Abstract Compelling evidence has implicated human gut microbiota in development of obesity and related cardiometabolic disorders. Recent studies identified a group of circulating metabolites that are functionally related to gut microbiota. These microbial metabolites act as ‘transducers’ in the microbiota-host communications and ‘sensors’ in gut-organ/tissue crosstalk, and affect ectopic fat distribution in various metabolic organs and tissues. Diet is among the most important regulators of gut microbiota; however, no study has assessed how weight-loss diets may affect the long-term changes of the newly identified microbial metabolites. The proposed systems study aims to fill this significant research gap. In two of the longest and largest comparator trials on the popular weight-loss diets - POUNDS LOST and CENTRAL, we propose to identify novel plasma microbial metabolites associated with long-term weight loss in response to diet interventions. We will examine relations of plasma levels of identified microbial metabolites and ectopic fat distribution in the liver, heart, pancreas, and kidney, as well as adipose and skeletal muscle tissues. In addition, we will assess whether changes in plasma microbial metabolites mediate the relations between weight loss and changes in cardiometabolic markers. Moreover, we will examine the dynamic associations between microbial metabolites and changes in plasma miRNAs related to obesity and cardiometabolic risk. The findings from our study will considerably improve development of novel prevention and treatment strategies for obesity and related cardiometabolic disorders.