共查询到20条相似文献，搜索用时 62 毫秒
1.
以临床疑似犬瘟热病犬外周血总RNA为模板,采用RT-PCR方法扩增犬瘟热病毒(CDV)血凝素(H)基因。序列测定和分析表明该病料犬瘟热病毒H基因序列与国内其他地区分离到的毒株同源性较高(94.6%~99.2%),而与Onderstepoort株、Convac株等疫苗毒株的同源性较低(90.4%~91.2%)。分子遗传进化分析显示所有毒株可分为7个大的分支,且各分支间具有一定的地域性,其中待检病料中CDV与大多数中国分离株一样处于Ⅰ型。进一步以H全基因为模板,截短表达其3′端816 bp、459 bp两个片段,并克隆入原核表达载体pET30 a进行表达。结果显示它们分别能表达大小约为36.4 ku和22.2 ku的融合蛋白。免疫转印表明该纯化蛋白均可与犬瘟热病毒抗血清发生阳性反应,说明重组蛋白具备抗原性,可用于进一步的血清学研究。 相似文献
2.
为了解1株圈养小熊猫源犬瘟热病毒（CDV）GD-1的遗传变异情况,通过RT-PCR方法对该株CDV进行H、F基因的克隆、测序及序列分析。结果显示：该分离株的H基因序列与GenBank中丹麦报道的登录号为GU266280的犬源CDV毒株的核苷酸序列相似性最高,为96%;F基因序列与巴西报道的登录号为KY057355的犬源CDV的核苷酸序列相似性最高,为95.7%。下载CDV代表毒株序列进行遗传演化、氨基酸序列比对及分子特征分析。结果显示：H蛋白共有8个潜在的N-糖基化位点,分别位于19、149、309、391、422、456、587、603位点;H蛋白的SLAM受体结合位点氨基酸序列与欧亚野生型毒株一致,与疫苗株相比,530、549位氨基酸不同,与其他CDV参考毒株H蛋白相比还存在24、41等9处氨基酸位点发生明显变异,与标准强毒株A75/17的氨基酸相似性为95.2%,与Onderstepoort、Convac等5株疫苗株的氨基酸序列相似性为88.2%~89.3%;F蛋白共有6个N-糖基化位点,分别位于62、108、141、173、179、517位,与Onderstepoort等疫苗株氨基酸相似性为89.1%~89.7%;与其他参考毒株相比还存在115、130等11处氨基酸发生变异;构建基于H、F基因的遗传进化树,结果显示：该毒株位于Asia-4型的一个小的进化分支,这与目前我国流行毒株主要位于Asia-1型存在明显不同。本研究首次报道了小熊猫源的Asia-4基因型CDV野毒株,并对毒株的H及F基因进行了序列分析,对于了解我国CDV流行株的遗传变异情况、流行病学调查、疾病防控及疫苗研发等具有重要意义。 相似文献
3.
为了解上海地区犬瘟热病毒（Canine distemper virus，CDV）遗传变异情况，本研究采用首尾重叠的11对特异性引物，对CDV上海株SH202003进行RT-PCR扩增，将扩增片段进行反复测序，序列拼接后最终获得了SH202003株全基因组序列，应用Lasergene 7.0和Mega 6.0软件对全基因及H基因进行序列分析，并构建系统进化树。结果显示，SH202003株基因组全长为15 690 bp，编码6种结构蛋白（N、P、M、F、H和L），H和L基因间隔序列为CUA，L和5'端尾随序列为CAA，与Hebei株核苷酸和氨基酸相似性最高，达到98.6%和96.6%，与疫苗株核苷酸相似性在92.2%~94.3%，氨基酸相似性只有82.7%~87.0%；全基因进化树中，SH202003株与流行野毒株在同一分支，与疫苗株在不同的分支；H基因同样与Hebei株亲缘关系最近，核苷酸和氨基酸相似性分别为98.7%和99.5%，与疫苗株Snyder Hill、CDV3、Convac及Onderstepoort亲缘关系较远；SH202003株处于Asia-1型分支，属于Asia-1型强毒株；SH202003株具有9个潜在N-糖基化位点，与强毒株Hebei株一致。研究表明，上海株SH202003属于CDV强毒株，为Asia-1型，其H基因序列相对保守，具有9个潜在N-糖基化位点，但是全基因序列存在较多突变，与疫苗株的匹配度较差，可能是免疫犬依然发生犬瘟热的主要原因。 相似文献
4.
