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1.
Strain HX2 of Rahnella aquatilis has been previously reported as a potential biological control agent (BCA) of grapevine crown gall. The production of an antibacterial substance (ABS) was suggested to be an important factor in the biocontrol process. This study was undertaken to determine the antibacterial properties and mode of action of ABS. Isolation and purification of ABS from culture broth of strain HX2 was achieved by methanol exaction, chromatography on macroporous resin and silica gel, and high-performance liquid chromatography (HPLC). Physical and chemical characteristic analysis revealed that ABS may be a thermostable and alkali-sensitive substance containing sugar(s) and an unknown 285-nm absorbed substance. Antibacterial activity assays revealed that ABS displayed a broad activity spectrum against all the 13 test isolates of phytopathogenic bacteria including the genera of Agrobacterium, Clavibacter, Pectobacterium, Pseudomonas, and Xanthomonas. Agrobacterium spp. strains were more sensitive to ABS than other tested strains, with larger inhibition zones and lower minimal inhibitory concentration (MIC). ABS exhibited a bactericidal effect against A. vitis both in vitro and in vivo. This compound did not cause bacterial cell lysis, as determined by morphological observation with an electron microscope. Also, no leakage of cytoplasmic materials from cells of A. vitis occurred after treatment with ABS at concentrations equivalent to the MIC. However, an inhibition of the incorporation of radiolabelled precursors into RNA and protein was observed after treatment with ABS. These results suggest that ABS inhibits RNA and protein syntheses in tumorigenic A. vitis.  相似文献   

2.
Erwinia amylovora is the bacterium responsible for fire blight, a necrotic disease affecting many rosaceous plants and especially pear tree and apple tree. A protein named harpin, secreted through the Hrp secretion pathway and able to elicit an hypersensitive reaction (HR) on tobacco has recently been isolated. Mutants inhrpN, the gene encoding harpin were described as non pathogenic on immature pear fruit and unable to elicit an HR on tobacco [Weiet al., 1992; Wei and Beer, 1993]. In this paper, the phenotype on plant ofhrpN mutants was carefully determined.hrpN mutants expressed a weak but significant virulence on host plants. Furthermore, when infiltrated into tobacco leaf mesophyll, thehrpN mutants elicited varied responses that fluctuated from null reaction to full necrosis of the infiltrated area. These results show that harpin is not absolutely required neither for pathogenicity on host plant nor for elicitation of an hypersensitive reaction on tobacco. Furthermore, in all the tests performed, mutant blocked in harpin secretion remained non pathogenic and unable to elicit an HR on tobacco. This suggests that factor(s), different from harpin, involved both in pathogenicity and HR eliciting ability is (are) secreted through the Hrp secretion pathway.Abbreviations HR hypersensitive reaction - NSI necrosis severity index - CFU colonie forming units  相似文献   

3.
建立了一种测定元胡、浙贝母和白芍3种中药材中嘧霉胺残留的气相色谱分析方法。样品经二氯甲烷提取,液-液分配,柱层析[m（中性氧化铝）:m（弗罗里硅土）=3:2]净化,气相色谱-氮磷检测器（GC-NPD）检测,基质匹配外标法定量,同时经气相色谱-串联质谱（GC-MS/MS）验证。结果表明:在0.02~10 mg/L范围内,分别以正己烷和3种空白基质溶液为溶剂配制嘧霉胺的标准工作溶液,峰面积与相应的质量浓度间呈良好的线性关系,R2均大于0.999;在0.01（元胡中为0.02）、0.1和0.5 mg/kg 3个添加水平下,添加回收率为75%~104%,日内相对标准偏差（RSD）为1.3%~10.2%（n=5）,日间RSD为1.2%~9.0%（n=3）。分析方法在元胡、浙贝母和白芍中的定量限（LOQ）分别为0.02、0.01和0.01 mg/kg。该方法的灵敏度、准确度及精密度均符合农药残留分析要求,且简单易行,净化效果好,适用于元胡、浙贝母和白芍中嘧霉胺分析。  相似文献   

