Acquired amphotericin B resistance attributed to a mutated ERG3 in Candidozyma auris

Abstract

First identified in 2009, Candidozyma auris (formerly Candida auris) is an emerging multidrug-resistant fungus that can cause invasive infections with a crude mortality rate ranging from 30 to 60%. Currently, 30-50% of C. auris isolates are intrinsically resistant to amphotericin B. In this study, we characterized a clinical case of acquired amphotericin B resistance using whole-genome sequencing, a large-scale phenotypic screen, comprehensive sterol profiling, and genotypic reversion using CRISPR. Data obtained in this study provide evidence that a deletion resulting in a frameshift in ERG3 significantly contributes to the observed resistant phenotype, and a nonsense mutation in ERG4 may more modestly contribute to resistance. Characterization of this isolate also revealed that a fitness cost is associated with the abrogation of ergosterol production and its replacement with other late-stage sterols. This article presents a clinical case description of amphotericin B resistance from a frameshift mutation in ERG3 in C. auris and marks an advancement in the understanding of antifungal resistance in this fungal pathogen.

Keywords: Candida auris; ERG3; amphotericin B; genetics; microbial; multidrug resistance; public health.

Fig 1

Dilution spot assay of amphotericin B-susceptible and resistant C. auris isolates. A ten-fold dilution series of LNV001 and LNV002 were plated on RPMI agar supplemented with varying concentrations of amphotericin B, and reading was observed every 24 h.

Fig 2

Amphotericin B MICs following ERG3 and ERG4 reversion to wild type. (A) Representative images of amphotericin B MICs at 24 h as determined by Etest (bioMérieux). (B) MICs determined by BMD in accordance with CLSI susceptibility testing. MICs were read visually for 100% growth inhibition at 24 h. Bars represent the modal MIC with points plotted for three biological replicates for each isolate and independent strain with MIC values for two independently derived LNV002_ERG3^WT and LNV002_ERG3^WT, ERG4^WT strains shown.

Fig 3

Significant growth of amphotericin B-susceptible isolate compared with resistant isolate Biolog Phenotypic plates PM1 and PM2a were inoculated with LNV001 and LNV002. Optical density was measured every 6–8 h for 72 h. The blue line represents LNV001, and the red line represents LNV002. Standard deviation is denoted by the blurring surrounding the lines. Significance was determined by a student’s t-test with the Bonferroni correction applied (α = 0.000263).