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1.
最近,一株无毒力的新城疫病毒(NDV)在鸡体内繁殖时,变成了强毒株,但致今尚未能证实,经鸡体传代的野生水禽新城疫病毒是否也具有变成速发型毒株的能力,为了通过实验证明从水禽中分离的非致病型NDV可以转变为速发型病毒,我们通过在鸡体内传播鹅源性无毒力株,经气囊接种连续传代9次,随后再在鸡脑内传代5次,结果显示,该病毒的毒力变得很强,致死率可达100%,通过致病性试验证实,其具有典型的速发型病毒特征;融合蛋白裂解位点的序列分析表明,原始的分离株含有无毒力型毒株共有的裂解序列;E-R-Q-E-R/L,而通过鸡体反复传代后,该序列变为致癞 性毒株共有的序列:K-R-Q-K-R/F,这些结果表明,野生水禽中自然存在的无毒力毒株,具有无毒力株相应的序列,但当其在鸡群中传播时,则具有变成高致病性病毒的能力,同时研究表明,鸡体提供了该病毒从非致病型向致病型转变的选择机制。  相似文献   

2.
A triple one-step RT-PCR was developed to screen and differentiate virulent from avirulent Newcastle disease virus (NDV) isolates. Three sets of oligonucleotides were designed, each specific for amplifying NDV fusion protein gene-specific RNA from virulent, avirulent or all isolates respectively. The sensitivity of one-step RT-PCR was determined using viral RNA extracted from serially diluted NDV-infected allantoic fluid and found to be 10(-5) HA units. Application of one-step RT-PCR to various NDV samples, including wild-type virulent isolates and avirulent vaccine strains, demonstrated the potential for rapid identification (3-4 h) of NDV isolates as well as the differentiation of virulent from avirulent strains.  相似文献   

3.
Hybridisation of PCR fragments with fluorogenic probes specific for pathotype allowed an estimation of pathogenicity of Newcastle disease virus (NDV) isolates using a modified TaqMan procedure. Six probes were used, designed to recognise nucleotide sequences in the fusion protein gene sequence corresponding to the precursor protein F0 cleavage site of both virulent and avirulent viruses. Forty-three of the 45 isolates tested, including 18 examined in a blind study were pathotyped successfully and rapidly, with close correlation between cleavage site nucleotide sequences, TaqMan results and intracerebral pathogenicity index (ICPI) values. One isolate, which could not be pathotyped by nucleotide sequencing, was shown using the TaqMan system to be a mixture of virulent and avirulent NDV. The results of this study suggest that using this modified TaqMan protocol, the likely virulence of most ND isolates can be determined rapidly and reproducibly.  相似文献   

4.
本实验以分子生物学和现代免疫学方法,对新城疫病毒(NDV)中国毒株F48E9和LaSota等毒株的糖蛋白的致病作用进行了探讨。结果表明,NDV F和HN蛋白为糖蛋白,强弱毒株之间F蛋白裂解位点的氨基酸序列有明显的不同,并决定了毒株的毒力,但NDV的致病作用尚需HN、F蛋白的协同作用,单一的F或HN蛋白不能构成NDV的致病作用。  相似文献   

5.
应用抗多肽抗体鉴别新城疫病毒强弱毒株   总被引:2,自引:0,他引:2  
在克隆我国流行的新城疫病毒(NDV)强毒株F48E8株、四平株及弱毒株长春株、V4株和HN和F基因并进行测序的基础上,针对我国流行的NDV强毒株F蛋白前体(F0)的F2片段的特异结构,人工合成特异性多肽,将其与小牛血清白蛋白(BSA)化学偶联制备成全抗原,免疫小鼠制备出抗多肽血清。经ELISA检测,该抗体与NDV强毒株呈强阳性反应,而与鸡痘病毒、鸡传染性法氏囊病病学、ND 弱毒株长春株和V4株呈阴  相似文献   

