Of all viruses causing liver pathology, Epstein-Barr virus (EBV) has been one of the most ubiquitous. Presumably, a co-infection of both EBV and hepatitis C virus (HCV) negatively influences the prognosis of HCV-infected patients. Hence, we endeavoredto estimate the impact of EBV infection on treatment with interferon plus ribavirin in chronic HCV patients. Eighty-four chronic HCV patients and sixteen control subjects (with no symptoms of HCV infection) were enrolled in this study. Viral loads were assessed using reverse transcriptase polymerase chain reaction, while anti-EBV IgM levels were determined by ELISA. Chronic HCV patients with negative EBV IgM exhibited higher response rates (85.7%) to interferon than those with positive EBV IgM (62%). Furthermore, HCV RNA load was remarkably higher in EBV-seropositive patients (with a P-value of 0.0001). Chronic HCV infection cannot be considered in isolation; as other pathogens such as EBV may have far-ranging effects against the disease prognosis. We concluded that EBV co-infection can deteriorate the response to interferon in patients with chronic HCV infection, and that EBV could enhance the replication of HCV. Accordingly, EBV treatment should be considered when designing specific protocols for treatment of such patients.