益生菌降解胆固醇的作用及机理研究进展

益生菌是来源于宿主并对宿主健康有一定促进作用的微生物。降胆固醇功能是某些益生菌的主要益生功能之一。近年来国际上对益生菌降胆固醇的体外和体内研究进展,主要包括菌体生长与降胆固醇的关系、pH的影响、胆酸盐的作用、胆酸盐水解酶活性、益生元的使用、人和动物喂养实验等方面。目前人们提出的益生菌降胆固醇作用机理的假说主要有共沉淀作用、酶对胆酸盐的分解作用、胆固醇掺入细胞膜、细菌对胆固醇的吸收等,这些机理假说尚有待进一步研究证实。降胆固醇益生菌制品研发的前景十分广阔。     

体外乳酸菌素促双歧杆菌生长实验初步观察

目的 在体外观察乳酸菌素是否有促双歧杆菌生长的作用。方法 将浓度为１００ｍｇ／ｍｌ、１０ｍｇ／ｍｌ、１ｍｇ／ｍｌ的乳酸菌素液按三角形对等距离分别滴种在接种了双歧杆菌（１０＾６ＣＦ／ｍｌ）的ＢＨＩ培养基上,然后置３５℃厌氧箱培养２ｄ后观察细菌生长情况。结果 不同浓度的乳酸菌素液促进双歧杆菌生长能力不同。浓度为１００ｍｇ／ｍｌ、１０ｍｇ／ｍｌ、１ｍｇ／ｍｌ的乳酸菌素液促双歧杆菌生长的阳性率（％）分别为：９６．７、５６．７、１．０。经统计学秩和检验方法将３种浓度乳酸菌素液促双歧杆菌生长作用进行两两比较得知：１００ｍｇ／ｍｌ与１０ｍｇ／ｍｌ,１００ｍｇ／ｍｌ与１ｍｇ／ｍｌ之间Ｐ〈０．０１,１０ｍｇ／ｍｌ与１ｍｇ／ｍｌ比较Ｐ〈０．０５。结论 乳酸菌素有促进益生菌－－双歧杆生长的作用,而且与其浓度高低有极密切关系,高浓度的     

益生菌体外增殖因子的初步研究

目的研究单糖、pH、温度及时间对青春双歧杆菌、长双歧杆菌和类干酪乳杆菌体外增殖的影响。方法用甘露糖、半乳糖、山梨醇及果糖代替MRS中的葡萄糖,筛选出每种细菌的最适碳源。以此为基础,选择其最佳初始pH、培养温度、碳源添加量及培养时间。结果青春双歧杆菌、长双歧杆菌和类干酪乳杆菌的最适碳源分别为葡萄糖、甘露糖和半乳糖;最佳初始pH为6.0、7.0和6.0;培养温度为42、30和30℃;碳源添加量为20、15和25 g/L;培养时间都为28-48 h。结论益生菌具有不同的最适增殖条件,本文研究结果为优化益生菌的生长条件提供了基础数据。     

肠道益生菌体外抑菌活性的研究

以6种常见肠道致病菌作为指示菌,研究了来自健康青年和儿童肠道的25株益生菌发酵浓缩液的体外抑菌效果.进一步研究了抑菌作用良好的LMa1和LMa3经pH调节、热处理、蛋白酶处理以及发酵过程中的抑菌活性变化.短链有机酸作为发酵终产物的主要成分之一,其抑菌效果也被测定.结果表明,抑菌物质包括短链有机酸（主要为乙酸和乳酸）和一类对热稳定、对部分蛋白酶敏感的物质,可能是某种细菌素.低pH是表现抑菌活性的必要条件,但仅有低pH（如低pH缓冲液）并无明显抑菌效果.     

我国微生态制剂的现状和发展设想

通过人外文献和市场的调查,并结合科研实践,指出了当前我国微生态制剂发展中存在的一些严峻问题,在介绍了人体肠道微生态规律、微生态制剂应有的质量标准,以及比较了多种产品的活菌含量和“亿活菌价”的基础上,作者认为,提高我国微生态制剂质量的４个关键应该是：选育优良菌种,提高活菌含量,延长保质期和添加有益菌的生长促进剂。为促进我国微生态制剂事业更快、更稳健地发展,作者提出了７项具体建议。     

