Diarrhea has been recognized as a frequent health problem among children enrolled in day-care centers. Thus, we evaluated the effect of a handwashing program in two day-care centers (HWC) on the incidence of diarrhea among children when compared to children in two control centers (CC). After the program was begun, the incidence of diarrhea at the HWC began to fall and after the second month of the study was consistently lower than that at the CC. The incidence of diarrhea in the HWC was approximately half that of the CC for the entire 35-week study period. Adenoviruses, rotavirus, Giardia lamblia, and enteropathogenic Escherichia coli were found in the stools of a small number of ill children, but not pathogen was identified in the stools of most children with diarrhea. These results suggest that a handwashing program will probably prevent at least some of the diarrhea in day-care centers.