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首页> 外文期刊>Journal of Clinical Microbiology >Multiplex PCR Targeting tpi (Triose Phosphate Isomerase), tcdA (Toxin A), and tcdB (Toxin B) Genes for Toxigenic Culture of Clostridium difficile
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Multiplex PCR Targeting tpi (Triose Phosphate Isomerase), tcdA (Toxin A), and tcdB (Toxin B) Genes for Toxigenic Culture of Clostridium difficile

机译：靶向tpi（磷酸三磷酸酯异构酶），tcdA（毒素A）和tcdB（毒素B）基因的多重PCR用于艰难梭菌的产毒培养

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A multiplex PCR toxigenic culture approach was designed for simultaneous identification and toxigenic type characterization of Clostridium difficile isolates. Three pairs of primers were designed for the amplification of (i) a species-specific internal fragment of the tpi (triose phosphate isomerase) gene, (ii) an internal fragment of the tcdB (toxin B) gene, and (iii) an internal fragment of the tcdA (toxin A) gene allowing distinction between toxin A-positive, toxin B-positive (A+B+) strains and toxin A-negative, toxin B-positive (A?B+) variant strains. The reliability of the multiplex PCR was established by using a panel of 72 C. difficile strains including A+B+, A?B?, and A?B+ toxigenic types and 11 other Clostridium species type strains. The multiplex PCR assay was then included in a toxigenic culture approach for the detection, identification, and toxigenic type characterization of C. difficile in 1,343 consecutive human and animal stool samples. Overall, 111 (15.4%) of 721 human samples were positive for C. difficile; 67 (60.4%) of these samples contained A+B+ toxigenic isolates, and none of them contained A?B+ variant strains. Fifty (8%) of 622 animal samples contained C. difficile strains, which were toxigenic in 27 (54%) cases, including 1 A?B+ variant isolate. Eighty of the 721 human stool samples (37 positive and 43 negative for C. difficile culture) were comparatively tested by Premier Toxins A&B (Meridian Bioscience) and Triage C. difficile Panel (Biosite) immunoassays, the results of which were found concordant with toxigenic culture for 82.5 and 92.5% of the samples, respectively. The multiplex PCR toxigenic culture scheme described here allows combined diagnosis and toxigenic type characterization for human and animal C. difficile intestinal infections.
机译：设计了多重PCR产毒培养方法，用于同时鉴定和鉴定艰难梭菌分离株的产毒类型。设计了三对引物，用于扩增（i） tpi （三糖磷酸异构酶）基因的物种特异性内部片段，（ii） tcdB 的内部片段。 em>（毒素B）基因，和（iii） tcdA （毒素A）基因的内部片段，可区分毒素A阳性，毒素B阳性（A + B +）菌株和毒素A阴性，毒素B阳性（A？B +）变异株。多重PCR的可靠性是通过使用72℃的面板确定的。艰难梭菌菌株，包括A + B +，A？B？和A？B +产毒类型以及其他11种物种类型菌株。然后将多重PCR分析包括在产毒培养方法中，用于检测，鉴定和鉴定C的产毒类型。在1,343个连续的人类和动物粪便样本中检测到艰难细菌。总体上，在721个人类样品中，有111个（15.4％）的C呈阳性。困难;这些样品中有67个（60.4％）含有A + B +产毒分离株，没有一个含有AβB+变异株。 622个动物样本中有50个（8％）含有C。艰难梭菌菌株，在27例（54％）病例中有毒，包括1个A？B +变异株。 Premier Toxins A＆B（Meridian Bioscience）和Triage C对721份人类粪便样本中的80份（难辨梭状芽孢杆菌培养阳性37例和阴性）进行了比较测试。艰难梭菌（Biosite）免疫测定，结果分别与82.5和92.5％的产毒培养一致。此处描述的多重PCR产毒培养方案可对人和动物C进行综合诊断和产毒类型表征。艰难的肠感染。

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