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1.
Ca2+,Mn2+等8种金属离子对软腐欧文氏菌一些致病因子的影响 总被引：1，自引：0，他引：1
在离体试验中,不同金属离子对马铃薯软腐欧文氏菌(Erwinia carotovora subsp.carotovora)菌株StEcc-12生长,产生胞外酶和胞外酶活性具有不同的影响。在含果胶酸钠的培养液中,Ca2+促进生长,并使培养液中的果胶裂解酶(PL)、果胶水解酶(PG)和蛋白酶3种胞外酶分别提高1.4~23.4倍,0.3~2.9倍和0.7~6.5倍。Mn2+对StEcc-12生长及产生3种胞外酶的影响与Ca2+相似。Ni2+显著抑制细菌的生长,但在含Ni2+培养液中3种胞外酶的单位活性比对照高1.7倍以上。在酶粗提液中分别加入上述3种离子对PL酶的活性均有抑制作用。Zn2+对细菌生长没有显著影响,高浓度Al3+抑制细菌生长,这2种离子均能促进3种胞外酶的产生。随着Fe2+和K+浓度的增高,细菌生长减少,两种果胶酶产量降低,蛋白酶产量增加,粗提液中PL酶活性下降。Mg2+对细菌生长和胞外PG产生没有明显影响,提高Mg2+浓度有利于PL的产生。酶粗提液中加入Mg2+后PL活性增高。所有以上离子都能使聚半乳糖醛酸发生不同程度的凝聚,但只有Ca2+和Mg2+的有效凝聚浓度与薯块的实际浓度接近。 相似文献
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从广东、福建、湖南、江西及浙江5省7市采集的14科33种观赏植物上分离到199株软腐欧文氏杆菌菌株,44项理化性状鉴定结果:126株为胡萝卜软腐欧文氏杆菌胡萝卜变种(Erwinia carotovora pv. carotovora简称Ecc)、7株为菊欧文氏杆菌(Erwinia chrysanthemi简称Ech)、66株介于Ecc和Ech之间的中间型(I、Ⅱ、Ⅲ、Ⅳ、V)。Ecc和中间型菌株是我国观赏植物的主要病原细菌。在本文中首次报道了竹芋科(竹芋)、苋科(千日红)两科及一些植物种:芦荟、白穗花、玉簪、黄精、富贵竹、兜兰、砂仁及饿术草是软腐欧文氏杆菌的新的寄主植物。血清学研究表明,Eca、Ecc及Ech这三个种之间不发生交叉反应,但同一个种内是可以起反应的。另外,Ecc与中间型相互之间有沉淀反应,表明它们之间的亲缘关系较近。 相似文献
3.
采用3,5一二硝基水杨酸法分析了莲子草假隔链格孢NimbyaalternantheraeSF一193菌株在活体内外产生的羧甲基纤维素酶（Cx）、口．葡萄糖苷酶、木聚糖酶、聚甲基半乳糖醛酸酶（PMG）和多聚半乳糖醛酸酶（PG）的活性。结果表明,SF一193可产生cx、伊葡萄糖苷酶、木聚糖酶和PG,且4种细胞壁降解酶在活体外和活体内的活性显著不同。其发酵滤液中以PG活性最高,其次为木聚糖酶、,β-葡萄糖苷酶和Cx;在空心莲子草叶片内以Cx活性最高,其次为β一葡萄糖苷酶、木聚糖酶和PG。在活体内,4种酶活性随接种天数和温度的变化而不同。Cx和卢．葡萄糖苷酶的活性在接种第3d达到最大,木聚糖酶和PG活性则分别在第2d和第5d达到最大;Cx、卢一葡萄糖苷酶和PG活性在30℃时最高,而木聚糖酶活性在35℃时最高。 相似文献
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番茄芝麻斑病原菌产生的细胞壁降解酶种类及其活性变化 总被引：3，自引：0，他引：3
番茄芝麻斑点病原菌能够产生一系列细胞壁降解酶,即果胶甲基酯酶(PE)、多聚半乳糖醛酸酶(PG)、果胶甲基半乳糖醛酸酶(PMG)、多聚半乳糖醛酸反式消除酶(PGTE)、果胶甲基反式消除酶(PMTE)和纤维素酶(Cx)。酶动力学研究表明:产生的各种细胞壁降解酶均有特定的最适反应条件。水解酶类的PG、PMG、Cx最大酶活的pH均为5.0,温度均为50℃;裂解酶类的PGTE和PMTE最大酶活的pH均为9.0,温度均为30℃,与其他病菌产生的细胞壁降解酶的特性基本相同。在活体外,随着培养天数的增加PG、PMG、PGTE、PMGE、Cx的活性都大幅度增加,但所有酶活性都明显低于活体内的酶活性。 相似文献
6.
