scholarly journals Reversible fluconazole resistance in Candida albicans: a potential in vitro model.

1997 ◽  
Vol 41 (3) ◽  
pp. 535-539 ◽  
Author(s):  
H M Calvet ◽  
M R Yeaman ◽  
S G Filler
Keyword(s):  
Candida Albicans ◽  
Molecular Mechanisms ◽  
Rabbit Model ◽  
Serial Passage ◽  
Yeast Nitrogen Base ◽  
Nitrogen Base ◽  
Fluconazole Resistance ◽  
Karyotypic Analysis

To study the development and potential mechanisms of antifungal resistance in relation to antifungal exposure, reversible fluconazole resistance was examined in vitro. Candida albicans ATCC 36082 blastospores were passed in liquid yeast nitrogen base medium containing either 4, 8, 16, or 128 micrograms of fluconazole per ml, and susceptibility testing was performed after each passage. High-level fluconazole resistance (50% inhibitory concentration, > 256 micrograms/ml) developed in the isolates after serial passage in medium containing 8, 16, or 128 micrograms of fluconazole per ml, but not in isolates passed in 4 micrograms of fluconazole per ml. Reduced susceptibility was noted within four to seven passages, which was equivalent to 14 to 19 days of exposure to the drug. However, all isolates returned to the susceptible phenotype after 8 to 15 passages in medium lacking the drug; thus, fluconazole resistance was reversible in vitro. In vivo, organisms retained the resistant phenotype after a single passage in the rabbit model of infective endocarditis. Restriction digest profiles and karyotypic analysis of the parent strain and selected fluconazole-resistant and -susceptible isolates from each group were identical. Investigations into the molecular mechanisms of this reversible resistance failed to reveal increased accumulation of mRNA for 14 alpha-demethylase, the target enzyme for fluconazole, or for the candidal multidrug transporters CDR1 and BENr. This process of continuous in vitro exposure to antifungal drug may be useful as a model for studying the effects of different antifungal agents and dosing regimens on the development of resistance and for defining the mechanism(s) of reversible resistance.

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.

scholarly journals Susceptibility to Thrombin-Induced Platelet Microbicidal Protein Is Associated with Increased Fluconazole Efficacy against Experimental Endocarditis Due to Candida albicans

2004 ◽  
Vol 48 (8) ◽  
pp. 3051-3056 ◽  
Author(s):  
Michael R. Yeaman ◽  
Darwin Cheng ◽  
Bhavesh Desai ◽  
Leon I. Kupferwasser ◽  
Yan-Qiong Xiong ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Host Defense ◽  
Rabbit Model ◽  
Fungal Burden ◽  
Treatment Groups ◽  
Time Points ◽  
Platelet Microbicidal Protein ◽  
The Relationship

ABSTRACT Platelet microbicidal proteins (PMPs) are believed to be integral to host defense against endovascular infection. We previously demonstrated that susceptibility to thrombin-induced PMP 1 (tPMP-1) in vitro negatively influences Candida albicans virulence in the rabbit model of infective endocarditis (IE). This study evaluated the relationship between in vitro tPMP-1 susceptibility (tPMP-1s) or resistance (tPMP-1r) and efficacy of fluconazole (FLU) therapy of IE due to C. albicans. Candida IE was established in rabbits with either tPMP-1s or tPMP-1r strains. Treatment groups received FLU (100 mg/kg/day) intraperitoneally for 7 or 14 days; control animals received no therapy. At these time points, cardiac vegetations, kidneys, and spleens were quantitatively cultured to assess fungal burden. At both 7 and 14 days and in all target tissues, the extent of candidal clearance by FLU was greater in animals infected with the tPMP-1s strain than in those infected with the tPMP-1r strain. These differences were statistically significant in the spleen and kidney. Corroborating these in vivo data, FLU (a candidastatic agent), in combination with tPMP-1, exerted an enhanced fungicidal effect in vitro against tPMP-1s and tPMP-1r C. albicans, with the extent of this effect greatest against the tPMP-1s strain. Collectively, these results support the concept that tPMP-1 susceptibility contributes to the net efficacy of FLU against C. albicans IE in vivo, particularly in tissues in which platelets and tPMP-1 likely play significant roles in host defense.

