The proportion of fecal samples culture-positive for Escherichia coli O157:H7 was determined for samples collected from 296 beef cows on pasture in a single Florida herd in October, November, and December 2001. The overall proportion of samples that cultured positive was 0.03. The proportion of cows that were culture-positive on at least one occasion was 0.091. No effect of pregnancy status or nutritional regimen on the proportion of culture-positive samples for E. coli O157:H7 was detected. We detected a breed effect on the shedding of E. coli O157, with Romosinuano cows having a lower (P < 0.01) proportion of samples culture-positive than Angus or Brahman cows. This difference might have resulted from the presence of confounding variables; however, it also might represent evidence of breed-to-breed genetic variation in E. coli O157 shedding. Further research is warranted to evaluate breed as a possible risk factor for shedding of this important foodborne pathogen. Further substantiated findings could indicate that breed is a cow-calf-level critical control point of E. coli O157:H7.