Letter

The major targets of acute norovirus infection are immune cells in the gut-associated lymphoid tissue

Received:
Accepted:
Published online:

Abstract

Noroviruses are the leading cause of food-borne gastroenteritis outbreaks and childhood diarrhoea globally, estimated to be responsible for 200,000 deaths in children each year1,2,3,4. Thus, reducing norovirus-associated disease is a critical priority. Development of vaccines and therapeutics has been hindered by the limited understanding of basic norovirus pathogenesis and cell tropism. While macrophages, dendritic cells, B cells and stem-cell-derived enteroids can all support infection of certain noroviruses in vitro5,6,7, efforts to define in vivo norovirus cell tropism have generated conflicting results. Some studies detected infected intestinal immune cells8,9,10,11,12, other studies detected epithelial cells13, and still others detected immune and epithelial cells14,15,16. Major limitations of these studies are that they were performed on tissue sections from immunocompromised or germ-free hosts, chronically infected hosts where the timing of infection was unknown, or following non-biologically relevant inoculation routes. Here, we report that the dominant cellular targets of a murine norovirus inoculated orally into immunocompetent mice are macrophages, dendritic cells, B cells and T cells in the gut-associated lymphoid tissue. Importantly, we also demonstrate that a norovirus can infect T cells, a previously unrecognized target, in vitro. These findings represent the most extensive analyses to date of in vivo norovirus cell tropism in orally inoculated, immunocompetent hosts at the peak of acute infection and thus they significantly advance our basic understanding of norovirus pathogenesis.

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References

  1. 1.

    Patel, M. M. Systematic literature review of role of noroviruses in sporadic gastroenteritis. Emerg. Infect. Dis. 14, 1224–1231 (2008).

  2. 2.

    Payne, D. C. et al. Norovirus and medically attended gastroenteritis in U.S. children. N. Engl. J. Med. 368, 1121–1130 (2013).

  3. 3.

    Koo, H. L., Ajami, N., Atmar, R. L. & DuPont, H. L. Noroviruses: the principal cause of foodborne disease worldwide. Discov. Med 10, 61–70 (2010).

  4. 4.

    Ahmed, S. M. et al. Global prevalence of norovirus in cases of gastroenteritis: a systematic review and meta-analysis. Lancet Infect. Dis. 14, 725–730 (2014).

  5. 5.

    Jones, M. K. et al. Enteric bacteria promote human and murine norovirus infection of B cells. Science 346, 755–759 (2014).

  6. 6.

    Wobus, C. E. et al. Replication of norovirus in cell culture reveals a tropism for dendritic cells and macrophages. PLoS Biol. 2, e432 (2004).

  7. 7.

    Ettayebi, K. et al. Replication of human noroviruses in stem cell-derived human enteroids. Science 353, 1387–1393 (2016).

  8. 8.

    Mumphrey, S. M. et al. Murine norovirus 1 infection is associated with histopathological changes in immunocompetent hosts, but clinical disease is prevented by STAT1-dependent interferon responses. J. Virol. 81, 3251–3263 (2007).

  9. 9.

    Bok, K. et al. Chimpanzees as an animal model for human norovirus infection and vaccine development. Proc. Natl Acad. Sci. USA 108, 325–330 (2011).

  10. 10.

    Taube, S. et al. A mouse model for human norovirus. mBio 4, e00450-13 (2013).

  11. 11.

    Ward, J. M. et al. Pathology of immunodeficient mice with naturally occurring murine norovirus infection. Toxicol. Pathol. 34, 708–715 (2006).

  12. 12.

    Lay, M. K. et al. Norwalk virus does not replicate in human macrophages or dendritic cells derived from the peripheral blood of susceptible humans. Virology 406, 1–11 (2010).

  13. 13.

    Cheetham, S. et al. Pathogenesis of a genogroup II human norovirus in gnotobiotic pigs. J. Virol. 80, 10372–10381 (2006).

  14. 14.

    Karandikar, U. C. et al. Detection of human norovirus in intestinal biopsies from immunocompromised transplant patients. J. Gen. Virol 97, 2291–2300 (2016).

  15. 15.

    Souza, M., Azevedo, M. S. P., Jung, K., Cheetham, S. & Saif, L. J. Pathogenesis and immune responses in gnotobiotic calves after infection with the genogroup II.4-HS66 strain of human norovirus. J. Virol. 82, 1777–1786 (2008).

  16. 16.

    Otto, P. H. et al. Infection of calves with bovine norovirus GIII.1 strain Jena virus: an experimental model to study the pathogenesis of norovirus infection. J. Virol. 85, 12013–12021 (2011).

  17. 17.

    Atmar, R. L. et al. Norwalk virus shedding after experimental human infection. Emerg. Infect. Dis. 14, 1553–1557 (2008).

  18. 18.

    Kirby, A. E., Shi, J., Montes, J., Lichtenstein, M. & Moe, C. L. Disease course and viral shedding in experimental Norwalk virus and Snow Mountain virus infection. J. Med. Virol. 86, 2055–2064 (2014).

  19. 19.

    Teunis, P. F. M. et al. Shedding of norovirus in symptomatic and asymptomatic infections. Epidemiol. Infect. 143, 1710–1717 (2015).

