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1.
综述了国内外在苜蓿细菌性病害方面的研究概况。全世界现已报道9种苜蓿细菌性病害,它们是由8种细菌病原引起的,这些细菌分属于6个属。其中由革兰氏阳性细菌引起的病害1种,即苜蓿细菌性萎蔫病(Clavibacter michiganensisi subsp. insidiosus);革兰氏阴性菌引起的病害有8种,即细菌性芽萎蔫病(Erwinia persicina),细菌性芽腐烂病(Erwinia chrysanthemi),根癌土壤杆菌(Agrobacterium tumefaciens),冠腐和根腐综合病(Pseudomonas viridiflava),细菌性叶斑病和细菌性猝倒病为同一病原(Xanthomonas campestris pv. alfalfae),细菌性茎疫病(Pseudomonas syringae pv. syringae)和矮化病(Xylella fastidiosa)。我国已报道危害较轻的细菌性叶斑病(X. campestris pv. alfalfae)和细菌性茎疫病(P. syringae pv. syringae)以及2012年新报道的细菌性芽萎蔫病(E. persicina)。本文归纳了苜蓿细菌性病害的种类及其特征;列举了病害的分布及其寄主范围;详述了它们的症状识别与病原表型特征;介绍了当前常见的苜蓿细菌病原鉴定方法,即形态、生理和生化表型特征鉴定,系统发育如16S rRNA序列分析以及自动化数值分析如Biolog自动鉴定系统。 相似文献
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细菌性干枯病是发生在广东杧果产区杧果上的一种新病害，为明确引起该病的病原菌种类，通过稀释涂布法分离和纯化、致病性测定、并结合形态学、生理生化特性及16S rDNA序列分析对该病原菌进行鉴定。结果表明，从杧果干枯病病样中分离获得的NY01菌株，革兰氏染色呈阴性、菌体杆状。最适温度28℃，最适pH为7.0～7.2。刺伤接种到健康杧果叶片，4天后在杧果的叶片呈现典型病斑。以NY01菌株的DNA为模板，扩增获得的16S rDNA片段，经BLAST分析，NY01菌株16SrDNA序列与Sphingomonas sanguinis（登录号KT766073）的亲缘关系最近，同源性达到99.63%。经形态学、生理生化及16S rDNA序列分析将杧果细菌性干枯病病原菌NY01菌株鉴定为血红鞘氨醇单胞菌（Sphingomonas sanguinis），血红鞘氨醇单胞菌能够引起杧果细菌性干枯病在国内属首次报道。 相似文献
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由柑橘黄单胞菌杧果致病变种(Xanthomonas citri pv．mangiferaeindicae, Xcm)引起的杧果细菌性角斑病是杧果上的一种重要细菌性病害,严重危害杧果产业发展。为了研究抗毒素phd基因的结构和功能,本试验通过在NCBI上选取已被注释的Xcm抗毒素phd基因的序列,设计出引物phd-F和phd-R,从Xcm003菌株中扩增出258 bp的DNA片段,提交至NCBI的登录号是MH401645。经序列分析该片段是一个完整的phd基因,编码85个氨基酸,其理论分子量9.68 kD,理论等电点为9.39,是一种亲水性蛋白,无信号肽,有6个磷酸化位点以及一个结构域。系统进化树结果显示,该基因的氨基酸序列与Xanthomonas citri pv.mangiferaeindicae LMG941的Phd蛋白聚为一类。通过克隆杧果细菌性角斑病菌phd基因并对其进行生物信息学分析,为深入研究phd基因做基础。 相似文献
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苜蓿假盘菌ISSR反应体系优化及指纹图谱构建 总被引：4，自引：1，他引：3