scholarly journals A Combination of Polybacterial MV140 and Candida albicans V132 as a Potential Novel Trained Immunity-Based Vaccine for Genitourinary Tract Infections

2021 ◽  
Vol 11 ◽  
Author(s):  
Leticia Martin-Cruz ◽  
Carmen Sevilla-Ortega ◽  
Cristina Benito-Villalvilla ◽  
Carmen M. Diez‐Rivero ◽  
Silvia Sanchez-Ramón ◽  
...  
Keyword(s):  
T Cells ◽  
Candida Albicans ◽  
Molecular Mechanisms ◽  
Genitourinary Tract ◽  
Vaccine Formulation ◽  
Trained Immunity ◽  
Immunological Mechanisms ◽  
Tract Infections

Recurrent urinary tract infections (RUTIs) and recurrent vulvovaginal candidiasis (RVVCs) represent major healthcare problems with high socio-economic impact worldwide. Antibiotic and antifungal prophylaxis remain the gold standard treatments for RUTIs and RVVCs, contributing to the massive rise of antimicrobial resistance, microbiota alterations and co-infections. Therefore, the development of novel vaccine strategies for these infections are sorely needed. The sublingual heat-inactivated polyvalent bacterial vaccine MV140 shows clinical efficacy for the prevention of RUTIs and promotes Th1/Th17 and IL-10 immune responses. V132 is a sublingual preparation of heat-inactivated Candida albicans developed against RVVCs. A vaccine formulation combining both MV140 and V132 might well represent a suitable approach for concomitant genitourinary tract infections (GUTIs), but detailed mechanistic preclinical studies are still needed. Herein, we showed that the combination of MV140 and V132 imprints human dendritic cells (DCs) with the capacity to polarize potent IFN-γ– and IL-17A–producing T cells and FOXP3+ regulatory T (Treg) cells. MV140/V132 activates mitogen-activated protein kinases (MAPK)-, nuclear factor-κB (NF-κB)- and mammalian target of rapamycin (mTOR)-mediated signaling pathways in human DCs. MV140/V132 also promotes metabolic and epigenetic reprogramming in human DCs, which are key molecular mechanisms involved in the induction of innate trained immunity. Splenocytes from mice sublingually immunized with MV140/V132 display enhanced proliferative responses of CD4+ T cells not only upon in vitro stimulation with the related antigens contained in the vaccine formulation but also upon stimulation with phytohaemagglutinin. Additionally, in vivo sublingual immunization with MV140/V132 induces the generation of IgG and IgA antibodies against all the components contained in the vaccine formulation. We uncover immunological mechanisms underlying the potential mode of action of a combination of MV140 and V132 as a novel promising trained immunity-based vaccine (TIbV) for GUTIs.

In Vitro and In Vivo Efficacy of the Triazole TAK-187 against Cryptococcus neoformans

1998 ◽  
Vol 42 (10) ◽  
pp. 2630-2632 ◽  
Author(s):  
Wiley A. Schell ◽  
Gisele Madeira Duboc De Almeida ◽  
Richard K. Dodge ◽  
Kenji Okonogi ◽  
John R. Perfect
Keyword(s):  
Cerebrospinal Fluid ◽  
Cryptococcus Neoformans ◽  
Cryptococcal Meningitis ◽  
Rabbit Model ◽  
Intermittent Therapy ◽  
In Vitro Activity ◽  
Fluconazole Resistance ◽  
In Vivo Efficacy

