一株引起严重急性呼吸道感染病例的A亚型人呼吸道合胞病毒全基因组基因特征分析
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摘要
人呼吸道合胞病毒(Human respiratory syncytial virus,HRSV)是导致儿童急性呼吸道感染的最重要的呼吸道病毒之一。根据对单克隆抗体的反应,HRSV分为A、B两个亚型。为探讨严重急性呼吸道感染(Severe acute respi-ratory infection,SARI)病例中HRSV全基因组基因特征,本研究对2017年河南省漯河市住院SARI病例中检测到的1株HRSV A亚型病毒通过Sanger测序方法对其全基因组序列进行了测定和分析。通过Sequencher 5.4.5、MEGA 5.05、BioEdit 7.0.5等生物信息学软件进行序列拼接和比对,进行了基因亲缘性关系分析、氨基酸变异和糖基化位点分析。基于HRSV全基因组序列和11个单个蛋白基因序列构建的亲缘性关系分析结果提示本研究中检测到的这株HRSVA病毒(RSVAs/Luohe.Henan/CHN/42.17)属于ON1基因型,该型是我国近年流行的优势基因型。该病毒全基因组序列与35条全球代表株的核苷酸和氨基酸同源性分别为92.69%～99.82%和93.63%～99.67%;G蛋白编码区氨基酸变异最高,而F蛋白相对保守。糖基化位点分析发现,该病毒的F蛋白有6个N-糖基化位点,未发现O-糖基化位点,此结果与原型株long株相同;G蛋白N-糖基化位点有6个,O-糖基化位点为82个,而原型株long株有11个N-糖基化位点,15个O-糖基化位点。本研究对2017年河南省漯河市SARI病例中一株HRSVA病毒全基因组序列进行了测定,与世界其他地区报道的HRSVA亚型病毒全基因组序列进行了对比分析,揭示了SARI病例中我国HRSV优势流行ON1基因型病毒全基因组的核苷酸和氨基酸变异特征,以及G蛋白和F蛋白编码区糖基化情况,丰富了我国HRSV基因数据库,也为HRSV的核酸检测方法的建立、疫苗研发和预防性单克隆抗体的评价提供了核苷酸和氨基酸的基础数据。
Human respiratory syncytial virus(HRSV) infection is one of the leading causes of acute respiratory tract infection in young children worldwide. HRSV could be divided into two subgroups, subgroup A(HRSVA) and subgroup B(HRSVB), based on its reactivity with monoclonal antibodies. We obtained one complete genome of HRSVA from hospitalized cases with severe acute respiratory infection(SARI) in Luohe City Henan Province, China, during 2017 using Sanger sequencing process. Sequence editing and alignment, phylogenetic analyses, amino-acid variation and glycosylation were conducted using Sequencher 5.4.5, MEGA 5.05 and BioEdit 7.0.5 software. The HRSVA strain in our study was the ON1 genotype, the most prevalent HRSV subgroup A circulating in China in recent years, based on phylogenetic analysis of the complete genomes and also with 11 individual protein-coding regions. Comparison between RSVAs/Luohe.Henan and 35 representative global strains demonstrated that the homology of nucleotides and amino acids was 92.69%～99.82% and 93.63%～99.67%, respectively; genetic variation was highest in the G-protein encoding region, whereas F protein was highly conserved. Six N-glycosylation sites were predicted for F protein, but no O-glycosylation was found, which was similar to the prototype long strain. In addition, different from the prototype long strain, 82 potential O-linked glycosylation sites and 6 N-glycosylation sites were predicted for G protein. In the present study, the whole-genome sequence was obtained directly from the clinical specimen of a SARI patient in Luohe City, China, during 2017.Compared with HRSVA viruses from other parts of the world, we illustrated the genetic characteristics of nucleotides and amino acids of ON1 genotype of HRSVA subtype virus, the most prevalent virus circulating in China in recent years, and glycosylation for F and G protein-coding regions were also analyzed. We obtained basic genetic data of nucleotides and amino acids, which enriched Chinese HRSV genetic database, will be useful for the development of nucleic acid detection assay, vaccine development and evaluation of prophylactic monoclonal antibodies.
