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1.
Natural infection of tospoviruses on three cucurbitaceous (Cucumis sativus, cucumber; Luffa acutangula, ridge gourd; Citrullus lanatus, watermelon) and three fabaceous (Vigna unguiculata, cowpea; Phaseolus vulgaris, French bean; Dolichos lablab, sem) vegetable crops in India was identified on the basis of nucleocapsid protein (NP) gene characteristics. The complete NP gene of the cowpea isolate from Kerala and the sem isolate from Tamil Nadu was 831 nucleotides long, encoding a protein of 276 amino acids. For other Tospovirus isolates from cucumber, French bean, ridge gourd and watermelon, the partial NP gene (291 nt) was sequenced. Comparative NP gene sequence analyses revealed that fabaceous isolates shared maximum identity both at the nucleotide (92–97%) and amino acid (93–97%) levels with the corresponding region of Groundnut bud necrosis virus (GBNV), whereas cucurbitaceous isolates shared maximum identity both at nucleotide (93–99%) and amino acid (95–98%) levels with the corresponding region of Watermelon bud necrosis virus (WBNV), results suggesting that the Tospovirus isolates infecting fabaceous hosts should be regarded as strain of GBNV, whereas those infecting cucurbitaceous hosts as a strain of WBNV. Nucleocapsid protein gene was conserved both in GBNV and WBNV isolates originating from different hosts and locations. 相似文献
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A S REDDY L J REDDY NALINI MALLIKARJUNA M D ABDURAHMAN Y V REDDY P J BRAMEL D V R REDDY 《The Annals of applied biology》2000,137(2):135-139
Eighty three wild Arachis germplasm accessions, belonging to 24 species of five sections and one natural hybrid derivative of a cross between the cultivated and a wild Arachis species, were evaluated along with a susceptible groundnut cultivar for resistance to Peanut bud necrosis virus (PBNV) in a replicated field trial at ICRISAT, Patancheru, India. Thirty days after sowing, the percentage of infected plants were recorded for all the accessions and subsequently young leaflets from all these accessions were tested for the presence of the virus by enzyme linked immunosorbent assay (ELISA). One accession each of A. benensis and A. cardenasii, and two accessions of A. villosa, in the section Arachis, two accessions of A. appressipila in the section Procumbentes, and one accession of A. triseminata under section Triseminatae were not infected by PBNV. These seven field‐resistant accessions were tested under glasshouse conditions for virus resistance by mechanical sap inoculations. One accession of A. cardenasii and two accessions of A. villosa did not show systemic infection. Similarly, in another glasshouse test, where 13 A. cardenasii accessions of section Arachis were evaluated, two accessions did not show systemic infection. In all these resistant accessions, the inoculated leaves showed infection, but the systemic leaves did not show the presence of virus in spite of repeated mechanical sap inoculations. So, the resistance in these accessions appears to be due to a block in systemic movement of the virus. To our knowledge this is the first report on the identification of resistance to PBNV in wild Arachis species. Since both A. cardenasii and A. villosa are the progenitors of cultivated groundnut and can be hybridised with the latter, the resistant accessions are being utilised in conventional breeding programmes to transfer PBNV resistance to widely adapted groundnut cultivars. 相似文献
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Virus‐like chlorotic symptoms were observed on tomato plants, cv. Velocity, grown in a greenhouse, region of Plovdiv. Samples collected from the leaves with interveinal yellowing and with initial interveinal chlorosis were tested for virus presence. Only the samples collected from the upper leaves with slight interveinal chlorosis were positive for Tomato infectious chlorosis virus (TICV) in indirect ELISA. Further, RT‐PCR analysis with specific primers for Tomato chlorosis virus (ToCV) heat shock protein 70, for TICV heat shock protein 70 and for TICV minor capsid protein was positive for TICV in all tested samples. No signals were obtained with primers for ToCV. Phylogenetic analysis showed that the Bulgarian sequence of Hsp70 and a sequence of Greek isolate clustered together having the highest resampling score. Regarding CPm, the Bulgarian isolate was more relevant to the French isolate. The obtained results from phylogenetic analysis supported the idea of a close relationship between the Bulgarian and Greek isolates. 相似文献