4.
Of the numerous fractions obtained from the crude plant extracts of Chenopodium ambrosioides, Conyza dioscordisis and Convolvulus arvensis, 11 were found to be active against stored grain pests. The chemical structures of compounds present in the 11 fractions were found by gas chromatography/mass spectrometry to include predominantly long-chain fatty acid esters of hexadecanoic, arachidonic, and octadecanoic acids.  相似文献   

5.
In 2005, characteristic symptoms of crown gall on grapevines (Vitis vinifera L. cv. Muscat of Alexandria, and cv. Seto Giants) were observed in a commercial greenhouse-orchard in Okayama Prefecture, Japan. Isolations from diseased tissues consistently yielded bacterial colonies that were white, glistening, and produced abundant polysaccharide on potato semi-synthetic agar (PSA) medium. Ten representative isolates were chosen for further characterization. A multiplex polymerase chain reaction (PCR) assay showed these strains were not Rhizobium vitis but did possess a Ti plasmid. The bacteriological characteristics of the isolates corresponded well with R. radiobacter. The almost complete 16S ribosomal DNA sequences of isolates AT06-1 and AT06-2, selected from 10 grapevine isolates, were determined and corresponded to sequences of R. radiobacter. The pathogenicity of the isolates was tested on young grapevine and tomato (Lycopersicon esculentum Mill.) plants. Gall symptoms developed on both plant species after inoculation, and bacteria with the same colony morphology as those inoculated were reisolated. Based on these results, the isolates were identified as R. radiobacter (Ti). This report is the first of the occurrence of R. radiobacter (Ti) on grapevine in Japan. Phylogenetic analyses using the partial nucleotide sequences of virC operon located on a Ti plasmid showed that the isolate of R. radiobacter (Ti) isolated from grapevine and some strains of R. vitis (Ti) belonged to the same monophyletic group, which differed from the groups of R. radiobacter (Ti) isolated from plants other than grapevine and of the majority of R. vitis (Ti) strains isolated from grapevine. The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under accessions AB306890, AB306891, and AB465432–AB465459.  相似文献   

6.
为明确茶园天敌与假眼小绿叶蝉Empoasca vitis之间关系的变化动态,用灰色关联度法和生态位法研究天敌与假眼小绿叶蝉在数量、时间、空间上的关系,综合分析假眼小绿叶蝉优势种天敌位次在茶树品种间和年份间的差异。结果显示,根据密切指数之和比较发现,2015年在乌牛早茶园与白毫早茶园中,与假眼小绿叶蝉跟随关系密切的前3位天敌中均有锥腹肖蛸Tetragnatha maxillosa和粽管巢蛛Clubiona japonicola,这2种天敌在2种茶园中的密切指数之和分别为5.493和5.784;2016年天敌中均有八斑球腹蛛Theridion octomaculatum和粽管巢蛛,这2种天敌在2种茶园中密切指数之和分别为5.296和5.448。2015年与2016年相比,乌牛早茶园前3位天敌中均有粽管巢蛛,白毫早茶园前3位天敌中均有锥腹肖蛸和粽管巢蛛。表明2个品种茶园不同年份假眼小绿叶蝉前3位优势种天敌为粽管巢蛛、锥腹肖蛸和八斑球腹蛛。  相似文献   

7.
Tumour tissue samples were collected from vines grown in various regions of Italy and other parts of Europe and extracted for detection of Agrobacterium vitis. Fifty strains were isolated on agar plates and screened by PCR with consensus primers from the virD2 gene. They were confirmed as A. vitis with a species-specific monoclonal antibody. The isolates were further analyzed by PCR for their opine synthase genes and ordered into octopine, nopaline and vitopine strains. Primers designed on the octopine synthase gene did not detect octopine strains of Agrobacterium tumefaciens. For quantitative PCR, virD2 fragments were sequenced: two classes of virD2 genes were found and two primer sets designed, which detected octopine and nopaline strains or only vitopine strains. For simultaneous identification of all opine-type strains, multiplex real-time PCR with either primer pair and SYBR Green was performed: the combined sets of primers gave signals with DNA from any A. vitis strain. Specificity of the new primers for real-time PCR was evaluated using several unidentified bacterial isolates from grapevines and other plant species. An elevated level of non-specific background was observed when the combined primer sets were used in multiplex PCR assays. The real-time PCR protocol was also used to detect A. vitis cells directly from grapevine tumours; avoiding direct isolation procedures a sensitivity in the range of one to ten cells per assay was found. Inhibition of the PCR reaction by plant material was overcome by treating tumour extracts with a DNA purification kit as a step for the isolation of nucleic acids.  相似文献   

