Dental caries is defined as a dynamic diet microbial disease of teeth, which results in localized dissolution and destruction of the mineralized tissues of the teeth. Dental caries develops when there is a susceptible tooth exposed to pathogenic bacteria in the presence of substrate. Under these conditions, the bacteria metabolize substrate to form acid, which decalcifies teeth. Dental caries is among the top oral health problem in both developing and developed nations affecting around 20-50% of the population globally. This study was conducted to assess the magnitude, associated factors, and antimicrobial susceptibility pattern of bacterial isolates among adult dental caries patients visiting Hiwot Fana specialized university hospital dental clinic from April 23 to-June 23, 2021. An institutional-based cross-sectional study was conducted among 290 study participants. Convenient sampling techniques were used to select the study participants. Data was entered into Epi-info version 7.2.4.0 and exported to Statistical Package for the Social Sciences version 20 for analysis. The result was explained by using summary measures of texts, tables, and graphs after analysis by using bivariate and multivariate logistic regression. Statistical significance was defined at a p-value of less than 0.05. The overall magnitude of bacteria among dental caries patients was 68.3%. S mutans 74(37.4%) and Lactobacillus spp 58(29.3%) were the most predominant isolated bacteria. Lack of teeth brushing (AOR: 2.8, 95% CI:1.6, 4.6), the habit of chewing khat always (AOR:4.8, 95%CI:2.10,8.80), the habit of chewing khat sometimes (AOR: 3.8: 95% CI: 2.520,9.48) and consumption of soft drinks (AOR: 1.9, 95% CI:1.2,3.1) were significantly associated with bacterial dental caries. Almost, all bacterial isolates were susceptible to ceftriaxone and ciprofloxacin compared to Amoxicillin, Azithromycin, chloramphenicol, clindamycin, doxycycline, erythromycin, gentamycin, penicillin, tetracycline, vancomycin and tobramycin. Teeth brushing habit, consumption of soft drink and a habit of chewing khat affects dental health and they are associated with bacterial dental caries. Harari regional health bureau better to focus by giving health education to the community about dental caries based on identified associated factors with dental caries.