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1.
Wheat blue dwarf(WBD) is a disease caused by phytoplasma and only reported from China. A fragment about 1.3 kb in protein translocation gene, secY was amplified by PCR from the total DNA of di-seased wheat sample with primer pair secYF/secYR, which was designed based on secY gene sequence of known 16SrI group members. Nucleotide acid sequence analysis of amplified fragment indicated that the length was 1 240 bp. A phylogenetic tree based on secY gene sequences was constructed and showed that wheat blue dwarf phytoplasma was clustered into the Candidatus Phytoplasma asteris, subgroup 16SrI-C. Wheat blue dwarf phytoplasma showed high homology with clover phyllody phytoplasma strains based on sequence comparison and phylogenetic analysis. 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
3.
Yusuke Takinami Kensaku Maejima Atsushi Takahashi Takuya Keima Takuya Shiraishi Yukari Okano Ken Komatsu Kenro Oshima Shigetou Namba 《Journal of General Plant Pathology》2013,79(3):209-213
Phytoplasma-induced floral malformations such as virescence, phyllody, and proliferation were observed on hydrangeas in Gunma Prefecture, Japan. Phylogenetic analyses based on 16S rRNA, secY, groEL, and amp gene sequences indicated that the affected hydrangea plants were associated with phytoplasmas belonging to ‘Candidatus Phytoplasma asteris’, but not to ‘Ca. P. japonicum’, which occurs in hydrangeas showing phyllody in Japan. This is the first molecular evidence of an association of ‘Ca. P. asteris’ with hydrangea plants in Japan. 相似文献
4.
紫花苜蓿丛枝病植原体的分子检测及鉴定 总被引：1，自引：0，他引：1
利用植原体16S rRNA基因通用引物对云南昆明发生的苜蓿丛枝病感病植株总DNA进行巢式PCR扩增,得到1.2kb的特异片段,从分子水平证实了苜蓿丛枝病的病原是植原体。从PCR产物的RFLP酶切图谱可看出,该植原体株系的酶切图谱与马里兰翠菊黄化植原体(AY1)相同。对扩增片段进行克隆及序列测定后,利用最小进化法做Bootstrap验证的系统进化树,表明苜蓿丛枝病植原体为Candidatus Phytoplasma asteris成员之一,与植原体16SrI-B亚组成员关系密切。 相似文献
5.
Test performance study of isothermal amplification tests for the detection of Grapevine flavescence dorée phytoplasma and ‘Candidatus Phytoplasma solani’ 
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下载免费PDF全文 N. Mehle P. Kogovšek F. Constable K. De Jonghe M. Loiseau F. Veratti C. Marzachi L. Ferretti E. Angelini L. Filippin E. Sousa E. Andrade J. Hodgetts M. Dickinson M. Dermastia 《EPPO Bulletin》2017,47(1):18-23
This test performance study (TPS) was carried out on DNA samples from grapevine, clematis, fungi and bacteria to compare and validate loop‐mediated isothermal amplification (LAMP) tests for detection of Grapevine flavescence dorée phytoplasma and ‘Candidatus Phytoplasma solani’ (Grapevine Bois noir phytoplasma). Two LAMP tests, for Grapevine flavescence dorée phytoplasma and ‘Candidatus Phytoplasma solani’ (as developed by Kogov?ek and colleagues), with proven applicability for rapid laboratory or on‐site detection were included in this study. They were performed in 10 laboratories. In addition, the commercial Qualiplante/Hyris isothermal amplification test for Grapevine flavescence dorée phytoplasma was performed in three laboratories. The accuracy of the three tests was shown to be over 98%. Moreover, the high accuracy of these tests, which used different devices across different laboratories, confirmed their reproducibility. 相似文献
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K. De Jonghe I. De Roo M. Maes 《European journal of plant pathology / European Foundation for Plant Pathology》2017,147(4):749-759
Over the years, real-time PCR outflanked endpoint PCR in phytopathogen diagnostics, mainly because of the increase in sensitivity and timesaving aspects of the technique. However, a time consuming 16S rRNA-based nested PCR method is still the gold standard for phytoplasma diagnosis. This is also the case for phytoplasma detection in Malus, Pyrus and Prunus, the three main host plants of apple proliferation (AP), pear decline (PD) and European stone fruit yellows (ESFY) phytoplasma, respectively. The last decade, loop-mediated isothermal amplification (LAMP) (Notomi et al. 2000) is gaining a lot in significance and is also for phytoplasmas expected to become a widely used reliable diagnostic tool. High specificity and sensitivity which also requires a less stringent need for DNA purification, and the short analysis time and the limited equipment requirements makes the LAMP method a fast and affordable alternative with great point-of-care diagnostic potential. In this paper, we present a LAMP primer set for the ribosomal group 16SrX, containing the important fruit tree phytoplasmas AP, PD and ESFY. The primers were developed and validated for fast and sensitive detection and general use for diagnosis. We foresee that the LAMP technique will also have its application in on-site diagnosis of the fruit tree phytoplasmas during inspections and surveys. 相似文献
8.