软腐欧氏杆菌的蛋白酶与致病作用的关系 总被引：1，自引：0，他引：1
胡罗卜软腐欧氏杆菌(Erwinia carotovora var.carotovora Dyc)在合成培养基MS中产生两种蛋白酶,但在无细胞滤液中测不到果胶裂解酶和果胶水解酶的活性。用等电聚焦电泳测得这两种蛋白酶都是碱性蛋白酶,等电点分别为8.3和8.9。变性温度为49-50℃,对酸或碱具有较高的稳定性。纯化的蛋白酶单独作用马铃薯块、能使其变软,切片观察表明细胞壁被明显降解。通过滤膜结合法,将供体菌1830/pJB4J1::Tn5的转座子转入到野生型StEcc-12中,从2000个接合子中获得了11个缺少产蛋白酶能力的突变体。这些突变体的分泌性蛋白质等电聚焦图谱与野生型菌株明显不同,其中一个变突体蛋白酶带完全缺失,另外一个则出现8条新的蛋白带,其他5个属部分缺失。这些突变体对马铃薯块组织离析、腐烂的能力都有不同程度的减弱。研究结果认为在由胡罗卜软腐欧氏杆菌引起的软腐病程中蛋白酶和果胶酶是协同作用的,果胶酶降解植物细胞间和胞壁中的果胶质,而蛋白酶则降解植物细胞壁和膜中的蛋白质。 相似文献
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通过平板稀释分离法，从多年生草本植物薄荷根际土壤中分离到一株对灰葡萄孢Botrytis cinerea抑菌活性良好的细菌TD-7。通过形态特征和16S rDNA、ITS序列分析，鉴定该菌为解淀粉芽胞杆菌Bacillus amyloliquefaciens，并对其发酵滤液抑菌机理、最优发酵条件、最优发酵培养液组分及防治效果进行了研究。研究发现，菌株TD-7发酵滤液对番茄灰霉菌孢子的萌发有强烈的抑制作用，对菌丝生长抑制率在95%以上。皿内酶分泌试验发现，该菌可分泌几丁质酶、纤维素酶和蛋白酶，未检测到β-1，3-葡聚糖酶活性。发酵滤液抑菌活性最强的条件为在250 mL三角瓶中，将种子液按3%的体积比接种到50~75 mL培养液中，初始pH 8.0，温度32~36℃，转速200 r/min，发酵5 d。通过正交优化，得到最佳发酵培养液配方为玉米粉3%、蛋白胨1.5%、KH2PO4 0.05%。经摇床发酵条件及发酵培养液组分优化，菌株TD-7发酵滤液抑菌活性提高了26.39%。该菌的发酵液对温室盆栽及田间大棚中的番茄灰霉病均具有良好的防治效果，分别为80.61%和87.70%。滤液稳定性试验结果表明，菌株TD-7发酵滤液中活性物质具有较广的pH适应范围（pH 4.0~9.0），在高温和紫外光照射下较稳定，有制成生物制剂应用于番茄灰霉病生物防治的潜力。 相似文献
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温湿度调控对番茄灰霉病菌产生的细胞壁降解酶的影响 总被引：6，自引：0，他引：6
番茄灰霉病菌在致病过程中能够产生4种细胞壁降解酶,以PMG酶活性最高,其次是β-葡萄糖苷酶和PG酶,Cx最少。灰霉病菌在不同温度下侵染番茄叶片时产生的致病酶活性不同,4种酶在20℃时表现了最高的活性,15℃次之,当温度达到25℃时,各种酶的活性都急剧下降,随着温度的再升高,酶活更低。随着湿度的增高,病菌产生的细胞壁降解酶的活性也增加,当相对湿度达到90%以上时,4种酶的活性也达到最高。温湿度对番茄灰霉病菌产生细胞壁降解酶的影响趋势,与其对发病的影响趋势是一致的。 相似文献
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玉米茎腐病菌产生的细胞壁降解酶种类及其活性分析 总被引：20，自引：0，他引：20
玉米茎腐病菌Pythium aphanidermatum、Fusarium graminearum能够产生一系列细胞壁降解酶,即果胶甲基酯酶(PE)、多聚半乳糖醛酸酶(PG)、果胶甲基半乳糖醛酸酶(PMG)、多聚半乳糖醛酸反式消除酶(PGTE)、果胶甲基反式消除酶(PMTE)和纤维素酶(Cx)。Fusarium graminearum产生的细胞壁降解酶活性明显高于Pythium aphanidermatum。病菌在活体内和活体外产生的细胞壁降解酶活性明显不同。酶动力学研究表明:Fusarium graminearum产生的各种细胞壁降解酶均有特定的最适反应条件。水解酶类的PG、PMG、Cx最大酶活的pH分别为6.0、5.0、6.0,温度分别为50、40、50℃;裂解酶类的PGTE和PMTE最大酶活的pH均为9.0,温度分别为30、20℃,与其它病菌产生的细胞壁降解酶的特性基本相同。 相似文献
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柑橘大实蝇Bactrocera minax(Enderlein)是我国西南和华中地区柑橘的主要害虫,以滞育蛹在土壤中越冬。为探讨温度对柑橘大实蝇滞育蛹生理状态的影响,本文测定了12、16、20℃和24℃下柑橘大实蝇滞育蛹海藻糖酶和山梨醇脱氢酶活性的变化。结果显示,低温(12℃)下,海藻糖酶活性逐渐上升,高温(24℃)下,海藻糖酶活性逐渐下降;不同温度下山梨醇脱氢酶活性变化不尽相同,在起始阶段(10 d时),柑橘大实蝇滞育蛹山梨醇脱氢酶活性普遍较强,随滞育时间的延长,酶活性出现不同的变化,12℃和20℃下,山梨醇脱氢酶活性呈现"U"形变化趋势,16℃下酶活性逐渐减弱,24℃下酶活性先升后降。16℃下,两种酶活性基本低于同时期其他温度处理。交互作用分析显示,温度和化蛹后天数的交互作用对滞育蛹海藻糖酶和山梨醇脱氢酶活性存在显著影响。上述结果表明,柑橘大实蝇自身生理状态应对不同越冬温度的反应不同,说明越冬温度与柑橘大实蝇滞育期生理水平有着密切的关系。本研究结果对深入了解温度调控柑橘大实蝇滞育期代谢生理机制有一定的参考价值。 相似文献
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J. W. L. Van Vuurde N. J. M. Roozen 《European journal of plant pathology / European Foundation for Plant Pathology》1990,96(2):75-89
Various isolation and serological techniques were compared for the detection ofErwinia carotovora subsp.atroseptica (Eca) andE. chrysanthemi (Ech) in cattle manure slurry containing c. 108 colony forming units (cfu) per ml. The slurry samples could be preserved at –80°C for 8 months without reduction in the number of bacteria but not at –20°C. Samples stored at –80°C were inoculated with concentrations of the target bacterium ranging from 102 to 108 per ml. Only immunofluorescence colony-staining (IFC) in combination with selective media was able to detect the target organism at a concentration of 100 cells per ml. No IFC-positive colonies were found in pour plates of the non-inoculated cattle slurry. The recovery of the target bacterium from slurry inoculated with 102 cfu of Ech per ml was 64% in PT medium (containing polygalacturonic acid) and 19% in crystal violet