用接合转移方法构建杀虫防病荧光假单胞菌

以经过改造的含有转座子Ｔｎ５的自杀质粒ｐＳＵＰ２０２１为载体，通过接合转移将苏云金杆菌杀虫蛋白基因ｃｒｙ Ｉ Ａ（ｃ）片段插入生防细菌荧光假单胞菌Ｐ３０３菌株染色体组。Ｓｏｕｔｈｅｒｎ和Ｗｅｓｔｅｒｎ印迹分析分别证实杀虫基因的导入和杀虫蛋白的表达。室内生物测定结果表明新构建的ＰＴ２１０、ＰＴ２１２等荧光假单胞菌工程菌菌株不仅保持了野生型自然菌株对小麦全蚀病良好的抑菌活性，而且表现出对小菜蛾和玉米暝     

用荧光假单孢菌防治番茄青枯病

土壤中存在着大量对植物病原菌有拮抗作用的细菌。据报道，番茄青枯病菌的拮抗菌占土壤细菌的６．５％－２８．６％。通过分析研究，番茄青枯病菌的拮抗菌多数属于荧光假单孢菌属。经多次分离筛选，从土壤中分离出了对作物无不良影响而对番茄青枯病菌有拮抗作用的菌株１０４个。经试验，利用这种拮抗菌防治番茄青枯病，当发病株率在２５％以下时，防治效果可达１００％；但当发病株率达４５％时，防治效果仅为５０％左右。这表明，利     

发光酶基因标记荧光假单胞菌X16L2的发光研究

通过光光光度仪和ＡＭＰＮ法定量测定荧光假单胞菌Ｘ１６Ｌ２,在液培和土壤中的发光情况及其数量,发现Ｘ１６Ｌ２在液培和土才壤中的发光动力学曲线相似,达到稳定发光强度的时间均在加入反应底物癸醛后约７分钟。Ｘ１６Ｌ２在液培中的发光强度与生物量吴正相关。土壤中Ｘ１６Ｌ２的发光量既与活菌数有关,也与土壤条件有关,土著微生物对Ｘ１６Ｌ２的光量,土壤中Ｘ１６Ｌ２的发光量既与活菌数有关,也与土壤条件有关。土著微生物     

产荧光假单胞菌分离鉴定及其质粒DNA复制区的克隆

从国内多个地区的65份土样中分离纯化了产荧光假单胞菌株146株,其中菌株DSC3和DKXC1含有质粒,细胞形态,菌落特征的观察和检测证明DKXC1菌株为恶臭假单胞菌（Pseudomonas putida),以其为出发菌株,克隆了该菌质粒DNA复制子1.7kb片段,序列分析结果表明该DNA片段由3个ORF组成,分别编码311、130、115个氨基酸;它与已知的Pf-E.coli穿梭质粒pUCP19的复制区同源性只有49%,该复制子DNA序列已在GenBank登记,登记号为AF273219,这是国内分离克隆的第一个Pseudomonas质粒DNA复制子,以此复制子构建了4.7kb的E.coli-Pf穿梭载体pYQSPE,该质粒在荧光假单胞菌P303中能够稳定复制和遗传。     

假单胞菌的生物防治作用研究

假单胞菌能产生多种抗生素 ,改善植物营养 ,促进植物生长 ,且能降解土壤中有毒物质 ,对植物促生长防病作用显著。部分假单胞菌株具有杀虫功能 ,由假单胞菌可开发一系列生物防治产品     

一株具有多途径氮代谢功能的荧光假单胞菌

为研究全自养脱氮过程，采用斯凯尔曼亚硝化单胞杆菌培养基，从南京某城市污水处理厂序批式处理系统（SBR）中分离得到一株具有多途径氮代谢功能的荧光假单胞菌Pseudomonas fluorescens dN13。该菌株在有氧条件下，无有机碳源存在，能进行氧化NH3-N产能代谢生长；有有机碳源存在且NO2^- -N和NO3^- -N的浓度较高时，它能将NO2^- -N和NO3^- -N还原为NH3-N。在厌氧条件下，只有乙酸钠作碳源时，该菌株仍能进行硝酸亚硝酸还原（反硝化）作用。在这个具有多途径氮代谢功能的菌株细胞内，硝化反硝化过程耦合成为可能。     

