Hepatitis virus infection, especially type C (hepatitis C virus [HCV]), has been suggested to be one of the important pathogenetic factors for low- and high-grade B-cell lymphoma, including splenic marginal zone lymphoma (SMZL), in southern Europe. Here, we analyzed the incidences of HCV and hepatitis B virus (HBV) infections, and the clinicopathologic features in 29 cases of splenic diffuse large B-cell lymphoma (DLBCL), 10 SMZL, 3 splenic mantle cell lymphoma, 1 hairy cell leukemia, 13 B-chronic lymphocytic leukemia, and 12 hepatosplenic T-cell and natural killer cell lymphoma. Fifteen (51.7%) splenic DLBCL cases were HCV antibody-positive, and another 6 (20.7%) had the HBsAg. The incidence of each was significantly (P < .01) higher than those of HCV (9.3%) and HBV (1.9%) infections in 54 node-based DLBCL cases. Four examined HCV-positive DLBCL cases showed no type II cryoglobulinemia. HCV RNA was detected in fresh tumor tissues from 6 of 7 examined DLBCL cases, and HBV DNA was present in another 2, as evaluated by real-time polymerase chain reaction. Immunohistologically, tumor cells in 5 of 7 examined DLBCL cases showed intracytoplasmic reactions for HCV NS3 and E2 proteins and the viral receptor CD81. Of 6 cases, 2 showed an intranuclear reaction for the HBV surface protein. By Southern blot analysis, no rearrangement of the Bcl2 gene was detected in the tumor tissue of 7 HCV-positive DLBCL cases. For the other types of malignant lymphoma, 1 case each of SMZL (10%) and hepatosplenic T-cell and natural killer cell lymphoma (8.3%) showed HCV infection. In conclusion, persistent human hepatitis virus infections, especially HCV, may play an important role in the tumorigenesis of splenic DLBCL in Japan.