以北京苜蓿假盘菌(Pseudopeziza medicaginis (Lib.) Sacc.)菌株为试材,用ISSR-23引物[序列为(AC)8T]研究PCR反应体系的主要成分及退火温度对假盘菌ISSR扩增的影响,以期为苜蓿假盘菌的深入研究奠定基础。结果表明:Primer、Mg2+、Taq酶和dNTP浓度对扩增均有影响,以Primer影响最为明显;优化的反应体系为:20μL反应体系中含0.1 ng模板DNA,2.0mmol/L MgCl2、0.5μmol/L Primer、1U Taq DNA聚合酶、0.2 mmol/LdNTP,2μL 10×PCR Buffer和2.5%去离子甲酰胺;在此基础上,建立6个不同地理来源苜蓿假盘菌菌株的指纹图谱并分析其亲缘关系;对44条ISSR引物进行筛选,22条可产生清晰稳定的多态性条带,其中16条可将6个供试菌株完全分开,建立了苜蓿假盘菌指纹图谱;经聚类分析,在0.55遗传相似水平下,6个供试菌株分为两个遗传谱系,菌株的遗传谱系与其地理来源之间不表现相关性。 相似文献
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番茄细菌性叶斑病菌的拮抗菌筛选、鉴定及其拮抗性能评价 总被引：5，自引：0，他引：5
采用含菌平板抑菌圈测定法,从120株高寒草地牧草内生细菌中分离筛选到一株对番茄细菌性叶斑病拮抗能力较强的菌株264ZY7,其抑菌圈直径为1.22 cm。264ZY7菌体短杆状,大小为1.534 μm×0.571 μm~3.210 μm×0.781 μm,革兰氏阳性菌,根据形态特征,并结合16S rDNA和gyrB基因序列相似性分析,将菌株264ZY7鉴定为解淀粉芽孢杆菌Bacillus amyloliquefaciens。通过室内盆栽防效试验,结果表明该菌株对番茄细菌性叶斑病的防效达69%,此外,该菌株对8种病原真菌具有抑菌能力,说明抑菌谱较宽,且抑菌率均大于30%,对番瓜根腐病菌(Fusarium sp.)效果最明显,抑菌率为52.84%。该研究结果为高寒草地紫花针茅内生细菌264ZY47的进一步开发利用提供了依据。 相似文献
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基于16S rRNA基因的枯草芽孢杆菌PCR快速检测方法的建立 总被引：1，自引：0，他引：1
参照GenBank中登录的枯草芽孢杆菌16S rRNA基因,设计1对引物,扩增片段大小为460bp的基因片段,该方法对枯草芽孢杆菌检测的灵敏性为1pg总DNA量,对枯草芽孢杆菌DNA的扩增结果为阳性,对照菌株扩增结果均为阴性,成功地建立了枯草芽孢杆菌PCR检测方法。该方法具有快速、灵敏度高、特异性强等优点,为枯草芽孢杆菌的分离及鉴定奠定了良好的基础。 相似文献
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《中国饲料》2016,(12)
为筛选调制优质动物苜蓿青贮饲料的优良乳酸菌,本试验以从郑州地区9个苜蓿样品中分离得到的乳酸菌为试验材料,经抗菌活性、生长特性及产酸特性筛选菌株,根据16S r DNA序列,结合生理生化特征进行菌种鉴定,并通过发酵苜蓿粉考察菌株的发酵特性。结果表明,从分离的19株乳酸菌中筛选到4株具有高抗菌活性的菌株,其中菌株ZZU 457和ZZU 466在MRS液体培养基中生长及产酸性能较好,均能够耐受6.5%的盐浓度环境;菌株ZZU 457耐受10~45℃,以及p H为3.5的环境,而菌株ZZU 466能够耐受5~50℃,以及p H为4.0的环境;苜蓿粉发酵结果表明接种ZZU 457或ZZU 466在发酵18 h内均能快速产酸,其中接种ZZU 466的苜蓿粉发酵36 h时p H值达到4.2;结合理化特征及16S r RNA序列分析结果,ZZU 466被鉴定为融合魏斯氏菌。优良菌株ZZU 466在苜蓿青贮饲料调制加工中具有潜在的应用价值。 相似文献
10.
采用16S rDNA序列分析及ERIC-PCR对从贵州修文贵长猕猴桃枝条、叶片、花蕾分离到的35株猕猴桃溃疡病菌进行遗传多样性分析。通过16S rDNA序列分析,鉴定所分离的病原菌为Pseudomonas syringae pv.actinidiae,并运用ERIC-PCR分析病菌群体的遗传多样性分析,相似系数为0.808时,35株菌株分为4个类群,85.7%的菌株属于第一个类群,且菌株无明显的采集地、采集部位的聚类;表明修文猕猴桃溃疡病菌具有丰富的遗传多样性;此研究为溃疡病的检测、防治提供科学依据。 相似文献
11.