pectate medium (CVP). Recoveries of Eca were 32% and 82%, respectively. Ech and Eca could be detected at levels of 103 cfu per ml of slurry by isolation on CVP. Crude filtration procedures were necessary for analysis of slurry samples with immunosorbent immunofluorescence (ISIF) cell staining. The detection level of ISIF for Ech was 105 cells per ml of slurry. IF-positive cells were incidentally observed in the non-inoculated slurry. Detection of Ech and Eca with ELISA was only possible in slurry inoculated with 108 cells of the target bacterium per ml.Samenvatting Verschillende isolatie- en serologische methoden werden vergeleken om de doelbacteriënErwinia carotovora subsp.atroseptica (Eca) enE. chrysanthemi (Ech) aan te tonen in runderdrijfmest met een natuurlijke bacterieflora van ca 108 kolonievormende eenheden(cfu) per ml. De mestmonsters konden gedurende de onderzoekperiode tenminste 8 maanden bij –80°C worden bewaard zonder dat er een afname van het aantal levende mestbacteriën werd geconstateerd, terwijl bij bewaring bij –20°C wel een afname werd gevonden. De ontdooide mestmonsters werden geïnoculeerd met de doelbacterie in concentraties tussen 102 en 108 per ml. De laagste concentratie van de doelbacterie, 102 cfu per ml, kon alleen worden aangetoond met de immunofluorescentie-kleuring van bacteriekolonies (IFC) in een selectief medium. Met deze techniek was het percentage herisolatie vanuit drijfmest geïnoculeerd met 102 Ech cfu per ml respectievelijk 64% in PT-medium (bevat polygalacturonzuur) en 19% in kristalviolet pectine medium (CVP). Voor Eca bedroegen deze percentages respectievelijk 82% en 32%. In de niet-geïnoculeerde mestmonsters werden geen IFC-positieve kolonies gevonden. Via isolatie op CVP konden 103 of meer cfu van Eca en Ech worden aangetoond. Ruwe filtratie van de mestmonsters was nodig voor het aantonen van Eca- en Ech-cellen met immunoadsorptie immunofluorescentie microscopie. De detectiedrempel lag voor deze techniek op 105 bacteriecellen per ml mestmonster. In niet-geïnoculeerde mest werden incidenteel IF-positieve bacteriën gevonden. Het aantonen van Ech en Eca met ELISA was slechts mogelijk in mest geïnoculeerd met 108 of meer cellen van de doelbacterie per ml. 相似文献
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B.A. Fraaije M. Appels S.H. De Boer J.W.L. Van Vuurde R.W. Van den Bulk 《European journal of plant pathology / European Foundation for Plant Pathology》1997,103(2):183-193
Automated conductance measurements in polypectate medium were used for the detection of pathogenic soft rot Erwinia spp. in potato peel extracts. The detection threshold for Erwinia carotovora subsp. atroseptica (Eca) in inoculated peel extracts was ca. 104 colony forming units (cfu) ml-1 when samples were considered positive on the basis of a response within 48 h at 20 °C. Detection of E. chrysanthemi (Ech) was less sensitive, only 105 cfu ml-1 peel extract were detected within 36 h at 25 °C. The linear correlation between detection times in conductimetry and inoculum levels of Eca and Ech in peel extracts was used for a quantitative estimation of Eca and Ech in naturally contaminated peel extracts. Samples giving a positive conductimetric response had to be confirmed with an enzyme-linked immunosorbent assay (ELISA) or polymerase chain reaction (PCR) for the presence of Eca and Ech, because E. carotovora subsp. carotovora (Ecc) also generated a conductance response. Conductimetry was sensitive and efficient for detection of contamination levels of Eca higher than 104 