益生菌对婴幼儿湿疹的预防作用观察

目的 探讨益生菌是否对婴幼儿湿疹有预防作用。方法 110例新生儿,随机双盲分组,治疗组52例,对照组58例,治疗组于新生儿期口服含有双歧杆菌与乳酸菌的益生菌制剂,连用3～6月,对照组仅作随防。对比2组湿疹的发生情况。结果 2组均于生后1个月开始随访,至6～12月,共有39例患婴幼儿湿疹,占总人数的35.4％,治疗组11例占28．2％,对照组28例占71．8％。结论 新生儿期口服益生菌对婴幼儿湿疹有预防作用。     

益生菌促进胃肠道健康的机制及应用

人体胃肠道内生活着大量微生物,它们影响着宿主的健康.益生菌是一种活的微生物,对维持肠黏膜屏障功能、调节免疫功能和促进营养物质的代谢吸收等具有重要作用;对肠道菌群紊乱、功能性消化不良、肠胃炎、腹泻、便秘、肠绞痛、肠易激综合征、炎症性肠病以及幽门螺杆菌感染等胃肠道疾病具有良好的应用.本文对益生菌与胃肠道健康的影响作简要概述...     

食物过敏患儿肠道菌群及sIgA含量的研究

目的通过研究儿童肠道菌群数量与粪便sIgA含量之间的相互关系,探讨菌群失调引起食物过敏的免疫致病机制。方法筛选2个月～6岁患儿49名和健康儿童57名,采用荧光定量PCR反应检测粪便中的双歧杆菌、乳酸菌、肠杆菌数量,并采用ELISA法检测粪便sIgA含量。结果与健康儿童比较,食物过敏患儿肠道双歧杆菌、乳酸菌显著下降,肠杆菌显著升高（P〈0．05）粪便sIgA含量显著下降（P〈0．05）。结论儿童肠道菌群失调是引起过敏性疾病的主要原因之一。     

保健食品双歧杆菌和乳酸菌计数方法的优化

目的通过稀释液和培养基的选择对保健食品双歧杆菌和乳酸菌计数方法进行优化。方法对2个品种6个批次的益生菌类保健食品,分别采用2种稀释液和3种培养基进行双歧杆菌和乳酸菌计数,并比较结果。结果选择CYS缓冲液为最优稀释液,M-TOS培养基为双歧杆菌计数的最优培养基,BBL培养基为乳酸菌总数计数的最优培养基。结论优化方法所得菌数均明显优于国标方法,可以用于保健食品的双歧杆菌和乳酸菌计数。     

双歧杆菌及微生态制剂

自 Tissier于 1899首先发现双歧杆菌 (Bif idobacterrium)以来 ,双歧杆菌及其制品与人们的日常生活间的关系日趋紧密。已报道双歧杆菌属的 3 2个种 [1]栖居于人和动物 (牛、羊、兔、鼠、猪、鸡和蜜蜂等 )的肠道、反刍动物的瘤胃、人的齿缝和阴道等部位。其中除齿双歧杆菌可能是病原菌外 ,其他种尚无致病性的报道。近年 ,含有一定数目活菌的微生态制剂的研究取得了很大的进展 ,尤其是含有双歧杆菌的微生态制剂。据19 94年统计 ,国内约有 60多种含双歧杆菌的产品投入市场。在国外 ,含双歧杆菌的食品也十分流行 ,特别是在日本、欧洲和北美。据…     

Viili中益生菌的分离及其发酵性质研究

目的 利用传统方法和分子生物学方法从viili中筛菌并对其特性进行研究.方法 采用MRS、YPD和脱脂乳营养琼脂3种平板从viili中分离出15株单菌,利用V3区通用引物和乳杆菌特异性引物对各单菌的PCR产物进行变性梯度凝胶电泳(denaturing gradient gel electrophoresis,DGGE)分析,将其归为5种不同菌.结果 16S DNA测序表明,5种单菌分别为Lactobacillus plantarum、Streptococcus thermophilus、Lactobacillus paracasei、Bacillus cereus和Lactobacillus delbrueckii subsp. bulgaricus.混合发酵实验表明:从viili中筛出的5种菌除Bacillus cereus外均有凝乳现象,其中Lactobacillus delbrueckii凝乳能力强,可在5 h内凝乳;Lactobacillus paracasei、Bacillus cereus和Lactobacillus plantarum组合产生的EPS量最多,高达186.71 mg/L,而Streptococcus thermophilus EPS产量仅为33.56 mg/L.结论 传统方法与分子生物学方法DGGE相结合,可以快速准确判断细菌种类;筛选菌特性研究结果表明,细菌之间的相互作用导致凝乳时间和EPS产量发生变化,其复合作用有待于进一步研究.     

