Objective: The aim of this study was to determine the antimicrobial susceptibility and cassette chromosome typing in Methicillin-Resistant Staphylococcus aureus (MRSA) isolated from healthcare workers. Methods:Two hundred thirty nasal swabs were collected from healthcare workers of three hospitals in Larestan, Iran. Antibiotic susceptibility was determined by disk diffusion method according to CLSI guideline. The minimum Inhibitory concentration (MIC) of vancomycin and screening test with methicillin were measured by E-test and agar screen plate procedures. Cefoxitin disc diffusion test was performed. MRSA isolates were selected and investigated for mecA gene and SCCmec typing by multiplex-PCR, then five methods were compared. Results:In total, 37 S. aureus were isolated, 28 were defined as MRSA. Frequency of SCCmec types were as nine SCCmec I, eight SCCmec IV, five SCCmec II, four SCCmec V and two SCCmec III. In this study, 75% of isolates were CA-MRSA, 25% were HA-MRSA. Evaluation of antibiotic resistance showed the greatest resistance to penicillin and the lowest resistance was observed to vancomycin and by E-test method 28.5% of isolates were intermediate resistance to vancomycin. However, screening test detected 92.8% resistance with oxacillin and 28 of isolates were resistant with Cefoxitin disc diffusion. For these 28 isolates mecA was positive. Conclusion:As a result, in our S. aureus isolates methicillin resistance was75.7%. The most frequent type was SCCmec I. Our result showed high rates of antibiotic resistance specially to methicillin in the S. aureus isolated hospitals that is a serious warning to the treatment of infection caused by this bacterium. J Microbiol Infect Dis 2018; 8(1):1-7