硫酸铵对两株钾矿物分解细菌生长代谢和风化钾长石的影响

【目的】为了明确钾矿物分解细菌Bacillus globisporus Q12和Rhizobium sp.Q32最合适的产酸和胞外多糖条件,并进一步阐明供试菌株对钾长石的溶解效应及其机制。【方法】分别向培养基中加入0-1.2 g/L(NH4)2SO4,选择菌株最适的产酸及合成胞外多糖条件,研究菌株对钾长石的溶解效果,并采用扫描电镜(SEM)观察钾长石表面形态及菌体分布特征。【结果】0.6、0和0.3 g/L(NH4)2SO4分别能使菌株Q12、Q32和混合菌株(Q12+Q32)产生较多的有机酸、胞外多糖以及有机酸和胞外多糖的复合物。菌株Q12、Q32及其混合菌株均能够显著地溶解钾长石,并释放出矿质元素,其中混合菌株的溶解效果要优于单一菌株;SEM分析表明,混合菌株对钾长石的溶蚀作用最强。【结论】(NH4)2SO4的含量能够影响供试菌株Q12和Q32的生长代谢及其对钾长石的风化作用,混合菌株可以通过产生的有机酸和胞外多糖的联合作用加速对钾长石的风化。     

不同风化程度钾长石表面矿物分解细菌的筛选及遗传多样性

【目的】不同风化程度钾长石表面矿物分解细菌生物多样性研究将有助于了解矿物生物风化、生物成矿和土壤形成的演化规律和机理。【方法】采用纯培养法自南平钾矿区高、中、低风化度钾长石以及矿区土壤样品中分离矿物分解细菌,通过摇瓶释硅实验比较不同菌株分解矿物能力,采用16SrDNA限制性酶切多态性分析(Amplified rDNA Restriction Analysis,ARDRA)研究了供试菌株的遗传多样性。【结果】分离筛选到35株生长良好的矿物分解细菌,与对照相比,接菌处理发酵液中有效硅增加了101～206%;所有供试菌株可分为11个OTU,分别属于5个门,6个科,7个属。多数菌株(74%)属于γ-变形杆菌纲(γ-Proteobacteria)。泛菌属(Pantoea),沙雷氏菌属(Serratia),假单胞菌属(Pseudomonas)为优势种群。【结论】南平钾矿区矿物分解细菌具有丰富的微生物种群多样性,且γ-变形杆菌纲(γ-Proteobacteria)细菌在钾长石风化过程中可能起了重要的作用。     

山东地区钾矿物分解细菌的分离及生物学特性

不同土壤类型钾矿物分解细菌资源调查和高效稳定释钾、促生细菌的筛选鉴定有助于丰富微生物资源库,发掘和利用钾矿物分解细菌以及探究矿物生物风化机理等。作者采用以钾长石为唯一钾源的选择性细菌培养基, 从山东地区不同土壤和不同植物根际土壤中分离纯化了23株生长势良好的钾矿物分解细菌, 通过测定细菌代谢产物IAA和铁载体,研究其产生促生物质的能力, 通过摇瓶试验筛选高效释钾菌株, 采用16S rDNA限制性酶切多态性分析(amplified rDNA restriction analysis, ARDRA)方法研究了钾矿物分解细菌的遗传多样性, 根据16S rDNA同源性对高效释钾菌株进行了鉴定。结果表明, 供试菌株均产吲哚乙酸或其衍生物, 43.5%的分离菌株产极高量铁载体。ARDRA结果表明供试菌株在60%相似性水平上可分为11个基因型, 同一类型土壤上不同作物根际或不同类型土壤上同一作物根际的钾矿物分解细菌存在明显的遗传差异。摇瓶试验结果表明供试菌株中具有较显著释钾能力的菌株占17%, 39%的供试菌株无释钾能力。筛选到2株高效释钾菌株AFM2、AC2, 分别使溶液中钾含量增加了29.8%和25.4%。16S rDNA同源性分析表明菌株AC2、AHZ1与Bacillus mucilaginosus聚为一群, 该群与包含菌株AZH4的Paenibacillus sp.中的种聚为一大发育分支, 该分支在细菌分类地位上隶属于Firmicutes; 菌株AFM2与Rhizobium sp. 和Agrobacterium tumefaciens聚为另一大发育分支, 该分支在细菌分类地位上隶属于Alphaproteobacteria。     

