CN112391304B - 副血链球菌f278及其用途 - Google Patents
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Abstract
本发明提供一种副血链球菌F278，其保藏号为CGMCC No.17578。副血链球菌F278为发明人采集的人母乳灌胃给在无菌包中生长的8周龄无菌小鼠，隔天用同样的母乳给每只小鼠进行二次灌胃。灌胃结束后继续在无菌包中饲养小鼠8周。用第8周的小鼠粪便进行细菌分离获得。副血链球菌F278，可以作为益生菌制剂，食品添加剂，或者药物的有效成份，尤其是针对婴幼儿母乳、母乳替代物、婴儿配方奶、乳制品或者相关产品中缺乏链球菌的情况。
Description
技术领域
本发明涉及一种生物工程菌，特别是涉及一种副血链球菌F278及其用途。
背景技术
人母乳不仅为新生儿提供营养，还提供人体共生细菌，已有研究表明健康母亲的乳汁中含有102–105CFU/毫升的细菌。按照每个纯母乳喂养的婴儿平均每天进食800毫升母乳计算，母乳每天给他们提供104–108个细菌细胞。母乳中的细菌在出生后立刻定植在新生儿肠道中，是新生儿肠道的“先锋菌群”，在新生儿的免疫系统发育中起到关键作用。
已有的研究主要集中在从母乳中分离双歧杆菌(Bifidobacterium)和乳杆菌(Lactobacillus)，并研究这些传统益生菌的免疫调节作用。但是，并不是所有母乳中都含有活的、可以分离到培养物的双歧杆菌和乳酸杆菌，对更、大规模人群的母乳菌群结构的研究表明，母乳中丰度最高、而且在不同人的母乳中都能检测到的细菌是链球菌(Streptococcus)。Jost等人采集了7个母亲的新鲜乳汁，只从其中2个母亲的乳汁中分离得到了双歧杆菌，只从1个母亲的乳汁中分离得到了乳酸杆菌，但从7个母亲的乳汁中都分离得到了链球菌；而且，文献表明在从人母乳中分离得到的细菌中，双歧杆菌只占到1.7％，而链球菌占到17.9％。此外，链球菌也是出生后2周内新生儿肠道中的优势细菌。这些结果说明，母乳链球菌是多数母亲通过乳汁传递给后代的人体共生细菌，具有重要的益生作用。因此，提供人母乳中链球菌的益生菌制剂十分有必要。
发明内容
鉴于以上所述现有技术的缺点，本发明的目的在于提供一种副血链球菌F278及其用途，用于解决现有技术中部分母乳、母乳替代物、婴儿配方奶、乳制品或者相关产品中缺乏链球菌的问题。
为实现上述目的及其他相关目的，本发明提供一种副血链球菌F278，其保藏号为CGMCCNo.17578。
本发明中副血链球菌F278为发明人采集的人母乳灌胃给在无菌包中生长的8周龄无菌小鼠，隔天用同样的母乳给每只小鼠进行二次灌胃。灌胃结束后继续在无菌包中饲养小鼠8周。用第8周的小鼠粪便进行细菌分离获得。
经16S rRNA基因扩增序列发育树分析，确定该菌株为副血链球菌，按照国际命名规则：属名+种名+株名对该菌株进行命名，属名、种名、株名分别为链球菌属、副血链球菌种和F278，命名为副血链球菌F278，该菌株于2019年4月28送至中国微生物保藏管理委员会普通微生物中心保藏，保藏号为：CGMCC No.17578。
进一步地，所述副血链球菌F278的核苷酸序列如SEQ ID NO.5所示。
本发明的另一方面提供了上述副血链球菌F278在制备激活哺乳动物免疫功能和/或提高哺乳动物免疫力的药品中的用途。
进一步地，所述药品可以专门针对婴幼儿，例如可以是益生菌制剂。
进一步地，上述副血链球菌F278可以是活的，也可以杀死后的。
进一步地，所述药品中含有副血链球菌F278或副血链球菌F278裂解物。
进一步地，所述药品中副血链球菌F278的含量为102CFU/ml-1010CFU/ml。
进一步地，所述药品具有促进哺乳动物发育、激活哺乳动物免疫功能和/或提高哺乳动物免疫力的作用。
进一步地，所述药品具有促进哺乳动物外周血单核细胞分泌白细胞介素(IL)-12和白细胞介素(IL)-10的作用。
如上所述，本发明的副血链球菌F278及其用途，具有以下有益效果：
采用本申请中的副血链球菌F278，可以作为益生菌制剂或者药物的有效成份，尤其是针对婴幼儿母乳、母乳替代物、婴儿配方奶、乳制品或者相关产品中缺乏链球菌的情况。副血链球菌F278安全有效。实验已经证实其能够有效的促进人外周血单核细胞分泌白细胞介素(IL)-12和白细胞介素(IL)-10，也可以促进线虫的免疫基因的表达并延长线虫寿命。
