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Archives of Virology

, Volume 159, Issue 2, pp 257–266 | Cite as

Failure of propagation of human norovirus in intestinal epithelial cells with microvilli grown in three-dimensional cultures

  • Sayaka Takanashi
  • Linda J. Saif
  • John H. Hughes
  • Tea Meulia
  • Kwonil Jung
  • Kelly A. Scheuer
  • Qiuhong WangEmail author
Original Article

Abstract

Human noroviruses (HuNoVs) are a leading cause of acute gastroenteritis. Establishment of a cell culture system for in vitro HuNoV growth remains challenging. Replication of HuNoVs in human intestinal cell lines (INT-407 and Caco-2) that differentiate to produce microvilli in rotation wall vessel (RWV) three-dimensional cultures has been reported (Straub et al. in Emerg Infect Dis 13:396–403, 2007; J Water Health 9:225–240, 2011, and Water Sci Technol 67:863–868, 2013). We used a similar RWV system, intestinal cell lines, and the same (Genogroup [G] I.1) plus additional (GII.4 and GII.12) HuNoV strains to test the system’s reproducibility and to expand the earlier findings. Apical microvilli were observed on the surface of both cell lines by light and electron microscopy. However, none of the cell types tested resulted in productive viral replication of any of the HuNoV strains, as confirmed by plateau or declining viral RNA titers in the supernatants and cell lysates of HuNoV-infected cells, determined by real-time reverse transcription PCR. These trends were the same when culture supplements were added that have been reported to be effective for replication of other fastidious enteric viruses in vitro. Additionally, by confocal microscopy and orthoslice analysis, viral capsid proteins were mainly observed above the actin filament signals, which suggested that the majority of viral antigens were on the cell surface. We conclude that even intestinal cells displaying microvilli were not sufficient to support HuNoV replication under the conditions tested.

Keywords

Cell Aggregate Rotate Wall Vessel Norwalk Virus Sapovirus Microcarrier Bead 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

We thank Juliette Hanson and Kingsly Berlin for assistance with animal care, Andrea Kaszas for assistance with microscopic examination, and Tomoichiro Oka for helpful discussions. This work was supported by a grant from the National Institute of Allergy and Infectious Diseases, National Institutes of Health (R21 AI081009-2). Salaries and research support were provided by state and federal funds provided to the Ohio Agricultural Research and Development Center (OARDC), The Ohio State University.

Conflict of interest

The authors declare that they have no conflict of interest.

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Copyright information

© Springer-Verlag Wien 2013

Authors and Affiliations

  • Sayaka Takanashi
    • 1
    • 2
    • 3
  • Linda J. Saif
    • 1
    • 2
  • John H. Hughes
    • 4
  • Tea Meulia
    • 5
  • Kwonil Jung
    • 1
    • 2
  • Kelly A. Scheuer
    • 1
    • 2
  • Qiuhong Wang
    • 1
    • 2
    Email author
  1. 1.Food Animal Health Research Program, Ohio Agricultural Research and Development CenterThe Ohio State UniversityWoosterUSA
  2. 2.Department of Veterinary Preventive Medicine, College of Veterinary MedicineThe Ohio State UniversityColumbusUSA
  3. 3.Department of Developmental Medical Sciences, Graduate School of MedicineThe University of TokyoTokyoJapan
  4. 4.Department of Molecular Virology, Immunology and Medical Genetics, College of MedicineThe Ohio State UniversityColumbusUSA
  5. 5.Molecular and Cellular Imaging Center, Ohio Agricultural Research and Development CenterThe Ohio State UniversityWoosterUSA

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