Use of a granular bioplastic formulation for carrying conidia of a non-aflatoxigenic strain of Aspergillus flavus.

Previous research demonstrated that aflatoxin contamination in corn is reduced by field application of wheat grains pre-inoculated with the non-aflatoxigenic Aspergillus flavus strain NRRL 30797. To facilitate field applications of this biocontrol isolate, a series of laboratory studies were conducted on the reliability and efficiency of replacing wheat grains with the novel bioplastic formulation Mater-Bi to serve as a carrier matrix to formulate this fungus. Mater-Bi granules were inoculated with a conidial suspension of NRRL 30797 to achieve a final cell density of approximately log 7 conidia/granule. Incubation of 20-g soil samples receiving a single Mater-Bi granule for 60-days resulted in log 4.2-5.3 propagules of A. flavus/g soil in microbiologically active and sterilized soil, respectively. Increasing the number of granules had no effect on the degree of soil colonization by the biocontrol fungus. In addition to the maintenance of rapid vegetative growth and colonization of soil samples, the bioplastic formulation was highly stable, indicating that Mater-Bi is a suitable substitute for biocontrol applications of A. flavus NRRL 30797.

Genes differentially expressed by Aspergillus flavus strains after loss of aflatoxin production by serial transfers.

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 because of the health hazard, and thus, aflatoxins are of 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 nonaflatoxigenic A. flavus strains were produced through serial transfers of aflatoxigenic parental strains. The three independent aflatoxigenic/nonaflatoxigenic pairs were compared via transcription profiling by microarray analyses. Cross comparisons identified 22 features in common between the aflatoxigenic/nonaflatoxigenic pairs. Physical mapping of the 22 features using the Aspergillus oryzae genome sequence for reference identified 16 unique genes. Aflatoxin biosynthetic and regulatory gene expression levels were not significantly different between the aflatoxigenic/nonaflatoxigenic pairs, which suggests that the inability to produce aflatoxins is not due to decreased expression of known biosynthetic or regulatory genes. Of the 16 in common genes, only one gene homologous to glutathione S-transferase genes showed higher expression in the nonaflatoxigenic progeny relative to the parental strains. This gene, named hcc, was selected for over-expression in an aflatoxigenic A. flavus strain to determine if it was directly responsible for loss of aflatoxin production. Although hcc transformants showed six- to ninefold 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.

Gene profiling for studying the mechanism of aflatoxin biosynthesis in Aspergillus flavus and A. parasiticus.

Aflatoxins are toxic and carcinogenic polyketide metabolites produced by certain fungal species, including Aspergillus flavus and A. parasiticus. Many internal and external factors, such as nutrition and environment affect aflatoxin biosynthesis; therefore, we analyzed the transcriptome of A. flavus using expressed sequence tags (ESTs) from a normalized cDNA expression library constructed from mycelia harvested under several conditions. A total of 7218 unique ESTs were identified from 26,110 sequenced cDNA clones. Functional classifications were assigned to these ESTs and genes, potentially involved in the aflatoxin contamination process, were identified. Based on this EST sequence information, a genomic DNA amplicon microarray was constructed at The Institute for Genomic Research (TIGR). To identify potential regulatory networks controlling aflatoxin contamination in food and feeds, gene expression profiles in aflatoxin-supportive media versus non-aflatoxin-supportive media were evaluated in A. flavus and A. parasiticus. Genes consistently expressed in several aflatoxin-supportive media are reported.