We isolated approximately 600 putative fungal endophytes from the roots of dominant grasses and forbs in a survey of five Long Term Ecological Research sites: Konza Prairie, Cedar Creek, Short Grass Steppe, Jornada Range, and H. J. Andrews Experimental Forest. Vegetation types sampled included tallgrass prairie, shortgrass steppe, arid grassland, oak savanna, and high montane meadow. Isolates were identified by restriction fragment length polymorphism (RFLP) analysis and sequencing of the ITS region within the rDNA. RFLP analysis suggested that root endophytic fungal communities were unique to each site and that shifts in endophyte community composition occurred as the growing seasons progressed. Sequence data indicated that Andropogon gerardii roots at Konza Prairie and Cedar Creek were dominated by species in the genus Periconia. Three sites were selected for seasonal comparisons of arbuscular mycorrhizal (AM) and dark septate endophyte (DSE) colonization over 2 years, H. J. Andrews, Jornada Range, and Konza Prairie. We observed significant seasonal differences in AM colonization at Jornada, year-to-year differences in AM and DSE colonization at the H. J. Andrews, and year-to-year differences in DSE colonization at Konza. DSE colonization was significantly higher in grasses than in forbs at all three sites. In contrast, there was no significant difference in arbuscular mycorrhizal colonization between grasses and forbs. In addition, presence of endophyte hyphae was lowest at 27% of root length examined in Achillea millefolium and 32% in Bromus carinatus at the highest elevation site, high montane meadows at the H. J. Andrews. Our results suggest that DSE are generally abundant and that different sites host distinct species.