益生菌的安全性

益生菌是指一类活的,摄入足够量就能够对人体产生有益作用的微生物,目前广泛应用于食品发酵、工业乳酸发酵以及医疗保健领域.随着市场上商品化益生菌的不断出现,它所带来的安全性问题也更加引起人们的关注.目前益生菌主要存在四个方面的安全问题:致病性和感染能力;有害的代谢活动:过度的免疫反应和可能的基因转移.传统的益生菌安全性评价方法具有一定的局限性.我们需要针对目前益生菌安全性存在的问题建立一套包含基因组学,代谢组学,蛋白质组学等研究内容的评估方法,对益生菌的安全性进行系统全面的评估.本文总结了一些对于益生菌安全性的研究进展和研究方法,以提示我国应尽快完善益生菌及其制品的安全性评价方法指标并建立安全性评价体系,使益生菌更好的为人们的健康服务.     

应用DGGE和Real-time PCR分析酸菜发酵液中乳酸菌的动态变化

目的 使用DGGE和实时荧光定量PCR分析酸菜发酵液中乳酸菌的动态改变。方法 采集传统天然发酵1~8周的酸菜发酵液50 mL,抽提基因组DNA,使用DGGE对乳酸菌菌群进行多样性、相似性研究和优势菌的鉴定,实时荧光定量PCR测定乳酸菌含量。结果 发酵周期加长,乳酸菌的含量随之也逐渐增大。清酒乳杆菌是酸菜自然发酵过程中的优势菌型,鼠李糖乳杆菌主要存在于发酵初期,发酵的中期（3~5周）、后期（6~8周）明显减少,同时发酵后期植物乳杆菌成为优势菌并完成全部发酵过程。发酵周期延长后,香农多样性指数和丰富度较高,出现先下降后上升的趋势,在第7周达到最大值。结论 在DNA水平上,DGGE和实时定量PCR检测了酸菜发酵液中乳酸菌的含量,进行菌群结构的动态分析,明确优势菌型,为实现酸菜标准化生产提供理论依据。     

Rapid lactic acid bacteria identification in dairy products by high-resolution melt analysis of DGGE bands

Aim: To investigate the application of high‐resolution melt (HRM) analysis for rapid species‐level identification of lactic acid bacteria (LAB) communities in dairy products, as well as for bacterial community profiling and monitoring. Methods and Results: First, comparisons of HRM profiles of known reference strains of LAB and their denaturing gradient gel electrophoresis (DGGE) bands showed very good agreement, allowing species recognition and identification from DGGE bands by HRM. Second, samples of cheese, kefir grains and kefir were characterized by PCR‐DGGE, and melting profiles of DGGE bands were compared with known reference strains. Of the 13 DGGE bands, ten were identified by HRM by comparison with the reference strains and only three required sequencing for identification. Use of HRM profiling for comparison and monitoring of total LAB communities from dairy products or starter cultures was also evaluated, and good agreement was found when comparing clustering of DGGE band profiles with clustering of HRM melting profiles. Conclusion: Identification of DGGE bands is possible by comparison of HRM melting profiles with known reference strains. Significance and Impact of the Study: HRM profiling is suggested as an additional approach for identification of DGGE bands.     

Chitosan oligosaccharides, dp 2-8, have prebiotic effect on the Bifidobacterium bifidium and Lactobacillus sp

In order to investigate the prebiotic potential of chitosan oligosaccharide (COS), prepared by enzymatic hydrolysis of fully deacetylated chitosan polymer, the effect of COS on bacterial growth was studied. The degree of polymerization (dp) of COS was determined by MALDI-ToF mass spectrometry, and the COS was found to be composed of dimer (33.6%), trimer (16.9%), tetramer (15.8%), pentamer (12.4%), hexamer (8.3%), heptamer (7.1%), and octamer (5.9%). The minimum inhibitory concentrations (MIC) of chitosan polymer against lactic acid bacteria and bifidobacteria were below 0.31%. However, this only applied to two strains, the other bacteria tested grew on MRS broth containing 5% COS. The effects of COS on the growth of bifidobacteria and lactic acid bacteria were compared with those of fructo-oligosaccharide (FOS). FOS was found to have a growth stimulatory effect on only three strains: Bifidobacterium bifidium, B. infantis and Lactobacillus casei. However, COS stimulated the growth of most Lactobacillus sp. and B. bifidium KCTC 3440. The amount of the growth and the specific growth rate of B. bifidium increased with increasing COS concentration. The cultivation time required to obtain maximum growth was reduced to about 25% in MRS broth supplemented with 0.2-0.4% COS. These results demonstrate that COS has considerable bifidogenic potential. Both cell growth and specific growth rates of L. brevis in MRS broth supplemented with 0.1% COS increased by 25%. The present study shows that COS stimulates the growth of some enteric bacteria, and that COS has potential use as a prebiotic health-food.     

