Insight into the Significance of Aspergillus fumigatus cyp51A Polymorphisms.

Abstract

Triazole antifungal compounds are the first treatment choice for invasive aspergillosis. However, in the last decade the rate of azole resistance among Aspergillus fumigatus strains has increased notoriously. The main resistance mechanisms are well defined and mostly related to point mutations of the azole target, 14-α sterol demethylase (cyp51A), with or without tandem repeat integrations in the cyp51A promoter. Furthermore, different combinations of five Cyp51A mutations (F46Y, M172V, N248T, D255E, and E427K) have been reported worldwide in about 10% of all A. fumigatus isolates tested. The azole susceptibility profile of these strains shows elevated azole MICs, although on the basis of the azole susceptibility breakpoints, these strains are not considered azole resistant. The purpose of the study was to determine whether these cyp51A polymorphisms (single nucleotide polymorphisms [SNPs]) are responsible for the azole susceptibility profile and whether they are reflected in a poorer azole treatment response in vivo that could compromise patient treatment and outcome. A mutant with a cyp51A deletion was generated and became fully susceptible to all azoles tested. Also, three cyp51A gene constructions with different combinations of SNPs were generated and reintroduced into an azole-susceptible wild-type (WT) strain (the ΔakuBKU80 strain). The alternative model host Galleria mellonella was used to compare the virulence and voriconazole response of G. mellonella larvae infected with A. fumigatus strains with WT cyp51A or cyp51A with SNPs. All strains were pathogenic in G. mellonella larvae, although they did not respond similarly to voriconazole therapeutic doses. Finally, the full genomes of these strains were sequenced and analyzed in comparison with those of A. fumigatus WT strains, revealing that they belong to different strain clusters or lineages.