Human respiratory syncytial virus(HRSV) infection is one of the leading causes of acute respiratory tract infection in young children worldwide. HRSV could be divided into two subgroups, subgroup A(HRSVA) and subgroup B(HRSVB), based on its reactivity with monoclonal antibodies. We obtained one complete genome of HRSVA from hospitalized cases with severe acute respiratory infection(SARI) in Luohe City Henan Province, China, during 2017 using Sanger sequencing process. Sequence editing and alignment, phylogenetic analyses, amino-acid variation and glycosylation were conducted using Sequencher 5.4.5, MEGA 5.05 and BioEdit 7.0.5 software. The HRSVA strain in our study was the ON1 genotype, the most prevalent HRSV subgroup A circulating in China in recent years, based on phylogenetic analysis of the complete genomes and also with 11 individual protein-coding regions. Comparison between RSVAs/Luohe.Henan and 35 representative global strains demonstrated that the homology of nucleotides and amino acids was 92.69%～99.82% and 93.63%～99.67%, respectively; genetic variation was highest in the G-protein encoding region, whereas F protein was highly conserved. Six N-glycosylation sites were predicted for F protein, but no O-glycosylation was found, which was similar to the prototype long strain. In addition, different from the prototype long strain, 82 potential O-linked glycosylation sites and 6 N-glycosylation sites were predicted for G protein. In the present study, the whole-genome sequence was obtained directly from the clinical specimen of a SARI patient in Luohe City, China, during 2017.Compared with HRSVA viruses from other parts of the world, we illustrated the genetic characteristics of nucleotides and amino acids of ON1 genotype of HRSVA subtype virus, the most prevalent virus circulating in China in recent years, and glycosylation for F and G protein-coding regions were also analyzed. We obtained basic genetic data of nucleotides and amino acids, which enriched Chinese HRSV genetic database, will be useful for the development of nucleic acid detection assay, vaccine development and evaluation of prophylactic monoclonal antibodies.
引文
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[10]彭质斌,郑建东,姜慧,秦颖,杨娟,余宏杰,冯录召.全国住院严重急性呼吸道感染病例哨点监测阶段性分析[J].疾病监测,2017, 32(1):3-5.
[11]江艳微,华军,吴婧,王丹,李莺,丁云芳,张涛,赵根明.苏州地区5岁以下严重急性呼吸道感染(SARI)住院患儿的病毒病原学和临床特征分析[J].复旦学报(医学版),2013, 40(4):407-412.
[12]吴琼,陈礼娟,黄新泉,欧书腾,刘子菁,范楚平.郴州地区5岁以下住院儿童严重急性呼吸道感染病毒病原学研究[J].医学理论与实践,2017, 30(7):943-945.
[13]谢嘉慧,华亮,钟家禹,张莹莹,陈翊,刘晓敏,连广琬,周珍文,朱冰.广州地区2011-2013年儿童呼吸道合胞病毒流行特征分析[J].中华生物医学工程杂志,2015, 21(3):270-275.
[14] van Elden L J, van Loon A M,van der Beek A, Hendriksen K A, Hoepelman A I, van Kraaij M G, Schipper P, Nijhuis M. Applicability of a real-time quantitative PCR assay for diagnosis of respiratory syncytial virus infection in immunocompromised adults[J]. J Clin Microbiol,2003, 41(9):4378-4381.
[15] Zhang Y, Xu W,Shen K,Xie Z,Sun L,Lu Q, Liu C,Liang G,Beeler J A,Anderson L J. Genetic variability of group A and B human respiratory syncytial viruses isolated from 3 provinces in China[J].Arch Virol,2007, 152(8):1425-1434.
[16] Peret T C, Hall C B, Schnabel K C, Golub J A, Anderson L J. Circulation patterns of genetically distinct group A and B strains of human respiratory syncytial virus in a community[J]. J Gen Virol, 1998, 79(Pt 9):2221-2229.
[17]Agoti C N,Otieno J R,Munywoki P K,Mwihuri A G,Cane P A,Nokes D J,Kellam P,Cotten M. Local evolutionary patterns of human respiratory syncytial virus derived from whole-genome sequencing[J]. J Virol, 2015,89(7):3444-3454.
[18] Do L A,Wilm A,van Doorn H R,Lam H M,Sim S,Sukumaran R,Tran A T,Nguyen B H,Tran T T,Tran Q H. Direct whole-genome deep-sequencing of human respiratory syncytial virus A and B from Vietnamese children identifies distinct patterns of inter-and intrahost evolution[J]. J Gen Virol, 2015, 96(12):3470.
[19] Kholy A A E,Mostafa N A,Ali A A,Soliman M M S, El-Sherbini S A, Ismail R I, Basha N E, Magdy R I, Rifai N E, Hamed D H. The use of multiplex PCR for the diagnosis of viral severe acute respiratory infection in children:a high rate of co-detection during the winter season[J]. Eur J Clin Microbiol, 2016, 35(10):1-7.
[20] Tang J W,Lam T T,Zaraket H,Lipkin W I,Drews S J,Hatchette T F, Heraud J M, Koopmans M P, investigators I. Global epidemiology of non-influenza RNA respiratory viruses:data gaps and a growing need for surveillance[J/OL]. Lancet Infect Dis, 2017, 17(10):e320-e326.
[21] Rey-Jurado E,Kalergis A M. Immunological features of respiratory syncytial virus-caused pneumonia-implications for vaccine design[J]. Int J Mol Sci, 2017, 18(3).pii:E556.
