动物营养学报    2022, Vol. 34 Issue (6): 3519-3528    PDF    
丁酸梭菌对肠道屏障的影响机制及其在畜禽养殖中应用的研究进展
王宗伟1,2 , 付建中3 , 李洪涛4 , 李艳青1     
1. 潍坊科技学院, 生物研发中心, 潍坊 262700;
2. 寿光市行政审批服务局, 寿光 262705;
3. 寿光市 稻田镇畜牧兽医工作站, 寿光 262706;
4. 临沂布恩生物科技有限公司, 临沂 276036
摘要: 丁酸梭菌作为饲料添加剂, 具有抑菌、促生长等多种生物学作用。本文综述了丁酸梭菌对肠道屏障的影响机制及其在生物安全性、畜禽抗病和促进肠道健康上的研究进展, 以期为丁酸梭菌在畜禽营养上发挥替抗作用提供参考。
关键词: 丁酸梭菌            肠道屏障    生物安全性    抗病    
Research Progress on Mechanism of Clostridium butyricum Affecting Intestinal Barrier and Its Application in Livestock and Poultry Breeding
WANG Zongwei1,2 , FU Jianzhong3 , LI Hongtao4 , LI Yanqing1     
1. Biological Research Centre, Weifang University of Science and Technology, Weifang 262700, China;
2. Shouguang Administrative Examination and Approval Service Bureau, Shouguang 262705, China;
3. Shouguang Daotian Town Animal Husbandry and Veterinary Workstation, Shouguang 262706, China;
4. Linyi Boone Biotechnology Co., Ltd., Linyi 276036, China
Abstract: As a feed additive, Clostridium butyricum has many biological functions such as bacteriostasis and growth promotion. This paper reviewed the mechanism of Clostridium butyricum on intestinal barrier and its research progress in biosafety, disease resistance and intestinal health promotion in livestock and poultry, in order to provide reference for the role of Clostridium butyricum to substitute antibiotics in nutrition of livestock and poultry.
Key words: Clostridium butyricum    pigs    poultry    intestinal barrier    biosafety    disease-resistant    

我国已经禁止在全价配合饲料中使用抗生素类生长促进剂[1],这将对畜禽的生长性能和健康产生负面影响。丁酸梭菌(Clostridium butyricum)具有修复肠道黏膜、增强免疫和抗氧化性能以及预防疾病等多种抗病和促进肠道健康的作用,是理想的饲料用抗生素替代品。本文综述了丁酸梭菌对肠道屏障的影响机制及其在生物安全性、畜禽抗病和促进肠道健康等畜禽养殖中应用的研究进展,以期为丁酸梭菌发挥替抗作用提供参考。

1 丁酸梭菌的发酵特性

丁酸梭菌又名酪酸梭菌,为革兰氏阳性菌,厌氧,周身鞭毛,能运动。丁酸梭菌(芽孢)能够耐受饲料制粒过程中的高温高压,对酸性胃液也有较好的耐受性,在过胃后pH大于5的肠道环境中才开始生长[2-3]。丁酸梭菌发酵产生短链脂肪酸(丁酸、乳酸和乙酸等)、消化酶(非淀粉多糖酶、脂肪酶和蛋白酶等)、抗菌肽、B族维生素和氢气(H2)等[4-5];个别菌种兼性好氧,且发酵产生E型肉毒毒素[6]。丁酸梭菌发酵时存在酸胁迫效应,工业化生产条件要求高,产量相对枯草芽孢杆菌等其他菌种要低。

2 丁酸梭菌对肠道屏障的影响机制

肠道屏障是指肠道防止肠腔内有害物质透过肠黏膜吸收进入畜禽体内的结构和功能的总和[7],根据功能的不同分为机械屏障、化学屏障、免疫屏障和生物屏障。丁酸梭菌对肠道屏障的影响机制主要是保护畜禽(尤其是幼年和肠道功能不全畜禽)胃肠道免受外源性(细菌、病毒等)刺激,提高抵抗力,促进其生长。

2.1 对肠道机械屏障的影响

肠道机械屏障是畜禽肠道最重要的物理屏障,主要包括完整的肠黏膜上皮细胞、细胞间紧密连接以及肠道的运动功能等。丁酸梭菌对肠道机械屏障的影响表现在以下3个方面:1)丁酸梭菌发酵产生的丁酸等短链脂肪酸直接为肠上皮细胞提供能量,促进肠绒毛生长,增强肠道消化吸收功能[8];2)发酵所产丁酸等通过提高肠上皮细胞的能量状态,激活蛋白激酶B(protein kinase B,AKT)/哺乳动物雷帕霉素靶蛋白(mammalian target protein of rapamycin,mTOR)信号通路介导的蛋白合成机制,上调紧密连接蛋白——密封蛋白(claudin)、闭合蛋白(occludin)和闭锁小带蛋白(zonula occlaudin,ZO)的表达,降低肠道通透性,维持肠道上皮屏障的完整性[9-11];3)丁酸梭菌通过上调肠道肌层结缔组织中卡哈尔间质细胞膜表面Toll样受体2(Toll-like receptor 2,TLR2)的表达,促进肠道蠕动[12],这有助于避免肠道内细菌的长时间黏附和促进菌群的更新。

2.2 对肠道化学屏障的影响

肠道的化学屏障主要包括覆盖在肠上皮表面起保护和润滑作用以黏蛋白(mucin,MUC)为主的黏液、消化液、胆汁以及肠道微生物产生的抑菌物质等。丁酸梭菌通过黏附人结肠癌细胞(HT-29细胞)表面MUC的黏液多糖位点,增加MUC的产生和糖基化[13];丁酸梭菌还显著上调断奶仔猪小肠MUC1、MUC4和MUC20的表达[14]。丁酸梭菌产生的丁酸等短链脂肪酸和CBP22等抗菌肽,能够抑制有害菌的生长[5, 15-17]。丁酸梭菌能显著提高肉鸡十二指肠淀粉酶和胰蛋白酶活性[4]以及肉鹅空肠脂肪酶活性[18],改变蛋鸡回肠胆汁酸组成[19],提高营养物质消化率,但这些消化酶活性的提高和胆汁酸组成的改变是否具有增强肠道化学屏障的功能有待进一步研究。