表面活性剂对Bacillus sp. ATCC 21616合成腺苷的影响

研究了三种不同的表面活性剂对芽孢杆菌ATCC21616的腺苷合成和菌体生长的影响.结果发现,非离子表面活性剂Tween 80对腺苷合成的促进作用最大,而阳离子表面活性剂CTAB对菌体抑制作用明显.添加低浓度阴离子表面活性剂SDS也能够促进腺苷产生.表面活性剂的最佳添加时间是发酵后期.     

细菌sRNA数据库北大核心CSCD

细菌sRNA是一类长度在50~500 nt的调控小RNA(small regulatory RNA),主要通过与靶标mRNA或靶标蛋白质结合发挥多种生物学功能。目前,随着生物信息学与高通量测序的应用,发现了越来越多的细菌sRNA,开发了多个相关数据库。为了sRNA工作者系统了解与应用这些数据,本文拟对包含细菌sRNA的综合数据库和细菌sRNA专业数据库作一概述,并对sRNA数据库的未来发展进行展望。     

枯草芽孢杆菌(Bacillus subtilis)Tpb55灌根与烟草根围细菌多样性变化的相关性北大核心CSCD

【目的】利用454高通量测序技术分析生防菌株枯草芽孢杆菌(Bacillus subtilis)Tpb55对烟草根围土壤细菌群落的影响;同时测定施加Tpb55处理对烟草黑胫病的大田防效。【方法】试验设置Tpb55菌剂108 CFU/m L灌根和空白对照2个处理,分别在处理0、10、22 d采集烟草根围土壤,提取土壤细菌总DNA,扩增细菌16S r DNA V1-V3区,对扩增产物进行454高通量测序,使用Qiime软件分析不同处理的细菌群落结构。【结果】对照和处理样品测序后共获得了41207个优质序列,鉴定属于细菌的25个门。所有样品中优势菌群均为放线菌门(Actinobacteria)、变形菌门(Proteobacteria)和酸杆菌门(Acidobacteria)。在病情发展过程中,放线菌门的细菌丰度逐渐下降,变形菌门含量呈上升趋势。施用Tpb55后,酸杆菌门含量明显上升并高于对照。对照中芽孢杆菌科(Bacillaceae)和多样性指示菌草酸杆菌科(Oxalobacteraceae)的细菌丰度均明显下降,Tpb55处理的样品中芽孢杆菌科含量不断上升,草酸杆菌科含量相对稳定。Chao1、ACE和Shannon指数分析表明,Tpb55处理的样品中细菌多样性和丰富度不断提高且高于对照。Tpb55处理后10 d和22 d,OTU序列数据库中与Tpb55 16S r DNA V1-V3区PCR产物高度同源OTU数目分别为31和45。Tpb55处理的烟草黑胫病病情指数(5.29)显著低于对照(38.52)。【结论】施用Tpb55可以提高根围土壤细菌多样性和群落结构稳定性,这可能是其发挥良好生防作用的重要机制之一。     