促生荧光假单胞菌对桃树根区土壤环境和植株生长的影响

【目的】探究荧光假单胞菌(Pseudomonas fluorescens)的促生特性及其对桃树根区土壤环境和桃树生长的影响,明确荧光假单胞菌在桃树根系–土壤微生态系统中的作用机制,为荧光假单胞菌在桃园的应用提供理论依据。【方法】采用不同固体培养及液体发酵试验鉴定荧光假单胞菌的促生特性。以1年生盆栽毛桃[Prunus persica (L.) Batsch]实生苗为试材进行预备试验,确定了荧光假单胞菌悬液的适宜施用浓度为4×10⁸ CFU/mL。以2年生‘瑞光39/毛桃’[P. persica (L.) Batsch]嫁接苗为试材进行盆栽试验,试验设土施荧光假单胞菌悬液(4×10⁸ CFU/mL)(PF)和清水对照(CK) 2个处理,在处理后1、2、4、6周,采集桃树根区土壤样品,测定荧光假单胞菌数量;在处理后40天采集根区土壤样品测定土壤微生物结构、酶活性、养分状况和pH。在处理后20、40天取植株样进行光合指标、生物量、根系生长发育的测定。【结果】1)荧光假单胞菌具有以下促生特性：吲哚-3-乙酸(IAA)产量为6.17 mg/L,溶无机磷和...     

两种浮萍根系分泌物对荧光假单胞菌脱氮影响的差异及其剂量效应研究

本研究对太湖地区高效除氮紫背浮萍Spirodela polyrrhiza(HZ1)和低效除氮青萍Lemna minor(WX3)的根系分泌物进行了连续收集和分离,并考察了根系分泌物剂量与成分对反硝化细菌荧光假单胞菌Pseudomonas fluorescens脱氮效率的影响。结果发现,两种浮萍根系分泌物总组分均能显著促进P.fluorescens的脱氮效率,但对P.fluorescens的生长无影响。HZ1根系分泌物总组分(HO)促进作用显著高于WX3根系分泌物总组分(WO),这与HZ1对水体的除氮效率高于WX3的结果一致。两种浮萍根系分泌物各组分也有不同的效果,其中仅WX3根系分泌物酸性组分(WA)显著抑制了P.fluorescens的脱氮效率,HZ1根系分泌物不同组分是促进或无作用。总体上,P.fluorescens的脱氮效率随浮萍根系分泌物及其各组分添加剂量的增加呈先上升后下降的趋势,而WA组分先下降后基本不变,各组分在剂量为1.0%(v/v)时起最大效果。总之,根系分泌物与浮萍–微生物耦合系统的除氮效率密切相关,酸性组分是两种浮萍根系分泌物促进P.fluorescens脱氮效果差异的主要原因。     

铜绿假单胞菌生物降解特性的研究进展

近年来在环境污染物的生物降解研究方面有了很大进展。铜绿假单胞菌（Pseudomon aslugtrlosa,PA）作为重要的降解菌株之一,具有较强的降解能力,可降解物质种类广泛,在环境污染物的生物降解中具有重要作用并占据重要地位。本文综述了PA的降解特性、代谢途径、遗传基础与酶系及促降解物质在生物降解方面的研究进展。     

假单胞菌HC5对马铃薯晚疫病菌的抑制作用研究

从土壤中分离获得对马铃薯晚疫病菌具有较强拮抗作用的假单胞菌HC5。采用皿内涂布法、平板对峙法和平板渗透法分别评价了HC5菌株发酵液对马铃薯晚疫病菌的抑制能力,采用平板渗透法进一步研究了菌株HC5发酵液在不同温度、pH以及紫外线照射时间下对马铃薯晚疫病菌抑制的稳定性。结果显示,不同方法下,菌株HC5对马铃薯晚疫病菌均具有较强的抑制作用,与对照组相比,病原菌几乎不生长。菌株HC5发酵液在温度50 ℃以下,pH为5～11时,紫外线照射120 min均对马铃薯晚疫病菌具有抑制作用,即菌株HC5的耐受稳定性较强。     

