Project 1 will identify and assign functions to conserved genes of unknown function in Acinetobacter baumannii, an emerging pathogen that causes hospital-acquired, antibiotic resistant infections. The project will focus on protein-coding genes that contribute to any of seven clinically relevant resistance traits. We hypothesize that such genes control the expression and activities of efflux pumps, membrane permeability determinants, inactivating enzymes, and stress responses. Genes required for resistance to four antibiotics, two biocides and desiccation will first be identified by genome-scale mutant screening using Tn-seq technology. A subset of the resistance genes of unknown function will then be characterized using a battery of genetic, genomic and proteomic approaches. The results will be interpreted in the context of current understanding of each trait in order to formulate potential molecular functions for the genes examined. Hypothesized functions will then be tested using approaches tailored to individual genes.