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The complete genome of a Potato virus X (PVX) isolate from India (ptDel‐9), which occurred symptomlessly in potato but induced ringspots on Nicotiana tabacum cv. Xanthi and necrotic mosaic on Nicotiana benthamiana, was sequenced. The genome was 6435 nucleotides long ( JF430080 ) and contained five open reading frames. The isolate was closely related to those reported from the Eurasian region (95.1–97.1% sequence similarity) and distantly related to those reported from South America (77.2–77.9%). The CP gene was expressed in Escherichia coli as a 76‐kDa fusion protein with maltose‐binding protein and used to generate polyclonal antibodies, which successfully detected PVX in field samples of potato by ELISA. In 20% of field samples, for which ELISA failed, the virus was successfully detected by RT‐PCR. This is the first report of molecular characterization of PVX occurring in India. 相似文献
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Yating Liu Xiuping Lu Long Zhi Yuanxian Zheng Xuejiao Chen Yue Xu Fajing Wu Yongzhong Li 《Journal of Phytopathology》2012,160(4):201-205
Leaf samples were collected from four spider lily and tobacco plants with tospovirus‐like symptoms in Yunnan province. The nucleocapsid ORFs of the four isolates were obtained by RT‐PCR, cloning and sequencing of the amplified products. Nucleotide sequences of the obtained amplicons were more than 90% identical to that of Calla lily chlorotic spot virus (CCSV) isolates deposited in the GenBank database. The virus induced systematic leaf necrosis, chlorotic lesions, rugosity and deformation on mechanically inoculated experimental hosts. This is the first report of CCSV in mainland China, and also on spider lily and tobacco. 相似文献
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West Nile (WN) virus is a mosquito-borne flavivirus that induces lethal encephalitis in humans and horses. Since an outbreak of WN encephalitis in humans and horses occurred in New York City in late August 1999, the possibility exists that WN virus will invade regions that have close links with the United States, such as Japan. We developed a genetic diagnostic method that discriminates between strains of WN virus and Japanese encephalitis (JE) virus. The method involves RT-PCR restriction fragment length polymorphism (RFLP) analysis with a RT-PCR primer set, a nested PCR primer set, and a restriction enzyme. We detected WN and JE viruses in experimentally infected animal brain, spleen, and serum samples. Our method is useful in distinguishing WN viruses from the endemic background of JE viruses, and in discriminating the highly virulent WN strain, which was isolated in New York in 1999, from other WN virus strains. 相似文献
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Kurokawa CS Araujo JP Soares AM Sugizaki MF Peraçoli MT 《Microbiology and immunology》2007,51(4):421-428
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Mingqiang Wang Fan Li Guohui Zhou Pingxiu Lan Donglin Xu Ruhui Li 《Journal of Phytopathology》2015,163(11-12):1036-1040
An improved RT‐PCR was developed and validated for the detection of Yam mild mosaic virus (YMMV). Sequences of the coat protein core region of 19 Chinese isolates were obtained, and analysis indicated the presence of different genetic variants. Phylogenetic analyses showed that the Chinese isolates were divided into two distinct clusters. Complete genomic sequences of two distinct Chinese variants were determined to be 9527 and 9529 nucleotides long, excluding the 3′ poly (A) tail. Their genomic structure and organization were virtually identical to that of a Brazilian isolate. The two variants shared identity of 87.3% to one another and 83.9–84.6% to the Brazilian variant at the genomic sequence level. Phylogenetic analyses supported that they represented two distinct YMMV lineages. 相似文献
9.
香茄环斑病毒HC—RT—PCR—ELISA检测 总被引：3，自引：0，他引：3
番茄环斑病毒（ToRSV)是我国对外检疫一类有害生物。目前国内尚无存在的报道。PCR技术是一种快速灵敏的植物病毒检测方法,但核酸内的聚合酶抑制物会导致漏检现象。而只通过凝胶电泳进行结果判定会出现假阳性,这两方面限制了PCR技术在对外检疫中的应用,利用共价结合在PCR管壁上的引物特异性杂交诱捕核酸粗提液中的靶标核酸,洗掉杂质及抑制物质,在同一管内作RT-PCR,凝胶电泳检测液相产物的同时对固相产物进行杂交检测。提高了结果的可靠性及灵敏度。利用所建立的HC-RT-PCR-ELISA成功地从法国进口葡萄苗中检出ToRSV。本方法可用于其他植物病毒及转基因产品的检测。 相似文献
10.
