The prevalence of Escherichia coli O157 associated with feedlot cattle in Saskatchewan was determined in a 10-month longitudinal study (3 feedlots) and a point prevalence study (20 feedlots). The prevalence of E. coli O157 at the three different sites in the horizontal study varied from 2.5 to 45%. The point prevalence of E. coli O157 among Saskatchewan cattle from 20 different feedlots ranged from 0% to a high of 57%. A statistically significant (P = 0.003) positive correlation was determined to exist between the density of cattle and the E. coli O157 prevalence rate. A significant correlation (P = 0.006) was also found between the E. coli O157 percent prevalence and the number of cattle housed/capacity ratio. All 194 E. coli O157 isolates obtained were highly virulent, and random amplified polymorphic DNA PCR analysis revealed that the isolates grouped into 39 different E. coli O157 subtypes, most of which were indigenous to specific feedlots. Two of the most predominant subtypes were detected in 11 different feedlots and formed distinct clusters in two geographic regions in the province. Antimicrobial susceptibility testing of the E. coli O157 isolates revealed that 10 were multidrug resistant and that 73 and 5 were resistant to sulfisoxazole and tetracycline, respectively.