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DOE OSTI · 3563285

Full ribosomal operon sequencing of anaerobic gut fungi (phylum Neocallimastigomycota ): insights on its markers and phylogenetic resolution

Young, Diana [Bavarian State Research Center for Agriculture, Freising (Germany)]·Stüer-Patowsky, Katrin [Technical University of Munich, Garching (Germany)]·Huang, Liren [Bielefeld University (Germany)]·Elshahed, Mostafa S. [Oklahoma State University, Stillwater, OK (United States)]·Youssef, Noha H. [Oklahoma State University, Stillwater, OK (United States)]·Hanafy, Radwa [Oklahoma State University, Stillwater, OK (United States)]·Cheng, Yanfen [Nanjing Agricultural University, Jiangsu (China)]·Moon, Christina D. [Bioeconomy Science Institute, Palmerston North (New Zealand)]·Soni, Priya [Bioeconomy Science Institute, Palmerston North (New Zealand)]·Joshi, Akshay [Zürich University of Applied Sciences, Wädenswil (Switzerland)]·Stabel, Marcus [Karlsruhe Institute of Technology (Germany)]·Ochsenreither, Katrin [University of Applied Sciences Kaiserslautern, Pirmasens (Germany)]·Dagar, Sumit Singh [Agharkar Research Institute, Pune (India)]·Hillman, Ethan [University of Michigan, Ann Arbor, MI (United States)]·Solomon, Kevin V. [University of Delaware, Newark, DE (United States)]·Fliegerová, Katerina O. [Czech Academy of Sciences, Prague (Czech Republic)]·Griffith, Gareth W. [Aberystwyth University (United Kingdom)]·Callaghan, Tony M. [SomaTech Ltd. Ballybay Food Hub (Ireland)]·Podmirseg, Sabine M. [University of Innsbruck (Austria)]·Sczyrba, Alexander [Bielefeld University (Germany); Forschungszentrum Jülich (Germany)]·Flad, Veronika [Bavarian State Research Center for Agriculture, Freising (Germany)]·Lebuhn, Michael [Bavarian State Research Center for Agriculture, Freising (Germany); bioBit GmbH, Munich (Germany)]·Wurzbacher, Christian [Technical University of Munich, Garching (Germany)]
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Abstract

The phylogenetic affiliations of anaerobic gut fungi (Neocallimastigomycota) are typically evaluated using single-gene markers. However, this approach often fails to resolve relationships between closely related lineages. To address this issue and identify alternative markers, we created a curated database comprising the complete ribosomal operon sequences of 156 isolates, representing 20 of the 22 recognized genera and two new genus-level clades. Using long-read sequencing, we obtained ~9 kbp operon sequences and developed a robust analysis pipeline. Incorporating both coding genes and non-coding regions (excluding IGS1) improved phylogenetic resolution. This phylogenetic approach successfully resolved the Cyllamyces and Caecomyces clades (hard-to-distinguish genetically), as well as seven analysed Piromyces species. We also scanned the operon for markers that are suitable for short-read sequencing platforms, with the aim of enhancing biodiversity and phylogenetic studies. Notably, the ETS1 genetic region also enabled the distinction between these lineages, indicating its phylogenetic value within the ribosomal operon. The resulting database is a valuable resource for expanding and strengthening phylogenetic frameworks.

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High-throughput sequencingNanopore and PacBiophylogenyrrn markers

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Young, Diana [Bavarian State Research Center for Agriculture, Freising (Germany)], Stüer-Patowsky, Katrin [Technical University of Munich, Garching (Germany)], Huang, Liren [Bielefeld University (Germany)], Elshahed, Mostafa S. [Oklahoma State University, Stillwater, OK (United States)], Youssef, Noha H. [Oklahoma State University, Stillwater, OK (United States)], Hanafy, Radwa [Oklahoma State University, Stillwater, OK (United States)], Cheng, Yanfen [Nanjing Agricultural University, Jiangsu (China)], Moon, Christina D. [Bioeconomy Science Institute, Palmerston North (New Zealand)], Soni, Priya [Bioeconomy Science Institute, Palmerston North (New Zealand)], Joshi, Akshay [Zürich University of Applied Sciences, Wädenswil (Switzerland)], Stabel, Marcus [Karlsruhe Institute of Technology (Germany)], Ochsenreither, Katrin [University of Applied Sciences Kaiserslautern, Pirmasens (Germany)], Dagar, Sumit Singh [Agharkar Research Institute, Pune (India)], Hillman, Ethan [University of Michigan, Ann Arbor, MI (United States)], Solomon, Kevin V. [University of Delaware, Newark, DE (United States)], Fliegerová, Katerina O. [Czech Academy of Sciences, Prague (Czech Republic)], Griffith, Gareth W. [Aberystwyth University (United Kingdom)], Callaghan, Tony M. [SomaTech Ltd. Ballybay Food Hub (Ireland)], Podmirseg, Sabine M. [University of Innsbruck (Austria)], Sczyrba, Alexander [Bielefeld University (Germany); Forschungszentrum Jülich (Germany)], Flad, Veronika [Bavarian State Research Center for Agriculture, Freising (Germany)], Lebuhn, Michael [Bavarian State Research Center for Agriculture, Freising (Germany); bioBit GmbH, Munich (Germany)], Wurzbacher, Christian [Technical University of Munich, Garching (Germany)]. 2026-08-26. Full ribosomal operon sequencing of anaerobic gut fungi (phylum Neocallimastigomycota ): insights on its markers and phylogenetic resolution. https://doi.org/10.3897/imafungus.17.195921

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