We have determined the nucleotide sequences of a highly conserved region of the RNA-dependent RNA polymerase of the prototype Snow Mountain agent (SMA) and of four other small, round-structured viruses (antigenically Norwalk virus [NV]-like or SMA-like) following reverse transcription-PCR amplification of viral RNA obtained from human stools. The stool samples were either from volunteers administered SMA or from sporadic cases and outbreaks of gastroenteritis that occurred in Japan and the United Kingdom between 1984 and 1992. The GLPSG and YGDD RNA polymerase motifs were in the proper locations in the sequences of the five SRSVs, but each sequence was distinct from the 8FIIa prototype NV sequence and from each other. Analysis of the sequences and reactivities in a new NV antigen enzyme-linked immunosorbent assay showed that the five viruses could be divided into two groups (serogroups) with NV and SMA, respectively, being the prototypes. The sequences of the capsid region and a nonstructural region (2C) were determined from one strain from each group. One virus (SRSV-KY-89/89/J), isolated in Japan and antigenically similar to the prototype NV (isolated 21 years earlier in Ohio), showed a remarkable level of sequence similarity to NV. KY-89 and the 8FIIa NV showed 87.2% nucleotide similarity over 2,516 continuous nucleotides amounting to 96 to 98.9% amino acid similarity in three distinct domains in two open reading frames. Between the prototype SMA and NV, the polymerase region showed 63% nucleotide and 59% amino acid similarity, respectively. Two other antigenically SMA-like isolates (SRSV-925/92/UK and SRSV-OTH-25/89/J), from the United Kingdom and Japan, showed 80% nucleotide and 88 to 92% amino acid similarity in the polymerase region to the prototype SMA isolated 16 and 13 years earlier in the United States. The capsid region of the antigenically SMA-like OTH-25 virus showed 53% nucleotide and 65% amino acid similarity to the prototype NV capsid region. Domains of sequence diversity and conservation were identified within the capsid protein of these two distinct prototype serotypes of virus. These results indicate that NV-like and SMA-like agents are still circulating, and sequence comparisons will be useful to identify and classify distinct viruses in the NV group.

Human rotavirus field isolates were characterized by direct sequence analysis of the gene encoding the serotype-specific major neutralization protein (VP7). Single-stranded RNA transcripts were prepared from virus particles obtained directly from stool specimens or after two or three passages in MA-104 cells. Two regions of the gene (nucleotides 307 through 351 and 670 through 711) which had previously been shown to contain regions of sequence divergence among rotavirus serotypes were sequenced by the dideoxynucleotide method with two different synthetic oligonucleotide primers. The resulting nucleotide sequences were compared with the corresponding sequences from rotaviruses of known serotype (serotype 1, 2, 3, or 4). A total of 25 field isolates and 10 laboratory strains examined by this method exhibited marked sequence identity in both areas of the gene with the corresponding regions of 1 of the 4 reference strains. In addition, the predicted serotype from the sequence analysis correlated in each case with the serotype determined when the rotaviruses were examined by plaque reduction neutralization or reactivity with serotype-specific monoclonal antibodies. These data suggest that as a result of the high degree of sequence conservation observed among rotaviruses of the same serotype, it is possible to predict the serotype of a rotavirus isolate by direct sequence analysis of its VP7 gene.