Hepatitis C virus (HCV) infection is a frequent feature in hemodialysis (HD) patients. The way of viral transmission is difficult to establish, but in previous studies the role of blood transfusions and of HD treatment duration, and the possibility of nosocomial transmission of the virus have been suggested. We present here the results of a virological follow-up of HCV infection in our HD unit in 1993-1994, and a molecular study of viral strains that led to a possible reconstruction of viral spreading. All patients in our unit were regularly tested for alanine aminotransferase, HCV antibodies and HCV RNA in serum. Seven seroconversions were detected during follow-up, and a high proportion of type 1b HCV strains was found in infected patients. Nucleotide sequences located in the envelope 1 (E1) viral coding region of type 1b strains were compared in our patients and numerous controls infected with the same HCV genotype. A high proportion of patients with antibodies to HCV were detected in our unit (32.5%). Blood transfusions and duration of HD treatment were risk factors for HCV infection. Seroconversions in patients never transfused and predominance of type 1b HCV strains suggested that infection had occurred via the nosocomial pathway in our unit. Similar sequences in the E1 region were found in four patients treated, forming a distinct cluster in a phylogenetic tree. Of these four patients, two had been infected before 1991, and the others made a seroconversion for HCV at the same period in 1994. In all other patients, including a nurse who had been in charge of some infected patients, distinct strains were found. Duration of HD treatment seems to be a major factor of risk for HCF infection in HD units. Contamination could occur during blood transfusion or via the nosocomial pathway through a crossinfection mechanism from patients already infected. The latter mechanism was formally demonstrated in this study.