本发明菌株保藏信息如下：
菌株名称：副血链球菌F278
保藏号为：CGMCC No.17578；
保藏日期：2019年4月28日；
保藏单位名称：中国微生物保藏管理委员会普通微生物中心；
保藏单位简称：CGMCC；
保藏单位地址：北京市朝阳区北辰西路1号院3号。
附图说明
图1显示为副血链球菌F278 ERIC图谱。
图2显示为基于16S rRNA基因全长序列建立的副血链球菌F278的进化树。
图3显示副血链球菌F278菌体细胞刺激人外周血单核细胞(PBMC)分泌白细胞介素(IL)-12和白细胞介素(IL)-10的量，且副血链球菌F278菌体细胞的刺激作用强于已有的市售益生菌鼠李糖乳杆菌GG(LGG)。
图4显示为副血链球菌F278菌体细胞显著延长秀丽隐杆线虫(Caenorhabditiselegans)寿命。
图5显示副血链球菌F278菌体细胞显著提高了秀丽隐杆线虫(Caenorhabditiselegans)免疫基因的表达水平。
具体实施方式
以下通过特定的具体实例说明本发明的实施方式，本领域技术人员可由本说明书所揭露的内容轻易地了解本发明的其他优点与功效。本发明还可以通过另外不同的具体实施方式加以实施或应用，本说明书中的各项细节也可以基于不同观点与应用，在没有背离本发明的精神下进行各种修饰或改变。
在进一步描述本发明具体实施方式之前，应理解，本发明的保护范围不局限于下述特定的具体实施方案；还应当理解，本发明实施例中使用的术语是为了描述特定的具体实施方案，而不是为了限制本发明的保护范围；在本发明说明书和权利要求书中，除非文中另外明确指出，单数形式“一个”、“一”和“这个”包括复数形式。
当实施例给出数值范围时，应理解，除非本发明另有说明，每个数值范围的两个端点以及两个端点之间任何一个数值均可选用。除非另外定义，本发明中使用的所有技术和科学术语与本技术领域技术人员通常理解的意义相同。除实施例中使用的具体方法、设备、材料外，根据本技术领域的技术人员对现有技术的掌握及本发明的记载，还可以使用与本发明实施例中所述的方法、设备、材料相似或等同的现有技术的任何方法、设备和材料来实现本发明。
除非另外说明，本发明中所公开的实验方法、检测方法、制备方法均采用本技术领域常规的分子生物学、生物化学、染色质结构和分析、分析化学、细胞培养、重组DNA技术及相关领域的常规技术。这些技术在现有文献中已有完善说明，具体可参见Sambrook等MOLECULAR CLONING：A LABORATORY MANUAL，Second edition，Cold Spring HarborLaboratory Press，1989and Third edition，2001；Ausubel等，CURRENT PROTOCOLS INMOLECULAR BIOLOGY，John Wiley&Sons，New York，1987and periodic updates；theseries METHODS IN ENZYMOLOGY，Academic Press，San Diego；Wolffe，CHROMATINSTRUCTURE AND FUNCTION，Third edition，Academic Press，San Diego，1998；METHODS INENZYMOLOGY，Vol.304，Chromatin(P.M.Wassarman and A.P.Wolffe，eds.)，AcademicPress，San Diego，1999；和METHODS IN MOLECULAR BIOLOGY，Vol.119，ChromatinProtocols(P.B.Becker，ed.)Humana Press，Totowa，1999等。
材料及其来源：
无菌小鼠：上海斯莱克实验动物中心，C57BL/6J小鼠
Wilkins-Chalgren(WCH)培养基：青岛，海博 货号：HB0261
M17培养基：青岛，海博 货号：HB0391
NGM培养基需要用以下试剂配制而成：
NaCl,Sigma S7653-1KG
