CN112126605A - 一种唾液乳杆菌及其应用 - Google Patents
一种唾液乳杆菌及其应用 Download PDFInfo
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- CN112126605A CN112126605A CN202011082982.5A CN202011082982A CN112126605A CN 112126605 A CN112126605 A CN 112126605A CN 202011082982 A CN202011082982 A CN 202011082982A CN 112126605 A CN112126605 A CN 112126605A
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- ger106
- lactobacillus
- lactobacillus salivarius
- salivariuse
- salivarius
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Abstract
本发明提供了唾液乳杆菌(Lactobacillus salivariuse)GER106,保藏在中国典型培养物保藏中心,保藏编号为CCTCC NO:M2020445,保藏地址为中华人民共和国湖北省武汉市武汉大学,保藏日期为2020年09月08日。还提供了该菌种的应用。本发明提供的唾液乳杆菌GER106来源于儿童口腔,可用于制备食品、发酵剂、功能食品或药物组合物,其具有提高免疫力、预防龋齿的作用。
Description
技术领域
本发明属于微生物领域,特别涉及一种唾液乳杆菌,还涉及该菌株的应用。
背景技术
免疫力低下导致人的免疫系统不能正常发挥保护作用,在此情况下,极易招致细菌、病毒、真菌等感染,因此免疫力低下最直接的表现就是容易生病。因经常患病,加重了机体的消耗,通常出现体虚感冒、气虚感冒、肺虚咳喘、脾虚池泻、气虚心悸、体质虚弱、营养不良、精神萎靡、疲乏无力、食欲降低、睡眠障碍等症状。此类人群生病的恢复期较长,而且常常反复发作,长此以往会导致身体和智力发育不良,还易诱发重大疾病。
龋齿是一种常见病、多发病,其发病率高,分布广,对人体危害大。经过科学家们不懈的努力,现在普遍认为细菌是龋齿发生的主要原因,其中变形链球菌是主要致病菌,其形成的菌斑粘附于牙齿表面,并在牙齿表面繁殖生长、产酸,腐蚀牙齿,最终形成龋齿。目前市场上应用的防龋产品大致分为以下几类:一为含氟制剂,其防龋原理为促进牙齿钙化,减少酸性腐蚀。此为间接作用,疗效有限;二是抗菌药物的应用,如含洗必汰、甲硝唑等的漱口水,因其有明显的副作用,应用受到限制;三是由植物提取物制成的防龋产品,如中药提取物制成的含漱液、口胶片等,其对龋齿防治有一定作用,但其均为药物,需在医生指导下使用,不宜长期使用;四是目前市场上被广泛接受的利用代糖生产的用于防龋护齿类的产品,如木糖醇等,它却没有从根本上解决防龋护齿的实质性问题。
人们对饮食与健康之间关系的认识日渐提高,这已使得他们对在提供基本营养之余还能增强健康的产品需求日益高涨,有研究表明摄入益生菌有益于维持身体微妙的微生物平衡,但是,目前对可预防龋齿的益生菌仍然没有成熟的产品。
发明内容
发明目的:为了解决保证人们的身体健康,本发明提供了一种唾液乳杆菌。
技术方案:本发明所采用的技术方案为:
唾液乳杆菌(Lactobacillus salivariuse)GER106,保藏在中国典型培养物保藏中心,保藏编号为CCTCC NO:M2020445,保藏地址为中华人民共和国湖北省武汉市武汉大学,保藏日期为2020年09月08日。
该菌株革兰氏染色结果显示:菌株在含CaCO3的MRS培养基上菌落直径1-2mm、白色、凸起、有溶钙圈;该菌株革兰氏阳性,圆端直杆状;菌体革兰氏染色特性如图1 所示。菌株GER106的16S rDNA的核苷酸序列如SEQ ID NO.1所示;经过16S rDNA 基因比对,与Genebank中的Lactobacillus salivarius菌株比对相似率达100%;结合微生物系统鉴定,GER106为一株唾液乳杆菌,命名为唾液乳杆菌GER106;唾液乳杆菌 GER106的16S rDNA如序列表所示,生物进化树如图2所示。
