Point Mutations in the 14-α Sterol Demethylase Cyp51A or Cyp51C Could Contribute to Azole Resistance in Aspergillus flavus

Abstract

Infections caused by <i>Aspergillus</i> species are being increasingly reported. <i>Aspergillus flavus</i> is the second most common species within this genus causing invasive infections in humans, and isolates showing azole resistance have been recently described. <i>A. flavus</i> has three <i>cyp</i>51-related genes (<i>cyp</i>51A, <i>cyp</i>51B, and <i>cyp</i>51C) encoding 14-α sterol demethylase-like enzymes which are the target of azole drugs. In order to study triazole drug resistance in <i>A. flavus</i>, three strains showing reduced azole susceptibility and 17 azole susceptible isolates were compared. The three <i>cyp</i>51-related genes were amplified and sequenced. A comparison of the deduced Cyp51A, Cyp51B, and Cyp51C protein sequences with other protein sequences from orthologous genes in different filamentous fungi led to a protein identity that ranged from 50% to 80%. Cyp51A and Cyp51C presented several synonymous and non-synonymous point mutations among both susceptible and non-susceptible strains. However, two amino acid mutations were present only in two resistant isolates: one strain harbored a P214L substitution in Cyp51A, and another a H349R in Cyp51C that also showed an increase of <i>cyp</i>51A and <i>cyp</i>51C gene expression compared to the susceptible strain ATCC2004304. Isolates that showed reduced in vitro susceptibility to clinical azoles exhibited a different susceptibility profile to demethylation inhibitors (DMIs). Although P214L substitution might contribute to azole resistance, the role of H349R substitution together with changes in gene expression remains unclear.