CN117327626A - 一种植物乳杆菌ts1及其培养方法与应用 - Google Patents
一种植物乳杆菌ts1及其培养方法与应用 Download PDFInfo
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- CN117327626A CN117327626A CN202311335181.9A CN202311335181A CN117327626A CN 117327626 A CN117327626 A CN 117327626A CN 202311335181 A CN202311335181 A CN 202311335181A CN 117327626 A CN117327626 A CN 117327626A
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- lactobacillus plantarum
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Classifications
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Abstract
本发明提供了一种植物乳杆菌TS1及其培养方法与应用,属于微生物技术领域,所述植物乳杆菌为植物乳杆菌Lactobacillusplantarum TS1,已于2023年5月22日保藏于中国典型培养物保藏中心,保藏编号为CCTCC NO:M 2023797。本发明通过从浆水中分离、纯化获得所述植物乳杆菌Lactobacillusplantarum TS1,研究发现,该菌株能够产生苯乳酸,对金黄色葡萄球菌、芽孢杆菌、铜绿假单胞菌、大肠杆菌和枯草芽孢杆菌具有较好的抑制作用,能够在人工胃液和肠液中生存,可用作抗生素替代品使用,将该菌株应用于发酵饲料的制备对于解决抗生素耐药性的问题具有重大意义。
Description
技术领域
本发明属于微生物技术领域,尤其涉及一种植物乳杆菌TS1及其培养方法与应用。
背景技术
抗生素作为饲料添加剂,在促进动物生长、预防疾病等方面发挥着重要作用。但抗生素的广泛应用却引出了抗生素残留、超级耐药菌等问题的产生,严重危害了动物源食品安全及公共卫生安全。因此,寻找一种可替代抗生素的天然产物对畜牧养殖行业极为重要。
苯乳酸(phenyllactic acid,PLA)是一种存在于天然蜂蜜中的酚酸,具有广谱的抗菌特性;其理化性质稳定,能够耐受高温和低pH值环境,在蛋白酶作用下可保持抗菌活性;且PLA是一种公认的、安全的微生物乳酸菌代谢产生的第三代抑菌物质,对人和动物的细胞均不会造成伤害;同时,PLA不但能够适应制粒和蒸煮等饲料加工的高温环境的同时,而且还能够适应青贮、生物发酵、动物胃肠道等环境。因此,PLA有望作为一种新的抗生素替代品使用。
目前,已知能够产生PLA的菌株包括枯草芽孢杆菌、植物乳杆菌、乳酸片球菌、凝结芽孢杆菌、副干酪乳杆菌等。而对于能够高产PLA的菌株还尚未发现,因此,获得一种高产PLA的菌株以替代抗生素的使用对于解决畜牧养殖业现有抗生素的问题极为重要。
发明内容
有鉴于此,本发明的目的在于提供一种植物乳杆菌及其培养方法与应用,本发明从浆水中分离、纯化获得了一株植物乳杆菌TS1,该菌株具有较好的益生性能,且能够产生苯乳酸,且苯乳酸的产生能够与有机酸发挥协同作用,产生更强的抑菌作用。
为实现上述目的,本发明提供了以下技术方案:
本发明提供了一种植物乳杆菌,所述植物乳杆菌为植物乳杆菌Lactobacillusplantarum TS1,于2023年5月22日保藏于中国典型培养物保藏中心,保藏编号为CCTCC NO:M 2023797。
本发明还提供了所述植物乳杆菌的培养方法,将植物乳杆菌Lactobacillusplantarum TS1接种于培养基上进行培养。
优选的,所述培养的温度为30~40℃,所述培养的时间为20~28h。
本发明还提供了所述植物乳杆菌在制备改善肠道菌群的药物中的应用。
本发明还提供了所述植物乳杆菌在制备改善肠道菌群的食品或保健品中的应用。
本发明还提供了所述植物乳杆菌在制备发酵食品中的应用。
本发明还提供了所述植物乳杆菌在制备抗生素替代品中的应用。
本发明还提供了所述植物乳杆菌在制备动物饲料中的应用。
优选的,所述动物饲料包括青贮饲料和饲料添加剂。
本发明还提供了所述植物乳杆菌在合成苯乳酸中的应用。
