产低温超氧化物歧化酶南极菌株的筛选、鉴定及其部分酶学性质

从421株南极细菌中筛选出产超氧化物歧化酶南极细菌NJ062、NJ379、NJ522和NJ548。它们的最适生长温度在10-15℃之间,在0℃均能生长,属于嗜冷菌。4株南极嗜冷菌均含有一种类型的SOD同工酶。菌株NJ062的超氧化物歧化酶为Mn-SOD;菌株NJ379,NJ522和NJ548的超氧化物歧化酶均为Fe-SOD。它们所产的超氧化物歧化酶最适酶活温度大约在40-50℃,在0℃仍有较高的活性,为最高活性的20%-35%,热稳定性较差,属低温酶。通过16S rDNA序列对4株产低温超氧化物歧化酶的南极嗜冷菌NJ062、NJ379、NJ522和NJ548进行鉴定。结果表明,它们均属于细菌域(Bacteria)中的紫细菌类群(Proteobacteria)中的γ-亚群。其中菌株NJ062属于假交替单胞菌属(Pseudoalteromonassp.);NJ379属于希瓦氏菌属(Shewanellasp.);NJ522和NJ548属于海单胞菌属(Marinomonassp.)。     

北极斯瓦尔巴群岛冰川流域可培养细菌的系统发育多样性及固氮菌初步研究

从北极斯瓦尔巴群岛冰川流域的上、中、下游采集了5种代表性土壤和冰川融水样品(MT、AT、TN、BT、MS),采用常规LB和严格无氮培养基低温培养方法,对其中的可培养细菌和潜在固氮细菌进行了总菌计数以及菌株的富集、分离与纯化。经菌落菌体形态观察、16S r RNA基因初步测序排重、固氮基因nif H的PCR检测,分别选取48株可培养分离细菌和31株潜在固氮菌进行了基于16S r RNA基因全序列测定的系统发育分析。目的是初步了解该特殊生境中可培养细菌(包括潜在固氮菌)的多样性及其分布状况,为进一步研究它们在此生境中的生态功能奠定基础。实验结果表明,可培养细菌在5个采集样品中的总菌数为BTTNATMT=MS,严格无氮培养基分离菌总菌数为BTATMTTNMS。48株可培养代表菌分属于Proteobacteria、Actinobacteria、Bacteroidetes、Firmicutes 4个门的7纲、14属之中,它们在5个样品中的多样性程度为BT(6纲、6属)MT(4纲、6属)AT、MS(均为3纲、3属)TN(2纲、3属),优势菌群主要分布在γ-proteobacteria纲的Pseudomonas属(20.8%,集中在AT、BT、MS样品中)和β-proteobacteria纲的Janthinobacterium属(35.4%,集中在MS样品中),表现出一定程度的系统发育多样性。潜在固氮菌实验结果表明,能在严格无氮培养基上生长且可检测到nif H的31株代表菌具有极为单一的系统发育多样性。其中90.32%菌株集中在γ-proteobacteria纲的Pseudomonas属,仅在MT和MS样品中有3株菌分布在α-proteobacteria纲的Rhizobium属、β-proteobacteria纲的Janthinobacterium属和Actinobacteria纲的Arthrobacter属,它们与分离自其他极端环境的相关标准菌株具有高度16S r RNA基因同源性。另外,本实验还发现有8株菌的16S r RNA基因序列与相关标准菌株的相似度低于98.50%,推测它们具有成为该特殊生境下新种的潜质。     

Phylogenetic analysis of bacteria in sea ice brine sampled from the Canada Basin,Arctic Ocean

Bacterial diversity in sea ice brine samples which collected from four stations located at the Canada Basin,Arctic Ocean was analyzed by PCR-DGGE.T wenty-three 16S rDNA sequences of bacteria obtained from DGGE bands were cloned and sequenced.Phylogenetic analysis clustered these sequences within γ-prote obacteria,Cytophaga-Flexibacter-Bacteroides(CFB)group,Firmicutes and A ctinobacteria.The phylotype of Pseudoalteromonas in the γ-proteobacteria was predominant and members of the CFB group and γ-proteobacteria were highly abundant in studied sea ice brine samples.     

