一株2013年云南省Ⅲ型疫苗相关脊髓灰质炎病毒全基因组分析
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摘要
目的分析一株疫苗相关脊髓灰质炎病毒(vaccine-related poliovirus,VRPV)云南分离株R1351/YN/CHN/2013的全基因组序列和遗传特性。方法采用RD细胞分离获得病毒,提取病毒RNA,通过RT-PCR扩增全基因组序列并测序。利用BioEdit 7.09、MEGA7.0、SimPlot3.5.1等软件对全基因序列进行分析。结果 R1351/YN/CHN/2013全基因序列由7 430个核苷酸构成,编码含2 206个氨基酸残基的多聚蛋白,5′UTR和3′UTR核苷酸长分别为742 nt和70 nt;与Sabin3疫苗株全基因核苷酸和氨基酸序列的同源性分别为99.7%和99.6%;在5′UTR的减毒位点(nt472)发生回复突变,在VP3减毒位点(nt2034)未发生改变;与Sabin3疫苗株的VP1序列比对,共发生3个碱基突变,变异率为0.33%。进化分析显示73株Ⅲ型脊髓灰质炎病毒株分为A、B和C三个分支,R1351/YN/CHN/2013属于C分支,Sabin1/2疫苗株独立形成两个分支。重组分析表明R1351/YN/CHN/2013基因组各区域并未与Sabin1/2疫苗株及其他肠道病毒发生重组。结论 R1351/YN/CHN/2013为Ⅲ型VRPV,并且在减毒位点nt472处发生了回复突变。
Objective To analyze the complete genome and genetic characteristics of a vaccine-associated poliovirus(VRPV) strain R1351/YN/CHN/2013 isolated from Yunnan province of China in 2013. Methods The virus was isolated from fecal samples in RD cells.The viral RNA was extracted and the complete genome was amplified by RT-PCR for sequencing.The whole genome sequence was analyzed using BioEdit 7.09, MEGA7.0, and SimPlot 3.5.1 software. Results The full-length genome of the R1351/YN/CHN/2013 strain contains 7 430 nucleotides including a 5′-UTR of 742 nucleotides followed by an ORF that encodes a polyprotein of 2 206 amino acids and a 3′-UTR of 70 nucleotides.The homology of nucleotide and amino acid sequences with Sabin3 vaccine strain was 99.7% and 99.6%, respectively.A back mutation occurred at the 5′UTR attenuating site(nt472), but not at the VP3 attenuating site(nt2034).Sequencing analysis of VP1 showed a mutation rate of 0.33% for the R1351/YN/CHN/2013 strain and the Sabin3 vaccine strain.Evolutionary analysis demonstrated that 73 strains PV3 isolates can be divided into three branches:A, B and C.R1351/YN/CHN/2013 belonged to the C branch, while the Sabin1/2 vaccine strain formed two branches independently.Recombinant analysis indicated that the regions of the R1351/YN/CHN/2013 genome were not recombined with the Sabin1/2 vaccine strain or other enteroviruses. Conclusions R1351/YN/CHN/2013 strain is type Ⅲ VRPV and a back mutation occurred at the attenuating site nt472.
Objective To analyze the complete genome and genetic characteristics of a vaccine-associated poliovirus(VRPV) strain R1351/YN/CHN/2013 isolated from Yunnan province of China in 2013. Methods The virus was isolated from fecal samples in RD cells.The viral RNA was extracted and the complete genome was amplified by RT-PCR for sequencing.The whole genome sequence was analyzed using BioEdit 7.09, MEGA7.0, and SimPlot 3.5.1 software. Results The full-length genome of the R1351/YN/CHN/2013 strain contains 7 430 nucleotides including a 5′-UTR of 742 nucleotides followed by an ORF that encodes a polyprotein of 2 206 amino acids and a 3′-UTR of 70 nucleotides.The homology of nucleotide and amino acid sequences with Sabin3 vaccine strain was 99.7% and 99.6%, respectively.A back mutation occurred at the 5′UTR attenuating site(nt472), but not at the VP3 attenuating site(nt2034).Sequencing analysis of VP1 showed a mutation rate of 0.33% for the R1351/YN/CHN/2013 strain and the Sabin3 vaccine strain.Evolutionary analysis demonstrated that 73 strains PV3 isolates can be divided into three branches:A, B and C.R1351/YN/CHN/2013 belonged to the C branch, while the Sabin1/2 vaccine strain formed two branches independently.Recombinant analysis indicated that the regions of the R1351/YN/CHN/2013 genome were not recombined with the Sabin1/2 vaccine strain or other enteroviruses. Conclusions R1351/YN/CHN/2013 strain is type Ⅲ VRPV and a back mutation occurred at the attenuating site nt472.
