Skip to content
1887

Abstract

Species A rotaviruses (RVs), which belong to the family and contain a genome of 11 segmented dsRNA segments, are a leading cause of severe acute gastroenteritis in infants and children younger than 5 years of age. We previously developed a strategy to recover rotavirus vaccine strain LLR from 11 cloned plasmids. Here, we report an improved reverse genetics system for LLR by combining two or three transcriptional cassettes in a single plasmid, which substantially enhances rescue efficiency from 66.7% (8/12) to 91.7 % (11/12). Furthermore, the recombinant LLR stably expressing NLuc was rescued based on the five-plasmid reverse genetics system. Improvements to the rotavirus reverse genetics system will enhance its applicability for studies of rotavirus biology and clinical use.

Funding
This study was supported by the:
  • the Postdoctoral Fellowship Program (Grade B) of China Postdoctoral Science Foundation (Award GZB20240207)
    • Principle Award Recipient: XiafeiLiu
  • the National Natural Science Foundation of China (Award 82402625)
    • Principle Award Recipient: XiafeiLiu
Loading

Article metrics loading...

/content/journal/jgv/10.1099/jgv.0.002064
2025-01-10
2025-01-18
Loading full text...

Full text loading...

References

  1. Desselberger U. Rotaviruses. Virus Res 2014; 190:75–96 [View Article] [PubMed]
    [Google Scholar]
  2. Komoto S, Sasaki J, Taniguchi K. Reverse genetics system for introduction of site-specific mutations into the double-stranded RNA genome of infectious rotavirus. Proc Natl Acad Sci USA 2006; 103:4646–4651 [View Article] [PubMed]
    [Google Scholar]
  3. Kanai Y, Komoto S, Kawagishi T, Nouda R, Nagasawa N et al. Entirely plasmid-based reverse genetics system for rotaviruses. Proc Natl Acad Sci USA 2017; 114:2349–2354 [View Article] [PubMed]
    [Google Scholar]
  4. Kobayashi T, Antar AAR, Boehme KW, Danthi P, Eby EA et al. A plasmid-based reverse genetics system for animal double-stranded RNA viruses. Cell Host Microbe 2007; 1:147–157 [View Article] [PubMed]
    [Google Scholar]
  5. Komoto S, Fukuda S, Ide T, Ito N, Sugiyama M et al. Generation of recombinant rotaviruses expressing fluorescent proteins by using an optimized reverse genetics system. J Virol 2018; 92:e00588–00518 [View Article] [PubMed]
    [Google Scholar]
  6. Philip AA, Dai J, Katen SP, Patton JT. Simplified reverse genetics method to recover recombinant rotaviruses expressing reporter proteins. J Vis Exp 2020; 2020: [View Article] [PubMed]
    [Google Scholar]
  7. Sánchez-Tacuba L, Feng N, Meade NJ, Mellits KH, Jaïs PH et al. An optimized reverse genetics system suitable for efficient recovery of simian, human, and murine-like rotaviruses. J Virol 2020; 94:e01294–01220 [View Article] [PubMed]
    [Google Scholar]
  8. Kobayashi T, Ooms LS, Ikizler M, Chappell JD, Dermody TS. An improved reverse genetics system for mammalian orthoreoviruses. Virology 2010; 398:194–200 [View Article] [PubMed]
    [Google Scholar]
  9. Soares-Weiser K, Bergman H, Henschke N, Pitan F, Cunliffe N. Vaccines for preventing rotavirus diarrhoea: vaccines in use. Cochrane Database Syst Rev 2019; 3:CD008521 [View Article] [PubMed]
    [Google Scholar]
  10. National Medical Products Administration of China National Institutes for Food and Drug Control (Center for Medical Device Standards Management of NMPA, National Institute for Drug Control). n.d https://english.nmpa.gov.cn/2019-07/19/c_389166.htm
  11. Liu X, Li S, Yu J, Chai P, Xie Z et al. Establishment of a reverse genetics system for rotavirus vaccine strain LLR and developing vaccine candidates carrying VP7 gene cloned from human strains circulating in China. J Med Virol 2024; 96:e70065 [View Article] [PubMed]
    [Google Scholar]
  12. Falkenhagen A, Patzina-Mehling C, Rückner A, Vahlenkamp TW, Johne R. Generation of simian rotavirus reassortants with diverse VP4 genes using reverse genetics. J Gen Virol 2019; 100:1595–1604 [View Article] [PubMed]
    [Google Scholar]
  13. Philip AA, Patton JT. Generation of recombinant rotaviruses expressing human norovirus capsid proteins. J Virol 2022; 96:e0126222 [View Article] [PubMed]
    [Google Scholar]
  14. Philip AA, Patton JT. Rotavirus as an expression platform of domains of the SARS-CoV-2 spike protein. Vaccines 2021; 9:449 [View Article] [PubMed]
    [Google Scholar]
  15. Goff A, Twenhafel N, Garrison A, Mucker E, Lawler J et al. In Vivo imaging of cidofovir treatment of cowpox virus infection. Virus Res 2007; 128:88–98 [View Article] [PubMed]
    [Google Scholar]
  16. Yamada K, Noguchi K, Kimitsuki K, Kaimori R, Saito N et al. Reevaluation of the efficacy of favipiravir against rabies virus using in Vivo imaging analysis. Antiviral Res 2019; 172:104641 [View Article] [PubMed]
    [Google Scholar]
  17. Heaton NS, Leyva-Grado VH, Tan GS, Eggink D, Hai R et al. In Vivo bioluminescent imaging of influenza a virus infection and characterization of novel cross-protective monoclonal antibodies. J Virol 2013; 87:8272–8281 [View Article] [PubMed]
    [Google Scholar]
  18. Zhu Y, Sánchez-Tacuba L, Hou G, Kawagishi T, Feng N et al. A recombinant murine-like rotavirus with Nano-Luciferase expression reveals tissue tropism, replication dynamics, and virus transmission. Front Immunol 2022; 13:911024 [View Article]
    [Google Scholar]
  19. Wang Y, Wu H, Wang B, Qi H, Jin Z et al. A NanoLuc luciferase reporter pseudorabies virus for live imaging and quantification of viral infection. Front Vet Sci 2020; 7:566446 [View Article] [PubMed]
    [Google Scholar]
  20. Pietrella D, Rachini A, Torosantucci A, Chiani P, Brown AJP et al. A beta-glucan-conjugate vaccine and anti-beta-glucan antibodies are effective against murine vaginal candidiasis as assessed by a novel in Vivo imaging technique. Vaccine 2010; 28:1717–1725 [View Article] [PubMed]
    [Google Scholar]
/content/journal/jgv/10.1099/jgv.0.002064
Loading
/content/journal/jgv/10.1099/jgv.0.002064
Loading

Data & Media loading...

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