Clostridium thermocellum与Bacillus licheniformis共培养分解纤维素的性质

【目的】采取人工构建复合菌系的方法探索微生物协同降解纤维素的机理及菌间关系。【方法】从一组高温发酵木质纤维素原料产沼气的菌群中分离获得若干菌株,其中一株细菌经16S rRNA基因全序列测序比对后鉴定为地衣芽孢杆菌(Bacillus licheniformis),将该菌株与厌氧纤维素分解菌Clostridium thermocellum CTL-6进行共培养,菌株组合表现出很强的滤纸纤维素分解能力。【结果】两菌共培养9 d,累计滤纸分解量为484.6 mg,滤纸相对分解率高达93.2%;pH变化呈先下降后逐步回升,培养3 d后pH由初始时的7.00降到最低值6.57,第9天升至7.73;菌株组合能同时产生纤维素酶和半纤维素酶,培养过程中两种酶活性大小均呈不断上升趋势,最大值分别为0.32 U/m L和0.57 U/m L。利用HPLC监测了乳酸、甲酸、乙酸、丙酸和丁酸5种有机酸含量的变化,其中丁酸、丙酸代谢量最高,分别为1 477.3 mg/L和1 068.8 mg/L;除丙酸外,其他4种有机酸含量变化趋势与滤纸降解的变化均无明显相关性。5种有机酸总含量的变化与p H的变化趋势一致,表明对pH变化起决定性作用的很可能是某种未检测的酸性较强的物质含量变化。【结论】Bacillus licheniformis能有效促进Clostridium thermocellum CTL-6的纤维素分解活性,且该菌株组合可作为后期进一步构建纤维素甲烷转化复合菌系的基础。     

芽孢杆菌Bacillus sp. S-1壳聚糖酶基因的克隆与序列分析

从连云港海滩晒虾蟹壳的泥土里筛选出一株产壳聚糖酶能力较高的菌株S-1,根据其形态特征、生理生化以及16S rDNA鉴定,初步认定该菌为芽孢杆菌属（Bacillus）。利用NCBI数据库中已经报道的Bacillus壳聚糖酶序列设计兼并引物,以菌株Bacillus sp. S-1的基因组DNA为模板进行聚合酶链式反应（PCR）,克隆到壳聚糖酶基因的部分序列;利用Clontech公司Universal GenomeWalker试剂盒构建该菌株的基因组步移文库,根据已测定的序列信息设计特异性引物,结合两步法PCR技术分别克隆两端未知序列,拼接获得壳聚糖酶基因的全长序列（该基因全长1362 bp编码453个氨基酸,注册号：EU924147）,并对该序列进行了生物信息学方面的分析。     

嗜碱芽孢杆菌Bacillus sp. N16-5表达载体的构建

【目的】构建可以在嗜碱芽孢杆菌Bacillus sp.N16-5中表达外源基因的表达载体。【方法】以枯草芽孢杆菌表达质粒pHCMC04为基本骨架,删除木糖诱导的启动子和木糖阻遏蛋白基因,分别插入枯草芽孢杆菌组成型强启动子P43和嗜碱芽孢杆菌N16-5的组成型启动子P EF,构建表达载体pABN165P43和pABN165P EF;将绿色荧光蛋白基因gfp作为报告基因连接至表达载体得到pABN165P43-gfp和pABN165P EF-gfp,利用原生质体转化法将其转化至Bacillus sp.N16-5,通过荧光显微镜和多功能荧光读板仪检测报告基因的表达情况。【结果】所构建的表达载体pABN165P43-gfp和pABN165P EF-gfp可以在Bacillus sp.N16-5中表达报告基因gfp,荧光定量数据显示,在7.5 h左右开始检测到绿色荧光蛋白的表达,从7.5 h到12 h荧光强度随时间增长迅速并在12 h左右达到最大,最大荧光值在7000左右。【结论】成功构建了2个嗜碱芽孢杆菌Bacillus sp.N16-5表达载体,实现了外源基因在嗜碱芽孢杆菌中的表达。     

具细菌群体感应抑制活性海洋细菌的筛选鉴定

目的:从海洋环境中筛选对细菌群体感应有抑制作用的活性菌株,为以致病菌群体感应系统为靶点的新型疗法提供新的药用资源。方法:从海水中分离纯化细菌菌株,采用根癌农杆菌平板筛选模型筛选细菌群体感应抑制活性细菌,对筛选出的海洋细菌进行生理生化和16S rDNA序列测定,根据《伯杰氏手册》进行菌种分类鉴定。结果:从217株海洋细菌中筛选出1株能显著抑制细菌群体感应效应的海洋细菌Y2,该海洋细菌具有蜡样芽孢杆菌(Bacillus cereus)的典型特征,其16S rDNA序列与GenBank中蜡样芽孢杆菌16S rDNA的部分序列有100%的同源性。结论:海洋环境中也存在具有抑制细菌群体感应活性的微生物。     

