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Fecal metatranscriptomics and virus isolation reveal picornavirus diversity and evolution in Japanese wild boars and pigs
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  • Published: 09 April 2026

Fecal metatranscriptomics and virus isolation reveal picornavirus diversity and evolution in Japanese wild boars and pigs

  • Shuntaro Mizuno1,2,
  • Hiroho Ishida1,
  • Ryou Konno1,
  • Natsuko Teshima3,
  • Tomoko Yokota3,
  • Shwe Thiri Maung Maung Khin3,4,5,
  • Hitoshi Takemae3,
  • Toru Oi6,
  • Fujiko Fukuda7,
  • Tsuneyuki Masuda8,
  • Yasuhiro Kikkawa9,
  • Tomoichiro Oka10,11,
  • Naoyuki Aihara1,
  • Takanori Shiga1,
  • Junichi Kamiie1,
  • Hironobu Murakami1,
  • Tetsuya Mizutani3,4,5,
  • Makoto Nagai1,3 &
  • …
  • Mami Oba  ORCID: orcid.org/0000-0002-4187-32171,3,4,5 

Scientific Reports , Article number:  (2026) Cite this article

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We are providing an unedited version of this manuscript to give early access to its findings. Before final publication, the manuscript will undergo further editing. Please note there may be errors present which affect the content, and all legal disclaimers apply.

Subjects

  • Evolution
  • Genetics
  • Microbiology
  • Molecular biology

Abstract

To elucidate the enteric virome and its relationship between wild boars and domestic pigs, metatranscriptomic analysis and virus isolation using a swine testis (ST) cell line were conducted on fecal samples collected from 300 wild boars and 363 pigs in Japan between 2017 and 2023. Inoculation onto ST cells induced cytopathic effects (CPEs) in samples from 47 wild boars and 178 pigs. Deep sequencing of fecal supernatants and CPE-positive culture supernatants revealed numerous viral sequences belonging to the order Picornavirales (picornaviruses; PVs), which became the focus of this study. A total of 42 (6 from wild boars and 36 from pigs) and 247 (39 from wild boars and 208 from pigs) PV sequences were obtained from fecal supernatants (wild-type; wt) and cell culture supernatants (tissue culture–adapted; tc), respectively. Among 67 porcine teschovirus (PTV) strains detected, 66 (3 from wild boars and 63 from pigs) were isolated in cell culture, except for one B1 serotype strain. The 63 porcine isolates were classified into 11 serotypes, and intertypic homologous recombination events were identified. Of the three wild boar strains, one showed high similarity to Japanese porcine strains, whereas another strain differed significantly, suggesting independent evolution. A total of 73 enterovirus G (EV-G) strains (25 from wild boars and 48 from pigs) were classified into nine genotypes. Genotypes G17 and G6 were predominant in both wild boar and pig populations, suggesting a close relationship between the two hosts. Papain-like cysteine protease (PL-CP) sequences were detected in six genotypes, and tc-type PL-CP–positive EV-G strains of genotypes G2, G8, and G18 are reported here for the first time. Although all previously reported G17 strains possess PL-CP, eight of thirteen G17 strains from wild boars in 2023 lacked PL-CP, highlighting the need to monitor future population dynamics. A total of 126 porcine sapelovirus (PSV) strains (14 from wild boars and 112 from pigs) were identified, most of which (13 and 107, respectively) were isolated in cell culture. PSV strains formed region-specific clusters, indicating geographic associations between wild boar and pig populations. Multiple genetically diverse groups coexisted within single districts, and numerous homologous recombination events were detected among them. Porcine kobuvirus (PKV) was detected in wild boars in Japan for the first time. For porcine sapovirus (SaV), genogroup GIII was the most frequently detected. The sole SaV strain identified from a wild boar clustered with genotype GXI but showed low VP1 sequence similarity to other GXI strains. The 3′-terminal region of this strain exhibited high identity to a Chinese GVII.5 strain, suggesting past cross-species transmission and homologous recombination events. These findings indicate that while some PVs in wild boars may have evolved independently, others share close genetic relationships with strains from pigs. PVs appear to be diversifying within wild boar and pig populations, with homologous recombination serving as a key driver of their genetic variability.

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Data availability

The GenBank/EMBL/DDBJ accession numbers for the sequences of the PTV, EV-G, PSV, PKV, and SaV strains determined in this study are [LC898734](https:/www.ncbi.nlm.nih.gov/nuccore/LC898734) to [LC900468](https:/www.ncbi.nlm.nih.gov/nuccore/LC900468) (Not consecutive. For details, see Supplementary Table 1.). Other datasets generated or analyzed during the current study are available from the corresponding author upon reasonable request.

