Skip to content
1887

Journal of General Virology header logo Journal of General Virology

Volume 106, Issue 12

Impact of cell type and species on RNA replication kinetics of Seoul virus Open Access

Abstract

Hantaviruses are zoonotic, tri-segmented, negative-sense RNA viruses and a significant public health threat. Viral pathogenesis varies between host species, with rodent reservoir infection being asymptomatic and human infection resulting in severe, immune-mediated disease. Viral pathogenesis is highly dependent on virus replication efficiency since it affects the virus’s ability to evade detection and determines the magnitude of the host immune response. However, the molecular replication kinetics for hantaviruses remain poorly defined. Therefore, we developed a sense- and segment-specific quantitative real-time PCR assay and an SYBR-based RT-qPCR assay, allowing us to quantify both negative-sense genome levels and total viral RNA synthesis of the small (S), medium (M), and large (L) segments of Seoul virus (SEOV). We then measured total viral RNA and genome accumulation in reservoir rat endothelial cells (RLMVEC), non-reservoir human endothelial cells (HUVEC-C), and Vero E6 epithelial cells. We also measured the ratio of each segment released into the culture supernatant, approximating the relative packaging efficiency. We found that, while the magnitude of viral RNA differed, RNA replication kinetics were largely similar between reservoir and non-reservoir endothelial cells. However, replication and release kinetics differed between infection of endothelial and Vero cells. We also found that the S, M, and L segments were not equally abundant during viral infection or release but instead followed a trend of M>L>S. Overall, this study validates two RT-qPCR assays to measure SEOV RNA, details the accumulation and release of each viral segment and demonstrates the impact of host cell type on hantavirus replication.

  • Received:
  • Accepted:
  • Published Online:
Keyword(s): negative-stranded segmented virus , RNA replication , RT-qPCR and Seoul virus
Funding
This study was supported by the:
  • National Institute of General Medical Sciences (Award T32AI007538)
    • Principal Award Recipient: StefanD. Klimaj
  • National Institute of General Medical Sciences (Award T32AI007538)
    • Principal Award Recipient: AutumnT. LaPointe
  • Division of Microbiology and Infectious Diseases, National Institute of Allergy and Infectious Diseases (Award 1R01AI171289-01)
    • Principal Award Recipient: AlisonM Kell
© 2025 The Authors
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License. This article was made open access via a Publish and Read agreement between the Microbiology Society and the corresponding author’s institution.
Loading

Article metrics loading...

/content/journal/jgv/10.1099/jgv.0.002189
2025-12-03
2025-12-16

Metrics

Loading full text...

Full text loading...

/deliver/fulltext/jgv/106/12/jgv002189.html?itemId=/content/journal/jgv/10.1099/jgv.0.002189&mimeType=html&fmt=ahah