本发明还提供了上述唾液乳杆菌(Lactobacillus salivariuse)GER106的规模化发酵方法,在发酵培养基中37℃恒温培养,发酵培养基为:10L纯水,蛋白胨100g,葡萄糖200g,酵母提取物50g,乙酸钠50g,磷酸氢二钾2g,柠檬酸氢二胺20g,硫酸镁5.8g,硫酸锰2.5g,牛肉膏100g,吐温80 10mL,121℃高温灭菌30min,调节pH6.0-6.5。
本发明还提供了唾液乳杆菌(Lactobacillus salivariuse)GER106在制备提高免疫力食品、药物或保健品中的应用。
优选地,唾液乳杆菌(Lactobacillus salivariuse)GER106的剂量为(0.5-3.0)×1010CFU/g。
本发明还提供了唾液乳杆菌(Lactobacillus salivariuse)GER106在制备预防龋齿的药物或保健品中的应用。
优选地,唾液乳杆菌(Lactobacillus salivariuse)GER106的剂量为(1.0-3.5)×1010CFU/g。
本发明还提供了一种药物组合物,所述组合物中包括唾液乳杆菌(Lactobacillussalivariuse)GER106,还包括至少一种药学上可接受的载体、稀释剂、赋形剂或辅助剂。
优选地,所述药物组合物用于提高免疫力和/或预防龋齿;所述唾液乳杆菌(Lactobacillus salivariuse)GER106的剂量为(0.5-3.5)×1010CFU/g。
有益效果:本发明提供的唾液乳杆菌GER106来源于儿童口腔,可用于制备食品、发酵剂、功能食品或药物组合物,其具有提高免疫力、预防龋齿的作用
附图说明
图1为唾液乳杆菌GER106的革兰氏染色光学显微镜照片。
图2为唾液乳杆菌GER106的生物进化树。
图3为唾液乳杆菌GER106发酵过程中OD值变化曲线。
图4为暴露于唾液乳杆菌GER106后,Caco-2/TC7细胞上清液中细胞因子和 b-defensin 2(HBD-2)的测量结果图。其中,(A)IL-6,(B)IL-8,(C)IL-10,(D)HBD-2。平均值±SE。与Caco-2/TC7对照相比,NS(无显著性),*P<0.05和**P<0.01。
图5为在不同培养条件下唾液乳杆菌GER106(109细胞/mL)的热灭活细胞存在下,变形链球菌对羟基磷灰石的相对粘附力测试方法和结果图。(A)测定方案:将羟基磷灰石圆片与唾液乳杆菌一起预孵育3小时;洗涤步骤后,将圆片与(C1)唾液乳杆菌和变形链球菌,(C2)变形链球菌或(C3)KCl缓冲液孵育2小时;再次洗涤最后的测定盘 (C3),并与变形链球菌再温育2小时;最后,进行最后的洗涤,并通过表面电镀法对变形链球菌进行定量。(B)附着力结果。考虑到阴性对照中细菌计数的平均值,将变形链球菌浓度数据标准化,标准偏差用误差线表示。
具体实施方式
下面结合附图对本发明做出进一步说明。
实施例1菌株的分离、纯化及鉴定
该菌种来源于健康儿童口腔。
唾液乳杆菌菌株分离纯化:取样品约0.2g,加入1ml MRS培养基,涡旋震荡1min,依次10倍稀释至10-6,每个稀释浓度取200μL涂布于含1%CaCO3乳酸菌选择性培养基MRS培养基上,37℃厌氧罐中培养48h,挑取平板上单菌落,在MRS固体培养基上划线纯化,37℃厌氧罐中培养48h,得到纯的菌落。
对纯化后菌落进行革兰氏染色鉴定;菌株在含CaCO3的MRS培养基上菌落直径 1-2mm、白色、凸起、有溶钙圈;该菌株革兰氏阳性,圆端直杆状;菌体革兰氏染色特性如图1所示。
唾液乳杆菌菌株筛选鉴定:对获得的菌株GER106采用16S rDNA测序进行鉴定;利用已发表的16S通用引物(引物序列为8F:5’-AGAGTTTGATCCTGGCTCA-3’;1492R: 5’-GGTTACCTTGTTACGACTT-3’)对菌株GER106的16S rDNA基因序列进行扩增和测序,PCR扩增产物送生工生物工程(上海)股份有限公司测序,菌株GER106的16S rDNA的核苷酸序列如SEQID NO.1所示;经过16S rDNA基因比对,与Gene bank中的Lactobacillus salivarius菌株比对相似率达100%;结合微生物系统鉴定,GER106为一株唾液乳杆菌,命名为唾液乳杆菌GER106;唾液乳杆菌GER106的16S rDNA如序列表所示,生物进化树如图2所示。