Peptone，Sigma P6713-500G
Agar，Sigma 05038-500G
CaCl2.2H2O,Sigma C3881-500G
MgSO4.7H2O，Sigma M1880-500G
KH2PO4,Sigma P0662-500G
K2HPO4.3H2O，Sigma,P9666-500G
Cholesterol，Sigma C8667-1G
NGM培养基配置方法：称取3g NaCl、2.5g Peptone、20g Agar，加单蒸水975mL溶解，121℃高压蒸汽灭菌20min，冷却至65℃左右。在无菌条件下，加入以下无菌溶液：1mL 1MCaCl2，1mL 1M MgSO4，25mL 1M磷酸钾缓冲液(PH 6.0)，1mL 5mg/mL Cholesterol。充分混匀后，使用无菌吸管分装至6厘米培养皿中，每块平皿15mL，室温静置凝固即制成NGM琼脂平板。
线虫：美国线虫保藏中心CGC(Caenorhabditis Genetics Centre,University ofMinnesota，Minneapolis)
大肠杆菌OP50：美国线虫保藏中心CGC(Caenorhabditis Genetics Centre,University of Minnesota,Minneapolis)
实施例1副血链球菌F278的分离
将100微升的新鲜采集的人母乳灌胃给在无菌包中生长的8周龄无菌小鼠，隔天用同样的母乳给每只小鼠进行二次灌胃。灌胃结束后继续在无菌包中饲养小鼠8周。用第8周的小鼠粪便进行细菌分离。
向小鼠粪便立即加入1mL的无菌的0.01mol/L磷酸盐缓冲液(PBS，添加0.1％ L-Cystei ne)，充分振荡使粪便样品成均匀浑浊。将粪悬液做10-2、10-3、10-4、10-5、10-6、10-7、10-8、10-9稀释梯度，将每个梯度的稀释液各100μL分别涂布于Wilkins-Chalgren(WCH)和M17固体培养基平板(1.2％琼脂)上，每个稀释梯度重复三块平板。将所有平板倒置于厌氧培养箱中，37℃培养。培养48小时后，选出菌落数量较为适中、出现较多可分开的单菌落的平板。根据不同的菌落形态和大小，在每种培养基的每个平板上随机挑取菌落，分别转接到相应新的WCH和M17固体培养基平板上，并编号。将每个单菌落进行三次划线纯化。把纯化的单菌落挑入液体M17培养基中，进行富集培养24小时，将获得的培养物进行保存和后续的鉴定。
实施例2副血链球菌F278的鉴定
(1)基因组DNA提取：厌氧液体培养24小时后，取3mL菌液，9000g离心5分钟收集菌体，加入475μL TE缓冲液(10mM Tris-HCl,1mM EDTA,pH 8.0)和25μL溶菌酶(lysozyme，50mg/mL)，37℃摇床震荡孵育1h之后，加入5μL(20μg/mL)蛋白酶K和50μL(20％)SDS，震荡混匀后55℃静置孵育30min。加入等体积约550μL酚氯仿异戊醇(体积比25：24：1)，震荡混匀，4℃静置，14000rpm离心15min，取上清，再用氯仿异戊醇(体积比24：1)重复抽提1-2次。上清中加入两倍体积(约800μL)的预先放在-20℃的无水乙醇和80μL(3M)醋酸钠，放入-20℃冰箱静置2小时沉淀DNA，14000rpm离心15min收集DNA。低温真空干燥后用50μLTE buffer(Tris-HCl，pH 8，10mM)溶解。加入20μLRNase(20mg/mL)，轻轻混匀，37℃温浴30min进行RNA消化。结合PicoGreen荧光染料(Thermo Fisher Scientific,Sunnyvale,USA)，利用酶标仪SpectraMax M5(Molecular Devices,San Francisco,USA)定量DNA浓度。