6.
Degenerate primers based RT-PCR (previously described by [Avian Dis 26 (1997) 837]) has been used for the detection and differentiation of Newcastle disease (ND) viruses. Two sets of primers (A+B and A+C), with common forward primer and distinct reverse degenerate primers, designed from fusion protein gene encoding for cleavage site, could differentiate virulent and avirulent Newcastle disease viruses (NDV). Both sets of primers amplified "F" gene sequence of virulent (velogenic and mesogenic) viruses, whereas in avirulent strains, amplification was only with primer set A+C. Total 10 NDV isolates and two clinical samples including both known and unknown pathotypes, were checked. Based on amplification results 5 viruses were found to be virulent type and 6 as avirulent with one of the two clinical samples, earlier positive by RT-PCR using non-degenerate "F" gene specific primers was found negative in this study. The technique has been found to be a simple and quick for the detection and differentiation of virulent and avirulent NDV, which is important for control of the disease in the events of the outbreaks.  相似文献   

7.
本实验选择最近分离的1株鸭源新城疫病毒,采用已经发表的鸡新城疫病毒序列设计引物,应用RT-PCR技术扩增鸭源新城疫病毒的F基因,并将其克隆至载体上,然后进行了核苷酸序列、氨基酸序列测定,结果表明该分离株为新城疫强毒株。  相似文献   

8.
从病死的鸡、鹅、鸽中分离到3株禽I型副黏病毒(CPMV、GPMV、PPMV)。毒力测定结果显示,CPMV、GPMV的毒力与鸡新城疫病毒(NDV)F48E9株的毒力相似,为强毒型;PPMV的毒力与鸡NDVLaSota株的毒力相似,为自然弱毒株。对该分离株F基因裂解位点的序列分析结果表明,CPMV、GPMVF裂解位点氨基酸序列为112K-R-Q-K-R-F117,符合强毒株的序列特征;PPMV F裂解位点氨基酸序列为112G-R-Q-G-R-L117,属于弱毒株的特征性序列。将该3株病毒制成灭活疫苗,分别免疫鸡,结果都能诱导鸡体产生较高水平的HI抗体;再分别以CPMV、GPMV攻击免疫鸡群,结果显示免疫鸡群可得到保护,而非免疫鸡群被攻击后全部致死。用HI方法检测3株病毒与Lasota株之间的交叉血凝抑制试验,结果表明LaSota与GPMV两毒株间的抗原性无明显差异,LaSota与CPMV、PPMV两毒株间的抗原性有较小的差异。本研究将有助于进一步开展禽Ⅰ型副黏病毒的致病机制和新型疫苗等方面的研究。  相似文献   

9.
During November 2002, six double-crested cormorants (DCCs; Phalacrocorax auritus) were found moribund in Big Pine Key, FL, exhibiting clinical signs indicative of neurologic disease. Postmortem diagnostic evaluations were performed on two adult birds. Virulent Newcastle disease virus (NDV) was isolated from a cloacal swab from cormorant 1. West Nile virus (WNV) was isolated from the brain and lung of cormorant 2. Nucleotide sequence analysis of a portion of the fusion (F) protein gene of the NDV cormorant isolate revealed it shared a 100% deduced amino acid identity with only two viruses: the 1992 epizootic cormorant isolate from Minnesota and the 1992 turkey isolate from North Dakota. The epidemiologic significance of the recognition of virulent NDV on cormorant wintering grounds during a nonepizootic period, in addition to the potential implications of the concurrent isolation of NDV and WNV from cormorants, is discussed.  相似文献   

10.
从山东济南某非典型新城疫发病鸡群中分离到一株新城疫病毒株(ShD-5—06),研究其生物学特性表明,该病毒具有新城疫强毒株的一些特征。从该分离株扩增出其F和HN基因,并与标准株进行同源性比较,为探讨NDV是否发生变异提供理论依据。本试验通过RT—PCR法特异性地扩增出F和HN基因全基因序列,并对其与已经发表的序列进行核苷酸序列测定和分析。结果表明,ShD-5—06株的F和HN基因开放性阅读框架(ORF)为1662bp和1716bp,分别编码489个和571个氨基酸。与国外发表的部分新城疫病毒强毒株和弱毒株之间相应序列进行比较,F基因核苷酸序列的同源性在84.1%~88.7%之间,氨基酸同源性在88.1%~93.3%之间;HN基因核苷酸序列的同源性在82.19,5~87.4%之间,氨基酸同源性在88.6%~90.9%之间;F蛋白裂解位点区(112~117)氨基酸组成与强毒株一致,说明NDV山东分离株(ShD-5—06)为新城疫强毒株。  相似文献   