8.
建立了QuEChERS-超高效液相色谱-串联质谱 (UPLC-MS/MS) 同时测定浙贝母鲜样和干样中吡虫啉、啶菌齅唑、氟唑菌酰胺、嘧菌环胺、吡唑醚菌酯和阿维菌素共6种农药残留量的方法。样品用V(乙腈) : V(水) = 4 : 1混合溶液提取,经C18 50 mg + PSA 50 mg + MgSO4 150 mg净化,采用UPLC-MS/MS电喷雾离子化正离子扫描和多反应监测模式 (MRM) 测定,基质匹配标准溶液外标法定量。结果表明:在0.001~0.2 mg/L 范围内,6种农药的质量浓度与相应的峰面积间呈良好的线性关系,相关系数均大于0.994。6种农药在浙贝母鲜样中的定量限 (LOQ) 为0.01 mg/kg,检出限 (LOD) 为2×10–5~3×10–4 mg/kg,在干样中的LOQ 为0.05 mg/kg,LOD 为1×10–4~1×10–3 mg/kg。在0.01、0.5 和 2 mg/kg 添加水平下,6种农药在浙贝母鲜样中的平均回收率均在80%~109%之间,相对标准偏差 (RSD) 在0.95%~13%之间 (n = 5);在0.05、0.5 和 2 mg/kg 添加水平下,6种农药在浙贝母干样中的平均回收率均在77%~101%之间,RSD在0.89%~7.5%之间 (n = 5)。经实际样品检测验证,该方法快速、简便、可靠、高效,适用于同时检测浙贝母中吡虫啉、啶菌齅唑、氟唑菌酰胺、嘧菌环胺、吡唑醚菌酯和阿维菌素等农药的残留。  相似文献   

9.
The tea green leafhopper Empoasca vitis Göthe (Homoptera: Cicadellidae) has become the key insect pest affecting tea plantations in southern China. The control of E. vitis with chemical insecticides is not only costly but also has serious undesirable effects. It is therefore imperative that environmentally sound and sustainable management alternatives be developed. The aim of this study was to evaluate the potential of using the non-host plant Catsia tora (Fabaceae) as a cover crop for the control of E. vitis in tea plantations. The volatile organic compounds derived from the C. tora plants were shown to be repellent to leafhoppers. These compounds were collected through headspace absorption, and its 19 components were identified through gas chromatography–mass spectrometry. Based on the behavioral responses of E. vitis to 11 of these volatile compounds (those with relative amounts greater than 1), three bioactive compounds (p-cymene, limonene, and 1,8-cineole) were observed to play a key role in the regulation of the E. vitis population and were significantly effective at repelling leafhoppers. The intercropping of C. tora in a tea field markedly reduced the E. vitis population levels and promoted an increase in the natural enemies of this pest, such as spiders, coccinellids, and lacewings. Our findings suggest that the sustainable pest management achieved using C. tora as a cover crop offers an interesting alternative strategy for the control of this important tea pest.  相似文献   