DGGE技术在森林土壤微生物多样性研究中的应用

微生物在森林土壤物质转化中扮演着重要角色,与森林的林型、土壤理化性质存在着密切关系.森林土壤微生物多样性及其变化在一定程度上反映了土壤环境的生产力和稳定性,对表征森林演替,土壤生态修复等有重要意义.变性梯度凝胶电泳(DGGE)技术测定微生物多样性具有快捷、高效和可重复性高等优点.简要介绍DGGE技术的原理,分析这一技术的局限性和优化方法.重点以实例说明该技术在森林土壤微生物多样性研究中的应用现状,展望该技术的发展前景,以期能为今后这一领域的研究提供科学依据.     

Progress and potential in the biotechnology of lactic acid bacteria

Abstract: Current activities and future prospects for the biotechnology of lactic acid bacteria are reviewed. Genetic engineering technology, including advances and limitations of plasmid vectors and chromosomal integration strategies are discussed together with the status of gene expression and the importance of in vivo gene transfer systems and transposition. Areas of biotechnological application considered include proteolysis and flavour generation, bacteriophage resistance, antimicrobials, metabolic engineering and the possible uses of lactic acid bacteria in relation to health.     

Characterization of d-amino acid aminotransferase from Lactobacillus salivarius

We searched a UniProt database of lactic acid bacteria in an effort to identify d-amino acid metabolizing enzymes other than alanine racemase. We found a d-amino acid aminotransferase (d-AAT) homologous gene (UniProt ID: Q1WRM6) in the genome of Lactobacillus salivarius. The gene was then expressed in Escherichia coli, and its product exhibited transaminase activity between d-alanine and α-ketoglutarate. This is the first characterization of a d-AAT from a lactic acid bacterium. L. salivarius d-AAT is a homodimer that uses pyridoxal-5′-phosphate (PLP) as a cofactor; it contains 0.91 molecules of PLP per subunit. Maximum activity was seen at a temperature of 60 °C and a pH of 6.0. However, the enzyme lost no activity when incubated for 30 min at 30 °C and pH 5.5 to 9.5, and retained half its activity when incubated at pH 4.5 or 11.0 under the same conditions. Double reciprocal plots of the initial velocity and d-alanine concentrations in the presence of several fixed concentrations of α-ketoglutarate gave a series of parallel lines, which is consistent with a Ping-Pong mechanism. The K_m values for d-alanine and α-ketoglutarate were 1.05 and 3.78 mM, respectively. With this enzyme, d-allo-isoleucine exhibited greater relative activity than d-alanine as the amino donor, while α-ketobutylate, glyoxylate and indole-3-pyruvate were all more preferable amino acceptors than α-ketoglutarate. The substrate specificity of L. salivarius d-AAT thus differs greatly from those of the other d-AATs so far reported.     

猪链球菌口服疫苗的制备及小鼠血清免疫效果的评价

为构建猪链球菌(Streptococcus suis)新型疫苗,给猪链球菌病的防治提供新思路,首先在大肠杆菌中表达猪链球菌毒力因子EF和MRP,以His-tag柱纯化重组蛋白,并制备EF和MRP鼠源多克隆抗体.随后将ef和mrp基因置于组成型启动子P59和信号肽Usp45下,克隆到乳酸菌表达载体pMG36e上,电击转化到干酪乳杆菌(Lactobacillus casei)ATCC27092中进行表达.Western blot 检测显示EF和MRP蛋白均能在乳酸菌中成功表达,且EF蛋白的表达量随着重组菌生长时间的增长而增大,而MRP蛋白的表达量在重组菌生长到OD600为0.8时达到最高,随后慢慢下降.用含有表达EF和MRP蛋白的重组干酪乳杆菌喂饲C57BL/6J小鼠,发现重组菌可在小鼠体内存活2d左右,并可有效刺激小鼠产生EF和MRP特异性抗体,为研制乳酸菌口服疫苗防治猪链球菌病的可行性进行了有益的探索.