CAJ | 학술논문

目的对柯萨奇病毒A组16型(coxsackievirus A16,CA16)毒株进行分离、鉴定,并分析其生物学特性,为CA16疫苗候选毒株的筛选奠定基础.方法从手足口病患者的咽拭子或疱疹液样本中分离CA16毒株,经噬斑纯化后,用逆转录-聚合酶链反应进行病毒鉴定.将病毒连续传代后,对其VP1基因进行序列测定,分析VP1基因和氨基酸序列的同源性和遗传稳定性.结果通过Vero细胞适应传代和噬斑纯化,获得遗传稳定的CA16毒株,感染Vero细胞后引起典型的细胞病变.病毒分离株经CA16特异性引物扩增后可见208 bp的特异性条带,经VP1基因测序进一步确定这些分离株为CA16.同源性和系统进化树分析表明,各毒株与shzh04-J31株的同源性最高,为92.4％～96.1％,均属于C基因亚型.病毒分离株在Vero细胞上连续传12代后,仅1株病毒的1个氨基酸发生改变.结论成功分离到CA16毒株,其生物学特性稳定,可用于CA 16疫苗的研发.
목적 대가살기병독A조16형(coxsackievirus A16,CA16)독주진행분리、감정,병분석기생물학특성,위CA16역묘후선독주적사선전정기출.방법 종수족구병환자적인식자혹포진액양본중분리CA16독주,경서반순화후,용역전록-취합매련반응진행병독감정.장병독련속전대후,대기VP1기인진행서렬측정,분석VP1기인화안기산서렬적동원성화유전은정성.결과 통과Vero세포괄응전대화서반순화,획득유전은정적CA16독주,감염Vero세포후인기전형적세포병변.병독분리주경CA16특이성인물확증후가견208 bp적특이성조대,경VP1기인측서진일보학정저사분리주위CA16.동원성화계통진화수분석표명,각독주여shzh04-J31주적동원성최고,위92.4％～96.1％,균속우C기인아형.병독분리주재Vero세포상련속전12대후,부1주병독적1개안기산발생개변.결론 성공분리도CA16독주,기생물학특성은정,가용우CA 16역묘적연발.
Objective To isolate and identify coxsackievirus A16 (CA16) strains,analyze their biological characteristics,and lay a foundation for screen of candidate CA16 vaccine strains.Methods The CA16 strains were isolated from throat swab or herpes liquid samples of patients with hand,foot and mouth disease,and then determined for specificity by reverse transcription (RT)-PCR after plaque cloning.After continuous subculture,the homology and genetic stability of the isolated strains was evaluated by analysis of VP 1 gene and amino acid sequences.Results Genetically stable CA16 strains were obtained by subculture and plaque cloning in Vero cells.The strains caused a typical cytopathic effect on Vero cells.The product of RT-PCR with CA16-specific primers showed a specific band at 208 bp and VP1 gene sequencing further confirmed that the isolated strains were CA16.The strains were identified as C subtype of CA16 by homology and phylogenetic analysis.The homology of VP1 gene between the isolated strains and shzh04-J31 strain was 92.4％-96.1％.After subculture in Vero cells for 12 passages,only one amino acid variation was found in one strain.Conclusion The CA16 strains with stable biological characteristics are isolated successfully and can be used for development of CA16 vaccine.