It was gradually realized that establishing and maintaining a beneficial balance between intestinal microbiota and the human body are necessary for the normal functions of the intestine9. The dysbiosis will contribute to the pathogenesis of functional gastrointestinal disorders10. Therefore, it is essential to study the role of intestinal microbiota from STC patients in gut motility. In this study, the enrolled STC donors received FMT effectively, and the symptoms in STC patients were improved during follow-up. Next, pseudo-germ-free mice receiving fecal microbiota from STC donors were more likely to present constipation symptoms, including the hardness of stools, infrequent passage of stools, and poor gastrointestinal motility. In addition, microbiota metabolites in mice from STC donors were also changed in SCFAs and BAs. After supplementation with butyrate and DCA, some symptoms in mice from STC donors were reversed. This might be attributed to the disturbed intestinal microbiota that could change the host metabolism and affect gastrointestinal motility. Therefore, we concluded that aberrant gut microbiota composition in constipation might affect gut motility by altering host metabolism.

Commensal bacteria play a key role in the development and maintenance of intestinal sensory and motor functions31. Studies on germ-free mice showed that gastric emptying and gastrointestinal transit were significantly delayed compared with those in conventionally raised mice31. Bacteroides thetaiotaomicron was reported to stimulate gut motility by increasing the expression of γ-aminobutyric, vesicle-associated protein-33 and enteric γ-actin32. Meanwhile, Lactobacillus acidophilus and Bifidobacterium bifidum could accelerate intestinal transit by releasing neuromessengers, while Micrococcus luteus and Escherichia coli had an inhibitory effect33. After antibiotics, mice in the weekly fecal transfer treatments were fed by gavage for 8 weeks with fecal material from their respective donors. Using mice from STC donors, we herein showed that the symptoms of constipation could be replicated in mice. Recently, a growing body of work has implicated that the intestinal microbiota of constipated patients is altered11,12,13,14; for example, unclassified_Ruminococcaceae, Alistipes, and Oscillibacter were negatively correlated with the mean stool frequency34. Additionally, it has also been proven that human microbiota could promote the biosynthesis of 5-HT from colonic ECs to modulate gastrointestinal motility23. Therefore, our study indicated that normal microbiota might exert modulatory effects on gut motility.

A compelling set of relationships between microbiota metabolism and host physiology has emerged. SCFAs derived from the microbial fermentation of dietary fiber can directly inhibit histone deacetylases, activate G-coupled-receptors, and serve as energy substrates, thus affecting various aspects of physiological processes35. It has been reported that SCFAs stimulate the contractions of the intestine through an enteric cholinergic reflex26. Butyrate, one product of SCFAs, plays a strong regulatory role in microbial TLR-dependent sensing, which is implicated in gut motility by secreting PYY and GLP-136. In our results, the butyrate levels were significantly lower in mice from STC donors than in mice from healthy donors. After supplementation with butyrate, the results of mice from STC donors were reversed in pellet frequency, water percentage, and colonic contractility. Therefore, fecal microbiota from STC donors might regulate gut motility by affecting the production of SCFAs.

Another particularly versatile class of microbial-produced metabolites, bile acids, is produced from cholesterol in hepatocytes and is metabolized by gut microbiota in the intestine30, 37. Evidence has demonstrated that BAs also impact gut motility by the release of serotonin from ECs38,39,40. When mice are transplanted with the microbiota from humans representing diverse culinary traditions, microbially deconjugated bile acid metabolites were correlated with faster gut transit41. Collectively, it is probable that BAs activate ECs to produce serotonin in the colon, and then alter gut motility. In our results, we found that the secondary BA level was lower in mice from STC donors, indicating that the microbial metabolism of BAs in the colon was disturbed. Decreased levels of secondary BAs could reduce the release of 5-HT from ECs, thereby affecting gut motility. After supplementation with DCA, the results of mice from STC donors were reversed in the water percentage and colonic contractility. However, the microbial metabolism of BAs is different between mice and humans. The primary BAs in humans are CA and CDCA, which in rodents are CA and muricholic acid (MCA). Next, deconjugated primary BAs that enter the colon are metabolized through 7-dehydroxylation into secondary BAs, including DCA from CA and LCA from CDCA. In rodents, the primary murine MCA also results in the formation of murideoxycholic acid (MDCA) in addition to DCA and LCA. In humans, ursodeoxycholic acid (UDCA) is a secondary BA that is formed by 7α/β-isomerization of CDCA, and this process can be performed by Clostridium absonum 30, 42. Therefore, altered bile acid profiles and a different metabolism process may affect various signaling through bile acid receptors between mice and humans. Thus, the results of our study warrant further discussion when translating findings from mice to humans.

FMT has shown efficacy for various gastrointestinal disorders, such as Clostridium difficile infection (CDI), inflammatory bowel disease (IBD), and irritable bowel syndrome (IBS). We previously suggested that FMT could increase bowel movements, improve the Wexner constipation score and GIQLI score in STC patients18. Studies have reported that in patients with recurrent CDI, the metabolism of bile salts and primary BAs to secondary BAs was disrupted, and FMT could result in the normalization of fecal bacterial community structure and could correct this abnormal metabolic composition43. FMT was also found to have a positive clinical response in ulcerative colitis, which was associated with successive colonization of donor-derived phylotypes44. Therefore, the mechanism of FMT might be the reestablishment of intestinal flora and abnormal microbiota metabolites in patients. Taken together, intestinal microbiota from STC donors was proven to affect gut motility, and the regulation of the intestinal microenvironment by FMT might be effective for constipated patients.

There are several limitations in this study. First, antibiotic treatment cannot completely remove the intestinal microbiota. Thus, it cannot be ruled out that some of the residual microbiota influence gut motility. Second, only six patients with slow transit constipation were enrolled, so we could not describe changes in the intestinal microbiota systematically. However, we plan to conduct a metagenomics analysis of intestinal microbiome in constipation in the future, with many more selected patients. Third, the observed changes in our study do not seem dramatic, possibly because gut motility is regulated by multiple signaling pathways. The intestinal microbiota might be just one of the causes. Thus, further work is necessary to clarify whether other pathways participate in the regulation of gut motility in constipation.

In summary, the results of this study have suggested that the colonization of the gut by microbiota from constipated donors might affect gut motility and stool consistency in mice through modulating host metabolism, and reestablishment of the intestinal microenvironment in constipated patients might be effective. The alterations of the microbiome in constipation, in turn, could affect the host, which might increase significant evidence for a novel treatment on fecal microbiota transplantation for constipated patients.