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Regarding viral infection of intestinal mucosa, there have been only a few studies on limited diseases, targeting a few herpes family viruses. In this study, we analyzed 12 kinds of DNA viruses including 8 species of herpes family viruses in the gastrointestinal mucosa of patients with hematologic malignancies, inflammatory bowel diseases, collagen diseases, or other miscellaneous forms of gastroenteritis using the multiplex virus PCR assay, which we recently developed. The virus PCR assay yielded positive results in 63 of 102 patients; Epstein-Barr virus (EBV) was the most frequently detected, followed by cytomegalovirus (CMV), human herpes virus 6 (HHV-6), HHV-7, parvovirus B19, and herpes simplex virus type 1. The frequencies of viral detection in the 4 diseases were similar involving these 6 viruses. Regarding CMV colitis, the multiplex virus PCR assay was superior to the immunohistopathologic method in detecting CMV. All viruses were more efficiently detected in the mucosa than in the blood in individual patients. These results suggest that CMV, EBV, and HHV-6 were commonly detected in the gastrointestinal mucosa of patients with these 4 diseases, and our multiplex virus PCR assay was useful for the early diagnosis of gastrointestinal virus infection, especially CMV colitis.