芽孢杆菌木聚糖酶的发酵条件研究

本文研究了芽孢杆菌Ｌ２３产木聚糖酶的时间曲线,碳源种类和浓度,添加物,发酵起始ｐｈ以及接种量对产酶的影响。该菌经３７℃培养５０小时,酶活力为３０ＩＵ／ｍｌ,酶最适反应温度为５７℃,最适ｐＨ值为７．０。     

Effect of manganese on alkaline phosphatase production in Bacillus sp. RK11

Summary Bacillus sp. RKll, an alkalophilic isolate from soil, produces an extracellular alkaline phosphatase. In the absence of Mn²⁺ in a complex medium, no alkaline phosphatase production or sporulation by the organism was detected. No other divalent metal could be substituted for Mn²⁺ in enzyme production or in sporulation. Manganous sulphate (70 mol) gave highest enzyme production although spore numbers continued to increase with manganous concentrations above this level. Maximum alkaline phosphatase production occurred when the metal was present at the time of inoculation but maximum spore numbers were detected when the metal was added 8–12h after inoculation. Inorganic pyrophosphatase was not associated with extracellular alkaline phosphatase, but it was detected intracellularly. Ninety-five percent of the alkaline phosphatase was detected extracellularly.     

A highly thermostable amylase from a newly isolated thermophilic Bacillus sp. WN11

A thermostable amylase-producing Bacillus sp. WN11 was isolated from Wondo Genet hot spring in Ethiopia. The enzyme had a temperature optimum of 75–80 °C. Over 80% of its peak activity was in the pH range of 5–8, with an optimum at 5·5. Thermal stability of the enzyme at 105 °C was higher with the addition of starch. The stabilizing effect of starch was concentration-dependent, showing better stability with increasing concentration of starch. At liquefying temperature (105 °C), addition of Ca²⁺ did not result in further improvement of the stabilizing effect of starch. This indicates that in the presence of starch, WN11 amylase does not require Ca²⁺ as a stabilizer at liquefying temperatures as high as 105 °C.     

Co-inoculation with Bacillus sp. CECT 450 improves nodulation in Phaseolus vulgaris L.

The strain Bacillus sp. CECT 450 increased nodulation on bean (Phaseolus vulgaris L.) when co-inoculated with Rhizobium tropici CIAT 899. This positive effect occured under controlled conditions on perlite-vermiculite, sand, or in a mixture of soil and sand. This increase was also observed in a field assay. Nodulation kinetic studies suggested that the synergistic effect is pronounced during the latter stages of cultivation. In contrast, the same bacteria co-inoculated with Bradyrhizobium japonicum USDA 110 reduced nodulation on soybean (Glycine max (L.) Merr.). Inoculation with Bacillus sp. CECT 450 alone had no effect on bean plants, but reduced root growth in soybean. The survival of Bacillus sp. CECT 450 on inoculated seeds was high, even when inoculated seeds were maintained for several months at room temperature.     

Bacillus alcalophilus n. sp.