丛枝菌根育苗缓解西瓜枯萎病的机制

西瓜易受尖孢镰刀菌侵害发生枯萎病.植物形成丛枝菌根(AM)可有效促进养分吸收和病害防控.在田间直接接种丛枝菌根菌时,侵染效率和生态效应多数会受到限制.采用育苗期接种丛枝菌根菌(Rhizophagus intraradices,R.i)培养菌根苗,成苗后移栽入盆钵,研究西瓜丛枝菌根苗在抗西瓜枯萎病中的作用及其机制.将西瓜...     

Rhizobacterium Bacillus amyloliquefaciens Strain SQRT3-Mediated Induced Systemic Resistance Controls Bacterial Wilt of Tomato

Tomato bacterial wilt caused by Ralstonia solanacearum seriously threats tomato growth in tropical and temperate regions around the world. This study reported an antagonistic bacterial strain, Bacillus amyloliquefaciens strain SQRT3, isolated from the rhizosphere soil of tomato plants, which strongly inhibited in vitro growth of pathogenic R. solanacearum. The suppression of tomato bacterial wilt by strain SQRT3 was demonstrated under greenhouse conditions. Additionally, induced systemic resistance (ISR) in tomato as one of the potential disease suppression mechanisms was investigated in the plants inoculated with the isolated bacterial strain SQRT3. The results showed that strain SQRT3 applied with R. solanacearum by drenching significantly reduced tomato bacterial wilt by 68.1% biocontrol efficiency (BE) and suppressed the R. solanacearum populations in the rhizosphere soil compared to the control only drenched with R. solanacearum. The BE of the isolated bacterial strain SQRT3 against tomato wilt increased to 84.1% by root-dipping. Tomato plants treated with both strain SQRT3 and R. solanacearum showed increases in activities of peroxidase and polyphenol oxidase compared with other treatments. The application of strain SQRT3 reduced membrane lipid peroxidation in tomato leaves. The expressions of marker genes for jasmonic acid-and salicylic acid-dependent signaling pathways were faster and stronger in tomato plants treated with both strain SQRT3 and R. solanacearum than in plants treated with either R. solanacearum or strain SQRT3 alone. Collectively, the findings indicated that strain SQRT3 can effectively control tomato wilt.     

胶冻样芽孢杆菌与生物质炭复配及对番茄产量和品质的影响

以提高胶冻样芽孢杆菌在土壤中的存活率、减缓其衰亡速率,延长存活时间,提高其对土壤难溶性钾的解钾效率为目的,利用不同种类的秸秆生物质炭与胶冻样芽孢杆菌进行复配筛选。通过室内培养试验发现,中性小麦秸秆炭可作为胶冻样芽孢杆菌较理想的载体。当土壤培养35天后,土壤速效钾含量在施用炭基解钾菌肥条件下较空白提高了28.53%,较小麦秸秆炭和纯菌液处理分别提高了20.75%、13.41%。纯菌液处理土壤中胶冻样芽孢杆菌的菌落数从1.1×10~9个/g土降为9.0×10~5个/g土,而炭基解钾菌肥处理则为1.4×10~7个/g土。田间施用炭基解钾菌肥能够提高番茄植株和果实中的全钾含量;提高番茄产量和品质,显著提高番茄Vc含量,降低硝酸盐含量。因此,中性小麦秸秆炭与胶冻样芽孢杆菌的复配能够提高菌剂的存活率,有效促进土壤中矿物态钾的释放。     

