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 30-60%. Currently, 30-50% of C. auris isolates are intrinsically resistant to amphotericin B. In this work, 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 work provides evidence that a deletion resulting in a frameshift in ERG3 contributes to the observed resistant phenotype. Characterization of this isolate also revealed 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.

Figure 1:

Dilution Spot Assay of Amphotericin B Susceptible and Resistant C. auris Isolates Ten-fold dilution series of LNV001 and LNV002 were plated on RPMI agar supplemented with varying concentration of amphotericin B and reading observed every 24 hours.

Figure 2:

Amphotericin B Minimum Inhibitory Concentrations following ERG3 and ERG4 reversion to wildtype. A) Representative images of Amphotericin B Minimum Inhibitory Concentrations (MICs) at 24 hours as determined by E-tests (bioMérieux). B) MICs determined by broth microdilution in accordance with CLSI susceptibility testing. MICs were read visually for 100% growth inhibition at 24 hours. 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.

Figure 3:

Significant Growth of Amphotericin B Susceptible Isolate Compared to Resistant Isolate Biolog Phenotypic plates PM1 and PM2a were inoculated with LNV001 and LNV002. Optical density was measured every 6–8 hours for 72 hours. Blue line is LNV001 and red line is LNV002. Standard deviation is denoted by the blurring surrounding the lines. Significance determined by a student’s T-test with the Bonferroni correction applied (α = 0.000263).