具有ACC脱氨酶活性的海滨锦葵(Kosteletzkya pentacarpos)内生细菌对小麦耐盐性的影响

从盐生植物海滨锦葵块根中分离内生细菌43株,经形态学特征和16S r DNA序列相结合的方法鉴定,分属10个种属,其中芽孢杆菌属是优势属,其次是假单胞菌属和农杆菌属,蜡样芽孢杆菌(Bacillus cereus)和地衣芽孢杆菌(Bacillus licheniformis)是优势种。对海滨锦葵内生细菌ACC脱氨酶活性的测定显示,其中5种菌明显具有ACC脱氨酶活性。用筛选到的5种细菌接种盐胁迫下小麦根系并测定其对于小麦耐盐性的影响。结果表明,蜡样芽孢杆菌、巨大芽孢杆菌(Bacillus megaterium)、短小芽孢杆菌(Bacillus pumilus)、地衣芽孢杆菌四种芽孢杆菌均能显著提高盐胁迫下小麦幼苗的干物质重和叶绿素含量,并能显著提高保护酶(SOD、POD、CAT)活性,对盐胁迫的毒害有一定的缓解作用。绿针假单胞菌(Pseudomonas chlororaphis)对小麦幼苗株高、根长、鲜重、干重、叶绿素含量和保护酶活性的提高也具有一定的作用。上述分析表明从海滨锦葵块根中分离出的5株具有ACC脱氨酶活性的内生细菌均能提高小麦幼苗的耐盐性。     

亳菊内生菌BN7的鉴定与生防促生功能研究

植物内生菌具有增强宿主植物抵抗生物和非生物胁迫的能力,也具有增强宿主植物对磷、钾等矿质元素的分解吸收,进而促进植物的生长。本文采用平板对峙法检测亳菊内生菌BN7对玉米弯孢病菌、黄瓜枯萎病菌、小麦赤霉病菌、串珠镰刀病菌、瓜炭疽病菌、茶叶轮斑病菌等6种植物病原菌的拮抗作用;测定菌株BN7的解磷、解钾、产生长素、降解纤维素能力和对DPPH的清除能力;并根据菌株形态特征、生理生化特征和16S rDNA序列分析对菌株BN7进行鉴定。结果表明:亳菊内生菌BN7对小麦赤霉菌等6种植物病原菌均有抑制作用;菌株BN7使有机磷和无机磷发酵液中可溶性磷分别增加了108.38和68.71 mg/L,使可溶性钾增加了32.03 mg/L,IAA的分泌量高达164.39 mg/L;培养3天菌株BN7羧甲基纤维素酶和滤纸酶的活力达到最高,分别为168.78和79.87 U/mL;培养7天的上清液对DPPH清除率可达82.13%;根据形态学特征、生理生化特征及16S rDNA序列分析,菌株BN7初步鉴定为巨大芽孢杆菌(Bacillus megaterium)。因此,亳菊内生菌BN7是一株具有广谱抗真菌、解磷、解钾、产IAA和降解纤维素能力的巨大芽孢杆菌。     

VC三菌种一步发酵方法的构建与研究

就维生素C微生物一步发酵方法进行了探索,构建了酮古龙酸杆菌、氧化葡萄糖酸杆菌和芽孢杆菌三菌混菌一步发酵的方法。研究发现,植物内生芽孢杆菌可以与酮古龙酸杆菌配合,促进酮古龙酸杆菌生长和产酸。在有山梨醇存在的条件下酮古龙酸杆菌及其伴生菌能够快速地生长增殖,植物内生芽孢杆菌在发酵的10h中不断消耗山梨醇。5L的发酵罐中,酮古龙酸杆菌、氧化葡萄糖酸杆菌和植物内生芽孢杆菌三菌混菌一步发酵在恒定的30℃温度,600r/min搅拌速度和1.5vvm通气条件下,补料发酵过程中醇酸质量转化率达到了81.89%,在分批发酵过程中,醇酸质量转化率达到了87.90%,进一步优化了维生素C生产工艺。     

