Astroviruses are single-stranded, positive-sense RNA viruses that are associated with gastroenteritis in humans and animals. We describe a reverse transcription-polymerase chain reaction (RT-PCR) assay using primers targeted to a nonstructural protein coding region that allowed sensitive detection and genetic typing of representative strains of seven astrovirus serotypes. Phylogenetic analysis of the nucleotide sequences of PCR products from the reference strains and several wild isolates indicated two distinct genogroups of sequences in open reading frame 1a (ORF 1a). These genogroups correlated with serotype: genogroup A included strains of types 1 to 5, while genogroup B included strains of types 6 and 7. This phylogenetic arrangement differs from the nearly equidistant clustering of serotypes seen when comparing nucleotide sequences from either ORF 1b or ORF 2. It is possible that recombination was responsible for the observed difference in genetic relationships.