【作者】 闫笑；

【导师】 李天宪；

【作者基本信息】 中国科学院研究生院（武汉病毒研究所） ， 微生物学， 2004， 硕士

【摘要】 鸡传染性法氏囊病（infection bursal disease, IBD）是由鸡传染性法氏囊病毒（Infectious Bursal Disease Virus，IBDV）引起的鸡和火鸡的一种高度接触性传染疾病。本病主要侵2-11周龄鸡,其中3-6周龄发病最为常见.感染该病后，鸡体法氏囊遭到严重破坏，重症者导致直接死亡，感染而不死亡者则发生免疫抑制,导致鸡对马立克氏疫苗、新城疫疫苗等常见疫病疫苗的免疫应答降低,导致免疫失败.此疾病给世界各国的禽养殖业带来了巨大损失.IBDV属于双节段dsRNA病毒科(Birnaviridae)禽双节段RNA病毒属(Avibirnavius),无囊膜,呈二十面体对称,病毒粒子直径60nm.其基因组由A和B两个双链RNA片段组成。VP2基因位于IBDV-A节段，编码形成37-40kda蛋白是IBDV的主要结构蛋白和保护性抗原成分。VP2基因内存在一个氨基酸残基高变区(206至305个氨基酸),不同毒株间在这段区域的变异很大.这段氨基酸残基高变区内主要包括一个七肽区(S-W-S-A-S-G-S)和两个亲水区(I (212-224)和Ⅱ(314-324))。七肽区的氨基酸位点特别是丝氨酸位点与IBDV的毒力和致病力密切相关,当这些位点发生变化时,IBDV发生致弱变异为弱毒株.亲水区对于超强毒株抗原性极为重要,其氨基酸残基的改变极可能导致病毒搞原性的改变.另外，高变区内有一些特征性氨基酸位点(Q249、G 254、N279和T284)对IBDV毒力强弱具有决定作用,发生变化Q249K、G 254S、N279D和T284A时,IBDV成为超强毒株（very virulent IBDV，vvIBDV）。本研究采用来源于江苏地区分离的六株IBDV毒株，针对其IBDV-VP2基因高变区在不同毒株间发生的不同变化,以RT-PCR法对其VP2基因高变区进行扩增，构建重组质粒pMD18T-VP2并测序，对测序结果进行分析，并选取较典型的一些已发表IBDV毒株，与其高变区进行序列比对。以ClustalX软件,得到基因序列及氨基酸序列同同源性,IBDV Y3、P2G、P8G、SZ、Y5和W04(六株)与D6984（荷兰）的同源性达到99.0%以上，与其它一些超强毒株的同源性也达到了97.8%以上,并在关键氨基酸位点符合vvIBDV特征,表明本实验采用的六株IBDV均为vvIBDV.采用Phylip3.5软件分析作出进化树，结果显示六个毒株与欧<WP=3>洲和日本的超强毒株有较近的亲缘关系，而与美洲株较远。从同源性比较及进化树分析结果,为IBDV分子流行病学研究和疫苗的研制提供了科学依据。更多还原

【Abstract】 Infectious bursal disease（IBD） is a highly touched infectious disease caused by Infectious bursal disease virus (IBDV) . Immunosuppression is caused when chichens are infected by IBDV.The double stranded RNA(dsRNA) genome, which is divided over two segments, is packed within a capsid. VP2 gene was located in the larger genome segment A. VP2 protein was the main structural protein and protective antigen of IBDV. A hypervariable region (AA206-AA305) including a seven-peptide sequence (S-W-S-A-S-G-S) and two hydrophilic regions (Ⅰ(AA212-AA224)、Ⅱ(AA314-AA324)) existed in the VP2 amino acid sequence. The serine residues in the seven-peptide sequence played an important role in the virulence of IBDV. The hydrophilic regions were very important for antigenic determinants of IBDV. Besides, IBDV strain would become very virulent (vvIBDV) when some amino acid residues changed (Q249K、G254S、N279D、T284A). The hypervariable region of VP2 gene of six isolates (Y3、P2G、P8G、SZ、Y5 and W04) from Jiangsu province was amplified by RT-PCR . six segments were cloned into pMD18T vector ，sequenced and analyzed. The results indicated that the six isolates were closely related with each other and had very high similarity with D6984 strain isolated from Dutch (99.0%) and other vvIBDV (more than 97.8%). Their amino acid sequences had the same characteristics with vvIBDV amino acid sequences. Based on 26 IBDV sequences, phylogenetic tree was constructed. The six isolates and vvIBDVs isolated from <WP=5>Europe and Japan were in the same branch. So it was suggested that the six isolates (Y3、P2G、P8G、SZ、Y5 and W04) isolated from Jiangsu province were vvIBDV.更多还原