实施例2唾液乳杆菌(Lactobacillus salivariuse)GER106耐酸性和胆汁性试验
将GER106菌液在37℃的MRS肉汤中生长过夜,并在调至pH 3的MRS肉汤中和 0.5%w/v胆汁中悬浮至大约108CFU/ml的细胞浓度(SigmaeAldrich,法国)4小时。选择这些条件以代表通过胃肠系统所花费的时间以及分别在鸡胃和肠中发现的pH值和胆汁浓度。通过在MRS琼脂平板上计数来评估细菌生存力,结果如表1所示。
表1模拟耐酸性和耐胆汁实验结果
测试标准 | 结果 |
耐酸性 | 是。在pH 3下2h后,95%的存活率 |
耐胆汁性 | 是。0.5%w/v胆汁4小时后,95%的存活率 |
对酸和胆汁的抵抗力通常被认为是益生菌评估的基本评估标准,因为菌株必须在胃和小肠中存活下来。因此,分别在pH 3下测试2h、在0.5%w/v的胆汁中测试4h唾液乳杆菌GER106的存活率。结果表明,该菌株能够耐受酸性条件和存在0.5%w/v胆汁,而没有任何显着的活力损失(95%存活率)。
实施例3唾液乳杆菌(Lactobacillus salivariuse)GER106的规模化发酵及冻干粉生产
规模化发酵培养基配制:按10L纯水(反渗透法),蛋白胨100g,葡萄糖200g,酵母提取物50g,乙酸钠50g,磷酸氢二钾2g,柠檬酸氢二胺20g,硫酸镁5.8g,硫酸锰2.5g,牛肉膏100g,吐温80 10mL,121℃高温高压灭菌20min,调定pH至6.5,加热搅拌均匀。121℃高温高压灭菌20min。
将菌株在固态MRS培养基上划线,37℃厌氧罐中培养24h后,挑选个体独立、凸起的菌斑接入500ml液体培养基中,37℃恒温箱中培养10-12h;OD值≥1.2时,转移至装5L液体培养基的摇瓶中,37℃恒温箱中培养8-12h;OD值≥1.2时,转移至装50L 液体培养基发酵罐中,稳定PH为6.0-6.5,37℃条件发酵8-12h;发酵过程中OD值变化趋势如图3所示;OD值≥1.2时,转入装500L液体培养基发酵罐中,稳定PH为6.0-6.5, 37℃条件发酵10-18h;OD值≥1.2时,结束发酵,将发酵罐温度快速降至10℃以下,对发酵液进行离心收获菌体。
收获的菌泥按1:1加入冻干保护剂(16%脱脂乳,10%糊精,4%聚乙烯吡咯烷酮,6%乳糖,0.5%谷氨酸钠,0.5%半胱氨酸,0.3 0.8%乙酸钠,其余为水),进行真空冷冻干燥32-50h,将冻干的菌饼进行气流粉碎,即获得唾液乳杆菌GER106冻干粉,活菌量在2.0×1011-8×1011CFU/g,用于实验、发酵、食品、保健品或药品等用途。
实施例4唾液乳杆菌(Lactobacillus salivariuse)GER106的粘附肠道细胞的能力
Caco-2/TC7细胞在第35到50代之间使用。细胞常规在DMEM培养基中生长,该培养基中添加了15%热灭活的胎牛血清,2mM的L-谷氨酰胺,青霉素和链霉素各100 U/ml和1%的非必需氨基酸。为了进行实验分析,将细胞以约105个细胞/cm2的密度播种在24孔组织培养板或允许上皮分化的插入物中。在37℃,5%CO2-95%空气气氛中培养细胞,并定期更换培养基。将在24孔组织培养板中生长的Caco-2/TC7孵育至早期汇合(未分化状态),而在插入物上生长的Caco-2/TC7细胞在汇合后21天(完全分化状态)使用。
通过离心收获唾液乳杆菌GER106,将其以108CFU/ml的浓度重悬于不含血清和抗生素的细胞培养基中,然后应用于融合的Caco-2/TC7单层。在37℃,5%CO2下温育4 小时后,用PBS洗涤单层膜以除去非粘附细菌,并通过与0.1%Triton X100温育15分钟进行裂解,然后将裂解物稀释并铺在MRS琼脂上以确定粘附细菌的数量。
对三个干预组粘附细菌数量进行统计,结果如表2所示。可以看出唾液乳杆菌GER106在肠道粘附细胞有明显的效果。
