Clostridium difficile is the most important cause of antibiotic-associated colitis, but its epidemiology remains unknown. Using a selective medium for the isolation of C. difficile, cultures were obtained from the environment and contacts of hospitalized patients carrying C. difficile in their stools. In areas where carriers had diarrhea, 85 (9.3%) of 910 cultures of floors and other surfaces, especially those subject to fecal contamination, were positive. In areas where there were no known carriers, only 13 (2.6%) of 497 cultures of similar sites were positive (P <0.005). C. difficile was isolated from hands and stools of asymptomatic hospital personnel, from sewage and soil, and from the home of a patient. Environmental isolates were toxigenic. C. difficile inoculated onto a floor persisted there for five months. Further studies are needed to document how often C. difficile shed by patients with antibiotic-associated colitis is acquired by other persons and whether isolation precautions are capable of limiting the organism's spread.