严重急性呼吸综合征实验室检测

Laboratory detection on severe acute respiratory syndrome

目的为严重急性呼吸综合征(SARS)病例的诊断提供科学依据.方法连续、系统地采集2003年12月至2004年1月广东省新发4例SARS怀疑病例不同发病时间的各类标本,采用酶联免疫吸附测定(ELISA)、免疫荧光测定(IFA)、中和试验等血清学检测技术和逆转录聚合酶链反应(RT-PCR)、荧光定量聚合酶链反应等分子生物学技术检测采集的标本,并对检测结果进行分析.结果血清学检测结果显示,4例病例在发病后6～8d,均可检测到SARS-CoV IgM和IgG抗体;前3例病例SARS-CoV抗体有4倍或4倍以上增长,第4例病例SARS-CoV IgG抗体虽无4倍增长,但却有明显的抗体阳转过程.采用ELISA、IFA和中和试验的检测结果基本一致.用荧光定量PCR检测咽拭等标本的SARS-CoV核酸,结果只有病例1的3份咽拭标本阳性,其他标本均阴性.对阳性标本,采用RT-PCR扩增出SARS-CoV的M、N、S基因并测序,S基因序列和种系发生分析提示,该病毒与从广东省果子狸中分离到的SARS毒株最接近.未能从这4例SARS病例的任何标本中分离到病毒.结论检测结果证明,该4例病例报告可以确认为SARS实验室确诊病例.

Objective To provide scientific evidence to identify and confirm severe acute respiratory syndrome （SARS） by laboratory detection. Methods Multiple clinical specimens were collected serially and systematically from the 4 suspected SARS patients, which occurred between Dec. 2003 to Jan. 2004 in Guangdong Province. The samples were tested by serologic and molecular methods. Results IgM or IgG antibodies against SARS-CoV were detectable after 6-8 days of the onset in four patients. The four-fold or greater rising in antibodies was clearly detected in three of the four patients, while the fourth patient＇s seroconversion was from negative to positive. The results analysed by enzyme-linked immunosorbent assay（ELISA）, immunoflourescence assay （IFA）, and neutralization test were highly correlated. SARS-CoV RNA was just detected in 3 throat swab specimens from case 1 by real-time PCR. M, N and S genes were amplified by reverse transcriptase polymerase chain reaction （RT-PCR） from the positive samples. Sequencing results showed that they were SARS-CoV gene segments, and most closely matched SARS-CoV gene sequences were isolated from civet cats in Guangdong Province. Nevertheless, SARS-CoV was not isolated from any samples of the 4 patients. Conclusion Based on these results, the 4 reported cases were laboratorily confirmed as SARS cases.