  20. 20.

    Thackray, L. B. et al. Murine noroviruses comprising a single genogroup exhibit biological diversity despite limited sequence divergence. J. Virol. 81, 10460–10473 (2007).

  21. 21.

    Karst, S. M. & Wobus, C. E. Viruses in rodent colonies: lessons learned from murine norovirus. Annu. Rev. Virol. 2, 525–548 (2015).

  22. 22.

    Liu, G., Kahan, S. M., Jia, Y. & Karst, S. M. Primary high-dose murine norovirus 1 infection fails to protect from secondary challenge with homologous virus. J. Virol. 83, 6963–6968 (2009).

  23. 23.

    Bonnardel, J. et al. Innate and adaptive immune functions of Peyer’s patch monocyte-derived cells. Cell Rep. 11, 770–784 (2015).

  24. 24.

    Gonzalez-Hernandez, M. B. et al. Murine norovirus transcytosis across an in vitro polarized murine intestinal epithelial monolayer is mediated by M-like cells. J. Virol. 87, 12685–12693 (2013).

  25. 25.

    Gonzalez-Hernandez, M. B. et al. Efficient norovirus and reovirus replication in the mouse intestine requires microfold (M) cells. J. Virol. 88, 6934–6943 (2014).

  26. 26.

    Orchard, R. C. et al. Discovery of a proteinaceous cellular receptor for a norovirus. Science 353, 933–936 (2016).

  27. 27.

    Haga, K. et al. Functional receptor molecules CD300lf and CD300ld within the CD300 family enable murine noroviruses to infect cells. Proc. Natl Acad. Sci. USA 113, E6248–E6255 (2016).

  28. 28.

    Miller, H. Intestinal M cells: the fallible sentinels? World J. Gastroenterol. 13, 1477–1486 (2007).

  29. 29.

    Taube, S., Jiang, M. & Wobus, C. E. Glycosphingolipids as receptors for non-enveloped viruses. Viruses 2, 1011–1049 (2010).

  30. 30.

    Zhu, S. et al. Identification of immune and viral correlates of norovirus protective immunity through comparative study of intra-cluster norovirus strains. PLoS Pathog. 9, e1003592 (2013).

  31. 31.

    Kahan, S. M. et al. Comparative murine norovirus studies reveal a lack of correlation between intestinal virus titers and enteric pathology. Virology 421, 202–210 (2011).

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Acknowledgements

The authors thank C. Jobin and X. Sun (University of Florida) for technical guidance in swiss rolling, H. Lelouard (Centre d’Immunologie de Marseille-Luminy) for discussions on Peyer’s patch cell types, J. Shirley (University of Florida) for technical guidance on multicolour flow cytometric analysis, and D.C. Machart and L. Schneider (University of Florida Molecular Pathology Core) for their assistance in processing histology samples. The authors also thank C. Fisher and T. Edwards (University of Florida) for their assistance with microscopic analyses, and D. Avram, D. Bloom, S. Tibbetts and F. Zhu for providing cell lines. This work was also supported by the technical guidance provided by ACDBio in terms of optimizing RNAscope assays. This work was funded by NIH R01AI116892 and NIH R01AI081921.

Author information

Author notes

  1. Katrina R. Grau, Alexa N. Roth and Shu Zhu contributed equally to this work.

Affiliations

  1. Department of Molecular Genetics & Microbiology, Emerging Pathogens Institute, Center for Inflammation and Mucosal Immunology, College of Medicine, University of Florida, Gainesville, FL, USA

    • Katrina R. Grau
    • , Alexa N. Roth
    • , Shu Zhu
    • , Abel Hernandez
    • , Drake T. Philip
    •  & Stephanie M. Karst
  2. Department of Infectious Diseases & Immunology, Center for Inflammation & Mucosal Immunology, College of Medicine, University of Florida, Gainesville, FL, USA

    • Natacha Colliou
    •  & Mansour Mohamadzadeh
  3. Department of Oral Biology, College of Dentistry, University of Florida, Gainesville, FL, USA

    • Bayli B. DiVita
    •  & Shannon M. Wallet
  4. Department of Neuroscience, Center for Translational Research in Neurodegenerative Disease, College of Medicine, University of Florida, Gainesville, FL, USA

    • Cara Riffe
    •  & Benoit Giasson

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Contributions

K.R.G., A.N.R., S.Z. and S.M.K. designed the study and analysed results. K.R.G. performed and analysed RNAscope-based FISH assays and quantified chromogenic assays. S.Z. and A.H. performed mouse infections, harvests and plaque assays, and S.Z. performed RNAscope-based chromogenic assays. A.N.R. performed and analysed in vitro infections and viability assays on cell lines as well as CD300lf expression on cell lines and Peyer’s patch cells. N.C. and M.M. assisted with fluorescence microscopy. N.C. and B.B.D. performed flow cytometric analyses of in vivo samples guided by the expertise of S.M.W. and M.M. D.T.P. performed TCID50 assays and analysed data. C.R. and B.G. assisted with analysing chromogenic assays using a slide scanner. K.R.G., A.N.R. and S.M.K. prepared the manuscript. M.M. and S.Z. edited the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Stephanie M. Karst.

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