山东省免疫猪场猪伪狂犬病病毒gC基因的序列分析
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摘要
为了调查山东省使用Bartha-K61疫苗免疫后,猪场猪伪狂犬病(PR)仍旧流行的原因,本研究对山东省动物疫病预防与控制中心在免疫猪场分离到的12株PRV的gC基因进行克隆测序与分析。核苷酸和氨基酸比对结果表明PRV欧美毒株与国内毒株亲缘关系较远,序列具有明显的地域特征,而大部分国内2011年后的分离毒株在氨基酸进化树上位于同一个分支,这表明gC基因的变异具有一定的时间特征。国内毒株gC蛋白的氨基酸与Bartha株的比对结果显示两者在24个位点上存在差异,并且国内毒株在63位之后增加了7个氨基酸AAASTPA。通过对LDZ01-2014毒株与Bratha株的gC蛋白抗原指数、亲水性和表面可及性分析发现两者的差异主要体现在gC蛋白的氨基端1/3部分,以期获得部分山东省Bartha-K61疫苗免疫失败的原因,并为以后的病原学研究和PR的有效防控提供参考。
In this research,gC gene of 12 pseudorabies virus(PRV) strains were isolated from Bartha-K61 vaccinated swine population of Shandong province in 2013 and 2014,and sequenced,in order to study the reasons for the prevalence of porcine pseudorabies(PR) in the use of Bartha-K61 vaccine in swine farms of Shandong province.The results showed that PRV strains isolated in China showed marked sequence divergence and phylogenetic analysis showed regional characteristics compared to Euramerican strains.In addition,most Chinese strains isolated after 2011 belonged to a relatively independent branch in phylogenetic analysis.Significant amino acid differences can be seen at 24 sites between Bartha and Chinese domestic strains and three were 7 amino acids of AAASTPA inserted at the 63 rd amino acid of the gC protein of the Chinese domestic strains.Prediction results of the antigenic index,hydrophilic and surface accessibility of gC protein demonstrated that the discrepancy of strain LDZ01-2014 and Bartha were mainly located in the N-terminal one third part of gC protein.From the results,we hope to obtain the reasons for the failure of some of the Bartha-K61 vaccines in Shandong province,and provide reference for future etiology studies and effective prevention and control of PRV.
In this research,gC gene of 12 pseudorabies virus(PRV) strains were isolated from Bartha-K61 vaccinated swine population of Shandong province in 2013 and 2014,and sequenced,in order to study the reasons for the prevalence of porcine pseudorabies(PR) in the use of Bartha-K61 vaccine in swine farms of Shandong province.The results showed that PRV strains isolated in China showed marked sequence divergence and phylogenetic analysis showed regional characteristics compared to Euramerican strains.In addition,most Chinese strains isolated after 2011 belonged to a relatively independent branch in phylogenetic analysis.Significant amino acid differences can be seen at 24 sites between Bartha and Chinese domestic strains and three were 7 amino acids of AAASTPA inserted at the 63 rd amino acid of the gC protein of the Chinese domestic strains.Prediction results of the antigenic index,hydrophilic and surface accessibility of gC protein demonstrated that the discrepancy of strain LDZ01-2014 and Bartha were mainly located in the N-terminal one third part of gC protein.From the results,we hope to obtain the reasons for the failure of some of the Bartha-K61 vaccines in Shandong province,and provide reference for future etiology studies and effective prevention and control of PRV.
引文
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[13] 陈懿,聂奎.伪狂犬病病毒囊膜蛋白gD研究进展[J].动物医学进展,2006,27(6):20-23.
[14] 陈超,刘存,李海娥,等.山东省免疫猪场猪伪狂犬病毒分离鉴定及gE毒力基因的序列分析[J].中国兽医学报,2016,36(6):902-907.
[15] 许雁峰.伪狂犬病病毒gD基因的生物信息学分析[D].四川雅安:四川农业大学,2006.
[16] 王键义.猪伪狂犬病病毒gC基因的克隆及生物信息学分析[D].四川雅安:四川农业大学,2008.
[17] YE C,ZHANG Q Z,TIAN ZJ,et al.Genomic characterization of emergent pseudorabies virus in China reveals marked sequence divergence:evidence for the existence of two major genotypes[J].Virology,2015,483:32-43.
[18] 彭金美,安同庆,赵鸿远,等.猪伪狂犬病病毒新流行株的分离鉴定及抗原差异性分析[J].中国预防兽医学报,2013,35(1):1-4.
