云南洱源牛街热泉原核微生物多样性分析

【目的】通过分析富含高分子有机物的云南洱源牛街热泉原核微生物16SrRNA基因克隆文库,丰富对高温热泉原核微生物多样性的认识,为进一步开发和利用该热泉微生物资源奠定基础。【方法】构建洱源牛街高温热泉原核微生物16SrRNA基因克隆文库,通过测序和序列相似性比对以及聚类分析研究该热泉原核微生物的多样性。【结果】该热泉原核微生物以细菌为主,包括变形菌门(Proteobacteria)、厚壁菌门(Firmicutes)、梭杆菌门(Fusobacteria)等在内的约10个细菌类群,其中变形菌门中的β-变形菌纲(β-Proteobacteria)为优势菌群,其次为拟杆菌门(Bacteroidetes)、绿菌门(Chlorobi);古菌的生物量和丰度较细菌少,分属广古菌(Euryarchaeota)和泉古菌(Crenarchaeota)两个类群,以广古菌为优势类群。     

新疆沙湾冷泉沉积物中免培养古菌多样性初步研究

【目的】了解新疆沙湾冷泉沉积物的古菌组成及多样性。【方法】采用免培养法,液氮研磨提取冷泉沉积物总DNA,使用古菌通用引物进行16S rRNA基因扩增,构建16S rRNA基因文库。对阳性克隆进行HhaI限制性酶切分型,选出具有不同酶切图谱的序列进行测序,将所得序列与GenBank数据库中序列比对并构建16S rRNA基因系统发育树。【结果】从冷泉沉积物古菌16S rRNA基因文库中随机挑选了121个阳性克隆,共得到22个不同的可操作分类单元,BLAST结果表明全部克隆子归属于泉古菌门(Crenarchaeote)中免培养类群。系统发育分析归类为Soil-Freshwater-subsurface group和MarinegroupI,2个亚群并且各占整个文库的50%。其中40%左右的克隆子与具有无机碳和硝酸盐同化能力的泉古菌有高的相似性。此外还发现40%的克隆子与低温泉古菌类群具有很高的相似性。【结论】新疆沙湾冷泉沉积物中古菌类群多样性较低,但存有大量高度适应此低温、贫营养环境的泉古菌类群。     

Bacterial,archaeal and eukaryotic diversity in Arctic sediment as revealed by 16S rRNA and 18S rRNA gene clone libraries analysis

We studied the microbial diversity in the sediment from the Kongsfjorden, Svalbard, Arctic, in the summer of 2005 based on the analysis of 16S rRNA and 18S rRNA gene clone libraries. The sequences of the cloned 16S rRNA and 18S rRNA gene inserts were used to determine the species identity or closest relatives by comparison with sequences of known species. Compared to the other samples acquired in Arctic and Antarctic, which are different from that of ours, the microbial diversity in our sediment is much higher. The bacterial sequences were grouped into 11 major lineages of the domain Bacteria: Proteobacteria (include α-, β-, γ-, δ-, and ε-Proteobacteria); Bacteroidetes; Fusobacteria; Firmicutes; Chloroflexi; Chlamydiae; Acidobacteria; Actinobacteria; Planctomycetes; Verrucomicrobiae and Lentisphaerae. Crenarchaeota were dominant in the archaeal clones containing inserts. In addition, six groups from eukaryotes including Cercozoa, Fungi, Telonema, Stramenopiles, Alveolata, and Metazoa were identified. Remarkably, the novel group Lentisphaerae was reported in Arctic sediment at the first time. Our study suggested that Arctic sediment as a unique habitat may contain substantial microbial diversity and novel species will be discovered.     

Bacterial diversity in deep-sea sediment from northeastern Pacific Ocean

16S rDNA sequencing method is one of the effectively used culture-independent techniques in recent years. In this study, 16S rDNA sequencing method was used to investigate the bacterial diversity in deep-sea sediment from northeastern Pacific polymetallic nodule province. Total DNAs were extracted by using 2 different methods (chemical method and DNA extracting kit method). After purification, genomic DNA was amplified by using 2 universal primers (27F and 1492R). Clones were selected and sequenced randomly. After the sequences were checked by using the Chimera Check Program of the RDP database, a bacterial 16S rRNA gene library of 79 clones was established. Phylogenetic analysis indicated that 79 clones could be divided into 11 phylotypes. Gamma Proteobacteria (22.8%) and alpha Proteobacteria (16.5%) were the dominant components of the sediment bacterial community, followed by Planctomycetacia (7.6%), delta Proteobacteria (6.3%), Nitrospira (6.3%), Actinobacteria (6.3%), beta Proteobacteria (5%), Acidobacteria (5.1%), Sphingobacteria (3.8%), Firmicutes (2.5%) and uncultured bacteria (17.7%). Gamma Proteobacteria also dominated at slices 0–2 cm and 4–6 cm. Different slices had different types of bacteria, alpha Proteobacteria, gamma Proteobacteria, delta Proteobacteria, Planctomycetacia, Nitrospira, Actinobacteria and Acidobacteria, however, appeared in all slices. Pseudomonas is common in many different deep-sea environments. In this study, it accounted for 22.2% of the total gamma Proteobacteria.     