Shiro Fukuta Reiko Takahashi Satoru Kuroyanagi Noriyuki Miyake Hirofumi Nagai Hirofumi Suzuki Fujio Hashizume Tomoko Tsuji Hiromi Taguchi Hideki Watanabe Koji Kageyama 《European journal of plant pathology / European Foundation for Plant Pathology》2013,136(4):689-701
A loop-mediated isothermal amplification (LAMP) reaction with a primer set designed from the rDNA ITS sequence of P. aphanidermatum was developed. Results of a specificity test using 57 strains of Pythium spp. indicated that the LAMP assay gave no cross reactions in other 39 Pythium species, 11 strains of Phytophthora spp. and eight other soil borne pathogens. The detection limit was 10 fg of genomic DNA, which was ten times the sensitivity of the polymerase chain reaction. The LAMP assay was applied to hydroponic solution samples from tomato fields, and the results were compared to those of the conventional plating method. LAMP was observed to be effective for the specific detection of P. aphanidermatum. Furthermore, P. aphanidermatum was detected directly in tomato roots infected with P. aphanidermatum without DNA extraction. The LAMP method established in this study is a simple, sensitive and rapid tool for the detection of P. aphanidermatum. 相似文献
9.
利用植原体16S rRNA基因及核糖体蛋白基因(ribosomal protein, rp)通用引物对发生在云南元谋的花生丛枝病病株DNA进行PCR扩增,并对扩增片段进行序列测定。扩增获得的云南元谋花生丛枝植原体(PnWB-YNym)16S rDNA、16S-23S rDNA和23S DNA片段总长1 806 bp,rp基因扩增片段长1 171 bp。云南株系与来源于台湾和海南的花生丛枝植原体均有较高同源性。比较16S rDNA片段,发现云南株系在5个位点上与来自台湾或海南的株系存在碱基差异,其中有1个位点的差异是云南元谋株系特异的;再分别比较核糖体蛋白rplV-rpsC 2个基因所编码的氨基酸序列,发现云南株系rpsC编码的第194位氨基酸与台湾和海南的株系存在差异。经16S rDNA片段系统进化及iPhyClassifier在线分析,表明PnWB-YNym在分类上属于16SrII-A亚组成员,与候选种‘Candidatus Phytoplasma australasiae’相关;基于rp基因构建的系统进化树表明,PnWB-YNym与16SrII-A亚组各成员聚为同一亚进化支(iii)。 相似文献
10.
Polona Kogovšek Nataša Mehle Anja Pugelj Tjaša Jakomin Hans-Josef Schroers Maja Ravnikar Marina Dermastia 《European journal of plant pathology / European Foundation for Plant Pathology》2017,148(1):75-84
A fluorescence-based real-time loop-mediated isothermal amplification (LAMP) assay for ‘Candidatus Phytoplasama solani’ (Bois noir phytoplasma; BNp) detection was developed and optimised for rapid laboratory and on-site BNp detection. This assay is highly specific, rapid and as sensitive as qPCR. It was validated according to European and Mediterranean Plant Protection Organisation recommendations. In addition, 286 grapevine leaf samples from the 2015 growing season were tested with this new real-time LAMP assay and an assay previously developed for detection of Flavescence dorée phytoplasma (FDp). These LAMP assays for detection of both BNp and FDp used without any DNA extraction step, which is a required step for qPCR analysis, were comparably effective to qPCR, and positive results were obtained in less than 35 min. 相似文献
11.
