硅酸盐细菌的分离及溶磷、解钾和脱硅活性研究

利用硅酸盐细菌分离培养基从云南省文山州砚山红舍克铝土矿区分离出一株硅酸盐细菌YSHSK13,通过形态、生理生化特征及16S rRNA基因测序分析,将其初步鉴定为节杆菌属(Arthrobacter sp.)的菌株;对其溶磷、解钾和脱硅活性的研究分析表明,在溶磷、解钾和脱硅培养基中培养10 d和20 d后,可溶性硅、钾和磷的含量分别比典型标准硅酸盐细菌菌株增加3.16、1.77、1.04倍和4.48、2.24、2.08倍,结果表明该菌株具有较强的溶磷、解钾和脱硅活性。     

高效钾长石分解菌株的筛选、鉴定及解钾活性研究

目的 筛选高效的钾长石分解细菌,并进行初步鉴定和解钾活性测定.方法 采集湖南省桂东县钾长石开采区土壤,利用钾长石为唯一钾源的选择性培养基筛选分离解钾细菌,通过摇瓶释钾试验复筛高效解钾菌株;同时,利用ICP-OES测定解钾菌的释钾效率.采用形态特征观察、生理生化特性检测和基于16S rRNA基因序列的系统发育分析初步鉴定高效解钾菌株.结果 分离获得11株生长良好的钾长石分解细菌,其中菌株JKC1、JKC2、JKC5和JKC7的解钾能力较强,解钾率分别为11.00％、11.50％、12.70％和11.70％.经初步鉴定JKC1为Bacillus megaterium,JKC2为Bacillus aryabhattai,JKC5为Azotobacter chroococcum,JKC7为Microbacterium trichothecenolyticum.结论 菌株JKC1、JKC2、JKC5和JKC7是高效的钾长石分解菌,可作为微生物浸矿(钾长石)机制研究的候选菌株.     

低氧条件适用性硅酸盐细菌的分离及对水稻幼苗生长影响

从湖南地区水稻田土样中分离适用于水田作物的硅酸盐细菌, 得到在亚历山大罗夫培养基上符合硅酸盐细菌菌落特征的菌株1 株, 编号为菌株s-3。通过一系列的生理生化试验, 菌体形态及菌落特征的观察以及16S rDNA 序列分析对菌株s-3 进行菌种鉴定, 初步鉴定菌株s-3 属于硅酸盐细菌的一种--环状芽孢杆菌(Bacillus circulans)。通过在不同供氧条件下测定菌株s-3 的解钾能力, 发现在微氧条件下菌株s-3 仍能保持较高的活性。通过水培试验, 测定菌株s-3 作为生物肥料对水稻植株幼苗的促生长能力, 发现施用菌株s-3 培养液后, 可大幅增加植株的钾总量, 促进植株生长, 具有作为生物肥料进一步研究的潜力。     

硅酸盐细菌HM8841菌株解钾作用的研究

早在三十年代苏联学者从土壤中分离到硅酸盐细菌,并测定解钾强度,培养5d分离出来的钾量为原硅酸盐中含量的15.9％,同时在不同土壤和不同农作物上进行了盆栽和田间试验,证明对农作物生长有较好的促进作用和增产效果。也有一些学者认为硅酸盐细菌的作用是刺激作用,为农作物补充钾素营养是微不足道的。 作者分离到硅酸盐细菌HM8841菌株,经工业发酵研制成菌剂。该菌剂经几年的大面积推广应用,在缺钾土壤上对各种农作物均表现出较好的增产效果。尤其在土壤中速效钾严重不足的情况下,施用该菌剂增产更加明显。这充分说明土壤缺少速效钾是限制农作物增产的重要原因。HM8841菌株能为农作物提供一定量的钾素使农作物提高产量。但其解钾能力有多高,能提供多少钾素,需要进一步研究。为此作者采取不同的方法,不同的底物对HM8841菌株的解钾效能进行了研究。     

