The gut microbiota has been linked with chronic diseases in humans such as obesity and diabetes. However, the demonstration of causality between constituents of the microbiota and specific diseases remains an important challenge in the field. In this presentation, using Koch’s postulates as a conceptual framework, I explore the chain of causation from alterations in the gut microbiota, particularly the endotoxin-producing members, to the development of obesity in both rodents and humans. Three components are essential for identifying the causative agents of obesity in the human microbiota: 1) microbiome-wide association studies; 2) isolation of the putative agents and disease reproduction in gnotobiotic animals; 3) mechanistic analysis of host responses to establish the molecular chain of causation. We employed this strategy in dietary therapy of morbid obesity/diabetes in human clinical trials to show that specific bacterial phylotypes, which are more relevant with MetS, can be identified, isolated and demonstrated in gnotobiotic models to be causatively contributing to MetS development in humans. For example, one endotoxin-producing bacterium was isolated from a morbidly obese human’s gut and demonstrated to be able to induce obesity and insulin resistance in germfree mice.