Using collections data to infer biogeographic, environmental, and host structure in communities of endophytic fungi.

Biodiversity collections contain a wealth of information encapsulated both in specimens and in their metadata, providing the foundation for diverse studies in fields such as ecology. Yet biodiversity repositories can present a challenge for ecological inferences because collections rarely are structured with ecological questions in mind: collections may be opportunistic in space or time, may focus on particular taxonomic groups, may reflect different collection strategies in different places or times, or may not be exhaustive in terms of retaining every specimen or having similar metadata for each record. In addition to its primary holdings, the Robert L. Gilbertson Mycological Herbarium at the University of Arizona holds a collection of living specimens of fungi isolated from the interior of healthy plants and lichens (i.e., endophytic and endolichenic fungi). Over the past decade, more than 7000 isolates from the southwestern United States were accessioned, including strains from diverse hosts in more than 50 localities across the biotically rich state of Arizona. This collection is distinctive in that metadata and barcode sequences are available for each specimen, many localities have been sampled with consistent methods, and all isolates obtained in surveys have been retained. Here, we use this herbarium collection to examine endophyte community structure in an ecological and evolutionary context. We then artificially restructure the collection to resemble collections more typical of biodiversity repositories, providing a case study for ecological insights that can be gleaned from collections that were not structured explicitly to address ecological questions. Overall, our analyses highlight the relevance of biogeography, climate, hosts, and geographic separation in endophyte community composition. This study showcases the importance of extensive metadata in collections and highlights the utility of biodiversity collections that can yield emergent insights from many surveys to answer ecological questions in mycology, ultimately providing information for understanding and conserving fungal biodiversity.

Montagnuphilones A-G, Azaphilones from Montagnulaceae sp. DM0194, a Fungal Endophyte of Submerged Roots of Persicaria amphibia.

Seven azaphilones, montagnuphilones A-G (1-7), together with previously known azaphilones 8-11, were encountered in Montagnulaceae sp. DM0194, an endophytic fungus isolated from submerged roots of Persicaria amphibia. The structures of 1-7 were elucidated on the basis of their MS and NMR spectroscopic analysis. Compounds 1-8 were evaluated for their cytotoxicity and ability to inhibit nitric oxide (NO) production in lipopolysaccharide-activated RAW264.7 macrophage cells. Among these, none were found to be cytotoxic to RAW264.7 cells up to 100.0 µM, but 8, 5, and 2 showed NO inhibitory activity with IC50 values of 9.2 ± 0.9, 25.5 ± 1.1, and 39.6 ± 1.8 µM, respectively.

Fungal endophytes in aboveground tissues of desert plants: infrequent in culture, but highly diverse and distinctive symbionts.

In hot deserts, plants cope with aridity, high temperatures, and nutrient-poor soils with morphological and biochemical adaptations that encompass intimate microbial symbioses. Whereas the root microbiomes of arid-land plants have received increasing attention, factors influencing assemblages of symbionts in aboveground tissues have not been evaluated for many woody plants that flourish in desert environments. We evaluated the diversity, host affiliations, and distributions of endophytic fungi associated with photosynthetic tissues of desert trees and shrubs, focusing on nonsucculent woody plants in the species-rich Sonoran Desert. To inform our strength of inference, we evaluated the effects of two different nutrient media, incubation temperatures, and collection seasons on the apparent structure of endophyte assemblages. Analysis of >22,000 tissue segments revealed that endophytes were isolated four times more frequently from photosynthetic stems than leaves. Isolation frequency was lower than expected given the latitude of the study region and varied among species a function of sampling site and abiotic factors. However, endophytes were very species-rich and phylogenetically diverse, consistent with less arid sites of a similar latitudinal position. Community composition differed among host species, but not as a function of tissue type, sampling site, sampling month, or exposure. Estimates of abundance, diversity, and composition were not influenced by isolation medium or incubation temperature. Phylogenetic analyses of the most commonly isolated genus (Preussia) revealed multiple evolutionary origins of desert-plant endophytism and little phylogenetic structure with regard to seasonality, tissue preference, or optimal temperatures and nutrients for growth in vitro. Together, these results provide insight into endophytic symbioses in desert-plant communities and can be used to optimize strategies for capturing endophyte biodiversity at regional scales.

Fungal endophytes of aquatic macrophytes: diverse host-generalists characterized by tissue preferences and geographic structure.

Most studies of endophytic symbionts have focused on terrestrial plants, neglecting the ecologically and economically important plants present in aquatic ecosystems. We evaluated the diversity, composition, host and tissue affiliations, and geographic structure of fungal endophytes associated with common aquatic plants in lentic waters in northern Arizona, USA. Endophytes were isolated in culture from roots and photosynthetic tissues during two growing seasons. A total of 226 isolates representing 60 putative species was recovered from 9,600 plant tissue segments. Although isolation frequency was low, endophytes were phylogenetically diverse and species-rich. Comparisons among the most thoroughly sampled species and reservoirs revealed that isolation frequency and diversity did not differ significantly between collection periods, among species, among reservoirs, or as a function of depth. However, community structure differed significantly among reservoirs and tissue types. Phylogenetic analyses of a focal genus (Penicillium) corroborated estimates of species boundaries and informed community analyses, highlighting clade- and genotype-level affiliations of aquatic endophytes with both sediment- and waterborne fungi, and endophytes of proximate terrestrial plants. Together these analyses provide a first quantitative examination of endophytic associations in roots and foliage of aquatic plants and can be used to optimize survey strategies for efficiently capturing fungal biodiversity at local and regional scales.