scholarly journals Candida albicans Biofilms Produce Antifungal-Tolerant Persister Cells

2006 ◽  
Vol 50 (11) ◽  
pp. 3839-3846 ◽  
Author(s):  
Michael D. LaFleur ◽  
Carol A. Kumamoto ◽  
Kim Lewis
Keyword(s):  
Candida Albicans ◽  
Amphotericin B ◽  
Biofilm Formation ◽  
Fungal Pathogens ◽  
Wild Type ◽  
Persister Cells ◽  
Dormant Cells ◽  
Multidrug Tolerance ◽  
Candida Albicans Biofilms

ABSTRACT Fungal pathogens form biofilms that are highly recalcitrant to antimicrobial therapy. The expression of multidrug resistance pumps in young biofilms has been linked to increased resistance to azoles, but this mechanism does not seem to underlie the resistance of mature biofilms that is a model of in vivo infection. The mechanism of drug resistance of mature biofilms remains largely unknown. We report that biofilms formed by the major human pathogen Candida albicans exhibited a strikingly biphasic killing pattern in response to two microbicidal agents, amphotericin B, a polyene antifungal, and chlorhexidine, an antiseptic, indicating that a subpopulation of highly tolerant cells, termed persisters, existed. The extent of killing with a combination of amphotericin B and chlorhexidine was similar to that observed with individually added antimicrobials. Thus, surviving persisters form a multidrug-tolerant subpopulation. Interestingly, surviving C. albicans persisters were detected only in biofilms and not in exponentially growing or stationary-phase planktonic populations. Reinoculation of cells that survived killing of the biofilm by amphotericin B produced a new biofilm with a new subpopulation of persisters. This suggests that C. albicans persisters are not mutants but phenotypic variants of the wild type. Using a stain for dead cells, rare dark cells were visible in a biofilm after amphotericin B treatment, and a bright and a dim population were physically sorted from this biofilm. Only the dim cells produced colonies, showing that this method allows the isolation of yeast persisters. Given that persisters formed only in biofilms, mutants defective in biofilm formation were examined for tolerance of amphotericin B. All of the known mutants affected in biofilm formation were able to produce normal levels of persisters. This finding indicates that attachment rather than formation of a complex biofilm architecture initiates persister formation. Bacteria produce multidrug-tolerant persister cells in both planktonic and biofilm populations, and it appears that yeasts and bacteria have evolved analogous strategies that assign the function of survival to a small part of the population. In bacteria, persisters are dormant cells. It remains to be seen whether attachment initiates dormancy that leads to the formation of fungal persisters. This study suggests that persisters may be largely responsible for the multidrug tolerance of fungal biofilms.

scholarly journals Patients with Long-Term Oral Carriage Harbor High-Persister Mutants of Candida albicans

2009 ◽  
Vol 54 (1) ◽  
pp. 39-44 ◽  
Author(s):  
Michael D. LaFleur ◽  
Qingguo Qi ◽  
Kim Lewis
Keyword(s):  
Candida Albicans ◽  
Amphotericin B ◽  
Antimicrobial Agents ◽  
Oral Candidiasis ◽  
Wild Type ◽  
Persister Cells ◽  
Microtiter Plates ◽  
High Concentrations

ABSTRACT Fungal biofilms produce a small number of persister cells which can tolerate high concentrations of fungicidal agents. Persisters form upon attachment to a surface, an important step in the pathogenesis of Candida strains. The periodic application of antimicrobial agents may select for strains with increased levels of persister cells. In order to test this possibility, 150 isolates of Candida albicans and C. glabrata were obtained from cancer patients who were at high risk for the development of oral candidiasis and who had been treated with topical chlorhexidine once a day. Persister levels were measured by exposing biofilms growing in the wells of microtiter plates to high concentrations of amphotericin B and plating for survivors. The persister levels of the isolates varied from 0.2 to 9%, and strains isolated from patients with long-term carriage had high levels of persisters. High-persister strains were isolated from every patient with Candida carriage of more than 8 consecutive weeks but from no patients with transient carriage. All of the high-persister isolates had an amphotericin B MIC that was the same as that for the wild type, indicating that these strains were drug-tolerant rather than drug-resistant mutants. Biofilms of the majority of high-persister strains also showed an increased tolerance to chlorhexidine and had the same MIC for this antimicrobial as the wild type. This study suggests that persister cells are clinically relevant, and antimicrobial therapy selects for high-persister strains in vivo. The drug tolerance of persisters may be a critical but overlooked component responsible for antimicrobial drug failure and relapsing infections.