Chinmay Biswas Piyali Dey Subrata Satpathy Manoj Kumar Pratik Satya B. S. Mahapatra 《Phytoparasitica》2013,41(5):539-543
Mesta (Hibiscus sabdariffa) is an important bast fiber crop. In August 2011, there was an outbreak of a phytoplasma-like disease on H. sabdariffa in different villages of the northern coastal mesta-growing region of Andhra Pradesh, India, covering mainly two districts – Srikakulam and Vijayanagaram. The infected plants showed characteristic symptoms such as phyllody and reddening of leaves. PCR with P1/P7 universal primer pair of 16 S rDNA yielded amplicons of 1850 bp from all symptomatic mesta leaf samples similar to samples of brinjal little leaf (phytoplasma positive reference control). However, asymptomatic samples were not amplified. Multiplex nested-PCR showed simultaneous amplification of DNA fragments with phytoplasma specific primers, viz., P1/P7 universal primer pair of 16 S rDNA, nested primer pair R16F2n/R2, uvrB and DegV gene-specific uvrB-degVF/R primer generating amplicons of 1850 bp, 1200 bp and 1023bp, respectively. However, SecY-map gene specific primer SecY-mapF/R was not amplified. The 1023 bp nucleotide sequence of uvrB and DegV gene of the phytoplasma was deposited in the GenBank (NCBI) with the accession no. JX975061. NCBI BLASTn analysis of the 1023 bp products showed that the phytoplasma strain belonged to elm yellows group (16SrV-D). This is the first report that Hibiscus sabdariffa is infected by a phytoplasma and we named it mesta phyllody disease (MPD). 相似文献
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A rapid DNA extraction and loop‐mediated isothermal amplification (LAMP) procedure was developed and evaluated for the detection of two specific groups of phytoplasmas from infected plant material. Primers based upon the 16–23S intergenic spacer (IGS) region were evaluated in LAMP assays for amplification of group 16SrI (aster yellows group) and group 16SrXXII (Cape St Paul wilt group) phytoplasma strains. DNA could be extracted from leaf material (16SrI phytoplasmas) or coconut trunk borings (16SrXXII phytoplasmas) onto the membranes of lateral flow devices, and small sections of these membranes were then added directly into the LAMP reaction mixture and incubated for 45 min at 65°C. Positive reactions were detected through the hydroxyl napthol blue colorimetric assay within 1 h of the start of DNA extraction, and were confirmed by subsequent agarose gel electrophoresis of the LAMP products. The level of detection was comparable to that obtained by nested PCR using conventional 16S rDNA phytoplasma‐specific primers. Furthermore, the assays were specific for the phytoplasmas they were designed to detect – the 16SrI assay only detected 16SrI phytoplasmas and not those from any other phylogenetic groups, whilst the 16SrXXII assay only detected 16SrXXII phytoplasmas. The DNA extractions and LAMP assay are easy to perform, requiring minimal equipment, and may therefore form the basis of a rapid and reliable field‐detection system for phytoplasmas. 相似文献
14.
Phytoplasma-induced leaf yellowing, stunting and rosetting were observed on Japanese spurge (Pachysandra terminalis) in Jung-gu of Daegu city, Korea. Molecular analyses based on PCR–RFLP and 16S rDNA sequences indicated that the phytoplasma
associated with Japanese spurge yellows disease is a new strain of Candidatus Phytoplasma asteris. This report is the first on the occurrence of phytoplasma disease on Japanese spurge in Korea. 相似文献
15.
Fusarium mangiferae is a major causal agent of mango malformation disease (MMD) worldwide. Rapid and accurate detection of the causal pathogen is the cornerstone of integrated disease management. In this paper, a real-time fluorescence loop-mediated isothermal amplification assay (RealAmp) was developed for quantitative detection of F. mangiferae in China. The LAMP primer set was designed based on a RAPD marker sequence and positive products were amplified only from F. mangiferae isolates, but not from any other species tested, showing a high specificity of the primer sets. The detection limit was approximately 2.26 × 10−4 ng/μl plasmid DNA when mixed with extracted mango DNA. Quantification of the pathogen DNA of MMD in naturally collected samples was no significant difference compared to classic real-time PCR Additionally, RealAmp assay was visual with an improved closed-tube visual detection system making the assay more convenient in diagnostics. 相似文献
16.
