Identification of predominant SNPs as a novel method for genotyping bovine Staphylococcus aureus isolates

Staphylococcus aureus is a formidable pathogen of both human and animal. Infection often gives rise to an economic loss resulting from the extended cost of treatment and hospitalization for humans, and loss of usable agriculture animal products from infected animals and treatment regiments. We describe here a protocol for the amplification and sequencing of predominant single nucleotide polymorphisms within the promoter region of hla (encoding alpha-toxin) that confers a hyper-producing alpha-toxin phenotype to S. aureus isolates associated with chronic and severe bovine mastitis infections. We validated our findings with a second round of analysis, confirming the SNPs as a valid genotypic marker for alpha-toxin hyper-producing bovine isolates. The identification of highly virulent isolates will allow for aggressive treatment of the infection and limit the disease and economic impact. With readily available reagents and facilities, this protocol can be completed in as little as 72 h once samples are isolated.