与现有技术相比,本发明具有如下有益效果:
本发明从浆水中分离纯化获得的菌株TS1的菌落大小为1~2mm,呈乳白色,表面湿润、中央凸起,镜检形态为短杆状;该菌株可产生苯乳酸,具有良好的耐酸、耐胆盐性能,能够在人工胃肠液中生存一段时间,因此,可应用于食品、保健品或药品中;同时,该菌株对金黄色葡萄球菌、芽孢杆菌、铜绿假单胞菌、大肠杆菌和枯草芽孢杆菌均具有很好的抑制能力,可应用于发酵饲料的制备。该菌株不但能够在动物肠道中存活,促进动物肠道蠕动,提高饲料的利用率;还可以替代饲料中抗生素的添加,使用绿色环保的方式用于饲料样品的保存,使得益生菌发酵饲料真正可以有效应用于饲料生产及家畜养殖中。
附图说明
图1为菌株16S rDNA基因扩增产物,其中,从左至右依次为菌株TS的PCR结果,菌株TS1的PCR结果,菌株TS2的PCR结果,菌株DBb4的PCR结果,菌株YB1的PCR结果,菌株QJS1.1的PCR结果,菌株QJS1.2的PCR结果,菌株QJS2.1的PCR结果,marker;
图2为菌株基于16S rDNA的系统发育树;
图3为菌株在平板上的菌落形态;
图4为菌株的革兰氏染色结果;
图5菌株耐酸试验的结果;
图6为菌株耐胆盐试验的结果;
图7为菌株耐人工胃液的试验结果;
图8为菌株耐人工肠液的试验结果;
图9为菌株耐药试验结果;
图10为菌株抑菌试验的结果。
生物保藏说明
植物乳杆菌Lactobacillusplantarum TS1,保藏于中国典型培养物保藏中心(CCTCC),保藏地址为中国.武汉.武汉大学,保藏日期为2023年5月22日,保藏编号为CCTCCNO:M 2023797。
具体实施方式
本发明提供了一种植物乳杆菌,所述植物乳杆菌为植物乳杆菌Lactobacillusplantarum TS1,于2023年5月22日保藏于中国典型培养物保藏中心,保藏编号为CCTCC NO:M 2023797。
本发明还提供了所述植物乳杆菌的培养方法,将植物乳杆菌Lactobacillusplantarum TS1接种于培养基上进行培养。
在本发明中,所述培养基优选为MRS琼脂固体培养基,所述MRS琼脂固体培养基的配方优选为葡萄糖18~22g、蛋白胨8~12g、牛肉膏8~12g、酵母浸粉3~7g、结晶乙酸钠3~7g、柠檬酸三铵1~3g、磷酸氢二钾1~3g、吐温-800.5~1.5mL、硫酸镁0.50~0.60g、硫酸锰0.2~0.3g,蒸馏水800~1200mL,pH值6.2~6.4;进一步优选为葡萄糖19~21g、蛋白胨9~11g、牛肉膏9~11g、酵母浸粉4~6g、结晶乙酸钠4~6g、柠檬酸三铵2g、磷酸氢二钾2g、吐温-800.7~1.3mL、硫酸镁0.54~0.56g、硫酸锰0.23~0.27g,蒸馏水900~1100mL,pH值6.25~6.35;更进一步优选为葡萄糖20g、蛋白胨10g、牛肉膏10g、酵母浸粉5g、结晶乙酸钠5g、柠檬酸三铵3g、磷酸氢二钾2g、吐温-80 1.0mL、硫酸镁0.58g、硫酸锰0.25g,蒸馏水1000mL,pH值6.3;所述培养的温度优选为30~40℃,进一步优选为33~37℃,更进一步优选为34~36℃,再进一步优选为35℃;所述培养的时间优选为20~28h,进一步优选为23~27h,更进一步优选为24~26h,再进一步优选为25h。
本发明还提供了所述植物乳杆菌在制备改善肠道菌群的药物中的应用。
本发明还提供了所述植物乳杆菌在制备改善肠道菌群的食品或保健品中的应用。
本发明还提供了所述植物乳杆菌在制备发酵食品中的应用。
本发明还提供了所述植物乳杆菌在制备抗生素替代品中的应用。
本发明还提供了所述植物乳杆菌在制备动物饲料中的应用。
在本发明中,所述动物饲料优选的包括青贮饲料和饲料添加剂。
本发明还提供了所述植物乳杆菌在合成苯乳酸中的应用。
下面结合实施例对本发明提供的技术方案进行详细的说明,但是不能把它们理解为对本发明保护范围的限定。
实施例1
1.1菌种的分离、纯化
取1mL甘肃省天水市秦安县兴国镇蔚林村浆水接入富集培养基中,35℃培养24h进行菌种富集。取0.1mL经稀释的菌体浓度为108CFU/mL的菌液接入初筛下层培养基,35℃培养24h后,注入初筛上层培养基,继续培养24h,选取大小为4~5mm的溶钙圈的菌落接种于MRS固体培养基中进行5次划线纯化,分离纯化后观察菌落形态,并置于4℃冰箱斜面保存。结果如图3所示。
