Bovine mastitis is the most important disease affecting the global dairy industry. Mastitis can be caused by an array of different bacterial pathogens including Staphylococcus aureus and Streptococcus agalactiae. Infections can be subclinical or clinical and are often chronic in nature and difficult to treat due to production of biofilm, an intracellular tropism and antibiotic resistance. Our previous work has identified pathogenic clones of S. aureus specialized for intramammary infection of dairy cows, that have evolved via host-switching events from humans followed by adaptive evolution by gene acquisition and diversification. We have identified adaptive traits associated with the ability of bovine S. aureus and S. agalactiae strains to survive and proliferate in the dairy niche including enhanced growth associated with induction of curdling and the capacity to produce a novel biofilm in the presence of milk. Here, we will employ complementary approaches including population genomics, transcriptomics, transposon mutagenesis and proteomics to determine the genetic and functional basis for these dairy-adaptive traits. In addition, we will employ an array of assays and a bovine mammary epithelial cell infection model to examine the relevance of the novel biofilm phenotype to pathogenesis and resilience to antibiotic treatment. An understanding of the mechanisms involved may inform the development of novel therapeutic approaches targeting key adaptations required for survival. This hypothesis will be tested using a specific antibody approach and newly discovered effectors produced by other bacterial species in the dairy bovine microbiota. These data will result in the comprehensive characterisation of a novel dairy adaptive phenotype from its evolutionary origin to pathogenic function, and therapeutic potential.