As Staphylococcus aureus strains evolve and gain resistance to antibiotics, the risk of bidirectional transmission of resistant strains between humans and animals increases. The objective of this study was to identify and type S. aureus among wild rodents in Boston, Massachusetts, to examine their genetic relationship to common human and animal isolates. A total of 168 bacterial isolates collected from 45 Brown rats (Rattus norvegicus) in Boston proper were analyzed. Polymerase chain reaction was used to detect the mecA and PVL genes. All S. aureus isolates were spa typed. A subset of isolates was characterized via multi-locus sequence typing (MLST). All S. aureus isolates were tested for antibiotic susceptibility. Overall prevalence of S. aureus was 11.9% (20/168). Of all S. aureus, 5.0% (1/20) were MRSA (based on detection of the mecA gene which encodes methicillin-resistance) and 95.0% (19/20) were methicillin-susceptible S. aureus (MSSA). All isolates tested resistant to benzylpenicillin. Two isolates were resistant to erythromycin and one isolate was resistant to four antibiotics, including oxacillin. Of the 20 isolates, 75.0% (15/20) were spa type t933. MLST results to date show that these are sequence type (ST) 1094. Additional molecular testing is ongoing. Our results indicate that wild rats from Boston, MA are carriers of S. aureus. Additional study is needed to examine the distribution of t933/ST1094, an uncommon strain previously found in ewes in Tunisia. Further research is warranted to identify and characterize lineages of S. aureus strains in order to minimize the risk of Staphylococcal infection from city rodents.