scholarly journals The Heat-Induced Molecular Disaggregase Hsp104 of Candida albicans Plays a Role in Biofilm Formation and Pathogenicity in a Worm Infection Model

2012 ◽  
Vol 11 (8) ◽  
pp. 1012-1020 ◽  
Author(s):  
Alessandro Fiori ◽  
Soňa Kucharíková ◽  
Gilmer Govaert ◽  
Bruno P. A. Cammue ◽  
Karin Thevissen ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Stress Responses ◽  
Biofilm Formation ◽  
Fungal Pathogens ◽  
Elevated Temperatures ◽  
Structural Defects ◽  
Infection Model ◽  
Content Type

ABSTRACT The consequences of deprivation of the molecular chaperone Hsp104 in the fungal pathogen Candida albicans were investigated. Mutants lacking HSP104 became hypersusceptible to lethally high temperatures, similarly to the corresponding mutants of Saccharomyces cerevisiae , whereas normal susceptibility was restored upon reintroduction of the gene. By use of a strain whose only copy of HSP104 is an ectopic gene under the control of a tetracycline-regulated promoter, expression of Hsp104 prior to the administration of heat shock could be demonstrated to be sufficient to confer protection from the subsequent temperature increase. This result points to a key role for Hsp104 in orchestrating the cell response to elevated temperatures. Despite their not showing evident growth or morphological defects, biofilm formation by cells lacking HSP104 proved to be defective in two established in vitro models that use polystyrene and polyurethane as the substrates. Biofilms formed by the wild-type and HSP104 -reconstituted strains showed patterns of intertwined hyphae in the extracellular matrix. In contrast, biofilm formed by the hsp104 Δ/ hsp104 Δ mutant showed structural defects and appeared patchy and loose. Decreased virulence of the hsp104 Δ/ hsp104 Δ mutant was observed in the Caenorhabditis elegans infection model, in which high in vivo temperature does not play a role. In agreement with the view that stress responses in fungal pathogens may have evolved to provide niche-specific adaptation to environmental conditions, these results provide an indication of a temperature-independent role for Hsp104 in support of Candida albicans virulence, in addition to its key role in governing thermotolerance.

scholarly journals Toll-Like Receptor 9 Modulates Macrophage Antifungal Effector Function during Innate Recognition of Candida albicans and Saccharomyces cerevisiae

2011 ◽  
Vol 79 (12) ◽  
pp. 4858-4867 ◽  
Author(s):  
Pia V. Kasperkovitz ◽  
Nida S. Khan ◽  
Jenny M. Tam ◽  
Michael K. Mansour ◽  
Peter J. Davids ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Candida Albicans ◽  
Fungal Pathogens ◽  
Toll Like Receptor ◽  
Wild Type ◽  
Necrosis Factor Alpha ◽  
Content Type ◽  
Tnf Α ◽  
Tlr9 Gene

ABSTRACTPhagocytic responses are critical for effective host defense against opportunistic fungal pathogens. Macrophages sample the phagosomal content and orchestrate the innate immune response. Toll-like receptor 9 (TLR9) recognizes unmethylated CpG DNA and is activated by fungal DNA. Here we demonstrate that specific triggering of TLR9 recruitment to the macrophage phagosomal membrane is a conserved feature of fungi of distinct phylogenetic origins, includingCandida albicans,Saccharomyces cerevisiae,Malassezia furfur, andCryptococcus neoformans. The capacity to trigger phagosomal TLR9 recruitment was not affected by a loss of fungal viability or cell wall integrity. TLR9 deficiency has been linked to increased resistance to murine candidiasis and to restriction of fungal growthin vivo. Macrophages lacking TLR9 demonstrate a comparable capacity for phagocytosis and normal phagosomal maturation compared to wild-type macrophages. We now show that TLR9 deficiency increases macrophage tumor necrosis factor alpha (TNF-α) production in response toC. albicansandS. cerevisiae, independent of yeast viability. The increase in TNF-α production was reversible by functional complementation of the TLR9 gene, confirming that TLR9 was responsible for negative modulation of the cytokine response. Consistently, TLR9 deficiency enhanced the macrophage effector response by increasing macrophage nitric oxide production. Moreover, microbicidal activity againstC. albicansandS. cerevisiaewas more efficient in TLR9 knockout (TLR9KO) macrophages than in wild-type macrophages. In conclusion, our data demonstrate that TLR9 is compartmentalized selectively to fungal phagosomes and negatively modulates macrophage antifungal effector functions. Our data support a model in which orchestration of antifungal innate immunity involves a complex interplay of fungal ligand combinations, host cell machinery rearrangements, and TLR cooperation and antagonism.

