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

Abstract

Infections caused by Aspergillus species are being increasingly reported. Aspergillus flavus is the second most common species within this genus causing invasive infections in humans, and isolates showing azole resistance have been recently described. A. flavus has three cyp51-related genes (cyp51A, cyp51B, and cyp51C) encoding 14-α sterol demethylase-like enzymes which are the target of azole drugs. In order to study triazole drug resistance in A. flavus, three strains showing reduced azole susceptibility and 17 azole susceptible isolates were compared. The three cyp51-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 cyp51A and cyp51C 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.

Dissection of the Activity of Agricultural Fungicides against Clinical Aspergillus Isolates with and without Environmentally and Medically Induced Azole Resistance

“…The mechanisms causing triazole resistance in A. flavus are less well characterised. However, recent studies have identified a number of azole resistant isolates and documented Cyp51A, Cyp51B or Cyp51C target enzyme alterations [ 24 , 25 , 26 , 27 , 28 ], upregulation of target gene expression [ 26 , 29 ], or efflux pump upregulation [ 26 , 29 , 30 ] in the resistant isolates. Moreover, resistance has been suggested among environmental A. flavus isolates in Vietnam [ 31 ] and Argentina [ 25 ].…”

CYP51 Mutations in the Fusarium solani Species Complex: First Clue to Understand the Low Susceptibility to Azoles of the Genus Fusarium

“…Liu et al showed that a T788G mutation in the CYP51C gene was responsible for voriconazole resistance in A. flavus . However, Paul et al contradicted this conclusion because this mutation was not found in their strains and another mutation (Y319H) has been highlighted and implicated in mediating voriconazole resistance [ 29 , 30 , 31 ]. CYP51A protein sequences from our panel of FSSC strains have been compared to other complexes in the genus Fusarium .…”

Bio function of Cytochrome P450 on fungus: a review

“…In humans, Aspergillus flavus is one of genus causing invasive infections. A. flavus has CYP51A, CYP51B, and CYP51C genes which are the target of azole drugs [67]. The result of gene expression analysis showed that CYP51A and CYP51B expression levels were greater than that of CYP51C.…”