玉米根系内生细菌种群及动态分析

2000-2002年,先后对辽宁省14个玉米主栽品种进行了根系内主要细菌种群分析．结果表明．玉米内生细菌的主要种群为芽孢杆菌属(Bucillus spp．),此外还包括肠杆菌属、沙雷氏杆菌属、假单胞菌属、黄单胞菌属和棍状杆菌属．其中Bacillus分布最广,已鉴定出8个种,包括枯草芽孢杆菌、巨大芽孢杆菌、蜡状芽孢杆菌、地衣芽孢杆菌、炭疽芽孢杆菌、蕈状芽孢杆菌、短小芽孢杆菌、环状芽孢杆菌．Bacillusspp．总量占根系内生细菌总量比苗期和成株期分别为75．5％和76．6％．内生细菌在不同玉米品种和不同生育期之间存在程度不同的差异．研究发现,品种的遗传背景与其内生细菌的种类和数量显著相关．     

两株具促生作用的苜蓿内生菌的分离纯化与鉴定

【目的】分离具有植物促生功能的苜蓿内生菌。【方法】从苜蓿的新鲜组织中分离纯化菌株,进行16S r RNA基因序列分析,生理生化鉴定并检测内生菌对苜蓿的促生特性。【结果】获得两株苜蓿内生菌,成团泛菌ASR16和短小芽孢杆菌ALR33。两株菌株均能产生植物生长激素IAA和嗜铁素,能溶解磷,对苜蓿的促生作用明显。【结论】获得两株具有植物促生特性的苜蓿内生菌ASR16和ALR33,可作为微生物菌肥的研究基础。     

甘草内生细菌的分离及拮抗菌株鉴定

从乌拉尔甘草健康植株的根茎叶中共分离到内生细菌98株,经初步鉴定芽孢杆菌属(Bacillus sp.)为优势种群,约占30%;从不同生长年份甘草的根、茎、叶组织中分离内生细菌种群密度从5.0×104cfu/g~2.9×107cfu/g鲜重不等。采用平板对峙方法筛选出6株对植物病原菌有明显体外拮抗活性的菌株,通过菌落、菌体形态观察、生理生化反应及16S rDNA序列分析,同时结合Biolog细菌自动鉴定系统验证,鉴定这6株拮抗菌分属萎缩芽孢杆菌(Bacillus atrophaeus)、多粘类芽孢杆菌(Paenibacillus polymyxa)、枯草芽孢杆菌(Bacillus subtilis)、Paenibacillus ehimensis。     

一株耐硒壶瓶碎米荠内生菌分离、鉴定及其体外硒代谢研究

【背景】壶瓶碎米荠对硒具有超积累能力,并主要以硒代胱氨酸的形式存在,与已有的硒超积累植物显著不同,其硒超积累机制不明。【目的】从硒超积累植物壶瓶碎米荠(Cardamine hupingshanensis)体内分离耐硒内生菌,并对其进行鉴定和体外硒代谢特征研究,为壶瓶碎米荠超积累硒的机制研究提供参考。【方法】从壶瓶碎米荠新鲜叶片中分离纯化耐硒内生菌株,对其进行生理生化特征及16S rRNA基因序列分析鉴定,并对其进行亚硒酸钠培养代谢。【结果】获得一株耐硒内生菌CSN-1,被鉴定为甲基营养型芽孢杆菌(Bacillus methylotrophicus),培养液中硒含量低(Se 1.5 mg/L)时其吸光度值较对照组高,硒含量高(Se 10 mg/L)时其吸光度值较对照组低;代谢后的上清液中硒主要以Se~(4+)存在,而菌体中硒主要是硒代胱氨酸(SeCys_2)。【结论】硒超积累植物壶瓶碎米荠叶片体内存在甲基营养型芽孢杆菌(Bacillus methylotrophicus)CSN-1,具有将亚硒酸钠转化为硒代胱氨酸的能力,低浓度的硒对该内生菌的生长具有一定的促进作用,而高浓度的硒则会抑制该内生菌的生长。     

