PROJECT SUMMARY Melioidosis is an often-fatal infection caused by inhalation, inoculation, or ingestion of the Gram-negative facultative intracellular pathogen and Tier 1 select agent Burkholderia pseudomallei (Bps). Bps has recently been isolated from the soil in the southern United States. Worldwide, 165,000 cases of melioidosis are estimated to occur each year; 85,000 (52%) of these patients die. Pneumonia is present in over 50% of melioidosis cases and more than doubles the risk of death. Yet, to develop novel, targeted therapeutics necessitates a deeper understanding of pulmonary and systemic mechanisms of host defense. Our team combines expertise in human and experimental melioidosis, pulmonary host defense, sepsis, and bioinformatics. We have developed a robust murine model of Bps pneumonia displaying mild and severe disease phenotypes. In parallel we have performed unbiased multi-omics analyses on a large prospective cohort of hospitalized patients with infection in NE Thailand with the goal of classifying melioidosis cases and understanding the host response to Bps. We identified distinct transcriptional and metabolomic profiles associated with melioidosis compared to other infected patients, and have built robust classifiers in each omics domain to predict death in human melioidosis. However, to comprehensively investigate mechanistic underpinnings requires a tractable experimental model with sufficient comparability to human disease. Moreover, to study the lethality of respiratory melioidosis requires sampling of lung tissue. We hypothesize that applying a comparative multi-omic approach to mice and humans with respiratory melioidosis will both a) yield critical insights into the pulmonary host defense mechanisms that fail to contain the infection and contribute to severe outcomes and b) establish comparability of the experimental murine model with human infection. We submit the following specific aims: 1). Define the temporal trajectory of multi-omic features of systemic host defense in murine respiratory melioidosis and identify perturbations representing success or failure of host defense. 2) Define lung cell-specific transcriptomic changes in murine respiratory melioidosis and identify signals that are associated with success or failure of pulmonary host defense. 3) Identify shared multi-omic signatures between murine and human respiratory melioidosis. The results of these studies will generate a rich compendium of data about the systemic and pulmonary host response to murine respiratory melioidosis and provide novel and comprehensive insights into the heterogeneity and key biological pathways underlying failed host response phenotypes of melioidosis pneumonia. Intersecting these findings with existing human melioidosis data will help to define clinically relevant targets for further investigation while simultaneously providing essential information about advantages and limitations of the animal model in recapitulating human infect...