Cryptic functional diversity within a grass mycobiome

PLOS ONE RESEARCH ARTICLE Cryptic functional diversity within a grass mycobiome Cedric Ndinga-Muniania ID1,2*, Nicholas Wornson3,4, Michael R. Fulcher5, Elizabeth T. Borer2, Eric W. Seabloom2, Linda Kinkel2,4, Georgiana May2 1 Plant and Microbial Biology Graduate Program, University of Minnesota, St. Paul, Minnesota, United States of America, 2 Department of Ecology, Evolution and Behavior, University of Minnesota, St. Paul, Minnesota, United States of America, 3 School of Statistics, University of Minnesota, Minneapolis, Minnesota, United States of America, 4 Department of Plant Pathology, University of Minnesota, Saint Paul, Minnesota, United States of America, 5 Foreign Disease–Weed Science Research Unit, United States Department of Agriculture, Frederick, Maryland, United States of America a1111111111 a1111111111 a1111111111 a1111111111 a1111111111 OPEN ACCESS Citation: Ndinga-Muniania C, Wornson N, Fulcher MR, Borer ET, Seabloom EW, Kinkel L, et al. (2023) Cryptic functional diversity within a grass mycobiome. PLoS ONE 18(7): e0287990. https:// doi.org/10.1371/journal.pone.0287990 Editor: Tzen-Yuh Chiang, National Cheng Kung University, TAIWAN Received: April 6, 2023 Accepted: June 17, 2023 Published: July 20, 2023 Peer Review History: PLOS recognizes the benefits of transparency in the peer review process; therefore, we enable the publication of all of the content of peer review and author responses alongside final, published articles. The editorial history of this article is available here: https://doi.org/10.1371/journal.pone.0287990 Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication. Data Availability Statement: All relevant data are within the paper and its Supporting Information files. All ITS sequences are available from the Genbank database. * Abstract Eukaryotic hosts harbor tremendously diverse microbiomes that affect host fitness and response to environmental challenges. Fungal endophytes are prominent members of plant microbiomes, but we lack information on the diversity in functional traits affecting their interactions with their host and environment. We used two culturing approaches to isolate fungal endophytes associated with the widespread, dominant prairie grass Andropogon gerardii and characterized their taxonomic diversity using rDNA barcode sequencing. A randomly chosen subset of fungi representing the diversity of each leaf was then evaluated for their use of different carbon compound resources and growth on those resources. Applying community phylogenetic analyses, we discovered that these fungal endophyte communities are comprised of phylogenetically distinct assemblages of slow- and fast-growing fungi that differ in their use and growth on differing carbon substrates. Our results demonstrate previously undescribed and cryptic functional diversity in carbon resource use and growth in fungal endophyte communities of A. gerardii. Introduction The growing understanding that microbiomes play important and diverse roles in host responses to changing environments has stimulated research in the taxonomic and functional diversity of microbiome communities [1–3]. Found in association with virtually all eukaryotic hosts, microbial symbionts maintain a wide variety of interactions with their host, ranging from beneficial to antagonistic, and affect host responses to the environment [4–8]. While next generation sequencing technologies have provided key insights into the taxonomic and functional genomic diversity of microbiomes [9–18], the processes generating and maintaining functional diversity in communities of many symbiotic organisms need further investigation. We investigated patterns of carbon resource use and growth of fungal endophytes associated with the prairie grass species, Andropogon gerardii, to better understand the evolutionary origins of taxonomic and functional diversity in this key group of plant symbiotic organisms. PLOS ONE | https://doi.org/10.1371/journal.pone.0287990 July 20, 2023 1 / 20 PLOS ONE Funding: This study was supported by a National Science Foundation (NSF) Macrosystems Biology grant (NSF-DEB 00037623) to co-PIs EB, ES, LK, GM. Support was also provided from the NSF Long Term Ecological Research (NSF-DEB-1234162 and NSF-DEB-1831944 to Cedar Creek LTER) and Research Coordination Network (NSF-DEB1042132) programs. Support to CNM was provided by the NSF-DEB, a Dissertation Fellowship from the Graduate School at University of Minnesota, and from the Graduate Program in Plant and Microbial Biology. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing interests: The authors have declared that no competing interests exist. Mycobiome functional diversity Patterns of resource use and growth largely define the ecological niche of microbiome species and inform processes structuring their communities [19–22]. For example, in saprobic communities, the most abundant fungal species on recalcitrant substrates such as lignin and hemicellulose are those able to break down these substrates as a main source of carbon [23–26]. Like saprobic communities, differing patterns of resource use by symbiotic microbes may also define different niches within the host [27] and mediate biotic interactions among co-occurring symbionts [28– 33]. For example, in plant symbiotic communities such as ectomycorrhizal fungi, the differing use of carbon and nitrogen resources by co-occurring taxa delimits differing niches, reducing competition and promoting coexistence of diverse species within these communities [34–37]. In contrast, overlap in resource use can intensify competition among co-occurring microbial symbionts, potentially contributing to lower diversity [38]. Understanding the functional diversity of resource use in microbial symbiont species will improve our understanding of how niche differences and biotic interactions affect the assembly of symbiotic microbiomes. Community phylogenetic approaches complement ecological studies as they inform the extent to which observed patterns of taxonomic and functional diversity may result from a shared evolutionary history [39, 40]. For microbial symbionts such as the fungal endophytes we study here, the endophytic trophic mode has evolved multiple times across the phylum Ascomycota [41, 42] contributing to the high level of taxonomic diversity observed in these communities [43, 44]. Although community phylogenetic studies often focus at or above the species level, adaptation to local environment and to biotic interactions may give rise to trait variation within and among closely related species and contribute to diversity [45–48]. In contrast, the constraints imposed by stressful environ (...truncated)