一株西沙群岛野生诺尼内生细菌NG14的分类鉴定及拮抗活性

植物的内生菌与植物生长健康及其功效性成分产生具有一定的相关性。以分离自我国西沙群岛野生诺尼成熟果实的内生细菌菌株NG14为对象,利用形态学观察,生理生化特征鉴定及系统发育学分析,对其菌种的分类地位进行研究,将其鉴定为多黏类芽孢杆菌(Paenibacillus polymyxa);并通过选择多种植物及人体病原菌作为生防对象进行拮抗试验,发现了该菌株具有广谱且良好的拮抗病原菌活性。     

一株引起马来甜龙竹组培污染内生菌的分离与鉴定

【目的】对一株引起马来甜龙竹组培污染内生菌的分离与鉴定。【方法】采用改良的NA培养基分离纯化菌株,并通过菌体的形态结构观察、生理生化试验及其16SrDNA序列同源性分析对其进行鉴定。【结果】菌株SWFU01的形态特征及生理生化试验结果与解淀粉芽孢杆菌[Bacillus amyloliquefaciens(Fukumoto)Priest et al.]的描述基本相同;16S rDNA序列分析表明,该菌株与解淀粉芽孢杆菌JS在同一系统发育分支,其同源性为99.28%。【结论】综合形态学特征、生理生化特征以及16S rDNA序列分析的研究结果,菌株SWFU01被鉴定为解淀粉芽孢杆菌。     

盐生植物内生菌多样性与募集研究进展

内生菌在盐生植物适应高盐环境中有重要作用,其类群组成受环境盐胁迫、宿主植物及微生物生物学特性的调控。环境胁迫会通过宿主生理状态传递到共生微生物,进而塑造特殊的内生菌类群,同时一种微生物是否可以成为内生菌又受宿主和微生物自身生物学特征的双重影响。植物只可以募集已经存在于环境在的微生物,由此为利用微生物改善植物的逆境适应提供了基础。该文章就盐胁迫如何塑造盐生植物内生菌类群及植物如何募集外源微生物进行了综述,为了解共生抗盐和利用微生物改良盐碱环境提供参考。     

一株番茄青枯菌拮抗菌的鉴定及抗病效果初探

从形态、生理生化、16S rDNA3个方面确定了番茄青枯菌拮抗菌株3-1-16的分类地位。光学显微镜下观察到菌体为杆状细胞,革兰氏染色均匀,并可见菌体染成蓝紫色。透射电镜进一步观察到细胞内有许多颗粒状物质,无伴胞晶体。Biolog鉴定,3-1-16与巨大芽孢杆菌(Bacillus megaterium)具有最高相似率为98%。16S rRNA分析,3-1-16与巨大芽孢杆菌MO31同源性最高为99.4%。聚类分析显示3-1-16与3株巨大芽孢杆菌聚成一支,支持度为100%。生理生化特征及培养特征测定结果表明,菌株3-1-16鉴定为巨大芽孢杆菌(Bacillus megaterium)。盆栽试验表明该菌株对番茄青枯病防病效果达到81.3%。     

小碎斑鱼蛉幼虫肠道细菌分离及鉴定研究

目的从微生态学角度研究小碎斑鱼蛉幼虫的营养生理活动。方法从小碎斑鱼蛉幼虫肠道环境中进行分离、纯化和培养,获得3个细菌菌株,对其菌体形态、染色反应、培养性状和生理生化反应进行系统研究。结果上述3个菌株均属于芽胞杆菌属(Bacillus),1号菌株为巨大芽胞杆菌(Bacillus megaterium),2号菌株为枯草芽胞杆菌(Bacillus subtilis),3号菌株为地衣芽胞杆菌(Bacillus licheniformis)。结论小碎斑鱼蛉幼虫肠道环境中的不同细菌,其数量存在明显差异。     

Efficient cloning in Bacillus megaterium: comparison to Bacillus subtilis and Escherichia coli cloning hosts

Quantitative cloning efficiencies for B. megaterium, B. subtilis , and E. coli were compared. Transformation of B. megaterium is less efficient than transformation of B. subtilis or E. coli . The frequency of recombinant clones was equal in E. coli and B. megaterium ; both somewhat higher than in B. subtilis . Equivalent average insert sizes were found in B. megaterium and E. coli clones, but significantly smaller inserts were obtained in B. subtilis clones. Clones obtained and propagated in B. megaterium were structurally stable when grown under plasmid selection.     

