明党参内生菌的分离鉴定及其对金黄色葡萄球菌的抑制作用

研究郑州市栽培明党参内生菌资源状况,筛选出对金黄色葡萄球菌具有拮抗作用的内生菌。采用平板培养法从明党参中分离内生细菌,通过混菌法和牛津杯双层平板扩散法筛选出对金黄色葡萄球菌具有明显抑制作用的菌株,并通过培养特征、形态特征、生化特征、全自动微生物质谱检测、16S rDNA基因序列分析和系统进化树分析对菌株进行了分类学鉴定。在明党参中共分离到29株内生细菌,通过初筛和复筛后,筛选出对金黄色葡萄球菌具有明显抑制作用的4株菌株NMY1、NMJ10、NMG10、PMG3,其中NMJ10对金黄色葡萄球菌的抑菌圈直径达20.3 mm,经鉴定菌株NMJ10和菌株PMG3为芽胞杆菌属新菌种(Bacillus spp.),菌株NMY1为枯草芽胞杆菌(B.subtilis),菌株NMG10为假蕈状芽胞杆菌(B.pseudomycoides)。明党参内生菌具有多样性,其中芽胞杆菌属为优势菌株,有拮抗金黄色葡萄球菌活性高的菌株存在,其代谢产物对金黄色葡萄球菌有较高抑制作用,为抗菌药源成分的开发提供了新的来源。     

关防风根腐病拮抗细菌筛选与鉴定

【背景】由木贼镰刀菌[Fusarium equiseti(Corda)Sacc.]引起的关防风根腐病,是近年来导致关防风产量及质量下降的主要土传病害之一。生物防治因其对环境安全及人畜无害等优势,成为目前植物病害防治的一种有效手段。【目的】挖掘关防风根际土壤中对木贼镰刀菌具有良好拮抗作用的生防菌株。【方法】采用稀释平板法分离根际土壤细菌;用滤纸片法和牛津杯法对拮抗细菌进行筛选和抑菌谱检测;用抗生素标记法标记拮抗细菌并测定其定殖能力;通过盆栽实验研究其对关防风根腐病的防效;通过形态学、生理生化特征及16S rRNA基因序列确定分类学地位。【结果】从健康关防风根际土壤中分离纯化了157株细菌,筛选获得对关防风根腐病菌具有显著抑菌作用的拮抗细菌SC-119,无菌滤液的抑菌率可达68.53%,而且兼具广谱抑菌能力和良好的定殖能力;盆栽实验表明,菌株SC-119对关防风根腐病的防治效果达到67.39%,其相对防效较接种哈茨木霉菌剂、枯草芽胞杆菌菌剂和代森锰锌分别提高了29.03%、32.26%和16.13%;对菌株SC-119进行分类学鉴定,确定其为萎缩芽胞杆菌(Bacillus atrophae...     

番木瓜果实内生细菌MGP3 菌株的鉴定及拮抗作用

【目的】从番木瓜果皮内筛选具有较强拮抗活性的内生细菌防治番木瓜采后炭疽病和疫霉病,以减少果实采后病害带来的损失。【方法】采用稀释分离和平板抑菌圈法进行内生细菌的分离筛选,结合形态特征、生理生化特性及16S rDNA部分序列同源性分析对菌株进行鉴定,菌株经利福平诱抗处理后田间接种到果树树干上,测定内生菌的定殖动态,采用采前和采后生防试验测定菌株对番木瓜炭疽病和疫霉病的生防效果。【结果】从番木瓜果皮中分离筛选到一株具有拮抗活性的内生细菌MGP3,对10种病原菌有较强的拮抗作用,鉴定该细菌为铜绿假单胞菌(Pseudomonas aeruginosa,登录号JF708186),MGP3可进入番木瓜叶片、叶柄和果皮中定殖。MGP3对采后番木瓜炭疽病和疫霉病的防治效果分别达到50%和71%;除苗期外,采前4个不同时期经MGP3菌液处理可以显著降低采收后果实炭疽菌的潜伏侵染率和炭疽病的病情指数。【结论】番木瓜内生拮抗细菌MGP3具有潜在的生防应用价值。     

