| The gut microbiota,including probiotics and pathogenic microorganisms,is involved in diarrhea by regulating pathogenic microorganisms and the production of intestinal mucosal antibodies.Mongolian medicine Baatar seven powder(BSP),a compound of Mongolian medicine listed in the Ministry of Health's Pharmaceutical Standards(Mongolian medicine sub-register),has been recognized as an effective drug for diarrhea,owing to its detoxification,analgesia,dysentery and immunity.Therefore,this study deeply explored the influence of intestinal microorganisms formed by early intervention of BSP on the mechanism calf diarrhea of intestinal mucosal barrier function,which laid the foundation for the clinical promotion of BSP.This study included four experiments:Experiment 1:To determine how BSP the structure and function of the gastrointestinal flora was regulated the structure and function of the gastrointestinal flora of calves infected with pathogenic E.coli O1,60 healthy Holstein calves were selected as control group(NC),infection group(EC),positive control group(CIP),and Mongolian medicine Baatar-7 at a high(5.0 g/kg),medium(2.50 g/kg)and low(1.25 g/kg)dosage.The results showed that(1)after 7 days of administration,in the cecum,colon and rumen contents,compared with the NC group,the levels of E.coli and Proteobacteria of calves in the EC group significantly increased.Compared with the EC group,BSP.M and BSP.H groups significantly increased Faecalibacterium and Alloprevotella in the cecum and colon.?Compared with the NC group,the levels of E.coli,Proteobacteria,and Clostridium perfringens in rectal feces of calves in the EC group significantly increased at days 3,5,7,and 15,30,60 and 90 days after ceasing BSP treatment.Experiment 2:To study how the intestinal microorganisms formed by early intervention of BSP was regulated intestinal mucosal barrier-related indicators of calves infected with pathogenic E.coli O1,followed by the detection of diarrhea calves small intestine morphology by HE,the observation of tight junction proteins and lymphocyte cytokines by immunofluorescence technique,the detection of anti-inflammatory and immune cytokines,ELISA.The results indicated that(1)the histological sections of E.coli induced that the small intestine villi shed and shrunk.The Claudin-1?Occludin and ZO-1 protein expression levels were upregulated,and the levels of CD8+and CD11c+ protein expression were downregulated in calves infected with pathogenic E.coli O1.(2)BSP.M group increased the highest levels of IL-2,IL-4,IFN-y,SIgA,IgA and IgG in serum,and significantly reduced the levels of IL-6,TNF-?,5-HT and Ach.(3)The relative abundance of increased bacterial after BSP administration,Lactobacillus,Faecalibacterium and Alloprevotella,were negatively correlated with the levels of CD8+ and IL-6,as well as positively correlated with CD4+.Experiment 3:Using cow-to-mouse fecal transplantations as an intervention to effectively colonized the intestinal microorganisms of mice,and reduced the emission of pathogenic bacteria,16S rRNA and RT-PCR were used to detect the levels of Proteobacteria and E.coli in the colon contents.The results showed that BSP.M-FMT significantly increased the relative abundance of Akkermansia and Bacteroidetes in the colon of mice,and significantly reduced the content of E.coli.In addition,it significantly increased the expression of AQP-3 protein in the intestine and reduced the diarrhea rate.Experiment 4:Based on Mongolian medicine is multi-level,multi-target drugs:(1)The highest protection rate(88,89%)in mice was obtained from effetive compounds of Baatar-7(A2B2C3D2E2F1G1),as same rate as ciprofloxacin treated mice.(2)Transcriptome and RT-PCR were used to detect the effect of BSEC on the antibacterial pathway of E.coli in vitro.The results showed that BSEC inhibited E.coli O1 by significantly up-regulating the genes in the sulfur metabolic pathway and microbial metabolism in diverse environments.(3)To study the effect of BSEC on the intestinal structure and mechanical barrier of diarrhea model mice,16S rRNA method and high performance liquid chromatography were used to detect the colonic microorganisms and some the relevant biochemical indicators in tryptophan metabolism pathway,the anti-inflammatory and immune factors by ELISA.?The results showed that the relative abundance of Proteobacteria and E.coli were significantly downregulated in mice infected with pathogenic E.coli O1 compared with EC group,the relatve abundance of Alloprevotella and Parabacteroides were upregulated.?The histological sections of mice with diarrhea induced by E.coli O1 indicated that the small intestine had intestinal villous shedding and edema.the content of Occludin,Claudin-1 and ZO-1 in the small intestine mucosa were significantly up-regulated compared with EC group.(4)Compared with the EC group,the BSEC.L,BSEC.M and BSEC.H groups reduced the levels of TNF-?,IL-1? and IL-6,increased the levels of IL-2,IL-4 and IFN-?,and enhanced the contents of SIgA,CD4+,CD3+,CD19+and CD4+/CD8+T ratio.(5)To study the effect of BSEC on the physiological and biochemical indexes of diarrhea model mice,the blood biochemical analyzer was used to detect the physiological and biochemical indexes.(6)The results showed that the levels of CYP1A1,IDO1,SLC6A4 protein,AhR and Kyn were up-regulated in BSP.M group compared with EC group,but the levels of TPH1 protein,5-HT and 5-HTP were down-regulated.In addition,the relative abundance of increased bacterial after BSP administration,Alloprevotella and Parabacteroides,were positively correlated with the levels of AhR and Kyn.Conclusion:The gastrointestinal flora environment formed by the early intervention of the Mongolian medicine Baatar-7 and its effetive compounds has an inhibitory effect on the growth of pathogenic E.coli,thereby improved the intestinal mucosa structure of animals and enhanced the digestive absorption function in the intestine,which maintain the body's physiological and biochemical functions,enhanced the tryptophan metabolism and the body's anti-inflammatory immune function. |
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