Paired serum and saliva samples were collected simultaneously from 50 intravenous drug users with serologically proven hepatitis C virus infection. The oral health of the volunteers was also assessed. Hepatitis C virus RNA was detected by nested PCR, employing primers from the 5' noncoding region. Positive PCR products were sequenced using the Sequenase PCR Product Sequencing Kit (Amersham Life Sciences). HCV RNA was detected in 33 (66%) of the 50 serum samples. HCV RNA was detected in 19 (57.6%) of the corresponding 33 saliva samples. There was no correlation between oral health status or HIV seropositivity and the detection of HCV in saliva. However, subjects with HCV in their saliva were significantly more likely to complain of xerostomia (P < 0.05). Isolate genotypes were identified in paired serum and saliva of 15 intravenous drug users. HCV genotypes 1, 2, 3 and 6 were detected in both specimens. In seven cases, a differing HCV genotype was found in serum compared to the paired saliva specimen. The distributions of genotypes in serum and saliva were very different, with genotype 2a more common in saliva than serum (P < 0.005). These data suggest that in some cases the source of salivary HCV may not be serum transudation along the periodontal membrane or across damaged mucosa, and that an alternative local source, possibly the salivary glands themselves, should be considered.