[22] Song J,Wang H,Ng T I,Cui A,Zhu S,Huang Y,Sun L,Yang Z,Yu D,Yu P,Zhang H,Zhang Y,Xu W. Sequence analysis of the fusion protein gene of human respiratory syncytial virus circulating in China from2003 to 2014[J]. Sci Rep, 2018, 8(1):17618.
[23] Chi H,Liu H F,Weng L C,Wang N Y,Chiu N C,Lai M J, Lin Y C, Chiu Y Y, Hsieh W S, Huang L M.Molecular epidemiology and phylodynamics of the human respiratory syncytial virus fusion protein in northern Taiwan[J/OL]. PLoS One, 2013, 8(5):e64012.
[24] Hause A M,Henke D M,Avadhanula V,Shaw C A,Tapia L I,Piedra P A. Sequence variability of the respiratory syncytial virus(RSV)fusion gene among contemporary and historical genotypes of RSV/A and RSV/B[J/OL].PLoS One, 2017, 12(4):e0175792.
[25]宋金华,王慧玲,石晶,崔爱利,杨子峰,孙利伟,于德山,张蕾,张红,张燕.2008-2014年中国8省市流行的人呼吸道合胞病毒A亚型G蛋白编码基因全长序列分析[J].病毒学报,2017(6):821-828. DOI:10.13242/j.cnki.bingduxuebao.003253.
[26] Canedo-Marroquin G,Acevedo-Acevedo O,Rey-Jurado E,Saavedra J M,Lay M K,Bueno S M,Riedel C A,Kalergis A M. Modulation of host immunity by human respiratory syncytial virus virulence factors:a synergic inhibition of both innate and adaptive immunity[J].Front Cell Infect Microbiol, 2017, 7:367.
[2] Griffiths C, Drews S J, Marchant D J. Respiratory syncytial virus:infection,detection, and new options for prevention and treatment[J]. Clin Microbiol Rev, 2017,30(1):277-319.
[3] Linder K A, Malani P N. Respiratory syncytial virus[J].Jama,2017, 317(1):98.
[4] Duvvuri V R,Granados A,Rosenfeld P, Bahl J,Eshaghi A, Gubbay J B. Genetic diversity and evolutionary insights of respiratory syncytial virus A ON1 genotype:global and local transmission dynamics[J]. Sci Rep,2015, 5:14268.
[5] Song J, Zhang Y, Wang H, Shi J, Sun L, Zhang X,Yang Z,Guan W,Zhang H,Yu P,Xie Z,Cui A,Ng T I,Xu W. Emergence of ON1 genotype of human respiratory syncytial virus subgroup A in China between2011 and 2015[J].Sci Rep,2017, 7(1):5501.
[6] Gimferrer L, Andres C, Campins M, Codina M G, Rodrigo J A, Melendo S, Martin M C, Fuentes F, Saiz M R, Esperalba J. Circulation of a novel human respiratory syncytial virus group B genotype during the 2014-2015season in Catalonia(Spain)[J/OL]. Clinical Microbiology&Infection, 2016, 22(1):97.e95-97.e98.
[7] LE A, Delfraro A, Franco D, Castillo J, Castillo M,Moreno B, Lopez-Verges S, Pascale J M, Arbiza J. Genetic variability of human respiratory syncytial virus group B in Panama reveals a novel genotype BA14[J]. J Med Virol, 2017, 89(10):1734.
[8] Song J, Wang H,Shi J, Cui A,Huang Y,Sun L,Xiang X,Ma G,Yu P,Yang Z,Li Q,Ng T I,Zhang Y,Zhang R,Xu W. Emergence of BA9 genotype of human respiratory syncytial virus subgroup B in China from2006 to 2014[J]. Sci Rep, 2017, 7(1):16765.
[9] Pale M,Nacoto A, Tivane A, Nguenha N,Machalele L,Gundane F,Muteto D,Chilundo J,Mavale S,Sema-Baltazar C,Pires G, Augusto O, Mussa T, Gudo E. Respiratory syncytial and influenza viruses in children under 2 years old with severe acute respiratory infection(SARI)in Maputo, 2015[J/OL]. PLoS One, 2017, 12(11):e0186735.
[10]彭质斌,郑建东,姜慧,秦颖,杨娟,余宏杰,冯录召.全国住院严重急性呼吸道感染病例哨点监测阶段性分析[J].疾病监测,2017, 32(1):3-5.
[11]江艳微,华军,吴婧,王丹,李莺,丁云芳,张涛,赵根明.苏州地区5岁以下严重急性呼吸道感染(SARI)住院患儿的病毒病原学和临床特征分析[J].复旦学报(医学版),2013, 40(4):407-412.
