Symbiosis establishment and ecological effects of endohyphal bacteria on foliar fungi

Plant microbiomes are increasingly appreciated as major drivers of plant health and ecosystem services, and are of ever-greater interest for their potential in human applications. However, plant-associated microorganisms often live in complex associations in nature. Here, I characterize one of these microbial associations: the symbiosis between foliar fungal endophytes and their bacterial endosymbionts (endohyphal bacteria, EHB). EHB influence fungal phenotypes and can shape the outcomes of plant-fungal interactions. EHB are thought to form facultative associations with many foliar fungi in the species-rich Ascomycota, but little is known about how these symbioses are initiated and maintained, or how EHB shape the ecology of their fungal hosts. In this study, I assessed factors mediating the relationships between two foliar fungi (Microdiplodia sp., Dothideomycetes; Pestalotiopsis sp. Sordariomycetes) and their EHB. I first established methods for introducing EHB into axenic mycelia of their fungal hosts, providing an important step forward for understanding the establishment of EHB associations and a critical tool for experimental tests of the effects of EHB on fungal phenotypes. Through experiments in vitro, I found that both the identity of the fungal host and the conditions under which fungi and bacteria are grown influence the establishment of EHB/fungal associations. Moreover, I showed EHB of foliar fungi can be transferred across fungal classes, thus creating experimental strains that could be used for the first time to examine the contribution of each symbiont to important fungal traits. Using these strains I evaluated how EHB influence the capacity of foliar fungi to degrade plant material as saprotrophs. I found that the presence and identity of EHB significantly influenced fungal growth on particular media, cellulase and ligninase activity, and mass loss from senescent tissue of their native host plant species in a partnership-specific manner. Because EHB can be acquired horizontally, they may help shape plant-fungal interactions, resultant ecosystem services, and the functional diversification of plant-associated fungi along the saprotroph-endophyte continuum. By manipulating EHB/fungal interactions in new ways, we can potentially influence fungal phenotypes for diverse human applications