Study On The Mechanism Of The Gut Microbiota Affects Growth Performance In Chickens By Regulating Fat Metabolism

Promoting the growth performance to improve the chicken production efficiency is crucial in poultry production.The gut microbiota coexists harmoniously with the host,which influences host' nutrient metabolism,immunity and growth performance.Fat metabolism plays an important role in maintaining host health and promoting host growth performance.The gut microbiota can regulate fat metabolism to improve host growth performance by affecting the expression of genes related to fat metabolism.However,few studies have been done on the relationship between gut microbiota in the content of different intestinal part and fat metabolism.Therefore,in this study,high-throughput sequencing technology was used to analyze the 16 S r RNA V4 gene sequence in different intestinal contents between high and low body weight chickens.Immunohistochemical stainings,fluorescent quantitative PCR and Western Blot techniques were used to verify the expression of genes related to fat metabolism,and the correlation analysis was carried out,to explore the machanism of the gut microbiota affecting the chicken growth performance by regulating fat metabolism.The main research contents and results are as follows:1.The test result of indicators related to growth performanceThere was a significant difference in body weight between high and low body weight chickens(P<0.01).Chest muscle index was significantly higher in high body weight chickens than that in low body weight chickens(P<0.05),leg muscle index was significantly higher than that in low body weight chickens(P<0.01),the cross-sectional area of chest muscle cell and leg muscle cell were significantly larger than that in low body weight chickens(P<0.05).The results suggested that there was a significant difference in growth performance between high and low body weight chickens.2.The test result of indicators related to fat metabolism2.1 The test result of blood biochemical indicatorsThe concentrations of serum triglyceride(TG),total cholesterol(TC),high-density lipoprotein cholesterol(HDL-C)and low-density lipoprotein cholesterol(LDL-C)were detected respectively in high and low body weight chickens.The concentration of HDL-C was significantly higher in high body weight chickens than that in low body weight chickens(P<0.05),the concentration of LDL-C was significantly lower than that in low body weight chickens(P<0.05),and there were no significant differences in the concentration of TG and TC in serum between high and low body weight chickens.The results suggested that there was a significant difference in fat metabolism between high and low body weight chickens.2.2 The morphological structure of abdominal fatThe abdominal fat tissue was utilized to make frozen slices.The result of HE staining showed that adipocytes were vacuoles,the number of adipocyte was significantly higher in high body weight chickens than that in low body weight chickens(P<0.05),the average diameter of adipocyte was significantly lower than that in low body weight chickens(P<0.01).The results suggested that there was more fat in the abdomen of low body weight chickens.2.3 The fat distribution in liverLiver index was significantly lower in high body weight chickens than that in low body weight chickens(P<0.05).The result of HE staining showed that there were more vacuolar fat in the liver of high body weight chickens.The results suggested that more fat was deposited in the liver of high body weight chickens.2.4 The expression of genes related to fat metabolismQ-PCR was used to detect the changes in the expression of genes involved in fat synthesis,fat transport,fat catabolism and adipocyte differentiation in liver,abdominal fat,chest muscle and leg muscle.The distribution and protein expression levels of phosphor-adenosine monophosphate activated protein kinase(P-AMPK)in liver,chest muscle and leg muscle were detected by immunohistochemical staining.Western Blot was used to detect the protein expression levels of adenosine monophosphate activated protein kinase(AMPK)and P-AMPK in liver,chest muscle and leg muscle.The results showed that the expression of fatty acid desaturase 1(FADS1),cytochrome P 450 2C45(CYP2C45),long-chain fatty acyl-Co A synthetase 1(ACSL1)related to fat synthesis,apolipoprotein AI(Apo AI)related to fat transport,peroxisome proliferators-activated receptor ?