ABSTRACT Multiple isolates of Cryptococcus neoformans, including those with fluconazole resistance, were tested to assess the in vitro activity of the new triazole TAK-187. MICs of TAK-187 were at least eightfold lower than those of fluconazole, and fungicidal concentrations for most isolates were 4 μg/ml or less. TAK-187 also was evaluated as intermittent therapy using two dosages in a rabbit model of experimental cryptococcal meningitis. Compared to daily treatment with fluconazole, as little as two doses of TAK-187 given 7 days apart were found to be effective. Plasma and cerebrospinal fluid TAK-187 concentrations were many times higher than MICs and fungicidal concentrations. Based upon its therapeutic efficacy and long half-life in the rabbit model, TAK-187 should be investigated for intermittent dosing in treatment or suppression of cryptococcal infections in humans.

scholarly journals Growth Media for Mixed Multispecies Oropharyngeal Biofilm Compositions on Silicone

2019 ◽  
Vol 2019 ◽  
pp. 1-8 ◽  
Author(s):  
Matthias Leonhard ◽  
Beata Zatorska ◽  
Doris Moser ◽  
Berit Schneider-Stickler
Keyword(s):  
Culture Conditions ◽  
Microbial Colonization ◽  
Future Research ◽  
Growth Media ◽  
Yeast Nitrogen Base ◽  
Microbial Composition ◽  
Nitrogen Base ◽  
Voice Prostheses

Aims. Microbial colonization of silicone voice prostheses by bacteria and Candida species limits the device lifetime of modern voice prostheses in laryngectomized patients. Thus, research focuses on biofilm inhibitive properties of novel materials, coatings, and surface enhancements. Goal of this in vitro study was the evaluation of seven commonly used growth media to simulate growth of mixed oropharyngeal species as mesoscale biofilms on prosthetic silicone for future research purposes. Methods and Results. Yeast Peptone Dextrose medium (YPD), Yeast Nitrogen Base medium (YNB), M199 medium, Spider medium, RPMI 1640 medium, Tryptic Soy Broth (TSB), and Fetal Bovine Serum (FBS) were used to culture combined mixed Candida strains and mixed bacterial-fungal compositions on silicone over the period of 22 days. The biofilm surface spread and the microscopic growth showed variations from in vivo biofilms depending on the microbial composition and growth medium. Conclusion. YPD and FBS prove to support continuous in vitro growth of mixed bacterial-fungal oropharyngeal biofilms deposits over weeks as needed for longterm in vitro testing with oropharyngeal biofilm compositions. Significance and Impact of Study. The study provides data on culture conditions for mixed multispecies biofilm compositions that can be used for future prosthesis designs.

scholarly journals Alcohol Dehydrogenase Restricts the Ability of the Pathogen Candida albicans To Form a Biofilm on Catheter Surfaces through an Ethanol-Based Mechanism

2006 ◽  
Vol 74 (7) ◽  
pp. 3804-3816 ◽  
Author(s):  
Pranab K. Mukherjee ◽  
Sotohy Mohamed ◽  
Jyotsna Chandra ◽  
Duncan Kuhn ◽  
Shuqing Liu ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Alcohol Dehydrogenase ◽  
Biofilm Formation ◽  
Rabbit Model ◽  
Treatment Strategies ◽  
Northern Blotting ◽  
Confocal Scanning Laser Microscopy ◽  
Biofilm Inhibition

