Submitted to: Annual Meeting of the UNJR Panel on Toxic Microorganisms
Publication Type: Abstract Only
Publication Acceptance Date: 11/24/2006
Publication Date: N/A
Technical Abstract: A 66-kb gene cluster in toxigenic Aspergillus flavus is responsible for the synthesis of aflatoxin (AF). Isolates of A. flavus can produce either, neither or both AF and cyclopiazonic acid (CPA). We used PCR to determine whether defects are present in the AF biosynthesis gene cluster of nonaflatoxigenic A. flavus isolates collected from agricultural soils of the southern United States. Sixteen CPA-positive and 24 CPA-negative isolates were grouped into four (A to D) and five (E to I) deletion patterns by AF-gene specific primers, respectively. Using an inverse PCR technique, we resolved the sequence breakpoints of patterns C, F and I. Pattern C, found in three isolates, has one breakpoint in the cypA 3' untranslated region, which is 217 nucleotides downstream of the TGA stop codon. Another breakpoint is in the coding region of the verA gene, at the first nucleotide encoding the amino acid residue #378 of VerA. The size of the deleted region is 40 kb. Pattern F, two isolates of different vegetative compatibility groups, has a breakpoint at the #300 nucleotide upstream of the start codon of the ver1 gene, which is identical to that found in the group 2 A. oryzae, such as RIB62. This evidence shows that group 2 A. oryzae isolates may have descended from certain nonaflatoxigenic A. flavus isolates. The breakpoints of the two isolates at the other end are identical, though the position in the chromosome is not known. Pattern I consists of 18 isolates belonging to different vegetative compatibility groups. For pattern I, in which the entire AF gene cluster is deleted, the breakpoint at the hypA end is in the hexA gene of the sugar utilization gene cluster, at the first nucleotide encoding the amino acid residue #62 of HexA. The breakpoint beyond the norB gene extends to an end of the chromosome as suggested by the presence of the 12 repeats of the telomeric sequence, CCCTAATGTTGA. Deletions in the AF gene cluster among nonaflatoxigenic A. flavus isolates thus are not rare, and the patterns are diverse. Genetic drift likely is a driving force that is responsible for the loss of the entire aflatoxin gene cluster in nonaflatoxigenic A. flavus isolates when aflatoxins have lost their adaptive value in nature.