Objective-To evaluate antimicrobial susceptibility of commensal Escherichia coli strains isolated from the feces of horses and investigate relationships with hospitalization and antimicrobial drug (AMD) administration. Design-Observational study. Animals-68 hospitalized horses that had been treated with AMDs for at least 3 days (HOSP-AMD group), 63 hospitalized horses that had not received AMDs for at least 4 days (HOSP-NOAMD group), and 85 healthy horses that had not been hospitalized or treated with AMDs (community group). Procedures-Fecal samples were submitted for bacterial culture, and up to 3 E coli colonies were recovered from each sample. Antimicrobial susceptibility of 724 isolates was evaluated. Prevalence of resistance was compared among groups by use of log-linear modeling. Results-For 12 of the 15 AMDs evaluated, prevalence of antimicrobial resistance differed significantly among groups, with prevalence being highest among isolates from the HOSP-AMD group and lowest among isolates from the community group. Isolates recovered from the HOSP-AMD and HOSP-NOAMD groups were also significantly more likely to be resistant to multiple AMDs. Resistance to sulfamethoxazole and resistance to trimethoprim-sulfamethoxazole were most common, followed by resistance to gentamicin and resistance to tetracycline. Use of a potentiated sulfonamide, aminoglycosides, cephalosporins, or metronidazole was positively associated with resistance to 1 or more AMDs, but use of penicillins was not associated with increased risk of resistance to AMDs. Conclusion and Clinical Relevance-Results suggest that both hospitalization and AMD administration were associated with prevalence of antimicrobial resistance among E coli strains isolated from the feces of horses.