BACKGROUND: External eye infection caused by bacteria can lead to reduced vision and blindness. Therefore, pathogen isolation and antimicrobial susceptibility testing are vital for the prevention and control of ocular diseases. OBJECTIVE: The main aim of this study was to assess bacterial isolates, their antimicrobial susceptibility pattern, and associated factors of external ocular infection (EOI) among patients attended eye clinic at Debre Markos Comprehensive Specialized Hospital (DMCSH), Northwest Ethiopia. METHODS: We conducted a cross-sectional study in patients with external ocular infections from January 1, 2021, to June 30, 2021, at DMCSH. Socio-demographic and clinical data were collected using semi-structured questionnaires. Following standard protocols, external ocular swabs were collected and inoculated onto blood agar, chocolate agar, MacConkey agar and mannitol salt agar (MSA). Finally, bacterial isolates were identified by Gram stain, colony morphology, and biochemical tests. Antimicrobial susceptibility testing was done by using the modified Kirby-Bauer disk diffusion technique according to Clinical and Laboratory Standards Institute (CLSI) guideline. Cleaned and coded data were entered into EpiData version 4.2 software and exported to Statistical Packages for Social Sciences (SPSS) version 22 for analysis. Bivariate logistic regression was applied to investigate the association between predictors and outcome variables. P-values ≤ 0.05 with 95% confidence interval were considered statistically significant. RESULTS: Two hundred seven study participants were enrolled in this study. More than half of them (57.5%, 119/207) were males, and 37.7% (78/207) of them were ≥ 65 years old. A total of 130 (62.8%) bacterial isolates were identified, with Gram-positive bacteria accounting for 78.5% (102/130) of the isolates. Staphylococcus aureus was the most common isolate with a 46.2% (60/130) prevalence. Ciprofloxacin was comparatively effective against Gram-positive and Gram-negative bacteria. The prevalence of culture-confirmed bacteria was significantly associated with age groups 15-24 (AOR: 9.18, 95%CI: 1.01-82.80; P = 0.049) and 25-64 (AOR: 7.47, 95%CI: 1.06-52.31; P = 0.043). Being farmer (AOR: 5.33, 95% CI: 1.04-37.33; P = 0.045), previous history of eye surgery (AOR: 5.39, 95% CI: 1.66-17.48; P = 0.005), less frequency of face washing (AOR: 5.32, 95% CI: 1.31-7.23; P = 0.010) and face washing once a day (AOR: 3.07, 95% CI: 1.13-25.13; P = 0.035) were also significantly associated with the prevalence of culture-confirmed bacteria. CONCLUSION: The prevalence of culture-confirmed bacteria among patients with EOI was high in the study area. A considerable proportion of bacterial isolates exhibited mono and/or multi-drug resistance. Age (15-64 years), being farmer, previous history of eye surgery and less frequency of face washing were significantly associated with the prevalence of culture-confirmed bacteria. Bacterial isolation and antibiotic susceptibility testing should be routinely performed in the study area to combat the emergence of antibiotic resistance.