伤寒、甲、乙、丙型副伤寒沙门菌多重荧光PCR方法的评估及临床应用

Assessment and Application of a Multiplex Real-time PCR Assay for Rapid Detection of S typhi,S. Paratyphi A,S. Paratyphi B and S. Paratyphi C

目的建立和评估多重荧光PCR方法快速鉴定伤寒、甲、乙、丙型副伤寒沙门菌的方法,并应用于临床血培养样品的检测。方法收集留取医院的可疑伤寒患者的血培养标本,用多重实时荧光PCR法和传统分离培养法同时进行分离鉴定,分析比较双盲实验结果,评价多重实时荧光PCR方法的灵敏度、特异度等检测性能指标。结果共检测临床样本538例,多重荧光PCR法检出阳性218例,比传统培养法多检出6例;阴性320例,与传统培养法一致。以传统检测方法为金标准,多重荧光PCR方法的灵敏度为100%,特异度为98.2%。结论建立的多重荧光PCR检测方法操作简便,可快速、特异、灵敏地检测出伤寒、甲、乙、丙型副伤寒沙门菌,可以应用于临床标本的早期快速分型诊断,提升重大传染病的应急处置能力和监测能力。

Objective To establish and evaluate a rapid multiplex real-time PCR（m-rt-PCR） assay for the simultaneous detection of S typhi, S. paratyphi A, S. paratyphi B and S. paratyphi C. The method could be applied to the rapid diagnosis of clinical blood samples. Methods Suspected blood cultures were collected to simultaneously detect S. Typhi, S. Paratyphi A, B and C by m-rt-PCR assay and standard culture methods. The results of the m-rt-PCR assay were compared with those obtained from traditional culture based methods using a 2×2 table to estimate indices of sensitivity and specificity. Results 538 unknown clinical samples were analyzed by culture, 212 were culture positive. Multiplex real-time PCR detected 218 positive samples. In comparison with culture method, clinical validation showed 100% sensitivity and 98.2% specificity. Conclusion The real -time PCR assay was simple, rapid, sensitive and specific. It could be applied to the rapid diagnosis of S. paratyphi A, S. paratyphi B, S. paratyphi C and S typhi. The established assay should increase the speed of detection and differentiation of typhoid fever causing organisms in a diagnostic setting.