The aim of this work is to set up a SYBR-green real time qPCR method, based on the use of specie-specific primers for the early detection and quantification of potential aflatoxigenic fungi Aspergillus flavus and Aspergillus parasiticus on whole maize kernels. A primer pair was used for amplifying a 352 bp fragment of aflR, gene regulator of the aflatoxin biosynthesis gene cluster. DNA amplification was achieved only with DNA extracted from fungal strains of A. parasiticus and A. flavus and from maize kernels inoculated with A. flavus or A. parasiticus, never with DNA of the other fungal species. Amplification was evident in maize artificially inoculated with A. flavus 3357 starting from 6 hours of incubation after inoculation, when mycelium was not visible by stereomicroscope analysis as yet. This real time qPCR method could be a real, effective method for the early detection and quantification of the most important aflatoxin-producing fungi in food commodities. The method proposed in this work represents a useful tool to evaluate the quality of raw material at different critical points of the food chain. It could be used to predict potential risk for the presence of potentially aflatoxigenic strains. The combination of this approach with the more expensive and laborious conventional chemical analysis of toxin, could be a real, effective alternative respect to traditional diagnostic methods for the early detection and quantification of aflatoxigenic fungi in food commodities.
Full conference title:
7th International Aspergillus Meeting
- Asperfest 7 (2010)