Uda A Tanabayashi K Mukai R Yachi M Nam KH Yamada A 《Journal of medical primatology》2001,30(3):141-147
Cynomolgus monkeys were divided into two groups in terms of the reactivity of their lymphocytes with the FN18 monoclonal antibody, which is directed to the CD3 of rhesus monkeys. It was shown that 24 (12.2%) out of 196 monkeys did not have lymphocytes that reacted with the FN18, although T cells from those animals responded well to mitogenic stimulation. We have determined the nucleotide sequences of the CD3delta, CD3gamma, and CD3epsilon chains and found that two amino acids of the CD3epsilon chain of the FN18 non-reactive monkeys were different when compared with the FN18 reactive monkeys. Our results indicated that the CD3epsilon molecule of cynomolgus monkeys is polymorphic at the epitope level, which is recognized by the FN18 monoclonal antibody. 相似文献
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Honma S Nakata S Kinoshita-Numata K Kogawa K Chiba S 《Microbiology and immunology》2000,44(5):411-419
Norwalk virus and Sapporo virus were approved as type species of the genus \"Norwalk-like viruses\" and the genus \"Sapporo-like viruses,\" respectively, in the family Caliciviridae. A total of 116 stool specimens containing Norwalk virus (NV) or Sapporo virus (SV) were tested by RT-PCR and Southern hybridization to evaluate nine sets of PCR primers and seven internal oligonucleotide probes in the RNA dependent RNA polymerase region of NV and SV for detection and differentiation of viruses in the NV and SV. Fifty-five stool samples were collected from 11 outbreaks of NV and/or SV gastroenteritis in an infant home, where residents were infants under 2 years of age, in Sapporo, Japan. Sixty specimens were obtained in Sapporo from sporadic cases in children, mainly under 6 years of age, of acute gastroenteritis due to small round structured viruses detected by EM. There is no single primer pair to detect all NV and SV, and at least three primer pairs, G1 set, G2 set and Sapp35/Sapp36, are required to detect viruses in the NV and SV clades. Many NV and SV strains were successfully classified into one of the NV/genogroup I, NV/genogroup II and SV by single-round RT-PCR and Southern hybridization. The new detection method for SV reported in this study combined with those for NV previously reported may elucidate the importance of Norwalk virus and Sapporo virus as a cause of viral gastroenteritis in all age groups in the world. 相似文献
13.
甲病毒指披膜病毒科(Togavirdae)甲病毒属(Alphavirus),以蚊虫等吸血昆虫为传播媒介,可引起如辛德毕斯热、基孔肯雅热、东方马脑炎、西方马脑炎等多种人畜共患性传染病[1,2],是医学上重要的虫媒病毒.甲病毒世界性分布,全世界已发现28种甲病毒,其中13种与人畜疾病有关,至少7种甲病毒发生过不同程度流行,造成巨大的经济损失,成为严重的公共卫生问题.张海林等(中国媒介生物学及控制杂志,1992,3(特7):419－421)从云南傣族一位发热病人血中分离到一株病毒,命名为YN87448病毒.经血清学鉴定该病毒符合披膜病毒科甲病毒属病毒特征.由于缺乏标准诊断血清,无法对它作进一步鉴定.该病毒直接分离自发热病人血清,病人主要症状为发热,寒颤,腰疼痛和全身酸痛.由于该病毒直接分离自病人血清,病毒鉴定对解释该病人的发病原因,甚至对于解释我国部分地区流行的无名热的病因均具有重要意义. 相似文献
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Yuki Matsushima Tomomi Shimizu Ikuko Doi Fuminori Mizukoshi Koo Nagasawa Akihide Ryo Hideaki Shimizu Masae Kobayashi Keiji Funatogawa Noriko Nagata Mariko Ishikawa Ayako Komane Nobuhiko Okabe Yoshio Mori Makoto Takeda Hirokazu Kimura 《Microbiology and immunology》2017,61(8):337-344
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将水泡性口炎病毒编码群特异性抗原的N基因片段克隆至pMD18-T克隆载体质粒中,构建N基因克隆重组质粒,进行核苷酸序列分析。然后亚克隆插入pBAD/Thio TOPO表达载体,经PCR限制性内切酶分析、测序鉴定,筛选获得N基因正向插入、有正确读码框的阳性克隆,成功构建了水泡性口炎病毒N基因重组表达载体。经L-Arabinose诱导表达,可稳定、高效地表达N蛋白抗原。SDS-PAGE、Western blotting及间接ELISA试验结果表明,表达蛋白为融合蛋白,质量约63.5 kD,其表达产量约占菌体总蛋白的16％,相当于92mg/L。融合蛋白中含有水泡性口炎病毒群特异性的核蛋白抗原,应用表达的VSV核蛋白抗原建立了酶联免疫吸附试验,通过对186份山羊、豚鼠实验动物人工感染VSV的血清样品和参考血清样品的检测,并与微量血清中和试验进行了比较,结果表明：以表达的VSV核蛋白为包被抗原的酶联免疫吸附试验是一种特异性强、敏感性高、快速、简单、安全的检测方法,抗原制备成本低。 相似文献
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K Hara T Kashiwagi Y Ohtsu K Masunaga Y Akasu-Tsuji N Tsumura H Kato J Iwahashi N Hamada M Toyoda T Toyoda 《Microbiology and immunology》2001,45(10):717-720
An epidemic of aseptic meningitis caused by human echovirus 9 (E-9) occurred in the summer of 1997 in northern Kyushu, Japan. Sequences of genome position 2504-3358, which encoded a part of VP1, of the nine isolated viruses were determined. An RGD motif and B-C loop were found in all. They were almost identical and closely related to the virulent strain Barty. 相似文献
17.