scholarly journals Visible Lights Combined with Photosensitizing Compounds Are Effective against Candida albicans Biofilms

2021 ◽  
Vol 9 (3) ◽  
pp. 500 ◽  
Author(s):  
Priyanka Bapat ◽  
Gurbinder Singh ◽  
Clarissa J. Nobile
Keyword(s):  
Candida Albicans ◽  
Photodynamic Therapy ◽  
Biofilm Formation ◽  
Fungal Pathogens ◽  
Fungal Infections ◽  
Therapeutic Strategies ◽  
Antifungal Drugs ◽  
Clinical Settings ◽  
Drug Resistant ◽  
Candida Albicans Biofilms

Fungal infections are increasing in prevalence worldwide, especially in immunocompromised individuals. Given the emergence of drug-resistant fungi and the fact that there are only three major classes of antifungal drugs available to treat invasive fungal infections, there is a need to develop alternative therapeutic strategies effective against fungal infections. Candida albicans is a commensal of the human microbiota that is also one of the most common fungal pathogens isolated from clinical settings. C. albicans possesses several virulence traits that contribute to its pathogenicity, including the ability to form drug-resistant biofilms, which can make C. albicans infections particularly challenging to treat. Here, we explored red, green, and blue visible lights alone and in combination with common photosensitizing compounds for their efficacies at inhibiting and disrupting C. albicans biofilms. We found that blue light inhibited biofilm formation and disrupted mature biofilms on its own and that the addition of photosensitizing compounds improved its antibiofilm potential. Red and green lights, however, inhibited biofilm formation only in combination with photosensitizing compounds but had no effects on disrupting mature biofilms. Taken together, these results suggest that photodynamic therapy may be an effective non-drug treatment for fungal biofilm infections that is worthy of further exploration.

scholarly journals Optimum Inhibition of Amphotericin-B-Resistant Candida albicans Strain in Single- and Mixed-Species Biofilms by Candida and Non-Candida Terpenoids

Biomolecules ◽  
2020 ◽  
Vol 10 (2) ◽  
pp. 342 ◽  
Author(s):  
Hidaya F. Z. Touil ◽  
Kebir Boucherit ◽  
Zahia Boucherit-Otmani ◽  
Ghalia Kohder ◽  
Mohamed Madkour ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Amphotericin B ◽  
Biofilm Formation ◽  
Fungal Pathogens ◽  
Care Center ◽  
Drug Resistant ◽  
Inhibitory Effects ◽  
Mixed Species ◽  
Surgical Devices ◽  
Opportunistic Mycoses

Candida albicans is one of the most common human fungal pathogens and represents the most important cause of opportunistic mycoses worldwide. Surgical devices including catheters are easily contaminated with C. albicans via its formation of drug-resistant biofilms. In this study, amphotericin-B-resistant C. albicans strains were isolated from surgical devices at an intensive care center. The objective of this study was to develop optimized effective inhibitory treatment of resistant C. albicans by terpenoids, known to be produced naturally as protective signals. Endogenously produced farnesol by C. albicans yeast and plant terpenoids, carvacrol, and cuminaldehyde were tested separately or in combination on amphotericin-B-resistant C. albicans in either single- or mixed-infections. The results showed that farnesol did not inhibit hyphae formation when associated with bacteria. Carvacrol and cuminaldehyde showed variable inhibitory effects on C. albicans yeast compared to hyphae formation. A combination of farnesol with carvacrol showed synergistic inhibitory activities not only on C. albicans yeast and hyphae, but also on biofilms formed from single- and mixed-species and at reduced doses. The combined terpenoids also showed biofilm-penetration capability. The aforementioned terpenoid combination will not only be useful in the treatment of different resistant Candida forms, but also in the safe prevention of biofilm formation.

scholarly journals Effect of Cold Atmospheric Plasma Jet Associated to Polyene Antifungals on Candida albicans Biofilms

Molecules ◽  
2021 ◽  
Vol 26 (19) ◽  
pp. 5815
Author(s):  
Lady Daiane Pereira Leite ◽  
Maria Alcionéia Carvalho de Oliveira ◽  
Mariana Raquel da Cruz Vegian ◽  
Aline da Graça Sampaio ◽  
Thalita Mayumi Castaldelli Nishime ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Amphotericin B ◽  
Fungal Infections ◽  
Negative Control ◽  
Atmospheric Plasma ◽  
Promising Alternative ◽  
Cold Atmospheric Plasma ◽  
Candida Albicans Biofilms