其中,MRS固体培养基的配方为:葡萄糖20g、蛋白胨10g、牛肉膏10g、酵母浸粉5g、结晶乙酸钠5g、柠檬酸三铵2g、磷酸氢二钾2.0g、吐温-801mL、硫酸镁0.58g、硫酸锰0.25g,琼脂18g,pH值6.2,蒸馏水1000mL,在121℃,灭菌20min;
MRS液体培养基的配方为:葡萄糖20g、蛋白胨10g、牛肉膏10g、酵母浸粉5g、结晶乙酸钠5g、柠檬酸三铵2g、磷酸氢二钾2.0g、吐温-80 1mL、硫酸镁0.58g、硫酸锰0.25g,pH值6.4,蒸馏水1000mL,于121℃灭菌20min;
MRS琼脂培养基的配方为:蛋白胨10.0g,牛肉浸粉8.0g,酵母浸粉4.0g,葡萄糖20.0g,磷酸氢二钾2.0g,柠檬酸三铵2.0g,乙酸钠5.0g,硫酸镁0.2g,硫酸锰0.04g,琼脂14.0g,吐温-80 1.0mL,pH值6.5,蒸馏水1000mL,25℃;
富集培养基的配方为MRS液体培养基中添加2.0g/L的苯乳酸;
初筛下层培养基的配方为:MRS琼脂培养基中添加2.0g/L的苯丙氨酸;
初筛上层培养基的配方为:MRS琼脂培养基中添加30g/L的碳酸钙。
结果显示,分离纯化获得的菌株的菌落大小约1-2mm,表面湿润、中央凸起,镜检形态为短杆状,单个或成对排列,将分离纯化获得的菌株命名为TS1。
1.2菌株生理生化鉴定
对分离纯化获得的菌株进行革兰氏染色,并使用细菌生化鉴定管进行生理生化鉴定试验。结果如表1和图4所示。
表1生理生化试验结果
由表1和图4可知,菌株为兼性厌氧菌,革兰氏染色结果呈蓝紫色,属于革兰氏阳性菌;该菌株能利用阿拉伯糖、甘露糖、葡萄糖等,硝酸盐还原为阴性,能在6.5%高盐肉汤上生长。
1.316S rDNA分子鉴定
采用16S rDNA法对所分纯的菌株进行分子鉴定。16S rDNA扩增及序列测定由上海生物工程股份有限公司完成。
菌株TS1的16S rDNA核苷酸序列为:AGGCTCAGGACGAACGCTGGCGGCGTGCCTAATACATGCAAGTCGAACGAACTCTGGTATTGATTGGTGCTTGCATCATGATTTACATTTGAGTGAGTGGCGAACTGGTGAGTAACACGTGGGAAACCTGCCCAGAAGCGGGGGATAACACCTGGAAACAGATGCTAATACCGCATAACAACTTGGACCGCATGGTCCGAGCTTGAAAGATGGCTTCGGCTATCACTTTTGGATGGTCCCGCGGCGTATTAGCTAGATGGTGGGGTAACGGCTCACCATGGCAATGATACGTAGCCGACCTGAGAGGGTAATCGGCCACATTGGGACTGAGACACGGCCCAAACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCACAATGGACGAAAGTCTGATGGAGCAACGCCGCGTGAGTGAAGAAGGGTTTCGGCTCGTAAAACTCTGTTGTTAAAGAAGAACATATCTGAGAGTAACTGTTCAGGTATTGACGGTATTTAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCGGTTTTTTAAGTCTGATGTGAAAGCCTTCGGCTCAACCGAAGAAGTGCATCGGAAACTGGGAAACTTGAGTGCAGAAGAGGACAGTGGAACTCCATGTGTAGCGGTGAAATGCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGGCTGTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGTATGGGTAGCAAACAGGATTAGATACCCTGGTAGTCCATACCGTAAACGATGAATGCTAAGTGTTGGAGGGTTTCCGCCCTTCAGTGCTGCAGCTAACGCATTAAGCATTCCGCCTGGGGAGTACGGCCGCAAGGCTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCTACGCGAAGAACCTTACCAGGTCTTGACATACTATGCAAATCTAAGAGATTAGACGTTCCCTTCGGGGACATGGATACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTATTATCAGTTGCCAGCATTAAGTTGGGCACTCTGGTGAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGATGGTACAACGAGTTGCGAACTCGCGAGAGTAAGCTAATCTCTTAAA