贵州独山、兴义不明原因发热病人登革病毒的分离和鉴定的初步研究  

Isolation and identification of dengue virus in the sera of patients with unknown fever in Dushan and Xingyi areas of Guizhou province

作　　者：周永兵[1] 左丽[1] 刘伟[1] 谢庭华 何成友 王定明[4] 

机构地区：[1]贵阳医学院免疫学教研室,550004 [2]贵州省独山县疾病预防控制中心 [3]贵州省兴义市疾病预防控制中心 [4]贵州省疾病预防控制中心

出　　处：《中华微生物学和免疫学杂志》2008年第3期221-225,共5页Chinese Journal of Microbiology and Immunology

基　　金：国家自然科学基金资助项目（30360101）;贵州省高层次人才基金[2005（11）];贵州省科学技术基金[（2006）2083]

摘　　要：目的对贵州独山、兴义两地夏秋季不明原因发热病人血清进行登革病毒（DEN）分离及鉴定，从病原学角度证实贵州省人群DEN的感染情况。方法于2005年6至10月份收集贵州独山县、兴义市两地不明原因发热病人血清356份，并接种于生长良好的单层C6／36细胞盲传3代，观察细胞病变，用抗DEN1～4型单克隆抗体通过间接免疫荧光法进行型别鉴定；用DENNS1基因区特异性通用引物进行RT-PCR检测分离的DEN毒株核酸，经序列测定并作系统发生树分析。结果3份病人血清标本可使C6／36发生细胞病变，用单克隆抗体、RT-PCR扩增和序列测定，鉴定为DEN2；系统发生树分析证明，分离的病毒与DEN2-43、DEN2-44株系统进化关系最近。结论贵州省独山、兴义两地人群中存在DEN感染。Objective To isolate and identify the Dengue virus （DEN） from the sera of patients with unknown fever in summer and autumh in Dushan and Xingyi areas of Guizhou province, China. Methods From June 2005 to October 2005, 356 blood samples were collected from patients with unknown fever in Dushan and Xingyi areas of Guizhou province. The serum samples were inoculated on the C6/36 cell monolayers. After three blind passages, the cytopathic effect （CPE） was observed. Identification of DEN antigen was carried out by indirect immunofluorescence assay （IFA） with the monoclonal antibody （McAb） against DEN1-4 virus. The total RNA was extracted from the serum and tested by RT-PCR with the universal primer for DEN NS region. And determination of the RNA sequence was performed, and phylogenetic analysis was carried out. Results Three serum samples caused CPE and were proved as DEN2 positive by IFA, RT-PCR and senquence determination. The phylogenetic tree analysis showed that the isolated virus strains had the closest relations with the systemic evolution of the strains DEN2-43 and DEN2-44. Conclusion DEN infections exist in the population of Dushan and Xingyi of Guizhou, China.

关 键 词：登革病毒 病毒分离 发热病人 血清 

分 类 号：R686[医药卫生—骨科学]