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首页> 外文期刊>Journal of Clinical Microbiology >Direct Detection of Erythromycin-Resistant Bordetella pertussis in Clinical Specimens by PCR
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Direct Detection of Erythromycin-Resistant Bordetella pertussis in Clinical Specimens by PCR

机译：PCR直接检测临床标本中抗红霉素的百日咳博德特氏菌

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Resistance of Bordetella pertussis to erythromycin has been increasingly reported. We developed an allele-specific PCR method for rapid detection of erythromycin-resistant B. pertussis directly from nasopharyngeal (NP) swab samples submitted for diagnostic PCR. Based on the proven association of erythromycin resistance with the A2047G mutation in the 23S rRNA of B. pertussis, four primers, two of which were designed to be specific for either the wild-type or the mutant allele, were used in two different versions of the allele-specific PCR assay. The methods were verified with results obtained by PCR-based sequencing of 16 recent B. pertussis isolates and 100 NP swab samples submitted for diagnostic PCR. The detection limits of the two PCR assays ranged from 10 to 100 fg per reaction for both erythromycin-susceptible and -resistant B. pertussis. Two amplified fragments of each PCR, of 286 and 112 bp, respectively, were obtained from a mutant allele of the isolates and/or NP swab samples containing B. pertussis DNAs. For the wild-type allele, only a 286-bp fragment was visible when the allele-specific PCR assay 1 was performed. No amplification was found when a number of non-Bordetella bacterial pathogens and NP swab samples that did not contain the DNAs of B. pertussis were examined. This assay can serve as an alternative for PCR-based sequencing, especially for local laboratories in resource-poor countries.
机译：百日咳博德特氏菌对红霉素的耐药性已有越来越多的报道。我们开发了一种等位基因特异性PCR方法，可直接从提交诊断性PCR的鼻咽（NP）拭子样本中快速检测抗红霉素的百日咳博德特氏菌。基于已证实的红霉素抗性与百日咳博德特氏菌23S rRNA中A2047G突变的关联，在两种不同版本的植物中使用了四种引物，其中两种被设计成对野生型或突变等位基因具有特异性。等位基因特异性PCR分析。通过对16株最近的百日咳博德特氏菌分离株和100份NP拭子样品进行基于PCR的测序获得的结果验证了方法，这些样品已提交诊断性PCR。对于易感红霉素和耐药百日咳博德特氏菌，两种反应的每个PCR检测的检出限为10至100 fg。从含有百日咳博德特氏菌DNA的分离株和/或NP拭子样品的突变等位基因中获得每个PCR的两个扩增片段，分别为286和112 bp。对于野生型等位基因，当进行等位基因特异性PCR分析1时，仅可见286 bp的片段。当检查许多不含百日咳博德特氏菌DNA的非贝雷氏杆菌细菌病原体和NP拭子样品时，未发现扩增。该测定法可以替代基于PCR的测序，特别是对于资源贫乏国家的本地实验室而言。

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