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Two genes atmG and ggsA are cloned from Aspergillus flavus using degenerate primers for conserved domains of fungal geranylgeranyl diphosphate synthases. The study broadens ones understanding of indole-diterpene metabolite biosynthesis in filamentous fungi and provides the first genetic evidence for the biosynthetic pathway of aflatrem in Aspergillus flavus.

In Aspergillus flavus and Aspergillus oryzae the genes for aflatrem biosynthesis have clustered at two discrete loci and reverse transcriptase PCR in A flavus has shown that aflatrem biosynthesis transcript levels have increased with the onset of aflatrem production. A scheme is proposed for aflatrem biosynthesis in A. flavus based on the reconstitution experiments in Penicillium paxilli and identification of putative intermediates in wild-type cultures of A. flavus.

A PCR and Southern analysis method was used to screen a broad taxonomic range of 44 endophyte isolates to determine the differences related to indole-diterpenes which are present in few endophyte-grass association compared to that of the other bioprotective alkaloids, which are more widespread among the endophytes. Analysis of toxin biosynthesis genes by PCR allows a diagnostic method to screen endopytes for both beneficial and detrimental alkaloids and could be applied as a resource for screening isolates required for forage improvement.

Cloning and analysis of a nonribosomal peptide synthetase (NRPS) gene from Neotyphodium lolii has disclosed a putative gene cluster for the biosynthesis of ergovaline containing a single-module NRPS gene, IpsB and other genes. A genetic foundation is provided for describing biochemical steps in the ergovaline pathways, the ecological role of individual ergot alkaloid compounds and the regulation of their synthesis in planta.

Clavicipitaceous fungal endophytes of the genera Epichloë and Neotyphodium form symbioses with grasses of the subfamily Pooideae, in which they can synthesize an array of bioprotective alkaloids. Some strains produce the ergopeptine alkaloid ergovaline, which is implicated in livestock toxicoses caused by ingestion of endophyte-infected grasses. Cloning and analysis of a nonribosomal peptide synthetase (NRPS) gene from Neotyphodium lolii revealed a putative gene cluster for ergovaline biosynthesis containing a single-module NRPS gene, lpsB, and other genes orthologous to genes in the ergopeptine gene cluster of Claviceps purpurea and the clavine cluster of Aspergillus fumigatus. Despite conservation of gene sequence, gene order is substantially different between the N. lolii, C. purpurea, and A. fumigatus ergot alkaloid gene clusters. Southern analysis indicated that the N. lolii cluster was linked with previously identified ergovaline biosynthetic genes dmaW and lpsA. The ergovaline genes are closely associated with transposon relics, including retrotransposons and autonomous and nonautonomous DNA transposons. All genes in the cluster were highly expressed in planta, but expression was very low or undetectable in mycelia from axenic culture. This work provides a genetic foundation for elucidating biochemical steps in the ergovaline pathway, the ecological role of individual ergot alkaloid compounds, and the regulation of their synthesis in planta. [ABSTRACT FROM AUTHOR]