Amin Daliri - Department of Mycology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
Masoomeh Shams-Ghahfarokhi - Department of Mycology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
Mehdi Razzaghi-Abyaneh - Department of Mycology, Pasteur Institute of Iran, Tehran, Iran

Background and Purpose: The current study aimed to report a multiplex polymerase chain reaction (PCR) assay as a monitoring technique to differentiate aflatoxigenic from non-aflatoxigenic strains of Aspergillus flavus isolated from pistachio orchards soil.Materials and Methods: In total, ۲۵ A. flavus strains were isolated from soil samples of pistachio orchards. To test the strains for Aflatoxin B۱(AFB۱)-producing ability, thin-layer chromatography (TLC) was used and the amounts of AFB۱ were measured by high performance liquid chromatography (HPLC). Multiplex PCR was used as a genome-based method to detect genes responsible for AFB۱ production by A. flavus and the results were analyzed in terms of speed and specificity of detection. A set of four primers was designed specifically for the omtA, omtB, ver-۱, and aflR genes which are commonly present in aflatoxin biosynthetic pathways.Results: The AFB۱ production by the A. flavus strains ranged from ۰ to ۳۲۱ ρg/μl. Four band patterns of the primer sets were observed only in AFB۱-producing A. flavus strains. Moreover, ۱۸ out of the ۲۵ strains showed all four bands belonging to omtA, omtB, ver-۱, and aflR, whereas ۷ strains did not display omtA, or aflR-related bands, in non-toxigenic and low toxin-producing A. flavus.Conclusion: The multiplex PCR is a supplementary strategy to current conventional mycotoxin analytical techniques, such as TLC and HPLC. It could be used as an efficient method to differentiate aflatoxigenic from non-aflatoxigenic strains of A. flavus. This achievement is crucial to minimize fungal contamination of food, feed, and agricultural commodities, thereby reducing the risk of subsequent aflatoxin consumption.

Aspergillus flavus, Aflatoxin B۱, Multiplex-PCR, Pistachio orchards