(PPAR?),carnitine palmitoyltransferase I(CPT-1)and fasting-induced adipose factor(fiaf)related to fat catabolism were significantly higher in the liver of high body weight chickens than that of low body weight chickens(P<0.05).The expression of sterol regulatory element-binding protein 1(SREBP1)and adiponectin(adiponetin)related to adipocyte differentiation were significantly lower in the abdominal fat of high body weight chickens than that of low body weight chickens(P<0.05).The expression of adipocyte fatty acid binding protein(A-FABP)related to fat transport was significantly lower in the chest muscle of high body weight chickens than that of low body weight chickens,the expression of CPT-1 was significantly higher in the chest and leg muscle of high body weight chickens than that of low body weight chickens(P<0.05).P-AMPK was mainly distributed in the cytoplasm and nucleus of hepatocytes in liver,connective tissue of the chest muscle and leg muscle,and the protein expression of P-AMPK was significantly higher in the liver,chest muscle and leg muscle of high body weight chickens than that of low body weight chickens(P<0.05).The results suggested that there were significant differences in the expression of genes related to fat metabolism between high and low body weight chickens.3.The test result of intestinal microbial compositional spectrumThe microbial diversity and composition of the duodenum,the jejunum,the ileum and the cecum in high and low body weight chickens were analyzed by 16 S r RNA high-throughput sequencing.The results showed that cecum had a richer and more diverse microbial community in the four intestinal segments between high and low body weight chickens.Firmicutes and Proteobacteria were the dominant bacteria in the small intestine.Lactobacillus was the most abundant genera in the small intestine.Firmicutes,Bacteroidetes and Proteobacteria were the most abundant phyla in cecum.Bacteroides was the dominant genera in cecum.The microbial community structure of duodenal,ileal and cecal contents was significantly different between high and low body weight chickens.In duodenum,the ratio of Firmicutes to Bacteroidetes was significantly lower in high body weight chickens than that in low body weight chickens(P<0.01).In cecum,the ratio of Frmicutes to Bacteroidetes was significantly lower in high body weight chickens than that in low body weight chickens(P<0.05).The linear discriminant analysis effect size analysis showed that the relative abundance of Microbacterium and Sphingomonas were significantly higher in the cecal contents of high body weight chickens than that of low body weight chickens,and the relative abundance of Slackia was significantly lower than that in low body weight chickens.Phylogenetic investigation of communities by reconstruction of unobserved states analysis showed that Adipocytokine signaling pathway and PPAR signaling pathway were significantly different in cecal contents between high and low body weight chickens.The results indicated that the microbial structure and composition of the cecal contents between high and low body weight chickens were significantly different,which was closely related to fat metabolism.4.Correlation analysis between the gut microbiota and growth performanceSpearman correlation analysis was utilized to explore the potential relationship between the microbiota of cecal content and growth performance.The results showed that Sphingomonas was significantly positively correlated with body weight,chest muscle weight,leg muscle weight,the cross-sectional area of chest muscle cell and leg muscle cell.Microbacterium was significantly positively correlated with body weight,chest muscle weight,the cross-sectional area of chest muscle cell and leg muscle cell.Slackia was significantly negatively correlated with body weight,chest muscle weight,leg muscle weight,the cross-sectional area of chest muscle cell and leg muscle cell.The results suggested that Sphingomonas and Microbacterium may be beneficial bacteria of the gut microbiota,which could promote the growth and development of animals.Slackia may be harmful bacteria of the gut microbiota,which could prevent the growth and development of animals.5.Correlation analysis between the gut microbiota and fat metabolismSpearman correlation analysis was used to analysis the relationship between the microbiota of cecal content and indicators related to fat metabolism in order to explore whether the gut microbiota affected the growth performance of chickens by regulating fat metabolism.The results showed that Sphingomonas was significantly positively correlated with the concentration of serum HDL-C,the expression of CPT-1 in liver,the expression of CPT-1 and P-AMPK in chest muscle.Microbacterium was significantly positively correlated with the expression of CPT-1 and P-AMPK in liver,chest muscle and leg muscle.Slackia was significantly negatively correlated with the concentration of serum TC and HDL-C,the expression of P-AMPK in liver,the expression of CPT-1 and P-AMPK in chest muscle,the expression of P-AMPK in leg muscle.The results showed that Sphingomonas and Microbacterium were significantly positively correlated with the oxidative decomposition of fatty acids,and the energy production to promote the growth and development of animals.Slackia was significantly positively correlated with the deposition of fat in animals,which was not conducive to the growth and development of the host.The above results showed that Sphingomonas,Microbacterium and Slackia in the cecal content can regulate the fat metabolism in liver,abdominal fat,chest muscle and leg muscle by regulating the expression of genes related to fat metabolism,thus affecting the growth performance of chickens.