表2干预期间粘附结果
粘附到肠上皮细胞是选择益生菌的另一个可靠标准。据报道,使用Caco-2细胞评估肠上皮细胞的粘附能力的体外测试结果与体内结果具有良好的相关性,并且该特征通常是菌株特异性的。唾液乳杆菌GER106(108CFU/ml)的粘附能力通过将细菌与汇合的 Caco-2/TC7单层孵育4小时来检查。在温育期结束时,通过冲洗除去非粘附细菌,并通过平板测定粘附细菌的数量。在这些条件下,在Caco-2/TC7单层裂解物中回收到105 CFU/ml粘附的唾液乳杆菌GER106,相当于初始种群的1%。
实施例4唾液乳杆菌(Lactobacillus salivariuse)GER106的免疫防御能力
用108CFU/ml唾液乳杆菌GER106处理24小时后,使用ELISA Quantikine试剂盒测量培养上清液中Caco-2/TC7细胞产生的细胞因子(IL-6,IL-8和IL-10)的水平。使用b-defensin 2,beta(Human)e ELISA试剂盒对b-defensin 2进行测量。
使用ELISA测定法对IL-6,IL-8,IL-10和b-defensin 2的测量见图4,唾液乳杆菌GER106诱导IL-8分泌增加1.8倍(图4中B图),b-defensin 2分泌增加2.6倍。与未处理的Caco-2/TC7细胞相比,b-defensin 2的分泌(图4中D图)。相反,对于IL-6(图 4中A图)和IL-10(图4中C图),暴露于细菌后,未观察到Caco-2/TC7细胞的基础分泌发生变化。
结果表明:唾液乳杆菌GER106通过诱导IL-8表现出体外免疫调节活性。我们还观察到唾液乳杆菌GER106诱导了b-defensin 2的分泌。已知诱导型bdefensins在肠屏障功能中起着重要作用,并且体外研究表明,临床上有效的益生菌可诱导产生抗菌性 b-defensin2。益生菌(包括唾液乳杆菌GER106)诱导b-defensin 2可能是增强先天防御机制的替代方法和补充新策略。
实施例5唾液乳杆菌(Lactobacillus salivariuse)GER106对引起龋齿的变形链球菌的抑制
唾液乳杆菌样品在KCl缓冲液中标准化为1010细胞/mL。通过离心浓缩变形链球菌细胞,用无菌KCl缓冲液洗涤3次,然后重悬于适当体积的KCl缓冲液中,以便获得等于3的OD600nm读数。两种细菌悬浮液均在室温下保存少于1小时或在4℃下最多持续6小时。
致密的羟基磷灰石圆片(5mm x 2mm)的处理如下:在水洗涤三个周期后,将圆片在37℃的溶菌酶溶液(20mg/mL)中孵育1小时,并再次在37℃下进行在胰蛋白酶溶液(20mg/mL)中孵育1h。用水彻底洗涤除去酶,并用0.2M NaOH处理圆片1h。
用水洗涤3个循环后,将圆片进行紫外线消毒(40分钟),然后将其在4℃的过滤器消毒KCl中保存。在平底96孔微量滴定板(每孔1个圆片)中进行粘附测定,在其中加入唾液乳杆菌样品(109个细胞/mL)的0.2mL KCl缓冲液。
在37℃的预温育期3小时后,随着轨道摇动(100rpm),用KCl缓冲液洗涤孔。然后,将变形链球菌(OD 600nm=0.3)加入到装有孔的圆片中,并在定轨摇动(100rpm) 下于37℃共同孵育2小时。最后,将圆片转移到1.5mL试管中,在其中用1mL KCl 缓冲液洗涤5次。变形链球菌定量后,结果表示为相对(%)附着力。阴性对照的平均变形链球菌浓度(粘附力为100%,无唾液乳杆菌GER106),使用以下公式对数据进行标准化:变形链球菌浓度×100/变形链球菌的平均值阴性对照中的浓度。
为了消除变形链球菌粘附的抑制是一种非特异性现象,由于在共孵育过程中对羟基磷灰石表面的暂时竞争,因此重复进行了分析,而没有将微生物与羟基磷灰石圆盘同时孵育,也没有进行洗涤唾液乳杆菌和变形链球菌与椎间盘接触的台阶(图5中A图)。在这些条件下,唾液乳杆菌CECT 5713抑制变形链球菌粘附的能力从洗涤后的70%降低到30%,但仍然可以检测到效果。羟基磷灰石与唾液乳杆菌细胞的预温育也确实在另外两个洗涤步骤后也抑制了病原体的粘附(图5中B图)。
实施例6唾液乳杆菌(Lactobacillus salivariuse)GER106应用实例
