ArticleViruses2026
Identification of Murine Rotavirus Virulence Determinants Using Bidirectional Selective Passaging and a Reverse Genetics System.
Article in Viruses, 2026. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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9 authors.
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Abstract
Live-attenuated rotavirus (RV) vaccines are the most effective interventions for preventing RV gastroenteritis (RVGE) in young children. However, the molecular basis of attenuation remains not well understood. Here, we describe a compact but comprehensive strategy to identify RV virulence determinants by combining low-passage bidirectional selection, sequence analysis, and segment-level phenotype testing via a reverse genetics infectious system. Using the virulent murine RV strain EW, virulence was quantified by diarrhea severity/duration and body-weight gain. Serial passaging in cell culture selected an attenuated population, which regained virulence after passaging in suckling mice. Sequence comparison of the virulent and attenuated EW populations revealed only seven amino acid differences. We summarized literature describing attenuation/virulence-associated mutations in various RV group A (RVA) strains and found previous findings identical or similar to four of the seven mutations: NSP4-T45M, VP4-S470L, VP4-T612A, and VP7-T75P. Virulent- and attenuated-type EW variants of VP2, VP4, VP7, and NSP4 were introduced individually, or as NSP4/VP7 or VP4/VP7 pairs, into a simian SA11-L2 backbone using an 11-plasmid reverse genetics system. Phenotyping of rescued viruses consistently linked cell-culture-adapted VP4 to enhanced replication in vitro and reduced virulence in suckling mice. In vivo passaging strongly favored VP4 residue S470 over cell-culture-selected L470. More generally, our findings (i) underscore VP4 and VP7 as key determinants of EW virulence, (ii) provide a practical framework for identifying driver mutations underlying RVA attenuation, and (iii) highlight attenuation-associated substitutions shared across diverse RVAs.
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