SUMMARY The gray leaf spot disease of perennial ryegrass and tall fescue is caused by the fungus Magnaporthe oryzae (anamorph = Pyricularia oryzae). A collection of single-copy and repetitive DNA markers was used to investigate genetic diversity among 22 isolates of the gray leaf spot pathogen. The single-copy DNA markers revealed only three polymorphisms among 95 restriction fragments spanning approximately 0.6% of the genome. In addition, Southern hybridization analysis and mating tests revealed that all isolates possessed the MAT1-2 mating-type allele. Fingerprinting of repetitive DNA loci using the Pot2 and MGR583 probes also revealed a high degree of genetic similarity (> 85%) among isolates. These data are consistent with the gray leaf spot pathogens having a recent evolutionary origin. In contrast to the results obtained with probes for internal chromosome loci, a telomere probe revealed that the chromosome ends of the very same isolates are highly divergent, with most isolates sharing less than 20% fingerprint similarity with any other isolate. Telomere mutations arise extremely frequently and changes in telomere fingerprint profiles were readily observed during vegetative growth and among cultures derived from single spores isolated from agar medium and from lesions on perennial ryegrass leaves.