ABSTRACT: BACKGROUND:Escherichia coli is bacteria that exist as commensal in the intestine of animals and humans, but pathogenic strains cause disease in chickens. The development of antimicrobial resistance in E. coli is one of major concern worldwide. A cross-sectional study was conducted from November, 2015 to April, 2016 in and around Ambo town on backyard chicken with the objectives of isolating E. coli from selected visceral organs, assessment of potential risk factor and determination of antimicrobial resistance pattern of the isolates. RESULTS:The overall isolation rate of E. coli was 11.5% (80/694) [95% CI: 9.64-14.61] and 32.5% (62/191) [95% CI: 25.39-39.09] at organ and chicken level, respectively. E. coli isolation rate was 15.2% (29/191), 13.6% (27/191), 6.3% (12/191) and 10.7% (13/121) from spleen, liver, kidney and ovary samples, respectively. The multivariable logistic regression analysis revealed higher probability of E. coli isolation from adult (adjusted Odds ratio [aOR] =2.5, P?=?0.013) than younger chickens, from clinically sick chickens (aOR?=?3.0, P?=?0.003) than apparently healthy. E. coli isolates were 100% susceptible to ciprofloxacin, norfloxacin and sulfamethoxazole-trimethoprim followed by 89-63.4% susceptibility to gentamicin, streptomycin, ceftazidime, nalidxic acid, nitrofurantoin, kanamycin, amikacin and chloramphenicol. Whereas, 100% resistance was observed against cloxacilin, cefotaxime and amoxicillin, whereas 92.7 and 46.3% were resistant to cefuroxime, and tetracycline, respectively. Multidrug resistant (MDR) was observed in 78.1% (64/82) of the isolates which exhibited 5 different MDR patterns to 7 antimicrobial classes. CONCLUSIONS:Higher isolation rate of E. coli was observed from visceral organs of chickens. Age and health status were predictors of E. coli isolation. Remarkable numbers of the isolates are resistant to different antimicrobials and multidrug resistant E coli isolates are widespread in the area.