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一种引起香石竹黄化病植原体的初步鉴定 总被引:1,自引:0,他引:1
植原体(phytoplasma)(原称类菌原体Myco- plasma-like Organism,简称MLO)是一类无细胞壁,存在于植物筛管细胞内的原核生物。迄今为止,世界上报道的植物植原体病害多达300余种, 且有不断增加的趋势。其主要症状包括丛枝、黄化、花变叶、花器褪化等。由于植原体至今未能分离培养成功,其培养性状、生化特性、营养需求等都无从得知,所以长期以来对其检测及鉴定技术的发展都没有突破性进展。近十多年来,PCR技术使植原体检测、鉴定与分类取得了令人瞩目的进展,对植原体16S rRNA基因的分析使人们从遗传本质上认识到了植原体与其它原核微生物间的差异,也使人们对植原体的检测达到了单拷贝基因水平。目前, 根据植原体16S rRNA和核糖体蛋白(rp)基因的序列分析形成了植原体分类和鉴定的基本框架。 相似文献
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植原体16S rDNA RFLP指纹图谱分析软件研究 总被引:1,自引:0,他引:1
植原体是一类寄生于植物韧皮部的原核生物。目前国际上主要根据植原体16S rDNA RFLP图谱的比对进行分类及鉴定。传统的PCR-RFLP试验步骤繁琐、重复性差,误差大,尤其在检测鉴定大量植原体病原时,耗时、耗力。通过对目前已公布的全部植原体16S rDNA序列进行比对整理,并生成16S rDNA RFLP电子指纹图谱库。在此基础上开发了指纹图谱分析软件DNA-FP Cluster1.0。该软件可基于序列或图谱形式进行比对,并自动输出比对结果。结果呈现形式为3种:即一种16S rDNA RFLP图谱与多种图谱间相似性分析;多种16S rDNA RFLP图谱间相似性分析;16S rDNA RFLP图谱间相似度树状图。该软件在不需要酶切试验操作的情况下实现了对植原体的快速分类与鉴定,并解决了因植原体材料不全而无法进行植原体酶切后比对的难题,为今后植原体候选种内的细化分类提供了方法与工具。 相似文献
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自从1967年首次在植物上发现类菌原体(Mycoplasmalike Organisms简称MLO是植物的病原菌以来,多种技术已应用于显现和证明感病植物中的类菌原体和螺原体(Spiroplasma)。电子显微镜观察是目前最重要和最普遍使用的类菌原体病害诊断方法,但此法昂贵且费时。近些年来,一些植病学家运用能与DNA结合的荧光染料 相似文献
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植原体TaqMan探针实时荧光PCR检测鉴定方法的建立 总被引:19,自引:1,他引:19
本研究成功建立了植原体分类鉴定和检测的TaqMan探针实时荧光PCR方法,该方法根据植原体16S rDNA保守区设计了1个TaqMan广谱探针和3个植原体组间点突变特异性探针,并对9种植原体和5种细菌以及3个植物样本进行实时荧光PCR。结果表明,用广谱探针可检测到所有植原体产生荧光信号,而细菌不产生荧光信号。当用植原体组间特异性探针检测时,仅能检测到该组植原体产生荧光信号,检测的敏感性比常规的PCR-电泳检测高约100倍、检测速度有较大提高。由于PCR产物是荧光探针检测,本方法特异性强,并可以用组特异探针直接确定植原体种类。实验采用完全闭管检测,降低了污染机会。本研究为其它原核生物、特别是不能培养菌、难培养菌的检测鉴定和分类提供了新方法。 相似文献
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通过透射电子显微镜,在表现卷叶、褪绿症状的丁香(Syringa oblata)样品的叶脉韧皮部筛管细胞内观察到大量植原体粒子。应用植原体16S rRNA基因通用引物对P1/P7和R16F2n/R16R2对表症丁香植株总DNA进行巢式PCR扩增,得到了约1.2 kb的目标片段,通过对扩增片段进行测序、系统发育分析和同源性分析,结果表明,该片段长度为1 246 bp,在系统发育进化树上与翠菊黄化组(Candidatus Phytoplasma asteris)成员是聚集在一起的,与该组成员同源性均在98%以上。用16Sr RNAⅠ组和Ⅴ组特异引物确定了该病害非混合侵染所致,相似性系数和RFLP分析表明该植原体属于16SrⅠ B亚组。这是国内关于翠菊黄化组植原体在丁香上感染的首次报道。 相似文献
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对仙人掌丛生幼嫩组织进行超薄切片电镜观察,在韧皮部筛管中存在大量植原体;根据植原体16S rRNA基因保守序列设计的通用引物对R16 F2/R2,应用PCR技术对仙人掌丛枝病进行分子检测,结果扩增到约1.2 kb的特异性片段,而在健康组织中却没有此特异片段;通过16S rDNA片段核酸序列同源性比较,结果表明仙人掌丛枝病植原体与花生丛枝病植原体亲缘关系最近,据此可初步判断仙人掌丛枝病植原体是一种属于16Sr Ⅱ组的植原体,基本确定了其分类地位。 相似文献
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Jaroslava Přibylová Karel Petrzik Josef Špak 《European journal of plant pathology / European Foundation for Plant Pathology》2009,124(1):181-185
Four Rhododendron hybridum plants (from cvs Moravanka and Don Juan), all exhibited symptoms of shortened axillary shoots, reduced leaves with
vein clearing and yellowing, undeveloped flowers, and general stunting in a rhododendron nursery garden in southern Bohemia
in 2007. Electron microscopy examination of ultra-thin sections revealed the presence of numerous polymorphic phytoplasma-like
bodies in the phloem tissue of leaf midribs and petioles. The phytoplasma etiology of this disease was further confirmed by
polymerase chain reaction (PCR) using universal phytoplasma primers. Restriction fragment length polymorphism (RFLP) analysis
of amplification products obtained with a R16F2/R16R2 primer pair from all symptomatic plants indicated the presence of phytoplasma
from the 16SrVI-A subgroup. A detailed comparison of the amplified sequences and phylogenetic analysis confirmed that the
phytoplasma belonged to the subgroup 16SrVI-A (clover proliferation phytoplasma group). This is the first report of the natural