铜绿假单胞菌对镉胁迫水稻苗期生长与镉积累的影响

稻米重金属污染是人们广泛关注的严重问题,微生物钝化是阻遏环境重金属进入生物循环的有效途径之一。为了解铜绿假单胞菌对苗期水稻镉污染的缓解效应,本文以无镉处理为对照,通过添加20μmol×L~(-1)镉的水培试验,研究了铜绿假单胞菌、载体A(硅藻土,粒径1~3 mm)、载体B(硅藻土,粒径3~6 mm)、载体C(活性炭,比表面积1 000 m~2×g-1)以及铜绿假单胞菌与载体制备的菌剂A、菌剂B和菌剂C对水稻生长、镉含量及镉积累量的影响。结果表明,20μmol×L~(-1)镉处理显著抑制了水稻根长、株高的生长和干物质积累,添加菌液及菌剂A、B、C后,水稻生长状况得到显著改善,总生物量比镉处理(0.523 g×株~(-1))提高38.5%~67.3%,以菌剂B处理的水稻生物量最高。水稻根、茎鞘、叶以及地上部镉含量显著降低,其中添加菌剂A、菌剂B、菌剂C及活性炭处理的水稻地上部镉含量分别比镉处理(101.3 mg×g~(-1))下降45.9%、47.9%、59.9%和59.9%,迁移系数降低16.7%、25.0%、33.3%和33.3%,富集系数减少48.1%、48.8%、58.8%和60.9%。添加活性炭、菌剂A和菌剂C处理的水稻单株镉积累量降低18.2%、9.5%和24.3%,添加菌剂B以及依次含有56.4 mL、45.3 mL、28.4 mL菌悬液的菌液A、菌液B和菌液C处理,其镉积累量依次增加15.0%、30.4%、14.9%、16.9%,说明菌株通过提高作物生物量增加了镉积累。综上,铜绿假单胞菌可显著促进镉胁迫水稻的生长,降低水稻的镉含量,抑制镉的迁移,降低水稻镉的有效性;菌剂A、B、C表现出良好的镉钝化能力,表明铜绿假单胞菌可为环境镉污染的生物修复提供新菌株。     

Meta分析评估中国木霉对枯萎病防控效果及其影响因素

由致病性尖孢镰刀菌(Fusarium oxysporum)侵染引起的植物枯萎病是一种真菌性土传病害,对多种经济作物产生严重危害。木霉生物防治作为一种安全、环保、经济效益高且长效的防治措施已成为世界范围内的发展趋势,但其确切防效及其影响因素尚不明确。基于中国范围内研究所发表的中英文文章,运用Meta分析(元分析)方法研究了木霉对土传枯萎病的防控效果,同时分析了影响木霉防控效果的多种因素。结果表明,施用木霉在整体上对枯萎病具有显著的防控效果,其中,作物科类、木霉形式、施用方法、是否与其他方式联合及气候类型(基于大田试验)5类因素对木霉防效有显著影响;木霉种类、试验类型、木霉菌剂施用剂量和木霉菌剂施用次数对结果无显著影响。本研究结果预计能为田间高效应用木霉防控土传枯萎病提供参考。     

Root colonization and growth promotion of wheat and maize by Pseudomonas aurantiaca SR1

Wheat (Triticum aestivum L.), rice (Oryza sativa) and maize (Zea mays L.) are the most important cereals worldwide. However, in the last few years, soil has been submitted to both use and handling pressures due to the increase in agricultural practices, which are leading to its degradation. The use of plant growth-promoting rhizobacteria (PGPR) as inoculants constitutes a biological alternative for sustainable production. Pseudomonas aurantiaca SR1 was formulated as an inoculant in order to evaluate its growth promotion effect in the field when applied on maize and wheat seeds at the sowing time. P and N fertilization treatments were also included in the assays. P. aurantiaca SR1 colonized the root system of both crops and it persisted at appropriate population densities. It also showed a significant plant growth-promoting effect that was reflected in the yield. Another relevant finding was that both crops, when inoculated with P. aurantiaca SR1, presented higher yields with fertilization doses lower than those conventionally applied. This indicated its potential use as a reasonable alternative for crop production, with a minimization of the ecological impact.     

