Contribution of Clinically Derived Mutations inERG11to Azole Resistance in Candida albicans

Abstract

ABSTRACTInCandida albicans, theERG11gene encodes lanosterol demethylase, the target of the azole antifungals. Mutations inERG11that result in an amino acid substitution alter the abilities of the azoles to bind to and inhibit Erg11, resulting in resistance. AlthoughERG11mutations have been observed in clinical isolates, the specific contributions of individualERG11mutations to azole resistance inC. albicanshave not been widely explored. We sequencedERG11in 63 fluconazole (FLC)-resistant clinical isolates. Fifty-five isolates carried at least one mutation inERG11, and we observed 26 distinct positions in which amino acid substitutions occurred. We mapped the 26 distinct variant positions in these alleles to four regions in the predicted structure for Erg11, including its predicted catalytic site, extended fungus-specific external loop, proximal surface, and proximal surface-to-heme region. In total, 31 distinctERG11alleles were recovered, with 10ERG11alleles containing a single amino acid substitution. We then characterized 19 distinctERG11alleles by introducing them into the wild-type azole-susceptibleC. albicansSC5314 strain and testing them for susceptibilities to FLC, itraconazole (ITC), and voriconazole (VRC). The strains that were homozygous for the single amino acid substitutions Y132F, K143R, F145L, S405F, D446E, G448E, F449V, G450E, and G464S had a ≥4-fold increase in FLC MIC. The strains that were homozygous for several double amino acid substitutions had decreased azole susceptibilities beyond those conferred by any single amino acid substitution. These findings indicate that mutations inERG11are prevalent among azole-resistant clinical isolates and that most mutations result in appreciable changes in FLC and VRC susceptibilities.