2.3 对肠道免疫屏障的影响

肠道免疫屏障主要由树突状细胞等淋巴细胞、分泌型免疫球蛋白A(sIgA)等抗菌物质,以及派伊尔结肠相关淋巴组织等组成。丁酸梭菌及其调节后产生的肠道有益菌通过脂磷壁酸、DNA等菌体成分激活肠道免疫和抗氧化功能,通过其代谢产物(如丁酸、抗菌肽和其他短链脂肪酸等)发挥肠道抗菌和消炎等作用。

2.3.1 激活免疫系统

丁酸梭菌及代谢产物通过激活肠道细胞Toll样受体(Toll-like receptor,TLR)、G蛋白偶联受体(G protein-coupled receptor,GPR)等,在不诱导肠道细胞凋亡和坏死[20]的前提下,激活多个信号通路,增强肠道黏膜的天然免疫系统和获得性免疫系统。首先,丁酸梭菌能利用脂磷壁酸等菌体成份激活肠道细胞TLR2(配体为脂磷壁酸等)-髓样分化因子88(myeloid differentiation factor 88,MyD88)或独立于MyD88的途径——核转录因子-κB(nuclear factor-kappa B,NF-κB)信号通路,低水平上调促炎因子——白细胞介素-8(IL-8)、白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)和肿瘤坏死因子-α(TNF-α)的表达,激活肠道天然免疫系统,帮助机体清除病原体[9, 21]。其次,丁酸等能激活短链脂肪酸受体(GPR43、GPR109a),增加小鼠炎症早期结肠黏膜固有层中性粒细胞、辅助性T淋巴细胞1和辅助性T淋巴细胞17的数量[22]。再次,丁酸梭菌能显著上调仔猪空肠上皮细胞内促进炎症发展的炎症小体——核苷酸结合寡聚化结构域(nucleotide-binding oligomerization domain,NOD)样受体家族含pyrin结构域蛋白(NOD-like receptor family, pyrin domain-containing protein,NLRP)3和抑制炎症发展的NLRP6的表达,极显著上调抑制炎症发展的NLRP12的表达[23]。最后,丁酸梭菌能显著提高肉鸡肠道黏膜固有层B淋巴细胞分泌的sIgA的含量,增强机体免疫功能[24]

2.3.2 降低炎症反应

丁酸梭菌及代谢产物能通过肠道上皮细胞、树突状细胞等细胞膜表面受体TLR、GPR,以及细胞内NLRP等,激活或抑制多个信号通路,调节细胞因子和免疫细胞,发挥抗炎、抗凋亡作用。TLR:1)丁酸梭菌通过脂磷壁酸等激活TLR2、TLR5信号通路,上调肠道细胞中β-防御素、抗菌肽以及抗炎因子——转化生长因子(TGF)-β2、TGF-β4等的表达[9, 25],缓解肠道炎症。其中,丁酸梭菌与金葡菌脂磷壁酸间的差异以及白细胞介素-10(IL-10)的表达,在丁酸梭菌介导的免疫保护中发挥重要作用[26-27]。丁酸梭菌通过激活TLR2-细胞内信号调节激酶(extracellular signal-regulated kinase,ERK)-激活蛋白-1(activated protein-1,AP-1)激酶通路诱导肠道固有层树突状细胞分泌TGF-β1,促进肠内调节性T细胞的生成,抑制炎症反应[28]。2)丁酸能显著抑制转录因子pu1与TLR4(配体为脂多糖等)基因启动子区域的结合亲和力[29]。丁酸梭菌通过抑制TLR4-MyD88-NF-κB信号通路,降低沙门氏菌感染无特定病原(SPF)鸡肠上皮细胞中促炎因子——干扰素-γ(IFN-γ)、IL-1β、IL-8和TNF-α的水平,缓解炎症反应[30]。GPR:1)丁酸等短链脂肪酸通过肠道CD4+ T细胞和先天性淋巴样细胞膜上GPR41,抑制组蛋白去乙酰化酶,提高白细胞介素-22(IL-22)(IL-10家族)的含量,发挥抗炎和免疫调节功能[31];2)丁酸通过上调小肠隐窝底部潘氏(Paneth)细胞GPR41、GPRx41和GPR109a mRNA的表达,分泌α-防御素,杀死鼠伤寒沙门氏菌[32]。NLRP炎症小体:丁酸梭菌能下调产肠毒素大肠杆菌仔猪肠道细胞中NLRP3和半胱天冬酶-1(Caspase-1)的表达,降低产肠毒素大肠杆菌K88感染断奶仔猪血清和肠道IL-1β和白细胞介素-18(IL-18)的水平[33]。此外,丁酸梭菌通过降低炎症小鼠肠道黏膜固有层树突状细胞数量,下调其膜表面T细胞免疫球蛋白和黏蛋白结构域3(T cell immunoglobulin and mucin domain 3,TIM3)的表达,减少IL-1β、IL-6的产生[34]。丁酸梭菌所产丁酸还通过上调小鼠胃黏膜细胞凋亡抑制因子——B淋巴细胞瘤-2(B cell lymphoma-2,Bcl-2)/细胞凋亡促进因子——Bcl-2相关X蛋白(Bcl-2-associated X protein,Bax)的表达比例,保护胃黏膜细胞[35]

2.3.3 提高抗氧化性能

丁酸梭菌通过黏附在猪肠上皮细胞上,激活Kelch样环氧氯丙烷相关蛋白-1(Kelch-like epichlorohydrin-associated protein-1,Keap1)-核因子E2相关因子2(nuclear factor erythroid-2 related factor 2,Nrf2)-抗氧化反应元件(antioxidant response element,ARE)信号通路,提高超氧化物歧化酶(superoxide dismutase,SOD)、谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)的表达,降低丙二醛(malondialdehyde,MDA)和脂质代谢物的水平,从而减少大肠杆菌对肠黏膜的损伤[36];丁酸梭菌还通过激活p38丝裂原活化蛋白激酶(p38 MAPK)/Nrf2信号通路,上调SOD等抗氧化酶基因的表达,缓解猪霍乱沙门氏菌造成的猪上皮细胞氧化损伤[37]。丁酸梭菌发酵产生的H2能迅速扩散到组织和细胞中,选择性地与细胞内的羟基自由基和亚硝酸阴离子等强氧化剂发生反应,并调节基因表达来降低氧化应激,发挥抗炎、抗凋亡作用[38]