(2)ERIC-PCR(Enterobacterial repetitive intergenic consensus sequence-PCR)指纹图谱分析：ERIC-PCR的扩增引物上游ERIC1(SEQ ID NO.1：5′-atgtaagctcctggggattcac-3′)，下游为ERIC2(SEQ ID NO.2：5′-aagtaagtgactggggtgagcg-3′)。25μLPCR扩增体系中含有20ng细菌基因组DNA，四种脱氧核苷酸(dNTP)的浓度各200Mm，2.5U of TaKaRa rTaq DNA聚合酶(Takara,Dalian,China),1×PCR buffer(Mg2+free),2mM MgCl2,引物各10pM。PCR程序为：95℃预变性7min；95℃变性30s，52℃退火1min,65℃延伸8min，循环30次；最后65℃延伸16min。取400ng ERIC-PCR产物进行1.5％(w/v)的琼脂糖电泳，使用UVI凝胶呈像系统(Tanon 3500,TanonScience&Technology Co.,Ltd.,China)拍照，得到指纹图谱(图1)。
该ERIC图谱是该菌株特定基因组的体现，可作为副血链球菌F278菌株的特征图谱。
(3)副血链球菌F278菌株16SrRNA基因全长序列的PCR扩增、克隆、测序和进化地位分析：
菌株16S rRNA基因全长序列的PCR扩增使用的引物为27f(SEQ ID NO.3：5'-agagtttgatcctggctcag-3')和1492r(SEQ ID NO.4：5'-cggcttaccttgttacgactt-3')。25μL体系中包括0.75U rTaq DNA聚合酶(Takara,Dalian,China)，1×PCR buffer(Mg2+free)，2mM MgCl2，每条引物各10pmol，四种脱氧核苷酸各200μM，细菌基因组DNA 10ng作为模板。扩增程序如下：95℃预变性7min；94℃变性30s，52℃退火1min，65℃延伸8min，如此循环25次，最后在65℃延伸16min。
根据Gel Extraction Kit 200(Omega，USA)的使用说明书对PCR产物进行纯化。根据连接试剂盒说明书，将纯化后的PCR产物与载体pGEM-T Easy Vector(Promega，Madison，USA)进行连接，连接产物用转化法转入到宿主菌E.coli DH5α感受态细胞(Transgen，Beijing，China)中。然后均匀涂布于预先添加了一定浓度的IPTG和X-Gal的LB-氨苄青霉素(100μg/mL)平板上，37℃培养12小时，随机挑选白斑阳性克隆，进行测序(LifeTechnologies，Shanghai，China)。副血链球菌F278 CGMCC No.17578菌株的16S rRNA基因的全长序列如SEQ ID NO.5所示：
gatgaacgctggcggcgtgcctaatacatgcaagtagaacgctgaagcttggtgcttgcaccgagcggatgagttgcgaacgggtgagtaacgcgtaggtaacctgcctcttagcgggggataactattggaaacgatagctaataccgcataaaagtcgacattgcatgatgttgacttgaaaggtgcgattgcatcactaagagatggacctgcgttgtattagctagttggtgaggtaacggctcaccaaggcaacgatacatagccgacctgagagggtgatcggccacactgggactgagacacggcccagactcctacgggaggcagcagtagggaatcttcggcaatgggggcaaccctgaccgagcaacgccgcgtgagtgaagaaggttttcggatcgtaaagctctgttgtaagagaagaacgagtgtgggagtggaaagttcacactgtgacggtaacttaccagaaagggacggctaactacgtgccagcagccgcggtaatacgtaggtcccgagcgttatccggatttattgggcgtaaagcgagcgcaggcggttagataagtctgaagttaaaggctgtggcttaaccatagtacgctttggaaactgtttaacttgagtgcaagaggggagagtggaattccatgtgtagcggtgaaatgcgtagatatatggaggaacaccggtggcgaaagcggctctctggcttgtaactgacgctgaggctcgaaagcgtggggagcaaacaggactagataccctggtagtccacgccgtaaacgatgagtgctaggtgttgggtcctttccgggactcagtgccgcagctaacgcattaagcactccgcctggggagtacgaccgcaaggttgaaactcaaaggaattgacgggggcccgcacaagcggtggagcatgtggtttaattcgaagcaacgcgaagaaccttaccaggtcttgacatccctctgaccgctctagagatagagttttccttcgggacagaggtgacaggtggtgcatggttgtcgtcagctcgtgtcgtgagatgttgggttaagtcccgcaacgagcgcaacccctattgttagttgccatcattgagttgggcactctagcgagactgccggtaataaaccggaggaaggtggggatgacgtcaaatcatcatgccccttatgacctgggctacacacgtgctacaatggctggtacaacgagtcgcgagtcggtgacggcaagctaatctcttaaagccagtctcagttcggattgtaggctgcaactcgcctacatgaagtcggaatcgctagtaatcgcggatcagcacgccgcggtgaatacgttcccgggccttgtacacaccgcccgtcacaccacgagagtttgtaacacccgaagtcggtgaggtaaccgtaaggagccagccgcctaaggtgggatagatgattggggtg
将获得16S rRNA基因序列在Genbank数据库中进行BLAST(http://www.ncbi.nlm.nih.gov/Blast)比对，与数据库的已知细菌最相似的序列的细菌为：Streptococcus parasanguinis ATCC 15912，相似性为98.98％。使用MEGA 5软件(Molecular Evolutionary Genetics Analysis package)构建系统进化树(Neighbor-joining phylogenetic tree)，显示F278菌株为副血链球菌(图2)。
实施例3热杀死的副血链球菌F278菌体细胞促进人外周血单核细胞(humanperipheral blood mononuclear cells，PBMCs)分泌白细胞介素12(IL-12)和白介素10(IL-10)
将副血链球菌F278和鼠李糖乳杆菌LGG分别在M17(中国，青岛，海博)和MRS(中国，青岛，海博)液体培养基中培养8小时，达到平台期。将培养物5,000g离心10min，收集副血链球菌F278和鼠李糖乳杆菌LGG菌体细胞。将菌体细胞用PBS重悬之后5,000g离心10min，如此洗涤2次，去除细菌培养基。用PBS将菌体细胞浓度调整为108CFU/ml和109CFU/ml，65℃水浴20min，对菌体细胞热致死，冻存到-80℃备用。
在24孔板中，每孔接种2×106个人外周血单核细胞(human peripheral bloodmononuclear cells，PBMC)后，加入20μl的1×108CFU/ml或者1×109CFU/ml的热杀死的副血链球菌F278和鼠李糖乳杆菌LGG菌体，最终体积为1ml，构建了细胞与细菌比例分别为1:1和1：10的共孵育体系。阴性对照组组中不加入细菌细胞，而加入20μl的PBS。。所用细胞培养基为含10％胎牛血清和1％链霉素和氨苄青霉素的1640细胞培养基。放置到37℃，5％二氧化碳培养箱中共孵育24h。收集细胞培养上清，用ELISA试剂盒测试其中白介素10和白介素12的浓度。
结果如图3显示，副血链球菌F278的菌体细胞能刺激人外周血单核细胞(PBMC)分泌较多的白细胞介素(IL)-12和白细胞介素(IL)-10(图3)，而鼠李糖乳杆菌LGG只能促进少量的IL-12和IL-10的分泌。
实施例4副血链球菌F278的菌体细胞提高线虫的免疫基因的表达水平并延长线虫的寿命
(1)副血链球菌F278的菌体细胞延长线虫的寿命
