Background There is growing evidence to implicate Staphylococcus aureus ( S. aureus) in the pathogenesis of recalcitrant chronic rhinosinusitis (CRS). Our group has demonstrated the ability of S. aureus to internalise within mast cells in nasal polyps and this may mediate disease recalcitrance. We investigated carriage of virulence genes in CRS-related S. aureus strains and its influence on the bacteria’s ability to localise and survive intracellularly. Methods S. aureus strains isolated from non-CRS controls (n=5), CRSsNP (n=4) and CRSwNP patients (n=4) were sequenced using short read paired sequencing and interrogated for carriage of virulence genes. A representative control and CRSwNP isolate were tested for intracellular survival in the LAD2 mast cell line to investigate phenotypic differences. Results Fifty percent of the CRSwNP group had deletion of the hld gene which may promote small colony variant formation or reduced enterotoxin production, and seventy-five percent expressed virulence genes associated with invasive disease. The CRSwNP isolate had a superior ability to localise intracellularly at 6 and 9 hours and showed a higher burden of S. aureus colony forming units at 24 hours. Conclusions When compared to non-CRS controls, CRS-related S. aureus strains demonstrate increased carriage of virulence genes. This appears to facilitate intracellular localisation of the bacteria conferring a survival advantage and enhancing pathogenicity. The latter may be partly due to a reduction in enterotoxin production and the acquisition of serine proteases splA and B and leukocidins E/D genes. This pathogenic S. aureus phenotype may manifest clinically with disease recalcitrance and refractoriness to antibiotics.