Gene dosage effect on the expression of the delta-endotoxin genes of Bacillus thuringiensis subsp. kurstaki in Bacillus subtilis and Bacillus megaterium

Significant differences in expression of the delta-endotoxin genes cryA1 and cryA2 of Bacillus thuringiensis subsp. kurstaki were observed in B. subtilis and B. megaterium. The cryA1 gene was expressed when present on a high-copy-number (hcn) vector in B. megaterium but not in B. subtilis. The cryA2 gene was expressed in both hosts, but at a higher level in B. megaterium. Expression of the cryA2 gene in B. megaterium was better from a hcn vector than from a low copy number vector. Inhibition of sporulation was observed when the toxin genes were present on hcn plasmids in B. subtilis while no such effect was evident in B. megaterium. In addition, there was a significant reduction in copy numbers in both B. subtilis and B. megaterium when delta-endotoxin genes or a spoVG promoter-containing fragment of DNA were cloned into hcn plasmids.     

黑粉虫与黄粉虫幼虫肠道细菌的比较

在黑粉虫和黄粉虫肠道中分别分离获得5株细菌,对其菌体形态、培养性状、染色反应、生理生化反应等进行了系统研究。鉴定结果表明,黑粉虫的5个细菌菌株分别属于金杆菌属(Aureobacterium)、李斯特氏菌属(Listeria)、微杆菌属(M i-crobacterium)、莫拉氏菌属(M oraxella)、短小杆菌属(Curtobacterium);黄粉虫的5个细菌菌株分别属于金杆菌属(Aureobacte-rium)、球形芽孢杆菌(Bacillus sphaericus)、微杆菌属(M icrobacterium)、巨大芽孢杆菌(B.m egaterium)、短小杆菌属(Curtobac-terium)。金杆菌属(Aureobacterium)、微杆菌属(M icrobacterium)和短小杆菌属(Curtobacterium)均在2种昆虫肠道中出现。     

Comparison of various properties of low-molecular-weight proteins from dormant spores of several Bacillus species.

Several properties of the major proteins degraded during germination of spores of Bacillus cereus, Bacillus megaterium, and Bacillus subtilis have been compared. All of the proteins had low molecular weights (6,000 to 13,000) and lacked cysteine, cystine, and tryptophan. The proteins could be subdivided into two groups: group I (B. megaterium A and C proteins, B. cereus A protein, and B. subtilis alpha and beta proteins) and group II (B. cereus and B. megaterium B proteins and B. subtilis gamma protein). Species in group II had lower levels of (or lacked) the amino acids isoleucine, leucine, methionine, and proline. Similarly, proteins in each group were more closely related immunologically. However, antisera against a B. megaterium group I protein cross-reacted more strongly with the B. megaterium group II protein than with group I proteins from other spore species, whereas antisera against the B. megaterium group II protein cross-reacted most strongly with B. megaterium group I proteins. Analysis of the primary sequences at the amino termini and in the regions of the B. cereus and B. subtilis proteins cleaved by the B. megaterium spore protease revealed that the B. cereus A protein was most similar to the B. megaterium A and C proteins, and the B. cereus B protein and the B. subtilis gamma protein were most similar to the B. megaterium B protein. However, amino terminal sequences within one group of proteins varied considerably, whereas the spore protease cleavage sites were more highly conserved.     

Metabolome profiling reveals metabolic cooperation between Bacillus megaterium and Ketogulonicigenium vulgare during induced swarm motility

