Methicillin-resistant Staphylococcus aureus (MRSA) is one of the major causes of nosocomial infections in our hospital. Therefore, we aimed to characterize MRSA isolates phenotypically from patients with nosocomial infections at Cumhuriyet University Hospital between December, 1999, and June, 2001, in Sivas by analysis of antibiotic patterns and genotypically using pulsed-field gel electrophoresis (PFGE) and repetitive element sequence-based polymerase chain reaction (rep-PCR). Forty-three nosocomial isolates were collected from various wards. All isolates were resistant to penicillin, tetracycline, oxacillin, and gentamicin. By rep-PCR and by separation of SmaI fragments of genomic DNA using PFGE, one major type (eight subtypes with PFGE) was identified among the strains. This clone was found to be different than some clones such as Iberian, Brazilian, and a major clone that was found in another Turkish University Hospital in Ankara. According to our results, there is a major MRSA clone with a potential to spread in our hospital. Infection control measures should be directed toward restricting the further spread of this clone. Therefore, in accordance with these findings, a surveillance culturing program should be established.