Screening and molecular classification of low-temperature protease from Antarctic microorganism and its characterization

<正> 107 strains producing protease were screened from 260 strains of Ant-arctic psychrophilic bacteria,among which proteolytic activity of five strains wasmore than 45 U ml~(-1).The 16S rRNA gene sequences homology and phylogenet-ic analysis of five Antarctic psychrophilic bacteria showed that NJ276、NJ5-9、NJ16-70、NJ345 belonged to the described genus Pseudoalteromonas and NJ341belonged to the genus Colwellia.The growth and the protease characteristic offour Antarctic psychrophilic bacteria had been studied,and the result showedthat the optimal temperature for growth and protease-producing of four strainswas about 10℃.Their growth and protease-producing were still high during in-cubating 2-5 days.The maximum proteolytic activity occurred at pH 9 for fourAntarctic psychrophilic bacteria.The optimal temperature of protease action ofboth strains NJ276 and NJ5-9 was about 50℃,however,the optimal temperatureof protease action of both strains NJ341 and NJ345 was about 40℃,and theirproteolytic activity under 0℃ exhibited nearly 30% of the maximum activity,but their thermal stabilities were weaker.These results indicated that proteasesfrom NJ341 and NJ345 were low-temperature proteases.     

南极酵母Rhodotorula sp.NJ298的鉴定及其产虾青素的研究

本研究对产虾青素的南极菌株NJ298进行了鉴定,同时对影响虾青素产量的主要环境因子进行了测试。形态特征、生理生化特征及ITS序列的系统发育分析表明,南极菌株NJ298属于红酵母属(Rhodotorula);虾青素积累主要在NJ298生长的对数期和稳定期;乙酸钠、蛋白胨是合成虾青素的最佳碳、氮源;在0-15℃之间,南极酵母NJ298均可以生长,但是低温更有利于虾青素的积累;pH值在6-8范围内NJ298均可生长,最适pH值为7.5;该菌对盐度要求不严格。     

古尔班通古特沙漠地衣结皮中可培养细菌多样性初探

为了解古尔班通古特沙漠优势结皮类型--地衣结皮中可培养细菌的多样性,利用3种培养基经稀释涂布和划线分离得到18株纯培养菌株。对分离的所有菌株提取基因组DNA,利用细菌通用引物扩增得到16S rDNA的序列并测序。结果表明：（1）基于Ribosomal Database Project (RDP)在线归类分析,18株菌可初步归为3门3纲3目6科6属,未见新属,未分离到固氮菌属（Azotobacter）。其中,6株归为变形菌门(Proteobacteria)、6株属于放线菌门(Actinobacteria)、6株属于厚壁菌门 (Firmicutes); 不动杆菌属（Acinetobacter）、链霉菌属（Streptomyces）、芽孢杆菌属（Bacillus）为优势属,分别占分离菌株数的33%、28%、22%。（2）在EzTaxon-e数据库中对各菌株序列进行进一步的BLAST同源性分析表明,18株菌可进一步归为14个OTUs（Operational Taxonomic Unit）,未见疑似新种（同源性均>97%）,其中链霉菌属OTU比例最高（35.71%）,依次为不动杆菌属和芽孢杆菌属（二者均为21.43%）。（3）古尔班通古特沙漠地衣结皮中的可培养细菌Shannon-Wiener多样性指数为2.5532,均匀度指数为0.9675。（4）上述可培养细菌在生物土壤结皮中具有固沙、加速矿化、生防等多重生态潜能。     

盐度对北极海冰细菌生长的影响

从中国第二次北极科学考察采集的海冰样品中分离到8种细菌,研究了盐度对其生长的影响。结果表明,8株海冰细菌对盐度有着广泛的适应性,在盐度为30-90的范围内均可较快生长,其中菌株BJ2、BJ3和BJ7的盐耐受性较强。进一步的研究表明,BJ2、BJ3和BJ7在0-150 的盐度范围内均有不同程度的生长,盐度为0-30时,菌株的生长最快;盐度为180时,这三株菌株也均有短暂的生长。随着盐度的增加,三株菌株生长的延迟期增加,到达生长稳定期的时间也相应增加。温度对菌株BJ2、BJ3和BJ7的盐耐受性也有着一定的影响,随着温度的升高, 菌株的生长速度变慢,但盐耐受性有所增加。     