2.4 对肠道生物屏障的影响

肠道的生物屏障主要包括益生菌在肠黏膜上皮定植形成的菌膜屏障,肠道内的菌群与机体形成的相互依赖、相互制约的微生态平衡系统。丁酸梭菌对肠道生物屏障的作用是指其定植于肠上皮细胞上形成的菌膜屏障、丁酸等短链脂肪酸对有害菌的抑制作用以及优化肠道菌群组成带来的有利影响。

丁酸梭菌通过黏附肠上皮细胞[13],竞争性阻挡有害菌与肠上皮细胞结合,形成有效的菌膜屏障,以阻止伤寒杆菌等革兰氏阴性菌黏附肠细胞进行繁殖和释放内毒素,以及阻止金黄色葡萄球菌等革兰氏阳性菌诱导的NF-κB和ERK信号通路引起的细胞凋亡[15, 39-40];丁酸梭菌还直接下调产肠毒素大肠杆菌黏附基因FaeG与肠毒素基因estAestB的表达,降低产肠毒素大肠杆菌对猪肠道上皮细胞的损伤作用[41]。丁酸梭菌优化肠道菌群组成,提高厚壁菌门数量,降低拟杆菌门数量,使双歧杆菌等产短链脂肪酸菌数量增加,致病菌数量减少[42-43],甚至具有改善小鼠后代肠道菌群平衡的作用[44]。丁酸梭菌所产丁酸等短链脂肪酸能够降低肠道内局部pH,抑制产气荚膜梭菌等有害菌的生长、代谢以及毒素的产生[5, 15-16];产生的CBP22等抗菌肽对大肠埃希氏菌等也有抑制作用[5, 17]。丁酸梭菌通过优化肠道菌群提高丁酸和油酸的含量激活结肠上皮细胞GPR120,催化15-脂氧合酶将油酸等转化为保护素D1(protectin D1,PD1),从而促进IL-10和TGF-β1的分泌,降低促炎因子的分泌[45-46]

3 丁酸梭菌的生物安全性

生物安全性是丁酸梭菌作为饲料添加剂首要考虑的因素,已筛选的丁酸梭菌在耐药性、耐药基因是否向致病菌水平转移以及丁酸梭菌产毒性能等生物安全性方面是安全的[47-49]。丁酸梭菌主要对氨基糖苷类抗生素耐药,对其他类型抗生素耐药性的研究结果不一[47-50],这可能与不同来源丁酸梭菌含有不同的耐药基因有关。丁酸梭菌CBM588对庆大霉素等3种氨基糖苷类抗生素耐药,对其他7类7种抗菌药均无耐药性,并认为其耐药性是由于该类药物无法通过其细胞膜主动转运至菌体发挥作用[47];但有研究认为,丁酸梭菌rpsL基因使氨基糖苷类抗生素失活[48],需采用基因敲除验证其耐药机制。丁酸梭菌耐药基因型与表型之间具有较高的一致性[41]。范伟祥等[49]对24株鸡源丁酸梭菌中8类24种耐药基因进行检测,发现mefA等16种耐药基因全部成阴性,而全部成阳性的sul2、florblaTEM3种耐药基因已经不同程度的存在于沙门氏菌、大肠杆菌等有害菌中。24株鸡源丁酸梭菌、CBM588均不含有typeAtypeBtypeEtypeF肉毒毒素基因和alphabetaepsilon梭菌毒素基因[47, 49];人源丁酸梭菌不含有艰难梭菌毒素、非溶血性肠毒素、肠毒素等毒素基因[51]。Isa等[47]认为,产E型肉毒毒素丁酸梭菌较少存在于食物和土壤中,并且typeE肉毒毒素基因体外转移率较低,因此CBM588通过基因转移获得产毒能力的几率很小;但2019年我国发现首例由丁酸梭菌引起婴儿E型肉毒毒素中毒[52],并且肉毒毒素基因(typeAtypeBtypeF)质粒能够从肉毒梭状芽孢杆菌水平转移到丁酸梭菌[53]。将来这些毒素基因是否会水平转移到作为饲料添加剂的丁酸梭菌菌体中需要进一步观察研究。同时建议在饲粮配制中尽量避免将丁酸梭菌与有可能含有肉毒梭状芽孢杆菌的动物源性饲料(例如肉骨粉)合用。

4 丁酸梭菌在畜禽养殖中的应用

丁酸梭菌通过提高畜禽胃肠道健康水平、增强免疫和抗氧化性能、调节抗病基因的表达、减少仔猪断奶等应激和预防疾病等,发挥抗病和促进肠道健康的作用。研究发现,丁酸梭菌有效添加量为2×107~2.7×109 CFU/kg饲粮。欧盟动物饲料添加剂和产品研究小组(EFSA panel on additives and products or substances used in animal feed,FEEDAP)认为,丁酸梭菌FERM BP-2789添加到饲粮中的最小有效添加量为2.5×108 CFU/kg[54]

