these structures are recognized by the innate immune system. Biofilms formed by Candida, Aspergillus, and Cryptococcus have received the most attention and are highlighted. We describe common themes involved in the resilience of fungal biofilms to host immunity and give examples of biofilm defenses that are pathogen-specific.

In vitro effects of glycyrrhetinic acid and hyaluronic acid on the growth of vulvovaginal Candida albicans and other yeasts

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Martina Stevan

2017-12-01

Full Text Available Aims. The present study aimed to test the in vitro activity against Candida albicans and non-albicans strains of 18-β glycyrrhetinic acid (18-β GA and hyaluronic acid (HA, both alone and in combination. This antimicrobial activity was assessed using the National Committee for Clinical Laboratory Standards (NCCLS method on Candida strains that were isolated from patients with recurrent vulvovaginal candidiasis (RVVC. Results. Our results demonstrate that the anti-Candida activity is independent from antifungal susceptibility level and the fact that the growth inhibition is stronger at acidic pH level makes the two drugs a promising biological alternative for the topical treatment of vulvovaginal candidiasis (VVC and RVVC. Conclusions. Furthermore, the reduction of both budding cells formation and germ tube elongation, on mammalian cell monolayers, may explain the observed growth inhibition and suggest a decreased virulence, respectively.

Characterization and evaluation of whey protein-based biofilms as substrates for in vitro cell cultures.

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Gilbert, Vanessa; Rouabhia, Mahmoud; Wang, Hongxum; Arnould, Anne-Lise; Remondetto, Gabriel; Subirade, Muriel

2005-12-01

Whey proteins-based biofilms were prepared using different plasticizers in order to obtain a biomaterial for the human keratinocytes and fibroblasts in vitro culture. The film properties were evaluated by Fourier Transform Infrared Spectroscopy (FTIR) technique and mechanical tests. A relationship was found between the decrease of intermolecular hydrogen bond strength and film mechanical behavior changes, expressed by a breaking stress and Young modulus values diminishing. These results allow stating that the film molecular configuration could induce dissimilarities in its mechanical properties. The films toxicity was assessed by evaluating the cutaneous cells adherence, growth, proliferation and structural stratification. Microscopic observation demonstrated that both keratinocytes and fibroblasts adhered to the biofilms. The trypan blue exclusion test showed that keratinocytes grew at a significantly high rate on all the biofilms. Structural analysis demonstrated that keratinocytes stratified when cultured on the whey protein-based biofilms and gave rise to multi-layered epidermal structures. The most organized epidermis was obtained with whey protein isolate/DEG biofilm. This structure had a well-organized basal layer under supra-basal and corneous layers. This study demonstrated that whey proteins, an inexpensive renewable resource which can be obtained readily, were non-toxic to cutaneous cells and thus they could be useful substrates for a variety of biomedical applications, including tissue engineering.

Reliability of Haemophilus influenzae biofilm measurement via static method, and determinants of in vitro biofilm production.

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Obaid, Najla A; Tristram, Stephen; Narkowicz, Christian K; Jacobson, Glenn A

2016-12-01

Information is lacking regarding the precision of microtitre plate (MTP) assays used to measure biofilm. This study investigated the precision of an MTP assay to measure biofilm production by nontypeable Haemophilus influenzae (NTHi) and the effects of frozen storage and inoculation technique on biofilm production. The density of bacterial final growth was determined by absorbance after 18-20 h incubation, and biofilm production was then measured by absorbance after crystal violet staining. Biofilm formation was categorised as high and low for each strain. For the high biofilm producing strains of NTHi, interday reproducibility of NTHi biofilm formation measured using the MTP assay was excellent and met the acceptance criteria, but higher variability was observed in low biofilm producers. Method of inoculum preparation was a determinant of biofilm formation with inoculum prepared directly from solid media showing increased biofilm production for at least one of the high producing strains. In general, storage of NTHi cultures at -80 °C for up to 48 weeks did not have any major effect on their ability to produce biofilm.

Quantitative real-time PCR combined with propidium monoazide for the selective quantification of viable periodontal pathogens in an in vitro subgingival biofilm model.

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Sánchez, M C; Marín, M J; Figuero, E; Llama-Palacios, A; León, R; Blanc, V; Herrera, D; Sanz, M

2014-02-01

Differentiation of live and dead cells is an important challenge when using molecular diagnosis for microbial identification. This is particularly relevant when bacteria have been exposed to antimicrobial agents. The objective of this study was to test a method using quantitative real-time polymerase chain reaction (qPCR) combined with propidium monoazide (PMA), developed for the selective quantification of viable P. gingivalis, A. actinomycetemcomitans, F. nucleatum and total bacteria in an in vitro biofilm model after antimicrobial treatment. PMA-qPCR method was tested in an in vitro biofilm model, using isopropyl alcohol as the antimicrobial agent. Matured biofilms were exposed for 1, 5, 10 and 30 min to isopropyl alcohol by immersion. Biofilms were disrupted and PMA added (final concentration of 100 μm). After DNA isolation, qPCR was carried out using specific primers and probes for the target bacteria. The differentiation of live and dead cells was tested by analysis of variance. When PMA was used in the presence of viable target bacterial cells, no statistically significant inhibition of qPCR amplification was detected (p > 0.05 in all cases). Conversely, after immersion in isopropyl alcohol of the biofilm, PMA resulted in a significant total reduction of qPCR amplification of about 4 log10 . P. gingivalis showed a vitality reduction in the biofilm of 3 log10 , while A. actinomycetemcomitans and F. nucleatum showed a 2 log10 reduction. These results demonstrate the efficiency of PMA for differentiating viable and dead P. gingivalis, A. actinomycetemcomitans and F. nucleatum cells, as well as total bacteria, in an in vitro biofilm model, after being exposed to an antimicrobial agent. Hence, this PMA-qPCR method may be useful for studying the effect of antimicrobial agents aimed at oral biofilms. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