香蕉根际土壤原生动物群落特征及与枯萎病病原菌的关系

香蕉枯萎病由病原真菌尖孢镰刀菌4号生理小种引起,其根际微生物组是抵御病原菌入侵的关键屏障。根际原生动物群落在根际微生物群落调控以及植物健康方面有不可忽视的作用。利用Illumina MiSeq高通量测序技术分析连作蕉园中植株发病前后根际土壤原生动物群落特征变化以及其与病原菌的相互关系。结果表明：相较于可培养细菌与真菌,原生动物的数量对病原菌的种群数量变化的影响更大;根际原生动物的群落多样性与丰富度都随植株生长而降低,且在发病植株中更低;抽蕾前后的健康与发病植株中根际原生动物群落结构存在显著差异;总体而言,吞噬型原生动物的相对丰度显著高于其他功能类型的原生动物,且在健康与发病植株中差异显著;吞噬型原生动物中与病原菌具有显著相关性的多为丝足虫门;在抽蕾期的健康植株中,丝足虫门的高丰度属中Group-Te与Cercomonas属与病原菌呈显著负相关;在抽蕾期的患病植株中,相对丰度较高的植物致病型原生动物Pythium属与病原菌呈显著正相关。综上所述,根际土壤中原生动物对病原菌的影响较大,在植株生长过程中,原生动物的群落特征发生改变,且在发病与健康植株中存在差异。与病原菌具有显著负相关的吞噬型原生动物,尤其是丝足虫门的部分高丰度属,可能会在防控香蕉枯萎病上产生一定潜力。     

乌兰布和沙区不同品种沙棘的抗旱性综合评价

[目的]了解乌兰布和沙区17个沙棘品种的抗旱能力,为选择耐旱沙棘树种提供理论依据。[方法]在不同的水分条件下,采用隶属函数值法对其抗旱性进行综合评价。[结果](1)随干旱胁迫的增加,各品种的自然水分饱和亏、束缚水及束缚水/自由水比值、细胞膜透性呈增加趋势。自然水分饱和亏在22.88%~64.85%之间,在胁迫后期,丘依斯克、欧11号、白丘的水分饱和亏接近,处于高水平状态;深秋红、欧11号在束缚水及束缚水/自由水比值处于较高水平;橙色与白丘的相对电导率显著高于其他品种。(2)随干旱胁迫的增加,各品种的叶绿素含量、保水力、光合速率、蒸腾速率及气孔导度呈降低趋势。亚历山大12号的叶绿素含量最高;橙色和向阳的失水速率显著高于其他品种;辽阜1号和深秋红的光合蒸腾速率下降幅度较大、气孔导度下降程度维持在中等水平。[结论]各沙棘品种的抗旱性由强到弱顺序依次为:辽阜1号欧11号深秋红状元黄棕丘植物园白丘西伯利亚红晕丘依斯克亚历山大12号橙色高加索向阳巨人无核卡图尼海滨沙棘。     

Impact of agronomic practices on populations of Fusarium and other fungi in cereal and noncereal crop residues on the Canadian Prairies

Fusarium head blight (FHB) is an important disease which has been causing damage to wheat and barley crops in western Canada. Because crop residues are an important source of inoculum, it is important to know the ability of Fusarium spp. to colonize and survive in different residue types, and how their populations might be affected by agronomic practices. Sampling of residue types on producers’ fields for quantification of Fusarium and other fungi was conducted in 2000–2001 in eastern Saskatchewan. Fusarium spp. were isolated from most fields, whereas their mean percentage isolation (MPI) was over 50% for cereal and pulse residues, and under 30% for oilseed residues. The most common Fusarium, F. avenaceum, had a higher MPI in pulse and flax (45–48%) than in cereal or canola (10–22%) residues. This was followed by F. equiseti, F. acuminatum, F. graminearum, F. culmorum and F. poae which were isolated from all, or most, residue types. Factors affecting Fusarium abundance in residues included the current crop, cropping history, and tillage system. In cereal residues, the MPI of F. avenaceum was higher when the current crop was another cereal (24%) versus a noncereal (4–8%). When the current crop was another cereal, the lowest MPI of F. avenaceum and F. culmorum occurred when the field had been in summerfallow (SF) two years previous (F. avenaceum: 17% for SF, 28% for a crop; F. culmorum: 1% for SF, 4% for a crop); in contrast, F. equiseti and Cochliobolus sativus were most common in residues of cereal crops preceded by SF (F. equiseti: 16% for SF, 10% for a crop; C. sativus: 22% for SF, 13% for a crop). The MPI of F. graminearum was higher when the crop two years previous was an oilseed (7%) versus a cereal (4%). In regards to tillage effects, when the current crop was a cereal, the MPI of F. avenaceum was higher under minimum (MT) and zero tillage (ZT) (22–37%) than conventional tillage (CT) (15%), that of F. graminearum was lowest under ZT (3% for ZT, 7–11% for CT-MT), whereas that of C. sativus was highest under CT (27% for CT, 6–11% for MT-ZT). Under ZT, previous glyphosate applications were correlated positively with F. avenaceum and negatively with F. equiseti and C. sativus. These observations generally agreed with results from previous FHB and root rot studies of wheat and barley in the same region. Percentage isolation of F. avenaceum from noncereal and of F. graminearum from cereal residues were positively correlated with FHB severity and percentage Fusarium-damaged kernels of barley and wheat caused by the same fungi.     

