Rapid detection of aflatoxin -associated genes in Aspergillus species using colony PCR based method

Abstract

Monitoring and controlling of food healthy is an indicator of scientific progress and social health in every country. Food contamination is caused by aflatoxin of saprophyte fungus associated with a number of pathological conditions including cancer and chromosomal anomalies. Aspergillus is defined as a group of fungi with various species. Identification of these fungi is of importance in terms of pathogenicity, toxicity and industry application. There are a number of laboratory methods for differentiation between Aspergillus species including physical macro and microscopic characteristics of colony. These methods are time-consuming and require highly trained personnel. In the present study, 27 samples of contaminated food and 15 samples of intact food were collected. Aspergillus flavus and Aspergillus parasiticus were selected as positive controls and examined using colony PCR reaction. All samples were re-cultured on medium culture of potato dextrose agar. Initial identification was performed based on microscopic features. Genomes of all strains were extracted using lyticase enzyme and lysis buffer containing: EDTA, Tris HCL, and NaCl. Then, aflatoxin genes of each sample were amplified through PCR. The results of colony PCR in intact food samples demonstrated that there is no used primer related to aflatoxin genes. In suspected contamination food samples, one sample with aflr and two samples with omt-1 were observed. The results of our study demonstrated that lyticase enzyme and lysis buffer are potential candidates for extraction of Aspergillus DNA.