Detection of aflatoxigenic molds in grains by PCR.
Aflatoxins are carcinogenic metabolites produced by several members of the Aspergillus flavus group in grains and floods. Three genes, ver-1, omt-1, and apa-2, coding for key enzymes and a regulatory factor in aflatoxin biosynthesis, respectively, have been identified, and their DNA sequences have been published. In the present study, three primer pairs, each complementing the coding portion of one of the genes, were generated. DNA extracted from mycelia of five Aspergillus species, four Penicillium species, and two Fusarium species was used as PCR template for each of the primer pairs. DNA extracted from peanut, corn, and three insect species commonly found in stored grains was also tested. Positive results (DNA amplification) were achieved only with DNA of the aflatoxigenic molds Aspergillus parasiticus and A. flavus in all three primer pairs. The detection limit of the PCR was determined by using the primer pairs complementing the omt-1 and ver-1 genes. Sterile corn flour was inoculated separately with six different molds, each at several spore concentrations. Positive results were obtained only after a 24-h incubation in enriched media, with extracts of corn inoculated with A. parasiticus or A. flavus, even at the lowest spore concentration applied (10(2) spores per g). No DNA spores per g). It is concluded that genes involved in the aflatoxin biosynthetic pathway may form the basis for an accurate, sensitive, and specific detection system, using PCR, for aflatoxigenic strains in grains and foods.
Shapira
R
R
Institute of Biochemistry, Food Science, and Nutrition, Faculty of Agriculture, Hebrew University of Jerusalem, Rehovot, Israel.
Paster
N
N
Eyal
O
O
Menasherov
M
M
Mett
A
A
Salomon
R
R
D000348:Aflatoxins / Q000633:toxicity*; D001483:Base Sequence; D004271:DNA, Fungal / Q000032:analysis; D002523:Edible Grain / Q000382:microbiology*; D005658:Fungi / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D003313:Zea mays / Q000382:microbiology
D000348:Aflatoxins; D004271:DNA, Fungal