Genetic polymorphism across regions of the three open reading frames of "Norwalk-like viruses"

Genomic characterization of Norwalk-like human caliciviruses (NLVs) origina ting from outbreaks and sporadic cases of acute gastroenteritis has reveale d surprisingly high levels of diversity, even in the RNA polymerase gene, w hich is anticipated to be highly conserved. Since information on antigenic relationship is limited, due to the lack of a tissue culture system for the se viruses, strains mostly are described on the basis of their genetic rela tedness. However, the lack of uniformly applied criteria has led to a confu sing array of strains with different groups employing different names for s imilar genetic lineages. Our goal was to conduct a structured analysis of g enomic relationships among NLV strains in an attempt to provide an interim framework for genotyping. We assembled a panel of 31 potentially distinct g enogroup I (GGI) and genogroup II (GGII) NLVs that reflected the diversity seen in strains detected by our laboratories and in published sequences. Ph ylogenetic analysis of sequences from regions of the open reading frames (O RF) 1, 2 and 3 was performed in order to investigate genomic relationships. The strains sequenced fell into seven phylogenetic groups in GGI and at le ast five phylogenetic groups in GGII, based on greater than 80% nucleotide identity in the region of ORF2 encoding the N-terminus of the capsid protei n, and consistent clustering with high bootstrap values irrespective of the method used. Analysis of the ORF1 and ORF3 regions supported for most stra ins the clustering as established for those derived from ORF2. In the ORF1 region, used by most laboratories for diagnostic RT-PCR, clustering was con sistent when a putative genotype border was set at 15% nucleotide mismatche s for viruses in GGI and at 10% for viruses in GGII. Two strains grouped wi thin different clusters based on ORF1 and ORF2 indicating that recombinatio n may have occurred. We discuss the implications of these observations for the classification and typing of NLVs.