成团泛菌YS19 symplasmata结构对菌体抵抗逆境的作用

成团泛菌（Pantoea agglomerans）YS19是从水稻“越富”品种中分离出的优势内生菌,其所形成的共质体（symplasmata）是一种与生物薄膜（biofilm）类似的多细胞聚集体结构,但细胞间联系比biofilm更加紧密。研究symplasmata结构对成团泛菌YS19抵抗逆境的贡献,有助于阐释内生菌与植物的相互作用的适应性。比较研究了symplasmata结构与散生菌体对于蔗糖渗透压冲击、重金属离子和干燥处理的抵抗能力差异,结果表明,与以散生状态存在的菌体相比,在面临逆境时形成symplasmata结构的菌体抗逆存活能力显著增强。     

水稻内生菠萝泛菌YJ76吲哚产量上调突变株的鉴定及生理性状

菠萝泛菌(Pantoea ananatis)YJ76是从水稻"越富"品种中分离的优势内生菌,与宿主水稻互作时具有多种促生作用,其分泌的吲哚作为细菌种内及种间的信号分子参与调控多种生理生化行为。[目的]筛选获得与吲哚调控相关的突变株,鉴定突变位点并研究突变基因对菌株的生存适应性以及对宿主水稻定殖和促生的影响,为研究吲哚调控通路奠定基础。[方法]用双亲本接合法构建YJ76的mTn5转座子插人突变文库,以染色体步移TAIL-PCR技术鉴定突变基因,最后探究基因突变对菌体产生的影响。[结果]筛选到1株吲哚产量大幅上升的YJ76突变株M04,鉴定突变位点为一个长度195 bp未报道过的新基因,将其命名为ipc(indole production control),基因突变后增强了YJ76对重金属、四环素和酸的抗性,也增强了菌体对宿主水稻定殖和促生的能力。[结论]吲哚产量上调的ipc突变株能够提高菌体生存适应性并增强其对宿主水稻定殖和促生的能力。     

水稻内生成团泛菌YS19共质体形成差异表达蛋白MalE及其兼职功能

成团泛菌YS19是从水稻“越富”品种中分离的一种优势内生细菌,与宿主水稻互作时具有多种促生作用,其形成的共质体(symplasmata)结构与菌体抗逆及与宿主互作有重要意义．研究发现了一种在YS19共质体形成阶段高表达的差异蛋白,对其用肽指纹图谱进行鉴定,发现其属于周质空间麦芽糖结合蛋白家族．克隆了该蛋白质的基因,重组表达并分离纯化了该蛋白质,发现它是一种兼职功能蛋白,其不仅参与麦芽糖的ABC运输系统,而且在强酸环境下不易发生变性沉淀,并可通过疏水面的显著暴露结合底物蛋白来发挥分子伴侣活性,这些兼职功能构成了菌体抗逆生存适应性的重要分子基础．     

水稻内生优势成团泛菌GFP标记菌株的性质与标记丢失动力学

为研究内生细菌对宿主植物侵染定殖的机理和其共生生物学作用 ,对水稻内生优势成团泛菌 (Pantoeaagglomerans)YS19与绿色荧光蛋白 (GFP)标记的YS19B ::gfp菌株的生长动力学进行了比较研究 ,探讨了成团泛菌YS19B ::gfp的标记稳定性和荧光性质 .标记菌株与野生型菌株相比 ,最大比生长速率和最大生物量仅减小 12 4 %和 6 % ,代时延长 14 0 % .成团泛菌YS19B ::gfp在指数期连续传代培养 10 0代后 ,GFP标记的保持率为 89 1% ,建立了标记菌株在有标记丢失存在时的生长动力学模型 :dX+ dt =μ+ (1-p)X+ ,解析出细胞分裂时标记丢失的概率p =9 75 6× 10 -7,确定了方程的模型参数 .标记菌株的荧光光谱在激发波长为 4 0 0nm时 ,最大发射波长为 5 0 8nm ,与供体菌株完全相同 .在LB培养基上生长时 ,成团泛菌YS19B ::gfp的GFP产生时间在指数期末期到稳定期较快 ,并于培养至 2 0h时达到最高 ,同时单位菌体生物量的荧光强度也达到最大 .结果说明 ,在GFP标记后成团泛菌YS19B ::gfp的生长仅受到较小影响 ,不致对成团泛菌的生理活动造成大的改变 ,同时由于该菌对宿主的侵染能力比其它内生细菌要强得多 ,因而该菌对植物的侵染活性影响也较小 ,该菌仍然可以保持其内生优势地位 .该标记的稳定性比较高 ,荧光产生正常 ,很适     

