Samples from cattle, other domestic and wild animals, flies, feeds, and water-troughs were collected from 12 cattle farms and tested for Escherichia coli O157. E. coli O157 was isolated from bovine fecal samples on all 12 farms with a within herd prevalence ranging from 1.1% to 6.1%. E. coli O157 was also found in 1 of 90 (1.1%) equine fecal samples, 2 of 65 (3.1%) canine fecal samples, 1 of 200 pooled bird samples (0.5%), 2 of 60 pooled fly samples (3.3%), and 10 of 320 (3.1%) water-trough sample sets (biofilm and water). No E. coli O157 were isolated from 300 rodents, 33 cats, 34 assorted wildlife, or 335 cattle feed samples. Indistinguishable pulsed-field gel electrophoresis patterns of XbaI digested chromosomal DNA and Shiga toxin types were observed for bovine and water-trough isolates from two farms and for one equine and two bovine isolates from one farm.