犬瘟热是犬瘟热病毒(Canine distemper virus,CDV)感染犬和其他食肉动物造成的多发性、致死性传染病,本文从分子水平上探讨CDV遗传进化特性、变异情况与流行规律之间的关系.通过收集2002-2010年在中国地区分离的14株CDV野毒株、2006-2007年在全球各地分离的12株CDV野毒株以及从不同宿主分离的12株CDV野毒株和4株疫苗株,将其分为4组,将前3组野毒株分别与国内外正在使用的4株疫苗株的H基因进行遗传变异分析.分析发现,CDV野毒株与疫苗株间H蛋白基因的核苷酸相似性为86.2％～92.1％,其氨基酸相似性为89.1％～91.9％;H基因的584位的天冬酰胺糖基化位点是Asia-Ⅰ型CDV所特有的;H蛋白3555区域的非同义氨基酸替换概率较高.作者推测H蛋白抗原变异可能造成弱毒疫苗免疫效力降低,不能为某些CDV株的感染提供完全有效的保护. 相似文献
5.
Hirama K Goto Y Uema M Endo Y Miura R Kai C 《The Journal of veterinary medical science / the Japanese Society of Veterinary Science》2004,66(12):1575-1578
Hemagglutinin (H) gene of two CDV isolates, the Haku93 and Haku00 strains, from masked palm civets was molecularly analyzed. H genes of both two CDVs contained one open reading frame encoding 607 amino acids. Nucleotide and predicted amino acid sequences of H gene of the CDV Haku93 and Haku00 revealed high similarity to those of recent field isolates such as the Yanaka and Tanu96, while they showed limited identity to those of old vaccine strains. Potential N-linked glycosylation sites in both Haku93 and Haku00 were identical to other recent CDV isolates. Phylogenetic analysis revealed that the CDV strains derived from masked palm civets were classified into the group of recent Japanese CDV isolates. 相似文献
6.
Nagao Y Nishio Y Shiomoda H Tamaru S Shimojima M Goto M Une Y Sato A Ikebe Y Maeda K 《The Journal of veterinary medical science / the Japanese Society of Veterinary Science》2012,74(6):699-705
Canine distemper virus (CDV), a morbillivirus that causes one of the most contagious and lethal viral diseases known in canids, has an expanding host range, including wild animals. Since December 2009, several dead or dying wild raccoon dogs (Nyctereutes procyonoides) were found in and around one safari-style zoo in Japan, and CDV was isolated from four of these animals. In the subsequent months (January to February 2010), 12 tigers (Panthera tigris) in the zoo developed respiratory and gastrointestinal diseases, and CDV RNA was detected in fecal samples of the examined tigers. In March 2010, one of the tigers developed a neurological disorder and died; CDV was isolated from the lung of this animal. Sequence analysis of the complete hemagglutinin (H) gene and the signal peptide region of the fusion (F) gene showed high homology among these isolates (99.8-100%), indicating that CDV might have been transmitted from raccoon dog to tiger. In addition, these isolates belonged to genotype Asia-1 and had lower homology (<90%) to the vaccine strain (Onderstepoort). Seropositivity of lions (Panthera leo) in the zoo and wild bears (Ursus thibetanus) captured around this area supported the theory that a CDV epidemic had occurred in many mammal species in and around the zoo. These results indicate a risk of CDV transmission among many animal species, including large felids and endangered species. 相似文献
7.
