Laboratory efforts to cultivate noroviruses

Erwin Duizer, Kellogg J. Schwab, Frederick H. Neill, Robert L. Atmar, Marion P.G. Koopmans, Mary K. Estes

Research output: Contribution to journalArticle

Abstract

Noroviruses (NoVs) are a leading cause of gastroenteritis worldwide and are recognized as the foremost cause of foodborne illness. Despite numerous efforts, routine cell cultures have failed to yield replicating NoV. This paper describes methods used to try to grow NoV in vitro in two laboratories. Cells (A549, AGS, Caco-2, CCD-18, CRFK, CR-PEC, Detroit 551, Detroit 562, FRhK-4, HCT-8, HeLa, HEC, HEp-2, Ht-29, HuTu-80, I-407, IEC-6, IEC-18, Kato-3, L20B, MA104, MDBK, MDCK, RD, TMK, Vero and 293) were cultured on solid or permeable surfaces. Differentiation was induced using cell culture supplements such as insulin, DMSO and butyric acid. In some cases, the cells and the NoV-containing stool samples were treated with bioactive digestive additives. Variables evaluated in cultivation experiments included the method of preparation of the virus inoculum, the genotype of the virus, conditions for maintenance of cell monolayers, additives in the maintenance medium and the method of inoculation of the cells. Serial blind passage studies were performed routinely. In addition to evaluation for CPE, evidence of virus replication was sought using immunofluorescent assays to detect newly produced viral capsid antigen and RT-PCR assays to detect the viral genome. Although some infected cultures remained NoV positive by RT-PCR for up to five passages and an occasional cell in a monolayer showed evidence of specific immunofluorescence, no reproducible NoV-incluced CPE was observed and all RT-PCR results that were positive initially were negative following continued passaging. Thus, attempts to develop a method for the cultivation of NoV were unsuccessful.

Original languageEnglish (US)
Pages (from-to)79-87
Number of pages9
JournalJournal of General Virology
Volume85
Issue number1
DOIs
StatePublished - Jan 1 2004

ASJC Scopus subject areas

  • Virology

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    Duizer, E., Schwab, K. J., Neill, F. H., Atmar, R. L., Koopmans, M. P. G., & Estes, M. K. (2004). Laboratory efforts to cultivate noroviruses. Journal of General Virology, 85(1), 79-87. https://doi.org/10.1099/vir.0.19478-0