钾长石矿区土壤解钾菌的分离与多样性

目的获得高效钾长石分解细菌及分析钾长石矿区土壤中解钾菌的多样性。方法采集湖南省吉首市钾长石矿区土壤,用钾长石为唯一钾源的选择性培养基筛选分离解钾细菌,采用形态特征观察、生理生化特性检测和基于16S rRNA基因序列的系统发育分析初步鉴定解钾菌株,并测定菌株摇瓶条件下的解钾能力。结果分离获得38株钾长石分解细菌,分别属于13个属,其中13株菌为剑菌属,4株菌为芽胞杆菌属。菌株L17的发酵液中有效钾含量最高,为87.66 mg/L,是对照组的15.8倍。结论钾长石矿区土壤细菌种类有较好的多样性,其中剑菌属和芽孢杆菌属为优势种群,Mitsuaria属的L17解钾能力最强。     

土壤中钾细菌的筛选及筛选底物加入量对其数量的影响

研究土壤中加入不同数量的钾长石粉后其解钾细菌数量的变化,同时初步筛选钾细菌,为进一步研究土壤中解钾细菌的分离、筛选及其鉴定提供参考。测试土壤中加入不同数量的钾长石粉后其解钾细菌数量,对结果进行比较分析;利用四苯硼钠法测试菌株解钾能力,并进行初步筛选。结果表明,按钾细菌数量由多到少的顺序,筛选底物加入量为0.2%0.1%0.4%空白对照;筛选菌株的解钾效率高于K01的共14株,高于K02的共9株。解钾效率最高的菌株5.15-2解钾效率为35.4%,较外购生产菌种K01高13倍,与本实验室保存菌种K02相比高3.6倍。     

土壤中解钾菌K02的筛选、鉴定及培养条件优化

解钾微生物是能够在土壤或纯培养条件下,将含钾矿物如长石、云母等不能被作物吸收利用的矿物态钾分解,并产生水溶性钾的微生物。利用以钾长石粉为唯一钾源的硅酸盐细菌选择性培养基,采用梯度稀释分离法平板划线对番茄土壤中的钾细菌进行筛选。利用原子吸收火焰分光光度法测定钾细菌培养液中可溶性钾的含量,筛选高效解钾菌株。通过形态观察和16S rDNA序列GenBank比对,同时利用clustalx和MEGA 4.0等相关软件构建系统进化树对解钾能力最强的菌株K02进行鉴定,初步鉴定该解钾菌为胶质类芽胞杆菌(Paenibacillus mucilaginosus);利用单因素试验法对K02菌株培养基组分及发酵条件进行优化,初步确定菌株K02最佳培养基组分以解钾复筛培养基为基础,其碳源、氮源及无机盐以可溶性淀粉1%、酵母膏0.2%、K_2HPO_40.05%为最佳;菌株K02最佳发酵条件:培养温度为30℃,培养时间为48 h,培养基装量为50 mL/250 mL,培养基初始pH值为7.5,接菌量为5%,这一结果为解钾菌肥的研制和生产提供参考。     

硅酸盐细菌LJP13的分离鉴定及生长特性

从‘凤丹’牡丹根际土壤中分离到一株硅酸盐细菌,编号为LJP13。在28℃、180 r/min条件下发酵培养7 d后,发酵液中水溶性速效钾含量达到1.3μg/mL,从而确定LJP13具有解钾活性,并定性为硅酸盐菌株。通过形态特征观察、生化特性测定和16S rDNA聚类分析,初步将菌株LJP13鉴定为芽孢杆菌属Bacillus菌株,暂命名为Bacillus sp.LJP13。对菌株LJP13的生长特性研究表明,菌株对几种底物的发酵产气能力顺序是麦芽糖葡萄糖直链淀粉乳糖甘露醇支链淀粉;在碳同化利用能力上蔗糖为最适底物。对不同氮源的同化能力顺序为酵母膏硝酸钾蛋白胨硫酸铵尿素。菌株的最适生长温度为28~32℃,适生pH为7。LJP13菌株可作为牡丹专用菌肥开发的备用菌株。     

一株硅酸盐细菌的鉴定及其紫外线诱变育种

从油用牡丹的根际土壤中分离到一株硅酸盐细菌,将其编号为CLM8。采用火焰光度法对该菌株在30℃、160 r/min条件下发酵培养7d的发酵上清液中水溶性钾素含量进行测定,最高含量达到0.9μg/mL,故将CLM8定性为硅酸盐菌株。综合CLM8的显微观察特征、生化特征和16S rRNA序列的克隆分析结果,确定菌株CLM8为成团泛菌属中的一个种,暂命名为Pantoea sp.CLM8。为提高野生菌株CLM8的解钾活性,通过对其紫外线诱变选育,筛选得到活性较高的突变株CLM8-B,其在30℃、160 r/min条件下发酵培养过程中,发酵上清液中水溶性钾素含量最高达到1.7μg/mL,是出发菌株CLM8的1.9倍。CLM8可作为油用牡丹及其它作物生物钾肥开发的备用菌株。     