1、利用唾液乳杆菌GER106制备乳制品、豆制品及果蔬制品的发酵剂
将唾液乳杆菌GER106在发酵罐发酵成熟后,离心获得菌泥后,加入保护剂进行乳化,将乳化后的悬浮液在温度37℃下静止30min,采用真空冷冻干燥法进行冻干,即得到乳制品、豆制品及果蔬制品的发酵剂。
2、利用唾液乳杆菌GER106制作含益生菌食品
将唾液乳杆菌GER106发酵液或冻干粉加入常温或低温条件下食用的固体饮料、茶饮料、蔬果汁、压片糖果等食品中,即可制成含有益生菌的食品。
3、利用唾液乳杆菌GER106提高人体免疫能力的益生菌饮料
将唾液乳杆菌GER106冻干粉与菊粉、抗性糊精等益生元进行混合,分装成2g-10g每袋的便装,每袋含唾液乳杆菌GER106量在50亿CFU-300亿CFU,即制作成有助于提高人体免疫能力的益生菌饮料食品,常温或冷藏保存,37℃以下温水冲服。
4、利用唾液乳杆菌GER106制作预防牙齿龋齿的益生菌饮料
将唾液乳杆菌GER106冻干粉与低聚果糖、膳食纤维等益生元进行混合,分装成2g-10g每袋的便装,每袋含唾液乳杆菌GER106量在100亿CFU-350亿CFU,即制作成有助于预防牙齿龋齿的益生菌饮料食品,常温或冷藏保存,37℃以下温水冲服,倒入口中嚼食最佳。
5、利用唾液乳杆菌GER106制作乳酸菌饮料
将脱脂乳粉与水按1:15混合后充分溶解,在95℃灭菌20min,然后降温至35℃,按10:1加入唾液乳杆菌GER106菌液进行发酵,实时监测PH值,当PH值下降速率最大时,温度降至4摄氏度进行冷藏或灌装,即得到含有唾液乳杆菌GER106活菌的乳酸菌饮料。
6、利用唾液乳杆菌GER106制作胶囊制品
将唾液乳杆菌GER106冻干粉装入药用微胶囊,即可得到用于干预肠道菌群、人体免疫力等作用的胶囊制品。
7、利用唾液乳杆菌GER106制作发酵乳
将鲜奶加入蔗糖溶解后进行低温灭菌(也可以是市场上销售的已灭菌鲜奶),在恒温容器中预热至37℃,唾液乳杆菌GER106、保加利亚乳杆菌和植物乳杆菌进行等比例混合作为发酵剂,加入预热好的鲜奶中,按1L鲜奶加入活菌量在1×108CFU-5×109CFU,混匀,在35-37℃条件下放置4-8h,发生凝乳后放置4℃冷藏12h,即得到发酵乳,食用是可根据个人口味加入蜂蜜等调整口味。此外,鲜奶发酵发生凝乳后,按1:10的比例加入鲜奶中,作为鲜奶发酵剂,以同样的方式进行发酵。
序列表
<110> 镇江市天益生物科技有限公司
<120> 一种唾液乳杆菌及其应用
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1050
<212> DNA
<213> 唾液乳杆菌(Lactobacillus salivarius)
<400> 1
cgaacccata tctgtcacct tagacggctg gctccttgcg gttaccccac cggctttggg 60
tgttacaaac tctcatggtg tgacgggcgg tgtgtacaag gcccgggaac gtattcaccg 120
cgacatgctg attcgcgatt actagcgatt ccgacttcat gtaggcgagt tgcagcctac 180
aatccgaact gagaacggct ttaagagatt agctaaacct cgcggtcttg cgactcgttg 240
taccgtccat tgtagcacgt gtgtagccca ggtcataagg ggcatgatga cttgacgtcg 300
tccccacctt cctccggttt gtcaccggca gtctcgccag agtgcccaac ttaatgctgg 360
caactgacaa caagggttgc gctcgttgcg ggacttaacc caacatctca cgacacgagc 420
tgacgacagc catgcaccac ctgtcacttt gtccccgaag ggaaagccta atctcttagg 480
tggtcaaagg atgtcaagac ctggtaaggt tcttcgcgtt gcttcgaatt aaaccacatg 540