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
17.
Zhan-Cui Yu Yang Cao Qun Zhang De-Fa Deng Zhen-Yu Liu 《Journal of General Plant Pathology》2012,78(4):298-300
Chinese scholar tree (Sophora japonica) with witches’ broom symptoms was observed in Shandong Province in China. Phytoplasmas were detected in the diseased plants using 16S rDNA amplification with phytoplasma-specific universal primer pairs. On the basis of the results of 16S rDNA sequencing, virtual restriction fragment length polymorphism patterns and phylogenetic analyses, the phytoplasma found in S. japonica with witches’ broom symptoms was confirmed as a ‘Candidatus Phytoplasma ziziphi’-related strain belonging to the Elm yellows group 16SrV. This is the first report of ‘Ca. P. ziziphi’ infecting S. japonica plant with witches’ broom symptoms. 相似文献
18.
Gulnara Balakishiyeva Madat Qurbanov Alamdar Mammadov Shaniyar Bayramov Jalal Aliyev Xavier Foissac 《European journal of plant pathology / European Foundation for Plant Pathology》2011,130(4):457-462
‘Candidatus Phytoplasma brasiliense’, a phytoplasma taxon associated with hibiscus witches’ broom disease was first described in 2001
in Brazil. In September 2007, a peach tree (Prunus persica) displaying yellowing symptoms reminiscent of phytoplasma infection was sampled in Guba region of Azerbaijan. A phytoplasma
was detected in the diseased peach tree by nested PCR amplification of its 16S rDNA with universal primers for phytoplasmas.
Phylogenetical analyses of the amplified 16S rDNA showed that the phytoplasma infecting the peach tree corresponded to ‘Ca. P. brasiliense’, a species never reported in Euro-Mediterranean area. To set up a detection assay, cloning of a ‘Ca. P. brasiliense’ DNA fragment was undertaken by comparative RAPD. The amplified dnaK-dnaJ genetic locus was used to design a nested PCR assay able to amplify all ‘Ca. P. brasiliense’ isolates of the subgroup 16SrXV-A without amplifying the related members of the group 16SrII. This assay
also allowed confirming the first detection of ‘Ca. P. brasiliense’ in diseased basil collected in south Lebanon. 相似文献
19.
Salvadora persica trees with symptoms of witches’-broom, little leaf and severe leaf curling were observed in the Biodiversity Park, North Delhi (India) during February 2010. Using a nested-PCR assay targeting two phytoplasma specific loci (16S ribosomal RNA and the ribosomal protein gene) and sequence analysis of the amplified products, we identified the associated phytopathogen as a phytoplasma of group 16SrI. RFLP-mediated characterization revealed that the phytoplasma belongs to subgroup 16SrI-B. Phylogenetic reconstruction also grouped the phytoplasma within group 16SrI. The study reports a new host of phytoplasma in India and worldwide. 相似文献
20.
香蕉条斑病毒LAMP快速检测方法的建立 总被引：1，自引：0，他引：1
环介导等温扩增（loop-mediated isothermal amplification,LAMP）是一种特异、灵敏、快速的新型基因检测技术。本研究以香蕉条斑病毒（Banana streak virus,BSV）ORF3保守区域为基础针对6个特定区域设计并筛选了4条LAMP扩增引物,通过对LAMP反应中MgSO4、dNTPs、Betaine等主要试剂浓度进行优化,建立了香蕉BSV的LAMP检测方法,63℃反应90 min后通过在反应产物中添加SYBR Green Ⅰ染料后颜色的变化,肉眼即可判断检测结果。LAMP具有极高的检测特异性和灵敏性,其检测下限约为3.2 ng·μL-1,是PCR检测灵敏度的25倍,能快速、准确地对疑似样品进行检测,本研究对华南地区部分疑似样品的检测结果显示LAMP阳性检出率比PCR检出率高。本文建立的BSV LAMP检测方法是对BSV检测方法的拓展和延伸,为香蕉病毒的快速检测提供技术保障。 相似文献