[19] 赵鸿远,彭金美,安同庆,等.猪伪狂犬病病毒变异株的分离鉴定及其gE基因的分子特征[J].中国预防兽医学报,2014,36(7):506-509.
[20] TRYBALA E,BERGSTR?M T,SPILLMANN D,et al.Interaction between pseudorabies virus and heparin/heparan sulfate.pseudorabies virus mutants differ in their interaction with heparin/heparan sulfate when altered for specific glycoprotein C heparin-binding domain[J].J Biol Chem,1998,273(9):5047-5052.
[2] 邓仕伟,汪勇,薛春芳.我国伪狂犬病流行现状及新特点[J].动物医学进展,2006,27(9):105-107.
[3] 蔡宝祥.家畜传染病学 [M].4版.北京:中国农业出版社,2001.
[4] 王钊哲,舒相华,杨亮宇,等.云南省部分地区72家规模化猪场猪伪狂犬病净化效果分析[J].中国兽医学报,2017,37(4):597-600.
[5] METTENLEITER T C.Immunobiology of pseudorabies (aujeszky?s disease)[J].Vet Immunol Immunopathol,1996,54(1/4):221-229.
[6] METTENLEITER T C.Aujeszky's disease (pseudorabies) virus:the virus and molecular pathogenesis-state of the art,June 1999[J].Vet Res,2000,31(1):99-115.
[7] 高泽文,范桂芳,赵婷婷,等.猪伪狂犬病病毒流行株遗传进化及序列比对分析[J].中国兽医学报,2018,38(1):1-10.
[8] MARCHIOLI C C,YANCEY R J,WARDLEY R C,et al.A vaccine strain of pseudorabies virus with deletions in the thymidine kinase and glycoprotein X genes[J].Am J Vet Res,1987,48(11):1577-1583.
[9] 周春陵,谭鑫,许拓,等.PRV变异株的分离及抗原差异性分析[J].中国兽医学报,2017,37(3):404-409.
[10] COE N E,MENGELING W L.Mapping and characterization of neutralizing epitopes of glycoproteins gIII and gp50 of the Indiana-Funkhauser strain of pseudorabies virus[J].Arch Virol,1990,110(1/2):137-142.
[11] ELOIT M,FARGEAUD D,L'HARIDON R,et al.Identification of the pseudorabies virus glycoprotein gp50 as a major target of neutralizing antibodies[J].Arch Virol,1988,99(1/2):45-56.
[12] PEETERS B,DE WIND N,HOOISMA M,et al.Pseudorabies virus envelope glyeoproteins gp50 and gⅡ are essential for virus penetration,but only gⅡ is involved in membrane fusion[J].J Virol,1992,66(2):894-905.
[13] 陈懿,聂奎.伪狂犬病病毒囊膜蛋白gD研究进展[J].动物医学进展,2006,27(6):20-23.
[14] 陈超,刘存,李海娥,等.山东省免疫猪场猪伪狂犬病毒分离鉴定及gE毒力基因的序列分析[J].中国兽医学报,2016,36(6):902-907.
[15] 许雁峰.伪狂犬病病毒gD基因的生物信息学分析[D].四川雅安:四川农业大学,2006.
[16] 王键义.猪伪狂犬病病毒gC基因的克隆及生物信息学分析[D].四川雅安:四川农业大学,2008.
[17] YE C,ZHANG Q Z,TIAN ZJ,et al.Genomic characterization of emergent pseudorabies virus in China reveals marked sequence divergence:evidence for the existence of two major genotypes[J].Virology,2015,483:32-43.
[18] 彭金美,安同庆,赵鸿远,等.猪伪狂犬病病毒新流行株的分离鉴定及抗原差异性分析[J].中国预防兽医学报,2013,35(1):1-4.
[19] 赵鸿远,彭金美,安同庆,等.猪伪狂犬病病毒变异株的分离鉴定及其gE基因的分子特征[J].中国预防兽医学报,2014,36(7):506-509.
[20] TRYBALA E,BERGSTR?M T,SPILLMANN D,et al.Interaction between pseudorabies virus and heparin/heparan sulfate.pseudorabies virus mutants differ in their interaction with heparin/heparan sulfate when altered for specific glycoprotein C heparin-binding domain[J].J Biol Chem,1998,273(9):5047-5052.
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