新疆沙湾冷泉沉积物的细菌系统发育多样性

为了解新疆沙湾冷泉沉积物的细菌群落组成与类群多样性,利用免培养方法直接从沙湾冷泉沉积物中提取环境总DNA,构建细菌16S rRNA基因文库。对随机挑选的241个细菌阳性克隆子进行HaeIII酶切分型得到86个可操作分类单元(OTUs),系统发育分析将其归为11个门:放线菌门(Actinobacteria),酸杆菌门(Acidobacteria),拟杆菌门(Bacteroidetes),绿菌门(Chlorobi),蓝细菌门(Cyanobacteria),厚壁菌门(Firmicutes),芽单胞菌门(Gemmatimonadetes),硝化螺旋菌门(Nitrospirae),变形菌门(Proteobacteria),浮霉菌门(Planctomycetes),疣微菌门(Verrucomicrobia)。其中酸杆菌门和变型菌门为优势类群,分别占细菌克隆文库的48%和25%。超过1/3的OTUs序列与GenBank中已存序列具有较低相似性(相似性小于95%)。此外20%左右的克隆子与固氮细菌和硝酸盐氧化细菌相关。研究结果表明,新疆沙湾冷泉沉积物中细菌种类丰富,代谢类型多样而且存在大量未知类群。     

山西左云县采煤区人工湿地冬季沉积物细菌群落多样性

【背景】人工湿地处理煤炭矿井废水效果显著,而关于人工湿地沉积物的细菌群落结构和特征研究较少。【目的】阐明人工湿地净化采煤区废水的效应和细菌群落结构及特征。【方法】对山西省左云县采煤区人工湿地不同采样点水质进行监测,利用高通量测序技术对人工湿地沉积物中细菌16S rRNA基因V3-V4区进行测序,分析沉积物中细菌群落组成。【结果】采煤区废水流经人工湿地后水质得到有效改善,其中生化需氧量(Biochemical oxygen demand,BOD5)、化学需氧量(Chemical oxygen demand,CODCr)、总氮(Total nitrogen,TN)、总磷(Total phosphorus,TP)去除率分别达到76.2%、93.4%、73.4%和99.3%;测序共获得2 832个操作分类单元(Operational taxonomic units,OTUs),共753个属,分布于51个门的150个纲;4个采样点中优势菌门为变形菌门(Proteobacteria,50%-64.7%)和拟杆菌门(Bacteroidetes,15.8%-21.2%),变形菌门中优势菌纲为β-变形菌纲和γ-变形菌纲,其中M1、M2、M3中富含氨氮氧化细菌的β-变形菌纲和γ-变形菌纲丰度最高;M4沉积物细菌OTU最多,多样性最高;沉积物特有优势菌属Sulfurimonas、Sulfuricurvum的相对丰度与NH4+-N的含量显著负相关,Azoarcus菌属相对丰度与TN存在显著正相关,Novosphingobium菌属相对丰度与TP含量存在显著正相关。【结论】煤矿废水流经人工湿地可得到有效的净化,细菌多样性在人工湿地生态功能中起到了决定作用。     

东北太平洋深海沉积物细菌多样性

采用两种方法提取中国结核合同区东区沉积物不同层次总DNA,通过克隆测序构建了含有79个克隆子的细菌16S rRNA基因文库,分析了该海域沉积物中细菌的多样性.79个克隆在系统发育树中形成了11个大分支,包括Gamma proteobacteria(22.8%),Alpha proteobacteria(16.5%),Planctomycetacia(7.6%),Delta proteobacteria(6.3%), Nitrospira(6.3%),Actinobacteria(6.3%),Beta proteobacteria(5%),Acidobacteria(5.1%),Sphingobacteria(3.8%),Firmicutes(2.5%),Other bacteria(17.7%),其中Gamma proteobacteria在总文库中所占比例最高,该分支细菌在0～2cm、4～6cm层也是优势菌种.Gamma proteobacteria中假单胞菌(Pseudomonas)为优势属(22.2%).各个层次中所含细菌类群有所不同,Alpha proteobacteria、Gamma proteobacteria、Delta proteobacteria 、Planctomycetacia、Nitrospira 、Actinobacteria和Acidobacteria为三层样品共有类群.     