Schrenzel M Nicolas M Witte C Papendick R Tucker T Keener L Sutherland-Smith M Lamberski N Orndorff D Heckard D Witman P Mace M Rimlinger D Reed S Rideout B 《Veterinary microbiology》2008,126(1-3):122-131
Mycobacterium avium subsp. avium and Mycobacterium intracellulare are primary causes of mycobacteriosis in captive birds throughout the world, but little is known about how they are transmitted. To define the local epidemiology of infection, we strain-typed 70 M. avium subsp. avium and 15 M. intracellulare culture isolates obtained over a 4-year period from captive birds. Typing was performed using randomly amplified polymorphic DNA (RAPD) PCR, amplified fragment length polymorphic (AFLP) fragment analyses, and for a subset of isolates, DNA sequencing of a segment of the 16S-23S rRNA internal transcribed spacer region. Six strain clusters comprising 43 M. avium subsp. avium, isolates were identified; 42 isolates had unique typing patterns, including all M. intracellulare isolates. Phylo-geographical analyses using RAPD and AFLP fingerprints and animal confinement histories showed no correlation between housing of infected birds and mycobacterial strain-type, except for two animals. The diversity of M. avium subsp. avium and M. intracellulare isolates and minimal evidence for bird-to-bird transmission suggest that environmental reservoirs may be important sources of infection in captivity. 相似文献
12.
The 16S rRNA gene of 39 S. equi subsp. zooepidemicus strains and two S. equi subsp. equi strains was amplified by polymerase chain reaction and subsequently digested with the restriction enzyme Hinc II. A restriction profile with two fragments with sizes of 1250 bp and 200 bp could be observed for both S. equi subsp. equi strains and for 30 of the 39 S. equi subsp. zooepidemicus strains indicating a sequence variation within the V2 region of the 16S rRNA gene of the remaining nine S. equi subsp. zooepidemicus isolates. A segment of the 16S rRNA gene including the hypervariable V2 region of 11 S. equi subsp. zooepidemicus and two S. equi subsp. equi could be amplified by PCR and sequenced. The sequence of the V2 region of eight S. equi subsp. zooepidemicus strains appeared to be identical or almost identical to the sequence of the two S. equi subsp. equi strains. The sequence of the remaining three S equi subsp. zooepidemicus strains differed significantly from the sequence of S. equi subsp. equi. These differences allowed a division of S. equi subsp. zooepidemicus strains into two 16S rRNA types and might possibly have consequences for the taxonomic position of these phenotypically indistinguishable strains of one subspecies. A molecular typing could additionally be performed by amplification of the gene encoding the 16S-23S rRNA spacer region. A single amplicon of the spacer gene of 1100 bp could be observed for one S. equi subsp. zooepidemicus, an amplicon of 950 bp for two S. equi subsp. equi strains and 10 S. equi subsp. zooepidemicus strains, a amplicon of 780 bp for 27 S. equi subsp. zooepidemicus strains and a single amplicon of 600 bp for one S. equi subsp. zooepidemicus strain. The variations of the V2 region of the 16S rRNA gene and the size variations of the 16S-23S rRNA spacer gene were not related to each other. Both variations could be used for molecular typing of this species, possibly useful in epidemiological aspects. 相似文献
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Yamashita K Shimizu A Kawano J Uchida E Haruna A Igimi S 《The Journal of veterinary medical science / the Japanese Society of Veterinary Science》2005,67(3):263-268
Staphylococci were isolated from the external auditory meatus in 14 (48.3%) of 29 dogs affected with otitis externa (OE dogs) and 28 (68.3%) of 41 dogs without OE (non-OE dogs). Twenty-two OE isolates were identified as belonging to 12 species, and 42 non-OE isolates were identified as belonging to 13 species. The predominant species found in both OE and non-OE isolates were S. intermedius, and S. epidermidis. Thirty-eight (59.4%) of 64 isolates were resistant to one or more of the 17 antimicrobial agents tested. Resistance to PCG and ABPC was most frequent. S. schleiferi subsp. coagulans, a recent etiologic agent of canine OE, was isolated from OE and non-OE dogs. All of the 5 S. schleiferi subsp. coagulans isolates showed typical characteristics. No clear difference in the extracellular enzyme or toxin profiles, nor in the PFGE patterns, was demonstrated between the OE and non-OE isolates of S. schleiferi subsp. coagulans. A new PCR primer set specific for 16S rDNA was designed to identify strains of S. schleiferi subsp. coagulans. The amplified fragment was detected in all of the 5 isolates as well as the type strain GA 211 (=JCM 7470) and a reference strain GA 11, but was not detected in any strains of the related species, S. aureus, S. intermedius and S. hyicus. The PCR may allow a simple, rapid and precise identification of S. schleiferi subsp. coagulans, in addition to the standard tube test for free coagulase. 相似文献
