Staphylococcus aureus is an important zoonotic pathogen which has significant consequences for animal and human health. S. aureus is one of the many foodborne pathogens that causes food poisoning through its diverse enterotoxins. It is believed that S. aureus strains that infect different host species are genetically distinct although some strains are thought to be infective to wide range of host species. Some reports also showed the presence of distinct strains of S. aureus with specific tissue-tropism that are adapted to infect specific tissues such as the mammary gland. However, there are no clearly defined clonal patterns of S. aureus isolates that cause infection in any given species of animals or human. The objectives of this study are to (1) evaluate clonal diversity of S. aureus isolates from cases of bovine mastitis to determine dominant clones responsible for bovine mastitis; and (2) determine enterotoxin gene patterns of S. aureus isolates from cases of bovine mastitis. Milk samples from cases of bovine mastitis collected from various farms in the eastern Tennessee area for diagnosis were evaluated at Tennessee Quality Milk Laboratory (TQML) for causative agents. Overall, 111 S. aureus isolates from these cases were evaluated for toxins genes such as toxic shock syndrome toxin gene 1 (tsst1) and enterotoxin genes by each gene-specific primer pair using PCR. All 111 S. aureus isolates were evaluated for seA, seB, and seC, seE, seJ, and tsst1 using PCR. Negative and positive controls were used in each PCR reaction. The clonal diversity of S. aureus isolates from cases of bovine mastitis has been evaluated by pulsed field gel electrophoresis (PFGE). These data were analyzed using GLIMX model for logistic regression in SAS 9.4. Results showed that few isolates were positive for the enterotoxins tested. The genetic diversity analysis shows 16 PFGE types throughout 11 farms. One type stood out, with half of the population of isolates being present among 9 farms. These results suggest a negative pattern for toxins and indicate that one main type predominates in the areas tested, which could help area specific control of S. aureus mastitis.