Nine sets of nested PCR primers from a 2.6-kb region of the hepatitis G virus (HGV) genome at nucleotide positions 5829 to 8421 were designed and used to analyze serum specimens obtained from patients with community-acquired non-A, non-B hepatitis who were HGV RNA positive. One set of primers was found to be most efficient in detecting HGV and was subsequently used to test 162 HCV-positive and 11 HCV-negative plasma units obtained from individual paid donors. HGV RNA was detected in 30 (17.3%) plasma units, 2 of which were found among the 11 HCV-negative specimens. A complete set of nine PCR fragments was obtained from two patients with community-acquired acute non-A, non-B hepatitis and from four paid donors. All PCR fragments were sequenced and were shown to have a nucleotide similarity of 85.9 to 92.3% and a derived amino acid similarity of 96.0 to 99.0%. The majority of nucleotide changes occurred in the third position of codons. The HGV nucleotide and protein sequences obtained in this study were compared with HCV sequences. Based on this analysis the 2.6-kb fragment was predicted to encode the C-terminal part of the putative NS4b, the entire NS5a, and almost the complete NS5b proteins. Putative protease cleavage sites separating these proteins were also predicted. In serial specimens obtained from the two HGV-infected patients, no significant variations were found in the HGV nucleotide and derived amino acid sequences over time. The HGV sequences obtained from one patient showed no changes over 6 months, whereas more than 99.0% homology was observed for sequences from the second patient over 2.5 years. Heterogeneity analysis performed on 10 sequences obtained in this study and corresponding regions from 6 known full-size sequences of the HGV genomes demonstrated notable discrete heterogeneity consistent with the existence of HGV genetic groups or types.