一株蜂粮源拮抗细菌的分离鉴定及其抑菌物质特性

【目的】为发掘和利用蜂粮中拮抗菌资源,对分离获得的拮抗细菌菌株PC2进行分类鉴定,并测定其发酵液抑菌物质基本特性。【方法】采用改良牛津杯双层平板法测定菌株发酵液抑菌谱及温度、p H、紫外线和蛋白酶对其抑菌活性稳定性的影响,菌株鉴定结合形态学、生理生化特征和16S r RNA基因序列分析,硫酸铵沉淀法和盐酸沉淀有机溶剂提取法进行抑菌活性物质的初步分离。【结果】从3种蜂粮中分离筛选得到17株拮抗菌株,其中1株细菌PC2以马铃薯葡萄糖液体培养基发酵制备的无菌发酵液对7种供试菌株具有较强抑制作用,经形态、生理生化特征及16S r RNA基因序列分析,将其初步鉴定为解淀粉芽胞杆菌(Bacillus amyloliquefaciens)。菌株发酵液抑菌活性对温度、酸和紫外线具有较强的稳定性,对蛋白酶K、胃蛋白酶、碱性蛋白酶处理敏感。菌株发酵液存在抑菌蛋白和脂肽类物质。【结论】菌株PC2在食品保鲜和农业生防中具有潜在的开发应用价值。     

Isolation of Paenibacillus sp. and Assessment of its Potential for Enhancing Mineral Weathering

One mineral-solubilizing bacterial strain designated KT was isolated from a soil in Henan Province, China. The full-length 16S rRNA gene sequence showed 95.2 to 96.7% similarity to the bacteria of the genus Paenibacillus, indicating that the strain KT belonged to the genus Paenibacillus. The potential of this strain to release potassium from silicate minerals was investigated using a potassium-bearing rock as the sole source of potassium to support its growth. After inoculation for 7 days, the concentrations of water-soluble Al, Ca and Fe released from the potassium-bearing rock in active bacterial culture were higher than those from the control with autoclaved inoculum, but the concentration of water-soluble K in active bacterial culture was similar to that in the control. The concentrations of HNO₃-extractable Al, Ca, Fe and K from the bacterial culture were also higher than those in the control. These results showed strain KT was able to release potassium from potassium-bearing rock. These results have important implications for extraction of potassium from rocks to support plant growth.     

Characterization of a novel thermophilic, cellulose-degrading bacterium Paenibacillus sp. strain B39

Aims:  The aims of this study were to identify and characterize the novel thermophilic, cellulose-degrading bacterium Paenibacillus sp. strain B39.
Methods and Results:  Strain B39 was closely related to Paenibacillus cookii in 16S rRNA gene sequence. Nonetheless, this isolate can be identified as a novel Paenibacillus sp. with respect to its physiological characteristics, biochemical reactions, and profiles of fatty acid compositions. A cellulase with both CMCase and avicelase activities was secreted from strain B39 and purified by ion-exchange chromatography. By sodium dodecyl sulfate–polyacrylamide gel electrophoresis analysis, the molecular weight of B39 cellulase was determined as 148 kDa, which was much higher than other cellulases currently reported from Paenibacillus species. The enzyme showed a maximum CMCase activity at 60°C and pH 6·5. Addition of 1 mmol l⁻¹ of Ca²⁺ markedly enhanced both CMCase and avicelase activities of the enzyme.
Conclusions:  We have identified and characterized a novel thermophilic Paenibacillus sp. strain B39 which produced a high-molecular weight cellulase with both CMCase and avicelase activities.
Significance and Impact of the Study:  Based on the ability to hydrolyse CMC and avicel, the cellulase produced by Paenibacillus sp. strain B39 would have potential applications in cellulose biodegradation.     