解淀粉芽胞杆菌HR-2的分离鉴定及对水稻稻瘟病菌的拮抗效果

稻瘟病是一种严重威胁全球粮食安全的水稻真菌病害。本研究采用平板对峙法从湖南岳阳地区筛选出1株对水稻稻瘟病菌具有高效拮抗活性的菌株HR-2。通过形态特征验证、16S rRNA、gyrA和tuf基因序列比对分析,鉴定该菌株为解淀粉芽胞杆菌(Bacillus amyloliquefaciens)。抑菌广谱性测定结果表明菌株HR-2对水稻稻瘟病菌、油菜菌核病菌和番茄青枯病菌均有较强的拮抗效果,抑制率分别为82.34%、72.32%和72.64%。盆栽试验结果显示,菌株HR-210倍稀释液和100倍稀释液对水稻稻瘟病的防治效果分别达到63.81%和40.77%,其中HR-210倍稀释液的防效与50 mg/L嘧菌酯的防效相当。试验结果表明解淀粉芽胞杆菌菌株HR-2抑菌谱广,尤其对水稻稻瘟病具有显著的防治效果,可为抗稻瘟病生物防治药剂的开发奠定基础。     

耐高温沼渣降解菌的筛选及复合菌剂降解效果研究

分离筛选耐高温纤维素降解细菌，构建可快速降解沼渣的复合菌剂，缩短堆肥腐熟时间。采用高温富集筛选法，从宁夏中卫市秸秆堆和畜禽粪便中分离筛选耐高温细菌，经羧甲基纤维素钠水解试验和滤纸条崩解试验复筛获得具有纤维素降解能力的菌株。通过纤维素酶和木质素酶活性比较，确定目标菌株并进行分类鉴定。开展拮抗性试验和菌种复配，构建复合型菌剂；经沼渣堆肥场地试验验证复合菌剂的促腐熟效果。本研究筛选获得2株耐高温纤维降解细菌——蜡样芽胞杆菌(Bacillus cereus)W44菌株和解蛋白芽胞杆菌(Bacillus proteolyticus)X51菌株，纤维素酶活性分别为34 812.69 U/L和34 159.10 U/L。比较菌株W44/X51配比分别为1:1、1:2和2:1的复合菌剂对沼渣堆肥的促腐熟效果，添加不同的复合菌剂均缩短了沼渣堆肥腐熟时间，其中1:2处理腐熟时间最短，较CK提前18 d。蜡样芽胞杆菌W44和解蛋白芽胞杆菌X51按1:2配比的复合菌剂可用于沼渣快速堆肥发酵。     

1株拮抗植物病原真菌的细菌筛选及其抑菌机理的初步研究

从上海市郊农作物根际土壤中共分离纯化得到276株细菌,利用平板对峙法筛选出1株对多种植物病原真菌具有较强拮抗作用的菌株。经过形态学观察、生理生化特征以及16S rDNA的同源性分析,初步鉴定该菌株为桔黄假单胞菌(Pseudomonas aurantiaca),该菌株的16S rDNA序列已在GenBank中注册,登录号为GQ358919。通过显微镜观察,发现JD37菌株的主要抑菌机理是通过产生拮抗物质造成病原真菌菌丝体断裂、扭曲、畸形等异常生长现象。该菌产生的拮抗物质对酸、碱较为稳定,但对高温敏感。     

