Effects Of Butyrate-producing Engineered Bacteria On Metabolism And Intestinal Flora In Mice Fed A High-fat Diet

Obesity is a major global health problem facing the world.Traditional medicine and surgical treatment have side effects,and new treatments are still exploring.The emergence of live biotherapeutisc provides new methods for the treatment of obesity.In this study,the butyrate-producing engineered bacteria were constructed to explore its effect on the metabolism and intestinal flora of obese mice induced by a high-fat diet.We hope it provides an experimental and scientific basis for the development of live biotherapeutisc for improving glucose and lipid metabolism.We knocked out autolysis regulated genes skf A and sdp C,the acetic acid synthesis regulatory genes acd A and ack A in Bacillus subtilis SCK6(BsS),respectively.And then we introduced the butyryl-Co A: acetate-Co A transferase coding gene in BsS to construct butyrate-producing bacteria.The results showed that compared with BsS,the butyrate-producing bacteria have significantly improved biomass,while the butyric acid production can reach 1.53±0.042 g/L.The constructed butyrate-producing bacteria were screened in the in vitro simulated colonic fermentation model,and the butyric acid-producing engineering bacteria BsS-5 was obtained.Butyric acid production could reach 0.507±0.001 g/L.Afterwards,we co-cultured BsS-5 with the obesity-derived bacterial community.We found that the concentration of butyric acid in the co-culture system significantly increased 1.8 times higher than that in obesity.Simultaneously,the 16 S amplicon sequencing results showed that BsS-5 could promote the diversity of the fecal flora of mice fed a high-fat diet,increased the abundance to the Bacteroides,and decrease the abundance Proteobacteria,Clostridium and Enterococcus that enriched in the intestine of obesity.We established an obsess mouse model by a high-fat diet.The obese mice were intervened by the intragastric administration of BsS-5 to evaluate its effect on metabolism and intestinal flora.They fed with inulin(HP),BsS(HS),BsS-5(HE)and BsS-5 prevention group(HEP)was added.The results showed compared with HFD,the weight of HE mice reduced by 18.8%,and the weight of HEP mice reduce by 28.4%after four weeks intervention of BsS-5.Besides,BsS-5 significantly reduced the weight of visceral fat and the levels of triglycerides,total cholesterol and low-density lipoprotein,and improved glucose tolerance and insulin resistance.The results of metabolomics showed that 22 metabolites significantly changed between HFD and HE and HEP groups,including urobilin,methionine and homogentisic acid,these metabolites play an important role in inhibiting obesity and regulating metabolism.The involved metabolic pathways include phenylalanine metabolism and synthesis,histidine metabolism and linoleic acid metabolism.Simultaneously,we found that BsS-5 can change the composition of the intestinal flora of HFD mice,reduced the abundance of Chlamydia and Firmicutes,increased the abundance of Bacteroides.Some vital intestinal microorganisms were identified,including Odoribacteraceae,Faecalibaculun and Allobaculum genera,Helicobacter bilis,Clotridium sp CAG 58,Biofidobacterium pseudolongum,Streptococcus caviae and Firmicutes bacterium?M10?2.Furthermore,BsS-5 can enhance the metabolic function of the intestinal flora.In summary,the constructed butyrate-producing engineered bacteria BsS-5 can inhibit weight gain caused by a high-fat diet,improve metabolism,and regulate intestinal flora.It has a prospect for the development of live biotherapeutisc.