cfu ml-1 peel extract. For Ech, conductimetric detection was less sensitive and inefficient due to low contamination levels of Ech and the presence of high numbers of Ecc in many samples after enrichment, which interfered with the test. Immunofluorescence cell staining (IF) combined with enrichment and immunofluorescence colony staining (IFC) were suited to detect and quantify low numbers of Eca and Ech at less than 104 cfu ml-1 in peel extracts. However, since false positive and negative reactions in serology were observed, the use of PCR after enrichment, or in combination with IFC to confirm positive results, was required for accurate detection. 相似文献
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M. C. M. Pérombelon 《European journal of plant pathology / European Foundation for Plant Pathology》1992,98(2):135-146
Blackleg is caused byErwinia carotovora subsp.atroseptica (Eca) andE. chrysanthemi (Echr) in cool and hot climates respectively. The bacteria are opportunistic pathogens and rely on their strong pectolytic character to infect plants when conditions favor their multiplication. Blackleg is a seedborne disease and the bacteria can survive in a quiescent form in lenticels and wounds during storage. The contaminated mother tuber and not the blackleg plant is the main source of progeny tuber contamination. Other sources of the pathogen are airborne (insects and aerosols) erwinias deposited on leaves and from there to the soil and progeny tubers, and erwinias in rotting tubers smeared into wounds incurred during mechanical crop handling.Most seed tubers are contaminated but blackleg incidence is related to seed contamination level modulated by soil water status. Competitiveness of the erwinias in the rotting mother tuber is affected by temperature, Eca is favored at <25°C and Echr at higher temperatures. The ubiquitousE. carotovora subsp.carotovora apparently fails to compete successfully with the other erwinias and saprophytic pectolytic bacteria in mother tubers and therefore does not cause blackleg.Disease control measures are based on avoiding tuber contamination by cultural means (early harvesting), reducing tuber contamination level (dry storage and hot water treatment) and planting ‘clean’ seed identified by quantifying tuber contamination rather than by visual crop inspection. Finally, recently identified highly resistant, even under anaerobic conditions, wildSolanum spp. could be used to breed for resistant cultivars by conventional methods or by genetic engineering. 相似文献
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J. M. Van der Wolf G. C. Gussenhoven 《European journal of plant pathology / European Foundation for Plant Pathology》1992,98(1):33-44
The specificity of two antisera raised to whole cells ofErwinia chrysanthemi (Ech), serogroup O1Ha, was studied in double antibody sandwich (DAS-) ELISA with 100 strains of different plant pathogenic bacteria (PPB), including 39 Ech strains, and of one of these antisera with 900 saprophytic bacteria isolated from extracts of potato peelings of Dutch seed potatoes grown in several production areas.All tested European Ech strains from potato reacted positively while no reactions were observed with any of the other plant pathogenic bacterial species. Two saprophytes (A254 and A256), both identified as pectinolyticPseudomonas fluorescens species, cross-reacted strongly with polyclonal antibodies against Ech. Non-specific reactions were found in DAS-ELISA with 16 saprophytes. The detection limits for the individual