11.
采用RT-PCR技术对Ⅰ类新城疫病毒(NDV)09-014分离株完整的融合蛋白(F)基因和血凝素-神经氨酸酶(HN)基因进行了扩增和遗传进化分析。F基因的序列测定结果表明:该分离株F基因全长为1 792 bp,可编码553个氨基酸,裂解位点的氨基酸组成为112E-R-Q-E-R-L117,具有典型的新城疫弱毒株特征。同源性分析表明本分离株的F基因与Ⅰ类新城疫病毒代表毒株之间核苷酸的同源性为93%~95.2%,而与Ⅱ类新城疫病毒代表毒株的同源性较低,介于70.6%~72.4%。HN基因的序列测定结果表明:HN基因全长2 001 bp,可编码616个氨基酸,同源性分析表明本分离株的HN基因与Ⅰ类新城疫病毒代表毒株之间核苷酸的同源性在92.7%~94.7%之间,而与Ⅱ类新城疫病毒同源性较低,为70.7%~71.5%。根据完整的F基因和HN基因构建的遗传进化树均表明:本分离株在分类地位上属于Ⅰ类新城疫病毒基因3型,因此Ⅰ类新城疫病毒的F基因和HN基因具有相似的进化速率。  相似文献   

12.
The nucleoprotein (NP) of Newcastle disease virus (NDV) was selected to study the relative importance of an internal structural protein in the avian immune response. The NP gene of the virulent, neurotropic NDV Texas GB (TGB) strain was cloned and sequenced. Nucleotide sequence data for the NP gene allowed comparison of the deduced amino acid sequences for the NP genes of NDV-TGB and the avirulent duck isolate NDV-D26. These comparisons demonstrated an 89% nucleotide sequence homology and a 97% homology between the deduced amino acid sequences. The NDV-TGB NP expressed in recombinant vaccinia virus (rVAC) was electrophoretically and immunologically identical to the wild-type NDV-TGB. Although inoculation of chickens with the recombinant vaccinia virus expressing the NDV NP gene elicited anti-NDV antibodies in higher titers than in birds inoculated with live LaSota NDV, this strong anti-NDV response did not protect against lethal challenge with NDV-TGB.  相似文献   

13.
一株Class Ⅰ新城疫病毒中国分离株分子特性的研究   总被引:1,自引:1,他引:1  
对从健康家鸭中分离到的一株Class I新城疫病毒分离株Duck/China/08-004/2008进行了遗传进化特性研究。利用RT-PCR扩增了该分离株F基因的主要功能区片段,并进行了克隆与序列分析。序列测定结果已经登录到GenBank,登录号为EU589149。该分离株F蛋白裂解位点的组成为112E-Q-Q-E-R-L117,具有典型新城疫弱毒株的分子特征。同源性分析表明其与国内普遍使用的弱毒疫苗株LaSota和V4核苷酸的同源性较低(分别为55.4%和57.7%)。通过构建F基因的遗传进化树,结果表明该分离株在分类地位上属于ClassI,与我国目前普遍使用的弱毒疫苗株LaSota(Class II中的基因II型)和V4(Class II中的基因I型)处在不同的进化分支。通过构建57株ClassI新城疫病毒的遗传进化树,表明本分离株与近年来香港活禽市场分离株较为类似,同属于基因3型。  相似文献   

14.
新城疫病毒F48E9株病毒糖蛋白的功能分析   总被引:4,自引:1,他引:3  
对抗新城疫病毒(NDV)HN和F糖蛋白的单克隆抗体(McAb)的生物学活性,应用ELISA、HI、HLI、Westernblot等进行了分析,结果证明,NDVF48E9株HN蛋白除有HA和NA功能区外,还有一个促进(启动)融合的功能区。此外,还筛选到4株NDV强毒株特异性McAb,为NDV强弱毒株的鉴别打下了基础  相似文献   