GCCATTCTCAGTTCGGATTGTAGGCTGCAACTCGCCTACATGAAGTCGGAATCGCTAGTAATCGCGGATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGAGAGTTTGTAACACCCAAAGTCGGT(SEQ ID No.1)
测序完成将菌株TS1的16S rDNA序列置于NCBI(National Center forBiotechnology Information,美国国立生物技术信息中心)数据库中进行比对,结果发现,菌株TS1的基因序列,在所有相似的序列中,与植物乳杆菌(Lactobacillus plantarum)K-J122在同一簇上。结合上述测得的生理生化特征,鉴定菌株TS1是植物乳杆菌。
其中,植物乳杆菌K-J122的核苷酸序列如下:AGTTTGGAACATGGCTCAGGACGAACGCTGGCGGCGTGCCTAATACATGCAAGTCGAACGAACTCTGGTATTGATTGGTGCTTGCATCATGATTTACATTTGAGTGAGTGGCGAACTGGTGAGTAACACGTGGGAAACCTGCCCAGAAGCGGGGGATAACACCTGGAAACAGATGCTAATACCGCATAACAACTTGGACCGCATGGTCCGAGCTTGAAAGATGGCTTCGGCTATCACTTTTGGATGGTCCCGCGGCGTATTAGCTAGATGGTGGGGTAACGGCTCACCATGGCAATGATACGTAGCCGACCTGAGAGGGTAATCGGCCACATTGGGACTGAGACACGGCCCAAACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCACAATGGACGAAAGTCTGATGGAGCAACGCCGCGTGAGTGAAGAAGGGTTTCGGCTCGTAAAACTCTGTTGTTAAAGAAGAACATATCTGAGAGTAACTGTTCAGGTATTGACGGTATTTAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCGGTTTTTTAAGTCTGATGTGAAAGCCTTCGGCTCAACCGAAGAAGTGCATCGGAAACTGGGAAACTTGAGTGCAGAAGAGGACAGTGGAACTCCATGTGTAGCGGTGAAATGCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGGCTGTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGTATGGGTAGCAAACAGGATTAGATACCCTGGTAGTCCATACCGTAAACGATGAATGCTAAGTGTTGGAGGGTTTCCGCCCTTCAGTGCTGCAGCTAACGCATTAAGCATTCCGCCTGGGGAGTACGGCCGCAAGGCTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGT GGAGCATGTGGTTTAATTCGAAGCTACGCGAAGAACCTTACCAGGTCTTGACATACTATGCAAATCTAAGAGATTAGACGTTCCCTTCGGGGACATGGATACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTATTATCAGTTGCCAGCATTAAGTTGGGCACTCTGGTGAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGATGGTACAACGAGTTGCGAACTCGCGAGAGTAAGCTAATCTCTTAAAGCCATTCTCAGTTCGGATTGTAGGCTGCAACTCGCCTACATGAAGTCGGAATCGCTAGTAATCGCGGATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGAGAGTTTGTAACACCCAAAGTCGGTGGGGTAACCTTTTAGAACCAGCCGCCTAAAGTGGGACAGATGATT(SEQ ID No.2)
