猪瘟病例的综合诊断及感染毒株遗传进化分析
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摘要
2018年5月,山东省菏泽市某育肥猪场新进仔猪突然出现急性发病死亡,死亡率高达30%以上。为确诊病原,分析病原的遗传演化规律,对病死猪进行了病理组织学诊断和病毒分离鉴定,并对分离株的E2基因进行了序列测定及遗传变异分析。结果显示,组织学病变符合猪瘟的病变特征,与参考毒株核苷酸和氨基酸的同源性分别为83.4%～97.9%和88.7%～98.7%,与本实验室从山东分离的流行毒株的核苷酸和氨基酸同源性均高于其他参考毒株。E2基因遗传进化分析表明,该分离株与本实验室分离的流行毒株遗传距离较近,同属于猪瘟病毒2.1d亚型分支。E2基因推导氨基酸突变位点分析表明,分离株的糖基化位点(T~(806)A)与所有参考株均不相同,其具体生物学意义有待研究。上述结果表明,该猪场仔猪的急性发病死亡是由猪瘟病毒感染所致,致病毒株属于猪瘟病毒2.1d亚型。
In May,2018,sudden deaths occurred in some new piglets in a fattening pig farm in Heze City of Shandong Province,and the mortality rate was more than 30%. In order to find out the pathogen and to analyze its genetic evolution status,pathological histological diagnosis and virus isolation and identification were carried out. Then the E2 gene of the isolated strain was sequenced and analyzed for genetic variation. According to the results,the histological lesions were consistent with the pathological features of swine fever,and the homologies of nucleotides and amino acids between the isolate and the reference strains were 83.4% to 97.9% and 88.7% to 98.7%,respectively,at the same time,the homologies were higher when compared with the strains isolated from Shandong than other provinces. According to the phylogenetic analysis of E2 gene,the isolate was close to the strain isolated by the laboratory of Shandong Agricultural University,and both of them belonged to the 2.1d subtype branch of classical swine fever virus(CSFV). Results of amino acid mutation site analysis showed that the glycosylation site(T~(806)A)of the isolate was different from all the reference strains,and its specific biological significance still needed to be studied. In conclusion,the epidemic was caused by swine fever virus and the virus strain belonged to the 2.1d subtype.
In May,2018,sudden deaths occurred in some new piglets in a fattening pig farm in Heze City of Shandong Province,and the mortality rate was more than 30%. In order to find out the pathogen and to analyze its genetic evolution status,pathological histological diagnosis and virus isolation and identification were carried out. Then the E2 gene of the isolated strain was sequenced and analyzed for genetic variation. According to the results,the histological lesions were consistent with the pathological features of swine fever,and the homologies of nucleotides and amino acids between the isolate and the reference strains were 83.4% to 97.9% and 88.7% to 98.7%,respectively,at the same time,the homologies were higher when compared with the strains isolated from Shandong than other provinces. According to the phylogenetic analysis of E2 gene,the isolate was close to the strain isolated by the laboratory of Shandong Agricultural University,and both of them belonged to the 2.1d subtype branch of classical swine fever virus(CSFV). Results of amino acid mutation site analysis showed that the glycosylation site(T~(806)A)of the isolate was different from all the reference strains,and its specific biological significance still needed to be studied. In conclusion,the epidemic was caused by swine fever virus and the virus strain belonged to the 2.1d subtype.
引文
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[15]DONG X N,CHEN Y H. Spying the neutralizing epitopes on E2 N-terminal by candidate epitope-vaccines against classical swine fever virus[J]. Vaccine,2006,24(19):4029-4034.
[16]RISATTI G,HOLINKA L,SAINZ I,et al. Mutations in the carboxyl terminal region of E2 glycoprotein of classical swine fever virus are responsible for viral attenuation in swine[J]. Virology,2007,364(2):371-382.
