耐盐异养硝化菌驯化方法及分离菌株鉴定

采用高C/N、逐渐降低DO、菌体对数生长期前期转接以及增加培养液中海水比例的方法完成了耐盐异养硝化菌的富集和驯化,并从中分离筛选出两株高效异养硝化菌qy37和gs2.通过对两株菌的形态观察、生理生化试验以及16S rRNA序列分析,确定菌株qy37和菌株gs2分别为假单胞菌属(Pseudomonas)和盐单胞菌属(Halomonas).     

植物内生菌与宿主关系研究进展

植物内生菌（endophyte）是指存活于健康植物组织内部,而又不引发宿主植物表现出明显感染症状的微生物类群,主要包括真菌、细菌和放线菌。其在植物体中的分布具有普遍性、多样性的特点。在目前研究过的所有植物中均发现有内生菌,它们可存在于植物的根、茎、叶、花、果实等各个部位。研究发现内生菌除了随植物遗传进行传播外,还可通过多种途径对宿主植物进行侵染,比如可以以内生菌体或孢子形式通过变形、吸器、或渗透等途径侵染植物,也可以通过分解植物表皮细胞壁或通过各种自然开口（包括侧根发生处,气孔,水孔等）或伤口（包括土壤对根的磨损,病虫对植物的损害及收割多年生植物造成的伤口等）等传播途经进入植物。内生菌由于与宿主植物长期共处,进而形成了一种复杂、特殊的关系。它们有的是互利共生关系,而有的是无害或微害寄生关系。两种关系可随多种因素变化而相互转化。本文主要从内生菌在宿主植物中的分布特点、侵染特性及与宿主植物共处方式等方面,对近十年来植物内生菌与宿主关系的研究进展进行综述和展望,以期为植物内生菌资源开发研究提供参考。     

贵州浓香型白酒大曲中霉菌的18S rDNA系统发育分析

从贵州某名酒厂大曲中分离到104株丝状真菌(霉菌),通过形态特征和生理特性检测等传统鉴定方法,初步把形态和生理生化特征一致的霉菌归类为14个大类群;并采用18S rDNA序列分析,进一步把菌株归类为MJ-I至MJ-V五个类群,随后构建系统发育树.结果表明,样品中的霉菌以曲霉属(Aspergillus)为主,占总分离菌数...     

华重楼内生菌Iun35的分离及其抗菌蛋白的性质

从华重楼(Paris polyphylla Smith var.chinensis)块茎中分离纯化得到一株具有较强抑菌活性的内生细菌Iun35,经形态特征、生理生化特性及16S rDNA鉴定为枯草芽孢杆菌(Bacillus subtilis).将分离得到的Iun35菌株发酵培养,培养液上清经硫酸铵分级沉淀、Sephadex G-75凝胶柱层析和DEAE-32纤维素柱层析分离纯化出一种抗菌蛋白UD35.用该蛋白对多种植物病原菌进行拮抗测定,结果显示,UD35对玉米纹枯病菌(Rhizoctonia solani Kuha)、小麦赤霉病菌[Gibberelle zeae(Schw.)]等多种菌有强烈抑制作用.测定其相对分子质量约为55 000,氨基酸分析结果表明其由17种氨基酸组成.图3表4参16     

紫花苜蓿根际氢氧化细菌的分离与鉴定

利用气体循环培养体系从陕西乾县HUP-豆科植物紫花苜蓿(Medicago sativa)根际土壤中分离获得37株细菌.菌株氧化氢能力测定结果表明,8株菌氧化氢和自养生长能力较强,初步确定为氢氧化细菌类群;根据其形态特征、培养特征和生理生化特性,鉴定为7个不同属:假单胞菌属(Pseudomonas)、邻单胞菌属(Plesiomonas)、脂肪杆菌属(Pimelobacter)、黄色杆菌属(Xanthobacter)、勒米诺氏菌属(Leminorella)、地杆菌属(Terrabacter)和稀有杆菌属(Rarobacter);其中氧化氢能力最强的优势菌株WMQ-7 16S rDNA序列(GenBank登录号为EU807744)长度为1 451bp,GC含量为53.8%,其核苷酸序列与假单胞菌属同源性高于99%,在系统发育树上位于同一分支,将WMQ-7菌株鉴定为假单胞菌属(Pseudomonas).图2表5参16     

