Characterization of SRSVs using RT-PCR and a new antigen ELISA

Stool samples from 451 patients involved in volunteer studies, 26 outbreaks and approximately 175 sporadic cases of acute gastroenteritis from different geographical locations in the world were tested for Norwalk virus (NV) using a newly developed antigen ELISA and RT-PCR. NV was detected in most outbreaks previously characterized as being of NV origin. Overall, a low number of positives for NV was obtained using either RT-PCR with primers that amplified a unique region of the genome, or an ELISA with hyperimmune antisera made to the baculovirus-expressed recombinant NV capsid. However, a significant number of positives was obtained when these samples were tested by RT-PCR using primers that amplified the more highly conserved regions of the genome. Sequence analysis of the amplified viral cDNAs indicated that small round structured viruses (SRSVs) with a wide range of variable genomic sequences (44-87% nucleotide and 31-99% amino acid similarity to the 8Flla NV genome sequence) were responsible for these outbreaks. Several recent outbreaks from the US, Japan and the UK were related to the Snow Mountain Agent (SMA) by sequence analyses. Continued accumulation of sequence information will facilitate the design of new primers for virus detection and increase our understanding of the relationships and epidemiology of these viruses from different sources.