内蒙古河套灌区湿地芦苇根际可培养细菌多样性

以内蒙古河套灌区湿地植物芦苇(Phragmites australis)根际土壤为材料,采用分离培养法对天然湿地和退化湿地的芦苇根际细菌多样性进行了研究。16SrRNA基因系统发育分析的结果显示,分离自天然湿地芦苇根际的45株细菌分属于γ-变形菌纲(44.44%)、厚壁菌门(Firmicutes)(42.22%)和放线菌门(Actinobacteria)(13.33%)3大类群,其中,γ-变形菌纲为最优势类群,芽孢杆菌属(Bacillus)(42.2%)为最优势菌属;分离自退化湿地芦苇根际的44株细菌分属γ-变形菌纲(13.63%)、β-变形菌纲(6.82%)、厚壁菌门(47.73%)及放线菌门(Actino-bacteria)(31.82%)4大类群,其中厚壁菌门为最优势类群,芽孢杆菌属(40.9%)为最优势菌属。研究结果表明,天然湿地和退化湿地芦苇根际细菌都具有较丰富的多样性,但天然湿地与退化湿地芦苇根际细菌群落结构存在一定差异。     

南极罗斯海沉积物中可培养菌株的分离及胞外水解酶活性检测

研究了南极罗斯海地区沉积物中可培养微生物多样性,并检测了菌株胞外酶活性,为开发利用南极资源奠定基础。采用Zobell2216E培养基、土豆培养基(PDA)及稀释平板划线法分离并获得纯培养的细菌和真菌,分别通过16SrDNA、18S/ITS序列鉴定分析了可培养微生物的多样性和系统发育。从南极罗斯海沉积物样品中获得78株细菌和54株真菌,细菌、真菌的优势菌株分别为嗜冷杆菌属(Psychrobacter)、枝孢属(Cladosporium);78株细菌菌株中有2株与模式菌株的16SrDNA基因序列相似性小于97%,可能是潜在的细菌新物种。细菌API20NE生理生化和真菌产胞外酶活性结果表明,获得的细菌和真菌大多数可检测到胞外水解酶活性。本研究丰富了南极可培养微生物资源,为南极罗斯海地区可培养微生物的多样性研究以及南极微生物资源的应用研究奠定了基础。     

一株产海藻糖合成酶南极海洋低温细菌的鉴定

从南极普里兹湾海域海水中分离到一株产海藻糖合成酶的海洋低温细菌BSw10041,革兰氏阴性,杆状,有极生鞭毛,能运动,菌落半透明。进行了常规生理生化和BIOLOG GN2细菌鉴定系统测试,结果表明菌株BSw10041与恶臭假单胞菌(Pseudomonas putida)的表型特征非常相似。为了进一步确定菌株BSw10041的分类学地位,测定了其16S rDNA序列,分析了相关细菌相应序列的同源性,构建了系统发育树,结果表明BSw10041与P．putida的亲缘关系最近。综合上述结果,菌株BSw10041可鉴定为Pseudomonas putida,定名为Pseudomonas putida BSw10041。     

北极海洋放线菌R-527F的抗菌活性筛选、鉴定及其抗菌物质发酵条件研究

对分离自北冰洋73°N以北海域沉积物中的一株放线菌R-527F进行分子鉴定及生物活性研究.通过对菌株的形态特征、培养性状、生理生化特征以及16S rDNA序列进行综合分析,确定该株菌为链霉菌属,暂定名为Streptomyces sp.R-527F.其含有聚酮类Ⅰ型(PKS Ⅰ)和非核糖体含硫多肽类(nonribosomal peptide synthetase,NRPS)化合物的合成基因.对其抗菌活性及活性物质发酵条件进行了研究,实验结果表明,该菌株所产生的抗菌物质对枯草芽孢杆菌具有显著抑制效果,抑菌圈直径达到18.2 mm.适合其抗菌物质的发酵条件:种子液接种量为4％,培养基初始pH为8.0,培养温度为28℃,培养时间为6d,摇床转速为180 r·min-1,装液量为40％,为该菌株抗菌物质的进一步研究奠定发酵条件参数基础.     