ctccaccgct tgtgcgggcc cccgtcaatt cctttgagtt tcaaccttgc ggtcgtactc 600
cccaggcgga atgcttattg cgttagctgc ggcactgaag ggcggaaacc ctccaacacc 660
tagcattcat cgtttacggc gtggactacc agggtatcta atcctgtttg ctacccacgc 720
tttcgaacct cagcgtcagt tacagaccag agagccgctt tcgccactgg tgttcttcca 780
tatatctacg catttcaccg ctacacatgg agttccactc tcctcttctg cactcaagtc 840
ttccagtttc caatgcacta ctccggttaa gccgaaggct ttcacatcag acttaaaaga 900
ccgcctgcgt tccctttacg cccaataaat ccggacaacg cttgccacct acgtattacc 960
gcggctgctg gcacgtagtt agccgtgact tgctgggtta gataccgtca tcgaatgaac 1020
agttactctc actcgtgttc ttctctaaca 1050
Claims (8)
1.唾液乳杆菌(Lactobacillus salivariuse)GER106,保藏在中国典型培养物保藏中心,保藏编号为CCTCC NO:M2020445,保藏地址为中华人民共和国湖北省武汉市武汉大学,保藏日期为2020年09月08日。
2.权利要求1所述的唾液乳杆菌(Lactobacillus salivariuse)GER106的规模化发酵方法,其特征在于:在发酵培养基中37℃恒温培养,发酵培养基为:10L纯水,蛋白胨100g,葡萄糖200g,酵母提取物50g,乙酸钠50g,磷酸氢二钾2g,柠檬酸氢二胺20g,硫酸镁5.8g,硫酸锰2.5g,牛肉膏100g,吐温80 10mL,121℃高温灭菌30min,调节pH6.0-6.5。
3.唾液乳杆菌(Lactobacillus salivariuse)GER106在制备提高免疫力药物或保健品中的应用。
4.如权利要求3所述的应用,唾液乳杆菌(Lactobacillus salivariuse)GER106的剂量为(0.5-3.0)×1010CFU/g。
5.唾液乳杆菌(Lactobacillus salivariuse)GER106在制备预防龋齿的食品、药物或保健品中的应用。
6.如权利要求5所述的应用,唾液乳杆菌(Lactobacillus salivariuse)GER106的剂量为(1.0-3.5)×1010CFU/g。
7.药物组合物,其特征在于,所述组合物中包括唾液乳杆菌(Lactobacillus salivariuse)GER106,还包括至少一种药学上可接受的载体、稀释剂、赋形剂或辅助剂。
8.权利要求7所述的药物组合物,其特征在于,所述药物组合物用于提高免疫力和/或预防龋齿;所述唾液乳杆菌(Lactobacillus salivariuse)GER106的剂量为(0.5-3.5)×1010CFU/g。
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Title |
---|
JEREMY P BURTON等: "Influence of the probiotic Streptococcus salivarius strain M18 on indices of dental health in children: a randomized double-blind, placebo-controlled trial", 《J. MED. MICROBIOL》 * |
张璇等: "唾液乳杆菌ZM06对变形链球菌生物膜形成的抑制作用", 《基因组学与应用生物学》 * |
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