丁酸梭菌早期干预对大肠杆菌攻毒肉鸡肠道炎症因子和抗菌肽基因表达的影响

Effect of early intervention by Clostridium butyricum on intestinal inflammatory factors and gene expression of antimicrobial peptides of boiler chicken challenged by Escherichia coli

为研究丁酸梭菌(CB)早期干预对大肠杆菌攻毒肉鸡肠道炎症因子和抗菌肽基因表达的影响,本试验选用健康的1日龄肉鸡200只,在1~3 d时,随机选择100只每天灌喂1 mL浓度为109 cfu/mL的丁酸梭菌,另外100只灌喂1 mL生理盐水。在7日龄阶段,生理盐水灌喂肉鸡分成:对照组和大肠杆菌组(EC);丁酸梭菌处理肉鸡分成:丁酸梭菌组(CB组)和丁酸梭菌+大肠杆菌组(CB+EC组),其中EC组和CB+EC组均分别灌喂1 mL 108 cfu/mL产肠毒素大肠杆菌菌液进行攻毒,而对照组和CB组则灌喂1 mL生理盐水。ETEC攻毒后记录鸡群的下痢情况,分别于攻毒后第3天(10日龄)和7天(14日龄)屠宰取回肠组织,实时荧光定量检测炎症因子和抗菌肽基因的表达。结果显示:(1)大肠杆菌攻毒后,丁酸梭菌灌喂的雏鸡均未出现死亡的情况,对照组和大肠杆菌组中雏鸡死亡率为3%。鸡群在灌喂大肠杆菌后出现了明显的下痢,大肠杆菌组下痢率为73.52%,而丁酸梭菌早期干预使得大肠杆菌攻毒引起的下痢率降低为29.41%。(2)在回肠组织中,攻毒3 d后,CB组与EC组和EC+CB组相比,IL-8和TLR4基因的表达量显著降低(P<0.05),而IL-10基因的表达量显著升高(P<0.05)。攻毒7 d后,CB组IL-10的表达量显著高于EC组(P<0.05),而对照组和CB组TLR4的表达量显著低于EC组与CB+EC组(P<0.05)。(3)对于肠道抗菌肽基因的表达水平,攻毒3 d后,CB组回肠组织AvBD10表达量极显著高于其他处理组(P<0.01)。攻毒7 d后,CB组与EC组AvBD9的表达量显著高于其他组(P<0.01),CathB1表达量各处理组均显著增加(P<0.01)。综上所述,丁酸梭菌早期干预可以在一定程度上降低大肠杆菌攻毒肉鸡肠道中的炎症反应和提高抗菌肽基因的表达,从而减少大肠杆菌引起的下痢。

The aim of this study was to investigate the effect of early intervention of Clostridium butyrate on intestinal inflammatory factors and gene expression of antibacterial peptide in broilers infected with Escherichia coli.In this study,200 healthy 1-day-old broilers were randomly selected to be fed with 1 mL of Clostridium butyrate with a concentration of 109 cfu/mL every day for 1~3 days,and the other 100 were fed with 1 mL saline.At the age of 7 days,the broilers were fed with saline and divided into control group and E.coli group(EC group).The broilers were fed with Clostridium butyric and divided into:Clostridium butyric group(CB group)and Clostridium butyric+E.coli group(CB+EC group),the EC group and CB+EC group were irrigation feed 1mL 108 cfu/mL enterotoxin Escherichia coli bacteria liquid poison,while the control group and CB group filled 1 mL saline.After ETEC challenge,the situation of dysentery was recorded in the broilers.Ileum tissues were slaughtered on the 3rd day(10 days old)and the 7th day(14 days old)after challenge,and the expression of inflammatory factors and antimicrobial peptide genes were detected by real-time fluorescence quantification.The results showed that:(1)After ETEC challenged,the broilers fed with Clostridium butyrate didn't die,the mortality rate in the control group and the EC group was 3%.The chickens showed obvious diarrhea after feeding E.coli,and the diarrhea rate of E.coli group was 73.52%,while the early intervention of Clostridium butyrate reduced the diarrhea rate caused by E.coli attack to 29.41%.(2)In ileal tissue,the expression of IL-8 and TLR4 genes in CB group was significantly lower than that in EC group and EC+CB group after ETEC challenged 3 days,and the expression of IL-10 in CB group was significantly higher(P<0.05).After ETEC challenged 7 days,the expression of IL-10 gene in CB group was significantly higher than that in EC group(P<0.05).The expression of TLR4 in control group and CB group was significantly lower than that in EC group and CB+EC group(P<0.05).(3)As for the expression level of intestinal antimicrobial peptide gene,3 days after challenge,AvBD10 in ileum tissues of CB group was significantly higher than that in other groups(P<0.01).7 days after challenge,AvBD9 in CB group and EC group was significantly higher than that in other groups(P<0.01).CathB1 expression in all treatment groups was significantly increased(P<0.01).In conclusion,the early intervention of Clostridium butyrate in vivo could decrease the inflammatory reaction and increase the expression of antibacterial peptide gene in the intestine of broilers infected by Escherichia coli to a certain extent,thus reducing the diarrhea caused by Escherichia coli.