Borrelia duttonii strain Ly, a causative agent of relapsing fever, contains a linear one megabase chromosome and 12 linear plasmid molecules. Here we report that the sequence of the 44-kb linear plasmid of strain Ly is found to contain variable major protein (vmp) genes for antigenic variation of relapsing fever borreliae. The determined sequence is of 44,010 bp except for both ends of the molecule. Of 39 open reading frames (ORFs) found in the sequence, 21 ORFs (named vmpA to U) showed moderate similarities with vmp genes for Borrelia hermsii. However, most of the vmp homologues are apparently nonfunctional because of their frameshifts within the sequence and/or absence of promoter and ribosome-binding signals upstream of their genes. RT-PCR experiments using the specific primer for each vmp gene revealed that vmpE, one of the vmp genes, was expressed at the location of the 44-kb plasmid molecule. The result suggests that the plasmid molecule may play a role in the preservation of the serotype switching of vmp genes in a mammalian host. 相似文献
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[背景]牛冠状病毒(Bovine coronavirus,BCoV)是引起新生犊牛死亡的主要病原之一,有效的检测手段是防治该病的前提。目前BCoV ELISA检测方法存在敏感性低、不稳定等缺陷。[目的]对原有BCoV ELISA方法进行改进,建立间接ELISA检测方法。[方法]应用我国BCoV流行毒株CD株n基因为模板,预测N蛋白抗原表位,通过原核表达制备可溶性的重组N蛋白作为抗原,建立间接ELISA方法,应用该方法对黑龙江省2010-2017年的BCoV感染进行血清流行病学调查。[结果]该ELISA方法最佳工作条件为:用50 mmol/LpH 9.6碳酸盐作为包被液,抗原包被浓度2.5μg/mL;用PBST作为样本稀释液,稀释浓度1:50,37℃孵育1.5 h;HRP-羊抗牛IgG稀释浓度1:7 500,37℃孵育1.0 h;用1%明胶37℃封闭30 min。阴阳性临界值为0.225。该方法与BRV、BRSV、BVDV、IBRV、BPIV3和E.coli阳性血清均无交叉反应。批内和批间变异系数均小于10%,与病毒中和试验的符合率高达93.5%。对黑龙江省部分地区共603份奶牛血清样品检测结果显示,BCoV抗体阳性率为98.84%。[结论]建立的ELISA方法特异性强、敏感性高、稳定性好,为进一步研发ELISA试剂盒提供了技术基础。 相似文献
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Vahid Rahimkhoei Jafar Rezaie Ali Akbari Aynaz Nourani Nassrollah Jabbari Zohreh Mehri Lighvan Mojtaba Amini 《Cell biochemistry and function》2021,39(1):29-34
Today, tremendous attention has been devoted to a new coronavirus, SARS‐CoV‐2 (2019‐nCoV), due to severe effects on the global public in all over the world. Rapid and accurate diagnosis of 2019‐nCoV are important for early treatment and cutting off epidemic transmission. In this regard, laboratory detection protocols, such as polymerase chain reaction (PCR) and computed tomography (CT) examination, have been utilized broadly for 2019‐nCoV detection. Recently, nano‐based methods for 2019‐nCoV diagnoses are rapidly expanding and declaring comparable results with PCR and CT. In this review, recent advances in nano‐based techniques have been highlighted and compared briefly with PCR and CT as well‐known methods for 2019‐nCoV detection. 相似文献