A cross sectional study was conducted in and around Motta town, East Gojjam zone, Ethiopia from November 2020 to May 2021 in order to isolate, identify and determine the antibiotic susceptibilities of S. aureus isolated from mastitic cows. A total of 109 lactating cows were examined to detect clinical and subclinical mastitis by physical examinations of the udder, milk, and California mastitis test, respectively. The total proportion of mastitis was 62 (56.88%). Out of this, 4 (3.67%) and 58 (53.21%) were clinical and subclinical mastitis respectively with significant difference (p<0.05). Staphylococcus aureus was isolated at a rate of 25 (40.32%) from mastitic cows, out of this 1(25%) and 24 (41.38%) in clinical and subclinical mastitic cows respectively. The result showed that the occurrence of Staphylococcus aureus was2.118 times more likely to occur in sub clinical than clinical mastitis (OR= 2.118). Among the potential risk factors that were found to be associated with the S. aureus mastitis, cross breed (OR=2.226), old age (OR= 3.055), parity with many calves (OR= 3.120), late lactation stage (OR= 3.391), poor housing conditions (OR= 8.889), previous mastitis history (OR= 0.227) and sub clinical mastitis type (OR= 2.118) had high effect on the occurrence of S. aureus mastitis. However, breed and mastitis type had not been a significant potential risk factor for the occurrence of S. aureus mastitis (p<0.05). The present study showed that S. aureus was totally not susceptible to penicillin (0%) and intermediate susceptible to tetracycline (50%).However, S. aureus was found to be highly susceptible to gentamycin (93.75%), erythromycin (84.38%), clindamycin (90.36%) and trimethoprim (78.13%). In conclusion, Mastitis was found to be one of the major constraining factors that affect milk production in dairy cows in the study area. S. aureus is the major etiological agent of mastitis especially in sub clinical mastitis. Various risk factors that are listed above were found to be critical for the occurrence of S. aureus mastitis. Also S. aureus was found to be resistant to penicillin. Key words: Antibiotic susceptibility, Clinical mastitis, Mastitic cows, Risk factors, Subclinical mastitis, S. aureus