内生菌与植物的相互作用：促生与生物薄膜的形成

植物内生菌由于其独特的生态学地位而广受关注,近年来有关植物内生菌与宿主相互作用的研究取得了很大进展.本文综述了植物内生菌通过分泌促生物质、拮抗病原菌等实现与宿主共生互作,同时植物为内生菌提供适宜的黏附表面,使其形成以生物薄膜(biofilm)为主要形式的多细胞聚集体结构以更好地适应周围的生存环境,从而更加高效地对植物产生促生作用.本文论述了内生菌在与植物的互作中形成的多细胞聚集结构在抵抗非生物胁迫方面的独特生理及生态学意义,结合水稻内生成团泛菌YS19形成多细胞聚集体symplasmata现象及其生物学效应,对未来有关植物内生菌的研究方向提出了一些看法.     

水稻种子内生泛菌(Pantoea spp.)系统发育多样性及其促生功能

【目的】Pantoea菌株是广泛分布在自然界中的一类功能多样的细菌。本研究对分离自水稻种子内生的Pantoea菌株进行系统发育分析及功能评价,从而确定分类地位、种类多样性、分布特征及功能特性。【方法】采用乙醇-次氯酸钠联合灭菌方法进行水稻种子的表面灭菌,进行内生细菌的分离与纯化;其次将纯化后的菌株进行16Sr RNA基因PCR扩增及序列分析,通过MEGA7软件构建系统发育树;将分离得到的菌株进行功能实验检测,如溶磷、产IAA、产铁载体、拮抗病原真菌等特性,最后检测菌株的溶血性;水稻分型采用SSR方法,并对水稻农学性状如分蘖数、株高、植株重及产量进行调查。【结果】本研究对分离自8个不同基因型水稻种子中的146株内生Pantoea菌株进行系统发育分析及功能评价,结果发现所分离到的泛菌菌株主要属于Pantoea dispersa、Pantoea agglomerans、Pantoea cypripedii以及Pantoea brenneri四个种,其中P. dispersa的菌株数量最多,分布最广,并且存在于所有的8个水稻种子样品中。对其中66株菌进行功能检测,发现86.3%和69.7%的菌株具有溶磷和产IAA能力,有7株菌具有产铁载体能力,未发现对真菌病害Fusarium moniliforme有拮抗作用的菌株,并发现3株菌具有溶血性;本实验未发现泛菌组成与水稻系统发育及农学性状存在明显的相关性。【结论】本研究首次对水稻种子中泛菌的多样性及其功能进行报道,发现不同基因型的水稻种子所含Pantoea种类及组成存在差异,种子选择性地积累了Pantoea类群,大部分菌株具有一定的促生特性。该研究结果有助于进一步探究微生物与植物的共进化、种子微生物的传播途径及作用方式。     

甘草内生细菌多样性研究

以分离培养的方法对内蒙古鄂尔多斯市甘草基地野生及栽培甘草内生细菌的多样性进行了初步研究.结果表明,野生及栽培甘草植株内存在大量种群丰富的内生细菌.经ERIC-PCR指纹图谱分析,共分离到120株内生细菌,野生及栽培甘草均表现出根和叶部位的内生细菌数量多于茎部.对其中82株进行16S rDNA片段测序分析,结果表明这些内生细菌分别与GenBank中α、β、γ-Proteobacteria、Firmicutes、Actinobacteria五类细菌中的19个已知属相似性达到97%～100%.内生细菌的主要优势种群为芽孢杆菌属(Bacillus sp.)、假单胞菌属(Pseudomonas sp.)、泛菌属(Pantoea sp.)和沙雷氏菌属(Serratia sp.).     