北极Ny-Ålesund地区真菌多样性分析及龙胆苦苷转化菌株的筛选

采用多种培养基对北极高纬地区（土壤及海洋沉积物）真菌进行分离纯化,共得真菌65株。通过分析菌株的18S rDNA序列研究了它们的系统发育多样性,18S rDNA序列分析表明,65株真菌分属25个属,其中数量上位居第一的是青霉属（Penicillium）,有菌株20株,其次为丛赤壳属（Nectria）和节枝孢属（Articulospora）,各有菌株6株。同时对这65株真菌开展转化龙胆苦苷阳性菌株的筛选。以β-葡萄糖苷酶作为筛选转化菌株的标志酶于65株真菌中筛选到阳性菌株28株,其中产酶能力强的9株菌作为转化实验的备选菌株,以龙胆苦苷为底物进行转化,将转化粗提物进行TLC及HPLC分析,最终确定将C-5作为转化龙胆苦苷的菌株。根据菌株形态特征和核酸序列分析结果,初步鉴定菌株C-5为青霉属真菌。暂定名为Penicillium sp.C-5。     

Screening on DHA-producing Antarctic bacteria N-6 and its cultural conditions

In Antarctic, the geography and climate differs from those in other places, and the bacteria there have adapted well to the environment there. Two hundred strains of bacteria were isolated from the sea ice in Antarctica. The bacteria were screened for DHA by means of GC, with fish oil as the standard. Seven strains containing DHA or EPA were obtained, among which the strain of No. N-6 was outstanding. And the component of DHA was identified by GC-MS. The relative content of DHA in N-6 was 8.72%, and total lipids in dry bacteria was 22.54%. The effects of some factors, including temperature, salinity and pH value, on the growth and DHA-content of strain N-6 were studied. The results show that the DHA-content is relatively high when in low temperature and high pH, and the bacterium is psychrophilic, alkalophilic.     

北极海洋放线菌R-527F的抗菌活性筛选、鉴定及其抗菌物质合成条件研究

对分离自北冰洋７３°Ｎ以北海域沉积物中的一株放线菌Ｒ５２７Ｆ进行分子鉴定及生物活性研究。通过对菌株的形态特征、培养性状、生理生化特征以及１６ＳｒＤＮＡ序列进行综合分析,确定该株菌为链霉菌属,暂定名为Ｓｔｒｅｐｔｏｍｙｃｅｓ ｓｐ．Ｒ５２７Ｆ。其含有聚酮类Ⅰ型（ＰＫＳⅠ）和非核糖体含硫多肽类（ｎｏｎｒｉｂｏｓｏｍａｌｐｅｐｔｉｄｅｓｙｎｔｈｅｔａｓｅ,ＮＲＰＳ）化合物的合成基因。对其抗菌活性及活性物质发酵条件进行了研究,实验结果表明,该菌株所产生的抗菌物质对枯草芽孢杆菌具有显著抑制效果,抑菌圈直径达到１８．２ｍｍ。适合其抗菌物质的发酵条件：种子液接种量为４％,培养基初始ｐＨ为８．０,培养温度为２８℃,培养时间为６ｄ,摇床转速为１８０ｒ?ｍｉｎ－１,装液量为４０％,为该菌株抗菌物质的进一步研究奠定发酵条件参数基础。     

