藏灵菇中Enterococcus faecalis RQ15的分离及鉴定

以民间家庭制作酸奶用藏灵菇为试材,通过有氧与厌氧相结合的分离方式,从中分离到1株乳酸球菌。经形态学、生理生化特性研究及分子生物学鉴定,该菌株为粪肠球菌,编号Enterococcus faecalis RQ15。在12%复原脱脂乳中延时培养及冷藏保存该菌株时,产生胨化现象。后续的研究表明Enterococcus faecalis RQ15产生的蛋白酶具有低温蛋白酶的特征。     

藏灵菇的生理特性与发酵性能研究

以纯牛乳为原料,藏灵菇为发酵剂,通过单因素试验和正交试验从藏灵菇的生理特性和发酵性能方面探讨藏灵菇酸奶发酵的最佳生产工艺。综合考虑藏灵菇菌发酵乳较优发酵温度为37℃,接种量为5%,发酵时间为16h。最后得出结论,在最适条件下,以藏灵菇为发酵剂发酵的酸奶具有良好的风味和口感。     

藏灵菇发酵乳乳清的抗氧化及免疫学活性研究

藏灵菇培养发酵后,将发酵液加热除去沉淀,获得乳清液,冷冻干燥后得到乳清粉,将其分别配成1%和5%溶液进行小鼠灌胃实验,15 d后,检测昆明种小白鼠力竭游泳时间、血液SOD活性、对鸡红细胞致敏小鼠血清溶血素水平、小鼠腹腔巨噬细胞吞噬活性及对二硝基氟苯诱导小鼠迟发型超敏反应的影响。结果表明:5%剂量组的小鼠力竭游泳时间明显延长、红细胞中SOD的活性也明显提高、并能提高血清溶血素水平(P0.05);而且1%和5%剂量组都具有提高腹腔巨噬细胞吞噬率的作用(P0.05),这为开发利用藏灵菇提供了新的思路。     

藏灵菇微生物种群结构的分子特性研究

用PCR—DGGE指纹技术,研究了藏灵菇中微生物多样性及藏灵菇发酵奶发酵过程微生物种群动力学。结果表明,藏灵菇中细菌的种群结构较酵母菌的复杂,不同来源的藏灵菇中细菌种群结构的相似性为78％-84％,酵母菌种群结构的相似性为80％-92％。发酵过程中细菌种群结构变化图谱中的条带B和条带E,以及酵母菌种群结构变化图谱中的条带N贯穿于整个发酵过程,是发酵过程的优势菌。序列分析表明,细菌种群结构的DGGE图谱中的绝大多数条带与乳酸菌相对应,其中最亮条带（条带E）的序列与乳酸乳球菌的相似性为100％。     

藏灵菇微生物种群结构的分子特性研究*

用PCR-DGGE指纹技术,研究了藏灵菇中微生物多样性及藏灵菇发酵奶发酵过程微生物种群动力学。结果表明,藏灵菇中细菌的种群结构较酵母菌的复杂,不同来源的藏灵菇中细菌种群结构的相似性为78%~84%,酵母菌种群结构的相似性为80%~92%。发酵过程中细菌种群结构变化图谱中的条带B和条带E,以及酵母菌种群结构变化图谱中的条带N贯穿于整个发酵过程,是发酵过程的优势菌。序列分析表明,细菌种群结构的DGGE图谱中的绝大多数条带与乳酸菌相对应,其中最亮条带(条带E)的序列与乳酸乳球菌的相似性为100%。     

16S rRNA和recA、groEL基因分类鉴定乳酸乳球菌乳酸亚种和乳脂亚种的比较

【目的】比较16S rRNA和recA、groEL基因部分序列用于乳酸乳球菌乳酸亚种和乳脂亚种分类鉴定的效果。【方法】对已鉴定的8株分离自传统发酵乳的乳酸乳球菌, 选取recA和groEL基因片段, 通过PCR扩增、测序, 将测序得到的序列比对后构建系统发育树, 并与16S rRNA基因序列分析技术进行比较。【结果】比较分析不同菌株16S rRNA和recA、groEL基因的亲缘关系, recA、groEL基因可以准确地完成乳酸乳球菌乳酸亚种和乳脂亚种的区分和鉴定。【结论】recA和groEL基因序列分析可以实现乳酸乳球菌乳酸亚种和乳脂亚种的区分, 因其具有快速、准确、稳定的特点, 可适合于乳酸乳球菌乳酸亚种和乳脂亚种间的快速分类鉴定。     