为建立一种鉴别犬瘟热病毒(CDV)野毒株与疫苗株的反转录-环介导等温扩增方法(RT-LAMP),本研究通过比对野毒株与疫苗株H基因设计特异性引物,对反应体系中的Mg2+、Betaine、Bst DNA Polymerase、dNTP和反应温度等条件分别进行优化,建立用于鉴别检测CDV野毒株与疫苗株的RT-LAMP。建立的RT-LAMP方法检测CDV野毒株时,在65℃水浴锅中反应40 min即可完成。该方法具有高度特异性,对犬细小病毒、犬腺病毒、狂犬病毒、犬冠状病毒无交叉反应,敏感度可达40 copies/μL,是常规RT-PCR方法的100倍。 相似文献
8.
根据Onderstepoort株H基因序列,设计1对引物建立RT-PCR-RFLP检测方法,对不同宿主来源的疑似犬瘟热临床样品进行检测,并对检出的犬瘟热病毒野毒山东株PCR产物进行克隆和序列分析,验证RT—PCR—RFLP检测方法。结果RT—PCR扩增片段为1921bp,产物经RFLP分析,野毒株的PCR产物能被NdeI酶切为1282、345和294bp3个片段,弱毒疫苗株则不能被切开;病毒RNA的最小检出量为2．15ng。临床检测共检出19份样品为犬瘟热阳性,其中15份为野毒株感染,其H基因编码区全长为1824bp,均在1279和1543处有NdeI酶切位点,推导氨基酸与野毒株的同源性在92．9％～96．9％,与疫苗株的同源性在89．0％～90．8％,进化树分析显示这些毒株在基因型上属于Asia-1型,为野毒株谱系。该方法的建立为临床上犬瘟热病毒的鉴别检测和诊断提供了依据。 相似文献
9.
从疑似犬瘟热(CD)病犬的脏器中分离到1株病毒,经间接免疫荧光试验、RT-PCR鉴定、红细胞凝集试验和对不同动物致病性试验,证实该病毒为犬瘟热病毒(CDV)强毒株,命名为CDV YD株。对H基因序列的测定和分析表明,YD株H基因与国内强毒株更接近,核苷酸同源性为98.4%~99.7%,氨基酸同源性为97.8%~99.7%,而与疫苗株核苷酸同源性较远为91.2%~91.5%,氨基酸同源性为90.3%~91.2%。推导的H蛋白氨基酸序列中有9个潜在的N连接糖基化位点(N-X-S/T),可能与病毒体外复制和中和抗体有关,另外有12个保守的半胱氨酸(Cys)残基,对H蛋白二级结构起重要作用;根据H基因核苷酸序列绘制的进化树表明,CDV YD株属于国内流行基因型:Asia-1型。该研究为了解当前中国CDV流行变异情况和犬瘟热疫苗的研发提供了数据。 相似文献
10.
11.
犬瘟热病毒小熊猫株H、F和N基因的克隆及表达 总被引：4，自引：0，他引：4
根据GenBank中发表的犬瘟热病毒（CDV）的核苷酸序列，设计并合成了扩增CDVH、F和N基因的3对引物，经RT—PCR分别扩增获得了CDV小熊猫株（LP株）H、F和N基因，并对H、F及N基因进行了克隆和序列测定。序列分析表明，CDV LP株属于强毒谱系，与CDV流行株的亲缘关系近．H基因含有较多潜在的糖基化位点．F和N基因相对比较保守。将CDV LP株H、F和N基因克隆入真核表达栽体pVAX1的CMV启动子下游，构建了CDV基因疫苗表达载体pVAXLPH、pVAXLPF、pVAXLPN，体外转染BHK-21细胞．用间接ELISA方法检测到目的蛋白的表达。用构建的3个表达质粒免疫小鼠，从小鼠血清中检测到了抗CDV抗体．初步证实用CDVH、F和N基因作为核酸疫苗免疫动物，可以激活机体的免疫应答。 相似文献
12.
