Staphylococcus aureus (S. aureus)is a significant foodborne bacterial pathogen resulting in food intoxication and other numerous human infections. The purpose of the present study was to ascertain the prevalence, virulence markers and antibiotic resistance genes of S. aureus isolates collected from some fish products and their handlers at retail markets in Cairo Governorate, Egypt. A total of 75 different fish products including sardine, herring and canned tuna (25 of each) and 100 hand swabs of fish handlers were screened for the presence of S. aureus utilizing 16S rRNA, nuc and clfA primers. The S. aureus strains were subsequently assessed for the potential of virulence (sea, seb, tst, pvl and lukM) and antimicrobial resistance (mecA, tetK and ermC) using particular primers. The findings showed that sardine had the highest bacterial count (4.25± 2.9log10 cfu/ g), followed by herring (3.79± 2.4log10 cfu/ g) and canned tuna (3.8± 2.7log10 cfu/ g). In addition, it was found that 8 out of 75 fish samples (10.7%) and 12 out of 100 fish handlers' specimens (12%) possessed coagulase-positive S. aureus. For fish samples, the molecular analysis revealed that all the examined S. aureus strains harbored pvl and tet K genes, while sea and tst genes were detected in 12.5% (1/8) of the recovered isolates. On the other hand, other genes such as seb, lukM and ermC were not identified. Concerning human specimens, the findings revealed that all the obtained S. aureus strains carried sea and tetK genes, while pvl, seb, ermC and tst genes were identified in 83.3 (10/ 12), 33.3 (4/ 12), 33.3 (4/ 12) and 8.3% (1/ 12) of the examined isolates, respectively. Interestingly, all isolates obtained from the fish handlers were categorized as methicillin-resistant S. aureus (MRSA), whereas no MRSA isolates were detected from the fish. The current study supported the existence of virulent antimicrobial-resistant S. aureus in the tested fish products and their handlers, generating a serious risk to food safety. Accordingly, strict preventive and control strategies should be implemented to tackle this major food safety issue.