分别用M17培养基(中国，青岛，海博)和MRS(中国，青岛，海博)培养基将副血链球菌F278和鼠李糖乳杆菌LGG在37℃厌氧工作站(DG500,DWS,United Kingdom)培养8小时。大肠杆菌OP50在好氧37℃条件下培养过夜。培养后的菌液15000×g 10min离心，去处上清液，收集副血链球菌F278，鼠李糖乳杆菌LGG和大肠杆菌OP50菌体，用无菌M9缓冲液,15000×g条件下，离心10min，将细菌菌体洗两次。洗过的菌体用M9缓冲液调节细菌菌体浓度为10mg/100μl(湿重/体积)，并混匀。在直径为6厘米不添加蛋白胨的NGM(mNGM)平板上滴加100μl含10mg菌体的的细菌重悬液。
在添加有蛋白胨的NGM平板上、以大肠杆菌OP50为食的线虫生长到L3期后，分别转移到mNGM平板上，分别以副血链球菌F278,鼠李糖乳杆菌LGG和大肠杆菌OP50的菌体为食，并放置到25℃条件下培养。副血链球菌F278，鼠李糖乳杆菌LGG和大肠杆菌OP50分别设置5个平行平板(20-25条/板)，共100-125条线虫测试其寿命并加以比较。实验过程中，线虫对铂丝轻微刺激无反应即视为死亡。使用OASIS 2(Online Application for SurvivalAnalysis 2)在线软件进行Kaplan-Meier生存分析，并用log-rank检验饲喂不同细菌菌体的线虫平均寿命的差异。
结果如图4显示，与线虫的标准食物大肠杆菌OP50相比，副血链球菌F278的菌体细胞和鼠李糖乳杆菌LGG细胞均显著延长了线虫的寿命，说明了副血链球菌F278的安全性和益生性。
(2)食用副血链球菌F278的菌体细胞的线虫的RNA的提取
饲喂副血链球菌F278、鼠李糖乳杆菌LGG、或大肠杆菌OP50第14天后，收集500条线虫，利用TRIZOL reagent(Invitrogen)将线虫裂解后提取RNA，利用RNeasy Mini Kit(Qiagen)纯化RNA，并用DNaseI(Invitrogen)试剂盒去除残留的DNA。
(3)用qPCR的方法测定副血链球菌F278的菌体细胞提高线虫免疫基因的表达水平纯化后的线虫RNA利用SuperScriptTM First-Strand Synthesis System for RT-PCR(Invitrogen)逆转录试剂盒，以oligo(dT)为引物合成第一条cDNA链。用SYBR GreenSupermix(BIO-RAD)对线虫的免疫基因进行定量检测(Roche，LightCycler96)。以看家基因act-1为参照基因，用2-ΔΔCt法计算免疫基因的相对表达量(如图5)。
结果显示，与线虫的标准食物大肠杆菌OP50相比，副血链球菌F278的菌体细胞显著提高了线虫免疫基因(cpr-1,cpr-5,lys-5,clec-60,C15C8.3)的表达水平。
以上的实施例是为了说明本发明公开的实施方案，并不能理解为对本发明的限制。此外，本文所列出的各种修改以及发明中方法、组合物的变化，在不脱离本发明的范围和精神的前提下对本领域内的技术人员来说是显而易见的。虽然已结合本发明的多种具体优选实施例对本发明进行了具体的描述，但应当理解，本发明不应仅限于这些具体实施例。事实上，各种如上所述的对本领域内的技术人员来说显而易见的修改来获取发明都应包括在本发明的范围内。
序列表
<110> 上海交通大学
<120> 副血链球菌F278及其用途
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1470
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 1
gatgaacgct ggcggcgtgc ctaatacatg caagtagaac gctgaagctt ggtgcttgca 60
ccgagcggat gagttgcgaa cgggtgagta acgcgtaggt aacctgcctc ttagcggggg 120
ataactattg gaaacgatag ctaataccgc ataaaagtcg acattgcatg atgttgactt 180
