水井坊窖底泥微生物群落结构的DGGE分析

为了补充和验证研究浓香型白酒不同窖泥的微生物群落特征分析,该研究采用DGGE技术对水井坊酒厂窖龄分别为50及150年相同窖池不同部位窖底泥（三点取样法）进行细菌、古菌和真菌微生物群落特征分析。研究结果显示：50年和150年窖龄窖泥的微生物群落的演替特征和分布特征具有一定相似性,进一步验证了50年可能是窖池微生态发生较大变化的一个分界线这一推测的准确性;不同部位的窖底泥的微生物群落结构具有一定的差异性,因此在今后对窖底泥的实验研究中,采用多点取样法进行分析。     

An assessment of factors characterising the microbiology of Grana Trentino cheese,a Grana‐type cheese

In this study, microbiological aspects of Grana Trentino, a variant of Grana Padano cheese, were defined by plate counts, random amplified polymorphic DNA (RAPD) PCR genotying, 16S rRNA gene sequencing of bacterial isolates and PCR–denaturing gradient gel electrophoresis (PCR–DGGE). Results showed variability in monthly fluctuations of whey culture counts, differences in the diffusion of bacterial genotypes among producers and dairy plant‐specific microbial associations. Moreover, the presence of bacteria not previously reported in this cheese type was highlighted, including coagulase‐negative staphylococci and Lactobacillus sanfranciscensis‐like micro‐organisms.     

超高压处理抑制低温烟熏火腿中的优势腐败菌

研究尝试应用微生物菌体总RNA提取代替DNA提取,进而通过反转录-PCR( RT-PCR),结合变性梯度凝胶电泳(denaturing gradient gel electrophoresis,DGGE)技术,以期揭示超高压处理后低温烟熏火腿中腐败微生物的存活情况,探索RNA-DGGE手段判定超高压处理后微生物存活状态的可行性.分别以400 MPa和600 MPa的压力在室温(22℃)条件下,对包装后的烟熏火腿进行10 min超高压处理,未经高压处理样品作对照,于4℃冷藏条件下,贮藏1、15、30、60、90 d,直接提取样品中微生物的总RNA,对其进行RT-PCR和DGGE指纹图谱分析.DGGE指纹图谱显示,超高压处理对烟熏火腿中的优势腐败菌具有较强的抑制作用,且随压力的升高抑菌效应增强;超高压处理后烟熏火腿微生物种群结构变得单一,Weissella viridescens和Leuconostoc mesenteroides是超高压处理后烟熏火腿中的优势腐败菌.基于菌体总RNA提取的DGGE手段能够有效检测超高压处理后微生物的存活状况,揭示超高压对低温烟熏火腿中优势腐败微生物的抑菌效果.     

浓香型白酒窖泥原核微生物DGGE电泳条件的优化

将DGGE方法应用于浓香型白酒窖泥原核微生物的研究,并对其电泳条件进行优化;对窖泥微生物16S rDNA的V6-V8高变区优化出的DGGE电泳条件为:胶浓度8%,变性范围40%～70%,电压条件先采用200 V、10min,再采用85V、15.5h,其电泳图谱具有较好的分离效果。     

Effect of pumping on the spatio-temporal distribution of microbial communities in a water well field

A water well field adjacent to the North Saskatchewan River (City of North Battleford, Saskatchewan, Canada) with a history of rapid deterioration of both well water quality and yield was selected to study the spatial and temporal distribution of subsurface microbial communities and their response to water pumping. A range of conventional cultural, microscopic and molecular techniques, including confocal laser scanning microscopy (CLSM), Biolog, qPCR and Denaturing Gradient Gel Electrophoresis (DGGE), was used during this study. Redox data and water and sediment chemistry showed that the aquifer was anoxic and harbored substantial amounts of Fe and Mn. CLSM analyses of incubated coupons indicated extensive biofilm growth in the zone immediately surrounding the well and was coincident with reduced water well yield. PCR screening and qPCR analyses showed that the potential for iron- and sulfate-reducing activity increased with proximity to the well. Bacterial communities inhabiting the zone closest to the well showed the greatest changes and differences in metabolic activities and composition as revealed by PCA (Principal Components Analysis) of the Biolog and DGGE data. The sequence analysis of all the samples revealed that Sulfuricurvum spp., Methylobacter spp., Geobacter spp. and Rhodobacter spp. were most commonly detected in this aquifer. Overall the findings demonstrated that the microbial numbers, metabolic activities and the community composition changed in response to water pumping but effects did not extend beyond 1-2 m zone from the well.     