4.1 猪

Han等[8]在28~56日龄“杜×长×大”断奶仔猪饲粮中分别添加2.5×108 CFU/kg丁酸梭菌与抗生素(金霉素0.075 g/kg、吉他霉素0.055 g/kg和维吉霉素0.01 g/kg),结果料重比无显著差异;2.5×109 CFU/kg丁酸梭菌能显著提高结肠短链脂肪酸含量、干物质等表观利用率。Chen等[55]在饲粮中添加4×108 CFU/kg丁酸梭菌,能显著提高21~56日龄断奶仔猪十二指肠、空肠和回肠绒毛高度和空肠绒毛高度/隐窝深度值,降低低消化率组(低消化率vs.高消化率=玉米vs.膨化玉米)腹泻率,促生长作用与抗生素组(恩拉霉素20 mg/kg、金霉素75 mg/kg)无显著差异。Lan等[56]在低营养水平饲粮(代谢能16.12 vs. 16.74 MJ/kg;蛋白质19% vs. 20%)中添加丁酸梭菌等益生菌复合物能更有效地提高28~70日龄“杜×长×大”断奶仔猪的平均日增重,降低粪便氨态氮和硫化氢浓度。本项目研究在商品教槽保育料中添加1×109 CFU/kg丁酸梭菌,能够显著降低仔猪(尤其5 kg以下弱仔猪)腹泻率;在养殖场将1×107 CFU/kg丁酸梭菌和抗菌肽(枯草芽孢杆菌发酵提纯)联合用于防治仔猪断奶后1周腹泻,效果较好。此外,饲粮中添加抗性麦芽糖糊精能够提高肠道丁酸水平,缓解肠道炎症[57],这提示是否可将抗性麦芽糖糊精与丁酸梭菌联合用于纤维含量较少的断奶仔猪饲粮。

Cao等[58]在妊娠90 d至断奶21 d“长×大”二元母猪饲粮中添加2×108 CFU/kg丁酸梭菌,能改善母猪抗氧化性能,缩短仔猪产程,提高哺乳仔猪生长性能。本项目研究在商品母猪妊娠和哺乳料中均添加3×108 CFU/kg丁酸梭菌,发现母猪便秘减少,这可能是因为丁酸梭菌能够优化肠道菌群,促进非淀粉多糖消化,增加肠道蠕动,从而加速粪便排出体外。

4.2 鸡

Zhang等[59]在饲粮中添加2×107 CFU/kg丁酸梭菌可提高1~28日龄大肠杆菌K88攻毒肉鸡的免疫应答,改善肠道屏障功能,提高平均日增重,促生长作用与硫酸黏菌素组(20 mg/kg)无显著差异。Abdel-Latif等[60]研究发现,在1~35日龄科布肉鸡饲粮中添加2.5×109 CFU/kg丁酸梭菌和0.25 g/kg酿酒酵母,比单独添加5×109 CFU/kg丁酸梭菌更能改善肉仔鸡的肠道健康和免疫功能,而且提高了生长性能。但Huang等[61]在21~23日龄罗斯308肉鸡饲粮(鱼粉含量占饲粮50%)中添加1×109 CFU/kg丁酸梭菌并没有改善产气荚膜梭菌引起的坏死性肠炎肉鸡的肠道菌群组成,对肠道病变也无显著影响。这可能是因为饲粮中纤维含量较低,不利于丁酸梭菌的生长繁殖。此外,甘露寡糖能够显著促进丁酸梭菌产生大量的丁酸[62],这提示是否可将甘露聚糖酶与丁酸梭菌联合用于肠道较短的禽类饲粮。

Zhan等[63]在48~58周龄京红1号品系蛋鸡饲粮中添加5×107和1×108 CFU/kg丁酸梭菌可优化盲肠菌群,增强免疫功能和抗氧化性能,改善蛋鸡产蛋性能和蛋品质。Wang等[19]在饲粮中添加丁酸梭菌2.7×109 CFU/kg可加速60~68周龄海兰褐蛋鸡肝脏脂肪酸氧化,降低脂肪在肝脏中的沉积。张茜等[64]研究发现,在饲粮中添加4.5×108 CFU/kg丁酸梭菌和0.5%竹醋液,比分别单独添加9×108 CFU/kg丁酸梭菌、1%竹醋液更能提高61~69周龄蛋鸡回肠总抗氧化能力和脂肪酶活性,降低回肠MDA含量,改善产蛋后期蛋鸡产蛋性能和蛋品质。

4.3 鸭

李娜等[65]给1日龄番鸭连续3 d灌服每次1 mL含2×109 CFU/mL丁酸梭菌菌液,可提高1~10日龄番鸭的体重、回肠绒毛高度和绒毛高度/隐窝深度值,上调回肠紧密连接蛋白——claudin-1、claudin-2和ZO-2的表达。赵文文等[66]在饲粮中添加5×107 CFU/kg丁酸梭菌和800 mg/kg黄芪多糖,可提高1~28日龄绍兴蛋雏鸭的免疫性能和抗氧化性能,改善肠道绒毛形态。Liu等[67]在饲粮中添加0.4×109~1.2×109 CFU/kg丁酸梭菌,提高了1~42日龄北京鸭肝脏抗氧化性能,降低了肝脂肪变性和肝细胞坏死,对皮质酮诱导的北京鸭肝损伤具有保护作用。

5 小结

综上所述,丁酸梭菌通过多种途径发挥抗病和促进肠道健康的作用,是一种优秀的绿色饲料添加剂,但是要取得与饲料用抗生素同样的杀菌、促生长效果,还有一定的差距。丁酸梭菌的研究重点应该放在:1)利用基因工程技术,改良丁酸梭菌菌株,使其既能够在肠道中快速产生大量丁酸,又能减少菌壁等自身成分对肠道的免疫刺激;2)深入研究丁酸梭菌抗病和促进肠道健康的主要作用机理;3)根据主要作用机理和各个品种畜禽的生理特点,寻找能够与丁酸梭菌搭配使用,完全替代饲料用抗生素的合适中草药、益生菌等,并合理配比使用;4)由于丁酸梭菌和肉毒梭状芽孢杆菌同为梭菌属,需深入研究丁酸梭菌将来可能出现的毒素基因水平转移等生物安全性问题。

