Objective: To determine epidemiologic features of an Escherichia coli O157:H7 outbreak occurred in Xuzhou, Jiangsu Province, China in 1999, and assess the incidence of E. coli O157:H7 in diarrhea patients and host animals and its relationship with disease onset, and provide a scientific basis for establishing prevention and control strategies. Methods: Epidemiological, microbiological, and molecular methods were performed to identify risk factors and describe the ecology of E. coli O157:H7 in the environment. Results: From May to September, in 1999, 99 cases of E. coli O157:H7 infection were confirmed. Fifty-six patients were enrolled in the case-control study. Bad personal health habits and poor sanitary conditions in the kitchen were associated with increased risks of infection, whereas hand washing was protective. The household survey indicated that residents in the epidemic region during the outbreak had higher than expected rates of diarrhea. The total E. coli O157:H7 carrier rate in the livestock was 12.36% (22/178), specifically 19.15% in cattle, 12.50% in goat, and 11.11% in swine. Numerical analysis of pulsed-field gel electrophoresis (PFGE) profiles divided strains into two clusters with 77.5% homology. One cluster contained 11 strains isolated from diarrheal patients, foods, and animals. The other cluster comprised 10 strains from patients and environment. Conclusions: In a large outbreak of E. coli O157:H7 infection among predominantly elderly residents in Xuzhou, high rates of carriage of E. coli O157: H7 among host animals most likely resulted in contamination of the environment, thereby leading to the outbreak. Effective and preventive control measures should be taken to avoid contamination, including environmental and family health improvement, good personal hygiene, and safe food handling practices.