ABSTRACT Candida biofilms formed on indwelling medical devices are increasingly associated with severe infections. In this study, we used proteomics and Western and Northern blotting analyses to demonstrate that alcohol dehydrogenase (ADH) is downregulated in Candida biofilms. Disruption of ADH1 significantly (P = 0.0046) enhanced the ability of Candida albicans to form biofilm. Confocal scanning laser microscopy showed that the adh1 mutant formed thicker biofilm than the parent strain (210 μm and 140 μm, respectively). These observations were extended to an engineered human oral mucosa and an in vivo rat model of catheter-associated biofilm. Inhibition of Candida ADH enzyme using disulfiram and 4-methylpyrazole resulted in thicker biofilm (P < 0.05). Moreover, biofilms formed by the adh1 mutant strain produced significantly smaller amounts of ethanol, but larger amounts of acetaldehyde, than biofilms formed by the parent and revertant strains (P < 0.0001), demonstrating that the effect of Adh1p on biofilm formation is mediated by its enzymatic activity. Furthermore, we found that 10% ethanol significantly inhibited biofilm formation in vitro, with complete inhibition of biofilm formation at ethanol concentrations of ≥20%. Similarly, using a clinically relevant rabbit model of catheter-associated biofilm, we found that ethanol treatment inhibited biofilm formation by C. albicans in vivo (P < 0.05) but not by Staphylococcus spp. (P > 0.05), indicating that ethanol specifically inhibits Candida biofilm formation. Taken together, our studies revealed that Adh1p contributes to the ability of C. albicans to form biofilms in vitro and in vivo and that the protein restricts biofilm formation through an ethanol-dependent mechanism. These results are clinically relevant and may suggest novel antibiofilm treatment strategies.

scholarly journals Pentamidine Is Active In Vitro against Fusarium Species

2003 ◽  
Vol 47 (10) ◽  
pp. 3252-3259 ◽  
Author(s):  
Michail S. Lionakis ◽  
Russell E. Lewis ◽  
George Samonis ◽  
Dimitrios P. Kontoyiannis
Keyword(s):  
Clinical Laboratory ◽  
Fusarium Species ◽  
Tissue Perfusion ◽  
In Vivo Studies ◽  
National Committee ◽  
Yeast Nitrogen Base ◽  
Nitrogen Base ◽  
Minimum Fungicidal Concentration

ABSTRACT Fusariosis is an emerging opportunistic mycosis against which currently used antifungals have limited activity. Here, we investigated the in vitro activities of pentamidine (PNT) against 10 clinical isolates of Fusarium species (five Fusarium solani isolates and five non-F. solani isolates) by using the National Committee for Clinical Laboratory Standards microdilution method in three different media (RPMI, RPMI-2, and a yeast nitrogen base medium), disk diffusion testing, and viability dye staining. PNT had significant activities against all 10 Fusarium isolates. Non-F. solani isolates were more susceptible than F. solani isolates (P < 0.05). Additionally, PNT was fungicidal against all non-F. solani isolates, whereas it had fungistatic effects against four of the five F. solani isolates. PNT also exhibited greater activity against conidial than against hyphal development of the fungus. This fungicidal activity against non-F. solani Fusarium isolates was confirmed microscopically after staining of PNT-treated Fusarium oxysporum hyphae with the fluorescent viability dyes 5,(6)-carboxyfluorescein diacetate (CFDA) and bis-(1,3-dibutylbarbituric acid) trimethine oxonol (DiBAC). The MICs at which 50% of the isolates were inhibited (2 μg/ml for non-F. solani isolates and 4 μg/ml for F. solani isolates) and the minimum fungicidal concentration at which 50% of the isolates were killed (8 μg/ml for non-F. solani isolates) were much lower than the PNT tissue concentrations previously reported in humans using conventional daily intravenous PNT dosing. Finally, PNT was more active against Fusarium isolates in a hypoxic environment of in vitro growth (P < 0.05). This finding may be clinically significant, because Fusarium, an angiotropic mold, causes tissue infarcts with resultant low tissue perfusion. Our findings suggest that PNT may have a role in the management of Fusarium infections. Future in vivo studies are needed to verify these in vitro findings.

scholarly journals Fluconazole Treatment Is Effective against a Candida albicans erg3/erg3 Mutant In Vivo Despite In Vitro Resistance

