Staphylococcus aureus (S. aureus) is a ubiquitous gram-positive pathogen that is one of the most frequent causes of skin and soft tissue infections. Frequently, these infections serve as a prelude to invasive life-threatening diseases. The prevalence of severe S. aureus infections have increased in healthy populations, associated with the development of both drug resistance and hypervirulence in methicillin-resistant S. aureus (MRSA) strains such USA 300. The spread of MRSA-induced disease during training and mission assignments is a well-documented threat to military public health. A wealth of both clinical and experimental evidence suggests that (1) neutrophils (PMNs) are essential mediators of anti-S. aureus host defense, and (2) emerging MRSA strains release toxins capable of killing PMNs, and consequently they can overwhelm host defense responses and rapidly cause disease in immune competent individuals. The rapidity of these events suggests a protective vaccine would be the best approach to disease control. Using a model of healed MRSA skin infection that elicits protective immunity against a second MRSA skin infection, we provide evidence that the appropriate induction of humoral and cellular immune responses can separately but synergistically support anti-MRSA effector responses and expedite pathogen clearance. The research plan described herein will test the hypotheses that active immunization against S. aureus is feasible, and that protective antigens include secreted bacterial virulence factors. The specific aims are designed to evaluate the pathogen and host-derived factors that are required to initiate and maintain ant-MRSA protective adaptive immune responses. In specific aim 1 we will identify antigens leading to protective immunity by systematically measuring the prophylactic effects of skin challenge with selected knockout strains of MRSA followed by rechallenge with WT MRSA. In aim 2, we will characterize T follicular helper and the germinal center responses that correspond with anti- MRSA protective immunity, and employ Langerin-DTR mice to delineate the skin DC requirements for these processes. In aim 3, we will characterize the cutaneous T cell signature corresponding with anti-MRSA protective immunity by measuring the kinetics and tissue distribution pathogen specific T cells following infection with strains of MRSA that have been engineered to express ova peptide OVA 323-339. After clarifying the adaptive T cell correlates of anti-MRSA protective immunity, we will use the Langerin-DTR system to elucidate the underlying DC requirements for their induction. Overall the program is designed to dissect the humoral and cellular immune components required for a protective immune response to MRSA. Once explored, these data will provide the basis to design and evaluate approaches to a preventive vaccine.