saprophytes varied between c. 105 and 109 cells.ml–1. The non-specific reactions were also found with monoclonal antibodies (mca 2A4) against a proteinase K resistent epitope of Ech and with antisera against other plant pathogens including an antiserum against potato virus YN. The non-specific reactions were observed in DAS-ELISA, but not in Ouchterlony double diffusion or immunofluorescence colonystaining, whereas A254 and A256 reacted in all tests, but only with antibodies against Ech. When in making dilution series potato peel extracts were used instead of phosphate buffered saline with 0.1% Tween 20, the 14 non-specifically reacting saprophytes only reacted at concentrations of 109 cells.ml–1 or higher. Only one of these 14 saprophytes was able to multiply on injured potato tuber tissue.In contrast to most saprophytic strains, the saprophytes A254 and A256 reacted strongly in ELISA in dilutions series made with potato peel extracts. A256 was able to grow on potato tuber tissue but only under low oxygen conditions; A254 did not grow at all on potato tissue.Defatted milk powder or bovine serum albumin added to the dilution buffer for the enzymeconjugated antibodies, drastically reduced the non-specific reactions, but not the reactions with A254 and A256.To reduce the cross-reaction with A254, an Ech antiserum was absorbed with A254. This resulted in a substantial drop in antibody reaction with the homologous antigen in Ouchterlony double diffusion. 相似文献
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采用Echandi方法,测定了从国内不同植物上分离,经鉴定的3个种和亚种的软腐欧氏杆菌(Erwiniacarotovora var.carotovora Dye,E.carotovora var.otroseptica Dye,E.chrysanthemi Burkholder.etal.)的224个菌株产生细菌素的能力。指示菌为代表3个不同种的菌株。测定结果表明:224个试验菌株中能产细菌素的有97个,占总数的43.3％。经测定产生细菌素的专化性有所不同,约各有1/3菌株分别对3种、2种、1种指示菌有抑制作用。有些广谱的细菌素甚至对其它属的植物病原细菌也有抑制作用。少数种内专化的产细菌素菌株可望用于软腐欧氏杆菌种鉴定的辅助手段。电镜观察初步表明,这些细菌素可能是分子大小不同的两种类型的细菌素:小分子细菌素在电镜下不可见,大分子细菌素具有短杆状结构,颇似无头壳的噬菌体的尾部。 相似文献
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17.
J. M. Van Der Wolf J. R. C. M. Van Beckhoven E. De Boef N. J. M. Roozen 《European journal of plant pathology / European Foundation for Plant Pathology》1993,99(2):51-60
Sixteen bacterial strains, cross-reacting with antibodies againstErwinia chrysanthemi (Ech), were isolated from potato peel extracts, ditch water, and the rhizosphere of wheat, onion, sugar beet and chicory using the immunofluorescence colony-staining procedure. Based on fatty acid profiles, isolates were classified as belonging to thePseudomonas fluorescens group.These strains, together with two previously isolated cross-reactingP. fluorescens strains, crossreacted with polyclonal antibodies against Ech in immunofluorescence cell-staining, Ouchterlony double diffusion, and ELISA. Seventeen strains also reacted strongly with monoclonal antibodies against the lipopolysaccharides (LPS) of Ech in ELISA.Cell envelopes (CE) and proteinase-K-treated CE (mainly LPS) of cross-reacting bacteria were further characterized with SDS-PAGE and Western blotting. Based on CE protein and LPS patterns, the cross-reacting bacteria were classified into two groups, each existing of two subgroups. Both CE and proteinase-K-resistant antigens strongly cross-reacted on immunoblots with antisera against a wild type strain of Ech. With an antiserum against a LPS O-chain lacking mutant of Ech only protein bands but no proteinase-K-resistant antigens were detected on immunoblots. These data suggest that in all cases the highly antigenic LPS O-chain is responsible for the cross-reactions. 相似文献