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In this study, by using the polymerase chain reaction (PCR) diagnosis for the detection and identification of Mycoplasma, we investigated mycoplasmas contaminating the semen of yearling bulls affected by seminal vesiculitis. The bulls presented neither subclinical nor clinical contagious bovine pleuropneumonia signs and the complement fixation test for specific antibodies was negative. Furthermore, we have investigated mycoplasmas isolated from semen of healthy breeding bulls of several breeds and origins, which routinely underwent breeding soundness examinations and presented no clinical signs of seminal vesiculitis. We were able to demonstrate mycoplasma infection in all tested samples by i) growth on mycoplasma-specific media and ii) a PCR-based method using a mycoplasma-specific MGSO/GPO1 primer set to amplify the 16S fragment rDNA. In addition, the identification of Mycoplasma species was made by PCR using the MSC1/MSC2 primer set that specifically amplifies M. mycoides subsp. mycoides SC or the MM450/MM451 primer set followed by AsnI digestion analysis in order to identify M. mycoides subsp. mycoides LC. The data presented herein clearly show that M. mycoides subsp. mycoides SC infection was associated with seminal vesiculitis while M. mycoides subsp. mycoides LC was only found in bull semen from healthy control animals. Our findings confirm that the M. mycoides subsp. mycoides SC is shed in the sperm making the ejaculate a valuable biological sample for the isolation of these bacteria from serologically negative animals. Although the pathogenic role of M. bovigenitalium in bull seminal vesiculitis has been established, our clinical findings, semen characteristics, microbiological and bacterial genomic analysis strongly suggest that M. mycoides subsp. mycoides SC may contribute to induce vesicular adenitis in the bull. 相似文献
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Woubit S Lorenzon S Peyraud A Manso-Silván L Thiaucourt F 《Veterinary microbiology》2004,104(1-2):125-132
Contagious caprine pleuropneumonia is a severe infectious disease of goats in Africa and the Middle East. It is caused by a fastidious mycoplasma, Mycoplasma capricolum subsp. capripneumoniae, a member of the "M. mycoides cluster". Members of this cluster share genomic and antigenic features, which result in common biochemical and serological properties, complicating species identification. Two species of this cluster, M. mycoides subsp. capri and M. mycoides subsp. mycoides large colony biotype, are very often isolated from clinical cases resembling contagious caprine pleuropneumonia. Furthermore, in the laboratory, M. capricolum subsp. capripneumoniae can be easily confused with the closely related capricolum subspecies. Considering these constraints and the scarcity of available methods for identification, a specific polymerase chain reaction was developed. A DNA fragment of 7109 bp containing genes coding for the arginine deiminase pathway (ADI) was chosen as target sequence for the selection of a specific primer pair. The full ADI operon from M. capricolum subsp. capripneumoniae strain GL100 was sequenced. Polymorphism within this locus was analyzed by comparison with the sequence from the closely related IPX strain (M. capricolum subsp. capricolum). It varied from 0.6% to 3.5%. The highest divergence was found in a region coding for arcD. Therefore, this gene was chosen as target for the specific amplification of a 316 bp-long DNA fragment. The specificity of this PCR was validated on 14 M. capricolum subsp. capripneumoniae strains and 27 heterologous strains belonging to the "M. mycoides cluster" and M. putrefaciens. This new PCR will be a valuable tool for the surveillance of contagious caprine pleuropneumonia. 相似文献
16.