引文
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[14] Wang FL,Liu SJ,Sun XY,et al.Analysis of a case of death of polio attenuated live vaccination [J].Yufang Yixue,2018,30 (9):943-945.(in Chinese)王富良,刘仕俊,孙向珏,等.一起脊髓灰质炎减毒活疫苗接种偶合死亡案例分析[J].预防医学,2018,30(9):943-945.
[2] Xu SF,Liu LQ,Miao LN,et al.Analysis of polio vaccine types and application status in Hebei Province of China[J].Yixue Dongwu Fangzhi,2018,34 (9):876-878.(in Chinese)徐书方,刘立秋,苗莉娜,等.脊髓灰质炎疫苗类型及河北省应用现状分析[J].医学动物防制,2018,34(9):876-878.
[3] Yan DM,Li XL,Zhang Y,et al.Analysis of genetic characteristics of type Ⅱ vaccine-derived poliovirus in Shanxi Province of China in 2014[J].Bingdu Xuebao,2015,31 (2):157-163.(in Chinese)严冬梅,李晓嫘,张勇,等.山西省2014年Ⅱ型疫苗衍生脊髓灰质炎病毒的基因特性分析[J].病毒学报,2015,31 (2):157-163.
[4] Nanteza MB,Bakamutumaho B,Kisakye A,et al.The detection of 3 ambiguous type 2 vaccine-derived polioviruses (VDPV2s) in Uganda[J].Virol J,2018,15 (1):77.
[5] Nix WA,Oberste MS,Pallansch MA.Sensitive,seminested PCR amplification of VP1 sequences for direct identification of all enterovirus serotypes from original clinical specimens[J].J Clin Microbiol,2006,44 (8):2698-2704.
[6] Tang G,Yin W,Cao Y,et al.Immunogenicity of sequential inactivated and oral poliovirus vaccines (OPV) versus inactivated poliovirus vaccine (IPV) alone in healthy infants:a systematic review and meta-analysis[J].Hum Vaccin Immunother,2018,14 (11):1-23.
[7] Ma S,Du Z,Feng M,et al.A severe case of co-infection with Enterovirus 71 and vaccine-derived Poliovirus type Ⅱ[J].J Clin Virol,2015,72 (72):25-29.
[8] Liu J,Zhang H,Zhao Y,et al.Characterization of four vaccine-related polioviruses including two intertypic type3/type2 recombinants associated with aseptic encephalitis[J].Virol J,2016,13 (1):162.
[9] Weeks-Levy C,Tatem JM,DiMichele SJ,et al.Identification and characterization of a new base substitution in the vaccine strain of Sabin 3 poliovirus[J].Virology,1991,185 (2):934-937.
[10] Mento SJ,Weeks-Levy C,Tatem JM,et al.Significance of a newly identified attenuating mutation in Sabin 3 oral poliovirus vaccine[J].Dev Biol Stand,1993,78 (78):93-99.
[11] Zhou B,Xu L,Zhu R,et al.A bispecific broadly neutralizing antibody against enterovirus 71 and coxsackievirus A16 with therapeutic potential[J].Antiviral Res,2018,161(161):28-35.
[12] Zhao Y,Zhang H,Liu H,et al.Molecular characteristics of hand,foot,and mouth disease for hospitalized pediatric patients in Yunnan,China[J].Medicine (Baltimore),2018,97 (31):e11610.
[13] Guan H,Wang J,Wang C,et al.Etiology of multiple non-EV71 and non-CVA16 enteroviruses associated with hand,foot and mouth disease in Jinan,China,2009-June 2013[J].Plos One,2015,10 (11):e142733.
[14] Wang FL,Liu SJ,Sun XY,et al.Analysis of a case of death of polio attenuated live vaccination [J].Yufang Yixue,2018,30 (9):943-945.(in Chinese)王富良,刘仕俊,孙向珏,等.一起脊髓灰质炎减毒活疫苗接种偶合死亡案例分析[J].预防医学,2018,30(9):943-945.