Identification of endophytic bacterial strain RSE1 from seeds of super hybrid rice Shenliangyou 5814 (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.,) and evaluation of its antagonistic activity

Endophytes are related with health and growth of plants. In this study, the endophytic bacterial strain RSE1 was isolated from seeds of super hybrid rice Shenliangyou 5814 (Oryza sativa L.,). Strain RSE1 was identified as Paenibacillus polymyxa by polyphasic taxonomy identification. Through the antagonistic test with the pathogenic strain of rice false smut, Ustilaginoidea oryzae CICC 2710, it showed that the strain RSE1 had an effective antagonistic activity against this pathogen. The draft genome of strain RSE1 was sequenced by Illumina HiSeq 2000, and 3 CDSs for glucanase gene were annotated and correlated to antagonistic activity. Using specific primers to amplify the biocontrol gene in glucanase family, β-1,3-1,4-glucanase gene (gluB) was found. This study laid a scientific foundation for developing and utilizing biological bio-control bacteria agent preventing the suffering from rice false smut.     

Survival and naphthalene-degrading activity of Rhodococcus sp. strain 1BN in soil microcosms

AIMS: The survival and activity of Rhodococcus sp. strain 1BN, inoculated into naphthalene-contaminated sandy-loam soil microcosms, were studied using classical and molecular methods. METHODS AND RESULTS: The naphthalene-degrading activity of 1BN in microcosms was examined through viable counts, CO2 production and naphthalene consumption, while its survival after inoculation was monitored by detecting the contemporary presence of alkane and naphthalene degradative genes and by analysing the 16S rDNA specific restriction profile. The inoculation of 1BN did not significantly enhance naphthalene degradation in the naphthalene-contaminated native soil, where 1BN maintained its catabolic activity also when in the presence of indigenous microflora. Instead the rate of naphthalene degradation by the inoculated 1BN was greater in sterile naphthalene-contaminated soil. The level of 1BN was only slightly higher after inoculation regardless of whether indigenous naphthalene-degrading bacteria were present or not and 1BN remained viable even when the substrate was depleted. CONCLUSIONS: This study documents the colonization and growth of 1BN in a non-sterile, naphthalene-added, sandy-loam soil having an active indigenous naphthalene-degrading population. SIGNIFICANCE AND IMPACT OF THE STUDY: An active and well-established naphthalene-degrading bacterial population in the native soil did not hamper the survival of the introduced 1BN that, through its activity, enhanced the mineralization rate of naphthalene.     

Purification and characterization of a 1,2-dihydroxynaphthalene dioxygenase from a bacterium that degrades naphthalenesulfonic acids.

1,2-Dihydroxynaphthalene dioxygenase was purified to homogeneity from a bacterium that degrades naphthalenesulfonic acids (strain BN6). The enzyme requires Fe2+ for maximal activity and consists of eight identical subunits with a molecular weight of about 33,000. Analysis of the NH2-terminal amino acid sequence revealed a high degree of homology (22 of 29 amino acids) with the NH2-terminal amino acid sequence of 2,3-dihydroxybiphenyl dioxygenase from strain Pseudomonas paucimobilis Q1. 1,2-Dihydroxynaphthalene dioxygenase from strain BN6 shows a wide substrate specificity and also cleaves 5-, 6-, and 7-hydroxy-1,2-dihydroxynaphthalene, 2,3- and 3,4-dihydroxybiphenyl, catechol, and 3-methyl- and 4-methylcatechol. Similar activities against the hydroxy-1,2-dihydroxynaphthalenes were also found in cell extracts from naphthalene-degrading bacteria.     

离子注入结合紫外线及~(60)Co-γ射线诱变选育碱性果胶酸裂解酶高产菌株的研究

以碱性果胶酸裂解酶产生菌芽孢杆菌WZ008为出发菌株,经形态鉴定和16S鉴定为类芽孢杆菌,命名为Paenibacillus sp.WZ008,通过N~+注入诱变、紫外线诱变、~(60)Co-γ射线诱变等多次反复诱变,选育得到一株产碱性果胶酸裂解酶性能稳定且酶活明显提高的突变株,其酶活为97.8U/mL,比出发菌株产碱性果胶酸裂解酶能力提高了1.04倍。     

Draft Genome Sequence of Paenibacillus polymyxa OSY-DF, Which Coproduces a Lantibiotic, Paenibacillin, and Polymyxin E1

Paenibacillus polymyxa OSY-DF is a Gram-positive rod-shaped bacterium isolated from a fermented vegetable food. This bacterial strain displays potent antimicrobial activities against Gram-positive and Gram-negative pathogenic bacteria, attributed to the production of the lantibiotic paenibacillin and the colistin peptide polymyxin E1. Here we report the draft genome sequence of Paenibacillus polymyxa OSY-DF.     