多位点测序分型技术在病原微生物分型鉴定中的应用

多位点测序分型(Multilocus sequence typing,MLST)技术是一种以核苷酸序列为基础的病原菌分型方法,它是高通量测序技术与成熟的群体遗传学相结合的产物。该方法简单易行,重复性强,可以通过国际互联网对某一致病菌株在全球范围内的传播分布情况进行追踪监控。目前,MLST技术已被广泛应用于原核病原菌及一些真核病原菌(如真菌)的分型鉴定中。主要对MLST技术的原理及其在一些常见病原菌分型鉴定中的应用进行了简要的阐述。     

16S rRNA序列分析及其相关分子生物学方法在乳酸菌分类、鉴定中的应用

乳酸菌是一类能在可利用的碳水化合物发酵过程中产生大量乳酸的细菌。目前,在自然界已发现至少43个属的这类菌。近年来,随着乳酸菌益生作用的相继发现,乳酸菌制品的生产及销售量也逐年呈上升趋势,正因如此,对乳酸菌进行正确的鉴定已成为乳酸菌产业中的重要内容之一。     

几何形态测量学方法在小哺乳动物化石分类鉴定中的应用——4种(鼠平)类化石大样本的个案研究

小哺乳动物化石在晚新生代生物地层学和生物年代学研究中具有重要作用,尤其是筛洗法在古生物调查中得到广泛应用后,通常可以采集到数量可观的标本,使得其地位较大型哺乳动物化石更加彰显.因此,小哺乳动物化石标本的分类鉴定也成了一项十分关键的工作.然而,传统形态学方法在对大量标本进行分类鉴定时,往往容易受主观因素影响而将不稳定的细微性状变异作为依据建立新种,或者忽视一些肉眼难于察觉的形态学差异而将两个甚至多个类群合并到一起,导致基于形态学的化石分类鉴定随意性增加,失去客观性.此外,对于不具鉴定意义的非关键性单个牙齿,很难凭借肉眼或显微镜观察进行区分.针对这些问题,本文选取了安徽繁昌人字洞早更新世早期三种(鼠平)类甘肃模鼠Mimomys gansunicus,郑氏异模鼠Heteromimomys zhengi和繁昌维蓝尼鼠Hllanyia fanchangensis的1284件臼齿并以早上新世内蒙古比例克的比例克模鼠Mimomys bilikeensis的163件臼齿作为参考,采用几何形态测量学方法在各个臼齿咬合面上分别选取了7～14个同源landmark对咬合面的形态特征进行了线性判别分析,建立了针对这4个种的臼齿咬合面形态差异判别函数.分析结果表明,根据这些landmark所提供的形态差异信息,人字洞的1284件臼齿标本中的确存在3个可以明显区分且形态学性状稳定的类群,先前的分类鉴定得到了验证.与之不同地点不同时代的比例克模鼠也可以很好区分.因此,建立在大样本基础上的这4种(鼠平)类臼齿咬合面同源形态特征的判别函数可以用来描述这些种类较为稳定的形态特征差异,并用来作为今后对样本较少的标本进行分类鉴定时的判别依据.因为几何形态测量学的方法不仅可以对二维的离散landmark数据以及重要形态学特征的轮廓线进行分析,甚至还可以扩展至三维空间,所以上述方法对于较为容易获得大样本标本的小哺乳物化石分类鉴定具有普遍意义.     

Lactic acid bacteria and yeasts in kefir grains and kefir made from them

In an investigation of the changes in the microflora along the pathway: kefir grains (A)→kefir made from kefir grains (B)→kefir made from kefir as inoculum (C), the following species of lactic acid bacteria (83–90%) of the microbial count in the grains) were identified: Lactococcus lactis subsp. lactis, Streptococcus thermophilus, Lactobacillus delbrueckii subsp. bulgaricus, Lactobacillus helveticus, Lactobacillus casei subsp. pseudoplantarum and Lactobacillus brevis. Yeasts (10–17%) identified were Kluyveromyces marxianus var. lactis, Saccharomyces cerevisiae, Candida inconspicua and Candida maris. In the microbial population of kefir grains and kefir made from them the homofermentative lactic streptococci (52–65% and 79–86%, respectively) predominated. Within the group of lactobacilli, the homofermentative thermophilic species L. delbrueckii subsp. bulgaricus and L. helveticus (70–87% of the isolated bacilli) predominated. Along the pathway A→B→C, the streptococcal proportion in the total kefir microflora increased by 26–30% whereas the lactobacilli decreased by 13–23%. K. marxianus var. lactis was permanently present in kefir grains and kefirs, whereas the dominant lactose-negative yeast in the total yeast flora of the kefir grains dramatically decreased in kefir C. Journal of Industrial Microbiology & Biotechnology (2002) 28, 1–6 DOI: 10.1038/sj/jim/7000186 Received 02 August 2000/ Accepted in revised form 15 July 2001     

