Epstein-Barr virus (EBV) was found in 7-17% of gastric adenocarcinomas, including lymphoepithelioma-like carcinomas, although its significance has not been clear. In addition, 20-30% of malignant lymphomas arising in the gastrointestinal tract have been known to express the EBV genome. Several lines of evidence indicate that EBV has been shown to infect both B lymphocytes and squamous epithelial cells via CD21 molecule in vivo and in vitro. The expression of CD21 in EBV-associated gastrointestinal tumors, however, has remained controversial. To determine the presence of CD21, an EBV receptor, in the EBV-associated gastrointestinal tumors, we, first, examined the EBV genome in sixty seven patients with either gastrointestinal adenocarcinomas or malignant lymphomas using in situ hybridization (ISH) for EBV-encoded small RNAs (EBERs) and PCR for EBNA-1. Then, the investigation of CD21 expression was performed only in the EBV-positive tumors with immunohistochemical method for CD21 antigen on paraffin sections. EBERs were detected in 6 out of 26 gastric adenocarcinomas, 2 of 24 colonic adenocarcinomas, and 8 of 17 malignant lymphomas. EBERs were more prevalent in the malignant lymphomas originating from the small and large intestine (6/6) than from the stomach (2/11), and were detected in both B and T cell phenotypes. EBNA-1 was amplified in 11 of 16 EBERs-positive cases. Interestingly, however, none of the EBV-positive six gastric adenocarcinomas and eight malignant lymphomas expressed the CD21 on the cell surfaces or cytoplasm of both tumor cells and adjacent normal epithelial cells. These results suggest that EBV infection in the gastrointestinal malignancies would be mediated via different routes besides the CD21 or a new receptor distinct from CD21.