Project Summary/Abstract Most bacteria are found living within complex polymicrobial environments where they must compete for nutrients, space, oxygen, and defend themselves against exogenous, host, or bacterial derived antimicrobials. Bacterial community dynamics and behavior can be greatly influenced by reactive nitrogen species (RNS), which are important signaling molecules, immune mediators, and antimicrobials. Unfortunately, there is limited understanding on how RNS regulate the structure and function of polymicrobial communities and what impact this interplay has on host immunity, which has created enormous gaps in knowledge. Previous studies that explore the role of RNS on microbial physiology predominantly use single species models instead of polymicrobial models, which better represent the lifestyle of microbes. Our preliminary findings indicate that commensal or beneficial bacteria induce the production of RNS to outcompete pathogenic bacteria and maintain homeostasis within a host. Yet, there are no studies that have clearly defined the molecular mechanisms that govern synergy between commensal bacteria and RNS, and how they dictate the pathogen- host relationship. Over the next five years, the studies outlined in this MIRA application will use molecular genetics, systems biology tools (metabolomics, proteomics, and RNA sequencing), and a Drosophila melanogaster invertebrate model to determine how RNS regulate interactions between commensals and pathogens and also elucidate what impact this has on the host. Critically the field of RNS as an effector of bacterial population dynamics is vastly understudied yet is vital to understand because it has significant health consequences. The findings of this research proposal will advance the field of bacterial-bacterial interactions, as well as the role of RNS responses in controlling and modulating the host-pathogen interface. Our studies could potentially provide valuable mechanistic data that can be harnessed for the development of commensal- mediated anti-infection therapies.