The increasing incidence of antifungal resistance represents a great challenge in the medical area and, for this reason, new therapeutic alternatives for the treatment of fungal infections are urgently required. Cold atmospheric plasma (CAP) has been proposed as a promising alternative technique for the treatment of superficial candidiasis, with inhibitory effect both in vitro and in vivo. However, little is known on the association of CAP with conventional antifungals. The aim of this study was to evaluate the effects of the association between CAP and conventional polyene antifungals on Candida albicans biofilms. C. albicans SC 5314 and a clinical isolate were used to grow 24 or 48 h biofilms, under standardized conditions. After that, the biofilms were exposed to nystatin, amphotericin B and CAP, separately or in combination. Different concentrations of the antifungals and sequences of treatment were evaluated to establish the most effective protocol. Biofilms viability after the treatments was compared to negative control. Data were compared by One-way ANOVA and post hoc Tukey (5%). The results demonstrate that 5 min exposure to CAP showed more effective antifungal effect on biofilms when compared to nystatin and amphotericin B. Additionally, it was detected that CAP showed similar (but smaller in magnitude) effects when applied in association with nystatin and amphotericin B at 40 µg/mL and 60 µg/mL. Therefore, it can be concluded that the application of CAP alone was more effective against C. albicans biofilms than in combination with conventional polyene antifungal agents.

scholarly journals A Screen for Small Molecules to Target Candida albicans Biofilms

2020 ◽  
Vol 7 (1) ◽  
pp. 9
Author(s):  
Matthew B. Lohse ◽  
Craig L. Ennis ◽  
Nairi Hartooni ◽  
Alexander D. Johnson ◽  
Clarissa J. Nobile
Keyword(s):  
Candida Albicans ◽  
Small Molecules ◽  
Amphotericin B ◽  
Biofilm Formation ◽  
Antifungal Drugs ◽  
Human Fungal Pathogen ◽  
Abiotic Surfaces ◽  
Candida Infections ◽  
Combinatorial Interactions ◽  
Candida Albicans Biofilms

The human fungal pathogen Candida albicans can form biofilms on biotic and abiotic surfaces, which are inherently resistant to antifungal drugs. We screened the Chembridge Small Molecule Diversity library containing 30,000 “drug-like” small molecules and identified 45 compounds that inhibited biofilm formation. These 45 compounds were then tested for their abilities to disrupt mature biofilms and for combinatorial interactions with fluconazole, amphotericin B, and caspofungin, the three antifungal drugs most commonly prescribed to treat Candida infections. In the end, we identified one compound that moderately disrupted biofilm formation on its own and four compounds that moderately inhibited biofilm formation and/or moderately disrupted mature biofilms only in combination with either caspofungin or fluconazole. No combinatorial interactions were observed between the compounds and amphotericin B. As members of a diversity library, the identified compounds contain “drug-like” chemical backbones, thus even seemingly “weak hits” could represent promising chemical starting points for the development and the optimization of new classes of therapeutics designed to target Candida biofilms.

scholarly journals Eap1p, an Adhesin That Mediates Candida albicans Biofilm Formation In Vitro and In Vivo

2007 ◽  
Vol 6 (6) ◽  
pp. 931-939 ◽  
Author(s):  
Fang Li ◽  
Michael J. Svarovsky ◽  
Amy J. Karlsson ◽  
Joel P. Wagner ◽  
Karen Marchillo ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Cell Adhesion ◽  
Biofilm Formation ◽  
Fungal Infections ◽  
Gene Dosage ◽  
Venous Catheter ◽  
Flow Chamber ◽  
Dependent Manner

ABSTRACT Candida albicans is the leading cause of systemic fungal infections in immunocompromised humans. The ability to form biofilms on surfaces in the host or on implanted medical devices enhances C. albicans virulence, leading to antimicrobial resistance and providing a reservoir for infection. Biofilm formation is a complex multicellular process consisting of cell adhesion, cell growth, morphogenic switching between yeast form and filamentous states, and quorum sensing. Here we describe the role of the C. albicans EAP1 gene, which encodes a glycosylphosphatidylinositol-anchored, glucan-cross-linked cell wall protein, in adhesion and biofilm formation in vitro and in vivo. Deleting EAP1 reduced cell adhesion to polystyrene and epithelial cells in a gene dosage-dependent manner. Furthermore, EAP1 expression was required for C. albicans biofilm formation in an in vitro parallel plate flow chamber model and in an in vivo rat central venous catheter model. EAP1 expression was upregulated in biofilm-associated cells in vitro and in vivo. Our results illustrate an association between Eap1p-mediated adhesion and biofilm formation in vitro and in vivo.

scholarly journals Abrogation of Neuraminidase Reduces Biofilm Formation, Capsule Biosynthesis, and Virulence of Porphyromonas gingivalis