广西典型高砷区蜈蚣草根围丛枝菌根真菌多样性研究

丛枝菌根真菌(arbuscular mycorrhizal fungi,AM真菌)在提高超富集植物蜈蚣草(Pteris vittata L.)修复砷污染土壤方面具有巨大的应用潜力,但是中国砷污染地区的AM真菌的分布和资源调查还不够全面。为探明砷污染地区AM真菌的多样性状况,采用野外调查和室内分析相结合的方法,通过形态学特征分离鉴定AM真菌种属,对广西南丹县高砷矿区及周边地区蜈蚣草根围土壤中AM真菌的多样性及群落组成进行调查研究。结果表明,不同程度砷污染土壤中生长的蜈蚣草均能被AM真菌侵染,随着砷浓度的增加物种多样性总体呈下降趋势。无污染土壤中的AM真菌侵染率和孢子密度主要受土壤磷和海拔的影响。低磷促进孢子萌发和菌丝生长,从而提高侵染率,而海拔升高会导致孢子密度降低。但随着土壤砷污染水平的增加,砷浓度逐渐成为主要影响因子,使侵染率上升,孢子密度下降。孢子密度和物种丰富的变化趋势一致。鉴定出4属10种AM真菌,其中7种属于球囊霉属(Glomus),近明囊霉属(Claroideoglomus)、管柄囊霉属(Funneliformis)和盾巨孢囊霉属(Scutellospora)各1种,球囊霉属是该调查区域的优势属。AM真菌孢子的形态直接影响其适应能力,颜色深、表面网纹间隔致密的AM真菌抗逆性更好,近明球囊霉(C.claroidem)、团集球囊霉(G.glomerulatum)和地表球囊霉(G.versiforme)等适应性较强的菌种分离频度达到了100%,相对丰度也较高。研究结果将会对砷污染地区AM真菌优良菌株的筛选及超富集植物-AM真菌联合修复重金属污染土壤提供有益的指导。     

青海湖嗜盐菌的分离与优势菌株QHL5的特性

采用高盐选择性培养基从20份青海湖水样中筛选分离获得35株青海湖嗜盐微生物.耐盐梯度实验分析表明:水体中以中度嗜盐菌为主,约占65.7%;弱嗜盐菌次之,约占25.7%;非嗜盐菌与耐盐菌相对较少,约占8.5%.其中优势菌QHL5的生长盐度耐受范围为0.04～3.0 mol/L,最适生长范围为0.3～0.9 mol/L,隶属于中度嗜盐菌.菌体形态和生长特性分析表明:QHL5菌落大小为4～6 mm,圆形或椭圆形,边缘整齐厚实,乳白色,中间微隆起,不透明,革兰氏染色阴性.显微形态呈杆状,最适生长pH为8.5～9.5,最适生长温度为35℃.生理生化、16S rRNA的全序列Blast和系统发育分析表明:QHL5菌株分类定位于γ变形菌纲海洋螺菌目盐单胞菌属樊氏盐单胞菌(Halomonas ventosae).通过80%乙醇法抽提胞内相溶物质,采用TLC法检测出该菌积聚四氢嘧啶.     

一株Ⅱ型甲烷氧化菌的筛选及培养条件

甲烷氧化菌能够以甲烷作为唯一碳源和能源物质生存.在甲烷氧化、氯代烃类污染物降解、相关化学品生产等方面具有重要的潜力.利用甲烷作为唯一碳源物质筛选分离得到一株甲烷氧化菌,并对其进行初步鉴定.在此基础上,利用单因子实验和Plackett_Burman实验等方法对获得菌株的培养条件:包括培养基条件(无机氮源种类和浓度、影响显著的金属离子及其浓度、p H值)以及培养温度进行优化.结果表明:经过16S r DNA鉴定,获得菌株与Methylocystis sp.SC2、Methylocystis hiersuta strain SV97等菌株的相似性达到99%,所以确定该菌株属于Methylocystis菌属(Ⅱ型甲烷氧化菌,甲基孢囊菌属),将本菌株命名为Methylocystis sp.M16.以1.0 g/L NH4NO3作氮源,1.0μmol/L铜离子,p H 7.00,温度30℃的条件下,甲烷去除率和菌体生物量最大.在各个因素的最优条件下,Methylocystis sp.M16菌液吸光值(A600 nm)均在0.5以上,甲烷去除率在95%以上.本研究分离得到一株Ⅱ型甲烷氧化菌Methylocystis sp.M16,并获得优化的培养基和培养条件,有望为M16菌应用提供理论参考.     