References

  1. Schountz T, Prescott J. Hantavirus immunology of rodent reservoirs: current status and future directions. Viruses 2014; 6:1317–1335 [View Article] [PubMed]
     [Google Scholar]
  2. Maas M, van Heteren M, de Vries A, Kuiken T, Hoornweg T et al. Seoul virus tropism and pathology in naturally infected feeder rats. Viruses 2019; 11:531 [View Article] [PubMed]
     [Google Scholar]
  3. Kell AM. Innate Immunity to orthohantaviruses: could divergent immune interactions explain host-specific disease outcomes?. J Mol Biol 2022; 434:167230 [View Article] [PubMed]
     [Google Scholar]
  4. Jonsson CB, Figueiredo LTM, Vapalahti O. A global perspective on hantavirus ecology, epidemiology, and disease. Clin Microbiol Rev 2010; 23:412–441 [View Article] [PubMed]
     [Google Scholar]
  5. Zhang Y-Z, Zou Y, Fu ZF, Plyusnin A. Hantavirus infections in humans and animals, China. Emerg Infect Dis 2010; 16:1195–1203 [View Article] [PubMed]
     [Google Scholar]
  6. Martínez VP, Di Paola N, Alonso DO, Pérez-Sautu U, Bellomo CM et al. “Super-spreaders” and person-to-person transmission of andes virus in Argentina. N Engl J Med 2020; 383:2230–2241 [View Article] [PubMed]
     [Google Scholar]
  7. Klimaj SD, LaPointe A, Martinez K, Acosta EH, Kell AM. Seoul Orthohantavirus evades innate immune activation by reservoir endothelial cells. PLoS Pathog 2024; 20:e1012728 [View Article] [PubMed]
     [Google Scholar]
  8. Valbuena G, Walker DH. The endothelium as a target for infections. Annu Rev Pathol 2006; 1:171–198 [View Article] [PubMed]
     [Google Scholar]
  9. Ermonval M, Baychelier F, Tordo N. What do we know about how hantaviruses interact with their different hosts?. Viruses 2016; 8:223 [View Article] [PubMed]
     [Google Scholar]
  10. Mackow ER, Dalrymple NA, Cimica V, Matthys V, Gorbunova E et al. Hantavirus interferon regulation and virulence determinants. Virus Res 2014; 187:65–71 [View Article] [PubMed]
     [Google Scholar]
  11. Kell AM, Hemann EA, Turnbull JB, Gale M Jr. RIG-I-like receptor activation drives type I IFN and antiviral signaling to limit Hantaan orthohantavirus replication. PLoS Pathog 2020; 16:e1008483 [View Article] [PubMed]
     [Google Scholar]
  12. LaPointe A, Gale M, Kell AM. Orthohantavirus replication in the context of innate immunity. Viruses 2023; 15:1130 [View Article]
     [Google Scholar]
  13. Easterbrook JD, Klein SL. Seoul virus enhances regulatory and reduces proinflammatory responses in male Norway rats. J Med Virol 2008; 80:1308–1318 [View Article] [PubMed]
     [Google Scholar]
  14. Strandin T, Smura T, Ahola P, Aaltonen K, Sironen T et al. Orthohantavirus isolated in reservoir host cells displays minimal genetic changes and retains wild-type infection properties. Viruses 2020; 12:457 [View Article] [PubMed]
     [Google Scholar]
  15. Li W, Klein SL. Seoul virus-infected rat lung endothelial cells and alveolar macrophages differ in their ability to support virus replication and induce regulatory T cell phenotypes. J Virol 2012; 86:11845–11855 [View Article] [PubMed]
     [Google Scholar]
  16. Garcin D, Lezzi M, Dobbs M, Elliott RM, Schmaljohn C et al. The 5’ ends of Hantaan virus (Bunyaviridae) RNAs suggest a prime-and-realign mechanism for the initiation of RNA synthesis. J Virol 1995; 69:5754–5762 [View Article] [PubMed]
     [Google Scholar]
  17. Jeeva S, Mir S, Velasquez A, Weathers BA, Leka A et al. Hantavirus RdRp requires a host cell factor for cap snatching. J Virol 2019; 93:e02088-18 [View Article] [PubMed]
     [Google Scholar]
  18. Hutchinson KL, Peters CJ, Nichol ST. Sin Nombre virus mRNA synthesis. Virology 1996; 224:139–149 [View Article] [PubMed]
     [Google Scholar]
  19. Tercero B, Terasaki K, Nakagawa K, Narayanan K, Makino S. A strand-specific real-time quantitative RT-PCR assay for distinguishing the genomic and antigenomic RNAs of Rift Valley fever phlebovirus. J Virol Methods 2019; 272:113701 [View Article] [PubMed]
     [Google Scholar]
  20. Barnard TR, Wang AB, Sagan SM. A highly sensitive strand-specific multiplex RT-qPCR assay for quantitation of Zika virus replication. J Virol Methods 2022; 307:114556 [View Article] [PubMed]
