This application seeks to address the impact of bacterial colonization and persistence in chronic wounds. The formation of biofilms has clearly been linked to chronic and persistent bacterial infections. This considerably delays and complicates wound healing. Unlike acute bacterial infections, which are often cleared by the host, biofilm-related chronic infections are not easily resolved even with high dose antibiotics and intact immunity. The bacterial pathogens Pseudomonas aeruginosa and Staphylococcus aureus, which are the focus of this application, cause an array of biofilm-related clinical diseases including persistent airway infections, burn wound infections, endocarditis, and surgical site infections. Unresolved infected wounds also contribute to nosocomial persistence and the spread of bacteria in health care settings. The abundance and persistence of chronic infections due to biofilm formation has led to the hypothesis that biofilms deploy directed mechanisms to subvert recognition, activation, and functions of the host immune system. This proposal will use state-of-the- art molecular and genomic approaches to better understand the impact of bacterial colonization in chronic wounds. Aim 1 will define mechanisms by which P. aeruginosa and S. aureus resist killing by host-derived antimicrobials and innate immune cells. Aim 2 will utilize a newly developed porcine full thickness burn chronic wound model and explanted human tissue from burn wound debridement to investigate biofilm persistent infections caused by P. aeruginosa and S. aureus. The development of a chronic infection model that can be sampled over time as well as access to materials derived from humans suffering burn-wound injury provides a relevant, unique, and novel approach for examining the effects of biofilm formation in the host. Conclusions made from in vitro data will be efficiently tested in the porcine model and human tissue for applicability in clinical infection. Ultimately information gathered will aid the treatment of an array of chronic infections including highly prevalent persistent wound infections. PUBLIC HEALTH RELEVANCE: Pseudomonas aeruginosa and Staphylococcus aureus are versatile opportunistic pathogens that can cause devastating persistent wound infections. Within the wound, these organisms transform into a resistant community called a biofilm. Biofilm-related chronic infections are not easily resolved even with high dose antibiotics and intact immunity. This application will use molecular and genomic approaches to define how biofilm-growing bacteria resist antibiotics and host immune cells. This will allow us to better understand the impact of bacterial colonization in chronic wounds and aid future management and treatment of chronic infections that significantly impact human health.