Background: SouthernTaiwan is a hepatitis B and C viruses (HBV, HCV) endemic area. SEN virus (SENV) infection has been suggested as transfusion-related hepatitis. Two variants of SENV (SENV-D and SENV-H) have been studied in non-transfused children and transfusion-dependent thalassemia patients. Methods: Sera of 67 non-transfused children and 55 pediatric thalassemia patients with multiple transfusions were tested for SENV-D and SENV-H DNAs, liver function, iron status, HBV and HCV markers. Results: The prevalence of SENV (D or H), SENV-D, SENV-H infection, and SENV-D/H coinfection was significantly lower in nontransfused children than in thalassemia patients (22.4, 20.9, 5.0 and 1.5%, respectively, versus 67.3, 52.7, 40.0 and 25.5%, respectively, p < 0.001). The serum alanine aminotransferase (ALT) levels in thalassemia patients with SENV infection alone were significantly lower than levels in patients with SENV/HCV co-infection (p < 0.05), but not different when compared with those without SENV/HCV infection. SENV viremia was not associated with elevated ALT levels in thalassemia patients. SENV viremia did not increase the risk of HCV infection in thalassemia patients. Conclusions: SENV infection is high among non-transfused controls in Taiwan. Transfusion significantly increases the relevance of SENV infection. SENV viremia was not associated with the ALT levels in thalassemia patients.