A

FIG. 2. Relationship of astrovirus strains based upon partial sequences from ORFla. Distances were calculated based on the 214 nucleotide

amplification products from ORFla by using the distances program of the GCG package. The country of origin of each strain is indicated by a three letter IS03166 code. All strains of serotypes 1-5 and 8 are clustered in genogroup A, while those of serotypes 6 and 7 are clustered in genogroup B. Within genogroup A, strains belonging to different serotypes (indicated by numbers) may have very similar, or identical ORFla sequences. '

FIG. 3. Relationship of astrovirus strains based upon complete amino acid sequences. Distances were calculated based on predicted amino acid sequences for the P2 and P3 proteins from twelve strains for which complete ORF2 sequences were available in GenBank. The dendrograms are plotted to the same scale with the bar representing 10 amino acid changes.

FIG. 3. Relationship of astrovirus strains based upon complete amino acid sequences. Distances were calculated based on predicted amino acid sequences for the P2 and P3 proteins from twelve strains for which complete ORF2 sequences were available in GenBank. The dendrograms are plotted to the same scale with the bar representing 10 amino acid changes.

assign strains to individual serotypes on the basis of sequences in ORFla. Furthermore, it appeared that sequences in this region of ORFla were more likely to cluster by geographic location than by serotype. For example, four samples from Egypt and three from Malawi had nearly identical sequences in their ORFla amplicons, despite clustering as different serotypes in ORF2 (Fig. 2). These results suggest that although conserved regions of ORFla are useful targets for RT-PCR assays designed to efficiently detect all astrovirus types, one cannot use analysis of sequence information from this region of the genome to assign genetic types that correlate with antigenic types.

The finding of multiple serotype 8 strains in these two collections prompted us to re-examine the issue of the genetic relationship between HAstV serotypes 4 and 8. Analysis of partial sequences from ORF2 indicates that strains from these serotypes fall into distinct clusters, but the pair-wise distances between the serotypes are less than some of the distances between strains within other serotypes (e.g. HAstV2, Fig. 1). To examine this relationship in more detail, we compared predicted amino acid sequences for 12 astrovirus strains for which complete ORF2 sequences were available. The close genetic relationship between serotypes 4 and 8 was observed when we compared predicted amino acid sequences for the P2 proteins (Fig. 3). In contrast, when we compared predicted amino acid sequences for the P3 proteins from the same 12 strains, the absolute distances between serotypes were greater, and those between serotypes 4 and 8 were comparable to those between the other serotypes (Fig. 3). These findings are consistent with a model wherein the relatively conserved P2 proteins of serotypes 4 and 8 contain shared epitopes, which were detected in the immunoelectron microscopy assay, while the divergent P3 proteins contain serotype-specific epitopes distinguishable on the basis of predicted amino acid sequences.

The recent application of molecular diagnostics has firmly established that astroviruses are a common cause of acute gastroenteritis in young children. Genetic analysis of amplification products can be a useful technique for determining astrovirus strain types and this approach has revealed a surprising diversity of strains co-circulating within relatively isolated populations. Additional studies are needed to assess the severity of astrovirus infections as the basis for a careful cost—benefit analysis of a potential vaccination program. Additionally, the role of astroviruses as a cause of outbreaks of gastroenteritis, both in children and adults, needs to be more fully investigated as the basis for formulating recommendations for public health interventions to interrupt transmission.

We would like to thank our collaborators Abdollah Naficy, Stephen Savarino, and John Clemens from the Pediatric Infectious Diseases and Vaccines Section, NIH, and their coworkers at NAMRU-2, and C. Anthony Hart and Nigel Cunliffe from the Department of Medical Microbiology, University of Liverpool.

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