一株假芝的分离鉴定与生物学特性

灵芝是我国著名传统药用真菌,假芝属(Amauroderma)真菌作为灵芝科重要属,我国已报道种共有22个.本研究从广州白云山采集到一株野生假芝子实体,经组织分离获得菌丝体纯培养,编号GZ-01菌株.利用形态鉴定和ITS鉴定手段,确定其分类学地位为假芝属,系统命名为A.subrugosum,为中国首次记录种,菌种保藏号ACCC52954.生物学特性研究表明,菌丝生长的最适碳源为甘露醇,最适氮源为酵母浸膏或大豆蛋白胨,最适C/N比为20/1,最适生长因子为维生素B6,最适生长温度为28℃,最适p H值范围为6.0-7.5.本研究获得新的药用真菌资源,其培养条件研究可为进一步开发利用奠定实验基础.     

青岛东风盐场中极端嗜盐古细菌的特性

从青岛东风盐场分离到3株嗜盐古细菌（编号：ZDA－1,ZDA－2和ZDA－5）,细胞为多形态杆状（0．7～15μm×2．0～50μm）,最适生长NaCl浓度为3．4mol／L,含有甘油二醚脂,类胡萝卜素色素和极性脂TGD－2（三糖基二醚脂）,x（C＋C）分别为63．7％,62．7％,和64．3％,据此,这3株菌可归入嗜盐小盒苗属（Hchoarcula）;但它们不水解淀粉,明胶不液化,不水解酪蛋白等生理生化特性不同于该属中现已正式承认的两个种,因此,本工作分离的3株菌可能是嗜盐小盒菌属一个新种.     

利用rep-PCR研究云南尼泊尔桤木根瘤内Frankia基因多样性

利用rep-PCR指纹技术对云南5个地区的尼泊尔桤木根瘤内Frankia菌基因多样性进行研究,实验发现Frankia菌呈现丰富的基因多样性.所有71个样品被分为11种不同的基因类型.除高黎贡山外,不同地域都有某种基因型占优势,显示Frankia菌基因型与地域有紧密关系.所观察的5个地区中,高黎贡山Frankia菌基因型种类最多,多样性指数最高,基因多样性最为丰富.结合高黎贡山地理资料及其它生物多样性相关研究,推测与高黎贡山尼泊尔桤木共生的Frankia可能作为种储备库,为其它地区的寄主植物提供了祖先菌株.图8表1参13     

固氮放线菌Frankia与放线菌根植物共生进化的研究进展

固氮放线菌Frankia是能够诱导大范围的放线菌根植物(actinorhizal plants)产生根瘤放线菌.放线菌根植物是植物受Frankia侵染后能够形成根瘤的植物[1].目前,它们之间的进化研究虽然取得了很大进展,但是仍然存在许多不清楚的东西.本文根据最近对植物和Frankia系统进化研究成果、根瘤中放线菌Frankia的多样性研究情况,综述放线菌根植物和共生Frankia进化的研究进展.     

植物根瘤内痕量Frankia DNA的提取及鉴定

建立了一种简捷、快速植物根瘤内痕量Frankia DNA的提取方法，即采集自然态下根瘤，剥离瘤瓣，取瘤瓣尖经液氮研主民粉末，以20g/L CTAB（十六烷基三甲基溴化铵）消化胞壁，然后以d=5-10μm的石英粉吸附DNA，再以无菌去离子蒸馏水释放DNA，所获DNA产率及纯度较酚／氯仿法好。经对辽宁沙棘、色赤杨根瘤的瘤内痕量Frankia DNA及体外培养的9种Frankia菌株DNA分纯，并用于酶切及PCR，获满意结果。图3参12     

Diversity of root associated microorganisms of selected medicinal plants and influence of rhizomicroorganisms on the antimicrobial property of Coriandrum sativum

