两株琼胶酶高产细菌的筛选和鉴定

从海洋环境中筛选得到两株琼胶酶的高产菌株，通过形态观察和生理生化反应，确定它们属于弧菌属，通过BIOLOG细菌鉴定系统鉴定，并同弧菌属标准菌株分析比较，确定上方宝剑两株菌都是塔式弧菌（Vibrio tubiashii)，这两株菌在碳源利用方面存在差别。     

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.     

北极海冰细菌产胞外酶及主要环境因子的初步研究

覆盖在北极海洋上的广阔海冰为其内部生存的微生物群落提供了一个独特的生境。研究表明 ,大量海冰细菌能够分泌产生胞外酶 ,其中产蛋白或脂质水解酶细菌的比例远高于产多糖水解酶的细菌。温度、盐度是直接影响海冰细菌生存与活力的 2个主要环境因子。 76%的产酪蛋白酶海冰细菌为低温菌 ,菌株只能在 <35°C条件下生长 ;而 98%的酪蛋白酶最适作用温度≥ 35°C ,其中 62 %的酶最适作用温度≥ 45°C。几乎所有的产酪蛋白酶海冰细菌都耐盐或嗜盐     

鞘藻属和毛鞘藻属营养细胞分裂方式的比较研究

于 1 999年 7月在湖北省仙桃市湿润土壤中采得一种鞘藻 ,经培养和电镜观察 ,发现该株鞘藻营养生长主要依赖位于丝状体最上面的细胞分裂 ,分生出的子细胞在该细胞亚顶端的胞壁破裂点掀开一个帽子 ,此帽子便附着于细胞侧壁上 ,这在鞘藻属中为首次报道 ,与 2 0 0 0年 1 2月在武汉纸坊青龙山水库采得的一种毛鞘藻的基细胞也附着相同的帽子结构。观察结果表明 ,该属基细胞亦具有明显的居间分裂能力。在观察这两属营养细胞居间分裂的同时 ,详细报道了这一种特殊的细胞分裂———顶端分裂 ,详细比较了这两属的营养细胞分裂方式 ,并对其分裂方式的机制进行探讨。结果表明 ,这两属细胞在细胞分裂方式上有极为亲密的亲缘关系 ,但毛鞘藻属更复杂、独特的营养细胞分裂显示它的进化地位比鞘藻属高。     

Extracellular enzymatic activities of cold-adapted bacteria from polar oceans and effect of temperature and salinity on cell growth

The potential of 324 bacteria isolated from different habitats in polar oceans to produce a variety of extracellular enzymatic activities at low temperature was investigated. By plate assay, lipase, protease, amylase, gelatinase, agarase, chitinase or cellulase were detected. Lipases were generally present by bacteria living in polar oceans. Protease-producing bacteria held the second highest proportion in culturable isolates. Strains producing amylase kept a relative stable proportion of around 30% in different polar marine habitats. All 50 Arctic sea-ice bacteria producing proteases were cold-adapted strains, however, only 20% were psychrophilic. 98% of them could grow at 3% NaCl, and 56% could grow without NaCl. On the other hand, 98% of these sea-ice bacteria produced extracellular proteases with optimum temperature at or higher than 35℃, well above the upper temperature limit of cell growth. Extracellular enzymes including amylase, agarase, cellulase and lipase released by bacteria from seawater or sediment in polar oceans, most expressed maximum activities between 25 and 35℃. Among extracellular enzymes released by bacterial strain BSw20308, protease expressed maximum activity at 40℃, higher than 35℃ of polysaccharide hydrolases and 25℃ of lipase.     

柄海鞘共附生细菌的分离培养与系统发育多样性研究

通过纯培养和16SrRNA基因序列的相似性比对,并构建系统发育树对威海海域柄海鞘共附生细菌的多样性进行了研究。用2216E培养基从柄海鞘样品中分离到78株细菌,通过形态学特征和TCBS培养基选择性筛选后,选取30株具有代表性的菌株进行了16SrRNA基因测序和基于16SrRNA基因序列的系统发育多样性分析。结果表明,30株菌株分别属于细菌域的4个系统发育类群(Actinobacteria,Bacteroidetes,Firmicutes,Gro-teobacteria)的11个科,11个属。根据16SrRNA基因序列,大多数菌株与其系统发育关系最密切的模式菌株之间存在一定的遗传差异(16SrRNA基因序列相似性为91.82%～98.93%)。其中菌株HQW7,HQYD1,HQWD4,HQE1和HQE2与相关模式菌株的16SrRNA基因最高相似度仅分别为91.82%,92.51%,92.99%,93.52%,93.87%,这5株菌可能代表新的分类单元。威海海域柄海鞘共附生细菌存在着较为丰富的物种多样性和系统发育多样性,并可能存在新的物种。     

南极菲尔德斯半岛表层土壤样品中细菌多样性的系统发育分析

通过PCR结合变性梯度凝胶电泳(DGGE)技术对从南极长城站附近表层土壤样品中获得16SrDNA序列特征片段V3区序列进行分离。对其中的主要12条DGGE条带进行胶回收,获得的DNA片段经测序以及计算机比对分析发现,它们分别属于β、γ、δ-变形细菌(Proteobac-teria)、噬纤维菌-屈挠杆菌-拟杆菌(Cytophaga-Flexibacter-Bacteroides,CFB)群细菌、放线细菌(Actinobacteria)、蓝细菌属(Cyanobacteria)、酸杆菌属(Acidobacteria)和绿屈挠菌属(Chlo-roflexi)等系统分类群。南极表层土壤样品中的大部分16S rDNA序列与从其他土壤或沉积物样品中直接获得的序列相似性较高(93%-100%)。     

弧菌拮抗菌的筛选

用平板划线或点种法对 6 0 2株海洋细菌进行筛选 ,得到 4株对海水养殖动物 (鱼、虾、贝 )的病原菌有拮抗作用的细菌。并对拮抗菌 QJ2的拮抗作用进行了研究 ,结果为 :QJ2有广泛的弧菌抗菌谱 ,对 37株弧菌的抗菌阳性率达到 89.2 % (33/ 37) ;用活菌平板计数法和 O.D.60 0 nm研究了QJ2培养物的去细胞上清液 (CFS)与病原菌 W- 1的作用动力学 ,15min时 W- 1的细菌数便开始减少 ,4 h时细菌数最少 ,6 h后开始增加 ,而对照组的细菌数呈逐渐上升趋势 ;QJ2对自身的拮抗物质不敏感 ;QJ2抗性物质的分子量不大于 80 0 0 Da;经常规生理生化方法和 API- 2 0细菌快速鉴定系统鉴定 ,QJ2为气味黄杆菌 (Flavobacterium odoratum)。     

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。     

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.