Peramine is an insect-feeding deterrent produced by Epichloë species in symbiotic association with C3 grasses. The perA gene responsible for peramine synthesis encodes a two-module nonribosomal peptide synthetase. Alleles of perA are found in most Epichloë species; however, peramine is not produced by many perA-containing Epichloë isolates. The genetic basis of these peramine-negative chemotypes is often unknown. Using PCR and DNA sequencing, we analyzed the perA genes from 72 Epichloë isolates and identified causative mutations of perA null alleles. We found nonfunctional perA-ΔR* alleles, which contain a transposon-associated deletion of the perA region encoding the C-terminal reductase domain, are widespread within the Epichloë genus and represent a prevalent mutation found in nonhybrid species. Disparate phylogenies of adjacent A2 and T2 domains indicated that the deletion of the reductase domain (R*) likely occurred once and early in the evolution of the genus, and subsequently there have been several recombinations between those domains. A number of novel point, deletion, and insertion mutations responsible for abolishing peramine production in full-length perA alleles were also identified. The regions encoding the first and second adenylation domains (A1 and A2, respectively) were common sites for such mutations. Using this information, a method was developed to predict peramine chemotypes by combining PCR product size polymorphism analysis with sequencing of the perA adenylation domains.

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Published in Applied and Environmental Microbiology, v. 81, no. 8, p. 2797-2807.

Copyright © 2014, American Society for Microbiology. All Rights Reserved.

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This research was supported by a Massey University Ph.D. scholarship and a grant from the Royal Society of New Zealand Marsden Fund (contract MAU1002). Genome sequencing was supported by United States Department of Agriculture grants 2012-67013-19384 and 2010-34457-21269, National Institutes of Health grants R01GM086888 and 2 P20 RR-16481, and The Samuel Roberts Noble Foundation.