青海牦牛源益生菌分离、鉴定及其益生特性的研究  

Study on Isolation,Identification and Probiotic Properties of Qinghai Yak Derived Probiotics

作　　者：谢林林 曹瑾 赵玉琪 张欣 郝力壮[2] 简莹娜 孙偲 李军训 宋洪宁 王楠[1] XIE Linlin;CAO Jin;ZHAO Yuqi;ZHANG Xin;HAO Lizhuang;JIAN Yingna;SUN Cai;LI Junxun;SONG Hongning;WANG Nan(Key Laboratory of Industrial Fermentation Microbiology,Ministry of Education,College of Biotechnology,Tianjin University of Science and Technology,Tianjin 300457,China;Qinghai Plateau Grazing Livestock Key Laboratory of Animal Nutrition and Feed Science,Qinghai University,Xining 810016,China;Qinghai Provincial Key Laboratory of Animal Disease Pathogen Diagnosis and Green Prevention and Control Technology Research,College of Animal Husbandry and Veterinary Science,Qinghai University,Xining 810016,China;Qinghai Baifenniu Biotechnology Co.,Ltd.,Xining 810016,China;Shandong Taishan Shengliyuan Group Co.,Ltd.,Tai’an 271000,China)

机构地区：[1]天津科技大学生物工程学院,教育部工业发酵微生物学重点实验室,天津300457 [2]青海大学,青海省高原放牧家畜动物营养与饲料科学重点实验室,西宁810016 [3]青海大学畜牧兽医科学院,青海省动物疾病病原诊断与绿色防控技术研究重点实验室,西宁810016 [4]青海百分牛生物科技有限公司,西宁810016 [5]山东泰山生力源集团股份有限公司,泰安271000

出　　处：《动物营养学报》2025年第1期658-671,共14页CHINESE JOURNAL OF ANIMAL NUTRITION

基　　金：青海省高端创新创业人才-柔性引进领军人才项目;国家重点研发项目子课题(2022YFD1302103);青海省高原放牧家畜动物营养与饲料科学重点实验室评估奖励项目;泰山产业领军人才工程专项。

摘　　要：本研究旨在从青海省牦牛源样品中分离筛选可用于改善牦牛腹泻的潜在益生菌菌株,并评价其体外益生特性。试验首先从健康牦牛粪便和酸奶中分离纯化出乳酸菌,通过菌落形态观察、16S rDNA以及对部分菌株进行保守基因rpoA测序,并进行BLAST比对,对筛选菌株进行鉴定。然后,通过评价菌株的生长、耐酸、耐胆盐、自动聚集能力和疏水性等特性筛选出优异菌株,重点考察其对来自腹泻致死牦牛中分离的致病菌的抑制能力,同时对其抗生素耐药性、细胞黏附能力、抗氧化能力[2,2-二苯基-1-苦基肼(DPPH)自由基清除率、羟自由基清除率及还原能力]及抗炎能力[巨噬细胞释放一氧化氮(NO)]进行测定。结果表明:试验共获得8株乳酸菌,其中1株为罗伊氏黏液乳杆菌,3株为植物乳植杆菌,4株为屎肠球菌。其中,罗伊氏黏液乳杆菌QM11具有较强的耐酸和耐胆盐能力;自动聚集率和疏水率分别为(64.07±1.36)%和(26.90±1.51)%;对牦牛源致病性大肠杆菌QH01、QH02和QH03的抑菌率分别为(87.29±0.98)%、(74.67±1.28)%和(86.43±0.21)%,对金黄色葡萄球菌、单增李斯特菌和鼠伤寒沙门氏菌等多种病原指示菌的抑菌率达74%以上;对氨苄西林、青霉素、庆大霉素和链霉素等多种抗生素敏感。与对照菌株鼠李糖乳杆菌GG相比,罗伊氏黏液乳杆菌QM11对结肠细胞的黏附能力更强,黏附率为(54.38±2.15)%;菌悬液的抗氧化能力更强,DPPH自由基清除率、羟自由基清除率和还原能力分别为(4.19±1.03)%、(180.29±9.28)%和(6.07±0.60)%;抑制巨噬细胞释放NO的能力与鼠李糖乳杆菌GG相当。综上所述,本试验筛选出的罗伊氏黏液乳杆菌QM11可作为后续研制用于防治牦牛腹泻益生菌制剂的备选菌株。The aim of this study was to isolate and screen potential probiotic strains for preventing and treating yak diarrhea from yak source samples in Qinghai Province,and to evaluate their probiotic properties in vitro.Firstly,lactic acid bacteria were isolated and purified from healthy yak feces and yogurt,and the selected strains were identified through colony morphology observation,16S rDNA sequencing,and conserved gene rpoA sequencing of some strains,followed by BLAST comparison.Then,excellent strains were selected by evaluating their growth,acid and bile salt resistance,auto-aggregation ability,hydrophobicity,and especially their inhibition ability against pathogenic bacteria isolated from yaks that died of diarrhea.At the same time,the antibiotic resistance,cell adhesion,antioxidant ability[2,2-diphenyl-1-picrylhydrazyl(DPPH)radical scavenging rate,hydroxyl radical scavenging rate and reducing ability]and anti-inflammatory ability[macrophage release of nitric oxide(NO)]were measured.The results showed that a total of 8 strains of lactic acid bacteria were obtained,in which 1 strain was Limosilactobacillus reuteri,3 strains were Lactiplantibacillus plantarum and 4 strains were Enterococcus faecium.Among them,Limosilactobacillus reuteri QM11 had strong acid and bile salt resistance;the auto-aggregation rate and hydrophobicity were(64.07±1.36)%and(26.90±1.51)%,respectively;the inhibition rates against yak pathogenic Escherichia coli QH01,QH02 and QH03 were(87.29±0.98)%,(74.67±1.28)%and(86.43±0.21)%,respectively;the inhibition rates against Staphylococcus aureus,Listeria monocytogenes,Salmonella typhimurium and other pathogenic indicator bacteria were more than 74%;it is sensitive to ampicillin,penicillin,gentamicin,streptomycin and many other antibiotics.Compared with the control strain Lactobacillus rhamnosus GG,Limosilactobacillus reuteri QM11 had stronger adhesion to colon cells,and the adhesion rate was(54.38±2.15)%;the antioxidant capacity of bacterial suspension was stronger,and the DPPH radical scavengi

关 键 词：牦牛 益生菌 筛选 益生特性 

分 类 号：S811[农业科学—畜牧学]