应用变性梯度凝胶电泳和16SrDNA序列分析对kefir粒中细菌多样性的研究

以开菲尔（Kefir）粒为材料,经过DNA抽提和16SrDNA V3区PCR扩增,扩增产物经变性梯度凝胶电泳（DGGE）分离并切割电泳条带进行序列测定,并与现有的数据库进行了比较,对Kefir粒的细菌多样性进行分析。结果表明,DGGE图谱中可检测到的8条带的16SrDNA基因序列中有7个基因序列与GenBank数据库登录的相关序列的相似性大于98％,余下的1个基因序列的相似性也大于96％。相似性大于98％的7个克隆中,有3个属于鞘氨醇杆菌属（Sphingobacterium）,2个属于乳杆菌属（Lactobacillus）,其它2个分别属于肠杆菌属（Errterobacter）和不动杆菌属（Acinetobacter）。首次报道了鞘氨醇杆菌作为优势菌群存在开菲尔Kefir粒中。     

应用变性梯度凝胶电泳和16S rDNA序列分析对kefir粒中细菌多样性的研究

以开菲尔(Kefir)粒为材料,经过DNA抽提和16S rDNA V3区PCR扩增,扩增产物经变性梯度凝胶电泳(DGGE)分离并切割电泳条带进行序列测定,并与现有的数据库进行了比较,对Kefir粒的细菌多样性进行分析。结果表明,DGGE图谱中可检测到的8条带的16S rDNA基因序列中有7个基因序列与GenBank数据库登录的相关序列的相似性大于98%,余下的1个基因序列的相似性也大于96%。相似性大于98%的7个克隆中,有3个属于鞘氨醇杆菌属(Sphingobacterium),2个属于乳杆菌属(Lactobacillus),其它2个分别属于肠杆菌属(Enterobacter)和不动杆菌属(Acinetobacter)。首次报道了鞘氨醇杆菌作为优势菌群存在开菲尔Kefir粒中。     

Detection of interactions between yeasts and lactic acid bacteria isolated from sugary kefir grains

Different techniques were tested for studying the synergism between the micro-organisms of sugary kefir grains. Agar cultures in Petri dishes did not give reproducible results. In sequential cultures, i.e. growing one organism, sterile filtering and then inoculating the other, 10 of 18 selected lactic acid bacteria/yeast pairs revealed stimulation of bacterial growth in a poor glucose medium. In mixed culture, Saccharomyces florentinus supported better survival of Lactobacillus hilgardii and a significant increase in lactic acid production; at the same time, the growth and alcoholic fermentation of S. florentinus were drastically reduced. The inter-relationships between these two strains were the same when immobilized in calcium alginate beads, even though total metabolite production was always lower than with free cells. The stimulation of Lact. hilgardii by Candida lambica in sequential culture was not confirmed in mixed culture, where the two organisms grew as in pure culture, and bacterial growth and lactic acid production were inhibited in the immobilized system.     

Numerical taxonomy of psychrotrophic lactic acid bacteria from prepacked meat and meat products