PCRDGGE技术分析混菌发酵乳中马克斯克鲁维酵母与乳酸菌的相互作用

利用变性梯度凝胶电泳(Denaturing Gradient Gel Electrophoresis,DGGE)技术,对混菌发酵及贮藏过程中马克斯克鲁维酵母与乳酸菌之间的相互作用进行分析,进而对整个过程中微生物优势菌群及其稳定性进行跟踪监测。结果表明:混菌发酵及贮藏过程中微生物组成比较稳定,优势菌为嗜热链球菌(Streptococcus thermophilus,ST);发酵过程中,马克斯克鲁维酵母(Kluyveromyces marxianus)的添加对乳酸菌生长起到促进作用,尤其是对保加利亚乳杆菌(Lactobacillus bulgaricus,LB)效果显著,贮藏期间该作用转变为抑制;整个过程乳酸菌的存在对酵母菌的生长具有一定的抑制作用。该研究可为深入探讨乳酸菌与酵母菌共同发酵机理及新型发酵乳制品的开发提供理论基础。     

Monitoring of the microbial communities involved in the soy sauce manufacturing process by PCR-denaturing gradient gel electrophoresis

Soy sauce is a traditional seasoning produced through the fermentation of soybeans and wheat using microbes. In this study, the microbial communities involved in the soy sauce manufacturing process were analyzed by PCR-Denaturing Gradient Gel Electrophoresis (PCR-DGGE). The bacterial DGGE profile indicated that the bacterial microbes in the koji were Weissella cibaria (Weissella confusa, Weissella kimchii, Weissella salipiscis, Lactobacillus fermentum, Lactobacillus plantarum, Lactobacillus iners, or Streptococcus thermophilus), Staphylococcus gallinarum (or Staphylococcus xylosus), and Staphylococcus kloosii. In addition to these bacteria, Tetragenococcus halophilus was also detected in the mash during lactic acid fermentation. The fungal DGGE profile indicated that the fungal microbes in the koji were not only Aspergillus oryzae but also several yeasts. In the mash, Zygosaccharomyces rouxii appeared in the early fermentation stage, Candida etchellsii (or Candida nodaensis) and Candida versatilis were detected at the middle fermentation stage, and Candida etchellsii was detected at the mature fermentation stage. These results suggest that the microbial communities present during the soy sauce manufacturing process change drastically throughout its production. This is the first report to reveal the microbial communities involved in the soy sauce manufacturing process using a culture-independent method.     

利用PCR-DGGE技术筛选分离海南黄帝椒产品微生物

黄帝椒是海南特产高辣度辣椒。该研究以黄帝椒产品为原料,采用聚合酶链式反应-变性梯度凝胶电泳(PCR-DGGE)技术分析了其微生物种群关系,并结合传统平板筛选法进行菌种分离及鉴定,获得了海南特辣黄帝椒产品优势微生物。研究结果显示,海南黄帝椒产品平板筛选分离到了Pseudomonas stutzeri,Lactobacillus plantarum,Lactobacillus brevis等可培养的菌株;DGGE图谱中检测到了6个条带,其中,乳酸菌占总菌数的65%,处于最优势地位,假单胞菌占总菌数的16%,处于次要地位。该研究也首次提及了不同的微生物对黄帝椒产品的颜色、脆性的影响。     

西藏Kefir粒中菌相的初步研究

利用扫描电镜、传统菌种分离及PCR-DGGE 3种技术相结合,对Kefir粒的菌相作了初步研究,试验结果显示,Kefir粒内层菌体密度较大,以短杆菌为主,有部分长杆菌,还有部分椭圆形酵母;对其中的乳酸菌和酵母菌进行了分离,共得到9株乳球菌、38株乳杆菌和9株酵母菌,描述了各菌株的细胞、菌落形态,研究了其牛乳发酵特性和半乳糖苷酶活性,并且对Kefir乳发酵过程中的菌数变化做了动态监测.     

冷藏带鱼贮藏期间主要微生物动态变化的PCR-DGGE分析

本文研究了冷藏带鱼贮藏期间的主要微生物种群演替规律,为延长水产品货架期提供参考。采用SDS化学裂解法,从冷藏带鱼中提取总细菌基因组DNA,并进行16SrRNA的V3可变区PCR扩增,再通过DGGE得到动态指纹图谱。结果表明,贮藏初期冷藏带鱼样品中的微生物群落丰富,嗜冷杆菌(Psychrobacter sp.)为前期的主要优势菌。随着贮藏时间的延长,仅有少数种类细菌存活并最终成为主导菌群。荧光假单胞菌(Pseudomonas fluorescens)在贮藏过程中比例逐渐增大,希瓦氏菌(Shewanella sp.)与弧菌(Vibrio sp.)比例也有明显增加,在贮藏后期发展成为优势腐败菌。