Genetic profiling via a novel PCR-RFLP method enabled identification of four genera of anaerobic gut fungi isolated from nyala, giraffe, and zebra hosts
Abstract
Herbivore gut microbiomes may contain a diversity of anaerobic gut fungi (AGF, phylum Neocallimastigomycota), important for fibre degradation. To perform functional studies and elucidate niches of AGF species, fungal isolates must be obtained into axenic culture, which is a resource-intensive process. Here, we leverage the integration of morphological and functional assessments of AGF isolates with a newly developed Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) strategy, to prioritize isolates of interest from faecal samples from zoo-housed animals for characterization and confirmatory internal transcribed spacer (ITS) or Large Subunit (LSU) sequencing. In silico prediction of PCR-RFLP profiles of cultured genera, followed by experimental validation, confirmed that LSU-based PCR-RFLP with AluI and Hyp188I digestion effectively identified fungi of distinct genera. Together, our workflow resulted in isolation of a so far uncultured Piromyces (NY08) species and Neocallimastix cameroonii from nyala samples, as well as Feramyces austinii from giraffe and Khoyollomyces ramosus from zebra. Amplicon sequencing confirmed these species dominated AGF communities in their hosts, likely benefiting isolation success, and we identified enrichment conditions which also affected cultivability. The workflow developed here aids efficient AGF isolations, which are instrumental in expanding opportunities for functional studies that provide insight into the physiology and ecology of these fungi and help realize applications in white and green biotechnology.
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BibTeX
@article{edge2026genetic,
title = {{Genetic profiling via a novel PCR-RFLP method enabled identification of four genera of anaerobic gut fungi isolated from nyala, giraffe, and zebra hosts}},
author = {Louisa Edge and Peiru Duan and Stephane Kerangart and Alice M Buckner and Jolanda M van Munster},
journal = {FEMS Microbiology Letters},
year = {2026},
month = oct,
doi = {10.1093/femsle/fnag131},
url = {https://doi.org/10.1093/femsle/fnag131},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Genetic profiling via a novel PCR-RFLP method enabled identification of four genera of anaerobic gut fungi isolated from nyala, giraffe, and zebra hosts
AU - Louisa Edge
AU - Peiru Duan
AU - Stephane Kerangart
AU - Alice M Buckner
AU - Jolanda M van Munster
PY - 2026
DA - 2026/10/29
JO - FEMS Microbiology Letters
DO - 10.1093/femsle/fnag131
UR - https://doi.org/10.1093/femsle/fnag131
AB - Herbivore gut microbiomes may contain a diversity of anaerobic gut fungi (AGF, phylum Neocallimastigomycota), important for fibre degradation. To perform functional studies and elucidate niches of AGF species, fungal isolates must be obtained into axenic culture, which is a resource-intensive process. Here, we leverage the integration of morphological and functional assessments of AGF isolates with a newly developed Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) strategy, to prioritize isolates of interest from faecal samples from zoo-housed animals for characterization and confirmatory internal transcribed spacer (ITS) or Large Subunit (LSU) sequencing. In silico prediction of PCR-RFLP profiles of cultured genera, followed by experimental validation, confirmed that LSU-based PCR-RFLP with AluI and Hyp188I digestion effectively identified fungi of distinct genera. Together, our workflow resulted in isolation of a so far uncultured Piromyces (NY08) species and Neocallimastix cameroonii from nyala samples, as well as Feramyces austinii from giraffe and Khoyollomyces ramosus from zebra. Amplicon sequencing confirmed these species dominated AGF communities in their hosts, likely benefiting isolation success, and we identified enrichment conditions which also affected cultivability. The workflow developed here aids efficient AGF isolations, which are instrumental in expanding opportunities for functional studies that provide insight into the physiology and ecology of these fungi and help realize applications in white and green biotechnology.
SN - 0378-1097
SN - 1574-6968
ER -