Acinetobacter baumannii, a Gram-negative bacterium, has emerged as an important opportunistic pathogen that poses a significant public health risk in the United States. Clinically, A. baumannii is emerging as a leading nosocomial pathogen, particularly in intensive care units specializing in respiratory care, trauma and burns. Despite having the complete genome sequences for three strains of A. baumannii, the identity of most genes that are essential for pathogenesis in a mammalian host is not known. We propose to identify which of the nearly 4000 bacterial genes are essential for survival in a mammalian host and to demonstrate the necessity for each gene during experimentally induced bacteremia. The result will represent a major step forward in understanding the pathobiology of A. baumannii infections and for devising strategies to treat these infections or vaccinate against their occurrence. Our long-term goal is to systematically establish a molecular model of pathogenesis for A. baumannii infection. The objective of this application is to identify all genes essential to establish infection within a mammalian host. Based on a successful pilot experiment with 109,000 mutant of A. baumannii ATCC17978 in a murine model of bacteremia, our central hypothesis is that genes of A. baumannii essential for development of bacteremia can be determined by combining global transposon mutagenesis, a murine model of infection, and high throughput sequencing. Our rationale for conducting this work is to establish a framework that will direct detailed mechanistic investigations of A. baumannii pathogenesis in compromised patients. Using a saturating transposon library of A. baumannii mutants, the animal model of bacteremia, and high- throughput DNA sequencing, we will accomplish the following specific aims: 1) identify virulence genes essential for A. baumannii to colonize and disseminate within a mammalian host; and 2) confirm A. baumannii virulence genes that contribute most significantly to colonization and dissemination in the murine model using defined mutations. This contribution will be significant because, in these comprehensive and unbiased experiments, we will determine for the first time the complete A. baumannii virulence gene set for two strains during a mammalian infection. This proposal should be considered innovative as it applies transposon-directed insertion-site sequencing (TraDIS) for genome-wide identification of A. baumannii virulence genes using a forward-genetic screen in a relevant animal model of infection.
The proposed research involving the determination of a complete virulence gene set of Acinetobacter baumannii in a mammalian model is relevant to public health because it focuses on an emerging bacterial pathogen that has gained a foothold in hospitals and convalescence centers and has developed multiple antibiotic resistances. The research community at large would benefit from the identification of the A. baumannii virulence gene set, to which they can apply their respective areas of expertise to tackle this overwhelming medical scourge. The proposal is relevant to NIH's mission because it fosters fundamental discovery using an innovative research strategy that improves the nation's capacity to protect and improve health.