occurrence of ‘Candidatus Phytoplasma trifolii’ in plants of Rhododendron hybridum. 相似文献
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Jam Nazeer Ahmad Samina J. N. Ahmad Muhammad Aslam Muhammad Aijaz Ahmad Nicoletta Contaldo Samanta Paltrinieri Assunta Bertaccini 《European journal of plant pathology / European Foundation for Plant Pathology》2017,149(1):117-125
A phytoplasma-associated disease was identified in Brassica campestris (sarson) plants during a survey conducted in Punjab province of Pakistan in 2014–2016. The symptomatic plants showed characteristic symptoms of phyllody and witches’ broom. Phytoplasma presence was detected by polymerase chain reaction on 16S ribosomal and tuf DNAs, followed by RFLP analysis and sequence comparison of the 16S rRNA and tuf genes. The phytoplasma detected was classified in a new ribosomal subgroup designed 16SrIX-H. The phytoplasma presence in phloem tissues of symptomatic sarson samples was also confirmed through light microscopy and transmission studies to healthy plants through dodder and the leafhopper Orosius albicinctus. This is the first report of identification of 16SrIX-H subgroup phytoplasma associated with sarson and its natural vector in Pakistan. 相似文献
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Transmission of ‘Candidatus Phytoplasma pyri’ by naturally infected Cacopsylla pyri to peach,an approach to the epidemiology of peach yellow leaf roll (PYLR) in Spain 下载免费PDF全文
Peach orchards in the northeast of Spain were severely affected in 2012 by a previously unreported disease in this area. The symptoms included early reddening, leaf curling, decline, abnormal fruits, and in some cases death of the peach trees. All the infected peach samples were positive for ‘Candidatus Phytoplasma pyri’, but none were infected by the ‘Ca. Phytoplasma prunorum’. In this work, potential vectors able to transmit ‘Ca. Phytoplasma pyri’ from pear to peach and between peach trees were studied and their infective potential was analysed at different times of the year. Transmission trials of the phytoplasma with potential vectors to an artificial feeding medium for insects and to healthy peach trees were conducted. Additionally, isolated phytoplasmas were genetically characterized to determine which isolates were able to infect peach trees. Results showed that the only insect species captured inside peach plots that was a carrier of the ‘Ca. Phytoplasma pyri’ phytoplasma was Cacopsylla pyri. Other insect species captured and known to be phytoplasma transmitters were present in very low numbers, and were not infected with ‘Ca. Phytoplasma pyri’ phytoplasma. A total of 1928 individuals of C. pyri were captured in the peach orchards, of which around 49% were phytoplasma carriers. All the peach trees exposed to C. pyri in 2014, and 65% in 2015, were infected by ‘Ca. Phytoplasma pyri’ 1 year after exposure, showing that this species is able to transmit the phytoplasma to peach. Molecular characterization showed that some genotypes are preferentially determined in peach. 相似文献
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A. Samad S. Panda Mahesh K. Gupta P. V. Ajayakumar Ashutosh K. Shukla 《European journal of plant pathology / European Foundation for Plant Pathology》2011,130(1):1-4
A new disease of Trachyspermum ammi, commonly known as Ajwain in India, was observed in Lucknow, India. The symptoms included small chlorotic leaves, highly