15.
A total of 38 Newcastle disease virus (NDV) isolates were obtained from 6060 fecal samples from northern pintail (Anas acuta) ducks collected in the Tohoku district in Japan during 2006-09. One isolate from each sampling location and date was selected for a total of 38 isolates, then 15 of these were characterized for their pathogenicity by mean death time of minimum lethal dose (MDT/MLD) using chicken embryos and by plaque formation on chicken embryo fibroblasts. Furthermore, nine isolates were randomly selected from these 15 isolates, and the fusion protein genes were sequenced to characterize amino acid sequences around the cleavage site. All 15 were confirmed to be nonvirulent by MDT/MLD test, and nine isolates were also confirmed as nonvirulent by the cleavage site of the fusion protein 112G/E-K/R-Q-G/E-R*L117 that was specific for nonvirulent NDVs. The characteristics of nine isolates identified by phylogenic analysis of the fusion protein gene indicated that the isolates belong to genotype I or II. In addition, we also isolated 68 avian influenza viruses and 28 other hemagglutinating viruses. Our data indicate that northern pintails are subclinically infected by, perpetuate, and distribute NDV along with different subtypes of avian influenza viruses and other hemagglutinating viruses during their migrations across vast areas over the Northern Hemisphere to Japan.  相似文献   

16.
A Newcastle disease virus (NDV) isolate designated IBS002 was isolated from a commercial broiler farm in Malaysia. The virus was characterised as a virulent strain based on the multiple basic amino acid motif of the fusion (F) cleavage site 112RRRKGF117 and length of the C-terminus extension of the hemagglutinin-neuraminidase (HN) gene. Furthermore, IBS002 was classified as a velogenic NDV with mean death time (MDT) of 51.2 h and intracerebral pathogenicity index (ICPI) of 1.76. A genetic distance analysis based on the full-length F and HN genes showed that both velogenic viruses used in this study, genotype VII NDV isolate IBS002 and genotype VIII NDV isolate AF2240-I, had high genetic variations with genotype II LaSota vaccine. In this study, the protection efficacy of the recombinant genotype VII NDV inactivated vaccine was also evaluated when added to an existing commercial vaccination program against challenge with velogenic NDV IBS002 and NDV AF2240-I in commercial broilers. The results indicated that both LaSota and recombinant genotype VII vaccines offered full protection against challenge with AF2240-I. However, the LaSota vaccine only conferred partial protection against IBS002. In addition, significantly reduced viral shedding was observed in the recombinant genotype VII-vaccinated chickens compared to LaSota-vaccinated chickens.  相似文献   

17.
从产蛋下降鸭群分离到两株新城疫病毒(NDV)(命名为Duck/China/SD6/2008和Duck/China/SD7/2009),经蚀斑纯化后测定其鸡胚平均死亡时间为77.6h,F蛋白裂解位点氨基酸序列为112-RRQKRF-117,符合强毒NDV的特征。根据F基因(374bp)对2个NDV分离株和30个NDV参考株进行基因分型结果表明,2个分离株均属于基因Ⅶ型。F基因和HN基因核苷酸序列同源性比较显示,2个鸭分离株与基因Ⅶ型NDV分离株同源性为94.1%~99.7%和94.4%~97.1%,与同期分离的鸡源NDV强毒株Chicken/China/SD4/2008同源性最高,分别为99.3%~99.5%和99.5%~99.7%,表明这两个鸭NDV毒株可能来源于感染NDV的鸡群。  相似文献   

18.
一株鸡副粘病毒F基因重要片段的扩增、克隆与分析   总被引:7,自引:0,他引:7  
应用RT-PCR技术对新城疫病毒(NDV)Y98株的F基因重要功能区片段扩增、克隆后进行序列测定,并与多株已报道的NDV参考株相应片段进行序列比较,经遗传基因进行化树分析后,首次报道了VII型DNV分离株在中国大陆的存在。  相似文献   

19.
20.
利用RT-PCR技术对番鸭源新城疫病毒FP1/02株的F蛋白基因进行分段扩增、定向克隆到pMD 18-T Simple Vector质粒载体,然后制定其核苷酸序列,拼接出F基因全序列.并推导出其相应的氨基酸序列。FP1/02株的F蛋白基因全长1690bp.编码553个氨基酸,其裂解位点的氨基酸序列为^112R-R-Q-K-R-F^112.具有强毒蛛特有的氨基酸序列结构特征。核苷酸序列分析结果表明,FP1/02株与其他不同源新城疫病毒毒株之间的核苷酸序列同源性为87.2%~93.3%.  相似文献   

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