<p>Constipation is a common gastrointestinal disorder, often associated with dysbiosis of the intestinal microecology. The treatments for constipation are various, and microbiome-based therapies provide supplementary or alternative options for its management. This study compared the therapeutic effects of three microbiome-based therapies, including probiotics, prebiotics, and traditional Chinese medicine (TCM), with the agonist mosapride on loperamide-induced constipation in mice, focusing on their restorative effects on intestinal motility and intestinal microecology. The results showed that mosapride had the most significant improvement in gut motility and key motility hormones, and also significantly restored gut microbiota. Probiotics significantly improved the levels of the motility hormones motilin (MTL) and substance P (SP) and significantly altered the composition of the microbiota. TCM significantly increased gut motility and was most effective in improving gut microbiota composition and short-chain fatty acid (SCFA) content. Inulin, on the other hand, had the least effect on defecation phenotype, intestinal motility, and restoration of intestinal microecology. Overall, this preclinical study in mice provides experimental insights into the therapeutic efficacy and potential mechanisms of these methods, which may contribute to the appropriate selection of candidate therapies for human constipation subtypes characterized by motility-microbiota axis dysfunction. In addition, the research highlights the critical importance of gut microbiota and their metabolites in the treatment of constipation and their potential as therapeutic indicators.</p>