2011 ◽  
Vol 80 (1) ◽  
pp. 3-13 ◽  
Author(s):  
Chen Li ◽  
Kurniyati ◽  
Bo Hu ◽  
Jiang Bian ◽  
Jianlan Sun ◽  
...  
Keyword(s):  
Porphyromonas Gingivalis ◽  
Biofilm Formation ◽  
Adult Population ◽  
Transcriptional Analysis ◽  
Wild Type ◽  
Content Type ◽  
Multiple Organs ◽  
Biochemical Analyses

ABSTRACTThe oral bacteriumPorphyromonas gingivalisis a key etiological agent of human periodontitis, a prevalent chronic disease that affects up to 80% of the adult population worldwide.P. gingivalisexhibits neuraminidase activity. However, the enzyme responsible for this activity, its biochemical features, and its role in the physiology and virulence ofP. gingivalisremain elusive. In this report, we found thatP. gingivalisencodes a neuraminidase, PG0352 (SiaPg). Transcriptional analysis showed thatPG0352is monocistronic and is regulated by a sigma70-like promoter. Biochemical analyses demonstrated that SiaPgis an exo-α-neuraminidase that cleaves glycosidic-linked sialic acids. Cryoelectron microscopy and tomography analyses revealed that thePG0352deletion mutant (ΔPG352) failed to produce an intact capsule layer. Compared to the wild type,in vitrostudies showed that ΔPG352 formed less biofilm and was less resistant to killing by the host complement.In vivostudies showed that while the wild type caused a spreading type of infection that affected multiple organs and all infected mice were killed, ΔPG352 only caused localized infection and all animals survived. Taken together, these results demonstrate that SiaPgis an important virulence factor that contributes to the biofilm formation, capsule biosynthesis, and pathogenicity ofP. gingivalis, and it can potentially serve as a new target for developing therapeutic agents againstP. gingivalisinfection.

scholarly journals Impact of the Order of Initiation of Fluconazole and Amphotericin B in Sequential or Combination Therapy on Killing of Candida albicans In Vitro and in a Rabbit Model of Endocarditis and Pyelonephritis

2001 ◽  
Vol 45 (2) ◽  
pp. 485-494 ◽  
Author(s):  
Arnold Louie ◽  
Pamela Kaw ◽  
Partha Banerjee ◽  
Weiguo Liu ◽  
George Chen ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Amphotericin B ◽  
Induced Resistance ◽  
Rabbit Model ◽  
In Vivo Studies ◽  
Infection Model ◽  
Simultaneous Exposure ◽  
The Impact

ABSTRACT In vitro time-kill studies and a rabbit model of endocarditis and pyelonephritis were used to define the impact that the order of exposure of Candida albicans to fluconazole (FLC) and amphotericin B (AMB), as sequential and combination therapies, had on the susceptibility of C. albicans to AMB and on the outcome. The contribution of FLC-induced resistance to AMB for C. albicans also was assessed. In vitro, AMB monotherapy rapidly killed each of four C. albicans strains; FLC alone was fungistatic. Preincubation of these fungi with FLC for 18 h prior to exposure to AMB decreased their susceptibilities to AMB for 8 to >40 h. Induced resistance to AMB was transient, but the duration of resistance increased with the length of FLC preincubation. Yeast sequentially incubated with FLC followed by AMB plus FLC (FLC→AMB+FLC) showed fungistatic growth kinetics similar to that of fungi that were exposed to FLC alone. This antagonistic effect persisted for at least 24 h. Simultaneous exposure of C. albicans to AMB and FLC [AMB+FLC(simult)] demonstrated activity similar to that with AMB alone for AMB concentrations of ≥1 μg/ml; antagonism was seen using an AMB concentration of 0.5 μg/ml. The in vitro findings accurately predicted outcomes in our rabbit infection model. In vivo, AMB monotherapy and treatment with AMB for 24 h followed by AMB plus FLC (AMB→AMB+FLC) rapidly sterilized kidneys and cardiac vegetations. AMB+FLC(simult) and FLC→AMB treatments were slower in clearing fungi from infected tissues. FLC monotherapy and FLC→AMB+FLC were both fungistatic and were the least active regimens. No adverse interaction was observed between AMB and FLC for the AMB→FLC regimen. However, FLC→AMB treatment was slower than AMB alone in clearing fungi from tissues. Thus, our in vitro and in vivo studies both demonstrate that preexposure of C. albicans to FLC reduces fungal susceptibility to AMB. The length of FLC preexposure and whether AMB is subsequently used alone or in combination with FLC determine the duration of induced resistance to AMB.

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