Functional roles of LaeA, polyketide synthase, and glucose oxidase in the regulation of ochratoxin A biosynthesis and virulence in Aspergillus carbonarius

Aspergillus carbonarius is the major producer of ochratoxin A (OTA) among Aspergillus species, but the contribution of this secondary metabolite to fungal virulence has not been assessed. We characterized the functions and addressed the roles of three factors in the regulation of OTA synthesis and pathogenicity in A. carbonarius: LaeA, a transcriptional factor regulating the production of secondary metabolites; polyketide synthase, required for OTA biosynthesis; and glucose oxidase (GOX), regulating gluconic acid (GLA) accumulation and acidification of the host tissue during fungal growth. Deletion of laeA in A. carbonarius resulted in significantly reduced OTA production in colonized nectarines and grapes. The increment laeA mutant was unable to efficiently acidify the colonized tissue, as a direct result of diminished GLA production, leading to attenuated virulence in infected fruit compared to the wild type (WT). The designed Acpks-knockout mutant resulted in complete inhibition of OTA production in vitro and in colonized fruit. Interestingly, physiological analysis revealed that the colonization pattern of the increment Acpks mutant was similar to that of the WT strain, with high production of GLA in the colonized tissue, suggesting that OTA accumulation does not contribute to A. carbonarius pathogenicity. Disruption of the Acgox gene inactivated GLA production in A. carbonarius, and this mutant showed attenuated virulence in infected fruit compared to the WT strain. These data identify the global regulator LaeA and GOX as critical factors modulating A. carbonarius pathogenicity by controlling transcription of genes important for fungal secondary metabolism and infection.