10.
A genomic library of Erwinia amylovora isolate T was constructed in the cosmid pLAFR3 and maintained in Escherichia coli. Clones were transferred individually by conjugation into the non-pathogenic isolate P66 of E. amylovora. Transconjugants were screened for restoration of pathogenicity to pear by stab inoculation into sections of immature pear fruits. Three clones complemented P66 restoring pathogenicity and ability to cause the hypersensitive reaction (HR) in Phaseolus vulgaris. Restriction mapping and hybridization experiments showed that the three clones had a common 3·7 kb fragment of E. amylovora DNA. Sub-cloning and insertion mutagenesis with Tn5-lac confirmed that a determinant of pathogenicity and ability to cause the HR (hrp gene) was located on a 2·1 kb HindIII/BamHI fragment within the common DNA. Hybridization experiments using the 2·1 kb HindIII/BamHI fragment as a probe demonstrated that the hrp gene was located in the chromosome of isolate T and that homologous sequences were present in the non-pathogenic isolates P66 and S. Clones which restored hrp function did not affect the growth of isolate P66 in minimal or nutrient-rich media. Transconjugants of Pseudomonas syringae pv. phaseolicola race 1 harbouring the hrp gene(s) cloned from E. amylovora did not cause the HR in susceptible cultivars of bean but symptoms developed more slowly than in the absence of the clones or with pLAFR3 alone.  相似文献   

11.
Several members of the bacterial genus Brenneria are pathogenic on different tree species. Cell-free extracts from the bacterial phytopathogens Brenneria rubrifaciens, B. salicis, and B. nigrifluens induced production of the red pigment rubrifacine in the B. rubrifaciens bruI insertional mutant Br-212. Analysis of the bruI locus identified an adjacent open reading frame, designated bruR, with homology to luxR. High-performance liquid chromatography and mass spectrometry analysis of ethyl acetate extracts from wild-type B. rubrifaciens and Escherichia coli expressing the bruI gene identified two acyl homoserine lactone (AHL) peaks, N-(3-oxohexanoyl)-homoserine lactone (3OC6HSL) and N-hexanoyl-homoserine lactone (C6HSL). Addition of synthetic 3OC6HSL and C6HSL at 10 μM to the bruI mutant, strain Br-212, induced rubrifacine production and the ability to elicit a hypersensitive reaction (HR) in tobacco leaves. Synthetic C6HSL was less effective at inducing pigment production than 3OC6HSL at 10 μM. The bruI mutant Br-212 did not produce detectable AHLs, indicating that C6HSL and 3OC6HSL are the major AHLs produced by this species. The AHLs N-heptanoyl-DL-homoserine lactone (C7HSL), N-octanoyl-DL-homoserine lactone (C8HSL), and N-(3-oxooctanoyl)-DL-homoserine lactone (3OC8HSL) also induced pigment production in Br-212 and restored its ability to elicit an HR in tobacco, suggesting that cross-talk with other bacterial species may be possible.  相似文献   

12.
Forty-one strains of Rhizobium vitis, either tumorigenic (Ti) or nonpathogenic, were characterized using multilocus sequence analysis (MLSA) of the partial nucleotide sequences of pyrG, recA, and rpoD. The strains separated into seven clades. Rhizobium vitis (Ti) strains isolated from Japan were divided into five genetic groups (A to E), and nonpathogenic R. vitis strains were divided into two genetic groups (F and G). This result suggests that there are new genetic groups of R. vitis in Japan. Among these groups, members of A and B groups are widely distributed throughout Japan.  相似文献   

13.
Intercellular washing fluid (IWF) obtained from the susceptibleArabidopsis accession Ws-eds1 inoculated withPeronospora parasitica isolate Emoy-2, contained an elicitor of necrosis with ecotype specificity towardsArabidopsis accessions with particular resistance genes. This elicitor caused necrosis on the highly resistant accessions La-er, Nd-1 and partly on Col-5, but not on the susceptible accessions Ws-eds1 and Oy-0. In resistant plants, injection of IWF caused hypersensitive reaction (HR)-like cell collapse which was associated with the accumulation of phenolics and lignin-like material in walls of cells undergoing cell death. The elicitor is sensitive to proteinase K and pronase enzymes, heating and autoclaving but insensitive to periodate oxidation, freezing and thawing, and is not dialyzable. Results suggest that the elicitor is a protein. Fractionation experiments using size-exclusion membranes revealed that elicitor activity has a molecular weight in excess of 100 kDa. http://www.phytoparasitica.org posting July 13, 2003.  相似文献   