Keawcharoen J Theamboonlers A Jantaradsamee P Rungsipipat A Poovorawan Y Oraveerakul K 《Veterinary microbiology》2005,105(2):137-142
The C-terminal part of the nucleocapsid protein gene of 13 canine distemper virus (CDV) isolates from Thailand, were analyzed. The nucleotide sequences were assigned to two clusters; cluster A exhibited a high degree of homology with the vaccine strain Onderstepoort, 99.10 and 97.61%, respectively, in the two isolates examined. Cluster B appeared closely related to virulent strains registered in the GeneBank database and to the virulent reference strain (A75/17); a total of 11 samples were analyzed, with 94.63-99.10% homology at the same position. The deduced amino acid sequences correlated with the two-nucleotide sequence clusters. However, there was no association among the CDV groups with histories of vaccination, sex, ages, clinical findings and evidence of viral antigen in tissues. 相似文献
13.
Wakasa C Iwatsuki K Ohashi K Nakamura K Kai C 《The Journal of veterinary medical science / the Japanese Society of Veterinary Science》2000,62(1):97-101
The nucleotide sequences of the phosphoprotein (P) of canine distemper virus (CDV) strains isolated between 1992 and 1996 in Japan were determined. This is the first report of the complete sequences of the P genes of recently prevalent CDV strains. The deduced amino acid sequences of the P, C and V proteins showed that in the new Japanese isolates, these proteins have approximately 93%, 90-91% and 92% identities with those of the Onderstepoort vaccine strain, respectively. The predicted functional regions were conserved. RNA editing resulting in a shift to the open reading frame (ORF) of the V protein was shown to occur with the same efficiency in both the field isolates and vaccine strain. 相似文献
14.
15.
从规模化养殖场鸭群气管和泄殖腔试子分离到新城疫病毒(NDV)27株,用2株针对NDV HN单抗进行抗原表位分析,并选择4个分离株进行F基因高变区(374bp)和HN基因全长序列分析。抗原表位分析结果显示,27个鸭分离株均能与其中一株单抗C3-B7反应,而与另外一株单抗1E5反应为阴性。F基因(374bp)序列分析结果显示,4个鸭分离株均属于NDV ClassⅠ分支,分离株之间核苷酸同源性为99.2%～100%;分离株与NDV ClassⅡ毒株遗传距离为0.9%～9.9%,与NDV ClassⅠ毒株遗传距离为38.5%～41.7%。根据核苷酸序列推导的氨基酸序列表明,4个鸭NDV分离株F蛋白裂解位点氨基酸模式为:112-EROERL-117。HN基因序列分析结果显示,4个鸭NDV分离株HN基因全长1851bp,编码585个氨基酸;同源性比较发现4个鸭NDV分离株之间核苷酸同源性为99.7%～99.8%,与NDV ClassⅡ毒株核苷酸同源性为68.4%～70.5%,与NDV ClassⅠ毒株核苷酸同源性为95.8%～98.0%。本研究结果显示,鸭分离毒均属于NDV ClassⅠ弱毒,在抗原表位和基因序列上与广泛应用的NDV弱毒疫苗株(LaSota)不同,这些毒株的来源有待进一步深入研究。 相似文献
16.
水貂犬瘟热CDV_3疫苗株基因组序列测定及H基因遗传稳定性分析 总被引：1，自引：0，他引：1
对水貂犬瘟热CDV3疫苗株基因组进行全序列测定与分析,以阐明该疫苗株的来源亲本毒株,基因特征及H基因遗传稳定性。将CDV3疫苗株基因组cDNA分9个重叠片段进行RT-PCR并克隆入pMD 18-T载体中,测定全长cDNA序列。并与CDV野毒参考株和疫苗株N、F和H基因氨基酸序列比对分析其基因变异情况。通过对CDV3疫苗株6个不同生产批次(Vero细胞培养代次)H基因序列测定以分析毒株的分子遗传稳定性。基因序列分析结果表明:CDV3疫苗株基因组全长15 690 bp,与CDV野毒株基因组同源性较低(93.0%~95.3%)。H基因核苷酸和氨基酸序列与Lederle疫苗株(EF418783)同源性最高,分别为99.6%和98.7%。而6个不同培养代次CDV3疫苗株H基因氨基酸序列无任何差异。由此证实CDV3疫苗株的亲本毒株与Lederle疫苗株有着最密切关系,不同培养代次的CDV3疫苗株H基因具有较高的遗传稳定性。 相似文献
17.
