Large Scale Screening of Epichloë Endophytes Infecting Schedonorus pratensis and Other Forage Grasses Reveals a Relation Between Microsatellite-Based Haplotypes and Loline Alkaloid Levels

Species belonging to the Festuca-Lolium complex are often naturally infected with endophytic fungi of genus Epichloë. Recent studies on endophytes have shown the beneficial roles of host-endophyte associations as protection against insect herbivores in agriculturally important grasses. However, large-scale screenings are crucial to identify animal friendly strains suitable for agricultural use. In this study we analyzed collected populations of meadow fescue (Schedonorus pratensis) from 135 different locations across Europe, 255 accessions from the United States Department of Agriculture and 96 accessions from The Nordic Genetic Resource Centre. The analysis also included representatives of S. arundinaceus, S. giganteus, and Lolium perenne. All plants were screened for the presence of Epichloë endophytes, resulting in a nursery of about 2500 infected plants from 176 different locations. Genetic diversity was investigated on 250 isolates using a microsatellite-based PCR fingerprinting assay at 7 loci, 5 of which were uncharacterized for these species. Phylogenetic and principal components analysis showed a strong interspecific genetic differentiation among isolates, and, with E. uncinata isolates, a small but significant correlation between genetic diversity and geographical effect (r = 0.227) was detected. Concentrations of loline alkaloids were measured in 218 infected meadow fescue plants. Average amount of total loline and the proportions of the single loline alkaloids differed significantly among endophyte haplotypes (P < 0.005). This study provides insight into endophyte genetic diversity and geographic variation in Europe and a reference database of allele sizes for fast discrimination of isolates. We also discuss the possibility of multiple hybridization events as a source of genetic and alkaloid variation observed in E. uncinata.