3株猪博卡病毒的全基因遗传进化及重组分析  

Genetic analysis and recombinant identification of three porcine bocaviruses

作　　者：覃绍敏[1] 王浩 刘金凤[1] 莫模双[3] 秦树英[1] 陈凤莲 马玲[1] 白安斌[1] 吴健敏[1] QIN Shao-min;WANG Hao;LIU Jin-feng;MO Mo-shuang;QIN Shu-ying;CHEN Feng-lian;MALing;BAI An-bin;WU Jian-min(Guangxi Veterinary Research Institute/Guangxi Key Laboratory of Veterinary Biotechnology,Nanning 530001,China;College of Animal Science and Technology,Guangxi University,Nanning 530004,China;Fangchenggang Center for Animal Disease Control and Prevention,Fangchenggang,Guangxi 538001,China)

机构地区：[1]广西兽医研究所/广西兽医生物技术重点实验室,南宁530001 [2]广西大学动物科学技术学院,南宁530004 [3]防城港市动物疫病预防控制中心,广西防城港538001

出　　处：《南方农业学报》2020年第6期1247-1255,共9页Journal of Southern Agriculture

基　　金：广西科技重大专项(桂科AA18118051);国家现代农业产业技术体系广西生猪创新团队建设项目(nycytxgxcxtd-15-01);广西水产畜牧兽医局科技计划项目(桂渔牧科1304524)。

摘　　要：【目的】了解猪博卡病毒(Porcine bocavirus,PBoV)在广西猪群中的流行状况及遗传学特征,为有效防控PBoV感染提供科学依据。【方法】采用PCR对采自广西境内养殖场的388份猪源样品进行PBoV检测,挑选3份阳性样品(1份为PBoVG1阳性样品,2份为PBoVG3阳性样品)进行全基因扩增,测序获得的序列使用LaserGene进行处理及多重比对分析,应用MEGA 5.2中的ClustalW进行遗传进化分析,通过邻接法(NJ)构建系统发育进化树,并以SimPlot 3.5.1进行潜在基因重组事件分析。【结果】猪内脏组织(肺脏、脾脏和淋巴结的混合样品)、扁桃体、脑组织和公猪精液的PBoV阳性率分别为25.19%、8.33%、2.94%和1.54%。在所有检测样品中,PBoVG3基因群的阳性率最高,为9.02%,PBoVG2和PBoVG1基因群的阳性率均为1.55%;不同基因群的PBoV存在混合感染现象,其中,PBoVG1+PBoVG3二重感染率为1.03%,PBoVG2+PBoVG3二重感染率为0.77%。从PBoV阳性样品中扩增获得3条PBoV全基因序列(毒株),命名为GXBH2014、GXBH2015和GXLC2014,对应的GenBanK登录号为MN747332、MN747333和MN747339。其中,GXBH2014株序列全长5055 bp,GXBH2015株序列全长5070 bp,GXLC2014株序列全长4313 bp。GXBH2014株和GXBH2015株与PBoVG3基因群参考毒株SH20F各编码基因的推导氨基酸序列同源性较高,为70.9%~98.5%;GXLC2014株与PBoVG1基因群参考毒株SX各编码基因的推导氨基酸序列同源性最高,为98.0%~99.1%。基于PBoV全基因编码区(CDS)全长序列构建的系统发育进化树也显示,GXBH2014株和GXBH2015株同处于PBoVG3基因群分支上,GXLC2014株处于PBoVG1基因群分支上。基因重组分析结果显示,在GXBH2014株的全基因序列中检测到1个潜在的重组断点,位于NP1基因的第391位核苷酸;而在GXBH2015株和GXLC2014株的全基因序列中均未检测到潜在的重组断点。【结论】广西猪群中普遍存在PBoV感染,且PBoVG1、PBoVG2和PBoVG3基因群毒株同时存在。其中,GXBH2014株可能是【Objective】The study was performed to survey the prevalence and genetic characterization of porcine bocavirus(PBoV)in Guangxi in order to provide scientific basis for effective prevention and control of PBoV infection.【Method】Three hundred and eighty eight porcine samples were collected from pig farms in Guangxi and were detected for PBoV by PCR.The genomic DNA of PBoVs were amplified,cloned and sequenced from three PCR positive samples(one PBoVG1 positive sample and two PBoVG3 positive samples).Multiple comparative analysis were conducted for obtained genomic sequences using LaserGene software.Genetic analysis were performed by ClustalW method using MEGA 5.2 software and neihgbor-joining(NJ)phylogenetic trees were constructed.Recombinant analysis were carried out using Sim-Plot 3.5.1 software.【Result】The positive rate of PBoV infection were 25.19%in the mixture(lung,spleen and lymph node)samples,8.33%in the tonsil samples,2.94%in the brain samples and 1.54%in the semen samples.All three PBoV genogroups were detected in the investigated samples with the infection rate of 9.02%for PBoVG3(the highest),1.55%for both G2 and G1,1.03%for co-infection of G1+G3,and 0.77%for co-infection of G2+G3.Additionally,the whole genomes sequences of three PBoV strains were obtained by amplification from PBoV positive samples and were designated as GXBH2014,GXBH2015 and GXLC2014 with genomic length of 5055,5070 and 4313 bp,respectively.The genomic sequences of PBoV strains GXBH2014,GXBH2015 and GXLC2014 were submitted to GenBank under accession numbers MN747332,MN747333 and MN747339.The deduced amino acid of GXBH2014 and GXBH2015 shared high homology with encoded genes of PBoVG3 genetic group reference strain SH20F,being 70.9%-98.5%.GXLC2014 and PBoVG1 genetic group reference strain SX shared maximum deduced amino acid identities,being 98.0%-99.1%.Phylogenetic trees based on the full-length coding sequence(CDS)of PBoV showed that PBoV strains GXBH2014 and GXBH2015 belonged to PBoVG3 genetic group and GXLC2014 belonged to

关 键 词：猪博卡病毒 全基因组 遗传进化分析 基因重组分析 

分 类 号：S852.659.2[农业科学—基础兽医学]