     [Google Scholar]
  21. Tercero B, Terasaki K, Narayanan K, Makino S. Mechanistic insight into the efficient packaging of antigenomic S RNA into rift valley fever virus particles. Front Cell Infect Microbiol 2023; 13:1132757 [View Article] [PubMed]
     [Google Scholar]
  22. Wigren Byström J, Näslund J, Trulsson F, Evander M, Wesula Lwande O et al. Quantification and kinetics of viral RNA transcripts produced in orthohantavirus infected cells. Virol J 2018; 15:18 [View Article] [PubMed]
     [Google Scholar]
  23. Thompson KAS, Yin J. Population dynamics of an RNA virus and its defective interfering particles in passage cultures. Virol J 2010; 7:257 [View Article] [PubMed]
     [Google Scholar]
  24. Brown CM, Bidle KD. Attenuation of virus production at high multiplicities of infection in Aureococcus anophagefferens. Virology 2014; 466–467:71–81 [View Article] [PubMed]
     [Google Scholar]
  25. Brennan JW, Sun Y. Defective viral genomes: advances in understanding their generation, function, and impact on infection outcomes. mBio 2024; 15:e0069224 [View Article] [PubMed]
     [Google Scholar]
  26. Vera-Otarola J, Solis L, Soto-Rifo R, Ricci EP, Pino K et al. The Andes hantavirus NSs protein is expressed from the viral small mRNA by a leaky scanning mechanism. J Virol 2012; 86:2176–2187 [View Article] [PubMed]
     [Google Scholar]
  27. Jääskeläinen KM, Kaukinen P, Minskaya ES, Plyusnina A, Vapalahti O et al. Tula and Puumala hantavirus NSs ORFs are functional and the products inhibit activation of the interferon-beta promoter. J Med Virol 2007; 79:1527–1536 [View Article] [PubMed]
     [Google Scholar]
  28. Binder F, Gallo G, Bendl E, Eckerle I, Ermonval M et al. Inhibition of interferon I induction by non-structural protein NSs of Puumala virus and other vole-associated orthohantaviruses: phenotypic plasticity of the protein and potential functional domains. Arch Virol 2021; 166:2999–3012 [View Article] [PubMed]
     [Google Scholar]
  29. Plyusnin A. Genetics of hantaviruses: implications to taxonomy. Arch Virol 2002; 147:665–682 [View Article] [PubMed]
     [Google Scholar]
  30. Rezelj VV, Mottram TJ, Hughes J, Elliott RM, Kohl A et al. M segment-based minigenomes and virus-like particle assays as an approach to assess the potential of tick-borne Phlebovirus genome reassortment. J Virol 2019; 93:e02068-18 [View Article] [PubMed]
     [Google Scholar]
  31. Shrivastava-Ranjan P, Jain S, Chatterjee P, Montgomery JM, Flint M et al. Development of a novel minigenome and recombinant VSV expressing Seoul hantavirus glycoprotein-based assays to identify anti-hantavirus therapeutics. Antiviral Res 2023; 214:105619 [View Article] [PubMed]
     [Google Scholar]
  32. Bermúdez-Méndez E, Katrukha EA, Spruit CM, Kortekaas J, Wichgers Schreur PJ. Visualizing the ribonucleoprotein content of single bunyavirus virions reveals more efficient genome packaging in the arthropod host. Commun Biol 2021; 4:345 [View Article] [PubMed]
     [Google Scholar]
  33. Noack D, van den Hout MCGN, Embregts CWE, van IJcken WFJ, Koopmans MPG et al. Species-specific responses during Seoul orthohantavirus infection in human and rat lung microvascular endothelial cells. PLoS Negl Trop Dis 2024; 18:e0012074 [View Article] [PubMed]
     [Google Scholar]
  34. Gavrilovskaya IN, Brown EJ, Ginsberg MH, Mackow ER. Cellular entry of hantaviruses which cause hemorrhagic fever with renal syndrome is mediated by beta3 integrins. J Virol 1999; 73:3951–3959 [View Article] [PubMed]
     [Google Scholar]
/content/journal/jgv/10.1099/jgv.0.002189
Loading
Impact of cell type and species on RNA replication kinetics of Seoul virus
J. Gen. Virol. 106, 002189 (2025); https://doi.org/10.1099/jgv.0.002189
/content/journal/jgv/10.1099/jgv.0.002189
/content/journal/jgv/10.1099/jgv.0.002189
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Most cited Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An error occurred
Approval was partially successful, following selected items could not be processed due to error