Short-term exposure to fluconazole induces chromosome loss in Candida albicans: an approach to produce haploid cells

Abstract

Candida albicans is considered to be an obligate diploid fungus. Here, we describe an approach to isolate aneuploids or haploids induced by the short-term (12-16 h) exposure of diploid reference strains SC5314 and CAI4 to the most commonly used antifungal drug, fluconazole, followed by repeated single-cell separation among small morphologically distinct colonies in the inhibition zone. The isolated strains had altered cell morphology and LOH events in the MTL and other marker alleles of the analyzed loci at 8 chromosomes of C. albicans with decreased DNA content. The present study employed next-generation sequencing (NGS) combined flow cytometry analysis of the DNA content to analyze the haploid, autodiploid, and aneuploid strains that arose from the fluconazole treatment instead of using the conventional single nucleotide polymorphism/comparative genome hybridization (SNP/CGH) method. A multiple-alignment tool was also developed based on sequenced data from NGS to establish haplotype mapping for each chromosome of the selected strains. These findings revealed that C. albicans experiences 'concerted chromosome loss' to form strains with homozygous alleles and that it even has a haploid status after short-term exposure to fluconazole. Additionally, we developed a new platform to analyze chromosome copy number using NGS.

Keywords: Candida albicans; Chromosome copy number; Fluconazole; Next-generation sequencing.