Description of Gopheromyces tardescens , gen. nov., sp.nov., Gigasporangiomyces pilosus, gen. nov., sp.nov., Kelyphomyces adhaerens, gen. nov., sp. nov., proposal of Testudinimycetales ord. nov. and Testudinimycetaceae fam. nov., and emended description of the order Neocallimastigales
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Anaerobic gut fungi (AGF, Neocallimastigomycota ) represent a phylum of zoospore-producing fungi inhabiting the gastrointestinal tracts of herbivores. Twenty mammalian-affiliated genera (M-AGF) and two tortoise-affiliated genera (T-AGF) have been described so far. Here, we report on three additional novel T-AGF isolates obtained from Texas and sulcata tortoises. Phylogenetic analysis using the D1-D2 regions of the large ribosomal RNA subunit (D1-D2 LSU), RNA polymerase II large subunit (RPB1), internal transcribed spacer-1 region (ITS1), and transcriptomics-enabled phylogenomic analysis clustered these strains into three distinct, deep-branching clades, closely related to previously described T-AGF genus Testudinimyces . All isolates displayed filamentous rhizoidal growth patterns and produced monoflagellated zoospores. Unique morphological characteristics included the production of elongated, thick, nucleated structures in GX isolates, the formation of thin hair-like projections on sporangial walls in SR isolates, and irregularly shaped sporangia in TM isolates. All strains grew optimally at 32-35 °C and showed distinct substrate utilization capacity (e.g., growth on pectin, chitin, galactose). LSU analyses revealed GX isolates as the first cultured representatives of tortoise-affiliated but previously uncultured lineage NY56, while SR and TM strains have not been encountered in prior culture-independent AGF surveys. We propose to accommodate these isolates in three new genera and species – Gopheromyces tardescens (GXA2), Gigasporangiomyces pilosus (SR0.6), and Kelyphomyces adhaerens (TM0.3). Further, based on the ecological, physiological, and phylogenetic distinctions between T-AGF and M-AGF, we propose to establish a new family ( Testudinimycetaceae ) to accommodate the genera Testudinimyces, Gopheromyces , Gigasporangiomyces, and Kelyphomyces , within a new order ( Testudinimycetales ), and amend the description of Neocallimastigales to circumscribe M-AGF genera only.
