Antibiotic cocktail followed by Clostridium butyricum (CBM588) supplementation improves colonic anastomotic healing in mice

• Preoperative Clostridium butyricum (CBM588) supplementation alone did not influence fecal bacterial diversity and without observable improvement of postoperative performance or anastomotic healing in a murine colonic anastomosis model. • Preoperative antibiotic cocktail therapy decreased fecal bacterial α-diversity and altered β-diversity, associated with improved postoperative performance, reduced areas of anastomotic necrosis, and increased anastomotic healing scores in a murine colonic anastomosis model. • Preoperative antibiotic cocktail therapy followed by Clostridium butyricum (CBM588) supplementation decreased fecal bacterial α-diversity, altered and modulated β-diversity, resulting in a remarkably increased percentage of Clostridiaceae . These findings were associated with improved postoperative performance, reduced areas of anastomotic necrosis, and increased anastomotic healing scores in a murine colonic anastomosis model. Microbiota manipulation through preoperative probiotic supplementation to reduce surgical site infection has shown promising results in recent years. However, the changes in gut microbiota resulting from this manipulation remain unclear. In eastern Asia, Clostridium butyricum (CBM588) has been demonstrated to have antimicrobial ability. This study aimed to explore postoperative outcomes associated with microbiota changes in a murine colonic anastomosis model. C57BL/6 mice were divided into four groups. All mice, including group A as the control, received standard oral neomycin and metronidazole before surgery. Group B received CBM588 before surgery. Group C received the antibiotic cocktail therapy before surgery. Group D received the antibiotic cocktail therapy followed by CBM588 supplementation before surgery. A 16S rRNA amplicon with SMS was used to show the impact of different treatments on the murine gut microbiota. The implementation of the antibiotic cocktail therapy resulted in improved postoperative performance, significantly better anastomotic healing scores, and reduced areas of anastomotic necrosis in both group C and D, but particularly prominently in group D. Metagenomic analysis revealed that group D was enriched with the order Clostridiales , the family Clostridiaceae , and the genus Clostridium , with low bacterial diversity and distinct bacterial composition. A preoperative antibiotic cocktail therapy followed by CBM588 supplementation modulates bacterial diversity and composition, resulting in an increased percentage of Clostridiaceae in a murine colonic anastomosis model. The associated outcomes include lower intra-abdominal adhesion scores, improved postoperative performance, improved anastomotic healing scores, and reduced areas of anastomotic necrosis.