Ninety-four strains of lactic acid bateria isolated from refrigerated, prepacked meat and meat products were together with 59 reference strains of Brochothrix, Lactobacillus, Leuconostoc, Pediococcus and Streptococcus phenotypically classfied, using 96 unit characters. Data were examined using Simple Matching (S_SM) or Jaccard coefficient (S_J), and unweighted pair group algorithm with arithmetic averages. Twenty-three clusters with two or more members were defined at the 84% S_SM-similarity level which corresponded to the S_J-similarity level of 61%. Based on S_SM, most field strains were included in nine clusters, and with three unsignificant exceptions these contained no reference strains. The field clusters were designated Carnobacterium piscicola (cluster 1; 5% of field isolates), Carnobacterium divergens(cluster 2; 9% of field isolates), Leuconostoc (cluster 9; 18% of field isolates) and Lactobacillus (cluster 4, 10, 11, 12, 13 and 14; together 60% of field isolates). The Lactobacillus clusters had many features in common with cluster II of Shaw & Harding (1984). Phenotypical characteristics of major clusters are given. The S_SM and S_J based classifications basically coincided for the field strains; the exception was cluster 4 which now were split in two parts. Fourteen clusters were made up of mainly reference strains (S_SM). Most of them included more than one type strain on species level; exceptions were Brochothrix thermosphacta (cluster 3), Lactobacillus salivarius (cluster 17) and Leuconostoc mesenteroides (cluster 18). Several rearrangements were seen amongst the clusters of the reference strains when S_J, instead of S_SM, was used for clustering.     

Identification of lactic acid bacteria isolated from human blood cultures

Fifteen lactic acid bacterial strains were isolated from blood cultures from 15 different patients in the Faculty Hospital in Brno, Czech Republic. All strains were identified using biochemical tests and repetitive PCR using the (GTG)5 primer. Doubtful identification results were confirmed by whole-cell protein analysis. The strains were assigned to the genera Lactobacillus (eight strains representing seven species), Leuconostoc (six strains representing four species) and Weissella (one strain). Antibiotic susceptibility testing was performed using the E-test and revealed high-level resistance to cotrimoxazol, metronidazole, vancomycin and teicoplanin, but nearly all strains were susceptible to erythromycin, clindamycin, ampicillin and penicillin.     

西藏小麦耐盐性鉴定及分析

在3种不同盐胁迫浓度下对161份来源于西藏的小麦材料进行了苗期耐盐性鉴定,其中有147份西藏半野生小麦及14份密穗小麦.以普通小麦耐盐品种茶淀红麦和盐敏感品种PI94341为对照.在147份西藏半野生小麦中有14份表现为耐盐,占9.5%.14份密穗小麦中没有发现耐盐材料.在这161份材料中还发现了3份对盐胁迫浓度梯度不敏感的品种.结果表明,我国特有的西藏半野生小麦中蕴藏有丰富的耐盐种质,它们可供小麦耐盐育种用做亲本.     

酸马奶中乳酸菌的鉴定及生物学特性的研究

[目的]对从酸马奶中分离出来的10株乳酸菌进行鉴定和生理生化特性研究,为工业生产筛选特性优良的菌种.[方法]通过形态学观察、生理生化特性、分子生物学特性及其对致病菌抑制作用的研究对其进行鉴定,并筛选特性优良菌株.[结果]10株乳酸菌分别为2株Lactobacillus plantarum、2株Enterococcus villorum、2株Enterococcus dispar、3株Enterococcus durans和1株Enterococcus raffinosus;其对Staphylococcus aureus、Escherichia coli和Enteritidis bacillus有不同程度的抑制作用.[结论]菌株HZ24、HZ25具有良好的生物学特性和益生功能,可以应用到食品发酵工业生产中.     

Lactobacillus tucceti sp. nov., a new lactic acid bacterium isolated from sausage

Following the application of several molecular techniques strain R 19c, isolated from sausage by Reuter in 1970 and deposited at the DSMZ as Lactobacillus sp., has been identified as pertaining to a new species. It showed singular ISR-DdeI and ISR-HaeIII profiles that allowed its differentiation from 68 lactic acid bacteria reference strains analyzed. Phylogenetic analysis based on 16S rRNA gene sequences places this strain in the genus Lactobacillus within the Lactobacillus alimentarius group. Species L. versmoldensis is the closest phylogenetic neighbor with 96.3% sequence similarity. DNA-DNA hybridization experiments confirmed the independent status at species level of this strain. Species-specific primers for PCR detection of this new species have been developed. Phenotypically it can be distinguished from the closest relative L. versmoldensis by several traits such as the peptidoglycan type (L-Lys-Gly-D-Asp), acid production from L-rhamnose, D-mannitol and L-fucose and its inability to ferment d-galactose, d-melibiose and d-sucrose. The name Lactobacillus tucceti sp. nov. is proposed with strain R 19c(T) (=DSM 20183(T)= CECT 5920(T)) as the type strain.