一株能水解巴卡亭ⅢC-10位乙酰基的成团泛菌的筛选和鉴定

10-脱乙酰巴卡亭Ⅲ是合成紫杉醇和多烯紫杉醇的前体。以巴卡亭Ⅲ为底物,结合TLC、HPLC、HPLC-MS分析方法,通过设计专门的筛选方法筛选产酶菌株,得到一株巴卡亭ⅢC-10位脱乙酰基酶产生菌株Z1-56。以形态特征、生理生化特征、16S rDNA序列分析作为菌株的鉴定手段,Z1-56被鉴定为成团泛菌(Pantoea agglomerans),首次发现成团泛菌产生巴卡亭ⅢC-10-脱乙酰酶。     

具溶磷能力的植物内生促生细菌的分离筛选及其生物多样性

【目的】具溶磷能力的植物内生促生细菌的分离筛选及生物多样性的研究将有助于扩大溶磷微生物来源、丰富功能内生细菌资源库及开发新的改善土壤磷素营养途径。【方法】结合内生细菌分离方法从油菜和玉米体内分离筛选具溶磷能力的内生细菌,测定菌株摇瓶条件下的溶磷能力,并研究其产生吲哚乙酸(IAA)、铁载体、1-氨基环丙烷-1-羧酸(ACC)脱氨酶等的特性,采用16SrDNA限制性酶切片段长度多态性分析(RFLP)研究了具溶磷能力的植物内生促生细菌的遗传多样性,并挑选典型菌株进行了鉴定。【结果】分离筛选到32株具稳定溶磷能力的植物内生细菌,所有菌株都能从磷酸钙中释放出有效磷并使培养液pH值降低,释放的有效磷浓度最高达到537.6mg/L。分离自油菜的供试细菌都能产生吲哚乙酸和铁载体,分离自玉米的供试细菌中有68.4%的菌株产生吲哚乙酸,63.2%的菌株产生铁载体,63.2%的菌株具ACC脱氨酶活性。分离菌株在76%相似性水平上可聚类分为8个群。11株典型菌株归属于泛菌属(Pantoea)、假单胞菌属(Pseudomonas)、伯克霍尔德氏菌属(Burkholderia)、不动杆菌属(Acinetobacter)及罗尔斯顿菌属(Ralstonia)等5个属。【结论】油菜和玉米体内溶磷细菌具有丰富多样的生物学特性和遗传多样性。     

桑疫病病原拮抗菌的分离、鉴定及发酵条件优化北大核心CSCD

【目的】从健康桑树内生菌中分离获得对桑疫病病原菌(Pseudomonas syringae pv.mori)具有显著拮抗作用的菌株,优化其产生抑菌活性物质的发酵条件,为其生防利用奠定基础。【方法】从严格表面消毒的桑树根茎中分离内生菌,采用平板划线法纯化内生菌,用抑菌圈法筛选拮抗菌;根据菌株的形态与培养特征、生理生化特性、16S rDNA序列分析对其进行鉴定。通过单因素试验和正交设计试验优化培养基组分及发酵条件。【结果】从健康桑树中分离获得内生菌77株,其中,编号为SWg2的菌株对桑疫病病原菌具有强而稳定的抑制作用。菌株SWg2的形态与培养特征、生理生化特性和泛菌属(Pantoea sp.)相符,而16S rDNA序列分析结果显示它与成团泛菌(P.agglomerans)的亲缘关系接近。研究表明其最佳发酵配方和培养条件为:甘油(2.00%)、硝酸铵(2.00%)、KH2PO4(0.10%)和MgSO4·7H2O(0.15%),起始pH为7.5,装瓶量20 mL/100 mL,最适培养温度为28℃,转速为170 r/min,种子液接种量为4%,摇瓶培养5 d。【结论】经鉴定,对桑疫病病原具拮抗作用的桑树内生菌SWg2为成团泛菌(P.agglomerans),命名为成团泛菌SWg2。对其发酵条件进行优化后对桑疫病病原菌显示出更强的拮抗作用。     

Presence of Rhizobium Etli bv . Phaseoli and Rhizobium Gallicum bv . Gallicum in Egyptian Soils

