The T788G Mutation in the cyp51C Gene Confers Voriconazole Resistance in Aspergillus flavus Causing Aspergillosis

Abstract

With voriconazole (VRC) being approved as the first choice in treating invasive aspergillosis (IA) and its increasing use in treatment, a VRC-resistant strain of Aspergillus flavus, the second leading cause of IA after Aspergillus fumigatus, has emerged. The VRC-resistant strain of A. flavus was isolated for the first time from the surgical lung specimen of an IA patient with no response to VRC therapy. In order to ascertain the mechanism of VRC resistance, the azole target enzyme genes in this strain of A. flavus were cloned and sequenced, and 4 mutations generating amino acid residue substitutions were found in the cyp51C gene. To further determine the role of this mutated gene for VRC resistance in A. flavus, an Agrobacterium tumefaciens-mediated gene replacement approach was applied. Consequently, the mutated cyp51C gene from this A. flavus strain was proven to confer the VRC resistance. Finally, to discern the one out of the four mutations in the cyp51C gene that is responsible for contributing to VRC resistance, a site-directed gene mutagenesis procedure combined with a gene replacement method was performed. As a result, the T788G missense mutation in the cyp51C gene was identified as responsible for VRC resistance in A. flavus. These findings indicated that the detection of this mutation in A. flavus could serve as an indicator for physicians to avoid the use of VRC during IA treatment. Further comprehensive surveillance for antifungal susceptibility, as well as intensive study on the mechanism of azole resistance in A. flavus causing IA, would be required to fully understand this mechanism.