解淀粉芽胞杆菌BaX030的分离鉴定及抗菌功能

摘要:【目的】芽胞杆菌是微生物活性物的重要来源,从全国各地采集的72份土壤样品中分离出339株芽胞杆菌,研究各菌株抑菌活性,分离纯化抑菌活性物,为丰富芽胞杆菌菌种资源和微生物次级代谢物的挖掘奠定实际应用基础。【方法】采用水浴加热和稀释平板涂布等方法从河南花生地采集的土壤中筛选得到一株具有很强抑菌活性的芽胞杆菌,结合形态观察、生理生化特征和16S rRNA基因序列同源性比对分析,对该菌株进行鉴定。丙酮沉淀、葡聚糖凝胶柱层析、C18反相柱层析得到Bacillus amyloliquefaciens X030抑菌活性物,LC-MS/MS鉴定其分子量。利用滤纸片扩散法和平板对峙培养法测定抑菌谱及拮抗性质。【结果】筛选分离得到一株解淀粉芽胞杆菌,归类并命名为解淀粉芽胞杆菌Bacillus amyloliquefaciens X030。BaX030对金黄色葡萄球菌(Staphylococcus aureus)、白色念珠菌(Candida albicans)、酵母菌( Saccharomycetes)有较强抑制效果,对水稻稻瘟病菌(Pyriculariaoryzae)、辣椒尖胞炭疽病菌(Chili pointed cell anthrax)、枇杷炭疽病菌(Gloeosporium eriobotryae speg)、烟草黑胫病菌(Phytophthora parasitica)有良好拮抗活性。初步确定BaX030产生的抑菌活性物为多肽类化合物。【结论】分离得到的B． amyloliquefaciens X030产生了一个对病原细菌具有较强抑制作用的多肽,同时该菌株在拮抗植物病原真菌方面也有明显的效果。     

一株抗水稻纹枯病菌的解淀粉芽胞杆菌分离与鉴定

【目的】筛选对水稻纹枯病菌(Rhizoctonia solani)具有强拮抗作用的细菌菌株。【方法】用指示菌法筛选拮抗菌株;通过形态观察、生理生化实验、Biolog及16S rDNA序列分析鉴定目标菌株;利用平板双向培养法和滤纸片扩散法测定抑菌谱及拮抗性质。【结果】分离到一株高活力的水稻纹枯病菌拮抗菌株YB-3,该菌株属于解淀粉芽胞杆菌(Bacillus amyloliquefaciens);菌株YB-3对常见的14株病原真菌和7株细菌具有较强的拮抗作用,并发现其对亲缘关系较近的芽孢菌属有较强的拮抗作用;该菌株的抑制活性具有温度稳定、耐酸、但对蛋白酶敏感的特点。【结论】通过指示菌法筛选到一株对水稻纹枯病菌有强拮抗作用的解淀粉芽胞杆菌(B.amyloliquefaciens)YB-3,它具有广谱、高效的植物病原菌拮抗活性。     

苦豆子根瘤内生细菌分离及其对棉花枯萎病的生物防治效果测定

对从塔里木盆地苦豆子中分离得到的内生细菌进行皿内涂布拮抗实验、对峙培养法拮抗实验、胞外分泌物拮抗性测定和盆栽控病实验等研究,结果表明塔里木盆地苦豆子根瘤中存在大量的棉花枯萎病菌拮抗性内生细菌。皿内涂布法筛选结果表明60株苦豆子根瘤内生细菌中有48株相对抑菌率超过50%。对峙培养法对48株拮抗性内生细菌进一步筛选的结果表明,对棉花枯萎病菌抑菌距离超过20 mm的菌株有40株。40株拮抗性内生细菌胞外分泌物对棉花枯萎病菌的抑菌距离超过5 mm的有26株。KDRE12和KDRE41对棉花枯萎病的盆栽控病平均防效分别为67.11%和72.65%,具有应用潜力。     

中药渣堆肥中解磷细菌的筛选、鉴定及其拮抗作用

为开发新型、安全及高效的微生物菌肥,以堆肥2个月后的中药渣为试材,筛选解磷拮抗细菌.用有机磷细菌培养基初步筛选5株解磷细菌;采用溶磷圈法和磷钼蓝比色法测定初筛菌株的解磷作用,解磷作用最优的菌株是YP5;通过形态学特征观察、生理生化试验和16SrDNA序列分析鉴定,确定菌株YP5为铜绿假单胞菌(Pseudomonas a...     

