Submitted to: Applied Microbiology and Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/24/2007
Publication Date: 10/23/2007
Citation: Chang, P., Wilkinson, J.R., Horn, B.W., Yu, J., Bhatnagar, D., Cleveland, T.E. 2007. Genes differentially expressed by Aspergillus flavus strains after loss of aflatoxin production by serial transfers. Applied Microbiology and Biotechnology. 77(4):917-925. Interpretive Summary: Aflatoxins are toxic and carcinogenic compounds produced by several molds. Aflatoxins pose a great risk to human and animal health and have caused significant economic loss due to rejection of contaminated agricultural commodities. Aspergillus flavus is a toxin-producing mold, but tends to lose the toxin-producing ability under laboratory conditions. A. flavus isolates collected in many parts of the world vary considerably in their ability to produce aflatoxins. To better understand the molecular mechanisms that regulate aflatoxin production, we used a whole-genome approach to examine the differences in genes expressed by toxigenic A. flavus parental strains and isogenic non-toxigenic progenies which were generated by serial transfers of the parental strains in rich medium. We found that the loss of aflatoxin production was not related to changes in aflatoxin biosynthesis gene expression and that more than a dozen of differentially expressed genes were associated with the loss of aflatoxin production. Some of the genes were located in very close physical proximity and likely subject to similar levels of regulation or selection. DNA microarray technology can identify gene targets for devising strategies to control aflatoxin contamination. Information obtained through fungal genomics could help in solving aflatoxin contamination in food and feed.
Technical Abstract: Aflatoxins are carcinogenic fungal secondary metabolites produced by Aspergillus flavus and other closely related species. Levels of aflatoxins in agricultural commodities are stringently regulated by many countries and thus aflatoxins are a major concern to both producers and consumers. A cluster of genes responsible for aflatoxin biosynthesis has been identified; however, expression of these genes is a complex and poorly understood phenomenon. To better understand the molecular events that are associated with aflatoxin production, three separate non-toxic A. flavus were produced through serial transfers. The three independent toxic/non-toxic pairs were compared via transcription profiling by microarray analyses. Cross comparison of these three experiments identified twenty-two features in common between the toxic/non-toxic pairs. Physical mapping of the twenty-two features using the A. oryzae genome sequence for reference identified sixteen unique genes. Aflatoxin biosynthetic and regulatory gene expression levels were not significantly different between the toxic/non-toxic pairs, which suggest that the inability to produce aflatoxins is not due to decreased expression of known biosynthetic or regulatory genes. Of the sixteen in common genes, only one, homologous to glutathione S-transferease genes, showed higher expression in the progeny relative to the parental strains. This gene, named hcc, was selected for over-expression in a non-toxigenic A. flavus to determine if it was directly responsible for loss of aflatoxin production. Though hcc transformants showed six- to nine-fold increase in expression, no discernible changes in colony morphology or aflatoxin production were detected. Possible roles of hcc and other identified genes are discussed in relation to regulation of aflatoxin biosynthesis.