Isolation of novel psychrophilic bacteria from Arctic sea ice

The phylogenetic diversity of culturable psychrophilic bacteria associ ated with sea ice from the high latitude regions of Canadian Basin and Chukchi S ea,Arctic,was investigated.A total of 34 psychropilic strains were isolated u sing three methods of(Ⅰ)dilution plating(at 4 ℃),(Ⅱ)bath culturing(at-1 ℃)and dilution plating,and(Ⅲ)cold shock(-20 ℃ for 24 h),bath culturin g and dilution plating under aerobic conditions.Sea-ice samples were exposed to-20 ℃ for 24 h that might reduce the number of common microorganisms and encou rag e outgrowth of psychrophilic strains.This process might be able to be introduce d to isolation psychrophilic bacteria from other environmental samples in future study.16S rDNA nearly full-length sequence analysis revealed that psychrophil i c strains felled in two phylogenetic divisions,γ-proteobacteria(in the gen era Colwellia、Marinobacter、Shewanella、Glaciecola、Marinomonas and Pseudoalt eromon as) and Cytophaga-Flexibacter-Bacteroides(Flavobacterium and Psychrof lexus).Fi fteen of bacterial isolates quite likely represented novel species(16S rDNA seq uence similarity below 98%).One of strains(BSi20002)from Canadian Basin showe d 100% sequence similarity to that of Marinobacter sp.ANT8277 isolated from the Antarctic Weddell sea ice,suggesting bacteria may have a bipolar distribution at the species level.     

吉尔吉斯斯坦西南部两栖爬行动物物种多样性

为了进一步了解吉尔吉斯斯坦两栖爬行动物物种多样性特征,于2012年9月15日～26日,在对吉尔吉斯斯坦西南部不同生境两栖爬行动物物种多样性进行野外实地调查和已有文献整理的基础上,初步整理出了分布在吉尔吉斯斯坦西南部的两栖爬行动物种类,并其物种多样性进行比较分析。本次考察共采集和记录实体标本621号,经鉴定共26种（亚种）,分隶4目10科18属,其中两栖纲1目2科2属2种,爬行纲3目8科16属22种。其中新疆蟾蜍（Bufo pewzowi）、亚洲林蛙（Rana asiatica）、赛氏沙蜥（Phrynocephalus saidalievi）三个种为优势种类,占总个体数的55.23%;快步麻蜥（Eremias velox）、喜山岩蜥（Laudakia himalayana）、草原蜥（Trapelus agilis）、荒漠无脸蜥（Ablepharus deserti）、施氏石龙子（Eumeces schneideri）、叶城麻蜥（Eremias yarkandensis）、棋斑游蛇（Natrix tessellata）、白条锦蛇（Elaphe dione）、中亚眼镜蛇（Naja oxiana）、四爪陆龟指名亚种（Testudo horsfieldii horsfieldii）11种类为常见种,占总个体数的36.88%;新疆岩蜥（Laudakia stoliczkana）、敏麻蜥（Eremias arguta）、捷蜥蜴（Lacerta agilis）、红沙蠎（Eryx miliaris）、东方沙蠎（Eryx tataricus）、草原蝰（Vipera renardi）等12各种为稀有种类,占总个体数的7.89%。在不同生境间均存在物种多样性指标差异显著（p<0.05）,物种多样性在草原最高（2.029 8）,其次为山地森林（1.920 4）,而荒漠最低（0.233 8）;丰富度均在山地森林最高（3.242 51）,其次为草原（2.540 0）,而在荒漠最低（0.288 5）;由于吉尔吉斯斯坦西南部的气候属于典型的大陆性气候,两栖爬行动物物种的组成主要以适应干旱环境的耐旱种类为主,区系组成较为简单,地方种类数量较少,在物种组成及起源等方面与我国西北地区的两栖爬行动物区系有着一定的联系。     

Effects of Elevated Atmospheric CO(2) on Rhizosphere Soil Microbial Communities in a Mojave Desert Ecosystem

The effects of elevated atmospheric carbon dioxide [CO₂] on microbial communities in arid rhizosphere soils beneath Larrea tridentata were examined. Roots of Larrea were harvested from plots fumigated with elevated or ambient levels of [CO₂] using Free-Air CO₂ Enrichment (FACE) technology. Twelve bacterial and fungal rRNA gene libraries were constructed, sequenced and categorized into operational taxonomical units (OTUs). There was a significant decrease in OTUs within the Firmicutes (bacteria) in elevated [CO₂], and increase in Basiomycota (fungi) in rhizosphere soils of plots exposed to ambient [CO₂]. Phylogenetic analyses indicated that OTUs belonged to a wide range of bacterial and fungal taxa. To further study changes in bacterial communities, Quantitative Polymerase Chain Reaction (QPCR) was used to quantify populations of bacteria in rhizosphere soil. The concentration of total bacteria 16S rDNA was similar in conditions of enriched and ambient [CO₂]. However, QPCR of Gram-positive microorganisms showed a 43% decrease in the population in elevated [CO₂]. The decrease in representation of Gram positives and the similar values for total bacterial DNA suggest that the representation of other bacterial taxa was promoted by elevated [CO₂]. These results indicate that elevated [CO₂] changes structure and representation of microorganisms associated with roots of desert plants.     

