Staphylococcus aureus is a leading cause of surgical site infections (SSIs). The association between S. aureus genotypes and the severity of illness is, however, incompletely understood. The aim of the study was to genotype S. aureus isolates from deep SSI in orthopaedic patients to identify molecular markers associated with invasive S. aureus infections. DNA microarray analysis was performed on S. aureus isolates collected from 60 patients with deep SSI following major orthopaedic surgery, while 57 isolates from nasal carriers served as controls. Genes associated with antibiotic resistance, adhesion, immune evasion, tissue invasion and toxin production were detected. The bone sialoprotein-binding protein gene (bbp) was more frequent in isolates from SSI patients compared to nasal carriers (95.0% vs. 82.5%), suggesting a role in invasive disease. No major differences in other molecular virulence markers could be distinguished among isolates from the two clinical groups, suggesting that any S. aureus strain may cause invasive infection. Our study reveals important genotypic information on isolates obtained from deep SSI following orthopaedic procedures.