摘要
采集急性期病人的咽拭子或漱口液,用Vero、Vero E6、MDCK、Hela、Hep-2等传代细胞,人胚肺二倍体细胞(HEL)和人胚肺(HP)细胞分离培养严重急性呼吸系统综合症(SARS)的病原体。结果用Vero、Vero E6、MDCK和HP细胞从标本中分离到一株病毒。间接免疫荧光试验发现,恢复期病人血清可与所分离的病毒起反应,在胞膜和胞浆中出现翠绿色荧光;中和试验结果表明,恢复期病人血清能中和病毒对细胞的致细胞病变作用:电镜下可观察到冠状病毒样颗粒:RT-PCR法可扩增到冠状病毒特异性基因片段,且其核苷酸序列与国内外发表的SARS冠状病毒(SARS-Cov)相应的基因序列相符,同源性达到100％。从传染性非典型肺炎病人的漱口液中分离到SARS冠状病毒,这种病毒与传染性非典型肺炎密切相关。
To isolate and identify the etiologic agent of severe acute respiratory syndrome(SARS), the throat swabs and gargle of patients with SARS were collected. We inoculated the specimens onto a number of cell lines including Vero, Vero E6, MDCK, Hela, Hep-2, HP and HEL cells. The isolates were identified by thin-section electron-microscopy, indirect-immunofluorescence, neutralization test, RT-PCR and nucleotide sequence analysis. Using cell culture techniques, a coronavirus was isolated from patient with SARS. Four cell lines, Vero, Vero E6, HP and MDCK cells, showed cytopathic effect. The infected cells showed strong cytoplasmic and membranous staining with a convalescent-phase serum from the patients with SARS in an indirect-immunofluorescence staining. The cytopathic effect of the viruses clould been neutralized by convalescent serum. Under electron-microscopy, a large number of coronavirus-like particles were obsewed in the infected cells. With specific primers, the specific cDNA fragments were amplified by reverse transcription-polymerase chain reaction(RT-PCR) from infected cells and the nucleotide sequence analysis showed that homology of the amplified fragments to the SARS-coronavirus (SARS-CoV) previously reported was 100%. A SARS-CoV was isolated from patients with SARS. This virus was closely associated with SARS.
出处
《中国病毒学》
CSCD
2003年第6期544-547,共4页
Virologica Sinica
基金
国家自然科学基金(30340013)
广东省科技攻关项目
关键词
传染性非典型肺炎
SARS冠状病毒
分离
培养
鉴定
序列分析
Severe acute respiratory syndrome(SARS)
SARS-CoV
Isolation and identification
Nucleotide sequence analysis.
