Heterothallism and sexual reproduction in the Iranian isolates of Aspergillus flavus

Ascomycota includes a large number of species that lack a known sexual stage but have a hidden potential for sexual reproduction, among which is Aspergillus flavus, an opportunistic aflatoxin-producing pathogen. The sexual stage of this heterothallic fungus results from crossing between strains with opposite mating types that belong to different vegetative compatibility groups. Here, twenty A. flavus Iranian isolates were investigated for heterothallism and sexual reproduction. The mating type genes MAT1-1 and MAT1-2 were explored using specific primers and multiplex PCR. The results indicated an equal frequency of mating types (1:1) in the investigated A. flavus isolates, indicating heterothallism, and possible sexual reproduction. The cross-cultures on Mixed Cereal Agar (MCA) and Aspergillus Complete Medium (ACM) induced the sexual phase. After 6 to 11 months, 33.16% of the crosses led to sexual reproduction and the sexual reproductive organ of the fungus appeared at the contact lines of the crosses. Scanning Electron Microscopy (SEM) analyses revealed multiple asci and ascospores, which were produced in the pseudoparenchymatous stromata. Ascospore production was more efficient in ACM than in the MCA. Our findings place the Iranian A. flavus in the genus Petromyces, as Petromyces flavus.