Isolation,identification and characterization of highly tellurite-resistant,tellurite-reducing bacteria from Antarctica

The tellurium oxyanion, tellurite, is extremely noxious to most living organisms. Its toxicity has been mainly related to the generation of reactive oxygen species (ROS) as well as to an unbalancing of the thiol:redox buffering system. Nevertheless, a few bacteria are capable of thriving at high tellurite concentrations. One mechanism of resistance is the enzymatic and non-enzymatic reduction of tellurite to the less toxic elemental tellurium. This reduction generates nano- to micrometric tellurium crystals that display different shapes and sizes.To date, a very limited number of highly tellurite-resistant and tellurite-reducing bacterial species are available from international culture collections. In this work, we decided to look for tellurite-reducing bacteria from an extreme environment, Antarctica. This environment exhibits a combination of several extreme factors such as high UV-radiation and desiccation and freezing conditions that impact directly on the local biodiversity. Since, as does, all these factors induce ROS formation, we hypothesized that Antarctic bacteria could also exhibit tellurite-resistance. In this context, we isolated 123 tellurite-resistant bacteria, and characterized six new tellurite-resistant and tellurite-reducing bacterial strains from samples collected in Antarctica. These strains were identified according to their 16S rRNA gene sequence as Staphylococcus hameolyticus, Staphylococcus sciuri, Acinetobacter haemolyticus, Pseudomonas lini, and two strains of Psychrobacter immobilis.The isolates display tellurite-resistance about 35- to 500-fold higher than Escherichia coli (Te-sensitive organism), and a high level of tellurite reduction which might be interesting for an application in the field of bioremediation or nanoparticle biosynthesis.     

白令海北部表层沉积物中细菌多样性的研究

提取白令海表层沉积物样品总DNA,构建沉积物中的细菌16SrDNA文库,通过RFLP酶切分型与序列测定,进行沉积物中的细菌多样性及系统发育分析。结果表明,该沉积物中细菌多样性丰富,获得的125条序列归属于10个细菌类群,包括占据优势地位的变形细菌(Pro-teobacteria,49.6%)、酸杆菌门(Acidobacteria,20.8%),以及拟杆菌门(Bacteroidetes,6.4%)、放线菌门(Actinobacteria,5.6%)、浮霉菌门(Planctomycetes,5.6%)、硝化螺旋菌门(Nitrospirae,4%)、疣微菌门(Verrucomicrobia,2.4%)、绿弯菌门(Chloroflexi,1.6%)、纤维杆菌门(Fibrobacteres,0.8%)、螺旋体(Spirochaetes,0.8%)。此外,还有一部分分类地位尚不明确的细菌(2.4%)。而在变形细菌中,δ、γ亚群为其中的优势类群。     

南极长城站空气微生物的分子生态学分析

微生物是气溶胶的重要组成部分。本研究通过收集南极长城站附近气溶胶,构建气溶胶中空气微生物16S rRNA基因文库,发现南极空气微生物具有很高的多样性,93个克隆子包含了53个OTU(Operational Taxonomic Unit),Shannon指数为3.58,Simpson指数为0.04。南极长城站空气微生物的主要细菌类群为Actinobacteria、Cyanobacteria/Chloroplasts、Bacteroidetes、Firmicutes、Planctomycetes、Proteobacteria及Verrucomicrobia等7个门,Proteobacteria门的细菌是优势类群,占长城站空气微生物细菌克隆文库的65.7%。克隆文库中海源微生物序列及陆源微生物序列各占30.1%,说明长城站附近具有较强的海陆大气交换。我们发现有15个克隆子序列与人类活动相关,占总克隆子数的16.1%,说明人类活动已经影响到了长城站的空气微生物群落结构。