gaaaggtgcg attgcatcac taagagatgg acctgcgttg tattagctag ttggtgaggt 240
aacggctcac caaggcaacg atacatagcc gacctgagag ggtgatcggc cacactggga 300
ctgagacacg gcccagactc ctacgggagg cagcagtagg gaatcttcgg caatgggggc 360
aaccctgacc gagcaacgcc gcgtgagtga agaaggtttt cggatcgtaa agctctgttg 420
taagagaaga acgagtgtgg gagtggaaag ttcacactgt gacggtaact taccagaaag 480
ggacggctaa ctacgtgcca gcagccgcgg taatacgtag gtcccgagcg ttatccggat 540
ttattgggcg taaagcgagc gcaggcggtt agataagtct gaagttaaag gctgtggctt 600
aaccatagta cgctttggaa actgtttaac ttgagtgcaa gaggggagag tggaattcca 660
tgtgtagcgg tgaaatgcgt agatatatgg aggaacaccg gtggcgaaag cggctctctg 720
gcttgtaact gacgctgagg ctcgaaagcg tggggagcaa acaggactag ataccctggt 780
agtccacgcc gtaaacgatg agtgctaggt gttgggtcct ttccgggact cagtgccgca 840
gctaacgcat taagcactcc gcctggggag tacgaccgca aggttgaaac tcaaaggaat 900
tgacgggggc ccgcacaagc ggtggagcat gtggtttaat tcgaagcaac gcgaagaacc 960
ttaccaggtc ttgacatccc tctgaccgct ctagagatag agttttcctt cgggacagag 1020
gtgacaggtg gtgcatggtt gtcgtcagct cgtgtcgtga gatgttgggt taagtcccgc 1080
aacgagcgca acccctattg ttagttgcca tcattgagtt gggcactcta gcgagactgc 1140
cggtaataaa ccggaggaag gtggggatga cgtcaaatca tcatgcccct tatgacctgg 1200
gctacacacg tgctacaatg gctggtacaa cgagtcgcga gtcggtgacg gcaagctaat 1260
ctcttaaagc cagtctcagt tcggattgta ggctgcaact cgcctacatg aagtcggaat 1320
cgctagtaat cgcggatcag cacgccgcgg tgaatacgtt cccgggcctt gtacacaccg 1380
cccgtcacac cacgagagtt tgtaacaccc gaagtcggtg aggtaaccgt aaggagccag 1440
ccgcctaagg tgggatagat gattggggtg 1470
Claims (4)
1.副血链球菌F278在制备药品中的用途，所述副血链球菌F278的保藏号为CGMCCNo.17578，所述药品具有促进动物外周血单核细胞分泌白细胞介素(IL)-12和白细胞介素(IL)-10的作用。
2.根据权利要求1所述的用途，其特征在于：所述副血链球菌F278是杀死的。
3.根据权利要求1所述的用途，其特征在于：所述药品中副血链球菌F278的含量为102CFU/ml-1010CFU/ml。
4.根据权利要求1所述的用途，其特征在于：所述药品为益生菌制剂。
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