14.
We have previously shown that flagellin of Pseudomonas syringae pv. tabaci is an elicitor that induces a hypersensitive reaction (HR) in nonhost tomato cells. Flagellin is the major HR elicitor produced by this pathogen, as shown by the inability of a flagellin-defective mutant, ΔfliC, to induce HR. Also, a ΔfliD mutant that secretes large amounts of monomer flagellins induces a strong HR in tomato. In this study, the possible involvement of an Hrp type III secretion system (TTSS) in flagellin-induced HR was investigated using flagella-defective mutants or Hrp TTSS-defective mutants. The hrcC gene encodes HrcC protein, which is required for Hrp pilus formation in the outer membrane. An hrcC mutation, introduced into the wild-type, ΔfliC, and ΔfliD mutants of P. syringae pv. tabaci did not affect swimming motility or flagellin secretion, whereas all ΔhrcC, ΔfliC, and ΔfliD mutants lost the ability to cause disease on host tobacco leaves. However, the ΔhrcC mutant and the ΔfliD/ΔhrcC double mutant were still able to induce HR cell death, expression of one of the defense-related genes hsr203J, and the generation of hydrogen peroxide in nonhost tomato cells. Thus, flagellin is required for both pathogenicity in host tobacco and HR in nonhost tomato. On the other hand, hrp TTSS is necessary for pathogenicity on host tobacco but is not indispensable to induce HR in nonhost tomato. These results clearly show that flagellin-induced HR is hrp-independent in tomato.The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under the accession number AB049570  相似文献   

15.
秦楠  郝林  李鑫 《植物保护学报》2015,42(5):813-819
为探究解淀粉芽胞杆菌Bacillus amyloliquefaciens HRH317菌株所产抗菌蛋白物质对禾谷镰孢菌Fusarium graminearum的抑制机理,采用硫酸铵分级沉淀菌株发酵液,DEAE Sepharose Fast Flow柱和Sephadex G-200柱层析分离纯化抗菌蛋白,聚丙烯酰胺凝胶电泳检测抗菌蛋白分子量;并采用平板计数法和扫描电子显微镜观察病原菌孢子萌发和菌丝生长情况。结果表明,硫酸铵饱和度为50%时HRH317菌株发酵液的抑菌活性最大,抑菌圈直径可达2.02 cm,显著大于其它饱和度处理;2次柱层析可纯化出抗菌蛋白,分子量约为36.0 k D。禾谷镰孢菌经该抗菌蛋白处理后12h,病原菌孢子萌发率为55.8%,显著低于对照,且2~12 h间其孢子萌发率平均下降了57.3%;该抗菌蛋白还可造成病原菌菌丝变细、畸形与扭曲,且末端膨大等现象。表明解淀粉芽胞杆菌HRH317菌株产生的抗菌蛋白对禾谷镰孢菌孢子萌发和菌丝发育具有较强的抑制作用。  相似文献   

16.
Tumorigenic and non-tumorigenic strains of Agrobacterium tumefaciens, A. rhizogenes, A.rubi, and A.vitis were examined using capillary isoelectric focusing, phenotypic determinative tests, PCR and fatty acid analysis. The isoelectric points (pI) of the 40 strains investigated clearly differentiated the strains according to their respective species. The different species were characterized with the following pI values: A. tumefaciens 2.2, A. rhizogenes 4.0, A. rubi 2.15, and A. vitis 2.6. This differentiation corresponded to the phenotypic, PCR and fatty acid characterizations. Strains with the similar chromosomal background but different plasmid content, e.g. A.vitis strain S4, and F2/5 gave the same pI values. Strains of Rhizobium species differed from Agrobacterium strains in their pI values. The advantage of capillary isoelectric focusing over the phenotypic determinative tests, PCR and fatty acid analysis is its speed (15 min), relative simplicity, and the very small amount of chemicals used. This rapid and simple method is a major improvement over the classical methods of separation of Agrobacterium species and should prove useful for rapid characterization of Agrobacterium-like colonies isolated from plant tumours for epidemiological and generic diversity studies.  相似文献   

