Contributions of Mutation and Subsequent Upregulation to Voriconazole Resistance in Aspergillus Flavus

is the second most significant pathogenic cause of invasive aspergillosis; however, its emergence risks and mechanisms of voriconazole (VRC) resistance have not yet been elucidated in detail. Here, we demonstrate that repeated exposure of to subinhibitory concentrations of VRC causes the emergence of a VRC-resistant mutant with a novel resistance mechanism. The VRC-resistant mutant shows a MIC of 16 μg/ml for VRC and of 0.5 μg/ml for itraconazole (ITC). Whole-genome sequencing analysis showed that the mutant possesses a point mutation in , which encodes a bZIP transcription factor working as the master regulator of the oxidative stress response, but no mutations in the genes. This point mutation in caused alteration of Leu558 to Trp (Yap1) in the putative nuclear export sequence in the carboxy-terminal cysteine-rich domain of Yap1. This Yap1 substitution was confirmed as being responsible for the VRC-resistant phenotype, but not for that of ITC, by the revertant to Yap1 with homologous gene replacement. Furthermore, Yap1 caused marked upregulation of the ATP-binding cassette transporter, and the deletion of restored susceptibility to VRC in These findings provide new insights into VRC resistance mechanisms via a transcriptional factor mutation that is independent of the gene mutation in .