Potential of rhizobacteria for prohibition of potato golden cyst nematode activity

Potato golden cyst nematode is an important causal agent of potato disease. Nematode cysts were isolated and identified by phenotypic and genotypic characteristics. Biocontrol activity of 120 isolated bacteria from rhizosphere towards Globodera rostochiensis, were evaluated. Nematode cysts were exposed to root extracts and released juveniles (J2) subjected to cultural filtrate of isolated bacteria and their mortality were recorded after 24 and 48 h. In total, 33 strains showed inhibitory activities which they caused mortality ranged from 61.10% to 78.74%. Five representatives of inhibitory strains were applied on potato cultivar Marfona under greenhouse conditions using tuber coating and soil drenching in a complete randomised block design with three replicates. The active bacterial strains were identified as Bacillus subtilis, Bacillus megaterium, Pseudomonas fluorescens bv. I, P. fluorescens bv. II and P. putida bv. B. In the greenhouse, significant differences were observed among the tested strains, where the highest disease severity reduction was 42.31%.     

微生物复合菌剂的制备

【背景】微生物复合菌剂比单一菌剂更能够在土壤中高效、稳定发挥作用促进作物生长,是微生物菌剂发展的趋势,但是目前对构建微生物复合菌剂的研究不够深入。【目的】研制可显著促进水稻生长的微生物复合菌剂,并构建微生物复合菌剂的数学模型。【方法】将解淀粉芽孢杆菌(Bacillus amyloliquefaciens) FH-1与7株植物促生细菌按照生物量1:1的比例复配成微生物复合菌剂,利用水稻盆栽实验筛选高效复合微生物菌剂。利用生化方法对各个菌株的促生特性进行测定。分析植物特征和菌株促生特性间的相关性关系,并据此利用一般线性方程构建复合菌剂的数学模型。【结果】与空白对照CK相比,复合菌剂FN显著提升水稻的苗长20.79%、根长26.67%和鲜重74.84%(P0.05),是7种微生物复合菌剂中综合效果最佳的微生物复合菌剂。偏相关分析结果表明解无机磷能力、产铁载体能力和产ACC脱氨酶能力在促进植物生长过程中可能发挥着更重要的作用。根据菌剂促生特性与植物特征的偏相关性结果,构建了微生物复合菌剂数学模型,预测准确率可达97%以上。【结论】成功研制了高效促进水稻生长的微生物复合菌剂,并构建了微生物复合菌剂的数学模型,可为微生物复合菌剂的研制提供一定的科学指导。     

Random-amplified polymorphic DNA (RAPD) analysis shows intraspecies differences among Xanthomonas albilineans strains

Five strains of Xanthomonas albilineans , causal agent of leaf scald disease in sugarcane from various geographical regions, were compared using random amplification of polymorphic DNA (RAPD) to determine whether they could be differentiated at the DNA level. CsC1-purified genomic DNA from these strains were amplified by the polymerase chain reaction (PCR) using arbitrary 10-mer primers according to standard RAPD conditions and the amplification product profiles analysed by conventional agarose gel electrophoresis. Although most RAPD markers were common to all five strains, unique profiles for each strain were discernible using four 10-mer arbitrary primers individually. Reproducible DNA fingerprints indicate that RAPD analysis can be used to identify and differentiate the X. albilineans strains. This technique has the potential for use in monitoring the appearance of foreign strains of X. albilineans in various geographical regions and could be used for the construction of phylogenetic trees.     

The modeled structure of the IgG Gar VL region and its implications for anti-flavin and anti-DNP fine specificities

The interaction of the purified C1q component of human C with synchronized, quiescent human gingival fibroblasts was investigated, and the presence of a specific binding site was demonstrated. Quantitative binding studies with radioiodinated C1q showed that binding was specific, saturable, and reversible upon addition of unlabeled C1q or by increasing the salt concentration. Scatchard plot analysis of the data yielded an affinity constant of 2 X 10(7) M-1 for all cell strains examined. The capacity for C1q binding varied among the eight cell strains examined. The number of binding sites per cell ranged from 2.6 to 17.7 X 10(6) with an average of 8.4 X 10(6). The receptor was insensitive to trypsin digestion, and it bound the collagen-like portion of the C1q molecule. Specific immunofluorescence staining showed that virtually all the viable cultured fibroblasts were able to bind added C1q. Flow cytometric analysis demonstrated a spectrum of fluorescence intensity among the cell strains, and there was a positive correlation between fluorescence intensity and the number of binding sites detected by using radiolabeled C1q.     