proliferating shoots, witches’ broom appearance, shortened internodes and an overall stunted growth. Poor flower heads and
fruit setting caused considerable yield losses for farmers. In transmission electron microscopy, pleomorphic bodies were detected
in the phloem cells of diseased plants but not in those of healthy plants. The disease etiology was investigated using direct
and nested polymerase chain reaction with phytoplasma-specific primers, DNA sequencing, and phylogenetic analysis. Phylogenetic
analysis indicated that this phytoplasma clustered in the 16SrVI group. A 1,249 bp sequence (FJ970035) of the 16S rRNA gene
from the phytoplasma showed 99% homology with the 16S rRNA gene, (FJ427295) of Ca. Phytoplasma trifolii strain A6 belonging to the phytoplasma group VI (reported from Iran). This is the first report of phytoplasma
infection affecting Ajwain (T. ammi). 相似文献
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通过透射电子显微镜,在从宁夏回族自治区固原市彭阳县红河镇采集的表现叶片上卷、红叶、气生薯症状的马铃薯样品叶脉韧皮部筛管细胞内观察到大量直径为500~700 nm的球形植原体粒子。以提取的感病和健康马铃薯叶片总DNA为模板,应用植原体16S rRNA基因和rp基因通用引物进行PCR扩增,从感病样品中扩增得到了长度均约为1.2 kb的片段。对获得基因核酸一致性比较分析表明,马铃薯僵顶植原体宁夏株系16S rRNA基因与‘Candidatus Phytoplasma fragariae’槭树株系(MK501642)16S rRNA基因核酸一致性最高,为99.7%,rp基因与‘Ca.P.fragariae’云南马铃薯YN-2G株系(KJ144889)rp基因核酸一致性最高,为100%;基于16S rRNA基因和rp基因构建系统进化树发现,马铃薯僵顶植原体宁夏株系与16SrⅫ-E亚组成员聚在一起。基于透射电镜观察和基因序列比较分析,证明宁夏发生的马铃薯僵顶病与植原体侵染相关,该植原体在分类地位上属于植原体16SrⅫ-E亚组。 相似文献
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小麦蓝矮病植原体16S rDNA基因片段的比较分析 总被引:21,自引:3,他引:18
小麦蓝矮病是陕西乃至西北冬麦麦区的一个重要病害,由介体条沙叶蝉专化性传播。对小麦蓝矮病株叶片和带毒条沙叶蝉进行超薄切片及电镜观察,在叶片韧皮部和叶蝉后肠中均观察到大量典型植原体。利用植原体16S rDNA基因保守序列通用引物对Rm16F2/Rm16R1,应用PCR技术从小麦蓝矮病株叶片中扩增到1.4 kb的特异片段。通过对16S rDNA基因片段序列同源性比较,结果表明小麦蓝矮病病原与三叶草绿变、翠菊黄化、绣球花绿变、草莓矮化和番茄巨芽植原体亲缘关系较近,其同源率为99.2%~99.9%。据此可以判定小麦蓝矮病植原体是属于植原体16SrⅠ组,确定了其分类地位。 相似文献
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Minoru Tanaka Koji Tanina Shinji Kasuyama Tomio Usugi 《Journal of General Plant Pathology》2007,73(4):286-289
In October 2001, a disease of rocket larkspur (Cosolida ambigua (L.) P. W. Ball et Heyw), characterized by witches’ broom, yellows and virescence of flowers, was found in Yakage Town in
Okayama Prefecture. Electron microscopy revealed the presence of phytoplasma-like bodies in the phloem of diseased plants.
The causal phytoplasma was identified as “Candidatus Phytoplasma asteris” based on 16S rDNA sequence analysis, and demonstrated to be acquired by the leafhopper Macrosteles striifrons.
The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under the accession number AB258330. 相似文献
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Hee-Young Jung Moo-Ung Chang Joon-Tak Lee Shigetou Namba 《Journal of General Plant Pathology》2006,72(4):261-263
Witches' broom disease in bamboo (Phyllostachys nigra Munro var. henonis) was found in Yeoungyang, Korea. In transmission electron micrographs, phytoplasma-like bodies were detected in the phloem
cells of diseased plants but not in those of healthy plants. The presence of phytoplasmas was confirmed by amplification of
a 1.8-kb DNA fragment using a primer pair specific for the region containing a 16S rRNA gene and an intergenic spacer region
between the 16S and 23S rRNA genes. Comparision of the 16S rRNA gene sequences showed that the causal phytoplasma belongs
to “Candidatus Phytoplasma asteris,” and shared the highest degree of similarity with the sequence of the onion yellows (OY) isolate in
Japan. This is the first phylogenetic identification of phytoplasma infection of bamboo in Korea. 相似文献