Aims: Three conventional methods and a multiplex PCR procedure with a set of four primers (Quadruplex‐PCR) were used to differentiate between aflatoxin‐producing and non‐producing strains of the Aspergillus flavus group. Methods and Results: By combining sets of primers for aflR, nor‐1, ver‐1 and omt‐A genes of the aflatoxin biosynthetic pathway, Quadruplex‐PCR showed that aflatoxinogenic strains gave a quadruplet pattern, indicating the presence of all the genes involved in the aflatoxin biosynthetic pathway which encode for functional products. Non‐aflatoxinogenic strains gave varying results with one, two, three or four banding patterns. A banding pattern in three non‐aflatoxinogenic strains resulted in non‐differentiation between these and aflatoxinogenic strains. Conclusion and Significance and Impact of the Study: Because conventional methods are time‐consuming, further studies are needed to develop a rapid and objective technique that permits complete differentiation between aflatoxin‐producing and non‐producing strains of the A. flavus group.

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A Real Time PCR system (Q-PCR) for the detection and quantification ofAspergillus flavus in black pepper has been used. Thenor1 gene, a gene of the aflatoxin biosynthetic pathway, served as target sequence. The determined copy numbers of thenor1 gene were compared to the cfu numbers obtained by plate counting. There was not a direct correlation of the results with both approaches, however the tendency was the same. In general both values increased with prolonged incubation time. The differences in copy numbers of thenor1 gene to cfu numbers decreased with increasing incubation time.