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Funding

This work was supported by JSPS KAKENHI (grant number 21K05947), and JSPS Program for Forming Japan’s Peak Research Universities (J-PEAKS) (grant number JPJS00420230003).

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Authors and Affiliations

  1. School of Veterinary Medicine, Azabu University, Sagamihara, Kanagawa, 252-5201, Japan

    Shuntaro Mizuno, Hiroho Ishida, Ryou Konno, Naoyuki Aihara, Takanori Shiga, Junichi Kamiie, Hironobu Murakami, Makoto Nagai & Mami Oba

  2. Institute of Agrobiological Sciences, National Agriculture and Food Research Organization, Tsukuba, Ibaraki, 305-8517, Japan

    Shuntaro Mizuno

  3. Center for Infectious Disease Epidemiology and Prevention Research, Tokyo University of Agriculture and Technology, Fuchu, Tokyo, 183-8509, Japan

    Natsuko Teshima, Tomoko Yokota, Shwe Thiri Maung Maung Khin, Hitoshi Takemae, Tetsuya Mizutani, Makoto Nagai & Mami Oba

  4. One Health Infectious Disease Control Research Hub, Tokyo University of Agriculture and Technology, Koganei, Tokyo, 184-0012, Japan

    Shwe Thiri Maung Maung Khin, Tetsuya Mizutani & Mami Oba

  5. Advanced Research Center for One Welfare, Research Unit of One Health Infectious Disease Control, Tokyo University of Agriculture and Technology, Koganei, Tokyo, 184-0012, Japan

    Shwe Thiri Maung Maung Khin, Tetsuya Mizutani & Mami Oba

  6. Faculty of Bioresources and Environmental Science, Ishikawa Prefectural University, Nonoichi, Ishikawa, 921-8836, Japan

    Toru Oi

  7. Ishikawa Hokubu Livestock Hygiene Service Center, Nanao, Ishikawa, Japan

    Fujiko Fukuda

  8. Faculty of Veterinary Medicine, Okayama University of Science, Imabari, Ehime, 794-0085, Japan

    Tsuneyuki Masuda

  9. Global Pig Farm, Inc, Shibukawa, Gunma, 377-0052, Japan

    Yasuhiro Kikkawa

  10. Division of Biomedical Food Research, National Institute of Health Sciences, Kawasaki, Kanagawa, 210-9501, Japan

    Tomoichiro Oka

  11. Department of Virology II, National Institute of Infectious Diseases, Japan Institute for Health Security, Musashimurayama, Tokyo, 208-0011, Japan

    Tomoichiro Oka

Authors
  1. Shuntaro Mizuno
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  2. Hiroho Ishida
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  3. Ryou Konno
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  4. Natsuko Teshima
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  11. Yasuhiro Kikkawa
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  12. Tomoichiro Oka
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  13. Naoyuki Aihara
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  14. Takanori Shiga
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  15. Junichi Kamiie
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  16. Hironobu Murakami
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  17. Tetsuya Mizutani
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  18. Makoto Nagai
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  19. Mami Oba
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Contributions

S.M., T.O., J.K., T.M., M.N. and M.O. conceived the experiments, S.M., H.I., R.K., N.T., T.Y., H.T., F.F., T.M., Y.K., T.O., N.A., T.S., M.N. and M.O. conducted the experiments, S.M., H.M., M.N. and M.O. analyzed the results. All authors reviewed the manuscript.

Corresponding author

Correspondence to Mami Oba.

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Competing interests

The authors declare no competing interests.

Ethics

The present study was carried out according to the Fundamental Guidelines for Proper Conduct of Animal Experiments and Related Activities in Academic Research Institutions under the jurisdiction of the Ministry of Education, Culture, Sports, Science and Technology of Japan. This study is reported in accordance with ARRIVE guidelines (https://arriveguidelines.org). All pathogen- and animal-related experiments were reviewed and approved by the Safety Management Regulations for Pathogens Committee of Azabu University (Approval No. 21–13) and the Animal Experiment Committee of Azabu University (Approval No. 240213–4), respectively, and were conducted in accordance with the Azabu University Safety Management for Pathogens Implementation Regulations and Manual, as well as the Azabu University Animal Experiment Implementation Regulations and Manual.

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Mizuno, S., Ishida, H., Konno, R. et al. Fecal metatranscriptomics and virus isolation reveal picornavirus diversity and evolution in Japanese wild boars and pigs. Sci Rep (2026). https://doi.org/10.1038/s41598-026-45835-0

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  • Received: 05 November 2025

  • Accepted: 23 March 2026

  • Published: 09 April 2026

  • DOI: https://doi.org/10.1038/s41598-026-45835-0

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Keywords

  • Picornavirales
  • Fecal viruses
  • Wild boar
  • Pig
  • Genetic diversity
  • Homologous recombination event
  • Population dynamics
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