孟凡红,于巍,张嵬,杨利伟,何玉君,高强,黄金凤,尹卫东
MENG Fan-hong,YU Wei,ZHANG Wei,YANG Li-wei,HE Yu-jun,GAO Qiang,HUANG Jin-feng,YIN Wei-dong

• 1:科兴(大连)疫苗技术有限公司
• 2:北京科兴生物制品有限公司

摘要(Abstract)：

目的研究水痘-带状疱疹病毒(varicella-zoster virus,VZV)Oka株在人二倍体细胞株SV-1上的遗传稳定性,并对不同代次毒株进行全基因测序及差异位点分析。方法将VZV Oka株在SV-1细胞株上连续传至48代,检测病毒滴度。对VZV Oka株33、35、38、39及48代病毒进行全基因测序,应用DNASTAR.Lasergene.v 7.1软件进行数据的拼接及序列分析。结果 VZV Oka株在SV-1细胞上连续传代获得的33~48代病毒滴度在4.000~4.625 lg CCID50/ml之间,病毒滴度的算数平均值为4.292 lg CCID50/ml;VZV Oka株基因组全长125 114 bp,GC含量46%;33、35、38、39、48代病毒的核苷酸序列同源性为99.99%,这5代病毒与标准疫苗株V-Oka的核苷酸序列同源性为99.96%;VZV Oka株在MRC-5、SV-1细胞上制备的38代病毒全基因序列有1个差异位点;本研究中的各代次病毒基因序列高度同源,与Varivax和Varilrix疫苗株、野毒株Dumas及亲本株p-Oka相比,与疫苗株V-Oka的同源性最高。结论 VZV Oka株在SV-1细胞上的适应性及遗传稳定性均良好。
Objective To investigate the genetic stability of varicella-zoster virus(VZV)Oka strain during subculture in human diploid cell SV-1 strain, and analyze the complete gene sequences and differential sites of Oka strain of various passages. Methods Oka strain was subcultured in SV-1 cells to passage 48, and determined for virus titer. The complete genes of passages 33, 35, 38, 39 and 48 were sequenced, of which the data were spliced and analyzed by using DNASTAR.Lasergene. v 7. 1 software. Results The virus titers of Oka strain of passages 33 ~ 48 were 4. 000 ~ 4. 625 lg CCID50/ ml,with an arithmetic mean of 4. 292 lg CCID50/ ml. The full-length of genome of Oka strain was 125 114 bp, while the GC content was 46%. The homology of nucleotide sequences of Oka strain of passages 33, 35, 38, 39 and 48 was 99. 99%,while that of the five passages to standard vaccine strain V-Oka was 99. 96%. One differential site was observed in the complete gene sequence of Oka strain subcultured to passage 38 in MRC-5 and SV-1 cells. High homologies were observed in the gene sequences of Oka strain of various passages. However, as compared with those of vaccine strains Vari-vax and Varilrix, wild strain Dumas and parent strain p-Oka, the homology of gene sequence of Oka strain to that of vac-cine strain V-Oka strain was high. Conclusion VZV Oka strain showed high adaptability and genetic stability in SV-1 cells.

关键词(KeyWords)： 水痘-带状疱疹病毒Oka株;遗传稳定性;差异位点
Varicella-zoster virus Oka strain;Genetic stability;Differential site

Abstract:

Keywords:

基金项目(Foundation):

作者(Author): 孟凡红,于巍,张嵬,杨利伟,何玉君,高强,黄金凤,尹卫东
MENG Fan-hong,YU Wei,ZHANG Wei,YANG Li-wei,HE Yu-jun,GAO Qiang,HUANG Jin-feng,YIN Wei-dong

DOI: 10.13200/j.cnki.cjb.000768

参考文献(References)：

• [1]Takahashi M,Otsuka T,Okuno Y,et al.Live vaccine used to prevent the spread of varicella in children in hospital[J].Lancet,1974,2(7892):1288-1290.
• [2]Weibel R,Neff BJ,Kuter BJ,et al.Live attenuated varicella virus vaccine:efficacy trial in healthy children[J].N Engl J Med,1984,310(22):1409-1415.
• [3]Varis T,Vesikari T.Efficacy of high-titer live attenuated varicella vaccine in healthy young children[J].J Infect Dis,1996,174(Suppl 3):S330-S334.
• [4]Gershon AA,Katz SL.Perspective on live varicella vaccine[J].J Infect Dis,2008,197(Suppl 2):S242-S245.
• [5]Lopez AS,Guris D,Zimmerman L,et al.One dose of varicella vaccine does not prevent school outbreaks:is it time for a second dose[J].Pediatrics,2006,117(6):e1070-e1077.
• [6]Wang GQ,Peng PX,Wang CY,et al.Isolation of varicellazoster virus from skin vesicle fluids[J].Chin J Virol,1986,2(4):303-305.(in Chinese)王桂秋,彭佩霞,王常玉,等.水痘-带状疱疹病毒分离和鉴定[J].病毒学报,1986,2(4):303-305.
• [7]Tillieux SL,Halsey WS,Thomas ES,et al.Complete DNA sequences of two Oka strain varicella-zoster virus genomes[J].J Virol,2008,82(22):11023-11044.
• [8]Sauerbrei A,Rubtcova E,Wutzler P,et al.Genetic profile of an Oka varicella vaccine virus variant isolated from an infant with zoster[J].J Clin Microbol,2004,42(12):5604-5608.
• [9]Schmidt M,Kress M,Heimemann S,et al.Varicella-zoster virus isolates,but not the vaccine strain OKA,induce sensitivity to alpha-1 and beta-l adrenergic stimulation of sensory neurones in culture[J].J Med Virol,2003,70(Suppl l):S82-S89.
• [10]Lu ZL,Wang GQ,Cheng B,et al.Analysis of point mutations in ORF62 of varicella-zoster virus(VZV)vaccine Oka strain and the transactivities of ORF62-encoding IE62 on VZV promoters[J].Chin J Microbiol Immunol,2009,29(10):953-957.(in Chinese)卢珍玲,王官清,程波,等.水痘-带状疱疹病毒疫苗株ORF62碱基突变及ORF62编码IE62反式激活力的分析[J].中华微生物学和免疫学杂志,2009,29(10):953-957.
• [11]Tyler SD,Peters GA,Grose C,et al.Genomic cartography of varicella-zoster virus:a complete genome-based analysis of strain variability with implications for attenuation and phenotypic differences[J].Virology,2007,359(2):447-458.
• [12]Oliver SL,Sommer MH,Reichelt M,et al.Mutagenesis of varicella-zoster virus glycoprotein I(g I)identifies a cysteine residue critical for g E/g I heterodimer formation,g I structure,and virulence in skin cells[J].J Virol,2011,85(9):4095-4110.
• [13]Berarducci B,Rajamani J,Zerboni L,et al.Functions of the unique N-terminal region of glycoprotein E in the pathogenesis of varicella-zoster virus infection[J].Proc Natl Acad Sci USA,107(1):282-287.