PROJECT SUMMARYApproximately 40000 infants are born each year in the United States with congenital heart defects (CHD) andheart defects are the leading cause of birth defect-related deaths in the United States(1). While advances insurgical treatment and post-operative care have reduced in-hospital mortality for children born with CHD 1/3 ofinfants continue to suffer morbidity from post-operative multiple organ dysfunction syndrome (MODS)(2 3).Emerging data suggests a role for intestinal dysbiosis and a pro-inflammatory metabolome in the pathogenesisof post-operative MODS(4 5). In two case reports of adult ICU patients with MODS fecal microbiometransplantation to correct dysbiosis resulted in resolution of MODS. A multi-omics approach may provideinsight into host-microbe interactions contributing to MODS. In children with CHD intestinal microbiomedysbiosis is prevalent pre-operatively and in pilot data from Dr. Typpo's NIDDK K23 award project we foundthat post-operative MODS was associated with a pattern of dysbiosis previously reported in adult ICU patients:decreased relative abundance of Firmicutes and Actinobacteria and an increased relative abundance ofBacteroidetes and Proteobacteria(5). Through PICRUSt analysis to predict functional composition of thedysbiotic metagenome(6) we identified twenty-five metabolic pathways associated with MODS and oneprotective pathway. The goal of this R03 proposal is to complement and expand Dr. Typpo's K23 program ofresearch from pure structural analysis (what microbes are present) of intestinal microbial populations tofunctional analysis (what they are doing) of microbial populations via the following 3 specific aims. SpecificAim 1: Characterize the pre- and post-operative fecal microbiome in infants with and without post-operativeMODS using whole genome shotgun metagenomics sequencing to identify if pre-operative microbiome isassociated with post-operative MODS. Specific Aim 2 Identify differentially abundant fecal and urinarymetabolites of infants with and without post-operative MODS to identify associations between the pre-operativemetabolome and post-operative MODS. Specific Aim 3: Derive mechanistic models to explain how observedpre-operative dysbiosis and host-microbe interactions contribute to the development of post-operative MODSby combining clinical metadata metagenomics (Aim1) and metabolomics (Aim2) output. This proposal issignificant because we will address gaps in knowledge regarding the functional consequences of intestinaldysbiosis in children who develop MODS after surgical correction of CHD. This proposal is innovative throughuse of combined metagenomics and metabolomics approaches to derive mechanistic pathways whichcontribute to MODS. We may identify novel microbial targets to reduce the incidence or severity of post-operative MODS. This proposal in combination with Dr Typpo's career development award (K23) will provideher with the training experience and preliminary data to test interventions designed to target host-microbeinteractions and prevent post-operative MODS in a successful future R01 proposal.