The Beneficial Effect of Equisetum giganteum L. against Candida Biofilm Formation: New Approaches to Denture Stomatitis

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Rafaela A. S. Alavarce

2015-01-01

Full Text Available Equisetum giganteum L. (E. giganteum, Equisetaceae, commonly called “giant horsetail,” is an endemic plant of Central and South America and is used in traditional medicine as diuretic and hemostatic in urinary disorders and in inflammatory conditions among other applications. The chemical composition of the extract EtOH 70% of E. giganteum has shown a clear presence of phenolic compounds derived from caffeic and ferulic acids and flavonoid heterosides derived from quercitin and kaempferol, in addition to styrylpyrones. E. giganteum, mainly at the highest concentrations, showed antimicrobial activity against the relevant microorganisms tested: Escherichia coli, Staphylococcus aureus, and Candida albicans. It also demonstrated antiadherent activity on C. albicans biofilms in an experimental model that is similar to dentures. Moreover, all concentrations tested showed anti-inflammatory activity. The extract did not show cytotoxicity in contact with human cells. These properties might qualify E. giganteum extract to be a promising alternative for the topic treatment and prevention of oral candidiasis and denture stomatitis.

Typing and virulence factors of food-borne Candida spp. isolates.

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Rajkowska, Katarzyna; Kunicka-Styczyńska, Alina

2018-08-20

Food-borne yeasts, excluding yeasts used as starter cultures, are commonly considered as food spoilage microorganisms. However, the incidence of non-C. albicans Candida (NCAC) infections has increased considerably over the past two decades. Although 15 Candida species are frequently identified as pathogens, a threat to human from food-borne Candida is poorly recognized. In the present study food-borne NCAC were characterized for the virulence factors, known to be associated with yeast pathogenicity. All food-borne strains in planktonic forms and 89% in biofilm structures represented biotypes established for C. albicans, and 61% demonstrated hemolytic activity. 56-94% of food-borne isolates formed biofilms on glass and biomaterials at a level comparable to clinical C. albicans. Nine out of eighteen tested food-borne NCAC strains (C. krusei, C. lusitaniae, C. famata, C. colliculosa, C. parapsilosis, C. tropicalis) showed similarity to clinical C. albicans in terms of their biotypes and the tested virulence factors, allocating them in a group of risk of potential pathogens. However, their capacity to grow at 37 °C seems to be the preliminary criterion in the study of potential virulence of food-borne yeasts. Copyright © 2018 Elsevier B.V. All rights reserved.

Determination of biofilm production by Candida tropicalis isolated from hospitalized patients and its relation to cellular surface hydrophobicity, plastic adherence and filamentation ability.

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Galán-Ladero, M A; Blanco-Blanco, M T; Hurtado, C; Pérez-Giraldo, C; Blanco, M T; Gómez-García, A C

2013-09-01

Candida tropicalis is an emerging virulent species. The aim of this study is to determine the biofilm-forming ability of 29 strains of C. tropicalis isolated from inpatients, and to examine its relation with other virulence factors such as cellular surface hydrophobicity (CSH), immediate (15 min, IA) and late (24 h, LA) plastic adherence and filamentation ability. The study was performed in parallel using two incubation temperatures - 37 and 22 °C - to determine the effect of growth temperature variations on these pathogenic attributes of C. tropicalis. Biofilm formation (BF) was measured by optical density (OD) and by XTT reduction (XTT); Slime index (SI), which includes growth as a correction factor in BF, was calculated in both methods. All strains were hydrophobic and adherent - at 15 min and 24 h - at both temperatures, with higher values for 22 °C; the adhered basal yeast layer appears to be necessary to achieve subsequent development of biofilm. Filamentation ability varied from 76.2% of strains at 37 °C to 26.6% at 22 °C. All C. tropicalis strains were biofilm producers, with similar results obtained using OD determination and XTT measurement to evaluation methods; SI is useful when good growth is not presented. BF at 37 °C was similar at 24 h and 96 h incubation; conversely, at 22 °C, the highest number of biofilm-producing strains was detected at 96 h. CSH is an important pathogenic factor which is involved in adherence, is influenced by the filamentation of yeast, and plays a critical role in BF. Copyright © 2013 John Wiley & Sons, Ltd.

Azasordarins: Susceptibility of Fluconazole-Susceptible and Fluconazole-Resistant Clinical Isolates of Candida spp. to GW 471558