聚合酶链反应检测O1群霍乱弧菌肠毒素A亚单位基因

Rapid detectiou of cbolera toxin gene of Vibrio cholerae strain O1 by polymerase chain reaction

为了加强对霍乱的防治，探索对霍乱弧菌的快速和敏感的检测方法，建立了一种快速标本处理和聚合酶链反应（ＰＣＲ）扩增Ｏ１群霍乱弧菌肠毒素Ａ亚单位（ｃｔｘＡ）基因区的诊断方法。结果表明：在对一组经常规培养、生化鉴定法诊断为霍乱的２６份腹泻患者粪便标本检测后，ＰＣＲ法有２４株检测到ｃｔｘＡ基因，培养阳性而ＰＣＲ阴性的２株为非Ｏ１群菌株。虽然现有的Ｏ１群血清学诊断方法无法检测新血清型Ｏ１３９，但研究表明Ｏ１３９与Ｏ１群霍乱弧菌肠毒素基因的核苷酸序列相同，本文引物序列也能扩增Ｏ１３９。结果提示：本方法具有简便、快速、特异和不需培养等特点，宜于临床应用。

Cholera toxin(CT)is the principal factor causing the large volume of intestinal fluidsecretion which is characteristic of cholera.The identification of cholera toxin is an important step inthe diagnosis of cholera.We established a diagnostic procedure employing rapid DNA templatespreparation and polymerase chain reaction(PCR)to detect the cholera toxin subunit A(ctxA)geneof Vibrio cholerae O1.Among the total 26 strains of Vibrio cholerap isolated from stooi samples ofpatients with acute secretory diarrhea in the summer of 1994,Shanghai by conventional biochemicalanalysis and enzyme-linked immunosorbent assay,24 were positive by PCR.The two negativestrains were cuIture-positive,but they did not belong to strain O1.Since the nucleotide sequence ofthe gene encoding the A and B subunits of CT of serotype O139 was identical to that of serotype O1,it could be amplified by the primer for strain O1 as well,while it could not be distinguished by com-mon serological diagnostic techniques.The results demonstrated that the detection of ctxA by PCRwas a rapid,sensitive,specific and culture-independent method for the diagnosis of cholera.