This application is a competitive renewal for 1R01HL103866, named “Gut flora metabolism of dietary carnitine and cardiovascular.” The overall proposal is predicated upon the recognition that gut microbiota contribute to both the development and adverse consequences associated with Cardiovascular Disease (CVD). Gut microbe dependent generation of trimethylamine (TMA), and its conversion in the host liver to trimethylamine N-oxide (TMAO), are recognized for being mechanistically and clinically linked to the development of CVD. During the past funding period, we investigated the microbial participants in conversion of dietary choline and phosphatidylcholine into TMA and TMAO, mechanisms through which TMAO contributes to CVD, and approaches for intervening and suppressing TMAO generation from choline. However, performance of these studies also revealed that microbial TMA/TMAO generation in some subjects, particularly omnivores, can originate from dietary carnitine. Moreover, the microbes and microbial enzymatic machinery involved in gut microbiota dependent TMAO elevation from either dietary carnitine, or a diet enriched in red meat, are distinct from the participants in choline/phosphatidylcholine conversion into TMA/TMAO. The overarching goal of this proposal is to better understand how gut microbial metabolism of dietary carnitine is linked to TMAO generation within a host, heightened CVD risks, and can serve as a therapeutic target for potential improvements in human health. The proposed research studies are multidisciplinary, and encompass a combination of basic, translational and human clinical investigations. In Aim 1 we will define the relevance of the metaorganismal carnitine → gamma-butyrobetaine (gBB) → TMA →TMAO pathway to CVD through both clinical and animal model studies. In Aim 2, we will explore therapeutic approaches for inhibiting dietary carnitine-dependent TMA and TMAO generation, and CVD-relevant phenotypes. Successful completion of the proposed studies will further reveal a role for gut microbes in CVD development and its major adverse complications and result in improved diagnostics and therapeutic approaches for the treatment and prevention of CVD.