Hepatitis C virus (HCV) RNA was detected in the sera of patients with non-A, non-B chronic liver disease by polymerase chain reaction (PCR). RNA was extracted from the serum, reverse transcribed to cDNA, and amplified by PCR. With this method, 30 patients with non-A, non-B chronic liver disease and 10 healthy subjects were tested. HCV RNA was detected in 13 of 16 (81%) anti-HCV-positive patients and also in 7 of 14 (50%) anti-HCV-negative patients, but in none of 10 anti-HCV-negative healthy subjects. Specificity of this method was confirmed by direct sequencing of amplified cDNA segment. The nucleotide sequences (37 nucleotides) obtained from 15 patients showed only 68-78% homology compared with the prototype HCV nucleotide sequence. In addition, of 15 nucleotide sequences, there were 12 different types. But the translated amino acid sequences (12 amino acids) showed 83-100% homology compared with the prototype HCV amino acid sequence. These data suggest the majority of anti-HCV-positive patients are carriers of HCV. But to detect all the viremic patients, the anti-HCV antibody testing may be insufficient. Direct detection of HCV RNA may be useful in the study of virus replication and its association with various liver diseases.