Phylogenetic and biological characterizations of a GI.3 norovirus

Noroviruses (NoVs) are a major cause of acute non-bacterial gastroenteritis worldwide. In this study, we report the isolation, near-complete genome sequencing, and expression and biological characterization of the major capsid protein (VP1) of a GI.3 NoV isolated from a child presenting acute gastroenteritis. The genome of the GI.3 NoV is 7746 bp in length, not including the poly-adenylation tail. Phylogenetic analysis based on the complete VP1 nucleotide sequences indicates that GI.3 NoVs could be divided into four clusters, with 4.6%, 5.3%, 6.6%, 1.9% intracluster variations in nucleotide and 4.8%, 3.8%, 6.1%, 1.7% intracluster variations in amino acid sequences, respectively. A Bayesian evolutionary analysis showed that GI.3 NoVs evolved at 2.44 × 10^-3, 2.78 × 10^-3, and 3.04 × 10^-3 nucleotide substitutions/site/year using a strict clock model, an uncorrelated log-normal model (UCLN), and an uncorrelated exponential derivation model (UCED), respectively. VP1 protein expression using a recombinant baculovirus expression system leads to the successful assembly of virus-like particles (VLPs). In vitro VLP-Histo-blood group antigen (HBGA) binding assay indicates that GI.3 NoV VLPs strongly bind to blood type A salivary HBGAs, moderately bind to blood type O salivary HBGAs, and weakly bind or do not bind to blood type B and AB salivary HBGAs. In vitro VLP-HBGA binding blockade assay indicated that the binding of GI.3 NoV VLPs to blood type A salivary HBGAs could only be blocked by anti-GI.3 NoV VLPs serum but not non-GI.3 NoV genotype-specific hyperimmune sera (GI.2, GI.7, GII.4, GII.6, GII.7, and GII.17). The detailed characterization of GI.3 NoV in this study provides evidence that GI.3 NoV undergoes rapid evolution and exhibits no cross-blocking effects, suggesting that GI.3 NoV may potentially be utilized in the development of multivalent NoV vaccines.