CAJ | 학술논문

目的对深圳市2005年从登革热患者急性期血液中分离到的1株登革病毒进行型别鉴定,从分子水平分析分离株的生物学特征,追踪其可能的地域来源.方法用C6/36细胞培养增殖病毒株SZ0524,收集病毒液.用逆转录-半套式PCR(RT-semi-nested-PCR)方法和荧光PCR方法对其进行型别鉴定.扩增病毒E基因后进行序列测定,并与不同国家和地区的登革病毒株进行同源性比较和进化树分析.结果深圳市登革病毒分离株用4型特异性引物扩增出392bp的特异性条带,荧光PCR进一步证实了分离的病毒株为登革4型病毒.SZ0524与登革病毒4型国际标准株H241株在E基因上核苷酸同源性为99.7%,而与登革病毒1、2、3型国际标准株HAWAII、NGC、H87同源性分别为57.0%、59.2%和56.2%.基因进化树显示SZ0524株与D4-73NIID株和D4-61NIID株亲缘关系最近,其次为H241,在进化树的同一分支上,属基因Ⅰ亚型.结论从分子水平证明从深圳市登革热患者血清中分离到的毒株确为DEN-4型病毒.结合流行病学调查资料,证实此病例为输入性感染病例,该毒株最有可能来源于东南亚一带.
목적 대심수시2005년종등혁열환자급성기혈액중분리도적1주등혁병독진행형별감정,종분자수평분석분리주적생물학특정,추종기가능적지역래원.방법 용C6/36세포배양증식병독주SZ0524,수집병독액.용역전록-반투식PCR(RT-semi-nested-PCR)방법화형광PCR방법대기진행형별감정.확증병독E기인후진행서렬측정,병여불동국가화지구적등혁병독주진행동원성비교화진화수분석.결과 심수시등혁병독분리주용4형특이성인물확증출392bp적특이성조대,형광PCR진일보증실료분리적병독주위등혁4형병독.SZ0524여등혁병독4형국제표준주H241주재E기인상핵감산동원성위99.7%,이여등혁병독1、2、3형국제표준주HAWAII、NGC、H87동원성분별위57.0%、59.2%화56.2%.기인진화수현시SZ0524주여D4-73NIID주화D4-61NIID주친연관계최근,기차위H241,재진화수적동일분지상,속기인Ⅰ아형.결론 종분자수평증명종심수시등혁열환자혈청중분리도적독주학위DEN-4형병독.결합류행병학조사자료,증실차병례위수입성감염병례,해독주최유가능래원우동남아일대.
To identify the genotype and analyze the molecular characteristics of dengue virus strain SZ0524 isolated from serum samples of patients with early stage of dengue fever in Shenzhen in 2005 so as to explore its possible origin. The C6/36 cell line was cultivated with virus strain SZ0524 and its suspension was harvested. The type of isolated virus strain was determined by RT-semi-nested PCR and fluorescent PCR. E gene of isolated virus strain was amplified by RT-PCR and sequenced. Homology and phylogenetic tree of E gene of this dengue virus with the strains isolated from other areas were constructed. This SZ0524 strain was further identified by fluorescent PCR, and confirmed to be the type 4 virus after obtaining the 392bp band with type 4 specific primers. The homology of nucleotide sequence of E gene of SZ0524 strain with the standard type 4 dengue virus H241 strain were 99.7%, but the homology with the standard dengue virus 1,2,3 in the same fragment were 57.0%, 59.2% and 56.2% respectively. Analysis of the phylogenetic tree indicated that SZ0524 was more close to D4-73NIID and D4-61NIID strain, next to H241 strain, and they lied in the same branch of phylogenetic tree. The isolated dengue virus type 4 belonged to genotype Ⅰand the SZ0524 strain was proved to be dengue virus type 4 in the molecular level. Combined with epidemiology information, it is suggested that this case can be classified as an imported case and the SZ0524 strain may be transferred from the southeast asian region.