Bovine mastitis (BM) has been causing great economic and financial losses in the dairy industry worldwide and one of its major pathogenic agents is S. aureus. BM treatment still relies on antibiotics. The extensive use of antimicrobials gives rise to bacterial strains such as methicillin-resistant S. aureus (MRSA) and mupirocin-resistant S. aureus (MuRSA). This study aimed to investigate and compare the prevalence of MRSA and MuRSA and antimicrobial susceptibility in S. aureus isolates from bovine mastitis milk in conventional and organic dairy farms. Milk samples were taken from mastitis-infected cattle in 6 conventional and 7 organic dairy farms from South Korea. According to the Food Codex, 163 suspected S. aureus colonies were isolated from the pooled milk samples from each farm. Further rapid coagulase test confirmed 11 out of 74 isolates from 4 conventional farms (CF1, CF2, CF3, CF4) while 17 out of 89 isolates from 3 organic farms (OF1, OF2, OF3) exhibited coagulase activity. Multiplex PCR amplification showed that the nuc gene marker for S. aureus was found in all coagulase-positive isolates from OF 1 and at least one isolate from CF1, CF2, CF3, and CF4. Conversely, only 2 isolates from CF2 contained mecA gene for MRSA while none had the mupA gene for MuRSA. Antimicrobial susceptibility testing revealed that conventional farm isolates were more resistant to antibiotics particularly, against ampicillin and tetracycline, and, in turn, could imply a potential to develop multidrug resistance if stringent measures to control antimicrobial use in dairy farms is not implemented properly.