2006 ◽  
Vol 50 (2) ◽  
pp. 580-586 ◽  
Author(s):  
Taiga Miyazaki ◽  
Yoshitsugu Miyazaki ◽  
Koichi Izumikawa ◽  
Hiroshi Kakeya ◽  
Shunichi Miyakoshi ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Resistance Mechanism ◽  
Resistance Mechanisms ◽  
In Vivo Studies ◽  
Azole Resistance ◽  
In Vitro Testing ◽  
Fluconazole Resistance ◽  
Hyphal Formation

ABSTRACT Candida albicans ERG3 encodes a sterol C5,6-desaturase which is essential for synthesis of ergosterol. Defective sterol C5,6 desaturation has been considered to be one of the azole resistance mechanisms in this species. However, the clinical relevance of this resistance mechanism is still unclear. In this study, we created a C. albicans erg3/erg3 mutant by the “Ura-blaster” method and confirmed the expected azole resistance using standard in vitro testing and the presence of ergosta-7,22-dien-3β-ol instead of ergosterol. For in vivo studies, a wild-type URA3 was placed back into its native locus in the erg3 homozygote to avoid positional effects on URA3 expression. Defective hyphal formation of the erg3 homozygote was observed not only in vitro but in kidney tissues. A marked attenuation of virulence was shown by the longer survival and the lower kidney burdens of mice inoculated with the reconstituted Ura+ erg3 homozygote relative to the control. To assess fluconazole efficacy in a murine model of disseminated candidiasis, inoculum sizes of the control and the erg3 homozygote were chosen which provided a similar organ burden. Under these conditions, fluconazole was highly effective in reducing the organ burden in both groups. This study demonstrates that an ERG3 mutation causing inactivation of sterol C5,6-desaturase cannot confer fluconazole resistance in vivo by itself regardless of resistance measured by standard in vitro testing. The finding questions the clinical significance of this resistance mechanism.

scholarly journals In Vitro Activity and In Vivo Efficacy of Icofungipen (PLD-118), a Novel Oral Antifungal Agent, against the Pathogenic Yeast Candida albicans

2006 ◽  
Vol 50 (9) ◽  
pp. 3011-3018 ◽  
Author(s):  
Andreja Hasenoehrl ◽  
Tatjana Galić ◽  
Gabrijela Ergović ◽  
Nataša Maršić ◽  
Mihael Skerlev ◽  
...  
Keyword(s):  
Amino Acids ◽  
Candida Albicans ◽  
Trna Synthetase ◽  
Vitro Activity ◽  
Growth Media ◽  
In Vitro Activity ◽  
Nitrogen Base ◽  
In Vivo Efficacy

ABSTRACT Icofungipen (PLD-118) is the representative of a novel class of antifungals, beta amino acids, active against Candida species. It has been taken through phase II clinical trials. The compound actively accumulates in yeast, competitively inhibiting isoleucyl-tRNA synthetase and consequently disrupting protein biosynthesis. As a result, in vitro activity can be studied only in chemically defined growth media without free amino acids that would compete with the uptake of the compound. The MIC of icofungipen was reproducibly measured in a microdilution assay using yeast nitrogen base medium at pH 6 to 7 after 24 h of incubation at 30 to 37°C using an inoculum of 50 to 100 CFU/well. The MICs for 69 Candida albicans strains ranged from 4 to 32 μg/ml. This modest in vitro activity contrasts with the strong in vivo efficacy in C. albicans infection. This was demonstrated in a lethal model of C. albicans infection in mice and rats in which icofungipen showed dose-dependent protection at oral doses of 10 to 20 mg/kg of body weight per day in mice and 2 to 10 mg/kg/day in rats. The in vivo efficacy was also demonstrated against C. albicans isolates with low susceptibility to fluconazole, indicating activity against azole-resistant strains. The efficacy of icofungipen in mice and rats was not influenced by concomitant administration of equimolar amounts of l-isoleucine, which was shown to antagonize its antifungal activity in vitro. Icofungipen shows nearly complete oral bioavailability in a variety of species, and its in vivo efficacy indicates its potential for the oral treatment of yeast infections.

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