MULTIGENE PROBING OF AFLATOXIGENIC MOLDS IN FOOD USING PCR AND IMMUNOLOGICAL TECHNIQUES

Aflatoxins are carcinogenic metabolites produced by several members of the Aspergillus flavus group in grains and foods. Three genes: ver-1, omt-1 and apa-2, coding for key enzymes and a regulatory factor in af latoxin biosynthesis respectively, have been identified and their DNA sequences published. In the present study, three primer pairs were gen erated, each complementing the coding portion of one of the genes. DNA extracted from mycelia of five Aspergillus species, four Penicillium species and two Fusarium species were used as PCR templates for each o f the primer pairs. DNA extracted from peanut, corn and three insect s pecies commonly found in stored grains was also tested. Positive resul ts (DNA amplification) were achieved only with DNA of the aflatoxigeni c molds A. parasiticus and A. flavus in all three primer pairs. The de tection limit of the PCR was determined using the primer pairs complem enting the omt-1 and ver-1 genes. Sterile corn flour was inoculated se parately with six different molds, each at several spore concentration s. Positive results were only obtained following a 24-h incubation in enriched media, with extracts of corn inoculated with A. parasiticus o r A. flavus, even at the lowest spore concentration applied (10(2) spo res/g). No DNA amplification was observed from corn inoculated with ot her molds, even at the highest inoculum level (10(6) spores/g).