The health consequences of hepatitis G virus (HGV) infection in humans remain to be determined. Inconsistent results of HGV and the risk of hepatocellular carcinoma (HCC) have been found in retrospective case-control studies. No prospective studies have addressed this issue. A prospective study including 18,244 men in Shanghai, China, was conducted to assess the role of HGV infection in HCC development. Serum HGV RNA and hepatitis B surface antigen (HBsAg) were assessed in 127 HCC case patients and 660 matched control subjects drawn from the cohort. Four patients with HCC (3.1%) and 21 control subjects (3.2%) were positive for serum HGV RNA. The presence of HGV RNA in prediagnostic serum was not associated with the risk of HCC (odds ratio, 1.0; 95% confidence interval, 0.3-2.9). No association between HGV RNA positivity and HCC risk was observed in either HBsAg-positive carriers or noncarriers. It is concluded that HGV infection plays no role in HCC development in this high-risk population.