参考文献
[1]
中华人民共和国农业农村部. 农业农村部关于印发《全国兽用抗菌药使用减量化行动方案(2021-2025年)》的通知[EB/OL]. (2021-10-25)[2021-10-25]. http://www.moa.gov.cn/govpublic/xmsyj/202110/t20211025_6380448.htm.
Ministry of Agriculture and Rural Affairs of the People's Republic of China. Notice of Ministry of Agriculture and Rural Affairs on printing and distributing "The national action plan for reducing the use of veterinary antimicrobial drugs (2021-2025)"[EB/OL]. (2021-10-25)[2021-10-25]. http://www.moa.gov.cn/govpublic/xmsyj/202110/t20211025_6380448.htm. (in Chinese)
[2]
贾丽楠. 丁酸梭菌抗逆性能及其对肉仔鸡益生性能的体外研究[D]. 硕士学位论文. 保定: 河北农业大学, 2018: 15-17.
JIA L N. Tolerance of Clostridium butyricum and its in vitro probiotic characteristics of broilers[D]. Master's Thesis. Baoding: Hebei Agricultural University, 2018: 15-17. (in Chinese)
[3]
GHODDUSI H B, SHERBURN R E, ABOABA O O. Growth limiting pH, water activity, and temperature for neurotoxigenic strains of Clostridium butyricum[J]. ISRN Microbiology, 2013, 2013: 731430.
[4]
李怀宇. 丁酸梭菌对肉鸡生长性能、屠体品质的影响及其机理研究[D]. 硕士学位论文. 杭州: 浙江大学, 2021: 6-9.
LI H Y. Effects of dietary supplementation of Clostridium butyricum on growth performance and carcass traits in broilers and its mechanism[D]. Master's Thesis. Hangzhou: Zhejiang University, 2021: 6-9. (in Chinese)
[5]
贾丽楠, 崔嘉, 李占一, 等. 丁酸梭菌对肉仔鸡肠道致病菌抑菌作用研究[J]. 中国家禽, 2017, 39(23): 22-25.
JIA L N, CUI J, LI Z Y, et al. Study on mechanism and inhibitory effect of Clostridium butyricum on bacteria in intestine of broilers[J]. China Poultry, 2017, 39(23): 22-25 (in Chinese).
[6]
CAMERINI S, MARCOCCI L, PICARAZZI L, et al. Type E botulinum neurotoxin-producing Clostridium butyricum strains are aerotolerant during vegetative growth[J]. mSystems, 2019, 4(2): e00299-18.
[7]
陈代文. 猪抗病营养理论与实践[M]. 北京: 中国农业大学出版社, 2012.
CHEN D W. Disease-resistant nutrition of swine: theory and practice[M]. Beijing: China Agricultural University Press, 2012 (in Chinese).
[8]
HAN Y S, TANG C H, LI Y, et al. Effects of dietary supplementation with Clostridium butyricum on growth performance, serum immunity, intestinal morphology, and microbiota as an antibiotic alternative in weaned piglets[J]. Animals, 2020, 10(12): 2887.
[9]
FU J, WANG T H, XIAO X, et al. Clostridium butyricum ZJU-F1 benefits the intestinal barrier function and immune response associated with its modulation of gut microbiota in weaned piglets[J]. Cells, 2021, 10(3): 527. DOI:10.3390/cells10030527
[10]
LYU W, YANG H, LI N, et al. Molecular characterization, developmental expression, and modulation of occludin by early intervention with Clostridium butyricum in Muscovy ducks[J]. Poultry Science, 2021, 100(8): 101271. DOI:10.1016/j.psj.2021.101271
[11]
YAN H, AJUWON K M. Butyrate modifies intestinal barrier function in IPEC-J2 cells through a selective upregulation of tight junction proteins and activation of the Akt signaling pathway[J]. PLoS One, 2017, 12(6): e0179586. DOI:10.1371/journal.pone.0179586
[12]
SUI S J, TIAN Z B, WANG Q C, et al. Clostridium butyricum promotes intestinal motility by regulation of TLR2 in interstitial cells of Cajal[J]. European Review for Medical and Pharmacological Sciences, 2018, 22(14): 4730-4738.
[13]
QI L L, LU X H, MAO H G, et al. Clostridium butyricum induces the production and glycosylation of mucins in HT-29 cells[J]. Frontiers in Cellular and Infection Microbiology, 2021, 11: 668766. DOI:10.3389/fcimb.2021.668766
[14]
王腾浩. 新型丁酸梭菌筛选及其对断奶仔猪生长性能和肠道功能影响的研究[D]. 博士学位论文. 杭州: 浙江大学, 2015: 83.
WANG T H. Screening of a novel Clostridium butyricum and its administration on growth performance and gut function in weaning piglets[D]. Ph. D. Thesis. Hangzhou: Zhejiang University, 2015: 83. (in Chinese)
[15]
GAO Q X, QI L L, WU T X, et al. Ability of Clostridium butyricum to inhibit Escherichia coli-induced apoptosis in chicken embryo intestinal cells[J]. Veterinary Microbiology, 2012, 160(3/4): 395-402.
[16]
WOO T D H, OKA K, TAKAHASHI M, et al. Inhibition of the cytotoxic effect of Clostridium difficile in vitro by Clostridium butyricum MIYAIRI 588 strain[J]. Journal of Medical Microbiology, 2011, 60(Pt 11): 1617-1625.
[17]
WANG T H, FU J, XIAO X, et al. CBP22, a novel bacteriocin isolated from Clostridium butyricum ZJU-F1, protects against LPS-induced intestinal injury through maintaining the tight junction complex[J]. Mediators of Inflammation, 2021, 2021: 8032125.
[18]
于洁, 范雪, 赵敏孟, 等. 丁酸梭菌和枯草芽孢杆菌对肉鹅生长性能、消化酶活性、抗氧化功能和肠道形态的影响[J]. 动物营养学报, 2021, 33(2): 860-868.
YU J, FAN X, ZHAO M M, et al. Effects of Clostridium butyricum and Bacillus subtilis on growth performance, digestive enzyme activities, antioxidant function and intestinal morphology of meat geese[J]. Chinese Journal of Animal Nutrition, 2021, 33(2): 860-868 (in Chinese). DOI:10.3969/j.issn.1006-267x.2021.02.027
