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首页> 外文期刊>Clinical and diagnostic laboratory immunology >Improved Repetitive-Element PCR Fingerprinting for Resolving Pathogenic and Nonpathogenic Phylogenetic Groups withinEscherichia coli
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Improved Repetitive-Element PCR Fingerprinting for Resolving Pathogenic and Nonpathogenic Phylogenetic Groups withinEscherichia coli

机译：改进的重复元素PCR指纹图谱，用于解决大肠杆菌内的致病性和非致病性系统发生基团

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Repetitive-element PCR (rep-PCR) fingerprinting is a promising molecular typing tool for Escherichia coli, including for discriminating between pathogenic and nonpathogenic clones, but is plagued by irreproducibility. Using the ERIC2 and BOXA1R primers and 15E. coli strains from the ECOR reference collection (three from each phylogenetic group, as defined by multilocus enzyme electrophoresis [MLEE], including virulence-associated group B2), we rigorously assessed the effect of extremely elevated annealing temperatures on rep-PCR's reproducibility, discriminating power, and ability to reveal MLEE-defined phylogenetic relationships. Modified cycling conditions significantly improved assay reproducibility and discriminating power, allowing fingerprints from different cyclers to be analyzed together with minimal loss of resolution. The correspondence of rep-PCR with MLEE with respect to tree structure and regression analysis of distances was substantially better with modified than with standard cycling conditions. Nonetheless, rep-PCR was only a fair surrogate for MLEE, and when fingerprints from different days were compared, it failed to distinguish between different clones within all-important phylogenetic group B2. These findings indicate that although the performance and phylogenetic fidelity of rep-PCR fingerprinting can be improved substantially with modified assay conditions, even when so improved rep-PCR cannot fully substitute for MLEE as a phylogenetic typing method for pathogenic E. coli.
机译：重复元素PCR（rep-PCR）指纹图谱是用于大肠杆菌（包括区分病原性和非病原性克隆）的一种很有前途的分子分型工具，但由于不可重复性而受到困扰。使用ERIC2和BOXA1R引物和15E。我们从ECOR参考库中收集了大肠杆菌（每个系统发育组中有3个，如多位点酶电泳[MLEE]所定义，包括与毒力相关的B2组），我们严格评估了退火温度过高对rep-PCR的影响重现性，区分能力和揭示MLEE定义的系统发育关系的能力。修改后的循环条件可以显着提高测定的可重复性和区分能力，从而可以分析不同循环仪的指纹，同时将分离度损失降至最低。修改后的rep-PCR与MLEE在树形结构和距离回归分析方面的对应性比标准自行车条件好得多。但是，rep-PCR只是MLEE的一个很好的替代品，当比较不同日期的指纹时，它无法区分所有重要系统发育组B2中的不同克隆。这些发现表明，尽管通过改进的测定条件可以显着改善rep-PCR指纹图谱的性能和系统发育保真度，但是即使如此改进的rep-PCR不能完全取代MLEE作为致病性 E的系统发育分型方法。大肠杆菌。

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