Single-cell detection of copy number changes reveals dynamic mechanisms of adaptation to antifungals in Candida albicans

Abstract

Genomic copy number changes are associated with antifungal drug resistance and virulence across diverse fungal pathogens, but the rate and dynamics of these genomic changes in the presence of antifungal drugs are unknown. Here we optimized a dual-fluorescent reporter system in the diploid pathogen Candida albicans to quantify haplotype-specific copy number variation (CNV) and loss of heterozygosity (LOH) at the single-cell level with flow cytometry. We followed the frequency and dynamics of CNV and LOH at two distinct genomic locations in the presence and absence of antifungal drugs in vitro and in a murine model of candidiasis. Copy number changes were rapid and dynamic during adaptation to fluconazole and frequently involved competing subpopulations with distinct genotypes. This study provides quantitative evidence for the rapid speed at which diverse genotypes arise and undergo dynamic population-level fluctuations during adaptation to antifungal drugs in vitro and in vivo. Development of a dual-fluorescent reporter system allows rapid quantification of diverse genomic copy number changes that arise in Candida albicans during adaptation to antifungal drugs.