[17]彭伍平,夏照和,侯强,等.猪瘟病毒石门强毒株和兔化弱毒疫苗株E2蛋白糖基化位点差异分析[J].病毒学报,2007,23(5):389-393.
[18]张志,李晓成.我国猪瘟流行现状和防控建议[J].中国动物检疫,2015,32(8):8-12.
[2]TU C,LU Z,LI H,et al. Phylogenetic comparison of classical swine fever virus in China[J]. Virus research,2001,81(1):29-37.
[3]KááNIGM,LENGSFELDT,PAULYT,etal.Classical swine fever virus:independent induction of protective immunity by two structural glycoproteins[J].Journal of virology,1995,69(10):6479-6486.
[4]RISATTI G R,BORCA M V,KUTISH G F,et al.The E2 glycoprotein of classical swine fever virus is a virulence determinant in swine[J]. Virology,2005,343(1):116-127.
[5]FERNANDEZ-SAINZ I,HOLINKA L G,GAVRILOV B K,et al. Alteration of the N-linked glycosylation condition in E1 glycoprotein of classical swine fever virus strain brescia alters virulence in swine[J]. Virology,2009,386(1):210-216.
[6]肖昌,余兴龙,张茂林,等.利用随机肽库鉴定猪瘟病毒E2蛋白抗原表位[J].中国兽医学报,2005,25(5):449-453.
[7]PENG W P,HOU Q,XIA Z H,et al. Identification of a conserved linear B-cell epitope at the N-terminus of the E2 glycoprotein of classical swine fever virus by phagedisplayed random peptide library[J]. Virus research,2008,135(2):267-272.
[8]LIN M,LIN F,MALLORY M,et al. Deletions of structural glycoprotein E2 of classical swine fever virus strain alfort/187 resolve a linear epitope of monoclonal antibody WH303 and the minimal N-terminal domain essential for binding immunoglobulin G antibodies of a pig hyperimmune serum[J]. Journal of virology,2000,74(24):11619-11625.
[9]YU M,WANG L F,SHIELL B J,et al. Fine mapping of a C-terminal linear epitope highly conserved among the major envelope glycoprotein E2(gp51 to gp54)of different pestiviruses[J]. Virology,1996,222(1):289-292.
[10]仇华吉,童光志,沈荣显.猪瘟兔化弱毒疫苗——半个世纪的回顾[J].中国农业科学, 2005,38(8):1675-1685.
[11]TU C,LU Z,LI H,et al. Phylogenetic comparison of classical swine fever virus in China[J]. Virus research,2001,81(1):29-37.
[12]LUO Y,SU L,YUAN S,et al. Classical swine fever in China:A minireview[J]. Veterinary microbiology,2014,172(1/2):1-6.
[13]ZIJL M V,WENSVOORT G,KLUYVER E D,et al.Live attenuated pseudorabies virus expressing envelope glycoprotein E1 of hog cholera virus protects swine against both pseudorabies and hog cholera[J]. Journal of virology,1991,65(5):2761-2765.
[14]刘思国,于辉,涂长春.猪瘟病毒E2蛋白抗原表位的研究[J].中国动物检疫,2004,21(2):42-44.
[15]DONG X N,CHEN Y H. Spying the neutralizing epitopes on E2 N-terminal by candidate epitope-vaccines against classical swine fever virus[J]. Vaccine,2006,24(19):4029-4034.
[16]RISATTI G,HOLINKA L,SAINZ I,et al. Mutations in the carboxyl terminal region of E2 glycoprotein of classical swine fever virus are responsible for viral attenuation in swine[J]. Virology,2007,364(2):371-382.
[17]彭伍平,夏照和,侯强,等.猪瘟病毒石门强毒株和兔化弱毒疫苗株E2蛋白糖基化位点差异分析[J].病毒学报,2007,23(5):389-393.
[18]张志,李晓成.我国猪瘟流行现状和防控建议[J].中国动物检疫,2015,32(8):8-12.