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通过多重聚合酶链反应检测艰难梭菌的毒力基因

Detection of virulence genes of Clostridium difficile by multiplex PCR.

作者信息

Antikainen Jenni, Pasanen Tanja, Mero Sointu, Tarkka Eveliina, Kirveskari Juha, Kotila Saara, Mentula Silja, Könönen Eija, Virolainen-Julkunen Anni-Riitta, Vaara Martti, Tissari Päivi

机构信息

Division of Clinical Microbiology, HUSLAB, Helsinki University Central Hospital, Helsinki, Finland.

出版信息

APMIS. 2009 Aug;117(8):607-13. doi: 10.1111/j.1600-0463.2009.02509.x.

Abstract

Clostridium difficile strains belonging to the PCR ribotype 027, pulse-field gel electrophoresis (PFGE) type NAP1, toxinotype III and restriction endonuclease analysis group BI harbouring mutations in the tcdC gene and possessing binary toxin components A and B have been described to cause epidemics with increased morbidity and mortality. In the present study we developed a conventional multiplex PCR designed to detect selected virulence associated markers of the hypervirulent C. difficile PCR ribotype 027. The multiplex PCR assay detected the major toxins A and B, binary toxin components A and B as well as a possible deletion in the tcdC gene: a characteristic pattern of amplification products for the PCR ribotype 027 strains was detected. This rather simple method was specific for the screening of this hypervirulent C. difficile strain. The correlation between the multiplex PCR and PCR ribotyping methods was excellent. The sensitivity and specificity were 100% in our epidemiological situation. In conclusion, this multiplex PCR was found useful in the preliminary screening for the hypervirulent C. difficile PCR ribotype 027.

摘要

已报道属于PCR核糖型027、脉冲场凝胶电泳(PFGE)型NAP1、毒素型III以及限制性内切酶分析组BI的艰难梭菌菌株,其tcdC基因存在突变并拥有二元毒素成分A和B,可引发发病率和死亡率增加的疫情。在本研究中,我们开发了一种传统的多重PCR方法,用于检测高毒力艰难梭菌PCR核糖型027的特定毒力相关标志物。该多重PCR检测法可检测主要毒素A和B、二元毒素成分A和B以及tcdC基因中可能存在的缺失:检测到了PCR核糖型027菌株的特征性扩增产物模式。这种相当简单的方法对筛选这种高毒力艰难梭菌菌株具有特异性。多重PCR与PCR核糖分型方法之间的相关性极佳。在我们的流行病学情况下,敏感性和特异性均为100%。总之,发现这种多重PCR在高毒力艰难梭菌PCR核糖型027的初步筛选中很有用。

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