Background: Patients on chronic hemodialysis are at high risk of HCV infection due to nosocomial transmission. The strict adhesion to universal precautions is the first step in prevention, but other simple tools such as systematic monitor disinfection and the use of separate machines for anti-HCV-positive patients need to be evaluated. Methods: A 5-year prospective study was carried out in 4 dialysis centers enrolling 135 patients. General precautions were adopted, but anti-HCV-positive patients were not isolated. In period A, lasting 24 months, monitor disinfection was performed after each dialysis session with sodium hypochlorite; peracetic acid was also used 3 times a week. In period B, lasting 36 months, 3 dialysis units (77 patients) prolonged the same preventive protocol of period A, while another unit (58 patients) also adopted the use of separate machines for anti-HCV-positive subjects. A third-generation ELISA anti-HCV test was performed every 2 months throughout the study. Results: Anti-HCV antibodies were initially detected in 43 patients (31.8%), prevalence rate ranging from 25 to 39.4%. One seroconversion occurred in period A, with an overall seroconversion rate of 0.54%/year. Also in period B one seroconversion occurred (unit 2), seroconversion rate of 0.36%/year. Therefore the mean seroconversion rate throughout the 5 years was 0.43%/year. Conclusion: Systematic monitor disinfection may be a simple and quite effective tool to avoid nosocomial transmission of HCV infection in the hemodialysis setting. In our opinion its use is mandatory. The use of separate machines for anti-HCV-positive patients seems unnecessary.