Accumulating evidence from recent studies links the gut microbiota to obesity, and microbiome therapy has been examined as a treatment.Clostridium butyricum(C. butyricum), an intestinal symbiont, protects the host from a range of diseases. Studies have shown a negative correlation between the relative abundance ofC. butyricumand a predisposition for obesity. However, the physiological function and material basis ofC. butyricumfor obesity are unclear. Here, fiveC. butyricumisolates were administered to mice on a high-fat diet (HFD) to determine their anti-obesity effects. All isolates suppressed the formation and inflammation of subcutaneous fat, and the two effective strains considerably reduced weight gain and ameliorated dyslipidemia, hepatic steatosis, and inflammation. These positive effects were not achieved by increasing the concentration of intestinal butyrate, and the effective strains could not be replaced by sodium butyrate (NaB). We also discovered that oral supplementation with the two most effective strains changed the metabolism of tryptophan and purine and altered the composition of the gut microbiota. In summary,C. butyricumimproved the metabolic phenotypes under the HFD by controlling the composition of the gut microbiota and modulating intestinal metabolites, thereby demonstrating its ability to fight obesity and providing a theoretical foundation for microbial preparations production.