该分离自浆水的植物乳杆菌(Lactobacillus plantarum)菌株已于2023年05月22日保藏于中国典型培养物保藏中心,菌种保藏号为CCTCC NO:M 2023797。
实施例2菌株益生性能的测定
1.1耐酸性试验
将菌株活化后制成菌体浓度为108CFU/mL的菌悬液,按4%的接种量分别接入pH为1.0、2.0、3.0的MRS液体培养基(同实施例1)中,于0、24h吸取1000μL培养基中的菌悬液,分别涂布至pH为1.0、2.0、3.0的固体培养基上,置于35℃恒温培养箱中倒置培养48h。计算培养基上的菌落数。结果如表2和图5所示。
其中,固体培养基的配方为:葡萄糖20g、蛋白胨10g、牛肉膏10g、酵母浸粉5g、结晶乙酸钠5g、柠檬酸三铵2g、磷酸氢二钾2g、吐温-80 1mL、硫酸镁0.58g、硫酸锰0.25g,琼脂18g,pH值6.4,蒸馏水1000mL,于121℃灭菌20min制得;
表2耐酸性试验结果
由表2可知,菌株TS1在pH3.0的培养基上的菌落数明显高于PH2.0的培养基上的菌落数,且明显高于pH1.0的培养基上的菌落数,具有很好的耐酸性能。
1.2耐胆盐试验
将菌株活化后制成菌体浓度为108CFU/mL的菌悬液,按4%的接种量分别接入胆盐浓度为0.3、1.5、3.0g/L的MRS液体培养基(同实施例1)上,分别于0、24h从中吸取1000μL菌悬液,分别涂布至胆盐浓度为0.3、1.5、3.0g/L的固体培养基上,置于35℃恒温培养箱倒置培养48h。计算菌株对胆盐的耐受力。结果如表3和图6所示。
所述菌株对胆盐的耐受力根据公式(1)进行计算:
胆盐耐受力(%)=(24h活菌数Nt/0h活菌数N0)×100%公式(1)
表3耐胆盐试验结果
由表3可知,菌株TS1在胆盐浓度为3.0g/L的培养基上较胆盐浓度为1.5g/L的培养基上的耐受力更强。
1.3耐人工胃液试验
将菌株活化后制成菌体浓度为108CFU/mL的菌悬液,按4%的接种量接入人工胃液中,35℃静置培养,分别于0h、2h、4h从中吸取1000μL菌悬液,进行4次梯度稀释,制成菌体浓度为108CFU/mL的菌悬液,涂布至固体培养基上,置于35℃恒温培养箱倒置培养48h。测定菌株在人工胃液中不同培养时间下的耐受力。结果如表4所示。
其中,人工胃液的制备方法为:取浓度为20%盐酸16.4mL,调节其pH值至3.0,加入胃蛋白酶(颗粒),使溶液的终浓度达到0.01g/mL,待胃蛋白酶充分溶解后,用0.22μm规格的微孔膜过滤除菌,即得。
所述菌株对人工胃液的耐受力根据公式(2)进行计算:
胃液耐受力(%)=(4h/2h活菌数Nt/0h活菌数N0)×100%公式(2)表4耐人工胃液试验结果
由表4可知,菌株在2h时对人工胃液的耐受力为75%,4h时对人工胃液的耐受力为24%,即,菌株TS1对人工胃液具有较好的耐受力。
1.4耐人工肠液试验
将菌株活化后制成菌体浓度为108CFU/mL的菌悬液,按4%的接种量接入人工肠液中,35℃静置培养,分别于0h、2h、4h从中吸取菌悬液,进行4次梯度稀释,制成菌体浓度为108CFU/mL的菌悬液,涂布至固体培养基上,置于35℃恒温培养箱倒置培养48h。测定菌株在人工肠液中不同培养时间下的耐受力。结果如表5所示。
其中,人工肠液的制备方法为:取磷酸二氢钾6.8g,加入500mL的去离子水充分溶解,调节pH值至6.8,然后加入胰蛋白酶(颗粒),使其终浓度为0.01g/mL,待胰蛋白酶充分溶解后,用规格为0.22μm的微孔膜过滤除菌,即得。
所述菌株对人工肠液的耐受力根据公式(3)进行计算:
肠液耐受力(%)=(4h/2h活菌数Nt/0h活菌数N0)×100%公式(3)表5耐人工肠液结果
由表5可知,菌株TS1在2h时对人工肠液的耐受力为70%,在4h时对人工肠液的耐受力为15%。即,该菌株TS1对人工肠液具有较好的耐受力。
实施例3药敏试验
采用K-B(药敏纸片琼脂扩散法)对菌株TS1进行药敏试验来评价菌株TS1的抗生素耐药性。具体方法如下:
将标准药敏纸片贴至含有菌液的固体培养基上,置于35℃恒温培养箱中培养24h。通过参照《谭瑶,赵清,舒为群等.K-B纸片扩散法药敏试验[J].检验医学与临床,2010,7(20):2290-2291.》中记载的方法判断菌株TS1对不同抗生素的耐药能力。其耐药性能的结果如表6和图9所示。(其中:R表示耐药;I表示中等敏感;S表示敏感)。