Gámiz C Martella V Ulloa R Fajardo R Quijano-Hernandéz I Martínez S 《Veterinary research communications》2011,35(6):381-390
Canine Distemper is a highly contagious viral systemic disease that affects a wide variety of terrestrial carnivores. Canine
Distemper virus (CDV) appears genetically heterogeneous, markedly in the hemagglutinin protein (H), showing geographic patterns
of diversification that are useful to monitor CDV molecular epidemiology. In Mexico the activity of canine distemper remains
high in dogs, likely because vaccine prophylaxis coverage in canine population is under the levels required to control effectively
the disease. By phylogenetic analysis based on the nucleoprotein (N) and on the H genes, Mexican CDV strains collected between
2007 and 2010 were distinguished into several genovariants, all which constituted a unique group, clearly distinct from field
and vaccine strains circulating worldwide, but resembling a CDV strain, 19876, identified in Missouri, USA, 2004, that was
genetically unrelated to other North-American CDV strains. Gathering information on the genetic heterogeneity of CDV on a
global scale appears pivotal in order to investigate the origin and modalities of introduction of unusual/novel CDV strains,
as well as to understand if vaccine breakthroughs or disease epidemics may be somewhat related to genetic/antigenic or biological
differences between field and vaccine strains. 相似文献
18.
Tiwari AK Kataria RS Indervesh Prasad N Gupta R 《Comparative immunology, microbiology and infectious diseases》2003,26(1):47-53
In order to differentiate infectious bursal disease virus (IBDV) isolates/strains, a quick method of RT-PCR followed by restriction enzyme analysis of VP1 gene sequence is being reported for the first time. A 480 bp fragment, comprising one of the RNA dependent RNA polymerase motifs of VP1 gene sequence of an Indian classical virus, an attenuated vaccine strain, Georgia and two Indian field isolates, genetically similar to reported very virulent strains of IBDV, was amplified by RT-PCR. Restriction enzyme digestion of PCR products with Taq1 enzyme generated distinct profile for field isolates, different from the classical and attenuated viruses, whereas restriction profile with BstNI restriction enzyme was similar in all the viruses, irrespective of the pathotype. Therefore, the present results suggest that Taq1 digestion can be taken up for the differentiation of field isolates from the classical and vaccine strains. The sequence analysis of VPI gene of reported very virulent IBD viruses from Europe and Japan, using 'MapDraw' programme of Lasergene software, revealed similar restriction enzyme profile as in Indian field isolates. 相似文献
19.
20.
Martella V Cirone F Elia G Lorusso E Decaro N Campolo M Desario C Lucente MS Bellacicco AL Blixenkrone-Møller M Carmichael LE Buonavoglia C 《Veterinary microbiology》2006,116(4):301-309
Canine distemper virus (CDV) is a highly contagious viral pathogen causing lethal disease in dogs and other mammalians. A high degree of genetic variation is found between recent CDV strains and the old CDV isolates used in the vaccines and such genetic variation is regarded as a possible cause of the increasing number of CDV-related diseases in dogs. The H gene shows the greatest extent of genetic variation that allows for distinction of various lineages, according to a geographical pattern of distribution and irrespective of the species of identification. In the present study, hemagglutinin (H) genes obtained from field strains detected from clinical specimens of Italian dogs were analyzed genetically. Phylogenetic analysis revealed that a homogeneous group of CDV strains is widespread in Italian dogs, all which are included into the European lineage. Unexpectedly, strains 179/04 and 48/05 clustered along with CDVs of the Arctic lineage, the highest identity being to strain GR88 (98.0 and 98.4%aa, respectively). The full-length sequence of a red fox CDV strain, 207/00 was also determined and analyzed. The H protein of the fox CDV strain was unrelated to strains within the major European lineage. These results suggest that at least three different CDV lineages are present in Italy. 相似文献