Suppression of root-knot disease by Pseudomonas fluorescens CHA0 in tomato: importance of bacterial secondary metabolite, 2,4-diacetylpholoroglucinol

The antimicrobial metabolites 2,4-diacetylphloroglucinol (2,4-DAPG) and pyoluteorin contribute to the ability of Pseudomonas fluorescens strain CHA0 to control plant diseases caused by soil-borne pathogens. P. fluorescens strain CHA0 and its derivatives CHA89 (antibiotics-deficient) and CHA0/pME3424 (antibiotics overproducing) were investigated as potential biocontrol agents against Meloidogyne javanica the root-knot nematode. Exposure of root-knot nematode to culture filtrates of P. fluorescens under in vitro conditions significantly reduced egg hatch and caused substantial mortality of M. javanica juveniles. Nutrient broth yeast extract (NBY) medium amended with 2% (w/v) glucose or 1 mM EDTA markedly repressed hatch inhibition activity of the strain CHA0 but not that of CHA0/pME3424 or CHA89. On the other hand, NBY medium amended with glucose significantly enhanced nematicidal activity of the strain CHA0/pME3424. Neither glucose nor EDTA had an influence on the nematicidal activity of the strains CHA0 and CHA89. Under in vitro conditions, antibiotic overproducing strain CHA0/pME3424 and CHA0 expressed phl‘-’lacZ reporter gene but strain CHA89 did not. Expression of the reporter gene reflects actual production of DAPG. In general, CHA0/pME3424 expressed reporter gene to a greater extent compared to its wild type counterpart CHA0. Regardless of the bacterial strains, reporter gene expression was markedly enhanced when NBY medium was amended with glucose but EDTA had no such effect. A positive correlation between the degree of juvenile mortality and extent of phl‘-’lacZ reporter gene expression was also observed in vitro. Strain CHA0 produced zones of 4-6 mm on MM medium containing gelatin while strain CHA0/pME3424 and CHA89 did not. When MM medium containing gelatin was amended with 2% glucose of 1 mM EDTA size of haloes produced by the strain CHA0 reduced to 2 mm. Under glasshouse conditions aqueous cell suspension of the strains CHA0 or CHA0/pME3424 at various inoculum levels (10⁷, 10⁸ or 10⁹ cfu ml⁻¹) significantly reduced root-knot development. CHA89 caused significant reduction in galling when applied at 10⁹ cfu ml⁻¹. To better understand the mechanism of nematode suppression, split root bioassay was performed. Split-root experiments, that guarantee a spatial separation of inducing agent and a challenging pathogen, showed that soil treatment of one half of the root system with cell suspension of CHA0 or CHA0/pME3424 resulted in a significant systemic induced resistance leading to reduction of M. javanica infection of tomato roots in the non-baterized nematode treated half. The results clearly suggest that the antibiotic 2,4-DAPG from P. fluorescens CHA0 act as the inducing agents of systemic resistance in tomato roots. Populations of CHA0 and its derivatives declined progressively by 10-fold between first and fourth harvests (0-21 days after inoculation). However, bacterial populations increased at final harvest (28 days after application).