Characterization of the depth-related changes in the microbial communities in Lake Hovsgol sediment by 16S rRNA gene-based approaches

The undisturbed sediment of Lake Hovsgol (Mongolia) is scientifically important because it represents a record of the environmental changes that took place between the Holocene (the present age) and Pleistocene (the last ice age; 12,000 14C years before present day). Here, we investigated how the current microbial communities change as the depth increases by PCR-denaturing gradient gel electrophoresis (DGGE) analysis of the 16S rRNA genes of the microbial communities. The microbial diversity, as estimated by the Shannon index, decreased as the depth increased. In particular, significant changes in archaeal diversity were observed in the middle depth (at 39-42 cm depth of total 60 cm depth) that marks the border between the Holocene and Pleistocene. Phylotype belonging to Beta-and Gamma-Proteobacteria were the predominant bacteria and most of these persisted throughout the depth examined. However, as the depth increased, some bacteria (some genera belonging to Beta-Proteobacteria, Nitrospira, and OP8-9) were not detectable while others (some genera belonging to Alpha-, Beta-, Gamma-Proteobacteria) newly detected by DGGE. Crenarchaea were the predominant archaea and only one phylotype belonging to Euryarchaea was found. Both the archaeal and bacterial profiles revealed by the DGGE band patterns could be grouped into four and three subsets, respectively, subsets that were largely divided by the border between the Holocene and Pleistocene. Thus, the diversity of the current microbial communities in Lake Hovsgol sediments decreases with increasing depth. These changes probably relate to the environmental conditions in the sediments, which were shaped by the paleoclimatic events taking place between the Holocene and Pleistocene.     

新疆冷泉沉积物葡萄糖利用细菌群落多样性的稳定同位素标记分析

新疆沙湾冷泉低盐且不含硫化物,为了解其沉积物细菌群落的碳源利用,以13C稳定同位素标记的葡萄糖作为底物培养沉积物中的细菌,通过提取和分离带有13C标记的总DNA,结合16S rDNA-PCR克隆文库法以及限制性片段长度多态性方法,对冷泉沉积物中葡萄糖利用细菌群落多样性进行分析.随机挑取417个阳性克隆,HaeⅢ酶切分型,共获得27个OTUs.经测序、序列同源性对比和系统发育学分析,归为细菌域中的9个门,即厚壁菌门(Firmicutes)、绿弯菌门(Chloroflexi)、硝化螺旋菌门(Nitrospirae)、放线菌门(Actinobacteria)、变形杆菌门(Proteobacteria)、拟杆菌门(Bacteroidetes)、浮霉菌门(Planctomycetes)、蓝细菌门(Cyanobacteria)和酸杆菌门(Acidobacteria),其中,厚壁菌门和变形杆菌门为优势类群,分别占克隆文库的28.3％和38.6％.与前期研究比较,以葡萄糖为碳源的细菌OTUs仅占总菌群的31％,表明该环境中可能存在其它碳源利用方式的细菌群落.     

高通量测序分析云南腾冲热海热泉真菌多样性

【目的】本研究揭示云南腾冲热海热泉真菌多样性及群落分布格局,探讨其理化因子对真菌群落结构的影响。【方法】利用Illumina Hi Seq2500高通量测序平台对腾冲热海热泉沉积物宏基因组ITS基因进行测序,并进行生物信息分析。【结果】从5个热泉样品中共检测到343484条有效序列,包括5个真菌门,20个纲,66个目。右姐妹泉(JMQR)、左姐妹泉(JMQL)、蛤蟆嘴泉(M)、桥泉(QQ)及鼓明泉(GMQP)分别以Agaricales、Eurotiales、Capnodiales和Hypocreales等为优势目。在属水平上,共获得365个属,从JMQR中检测到212个属,以裂褶菌属(Schizophyllum)为最优势;从JMQL中挖掘到197个属,以青霉属(Penicillium)为最优势类群;从M和QQ中分别获得222个和270个属,均以枝孢属(Cladosporium)为最优势;从GMQP中发现179个属,以侧齿霉属(Engyodontium)丰度最高。NH4+含量、温度及pH影响不同优势真菌的分布,其中以pH与优势类群(OTU1%)结构变化显著性最高(P=0.05)。【结论】云南腾冲热海高温热泉蕴藏着极其丰富的真菌物种,其不同样品真菌分布具有差异性,pH可能是影响热泉真菌群落分布的重要因素之一。