Effect of ultraviolet light irradiation and nitrosoguanidine on viability of 46 strains of Arthrinium and their antibiotic production.

The ability of strains of the Arthrinium genus to inhibit microbial development has been previously described. In the present work different periods of mutagenic treatment using ultraviolet light, and of nitrosoguanidine treatment, on strains of Arthrinium were investigated. With nitrosoguanidine treatment the survival rate ranged from 2.17 to 8.78%. Mutant strains were only obtained with a higher antibiotic production in comparison with the wild-strain, when the mutagenic agent was UV light.     

西葫芦根腐病菌拮抗细菌的防病促生作用

【目的】获取促生的同时可抑制尖孢镰刀菌(Fusarium oxysporum)的拮抗菌并明确其防治效果。【方法】平板对峙生长法测定前期分离的19株促生菌对尖孢镰刀菌的抑制作用,温室条件下接种微生物菌剂对西葫芦根腐病的防治作用;田间条件下接种复合菌剂代替部分化肥对西葫芦生长的影响。【结果】可有效拮抗尖孢镰刀菌的菌株有9株,其中,菌株FX2的抑菌活性较好,抑制率达到66.80%。在温室盆栽中,接种微生物菌剂(LHS11+FX2)对西葫芦根腐病抑制率达到57.14%。在田间试验中,微生物菌剂配施化肥对西葫芦的生物量和根系形态影响显著,以85%化肥+复合菌剂处理效果较优,其对西葫芦成熟期的产量显著提高27.13%。【结论】复合菌剂(LHS11+FX2)对西葫芦根腐病具有较好的防治作用;85%化肥+复合菌剂对西葫芦的促生作用明显,在一定程度上节约了化肥投入成本,提高了增产效益。     

Putative polymerase chain reaction markers for insecticide resistance in the leafminer Liriomyza trifolii (Diptera: Agromyzidae) to cyromazine and abamectin

In this study, random amplified polymorphic DNA polymerase chain reaction (RAPD-PCR) was used to identify polymorphic genomic DNA that would discriminate among cyromazine-resistant, abamectin-resistant, and susceptible Liriomyza trifolii (Burgess) (Diptera: Agromyzidae) leafminers. Using a reference strain that was susceptible to both cyromazine and abamectin, and a cyromazine-resistant strain and an abamectin-resistant strain, 400 oligonucleotides were assayed using RAPD-PCR. We found that two oligonucleotides, B10 and G16, amplified unique bands in the cyromazine-resistant strain but not in the reference or abamectin-resistant strains. Three oligonucleotides, K04, J13, and I02, showed polymorphisms unique to the abamectin-resistant strain but not in the reference or cyromazine-resistant strain. Leaf dip bioassays and RAPD-PCR were performed on two additional reference strains, seven strains from commercial ornamental production greenhouses, and one field strain. The two reference strains were negative for the resistance-correlated oligonucleotides. Of the seven strains from ornamental greenhouses, leaf dip bioassays showed that five had some level of resistance to both abamectin and cyromazine, whereas two were susceptible. The field strain was susceptible to both cyromazine and abamectin. In RAPD-DNA analyses, the five strains with abamectin resistance were positive for the three abamectin resistance-correlated oligonucleotides K04, J13, and I02. In the cases of cyromazine resistance, the five strains with cyromazine resistance were positive for the two cyromazine resistance-correlated oligonucleotides B10 and G16. The field strain and two greenhouse strains that were susceptible in leaf dip bioassays were negative for all three abamectin resistance-correlated oligonucleotides. The field strain and one greenhouse strain were negative for the two cyromazine resistance-correlated oligonucleotides; however, one greenhouse strain that was susceptible to cyromazine in leaf dip bioassay tested positive for one of the cyromazine resistance-correlated oligonucleotides. This method can be used to quickly identify cyromazine resistance, abamectin resistance, or both in leafminers, enabling a grower to choose an effective insecticide for leafminer control in a timely manner.     

