The role of gut microbiota in human health and disease processes is increasingly recognized. Moreover, the gut microbiome is now recognized as a large endocrine organ that can generate biologically active metabolites, which upon absorption into the host, often have dedicated receptors, and impact physiological processes, and disease susceptibility. In this Project, we examine the role of specific gut microbiota pathways in cardiovascular disease (CVD) pathogenesis. We have employed untargeted metabolomics as a discovery platform, coupled with cellular and animal model studies, to identify additional/new potential gut microbiota- dependent pathways enhanced in type 2 diabetes mellitus that are both associated with, and potentially a contributor to CVD. Subjects with type 2 diabetes are at a disproportionately increased risk for development of CVD, and yet, the overall level of glycemic control is not necessarily related to CVD risks. There thus are metabolic pathways beyond the glucocentric view of diabetes that contribute to CVD in this at risk population. Large-scale clinical investigations are used to focus research attention on candidate metabolites whose circulating levels are reproducibly associated with incident development of adverse cardiovascular events like heart attack, stroke and death in this at risk population. After validating the associations (and often typically observing the candidate metabolite predicts risks in diabetic and non-diabetic subject alike), we move forward with mechanistic studies aimed at defining whether observed associations are linked to CVD and metabolic disease relevant phenotypes, through cellular and animal model studies. Preliminary cell, animal model and microbial transplantation related studies using genetically engineered human commensals (gain and loss of function mutants) are used to interrogate the role of specific microbiota derived metabolites, and the microbial enzyme(s) responsible for their generation, in eliciting relevant phenotypes in the host like enhanced thrombosis potential, or susceptibility to atherosclerosis. Our proposed studies promise to identify new mechanisms through which gut microbiota may contribute to CVD. They also will help improve identification of those at risk for CVD and its adverse events who otherwise might not be identified. They also will provide enabling discoveries that will foster potential development of novel treatments for CVD.