巢式聚合酶链反应快速检测血液标本中伤寒沙门菌的实验研究

Study of the nested PCR detection salmonella typhi in the blood specimens

目的建立一种检测血液中伤寒沙门菌的方法。方法根据伤寒沙门菌特异鞭毛基因设计两对引物 ,通过巢式聚合酶链反应 (PCR)扩增伤寒沙门菌DNA片断 ,扩增产物通过凝胶电泳进行分析。结果用伤寒沙门菌DNA系列稀释液进行试验 ,巢式PCR能够检出 10个伤寒沙门菌。 36份培养阳性标本 ,33份PCR阳性 ;6份培养阴性但临床高度可疑的伤寒病人标本PCR阳性 ;10份其它原因引起发热的临床标本均为阴性。结论巢式PCR能够快速、特异、准确地检出血液中的伤寒沙门菌。

Objective To develop nested PCR method for the detection of Salmonella typhi in the blood specimens from patients with typhoid fever. Methods Two pairs of oligonucleotide primers were designed to amplify a 345bp fragment of the flagellin gene of S. Typhi. Amplified products were analyzed by agarose gel electroplioresis, Results The nested PCR could detect 10 organisms of S. typhi as 10 determined by serial dilutions of DNA from S. typhi. The peripheral mononuclear cells from 33/36 patients with typhoid fever confirmed by blood culture and from 6 patients with suspected typhoid fever on the clinical features were positive for DNA fragment of the flagellin gene of S. typhi, whereas 10 blood specimens of patients with other febric diseases were negative Conclusion The nested PCR could detect rapidly and accurately S. typhi in the blood specimens