Fungal endophytes and fungi that live within healthy plant tissues without causing disease are known from all major lineages of land plants. These diverse but little-known symbionts are united by the absence of disease symptoms on their hosts during a significant phase of the infection cycle, and in many cases have beneficial effects on the plants they inhabit (e.g., pest deterrence). Recent work has shown that many plant-associated fungi are inhabited by bacterial endosymbionts, which can influence pathogenicity of those fungi. Endophytes are closely related to plant pathogens, but the evolutionary, ecological, or genetic factors that may predispose them to pathogenicity or avirulence are unknown. In preliminary studies, bacterial endosymbionts were shown to be present in apparently pure cultures of endophytic fungi, but the diversity, taxon composition, and ecological roles of these bacteria have never been explored. The proposed work will take advantage of a unique library of over 8000 living endophyte cultures to (1) determine the frequency with which bacterial endosymbionts occur among endophytic fungi; (2) compare the diversity, geographic distribution, and host specificity of endosymbiotic bacteria with those of endophytic fungi; and (3) experimentally assess the ecological roles of bacterial endosymbionts in endophytic and related plant-pathogenic fungi. This project will shed light on the ways in which bacteria mediate virulence in fungi, providing an important tool in the study of fungal-plant interactions. The intellectual merit of this study lies in disentangling a newly discovered level of complexity in plant-microbe symbioses, and in capturing the diversity of endosymbionts that themselves live within a hyperdivese guild of symbiotic microbes. This project will contribute to the training of one to three undergraduates and one graduate student, with a special focus on training individuals from groups that are traditionally under-represented in the sciences.