[19]
WANG W W, WANG J, ZHANG H J, et al. Supplemental Clostridium butyricum modulates lipid metabolism through shaping gut microbiota and bile acid profile of aged laying hens[J]. Frontiers in Microbiology, 2020, 11: 600. DOI:10.3389/fmicb.2020.00600
[20]
GAO Q X, XIAO Y P, ZHANG C J, et al. Molecular characterization and expression analysis of Toll-like receptor 2 in response to bacteria in silvery pomfret intestinal epithelial cells[J]. Fish & Shellfish Immunology, 2016, 58: 1-9.
[21]
GAO Q X, QI L L, WU T X, et al. Clostridium butyricum activates TLR2-mediated MyD88-independent signaling pathway in HT-29 cells[J]. Molecular and Cellular Biochemistry, 2012, 361(1/2): 31-37.
[22]
HAYASHI A, NAGAO-KITAMOTO H, KITAMOTO S, et al. The butyrate-producing bacterium Clostridium butyricum suppresses Clostridioides difficile infection via neutrophil-and antimicrobial cytokine-dependent but GPR43/109a-independent mechanisms[J]. Journal of Immunology, 2021, 206(7): 1576-1585. DOI:10.4049/jimmunol.2000353
[23]
李玉鹏, 李海花, 王柳懿, 等. 丁酸梭菌对断奶仔猪生长性能、肠道屏障功能和血清细胞因子含量的影响[J]. 动物营养学报, 2017, 29(8): 2961-2968.
LI Y P, LI H H, WANG L Y, et al. Effects of Clostridium butyricum on growth performance, intestinal barrier function and serum cytokine contents of weaned piglets[J]. Chinese Journal of Animal Nutrition, 2017, 29(8): 2961-2968 (in Chinese). DOI:10.3969/j.issn.1006-267x.2017.08.041
[24]
何菊, 胡迪, 郭云清, 等. 丁酸梭菌CB1对肉鸡免疫器官指数、黏膜SIgA抗体和血清生化指标的影响[J]. 中国兽医学报, 2018, 38(5): 998-1002, 1007.
HE J, HU D, GUO Y Q, et al. Influence of Clostridium butyricum CB1 preparations on immune organ index, mucous membrane SIgA and serum biochemical parameters of broiler chickens[J]. Chinese Journal of Veterinary Science, 2018, 38(5): 998-1002, 1007 (in Chinese).
[25]
TERADA T, NⅡ T, ISOBE N, et al. Effects of probiotics Lactobacillus reuteri and Clostridium butyricum on the expression of Toll-like receptors, pro-and anti-inflammatory cytokines, and antimicrobial peptides in broiler chick intestine[J]. Journal of Poultry Science, 2020, 57(4): 310-318. DOI:10.2141/jpsa.0190098
[26]
WANG J B, QI L L, WU Z G, et al. Different effects of lipoteichoic acid from C. butyricum and S. aureus on inflammatory responses of HT-29 cells[J]. International Journal of Biological Macromolecules, 2016, 87: 481-487. DOI:10.1016/j.ijbiomac.2016.03.010
[27]
GAO Q X, QI L L, WU T X, et al. An important role of interleukin-10 in counteracting excessive immune response in HT-29 cells exposed to Clostridium butyricum[J]. BMC Microbiology, 2012, 12: 100. DOI:10.1186/1471-2180-12-100
[28]
KASHIWAGI I, MORITA R, SCHICHITA T, et al. Smad2 and Smad3 inversely regulate TGF-β autoinduction in Clostridium butyricum-activated dendritic cells[J]. Immunity, 2015, 43(1): 65-79. DOI:10.1016/j.immuni.2015.06.010
[29]
ISONO A, KATSUNO T, SATO T, et al. Clostridium butyricum TO-A culture supernatant downregulates TLR4 in human colonic epithelial cells[J]. Digestive Diseases and Sciences, 2007, 52(11): 2963-2971. DOI:10.1007/s10620-006-9593-3
[30]
ZHAO X N, YANG J, JU Z J, et al. Clostridium butyricum ameliorates Salmonella enteritis induced inflammation by enhancing and improving immunity of the intestinal epithelial barrier at the intestinal mucosal level[J]. Frontiers in Microbiology, 2020, 11: 299. DOI:10.3389/fmicb.2020.00299
[31]
YANG W J, YU T M, HUANG X S, et al. Intestinal microbiota-derived short-chain fatty acids regulation of immune cell IL-22 production and gut immunity[J]. Nature Communications, 2020, 11(1): 4457. DOI:10.1038/s41467-020-18262-6
[32]
TAKAKUWA A, NAKAMURA K, KIKUCHI M, et al. Butyric acid and leucine induce α-defensin secretion from small intestinal Paneth cells[J]. Nutrients, 2019, 11(11): 2817. DOI:10.3390/nu11112817
[33]
LI H H, LI Y P, ZHU Q, et al. Dietary supplementation with Clostridium butyricum helps to improve the intestinal barrier function of weaned piglets challenged with enterotoxigenic Escherichia coli K88[J]. Journal of Applied Microbiology, 2018, 125(4): 964-975. DOI:10.1111/jam.13936
[34]
ZHAO Q, YANG W R, WANG X H, et al. Clostridium butyricum alleviates intestinal low-grade inflammation in TNBS-induced irritable bowel syndrome in mice by regulating functional status of lamina propria dendritic cells[J]. World Journal of Gastroenterology, 2019, 25(36): 5469-5482. DOI:10.3748/wjg.v25.i36.5469
[35]
王金丹, 乔莞宁, 朱静, 等. 酪酸梭菌代谢产物丁酸和氢气对急性胃黏膜损伤的作用[J]. 中国病理生理杂志, 2017, 33(10): 1906-1911.
WANG J D, QIAO G N, ZHU J, et al. Effect of metabolites of Clostridium butyricum, butyric acid and hydrogen, on acute gastric mucosal lesion[J]. Chinese Journal of Pathophysiology, 2017, 33(10): 1906-1911 (in Chinese). DOI:10.3969/j.issn.1000-4718.2017.10.029
[36]
