Loss of C-5 Sterol Desaturase Activity Results in Increased Resistance to Azole and Echinocandin Antifungals in a Clinical Isolate of Candida parapsilosis

Rybak, Jeffrey M.; Dickens, C. Michael; Parker, Josie E.; Caudle, Kelly E.; Manigaba, Kayihura; Whaley, Sarah G.; Nishimoto, Andrew T.; Luna-Tapia, Arturo; Roy, Sujoy; Zhang, Qing; Barker, Katherine S.; Palmer, Glen E.; Sutter, Thomas R.; Homayouni, Ramin; Wiederhold, Nathan P.; Kelly, Steven L.; Rogers, P. David

doi:10.1128/aac.00651-17

Cited by 47 publications

(37 citation statements)

References 34 publications

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“…To assay for other changes to the ergosterol biosynthesis pathway which may have been contributing to fluconazole resistance, each of the fluconazoleevolved strains and the parental AR0387 were subsequently subjected to comprehensive sterol profiling. Briefly, each strain was grown to the exponential-growth phase in RPMI liquid media with or without 16 mg/liter of fluconazole (a concentration approximating the average serum concentration achieved in patients being treated for candidemia) (22,23).…”

Section: Resultsmentioning

confidence: 99%

Mutations in TAC1B : a Novel Genetic Determinant of Clinical Fluconazole Resistance in Candida auris

Rybak

Muñoz

Barker

et al. 2020

mBio

Self Cite

135

151

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Candida auris has emerged as a multidrug-resistant pathogen of great clinical concern. Approximately 90% of clinical C. auris isolates are resistant to fluconazole, the most commonly prescribed antifungal agent, and yet it remains unknown what mechanisms underpin this fluconazole resistance. To identify novel mechanisms contributing to fluconazole resistance in C. auris, fluconazole-susceptible C. auris clinical isolate AR0387 was passaged in media supplemented with fluconazole to generate derivative strains which had acquired increased fluconazole resistance in vitro. Comparative analyses of comprehensive sterol profiles, [3H]fluconazole uptake, sequencing of C. auris genes homologous to genes known to contribute to fluconazole resistance in other species of Candida, and relative expression levels of C. auris ERG11, CDR1, and MDR1 were performed. All fluconazole-evolved derivative strains were found to have acquired mutations in the zinc-cluster transcription factor-encoding gene TAC1B and to show a corresponding increase in CDR1 expression relative to the parental clinical isolate, AR0387. Mutations in TAC1B were also identified in a set of 304 globally distributed C. auris clinical isolates representing each of the four major clades. Introduction of the most common mutation found among fluconazole-resistant clinical isolates of C. auris into fluconazole-susceptible isolate AR0387 was confirmed to increase fluconazole resistance by 8-fold, and the correction of the same mutation in a fluconazole-resistant isolate, AR0390, decreased fluconazole MIC by 16-fold. Taken together, these data demonstrate that C. auris can rapidly acquire resistance to fluconazole in vitro and that mutations in TAC1B significantly contribute to clinical fluconazole resistance. IMPORTANCE Candida auris is an emerging multidrug-resistant pathogen of global concern, known to be responsible for outbreaks on six continents and to be commonly resistant to antifungals. While the vast majority of clinical C. auris isolates are highly resistant to fluconazole, an essential part of the available antifungal arsenal, very little is known about the mechanisms contributing to resistance. In this work, we show that mutations in the transcription factor TAC1B significantly contribute to clinical fluconazole resistance. These studies demonstrated that mutations in TAC1B can arise rapidly in vitro upon exposure to fluconazole and that a multitude of resistance-associated TAC1B mutations are present among the majority of fluconazole-resistant C. auris isolates from a global collection and appear specific to a subset of lineages or clades. Thus, identification of this novel genetic determinant of resistance significantly adds to the understanding of clinical antifungal resistance in C. auris.

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Section: Resultsmentioning

confidence: 99%

Mutations in TAC1B : a Novel Genetic Determinant of Clinical Fluconazole Resistance in Candida auris

Rybak

Muñoz

Barker

et al. 2020

mBio

Self Cite

135

151

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“…Interestingly, resistance to these agents can be linked to single pathways. For example, a recent publication reveals that C-5 desaturase activity regulates susceptibility to both azoles and echinocandins, as disruption of ERG3 leads to high-level azole and intermediate-to high-level echinocandin resistance (319).…”

