曲梅, 黄芳, 刘园, 窦相峰, 刘桂荣, 严寒秋, 高志勇, 王全意. 应用TaqMan实时荧光-聚合酶链反应方法快速检测粪便标本空肠弯曲菌的研究[J]. 疾病监测, 2009, 24(5): 343-345. DOI: 10.3784/j.issn.1003-9961.2009.05.011
引用本文: 曲梅, 黄芳, 刘园, 窦相峰, 刘桂荣, 严寒秋, 高志勇, 王全意. 应用TaqMan实时荧光-聚合酶链反应方法快速检测粪便标本空肠弯曲菌的研究[J]. 疾病监测, 2009, 24(5): 343-345. DOI: 10.3784/j.issn.1003-9961.2009.05.011
QU Mei, HUANG Fang, LIU Yuan, DOU Xiang-feng, LIU Gui-rong, YAN Han-qiu, GAO Zhi-yong, WANG Quan-yi. Application of real-time PCR TaqMan assay for rapid detection of Campylobacter jejuni from stool samples[J]. Disease Surveillance, 2009, 24(5): 343-345. DOI: 10.3784/j.issn.1003-9961.2009.05.011
Citation: QU Mei, HUANG Fang, LIU Yuan, DOU Xiang-feng, LIU Gui-rong, YAN Han-qiu, GAO Zhi-yong, WANG Quan-yi. Application of real-time PCR TaqMan assay for rapid detection of Campylobacter jejuni from stool samples[J]. Disease Surveillance, 2009, 24(5): 343-345. DOI: 10.3784/j.issn.1003-9961.2009.05.011

应用TaqMan实时荧光-聚合酶链反应方法快速检测粪便标本空肠弯曲菌的研究

Application of real-time PCR TaqMan assay for rapid detection of Campylobacter jejuni from stool samples

  • 摘要: 目的建立空肠弯曲菌TaqMan实时荧光-PCR方法,用于粪便标本的直接检测。方法根据空肠弯曲菌特异性基因hipO和mapA分别设计引物和探针,在对2组引物和探针进行灵敏度、特异性和重复性评价的基础上,对45例临床腹泻患者粪便标本提取DNA之后,荧光PCR检测,同时进行分离培养。 结果两组引物和探针能准确检测空肠弯曲菌菌株2株,检测限可达到10~20 cfu/ml,并与其他肠道致病菌无交叉反应。检测45份腹泻病例粪便标本,该方法检测到3份为阳性,同时进行的传统培养方法仅从该3份标本中的两份中分离到空肠弯曲菌。 结论本研究建立的TaqMan荧光PCR检测粪便标本中所携带的空肠弯曲菌灵敏度高,特异性好,能够提高粪便中空肠弯曲菌的阳性检出率和缩短检测时限。

     

    Abstract: ObjectiveTo establish real-time PCR TaqMan assay for the direct detection of ICampylobacter jejuni/I from stool samples. MethodsTwo pairs of specific primers and probes were designed by IhipO/I and ImapA/I genes respectively. On the basis of evaluation of sensitivity, specificity and reproducibility of two systems, real-time PCR was performed to identify DNA extracted from 45 stool samples of clinical diarrhea patients and conventional culture was conducted too. ResultsIC. jujeni/I strain could be detected by the 2 pairs of primers and probes with the detection limit of 10-20 cfu/ml. There was no cross-reactions with other enteric pathogens. Of 45 stool sample, 3 were positive by real-time PCR, and 2 of which were positive by conventional pure culture. ConclusionReal-time PCR TaqMan assay in this study was highly sensitive and specific, which could rapidly detect IC. jejuni/I from stool samples with high positive rate and in a shorter time.

     

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