[12]吴琼,陈礼娟,黄新泉,欧书腾,刘子菁,范楚平.郴州地区5岁以下住院儿童严重急性呼吸道感染病毒病原学研究[J].医学理论与实践,2017, 30(7):943-945.
[13]谢嘉慧,华亮,钟家禹,张莹莹,陈翊,刘晓敏,连广琬,周珍文,朱冰.广州地区2011-2013年儿童呼吸道合胞病毒流行特征分析[J].中华生物医学工程杂志,2015, 21(3):270-275.
[14] van Elden L J, van Loon A M,van der Beek A, Hendriksen K A, Hoepelman A I, van Kraaij M G, Schipper P, Nijhuis M. Applicability of a real-time quantitative PCR assay for diagnosis of respiratory syncytial virus infection in immunocompromised adults[J]. J Clin Microbiol,2003, 41(9):4378-4381.
[15] Zhang Y, Xu W,Shen K,Xie Z,Sun L,Lu Q, Liu C,Liang G,Beeler J A,Anderson L J. Genetic variability of group A and B human respiratory syncytial viruses isolated from 3 provinces in China[J].Arch Virol,2007, 152(8):1425-1434.
[16] Peret T C, Hall C B, Schnabel K C, Golub J A, Anderson L J. Circulation patterns of genetically distinct group A and B strains of human respiratory syncytial virus in a community[J]. J Gen Virol, 1998, 79(Pt 9):2221-2229.
[17]Agoti C N,Otieno J R,Munywoki P K,Mwihuri A G,Cane P A,Nokes D J,Kellam P,Cotten M. Local evolutionary patterns of human respiratory syncytial virus derived from whole-genome sequencing[J]. J Virol, 2015,89(7):3444-3454.
[18] Do L A,Wilm A,van Doorn H R,Lam H M,Sim S,Sukumaran R,Tran A T,Nguyen B H,Tran T T,Tran Q H. Direct whole-genome deep-sequencing of human respiratory syncytial virus A and B from Vietnamese children identifies distinct patterns of inter-and intrahost evolution[J]. J Gen Virol, 2015, 96(12):3470.
[19] Kholy A A E,Mostafa N A,Ali A A,Soliman M M S, El-Sherbini S A, Ismail R I, Basha N E, Magdy R I, Rifai N E, Hamed D H. The use of multiplex PCR for the diagnosis of viral severe acute respiratory infection in children:a high rate of co-detection during the winter season[J]. Eur J Clin Microbiol, 2016, 35(10):1-7.
[20] Tang J W,Lam T T,Zaraket H,Lipkin W I,Drews S J,Hatchette T F, Heraud J M, Koopmans M P, investigators I. Global epidemiology of non-influenza RNA respiratory viruses:data gaps and a growing need for surveillance[J/OL]. Lancet Infect Dis, 2017, 17(10):e320-e326.
[21] Rey-Jurado E,Kalergis A M. Immunological features of respiratory syncytial virus-caused pneumonia-implications for vaccine design[J]. Int J Mol Sci, 2017, 18(3).pii:E556.
[22] Song J,Wang H,Ng T I,Cui A,Zhu S,Huang Y,Sun L,Yang Z,Yu D,Yu P,Zhang H,Zhang Y,Xu W. Sequence analysis of the fusion protein gene of human respiratory syncytial virus circulating in China from2003 to 2014[J]. Sci Rep, 2018, 8(1):17618.
[23] Chi H,Liu H F,Weng L C,Wang N Y,Chiu N C,Lai M J, Lin Y C, Chiu Y Y, Hsieh W S, Huang L M.Molecular epidemiology and phylodynamics of the human respiratory syncytial virus fusion protein in northern Taiwan[J/OL]. PLoS One, 2013, 8(5):e64012.
[24] Hause A M,Henke D M,Avadhanula V,Shaw C A,Tapia L I,Piedra P A. Sequence variability of the respiratory syncytial virus(RSV)fusion gene among contemporary and historical genotypes of RSV/A and RSV/B[J/OL].PLoS One, 2017, 12(4):e0175792.
[25]宋金华,王慧玲,石晶,崔爱利,杨子峰,孙利伟,于德山,张蕾,张红,张燕.2008-2014年中国8省市流行的人呼吸道合胞病毒A亚型G蛋白编码基因全长序列分析[J].病毒学报,2017(6):821-828. DOI:10.13242/j.cnki.bingduxuebao.003253.
[26] Canedo-Marroquin G,Acevedo-Acevedo O,Rey-Jurado E,Saavedra J M,Lay M K,Bueno S M,Riedel C A,Kalergis A M. Modulation of host immunity by human respiratory syncytial virus virulence factors:a synergic inhibition of both innate and adaptive immunity[J].Front Cell Infect Microbiol, 2017, 7:367.