Prevalence of aflatoxigenic fungi in the animal feed produces aflatoxins in the feed posing serious health complications. Aflatoxins are toxic, carcinogenic and mutagenic for livestock and human. In the current study, animal feed was evaluated for the presence of aflatoxigenic filamentous fungi and the isolates were subjected to identification using microbiological, biochemical and molecular biology methods. Based on the DNA sequences of Internally Transcribed Spacer (ITS) regions, the isolates were recognized as Aspergillus unguis, A. niger and A. flavus. Phylogenetic relationship was deduced by constructing a phylogenetic tree using Neighbor-Joining method. Biochemical characterization revealed that isolated Aspergillus flavus is aflatoxigenic. The genes responsible for the aflatoxins biosynthesis in A. flavus were amplified using polymerase chain reaction. The genome of A. flavus was shown to harbor the aflatoxin synthesizing genes namely aflO, aflR, aflS, aflP, aflD, aflM and aflQ. The amplicons were sequenced and submitted to GenBank at NCBI. It was observed that A. flavus was the only aflatoxigenic species responsible for feed contamination in collected samples and all the seven targeted aflatoxin biosynthesis genes were found in the studied aflatoxigenic species. The sequencing of ITS regions has shown to be a reliable tool for the identification of atoxigenic and toxigenic fungal species. These findings on gene cluster variability may be useful for better understanding of genetic control as well as toxicological risks of aflatoxins.

To Cite This Article: Usman M, Javed MR, Mehmood MA, Huma T and Ijaz A, 2019. Isolation of aflatoxigenic Aspergillus flavus from animal-feed and exploration of the genetic basis of aflatoxin biosynthesis. Pak Vet J, 39(4): 541-547. http://dx.doi.org/10.29261/pakvetj/2019.078