The total heterotrophic bacteria, actinomycetes and fungus were enumerated from the rhizosphere and non-rhizosphere soil of 50 selected locally available medicinal plants in and around Bharathiar University. In all the plants, population of microorganism were higher in the rhizosphere soil than in the non rhizosphere soil. Among the microorganisms, bacterial population was higher in number followed by fungus and actinomycetes. Of the medicinal plants, the maximum rhizosphere effect was observed in Annona squamosa and the minimum effect was seen in Eclipta alba and Cassia auriculata. Among the bacteria the dominant species was Bacillus followed by Pseudomonas, Enterobacter, Corynebacterium, Micrococcus and Serratia. The Streptomyces species was found to be dominant followed by Deuteromycetes and Frankia among the actinomycetes. Among the fungal isolates Rhizopus was found to be higher in number followed by Aspergillus, Penicillium, Mucor and Fusarium. About 70.96% of the bacterial isolates were found to be nitrate reducers and 90.60% of the bacteria solubilised phosphate. The rhizosphere bacterial isolates were also capable of hydrolyzing starch, cellulose, casein, urea and gelatin. The isolates of bacteria, actinomycetes and fungus were also able to produce phytohormone Indole-3-acetic acid (IAA). The maximum IAA production was recorded by Fusarium sp (5.8 mg/l). The rhizosphere bacterial isolates showed resistance to 14 commercially used antibiotics. In an attempt to check the influence of these plant growth promoting microorganisms on the antimicrobial property of Coriandrum sativum against Escherichia coli MTCC-443 and Aeromonas hydrophila MTCC-646, the results observed was not encouraging since the inoculants did not influence the antibacterial property. However extensive and in depth study is required to find out the influence of rhizomicroorganisms on the antibacterial property of medicinal plants. The other results clearly indicated that the rhizosphere microorganisms could be exploited for its innumerable properties and active metabolites.     

不同林龄木麻黄人工林生物多样性与土壤养分状况研究

木麻黄（Casuarina equisetifolia）是我国东南沿海海岸防护林骨干树种。本文对广东省茂名市木麻黄防护林开展了不同林龄的种群结构、植物多样性、以及土壤养分特征的研究。结果表明,滨海沙地木麻黄群落在生长过程中有明显的自疏现象,18年林个体数（975株·hm-2）不足3年林个体数一半（2350株·hm-2）,3年至6年龄木麻黄生长最快。调查林地内共有植物27种,其中灌木8种,草本植物18种。随林龄的增长,林下植物种数显著增加,多样性指数、均匀度指数逐步增加,优势度指数下降。林地土壤有机碳、全氮与速效氮供应水平极低;土壤磷供应相对较好,表层有效磷一般高于3.5mg·kg-1。土壤有效养分与植物多样性显著相关,显示养分是植物定居的主要限制因素。结果表明,木麻黄林结构简单,生物多样性低,土壤养分贫乏。     

木麻黄根区土壤理化特性及酶活性与青枯病发生关联分析

为探究木麻黄(Casuarina spp.)根区土壤理化特性及酶活性与青枯病发生的关系,进而为加强生物防治提供参考.选取位于广东省潮州市饶平县大埕镇、湛江市吴川市塘尾镇和湛江市徐闻县前山镇的3个木麻黄人工林为研究对象,在每个林分下分别设置3个样地,每个样地采集木麻黄青枯病感病植株和健康植株根区土壤各3份,共54个土壤样...     

Morphological variation and life history in cylindrical forms of Scytosiphon lomentaria (Scytosiphonaceae: Phaeophyta) from southern Australia

Variation amongst wild populations of Scytosiphon lomentaria (Lyngbye) Link is shown to be of a continuous nature, and the species cannot be readily segregated into distinct subspecific taxa on the basis of morphological characteristics. Zoospores derived from plurilocular sporangia developed asexually into cylindrical thalli under long and short days at 16°C. The progeny of some cylindrical thalli included microthalli bearing plurilocular sporangia or unilocular sporangia. Cylindrical forms of S. lomentaria were obtained in culture from wild microthalli of two kinds, bearing either plurilocular or unilocular sporangia. The microthalli differ morphologically from the ralfsioid stages commonly known to be plethysmothalli and sporophytes of S. lomentaria, but closely resemble the microthalli derived from cultures of wild S. lomentaria.     

