Apply PCR method in detection of Aspergillus flavus in infected corn and peanut kernels

Abstract

In this research, a protocol for detection of Aspergillus flavus was developed focusing on sample treatment methods, and evaluating PCR method to detection of target on dried peanut and corn kernels. The effective protocol was produced for detection of A. flavus in stored food or feed with high specificity and high sensitivity. In general, samples were collected and prepared to ensure that they were infected by A. flavus. These infected samples were then used for sample treatment after 1-day inoculation for peanuts and 3-day inoculation for corn kernels at room temperature. Two sample treatment methods were applied in order to obtain the highest number of microorganisms infecting in the kernels and to avoid inhibitions of host’s compound or other environmental factors to the results. Direct method was better in detecting A. flavus by using PCR assay because it was time-saving, and its DNA quality and quantity were satisfied for the detection. Detection limit of PCR method on 0.5 – 1.5g of infected kernels was approximately 40 ng of isolated DNA. The sensitivity and specificity of the optimized protocol were twenty times higher than the conventional morphological method. Therefore, this protocol can be used for monitoring A. flavus on stored food or feed, especially in the detection of potential aflatoxigenic risk in stored peanut and corn kernels, even in case of low-level infections, and in a rapid, effective and high-throughput manner. Key words: Aspergillus flavus Peanuts, corn kernels Direct method, indirect method PCR DNA