Samenvatting Mede naar aanleiding van de gunstige ervaringen, die Doorenbos in de practijk opdeed met èèn der door ons voorgestelde nieuwe electieve cholera-voedingsbodems (“glycocollplaat”), wordt getracht ook een aankweek-voedingsbodem, electief voor vibrionen, samen te stellen. Hierbij worden echter bezwaren, o.a. van physico-chemischen aard, ondervonden, terwijl bovendien soms uit faeces van normale individuen in enkele der onderzochte vloeibare aankweek-voedingsbodems (sterk alcalisch haemoglobine-carbonaat-peptonwater, sterk alcalisch carbonaat-peptonwater van pH-ca. 10) electief ook een Gram-positieve sporevormende bacterie wordt aangekweekt. Hieruit kan deze bacil dan op de door ons beschreven choleravoedingsbodems en op Dieudonnè-platen worden ge?soleerd. Dadelijk uit faeces gelukt de isolatie dezer microbe op choleravoedingsbodems niet; evenmin door aankweek op peptonwater van “gewone” pH. In de praktijk geeft deze bacil dus geen moeilijkheden bij het onderzoeken van faeces op vibrionen. Kenmerkend voor deze bacil is, dat deze alleen op en in sterk alcalische voedingsbodems groeit, niet bij zwakke alcaliteit of nog lagere pH. Daar dit micro-organisme, voor zoover ons bekend nog niet eerder beschreven is, stellen wij voor hem “Bacillus alcalophilus n. sp.” to noemen. De bacil is beweeglijk en splitst eiwit (gelatine, haemoglobine) in sterk alcalisch milieu. Summary Also in view of the favourable results whichDoorenbos has obtained in practical cases with one of the new elective cholera media (glycocoll-plate) proposed by us, an attempt has been made to compose a culture medium for the elective enrichment of elective vibrions. Difficulties of a physico-chemical nature are, however, met with in doing so moreover in some of the liquid culture media (highly alkaline haemoglobine-carbonate-peptone water, highly alkaline carbonate-peptone water of pH ca. 10) from the faeces of normal individuals which have been examined, a Gram-positive sporeforming bacterium is cultivated. From this the bacillus may then be isolated on the cholera cultivation media described by us and on Dieudonnè plates. Directly from faeces this microbe does not grow on elective cholera culture media, nor by enrichment in peptone water of “ordinary” pH. So in practice this bacillus does not present any difficulty when examining faeces for vibrions. It is typical of this bacillus that it only grows in and on highly alkaline culture media, and not in cases where the medium is of weak alkalinity or still lower pH. As this micro-organism has not, as far as we know, been previously described, we propose to name it “Bacillus alcalophilus n.sp.” The bacillus is motile and digests albumen (gelatine, haemoglobine) in a strong alkaline medium.     

A novel enzyme of Bacillus sp. 217C-11 that produces inulin from sucrose

We found a bacterium that converts sucrose to a useful material, using about 6,000 samples of bacteria isolated from soil. This bacterium, Bacillus sp. 217C-11, was identified according to Bergey's manual, and produced a highly efficient enzyme that converted sucrose into inulin. So, the enzyme was purified to homogeneity through five chromatographic steps, to identify its enzymatic properties. The molecular mass of the enzyme was estimated to be 45,000, and this enzyme was a monomer protein (by SDS-PAGE). The optimum pH and temperature of this enzyme were 7-8 and 45-50 degrees C, respectively. The enzyme reacted only with sucrose, but did not with other disaccharides, fructooligosaccharides and inulin. This paper will show that our enzyme is a novel one, which is different from the other well-known enzymes concerned in inulin production.     

Characterization of a novel endo-levanase and its gene from Bacillus sp. L7

A novel endo-levanase producing bacterium belonging to the Bacillus family has been isolated from soil. The enzyme was characterized and found to have no exo-β-fructofuranosidase activity. The endo-levanase gene was cloned and sequenced. Homology searches have shown that the C-terminal domain of the enzyme is homologous to a number of known β-fructofuranosidases, including exo-levanase from Bacillus subtilis and yeast invertases. The N-terminal region of the endo-levanase which is homologous to the C-terminal sequence of the B. subtilis levanase appears to be a levan-binding domain.     