人参内生细菌ge21菌株的鉴定及抑菌活性测定

从人参根内分离到一株细菌ge21,经形态特征观察、生理生化特性测定和16SrDNA序列分析,表明该菌株同已知菌Paenibacillus granivorans的相似性最高,为96.89%,初步认定为类芽孢杆菌属的一个潜在新种。抑菌试验结果表明,该菌株对尖孢镰刀菌(Fusarium oxysporium)、寄生疫霉菌(Phytophthora parasitica)、剑麻炭疽病菌(Colletotrichum agaves)、烟草赤星病菌(Alternaria longipes)、稻瘟病菌(Pyricularia oryzae)、人参立枯病菌(Rhizoctonia solani)、人参疫病菌(Phytophthora cactorum)和人参菌核病菌(Sclerotinia schinseng)均有一定的抑菌活性。     

Isolation and identification of a Paenibacillus polymyxa strain that coproduces a novel lantibiotic and polymyxin

A new bacterial strain, displaying potent antimicrobial properties against gram-negative and gram-positive pathogenic bacteria, was isolated from food. Based on its phenotypical and biochemical properties as well as its 16S rRNA gene sequence, the bacterium was identified as Paenibacillus polymyxa and it was designated as strain OSY-DF. The antimicrobials produced by this strain were isolated from the fermentation broth and subsequently analyzed by liquid chromatography-mass spectrometry. Two antimicrobials were found: a known antibiotic, polymyxin E1, which is active against gram-negative bacteria, and an unknown 2,983-Da compound showing activity against gram-positive bacteria. The latter was purified to homogeneity, and its antimicrobial potency and proteinaceous nature were confirmed. The antimicrobial peptide, designated paenibacillin, is active against a broad range of food-borne pathogenic and spoilage bacteria, including Bacillus spp., Clostridium sporogenes, Lactobacillus spp., Lactococcus lactis, Leuconostoc mesenteroides, Listeria spp., Pediococcus cerevisiae, Staphylococcus aureus, and Streptococcus agalactiae. Furthermore, it possesses the physico-chemical properties of an ideal antimicrobial agent in terms of water solubility, thermal resistance, and stability against acid/alkali (pH 2.0 to 9.0) treatment. Edman degradation, mass spectroscopy, and nuclear magnetic resonance were used to sequence native and chemically modified paenibacillin. While details of the tentative sequence need to be elucidated in future work, the peptide was unequivocally characterized as a novel lantibiotic, with a high degree of posttranslational modifications. The coproduction of polymyxin E1 and a lantibiotic is a finding that has not been reported earlier. The new strain and associated peptide are potentially useful in food and medical applications.     

Characterization of pectin lyase produced by an endophytic strain isolated from coffee cherries

AIMS: The effect of endophytic bacterial activity on the quality of coffee beverage was studied. METHODS AND RESULTS: A survey of the micro-organisms in coffee cherries was performed before harvesting, and their growth on the main nutrients available in coffee cherries was determined in vitro. CONCLUSION: Many endophytic bacteria were isolated from surface-sterilized coffee cherries. One of the pectinolytic strains was physiologically and phenotypically characterized, and was tentatively identified by partial 16S rDNA sequencing as Paenibacillus amylolyticus. This endophytic strain produced an extracellular pectinase with maximal activity at 40 degrees C and pH 7.9, and was thermostable up to 45 degrees C. EDTA and metal ions had little effect on pectin lyase activity. Km and Vmax values were 4.6 mg ml(-1) and 94.0 10(-8) mol min(-1) ml(-1), respectively. SIGNIFICANCE AND IMPACT OF THE STUDY: Pectin lyases have been found in fungi but rarely in bacteria, and this isolate is a promising tool for regulation studies of these enzymes.