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首页> 美国卫生研究院文献>Applied and Environmental Microbiology >Comparison of Rapid Quantitative PCR-Based and Conventional Culture-Based Methods for Enumeration of Enterococcus spp. and Escherichia coli in Recreational Waters
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Comparison of Rapid Quantitative PCR-Based and Conventional Culture-Based Methods for Enumeration of Enterococcus spp. and Escherichia coli in Recreational Waters

机译：基于快速定量PCR的方法与基于常规培养物的肠球菌计数方法的比较。娱乐水域中的大肠杆菌和大肠杆菌

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摘要

Recreational water quality is currently monitored using culture-based methods that require 18 to 96 h for results. Quantitative PCR (QPCR) methods that can be completed in less than 2 h have been developed, but they could yield different results than the conventional methods. We present two studies in which samples were processed simultaneously for Enterococcus spp. and Escherichia coli using two culture-based methods (EPA method 1600 and Enterolert/Colilert-18) and QPCR. The proprietary QPCR assays targeted the 23S rRNA (Enterococcus spp.) and uidA (E. coli) genes and were conducted using lyophilized beads containing all reagents. In the first study, the QPCR method developers processed 54 blind samples that were inoculated with sewage or pure cultures or were ambient beach samples. The second study involved 163 samples processed by water quality personnel. The correlation between results of QPCR and EPA 1600 during the first study (r2) was 0.69 for Enterococcus spp., which was less than that observed between the culture-based methods (r2, 0.87). During the second study, the correlations were similar. No false positives occurred in either study when QPCR-based assays were used with blank samples. Levels of reproducibility measured through coefficients of variation were similar for results by Enterococcus QPCR and culture-based methods during both studies but were higher for E. coli QPCR results in the first study. Regarding the concentration at which beach management decisions are issued in the State of California, the agreement between results of Enterococcus QPCR and EPA method 1600 was 88%, compared to 94% agreement between EPA method 1600 and Enterolert. The beach management decision agreement between E. coli QPCR and Colilert-18 was 94%. The samples showing disagreement suggested an underestimation bias for QPCR.
机译：目前，休闲娱乐的水质使用基于培养的方法进行监控，需要18到96小时才能获得结果。已经开发了可以在不到2小时内完成的定量PCR（QPCR）方法，但是它们可以产生与常规方法不同的结果。我们目前进行两项研究，其中样本同时针对肠球菌进行了处理。和大肠杆菌使用两种基于培养的方法（EPA方法1600和Enterolert / Colilert-18）和QPCR。专有的QPCR分析针对23S rRNA（Enterococcus spp。）和uidA（E. coli）基因，并使用含有所有试剂的冻干珠进行。在第一项研究中，QPCR方法开发人员处理了54个盲样本，这些样本接种了污水或纯培养物，或者是环境海滩样本。第二项研究涉及水质人员处理的163个样品。第一次研究（r 2 ）对肠球菌的QPCR结果与EPA 1600之间的相关性为0.69，低于在基于培养的方法之间观察到的相关性（r 2 < /sup>，0.87）。在第二项研究中，相关性相似。当将基于QPCR的分析与空白样品一起使用时，两项研究均未出现假阳性。在两个研究中，肠球菌QPCR和基于培养物的方法通过变异系数测得的再现性水平相似，但在第一项研究中，大肠杆菌QPCR结果更高。关于在加利福尼亚州发布海滩管理决定的集中程度，肠球菌QPCR结果与EPA方法1600之间的一致率为88％，而EPA方法1600与Enterolert之间的一致性为94％。大肠杆菌QPCR与Colilert-18之间的海滩管理决策协议为94％。显示出分歧的样本表明QPCR的估计偏低。

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