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Evidence for the circulation of genotype 4 Eurasian avian-like lineage swine H1N1 influenza A viruses on Korean swine farms, obtained using a newly developed one-step multiplex RT-qPCR assay

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Abstract

Genotype 4 (G4) Eurasian avian-like lineage swine H1N1 influenza A viruses, which are reassortants containing sequences from the pandemic 2009 H1N1 virus lineage, triple-reassortant-lineage internal genes, and EA-lineage external genes, have been reported in China since 2013. These have been predominant in pig populations since 2016 and have exhibited pandemic potential. In this study, we developed a one-step multiplex RT-qPCR assay targeting the M, HA1, and PB2 genes to detect G4 and related EA H1N1 viruses, with detection limits of 1.5 × 101 copies/μL and 1.15 × 10−2 ng/μL for the purified PCR products and RNA templates, respectively. The specificity of the detection method was confirmed using various influenza virus subtypes. When the one-step multiplex RT-qPCR assay was applied to swine respiratory samples collected between 2020 and 2022 in Korea, a virus related to G4 EA H1N1 strains was detected. Phylogenetic analysis based on portions of all eight genome segments showed that the positive sample contained HA, NA, PB2, NS, and NP genes closely related to those of G4 EA H1N1 viruses, confirming the ability of our assay to accurately detect G4 EA H1N1 viruses in the field.

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Data availability

The sequences generated in this study were deposited in the GenBank database under the accession numbers listed in Supplementary Table S1.

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Acknowledgements

This work was supported by the Bio & Medical Technology Development Program through the National Research Foundation of Korea (NRF), funded by the Ministry of Science & ICT (2021M3E5E3083402), and the Basic Science Research Program through the National Research Foundation of Korea (NRF), funded by the Ministry of Education (2020R1A6A1A06046235).

Funding

This article was funded by Ministry of Science and ICT, South Korea (2021M3E5E3083402), and Ministry of Education (2020R1A6A1A06046235).

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Author notes

  1. Soo Hyun Moon and Woonsung Na contributed equally to this work.

Authors and Affiliations

  1. Department of Biological Sciences and Biotechnology, College of Natural Sciences, Chungbuk National University, Cheongju, 28644, South Korea

    Soo Hyun Moon, Ji Yeong Noh, Seong Sik Jang, Min Chan Kim, Hyun A. Lim, Ha Yeon Kim, Da Young Mun & Hye Kwon Kim

  2. College of Veterinary Medicine, Chonnam National University, Gwangju, 61186, South Korea

    Woonsung Na

  3. Optipharm Animal Disease Diagnostic Center, Cheongju, 28158, South Korea

    Seongho Shin

  4. Optipharm, Inc., Cheongju, 28158, South Korea

    Hyunil Kim

  5. Infectious Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, South Korea

    Sun-Woo Yoon

  6. Department of Vaccine Biotechnology, Andong National University, Andong, 36729, South Korea

    Sun-Woo Yoon

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  1. Soo Hyun Moon
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Correspondence to Sun-Woo Yoon or Hye Kwon Kim.

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Moon, S.H., Na, W., Shin, S. et al. Evidence for the circulation of genotype 4 Eurasian avian-like lineage swine H1N1 influenza A viruses on Korean swine farms, obtained using a newly developed one-step multiplex RT-qPCR assay. Arch Virol 168, 267 (2023). https://doi.org/10.1007/s00705-023-05887-3

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