表6药敏试验结果
序号 | 药敏试纸名称 | 药敏圈直径(cm) | 敏感性 |
1 | 万古霉素 | - | R |
2 | 多黏菌素 | - | R |
3 | 美满霉素 | 1.2 | I |
4 | 丁胺卡那霉素 | 2 | I |
5 | 先锋霉素VI | 3 | I |
6 | 强力霉素 | 2 | I |
7 | 新霉素 | 2 | S |
8 | 庆大霉素 | 2 | I |
9 | 红霉素 | 3.5 | S |
10 | 青霉素 | 2.3 | I |
11 | 四环素 | 2.7 | S |
12 | 头孢唑林 | 3 | S |
13 | 头孢哌酮 | 3 | S |
14 | 头孢呋辛 | 3.9 | S |
15 | 哌拉西林 | 4.8 | S |
16 | 羧苄西林 | 3.9 | S |
17 | 诺氟沙星 | - | R |
18 | 复方新诺酮 | 2.9 | S |
19 | 奥复星 | 1.4 | R |
20 | 痢特灵 | 2.5 | I |
由表6和表7可知,菌株TS1对四环素、头孢唑林、哌拉西林、红霉素等表现为敏感,对庆大霉素、青霉素、美满霉素等表现为中等敏感。即,该菌株具有较好的药物敏感性,安全性高。
实施例4抑菌试验
抑菌试验采取牛津杯法进行,加入菌液时,分别在已含有指示菌金黄色葡萄球菌、枯草芽孢杆菌、芽孢杆菌、铜绿假单胞菌、大肠杆菌的5个MRS琼脂固体培养基上放5个牛津杯,注入0.1mL菌体浓度为108CFU/mL菌液,置于35℃恒温培养箱中培养24h,重复3次,测定结果,并求出其平均值。测定结果如表7所示。
测定结果参照张素辉等在《仔猪粪便中乳酸菌的分离、鉴定及其抑菌特性的研究》一文中记载的标准(张素辉,曹国文,徐登峰,等.仔猪粪便中乳酸菌的分离、鉴定及其抑菌特性的研究[J].饲料工业,2007(14):31-33.)进行判定,具体为:抑菌圈直径<4mm为不敏感,5~10mm为低度敏感,11~15mm为中度敏感,15mm以上为高度敏感。
表7抑菌试验结果
由表7可知,菌株TS1对金黄色葡萄球菌、芽孢杆菌、铜绿假单胞菌的抑制作用均达到“高敏感”(抑菌圈直径>15.00mm),对大肠杆菌、枯草芽孢杆菌的抑制作用为“中度敏感”(11.00mm<抑菌圈直径<15.00mm)。可见,培养过程中产生的有机酸如乳酸等类物质可使菌液呈现酸性,对指示菌的生长产生酸抑制,同时产生的一类具有抑菌效果的苯乳酸能够与有机酸发挥协同作用,产生更强的抑菌作用。
结论:本发明提供的菌株具较好的耐酸、耐胆盐及耐胃液和肠液能力,进入动物肠道后,有较多的活菌存在,菌株作为益生菌,还可以产生苯乳酸,在改善动物肠道健康的同时,能够保证自身的存活,进一步确保了菌株能够在肠道内发挥作用。本发明提供的植物乳杆菌的上述特性使其能够应用于制备发酵食品和动物饲料中。
以上所述仅是本发明的优选实施方式,应当指出,对于本技术领域的普通技术人员来说,在不脱离本发明原理的前提下,还可以做出若干改进和润饰,这些改进和润饰也应视为本发明的保护范围。
Claims (10)
1.一种植物乳杆菌,其特征在于,所述植物乳杆菌为植物乳杆菌Lactobacillusplantarum TS1,于2023年5月22日保藏于中国典型培养物保藏中心,保藏编号为CCTCC NO:M 2023797。
2.如权利要求1所述的植物乳杆菌的培养方法,其特征在于,将植物乳杆菌Lactobacillusplantarum TS1接种于培养基上进行培养。
3.根据权利要求2所述的培养方法,其特征在于,所述培养的温度为30~40℃,所述培养的时间为20~28h。
4.如权利要求1所述的植物乳杆菌在制备改善肠道菌群的药物中的应用。
5.如权利要求1所述的植物乳杆菌在制备改善肠道菌群的食品或保健品中的应用。
6.如权利要求1所述的植物乳杆菌在制备发酵食品中的应用。
7.如权利要求1所述的植物乳杆菌在制备抗生素替代品中的应用。
8.如权利要求1所述的植物乳杆菌在制备动物饲料中的应用。
9.根据权利要求8所述的应用,其特征在于,所述动物饲料包括青贮饲料和饲料添加剂。
10.如权利要求1所述的植物乳杆菌在合成苯乳酸中的应用。
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