芽孢杆菌WS-3L淀粉酶的纯化和特性的研究

芽孢杆菌WS-3L产生的α-淀粉酶AmyL经过多步纯化,酶的回收率为15.5%,比活提高了345倍.该淀粉酶能够有效水解淀粉生成麦芽寡糖.酶的最适反应温度为45℃,最适反应pH为6.5,在pH 7.0～8.0,40℃以下酶活较稳定;离子Cu2 、NH4 、Ag 、Hg 和EDTA、SDS对酶活力有显著抑制作用,而其他一些常见金属离子如Na 、K 则对酶活影响不大.AmyL对可溶性淀粉、直链淀粉、支链淀粉的Km值和Vmax分别为2.81 mg/mL、8.37 mg/mL、1.80 mg/mL和11.67 μmol/(min·mL)、10.00μmol/(min·mL)、13.33 μmol/(min·mL),表明支链淀粉是该酶的理想水解底物.玉米淀粉对AmyL有很高的吸附率,预示这可以作为酶快速固定的一个简易方法应用到实际生产中.     

Weathering of Basalts by Aspergillus sp. FS-4 Strain: Glass Compositions are Prone to Weathering

Abstract

Mechanisms underlying microbe-induced dissolution of basalt and its effect on glass composition remain unclear. Two powdered basalts, with a glass composition of 27?vol% (Basalt-T) and 18?vol% (Basalt-F), respectively, were inoculated with Aspergillus sp. FS-4 in Czapek medium. Changes in pH, siderophores and main dissolved elements were examined. Basalt-F led to a greater decrease in pH and higher production of siderophores than Basalt-T mainly due to the fungi activity. Element dissolution was 2 to 3 fold higher in Basalt-T than in Basalt-F, and the dissolution was non-stoichiometric for both basalts. FS-4 appears important for basalt weathering. Composition of geologic material had a considerable impact on weathering. Materials with a higher glass percentage are prone to weathering. Further quantitative evaluation is required.     

Molecular imprinting of cyclodextrin glycosyltransferases from Paenibacillus sp. A11 and Bacillus macerans with gamma-cyclodextrin

Cyclodextrin glycosyltransferase catalyzes the formation of a mixture of cyclodextrins from starch by an intramolecular transglycosylation reaction. To manipulate the product specificity of the Paenibacillus sp. A11 and Bacillus macerans cyclodextrin glycosyltransferases towards the preferential formation of gamma-cyclodextrin (CD(8)), crosslinked imprinted proteins of both cyclodextrin glycosyltransferases were prepared by applying enzyme imprinting and immobilization methodologies. The crosslinked imprinted cyclodextrin glycosyltransferases obtained by imprinting with CD(8) showed pH and temperature optima similar to those of the native and immobilized cyclodextrin glycosyltransferases. However, the pH and temperature stability of the immobilized and crosslinked imprinted cyclodextrin glycosyltransferases were higher than those of the native cyclodextrin glycosyltransferases. When the catalytic activities of the native, immobilized and crosslinked imprinted cyclodextrin glycosyltransferases were compared, the efficiency of the crosslinked imprinted enzymes for CD(8) synthesis was increased 10-fold, whereas that for cyclodextrin hydrolysis was decreased. Comparison of the product ratios by high-performance anion exchange chromatography showed that the native cyclodextrin glycosyltransferases from Paenibacillus sp. A11 and Bacillus macerans produced CD(6) : CD(7) : CD(8) : > or = CD(9) ratios of 15 : 65 : 20 : 0 and 43 : 36 : 21 : 0 after 24 h of reaction at 40 degrees C with starch substrates. In contrast, the crosslinked imprinted cyclodextrin glycosyltransferases from Paenibacillus sp. A11 and Bacillus macerans produced cyclodextrin in ratios of 15 : 20 : 50 : 15 and 17 : 14 : 49 : 20, respectively. The size of the synthesis products formed by the crosslinked imprinted cyclodextrin glycosyltransferases was shifted towards CD(8) and > or = CD(9), and the overall cyclodextrin yield was increased by 12% compared to the native enzymes. The crosslinked imprinted cyclodextrin glycosyltransferases also showed higher stability in organic solvents, retaining 85% of their initial activity after five cycles of synthesis reactions.