Clostridium difficile is the major identifiable infectious cause of nosocomial diarrhea. A prospective study was conducted at New England Deaconess Hospital (Boston) to examine risk factors for C. difficile carriage at both admission and follow-up. Specimens from patients admitted to two wards (one medical, one surgical) and three intensive care units (two surgical, one medical) were cultured weekly until discharge. For 89 (18%) of 496 patient admissions, at least one culture was positive. The prevalence of culture positivity within 72 hours of admission was 11%. Risk factors for culture positivity at admission were prior C. difficile diarrhea (adjusted odds ratio [OR] = 9.5), renal insufficiency (OR = 6.7), and recent hospitalization elsewhere (OR = 3.1). Fifteen percent of patients for whom initial cultures were negative and for whom follow-up cultures were performed acquired C. difficile. Admission to the vascular surgery service (relative risk [RR] = 2.3) and liver transplantation (RR = 4.2) were significant risk factors for C. difficile acquisition. Patients asymptomatically colonized on admission had very low risk (1 in 44) for subsequent development of C. difficile diarrhea. In contrast, nine (47%) of 19 patients who acquired toxigenic strains developed C. difficile diarrhea, a finding suggesting that progression to diarrhea occurs early after acquisition or does not occur at all. The relatively high prevalence of culture positivity at admission may be characteristic of tertiary care hospitals and adds to the difficulty of controlling this nosocomial pathogen.