The metabolic cooperation in the ecosystem of Bacillus megaterium and Ketogulonicigenium vulgare was investigated by cultivating them spatially on a soft agar plate. We found that B. megaterium swarmed in a direction along the trace of K. vulgare on the agar plate. Metabolomics based on gas chromatography coupled with time-of-flight mass spectrometry (GC-TOF-MS) was employed to analyze the interaction mechanism between the two microorganisms. We found that the microorganisms interact by exchanging a number of metabolites. Both intracellular metabolism and cell-cell communication via metabolic cooperation were essential in determining the population dynamics of the ecosystem. The contents of amino acids and other nutritional compounds in K. vulgare were rather low in comparison to those in B. megaterium, but the levels of these compounds in the medium surrounding K. vulgare were fairly high, even higher than in fresh medium. Erythrose, erythritol, guanine, and inositol accumulated around B. megaterium were consumed by K. vulgare upon its migration. The oxidization products of K. vulgare, including 2-keto-gulonic acids (2KGA), were sharply increased. Upon coculturing of B. megaterium and K. vulgare, 2,6-dipicolinic acid (the biomarker of sporulation of B. megaterium), was remarkably increased compared with those in the monocultures. Therefore, the interactions between B. megaterium and K. vulgare were a synergistic combination of mutualism and antagonism. This paper is the first to systematically identify a symbiotic interaction mechanism via metabolites in the ecosystem established by two isolated colonies of B. megaterium and K. vulgare.     

Can immobilization of Bacillus megaterium cells in alginate beads protect them against bacteriophages?

The ability of free and alginate-immobilized cells of Bacillus megaterium to dissolve tricalcium phosphate as well as their susceptibility to phages were compared in pure liquid cultures and in pot experiment with maize. In both liquid culture and cultivated soil, alginate-immobilized cells of B. megaterium exhibited much higher efficiency in increasing the availability of phosphorus than the free cells. Bacteriophages of B. megaterium were found to be common in soil. Specific bacteriophages, in the presence of free cells of B. megaterium, completely inhibited the phosphate-dissolving activity of the bacteria in pure liquid culture and markedly decreased their number in rhizosphere of maize plants. The phosphorus content of maize plants inoculated with free cells of B. megaterium decreased in the presence of their specific bacteriophages, whereas, when alginate-immobilized cells were used as inoculum, no effect of the presence of bacteriophages on phosphate-dissolving activity of bacterial cells was detected.     

Methylated Bases of Bacillus megaterium KM Nucleic Acids: Comparison with Escherichia coli

Parallel studies were performed with methionineless derivatives of Escherichia coli 15 T(-) and Bacillus megaterium KM: T(-). Methylated bases are present in the total cell ribonucleic acid (RNA) of B. megaterium. The level of RNA methylation in E. coli is about 60% greater than that in B. megaterium. Although E. coli deoxyribonucleic acid (DNA) was found to contain 0.12% 5-methylcytosine (5-MC) and 0.24% 6-methylaminopurine (6-MA), methylated bases were not detected in the DNA of B. megaterium. Assuming a molecular weight of 7 x 10(9) daltons for B. megaterium DNA, it was calculated that this organism could not contain more than one molecule of 5-MC or 6-MA per genome, and that possibly no methylated bases were present. Methylated bases were also not detected in the DNA of thymine-starved B. megaterium. Crude extracts of this organism possess RNA methylase activity but no detectable DNA methylase activity.     

Comparative antigenicity of spore coat proteins from Bacillus species using antibody to spore coat proteins of Bacillus megaterium

Spore coat proteins obtained by extraction with sodium dodecylsulfate/dithiothreitol from six Bacillus spores were compared by immunoblot analysis using antibodies to spore coat proteins from two strains of B. megaterium. Although the extract from spores of each strain had heterogenous proteins with various molecular weights, there were some bands which cross-reacted with specific antibodies from B. megaterium spores. Specific antibody to 48K protein from B. megaterium ATCC 12872 cross-reacted with 17K protein from B. megaterium ATCC 19213, 13K protein from B. cereus and 50K protein from B. subtilis 60015 and B. subtilis NRRL B558. Also, specific antibody to 22K protein from the same strain cross-reacted with 22K and 17K proteins from B. megaterium ATCC 19213 and 13K protein from B. cereus T. Specific antibody to 17K protein from B. megaterium ATCC 19213 reacted with 22K and 19K proteins in addition to 17K protein of own strain, and it was cross-reactive with 16K protein from B. megaterium ATCC 12872, 19K and 27K proteins from B. thiaminolyticus, 13K protein from B. cereus.