Seven bean rhizobial strains EBRI 2, 3, 21, 24, 26, 27 and 29 identified as Rhizobium etli, and EBRI 32 identified as Rhizobium gallicum, isolated from Egyptian soils and which nodulated Phaseolus vulgaris efficiently, were subjected to hybridization with a nifH probe in order to estimate the copy number of this gene. Seven strains (EBRI 2, 3, 21, 24, 26, 27 and 29) which were only able to nodulate Phaseolus vulgaris, contained three copies of the nifH gene, consistent with their identification as Rhizobium etli bv. phaseoli. Only one strain (EBRI 32) which nodulated both Phaseolus vulgaris and Leucaena leucocephala, had one copy of nifH gene. This confirmed the classification of this strain as Rhizobium gallicum bv. gallicum.     

Effects of MULTIFOLIATE-PINNA, AFILA, TENDRIL-LESS and UNIFOLIATA genes on leafblade architecture in Pisum sativum

In order to dissect the genetic regulation of leafblade morphogenesis, 16 genotypes of pea, constructed by combining the wild-type and mutant alleles of MFP, AF, TL and UNI genes, were quantitatively phenotyped. The morphological features of the three domains of leafblades of four genotypes, unknown earlier, were described. All the genotypes were found to differ in leafblade morphology. It was evident that MFP and TL functions acted as repressor of pinna ramification, in the distal domain. These functions, with and without interaction with UNI, also repressed the ramification of proximal pinnae in the absence of AF function. The expression of MFP and TL required UNI function. AF function was found to control leafblade architecture multifariously. The earlier identified role of AF as a repressor of UNI in the proximal domain was confirmed. Negative control of AF on the UNI-dependent pinna ramification in the distal domain was revealed. It was found that AF establishes a boundary between proximal and distal domains and activates formation of leaflet pinnae in the proximal domain.     

Prosthecium species with Stegonsporium anamorphs on Acer

Data from microscopic morphology, single-spore cultures, and DNA analyses of teleomorphs and anamorphs support the recognition of five species of Prosthecium with Stegonsporium anamorphs on Acer: P. acerinum sp. nov., the teleomorph of S. acerinum; P. acerophilum comb. nov., formerly known as Dictyoporthe acerophila; P. galeatum comb. nov., originally described as Massaria galeata; P. opalus sp. nov.; and P. pyriforme sp. nov., the teleomorph of S. pyriforme s. str. The morphology of both type specimens and freshly collected material was investigated. The teleomorphs have brown ellipsoidal ascospores with five distosepta and often a longitudinal distoseptum. The anamorphs of all species described here belong to Stegonsporium; their connection to the Prosthecium teleomorphs was demonstrated by morphology and DNA sequences of single spore cultures derived from both ascospores and conidia. The anamorphs and teleomorphs of all five Prosthecium species are described and illustrated by LM images, and a key to these species is provided. As perceived from this work, S. pyriforme is restricted to Europe and does not occur in North America, whereas S. acerinum is restricted to North America, not found in Europe. The host associations given in the literature are revised and evidence is provided that only A. opalus, A. pseudoplatanus, and A. saccharum are confirmed hosts of Prosthecium with Stegonsporium anamorphs. Molecular phylogenetic analyses of tef1, ITS rDNA, and partial nuLSU rDNA sequences confirm that the species with Stegonsporium anamorphs are closely related to P. ellipsosporum, the generic type species. Stilbospora macrosperma is confirmed as the anamorph of P. ellipsosporum by DNA data of single spore isolates obtained from both ascospores and conidia.     

Characterisation of cell wall polysaccharides, arabinogalactans-proteins (AGPs) and phenolics of Cola nitida, Cola acuminata and Garcinia kola seeds

