The specificity and strength of the reported association between cytomegalovirus (CMV) and colonic adenocarcinoma were tested by analysis of a consecutive series of surgically resected colons for: CMV‐DNA by a DNA‐DNA reassociation kinetics (hybridization) procedure; latent virus by co‐cultivation of fresh tissue with indicator fibroblasts; and CMV viral antigens by immunofluorescence. Ten of 13 cancer patients whose colonic tissue was able to be examined by all techniques showed some evidence of active or prior CMV infection. Four cancer specimens were CMV‐DNA (hybridization)‐positive; an additional specimen from a cancerous colon was culture‐positive. In six instances, CMV DNA was detected in mucosal cells adjacent to colon adenocarcinoma. In tissue from one of three patients with ulcerative colitis and two of seven patients with other non‐neoplastic colonic disease, CMV DNA was also detected. No fresh colonic tissues were demonstrated to have CMV surface or nuclear antigens when examined by immunofluorescence. Culture of peripheral lymphocytes was positive for CMV in three of 14 cancer patients. A CMV‐specific defect in humoral immunity could not be documented in that most cancer patients, as well as cancer‐free patients, exhibited circulating specific antibody to CMV and had a normal capacity for CMV‐specific antibody‐dependent cellular cytotoxicity. We conclude that CMV, probably in a latent form, is readily detectable in colonic cells of man, including those derived from malignant, pre‐malignant and non‐malignant tissues. Neither preferential replication in damaged tissue nor carriage of CMV by peripheral lymphocytes homing to gut appear to explain the presence of CMV in colon cells.