Staphylococcus aureus nasal carriage is a common source of nosocomial infection and colonization. Many S. aureus isolates showed resistant to methicillin all over the world and its prevalence as an endemic nosocomial pathogen has been increasing. The aim of the present study was to assess the burden of methicillin-resistant S. aureus nasal carriage, its association with factors of interest including its genetic relationships, and its antimicrobial susceptibility. This was a cross-sectional study that was carried out from March 2008 to September 2008 among hospitalized patients, patients’ relatives, and healthcare workers in all the wards of Hospital Universiti Sains Malaysia. The prevalence of S. aureus nasal carriage was found to be 28.7%. The highest numbers of MRSA nasal carriers were found in surgical wards (35.3%), followed by intensive care units (29%). This study showed that patients with a history of previous antibiotic intake within the previous month, nasogastric tube in situ, and longer hospitalization had a significantly high risk of being MRSA nasal carriers. The excess length of hospital stay was significantly (P=0.006) longer among MRSA carriers (mean±SD: 27±22.32 days) than methicillin-sensitive S. aureus carriers (13.8±16.73 days). The genetic relationship of all 34 nasal MRSA isolates revealed four major clusters of isolates, designated as pulsed-field type A–D. All MRSA isolates encountered in this study were resistant to penicillin, oxacillin, ciprofloxacin, gentamicin and erythromycin but sensitive to tigecyclin, linezolid, teicoplanin and vancomycin. The most significant associated factors with acquisition of MRSA nasal carriage were duration of hospitalization, antibiotic used, and nasogastric intubation, and there was a relationship between MRSA isolated from inpatients and healthcare workers. Thus, linezolid and tigecyclin might be used instead of the empirical antibiotics to which MRSA is resistant.