DOU C X, SHANG Z Y, QIAO J Y, et al. Clostridium butyricum protects IPEC-J2 cells from ETEC K88-induced oxidative damage by activating the Nrf2/ARE signaling pathway[J]. Oxidative Medicine and Cellular Longevity, 2021, 2021: 4464002.
[37]
尚智援, 窦彩霞, 王克玮, 等. 丁酸梭菌通过激活p38丝裂原活化蛋白激酶/核因子E2相关因子信号通路减轻猪霍乱沙门氏菌导致的猪小肠上皮细胞氧化损伤[J]. 动物营养学报, 2021, 33(12): 7105-7117.
SHANG Z Y, DOU C X, WANG K W, et al. Clostridium butyricum alleviates porcine intestinal epithelial cells oxidative damage induced by Salmonella cholerae through activating p38 mitogen activated protein kinase/nuclear factor E2 related factor signaling pathway[J]. Chinese Journal of Animal Nutrition, 2021, 33(12): 7105-7117 (in Chinese).
[38]
SLEZÁK J, KURA B, FRIMMEL K, et al. Preventive and therapeutic application of molecular hydrogen in situations with excessive production of free radicals[J]. Physiological Research, 2016, 65(Suppl.1): S11-S28.
[39]
WANG J B, QI L L, MEI L H, et al. C. butyricum lipoteichoic acid inhibits the inflammatory response and apoptosis in HT-29 cells induced by S. aureus lipoteichoic acid[J]. International Journal of Biological Macromolecules, 2016, 88: 81-87. DOI:10.1016/j.ijbiomac.2016.03.054
[40]
王倩, 李海花, 窦彩霞, 等. 一株丁酸梭菌的分离鉴定及其益生特性研究[J]. 中国畜牧兽医, 2020, 47(1): 258-266.
WANG Q, LI H H, DOU C X, et al. Isolation, identification and probiotic characteristics of a Clostridium butyricum[J]. China Animal Husbandry & Veterinary Medicine, 2020, 47(1): 258-266 (in Chinese).
[41]
杨华, 施杏芬, 桂国弘, 等. 丁酸梭菌对产肠毒素大肠杆菌刺激猪肠道上皮细胞炎症反应的抑制效果[J]. 动物营养学报, 2019, 31(12): 5688-5695.
YANG H, SHI X F, GUI G H, et al. Inhibitory effect of Clostridium butyricum on inflammatory reaction of porcine intestinal epithelial cells induced by enterotoxigenic Escherichia coli[J]. Chinese Journal of Animal Nutrition, 2019, 31(12): 5688-5695 (in Chinese). DOI:10.3969/j.issn.1006-267x.2019.12.035
[42]
HAGIHARA M, KUROKI Y, ARIYOSHI T, et al. Clostridium butyricum modulates the microbiome to protect intestinal barrier function in mice with antibiotic-induced dysbiosis[J]. iScience, 2020, 23(1): 100772. DOI:10.1016/j.isci.2019.100772
[43]
LUO X S, KONG Q, WANG Y M, et al. Colonization of Clostridium butyricum in rats and its effect on intestinal microbial composition[J]. Microorganisms, 2021, 9(8): 1573. DOI:10.3390/microorganisms9081573
[44]
MIAO R X, ZHU X X, WAN C M, et al. Effect of Clostridium butyricum supplementation on the development of intestinal flora and the immune system of neonatal mice[J]. Experimental and Therapeutic Medicine, 2018, 15(1): 1081-1086.
[45]
ARIYOSHI T, HAGIHARA M, TOMONO S, et al. Clostridium butyricum MIYAIRI 588 modifies bacterial composition under antibiotic-induced dysbiosis for the activation of interactions via lipid metabolism between the gut microbiome and the host[J]. Biomedicines, 2021, 9(8): 1065. DOI:10.3390/biomedicines9081065
[46]
ARIYOSHI T, HAGIHARA M, EGUCHI S, et al. Clostridium butyricum MIYAIRI 588-induced protectin D1 has an anti-inflammatory effect on antibiotic-induced intestinal disorder[J]. Frontiers in Microbiology, 2020, 11: 587725. DOI:10.3389/fmicb.2020.587725
[47]
ISA K, OKA K, BEAUCHAMP N, et al. Safety assessment of the Clostridium butyricum MIYAIRI 588® probiotic strain including evaluation of antimicrobial sensitivity and presence of Clostridium toxin genes in vitro and teratogenicity in vivo[J]. Human & Experimental Toxicology, 2016, 35(8): 818-832.
[48]
易至, 丁洁琼, 王鸿超, 等. 基于比较基因组学的丁酸梭菌遗传多样性及生物学特性[J]. 食品与发酵工业, 2020, 46(10): 1-7.
YI Z, DING J Q, WANG H C, et al. Genetic diversity and biological characteristics of Clostridium butyricum based on comparative genomics[J]. Food and Fermentation Industries, 2020, 46(10): 1-7 (in Chinese).
[49]
范伟祥, 曹艳丽, 崔璐璐, 等. 24株鸡源丁酸梭菌的分离鉴定及耐药基因与毒力基因携带情况[J]. 微生物学报, 2021, 61(1): 115-126.
FAN W X, CAO Y L, CUI L L, et al. Detection of resistance and virulence genes from 24 Clostridium butyricum strains isolated from chickens[J]. Acta Microbiologica Sinica, 2021, 61(1): 115-126 (in Chinese).
[50]
高文文, 尚佳萃, 周雪, 等. 一株高产丁酸的丁酸梭菌分离鉴定及其生物学性质研究[J]. 食品工业科技, 2020, 41(7): 82-88, 101.
GAO W W, SHANG J C, ZHOU X, et al. Isolation and identification of a strain of Clostridium butyricum with high yield of butyric acid and its biological characteristics[J]. Science and Technology of Food Industry, 2020, 41(7): 82-88, 101 (in Chinese).
[51]
SULTHANA A, THORRAMAMIDI A, LAKSHMI S G, et al. Whole-genome shotgun sequencing and characterization of probiotic strain Clostridium butyricum UBCB 70 to assess its safety[J]. Microbiology Resource Announcements, 2019, 8(5): e01732-18.
[52]
董银苹, 江涛, 赵帅, 等. 我国首例由丁酸梭菌引起婴儿E型肉毒中毒实验室诊断研究[J]. 中国食品卫生杂志, 2020, 32(5): 499-503.