Section: Antifungal Susceptibility and Resistancementioning

confidence: 99%

Candida parapsilosis: from Genes to the Bedside

et al. 2019

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SUMMARY Patients with suppressed immunity are at the highest risk for hospital-acquired infections. Among these, invasive candidiasis is the most prevalent systemic fungal nosocomial infection. Over recent decades, the combined prevalence of non-albicans Candida species outranked Candida albicans infections in several geographical regions worldwide, highlighting the need to understand their pathobiology in order to develop effective treatment and to prevent future outbreaks. Candida parapsilosis is the second or third most frequently isolated Candida species from patients. Besides being highly prevalent, its biology differs markedly from that of C. albicans, which may be associated with C. parapsilosis’ increased incidence. Differences in virulence, regulatory and antifungal drug resistance mechanisms, and the patient groups at risk indicate that conclusions drawn from C. albicans pathobiology cannot be simply extrapolated to C. parapsilosis. Such species-specific characteristics may also influence their recognition and elimination by the host and the efficacy of antifungal drugs. Due to the availability of high-throughput, state-of-the-art experimental tools and molecular genetic methods adapted to C. parapsilosis, genome and transcriptome studies are now available that greatly contribute to our understanding of what makes this species a threat. In this review, we summarize 10 years of findings on C. parapsilosis pathogenesis, including the species’ genetic properties, transcriptome studies, host responses, and molecular mechanisms of virulence. Antifungal susceptibility studies and clinician perspectives are discussed. We also present regional incidence reports in order to provide an updated worldwide epidemiology summary.

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“…Briefly, each strain was grown to exponential growth phase in RPMI liquid media with or without 16mg/L of fluconazole (a concentration approximating the average serum concentration achieved in patients being treated for candidemia). (23)…”

Section: Resultsmentioning

confidence: 99%

“…A vacuum centrifuge (Heto) was used to dry samples, which were then derivatized by adding 100 µl 90% N,O-bis(trimethylsilyl)-trifluoroacetamide–10% tetramethylsilane (TMS) (Sigma) and 200 µl anhydrous pyridine (Sigma) while heating at 80°C for 2 hours as previously described. (23, 29) GC-MS (with a Thermo 1300 GC coupled to a Thermo ISQ mass spectrometer; Thermo Scientific) was used to analyze and identify TMS-derivatized sterols through comparison of the retention times and fragmentation spectra for known standards. Sterol profiles for each sample were determined by analyzing the integrated peak areas from GC-MS data files using Xcalibur software (Thermo Scientific).…”

Section: Methodsmentioning

confidence: 99%

Mutations inTAC1B: a novel genetic determinant of clinical fluconazole resistance inC. auris

Rybak

Muñoz

Barker

et al. 2020

Preprint

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View full text Add to dashboard Cite

Candida auris has emerged as a multidrug-resistant pathogen of great clinical concern. Approximately 90% of clinical C. auris isolates are resistant to fluconazole, the most commonly prescribed antifungal agent, yet it remains unknown what mechanisms underpin this fluconazole resistance. To identify novel mechanisms contributing to fluconazole resistance in C. auris, the fluconazole-susceptible C. auris clinical isolate AR0387 was passaged in media supplemented with fluconazole to generate derivative strains which had acquired increased fluconazole resistance in vitro. Comparative analysis of comprehensive sterol profiles, [3H]-fluconazole uptake, sequencing of C. auris genes homologous to genes known to contribute to fluconazole resistance in other species of Candida, and the relative expression of C. auris ERG11, CDR1, and MDR1 were performed. All fluconazole-evolved derivative strains were found to have acquired mutations in the zinc-cluster transcription factor-encoding gene, TAC1B, and a corresponding increase in CDR1 expression relative to the parental clinical isolate, AR0387. Mutations in TAC1B were also identified in a set of 304 globally distributed C. auris clinical isolates representing each of the four major clades. Introduction of the most common mutation found among fluconazole-resistant clinical isolates of C. auris into the fluconazole-susceptible isolate AR0387, was confirmed to increase fluconazole resistance by 8-fold, and the correction of the same mutation in a fluconazole-resistant isolate, AR0390, decreased fluconazole MIC by 16-fold. Taken together, these data demonstrate that C. auris can rapidly acquire resistance to fluconazole in-vitro, and that mutations in TAC1B significantly contribute to clinical fluconazole resistance.IMPORTANCECandida auris is an emerging multidrug-resistant pathogen of global concern, known to be responsible for outbreaks on six continents and commonly resistant to antifungals. While the vast majority of clinical C. auris isolates are highly resistant to fluconazole, an essential part of the available antifungal arsenal, very little is known about the mechanisms contributing to resistance. In this work, we show that mutations in the transcription factor TAC1B significantly contribute to clinical fluconazole resistance. These studies demonstrate that mutations in TAC1B can arise rapidly in vitro upon exposure to fluconazole, and that a multitude of resistance-associated TAC1B mutations are present among the majority of fluconazole-resistant C. auris isolates from a global collection and appear specific to a subset of lineages or clades. Thus, identification of this novel genetic determinant of resistance significantly adds to the understanding of clinical antifungal resistance in C. auris.

show abstract

Loss of C-5 Sterol Desaturase Activity Results in Increased Resistance to Azole and Echinocandin Antifungals in a Clinical Isolate of Candida parapsilosis

Cited by 47 publications

References 34 publications

Mutations in TAC1B : a Novel Genetic Determinant of Clinical Fluconazole Resistance in Candida auris

Mutations in TAC1B : a Novel Genetic Determinant of Clinical Fluconazole Resistance in Candida auris

Candida parapsilosis: from Genes to the Bedside

Mutations inTAC1B: a novel genetic determinant of clinical fluconazole resistance inC. auris

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