毛竹根际可培养微生物种群多样性分析

[目的]为了了解天然毛竹林根际可培养微生物种群的多样性信息,[方法]采用稀释平板法,对浙江天目山和重庆缙云山天然毛竹林根际细菌和放线菌进行了分离,并对其16S rDNA序列进行了分析.[结果]分别从天目山和缙云山天然毛竹林根际分离得到51株和31株菌落形态差异的细菌和放线菌.16S rDNA序列分析表明,天目山和缙云山毛竹根际细菌主要包括厚壁菌门(Firmicutes,分别为40%和58%)、放线菌门(Actinobacteria,分别为36.7%和10.52%)、变形菌门-亚群(Alphaproteobacteria,分别为10%和5.26%)和变形菌门 --亚群(Gammaproteobacteria,分别为10%和26.32%),其中芽孢杆菌属(Bacillus sp.)为共同的优势菌属(分别为34.38%和42.11%).分离的菌株中,B188、B171和B152等6株与GenBank中已报道16S rRNA基因序列的相似性从90%到96%不等,可能代表着新属或种.[结论]这表明,天然毛竹林根际具有较为丰富的可培养微生物种群多样性,并存在一些潜在的新的微生物菌种资源.     

Selection and evaluation of the potential of choline-metabolizing microbial strains to reduce Fusarium head blight

Choline and betaine are found in wheat flower tissues and have been implicated in stimulating hyphal growth of the primary causal agent of Fusarium head blight (FHB), Gibberella zeae. Choline metabolizing strains (CMS) from wheat anthers may therefore be a useful source of antagonists of G. zeae. One-hundred twenty-three of 738 microbial strains that were recovered from wheat anthers collected from plants grown in Illinois and Ohio were CMS as determined by growth in a liquid medium containing choline as a sole carbon and nitrogen source and a colorimetric, choline oxidase-based assay of culture filtrate. Thirty-one out of 123 CMS reduced FHB disease severity by at least 25% in greenhouse tests on wheat and 17 reduced FHB severity by at least 50%. All five CMS selected for field testing in 2003 reduced disease severity compared to the untreated check at both field locations on moderately resistant cultivar Freedom. Freedom wheat treated with Pseudomonas sp. AS 64.4 had 63% and 46% less FHB severity than untreated wheat at the two sites. Three of five CMS reduced severity at both locations on susceptible cultivar Pioneer Brand 2545. Disease control was comparable to that obtained using the fungicide Folicur 3.6F. Selection of wheat anther colonists for ability to utilize choline as a sole carbon and nitrogen source has utility as a screening tool in the search for efficacious antagonists of G. zeae although choline utilization does not insure that an isolate will be an effective biocontrol agent against Fusarium head blight.     

Deoxyribonucleic acid reassociation in the classification of flavobacteria.

DNA-DNA reassociation studies showed that Flavobacterium emningosepticum strains had a genetic relatedness of 91 to 100% with inter-strain duplexes having high thermal stabilities. The only exception, strain NCTC10016, had an average relatedness of only 43% to other strains of F. meningosepticum. The apparent divergence in DNA base sequence of this strain was reflected in the structural differences of some enzymes. There was a gradation of DNA relatedness among the Flavobacterium group II-b strains, but three strains were sufficiently related to constitute a species. Low levels of genetic relatedness were confirmed between F. meningosepticum and strains of Flavobacterium group II-b, group II-f, F. aquatile, F. breve, F. heparinum, F. pectinovorum, F. odoratum and Moraxella saccharolytica. All strains had base compositions in the range 32 to 46% guanine plus cytosine. The genome sizes of representative strains of F. meningosepticum and Flavobacterium group II-b were 2-50 X 10(9) to 3-52 X 10(9) daltons. The taxonomic implications of these findings are discussed.