DNA extracts from hepatocellular carcinomas of 13 patients from South Africa were examined for hepatitis B virus (HBV) DNA sequences by molecular hybridization using [32P]-labeled recombinant, cloned, and purified HBV-DNA. Eight patients were HBV carriers as demonstrated by the presence of hepatitis B surface antigen (HBsAg) in their serum, and each of these patients and HBV-DNA sequences in hepatocellular carcinoma tissue. Five patients who were not HBsAg carriers, did not have HBV-DNA in their tumors. In DNA extracts from all tumors of patients who were HBsAg-positive, the HBV-DNA was integrated into the host genome. The integration pattern was unique for each tumor, but HBV-DNA bands of a given length were present in more than one specimen and in a human hepatocellular carcinoma cell line (PLC/PRF/5). These results suggest that integration of HBV-DNA into the human genome occurs in conjunction with malignant transformation.