Background: Methicillin-resistant Staphylococcus aureus (MRSA) has been considered as an important pathogen with a variety of virulence factors in communities and hospitals worldwide. Objectives: In this study, we focused on the detection of different virulence factors and enterotoxin genes of MRSA strains isolated from a referral hospital in Tehran, Iran. Moreover, the presence of different prophage types was studied. Methods: A total of 491 MRSA strains were isolated during three years from a referral hospital in Tehran. The staphylococcal enterotoxin (sea-seq) and pvl, hlb, sak, eta and tst genes were detected. A multiplex-polymerase chain reaction (PCR) assay was used for prophage typing of MRSA isolates. Results: Totally, 11 enterotoxin and 5 virulence factor genes were detected in MRSA strains. The sea, sek, seq, and hlb genes were present in all the MRSA and other enterotoxin genes. sel, seg, sem, sei, sen, seo, sec and sep were detected in 32.8, 20.3, 12.6, 8.3, 4.1, 2.6, 1.6 and 0.4 of the strains, respectively. A total of 93, 81, 15.9 and 5.7 of the strains harbored the sak, eta, tst and pvl genes, respectively. SGF, SGFa and SGFb proghage type genes were detected in 100 of the MRSA strains, and four different prophage patterns were identified among the strains. Conclusions: The presence of different prophage-encoded virulence factors among MRSA strains enable MRSA to produce a broad range of diseases, indicating MRSA strains as a potential threat to patients’ health. © 2018, Archives of Clinical Infectious Diseases.