Phylogeny of the Mycoplasma mycoides cluster as shown by sequencing of a putative membrane protein gene 总被引：2，自引：0，他引：2
The Mycoplasma mycoides cluster is made of six species that are closely related both genetically and phenotypically. Two are of particular importance, M. mycoides subsp. mycoides SC causing contagious bovine pleuropneumonia and M. capricolum subsp. capripneumoniae causing contagious caprine pleuropneumonia. The sequences of a putative membrane protein gene and partial flanking open reading frames have been obtained from various strains in this cluster, including all reference strains. Sequence analysis showed this locus is present and fully conserved in all strains of M. mycoides subsp. mycoides SC isolated from geographically most distant places worldwide. In M. capricolum subsp. capripneumoniae polymorphism in this locus has been found at seven positions and revealed that they can be used as epidemiological markers. Conserved regions were used to define a primer pair that enables the amplification by PCR of two fragments 302 and 1298bp long, respectively. The 302bp long fragment contains an intergenic sequence that can be used for phylogenetic studies or for identification purposes. Parsimony analysis on an alignment of 49 DNA sequences show a subdivision of the M. mycoides cluster into two subgroups that is in accordance with results obtained by phenotypic methods. Two lineages exist within the capricolum subgroup, one of them clustering strains identified as M. capricolum subsp. capricolum, M. capricolum subsp. capricolum and M. sp Bovine Group 7. However M. capricolum subsp. capripneumoniae strains can readily be identified by three specific nucleotide positions or by sequencing the 1298bp long fragment. There is no clear subdivision within the mycoides subgroup, supporting the idea that M. mycoides subsp. mycoides LC and M. mycoides subsp. capri should not be separated into two subspecies. Mycoplasma mycoides subsp. mycoides SC strains can easily be distinguished as they bear an insertion sequence 15bp downstream from the stop codon of the membrane protein gene. 相似文献
17.
Akineden O Hassan A Alber J El-Sayed A Estoepangestie AT Lämmler C Weiss R Siebert U 《Veterinary microbiology》2005,110(1-2):147-152
The present study was designed to identify and compare 32 beta-hemolytic streptococci isolated from 28 different harbor seals of the German North Sea during the phocine distemper outbreak in 2002. The bacteria were identified as Streptococcus equi subsp. zooepidemicus based on cultural, biochemical, serological and molecular studies. Epidemiological investigations by PCR restriction fragment length polymorphism analysis of the 16S-23S rDNA intergenic spacer region and gene szp and by macrorestriction analysis of the chromosomal DNA of the strains by pulsed field gel electrophoresis revealed that all 32 strains appeared to be identical. These results indicate that a single bacterial clone seemed to be distributed among the harbor seal population of the German North Sea during this outbreak. 相似文献
18.
Akineden O Alber J Lämmler C Weiss R Siebert U Foster G Tougaard S Brasseur SM Reijnders PJ 《Veterinary microbiology》2007,121(1-2):158-162
The present study was designed to identify 15 beta-hemolytic streptococci isolated during a period between 1988 and 2005 from nine harbour seals and six grey seals from various origins of the North Sea. All isolates were identified as Streptococcus equi subsp. zooepidemicus. The bacteria were additionally investigated for relatedness by restriction fragment length polymorphism analysis of PCR amplified 16S-23S rDNA intergenic spacer region and gene szp and by macrorestriction analysis of chromosomal DNA of the strains by pulsed field gel electrophoresis. The molecular analysis yielded identical or closely related patterns within the strains of the present study and with the S. equi subsp. zooepidemicus strains isolated from harbour seals of German North Sea which were investigated previously [Akineden, O., Hassan, A.A., Alber, J., El-Sayed, A., Estoepangestie, A.T.S., L?mmler, C., Weiss, R., Siebert, U., 2005. Phenotypic and genotypic properties of S. equi subsp. zooepidemicus isolated from harbor seals (Phoca vitulina) from the German North Sea during the phocine distemper outbreak in 2002. Vet. Microbiol. 110, 147-152]. This indicates that this single or closely related bacterial clone existed during both phocine distemper virus epidemics in 1988 and 2002 and that a direct transmission of the strains has occurred between two seal species and between seal populations of far distant regions possibly with grey seals as a vector. 相似文献