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首页> 外文期刊>International Journal of Biomedical Science >A New Lab Developed Real Time PCR Assay for Direct Detection of C. Difficle from Stool Sample without DNA Extraction
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A New Lab Developed Real Time PCR Assay for Direct Detection of C. Difficle from Stool Sample without DNA Extraction

机译：一个新实验室开发的实时PCR检测方法，无需DNA提取即可直接检测粪便样品中的艰难梭菌

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Clostridium difficile is a major cause of nosocomial antibiotic-associated infectious diarrhea and pseudomembranous colitis. Detection of C. difficile by anaerobic bacterial culture and/or cytotoxicity assays has been largely replaced by rapid enzyme immunoassays (EIA). However, due to the lack of sensitivity of stool EIA, we developed a multiplex real-time PCR assay targeting the C. difficile toxin genes tcdB. stool samples from hospitalized pediatric patients suspected of having C. difficile-associated disease were prospectively collected. Three testing modalities were evaluated, including enriched culture, cepheid Xpert and real-time Pcr (tcdB) on stool samples performed with tcdB gene-specific primers and hydrolysis probes. A total of 150 de-identified clinical specimen were analyzed. The sensitivities of stool real-time Pcr were 95% against cepheid Xpert C. difficile and 93% against enriched culture respectively, with a specificity of 97% and 94%. The lower limit of detection of the stool real-time PCR was 0.5 cFU/ml of per reaction for tcdB. Direct detection of C. difficile toxin genes in stool samples by real-time Pcr showed performance comparable to enriched culture. Real-time PCR of DNA from stool samples is a rapid and cost-effective diagnostic modality for patients that should facilitate appropriate patient management.
机译：艰难梭菌是引起医院内抗生素相关感染性腹泻和假膜性结肠炎的主要原因。通过厌氧细菌培养和/或细胞毒性测定法检测艰难梭菌已被快速酶免疫测定法（EIA）取代。然而，由于缺乏粪便EIA的敏感性，我们开发了针对艰难梭菌毒素基因tcdB的多重实时PCR分析。前瞻性地收集了疑似艰难梭菌相关疾病住院儿科患者的粪便样本。评估了三种测试方式，包括对富含tcdB基因特异性引物和水解探针的粪便样品进行的富集培养，造父变星Xpert和实时Pcr（tcdB）。总共分析了150个身份不明的临床标本。粪便实时Pcr对造父变种艰难梭菌的敏感性分别为95％和93％，对富集培养的敏感性为97％和94％。粪便实时PCR检测的下限为tcdB每反应0.5 cFU / ml。实时Pcr直接检测粪便样品中的艰难梭菌毒素基因显示出与浓缩培养相当的性能。粪便样本中DNA的实时PCR是对患者的一种快速且具有成本效益的诊断方法，应有助于适当的患者管理。

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