Riparian forest composition affects stream litter decomposition despite similar microbial and invertebrate communities

Cross-boundary flows of energy and nutrients link biodiversity and functioning in adjacent ecosystems. The composition of forest tree species can affect the structure and functioning of stream ecosystems due to physical and chemical attributes, as well as changes in terrestrial resource subsidies. We examined how variation in riparian canopy composition (coniferous, deciduous, mixed) affects adjacent trophic levels (invertebrate and microbial consumers) and decomposition of organic matter in small, coastal rainforest streams in southwestern British Columbia. Breakdown rates of higher-quality red alder (Alnus rubra) litter were faster in streams with a greater percentage of deciduous than coniferous riparian canopy, whereas breakdown rates of lower-quality western hemlock (Tsuga heterophylla) litter were independent of riparian forest composition. When invertebrates were excluded using fine mesh, breakdown rates of both litter species were an order of magnitude less and were not significantly affected by riparian forest composition. Stream invertebrate and microbial communities were similar among riparian forest composition, with most variation attributed to leaf litter species. Invertebrate taxa richness and shredder biomass were higher in A. rubra litter; however, taxa evenness was greatest for T. heterophylla litter and both litter species in coniferous streams. Microbial community diversity (determined from terminal restriction fragment length polymorphisms) was unaffected by riparian forest or litter species. Fungal allele richness was higher than bacterial allele richness, and microbial communities associated with lower-quality T. heterophylla litter had higher diversity (allele uniqueness and richness) than those associated with higher-quality A. rubra litter. Percent variation in breakdown rates was mostly attributed to riparian forest composition in the presence of invertebrates and microbes; however, stream consumer biodiversity at adjacent trophic levels did not explain these patterns. Riparian and stream ecosystems and their biotic communities are linked through exchange and decomposition of detrital resources, and we provide evidence that riparian forest composition affects stream ecosystem catabolism despite similarities in microbial and invertebrate communities.     

应用全细胞蛋白SDS-PAGE分子标记技术验证含羞草根瘤菌的结瘤能力

对云南省热带及亚热带地区的含羞草根瘤菌进行了分离,选择其中40株菌为接种菌株,通过结瘤试验并采用全细胞蛋白SDS-PAGE分子标记方法研究了其结瘤能力.经过结瘤试验,发现除菌株SWF66075和SWF66093没有结瘤外,其它38株菌株均与含羞草植物结瘤.结瘤率为95%.从结瘤试验所获根瘤中,分离得到结瘤菌株,采用全细胞蛋白SDS-PAGE分子标记对结瘤菌株与接种菌株进行了比较研究.蛋白图谱及聚类分析显示,26株接种菌株与其结瘤菌株的全细胞蛋白分子图谱完全相同,在100%的相似水平上与其结瘤菌株聚在一起,说明宿主植物所形成的根瘤确系接种菌株侵入所致,因而可将这些菌株确认为根瘤菌菌株;而SWF66012、SWF66029、SWF66044和SWF66058等12株菌株的结瘤菌株与其各自接种菌株的全细胞蛋白图谱存在较大差异,推测这12株接种菌株与其结瘤菌株可能不是同一菌株,尚不能确定它们与含羞草植物的结瘤能力,这些菌株是否为根瘤菌菌株仍需进一步验证.研究结果表明,全细胞蛋向SDS-PAGE分子标记技术是一种快速、准确地验证根瘤菌结瘤能力的方法.该方法进一步完善了结瘤试验,并初步揭示了根瘤菌的竞争结瘤能力,适用于对大量根瘤内分离菌株进行根瘤菌的证实研究.图2参28     

Relationships among multiple trace elements in coastal Casuarina equisetifolia ecosystems on Hainan Island, South China

Forty-six trace elements in coastal Casuarina equisetifolia plant–soil systems at nine sampling sites on Hainan Island were analyzed using ICP-MS. The relationships among the trace elements of the same group or the same periodicity of the Periodic Table in the plants and soils were complex and no consistent patterns were found. More combinations of elements occurred with high positive correlation coefficients within the same periodicity than within the same group of the Periodic Table, and there were more high positive correlations in soils than in plants. However, there were many element combinations in Block d (transition elements) with high positive correlation coefficients in plants. Markedly high positive correlation coefficients between individual rare earth elements and Y and among Zr, Nb, Cd existed in both plants and soils. The dendrograms obtained by cluster analysis show that rare earth elements had very similar occurrence and distribution in both soils and plants. Thus, they behaved as a coherent group of elements both geochemically and biogeochemically. The transition elements were more coherent in plants than in soils.