DONG Y P, JIANG T, ZHAO S, et al. Laboratory investigation of the first infant botulism case caused by type E botulinum neurotoxin producing Clostridium butyricum in China[J]. Chinese Journal of Food Hygiene, 2020, 32(5): 499-503 (in Chinese).
[53]
NAWROCKI E M, BRADSHAW M, JOHNSON E A. Botulinum neurotoxin-encoding plasmids can be conjugatively transferred to diverse clostridial strains[J]. Scientific Reports, 2018, 8(1): 3100. DOI:10.1038/s41598-018-21342-9
[54]
EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP), BAMPIDIS V, AZIMONTI G, et al. Safety and efficacy of the feed additive consisting of Clostridium butyricum FERM BP-2789 (Miya-Gold® S) for chickens for fattening, chickens reared for laying, turkeys for fattening, turkeys reared for breeding, minor avian species (excluding laying birds), piglets (suckling and weaned) and minor porcine species (Miyarisan Pharmaceutical Co. Ltd. )[J]. EFSA Journal. European Food Safety Authority, 2021, 19(3): e06450.
[55]
CHEN L, LI S, ZHENG J, et al. Effects of dietary Clostridium butyricum supplementation on growth performance, intestinal development, and immune response of weaned piglets challenged with lipopolysaccharide[J]. Journal of Animal Science and Biotechnology, 2018, 9: 62. DOI:10.1186/s40104-018-0275-8
[56]
LAN R X, TRAN H, KIM I. Effects of probiotic supplementation in different nutrient density diets on growth performance, nutrient digestibility, blood profiles, fecal microflora and noxious gas emission in weaning pig[J]. Journal of the Science of Food and Agriculture, 2017, 97(4): 1335-1341. DOI:10.1002/jsfa.7871
[57]
WANG S L, ZHANG S Y, HUANG S M, et al. Resistant maltodextrin alleviates dextran sulfate sodium-induced intestinal inflammatory injury by increasing butyric acid to inhibit proinflammatory cytokine levels[J]. BioMed Research International, 2020, 2020: 7694734.
[58]
CAO M, LI Y, WU Q J, et al. Effects of dietary Clostridium butyricum addition to sows in late gestation and lactation on reproductive performance and intestinal microbiota1[J]. Journal of Animal Science, 2019, 97(8): 3426-3439. DOI:10.1093/jas/skz186
[59]
ZHANG L, ZHANG L L, ZHAN X A, et al. Effects of dietary supplementation of probiotic, Clostridium butyricum, on growth performance, immune response, intestinal barrier function, and digestive enzyme activity in broiler chickens challenged with Escherichia coli K88[J]. Journal of Animal Science and Biotechnology, 2016, 7: 3. DOI:10.1186/s40104-016-0061-4
[60]
ABDEL-LATIF M A, ABD EL-HACK M E, SWELUM A A, et al. Single and combined effects of Clostridium butyricum and Saccharomyces cerevisiae on growth indices, intestinal health, and immunity of broilers[J]. Animals, 2018, 8(10): 184. DOI:10.3390/ani8100184
[61]
HUANG T, PENG X Y, GAO B, et al. The effect of Clostridium butyricum on gut microbiota, immune response and intestinal barrier function during the development of necrotic enteritis in chickens[J]. Frontiers in Microbiology, 2019, 10: 2309. DOI:10.3389/fmicb.2019.02309
[62]
WEI X Y, FU X D, XIAO M S, et al. Dietary galactosyl and mannosyl carbohydrates: in-vitro assessment of prebiotic effects[J]. Food Chemistry, 2020, 329: 127179. DOI:10.1016/j.foodchem.2020.127179
[63]
ZHAN H Q, DONG X Y, LI L L, et al. Effects of dietary supplementation with Clostridium butyricum on laying performance, egg quality, serum parameters, and cecal microflora of laying hens in the late phase of production[J]. Poultry Science, 2019, 98(2): 896-903. DOI:10.3382/ps/pey436
[64]
张茜, 潘翠, 张钊. 丁酸梭菌联合竹醋液对产蛋后期蛋鸡蛋品质及肠道功能的影响[J]. 饲料研究, 2021, 44(13): 57-60.
ZHANG Q, PAN C, ZHANG Z. Effect of Clostridium butyricum combined with bamboo vinegar on egg quality and intestinal function of laying hens in later laying period[J]. Feed Research, 2021, 44(13): 57-60 (in Chinese).
[65]
李娜, 杨华, 吕文涛, 等. 丁酸梭菌早期干预对幼龄番鸭生长性能、肠道黏膜形态及肠屏障功能的影响[J]. 中国畜牧杂志, 2021, 57(4): 200-206.
LI N, YANG H, LV W T, et al. Effects of early intervention of Clostridium butyricum on growth performance, intestinal mucosal morphology and intestinal barrier function of young Muscovy duck[J]. Chinese Journal of Animal Science, 2021, 57(4): 200-206 (in Chinese).
[66]
赵文文, 袁文华, 沈军达, 等. 黄芪多糖和丁酸梭菌对蛋雏鸭免疫性能、抗氧化性能以及肠道形态的影响[J]. 动物营养学报, 2018, 30(10): 4143-4150.
ZHAO W W, YUAN W H, SHEN J D, et al. Effects of Astragalus polysaccharides and Clostridium butyricum on immune function, antioxidant ability and intestinal morphology of laying ducklings[J]. Chinese Journal of Animal Nutrition, 2018, 30(10): 4143-4150 (in Chinese). DOI:10.3969/j.issn.1006-267x.2018.10.040
[67]
LIU Y H, LIU C, HUANG L Q, et al. A discovery of relevant hepatoprotective effects and underlying mechanisms of dietary Clostridium butyricum against corticosterone-induced liver injury in Pekin ducks[J]. Microorganisms, 2019, 7(9): 358. DOI:10.3390/microorganisms7090358