Many Cola plant species are endemic to West and Central Africa. Cola acuminata and Cola nitida are used as masticatory when fresh, while the dried nuts are used for beverages and pharmaceutical purposes in Europe and North America. Garcinia kola seeds, that serve as a substitute for the true kola nuts, are used in African traditional medicine for the treatment of various diseases, including colic, headache and liver cirrhosis. Seeds extracts of G. kola are also known for their anti-inflammatory, antimicrobial and antiviral properties. To gain information on the chemical properties of the kolas, we have isolated and analyzed cell wall polysaccharides, arabinogalactan-proteins and phenolic substances from the seeds of the three kola species. The sugar composition of cell wall material of C. acuminata, C. nitida and G. kola revealed that Gal (up to 30%), Ara, GalA and Glc as the predominant monosaccharides, representing approximately 90% by mol of the total hydrolysable sugar present in this material. In Ammonium oxalate cell wall fraction, GalA was found to be the major sugar present in all kola species. In the alkali-soluble fraction, there were significant differences in the level of Glc and Gal. The level of Glc was high in C. acuminata and C. nitida while the level of Gal and Xyl were high in C. nitida and G. cola. Isolation and quantification of arabinogalactan-proteins demonstrate that G. kola seeds contained four to eight times more of these proteoglycans than the seeds of the other two species. Finally, analysis of soluble phenolic substances shows that caffeine and catechin were largely represented in C. acumina and C. nitida seeds, with caffeine accounting for 50% of all soluble phenolics. These findings indicate that the three Kola seeds are highly enriched in pectins and proteoglycans and that C. acuminata and C. nitida can be used as a possible source of caffeine and catechin.     

中国黑粉菌属和利罗黑粉菌属

黑粉菌属是Roussel 1806年建立的,全世界记载有三百余种,主要寄生于禾本科,是经济作物及牧草的重要致病菌·长期以来,对黑粉菌的邢子使用过各种名称,如厚垣孢子,冬孢子及黑粉孢子等.本文采用黑粉孢子以区别锈菌的冬孢子. 芳’(1979)在《中国真菌总汇》中列出黑粉菌属五十种及一个变型.作者经过显微结构和超显微结构的研究,承认其中二十九种为正确名称,八种及一变型为异名,顶黑粉菌(Ustilago acrearus Berk.)由于错拼而被废弃.埃地黑粉菌(Ustilago emodensis Berk.)被转移至利罗粉菌属(Liroa).另有十一种黑粉菌因缺少标本留待今后订正.自1979年以后,杨信东(1983)增加黑粉菌属二种我国新纪录,K.范基和郭林(1986)描述一新种,四种新纪录.在本文中,作者描述一新种:鸢尾蒜黑粉(Ustilago ixiolirii Guo L) ,孢子堆生在蒴果内,不开裂,黑色,粉末状.黑粉孢子球形,近球形,稀椭圆形, 12.5-21×10-21μm,黑褐色,壁厚1-1.Sμm,纹饰脑状.是迄今生在石蒜科植物上唯一黑粉菌的种,其它几种黑粉菌均属条黑粉菌属.本文增加七种我国新纪录.共计四十九种,寄生于六科四十四属植物,主要是禾本科和蓼科.这仅是黑粉菌属研究的初步报告,在全国范围内大量采集黑粉菌标本后,作者相信会有更多新种和我国新纪录被发现.利罗黑粉菌属(Liroa)是从黑粉菌属(Ustaligo)分出的,此属为单种属.     

Comparative study of Bifidobacteriumanimalis, Escherichiacoli, Lactobacilluscasei and Saccharomycesboulardii probiotic properties

The present work investigates some probiotic properties of four different microorganisms (Bifidobacterium animalis var. lactis BB-12, Escherichia coli EMO, Lactobacillus casei and Saccharomyces boulardii). In vitro and in vivo tests were carried out to compare cell wall hydrophobicity, production of antagonistic substances, survival capacity in the gastrointestinal tract of germ-free mice without pathological consequence, and immune modulation by stimulation of Küpffer cells, intestinal sIgA and IL-10 levels. In vitro antagonism against pathogenic bacteria and yeast was only observed for the probiotic bacteria B. animalis and L. casei. The hydrophobic property of the cell wall was higher for B. animalis and E. coli EMO, and this property could be responsible for a better ability to colonize the gastrointestinal tract of germ-free mice. Higher levels of sIgA were observed mainly for S. boulardii, followed by E. coli EMO and B. animalis, and only S. boulardii induced a significant higher level of IL-10. In conclusion, for a probiotic use, S. boulardii presented better characteristics in terms of immunomodulation, and B. animalis and L. casei for antagonistic substance production. The knowledge of the different probiotic properties could be used to choice the better microorganism depending on the therapeutic or prophylactic application.