17.
建立了超高效液相色谱-串联质谱 (UPLC-MS/MS) 检测氟虫双酰胺和噻嗪酮在茭白中残留的方法。样品采用乙腈提取,乙二胺-N-丙基硅烷 (PSA) 净化,0.1%甲酸-甲醇梯度洗脱,电喷雾正离子扫描,多反应监测模式,超高效液相色谱-串联质谱测定,外标法定量。结果表明:在0.005~1 mg/kg添加水平下,氟虫双酰胺和噻嗪酮在茭白植株和茭白中的平均回收率在81%~107%之间,相对标准偏差在4.2%~11%之间。消解动态规律符合一级动力学方程,氟虫双酰胺和噻嗪酮的半衰期分别为2.3 d和2.8 d,属易降解农药。最终残留试验结果表明:10% 阿维·氟酰胺悬浮剂按制剂用量450~675 g/hm2分别施药2和3次,间隔期5 d,距最后一次施药后7、14和21 d采样,氟虫双酰胺在茭白中的残留量均<0.01 mg/kg;25% 噻嗪酮可湿性粉剂按制剂用量600~900 g/hm2分别施药2和3次,间隔期5 d,距最后一次施药后7、14和21 d采样,噻嗪酮在茭白中的残留量为<0.005~0.078 mg/kg。建议10%阿维·氟酰胺悬浮剂最高制剂用量为450 g/hm2,最多施药2次,安全间隔期以7 d为宜;25%噻嗪酮可湿性粉剂最高制剂用量为675 g /hm2,最多施药2次,安全间隔期以21 d为宜。  相似文献   

18.
Modes of passive dispersal of the grape rust mite Calepitrimerus vitis (Eriophyoidea) were investigated in a vineyard of South-Western Germany. More than 200 Eriophyoidea per month were trapped in a wind chamber during summer (32,1% C.vitis) suggesting long-distance dispersal by air currents. Rain washed part of the adult C.vitis population from the foliage. SE micrographs suggest that quiescent nymphs are affixed to the leaf by a substance of unknown nature. However, the role of rain in C.vitis colonisation of uninfested vineyards is still unclear, as is the role of phoretic transport by arthropods. For the first time, evidence of rust mite dispersal by human activity is presented. A large number of C.vitis was found adhering to clothes and hands of workers carrying out customary cultural practices in the vineyard. Other arthropods, including Typhlodromus pyri, the main predator of C.vitis, were also passively transported by wind, rain and human activity.  相似文献   

19.
为获得对马铃薯晚疫病菌Phytophthora infestans具有拮抗效果的内生细菌,采用平板对峙法对21株分离自东祁连山高寒草地牧草的内生细菌进行抑菌能力测定,并结合形态学特征和16S r DNA基因序列同源性分析对拮抗菌株进行鉴定。结果表明,分离的21株内生细菌对马铃薯晚疫菌均有一定的抑制作用,其中262AY11、262AY6和264AY2抑菌效果最好,抑菌率分别为78.41%、78.03%和75.38%,根据16S r DNA基因序列分析将其分别鉴定为韦氏芽胞杆菌Bacillus weihenstephanensis、解淀粉芽胞杆菌B.amyloliquefaciens和枯草芽胞杆菌B.subtilis,在Gen Bank中的登陆号分别为KC414703、KC441759和HQ202817。262AY11和262AY6具有固氮能力;262AY6具有分泌IAA能力,在不含和含色氨酸的金氏培养基中产IAA的量分别为9.94 mg/L和14.79mg/L,且对马铃薯坏疽病菌Phoma foveata、马铃薯枯萎病菌Fusarium avenaceum和马铃薯炭疽病菌Colletotrichum coccodes的抑菌率分别为70.25%、62.38%和81.75%;264AY2对3种病原菌的抑菌率分别为67.09%、50.75%和75.99%。表明芽胞杆菌262AY11、262AY6和264AY2能有效抑制马铃薯晚疫病菌的生长,具有固氮和产IAA的生物学功能,且对另外3种马铃薯病原真菌也具有一定的生防潜力,可作为生防菌进行开发和利用。  相似文献   

20.
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