Department of Agriculture and Environmental Sciences and Cooperative Research Programs, Lincoln University in Missouri, Jefferson City, MO 65101, USA.
Department of Computer Science, Bioinformatics Institute, and C. S. Bond Life Sciences Center, University of Missouri, Columbia, MO 65211, USA.
Water Res. 2014 Sep 15;61:224-31. doi: 10.1016/j.watres.2014.05.015. Epub 2014 May 22.
This study is to tackle the challenge posed by the "naturalized" Escherichia coli population against the worldwide practice of E. coli-based water quality monitoring. In the literature, the putative glucosyltransferase gene (ycjM) of E. coli has been identified in silico to be one of the 114 genes specific to enteric E. coli. Based on the sequence of E. coli K-12 MG1655, a PCR assay (ycjPCR) targeting ycjM was developed in this study. As demonstrated by the ycjPCR assay using 367 E. coli strains isolated from animal feces, 97.2% of the isolates carried the ycjM with variations from 93.9% to 100% among nine different host sources, but none of the 17 strains of non-E. coli bacteria and only 23.0% of the environment-isolated cryptic Escherichia strains contained the ycjM. These data experimentally confirmed ycjM to be enteric specific. Our study also showed that the ycjPCR assay was superior to the commonly used tuf- or uidA-based PCR methods in differentiating enteric E. coli from ß-D-glucuronidase-positive environmental bacteria. Furthermore, study on 190 E. coli isolates from water samples, using EPA Method 1603 followed by bacterial identification with Biolog MicroStation™ and ycjPCR assay, indicated that the prevalence of ycjM in the E. coli water isolates had a significant (p < 0.05, odds ratio ) spatial variation from 69.6% to 93.8%. These data suggest that E. coli profile using EPA Method 1603 or other ß-D-glucuronidase-activity-based methods may need further analysis using the ycjM profile to accurately determinate fecal pollution in water.
本研究旨在应对“自然化”大肠杆菌种群对全球基于大肠杆菌的水质监测实践带来的挑战。在文献中，已通过计算机鉴定出大肠杆菌中假定的葡糖基转移酶基因（ycjM）是 114 种肠道大肠杆菌特异性基因之一。基于大肠杆菌 K-12 MG1655 的序列，本研究中开发了针对 ycjM 的 PCR 检测方法（ycjPCR）。通过使用 367 株从动物粪便中分离的大肠杆菌菌株进行的 ycjPCR 检测证实，97.2%的分离株携带 ycjM，9 种不同宿主来源的菌株之间的变异率为 93.9%至 100%，但非大肠杆菌的 17 株细菌和仅 23.0%的环境分离的隐生大肠杆菌都没有携带 ycjM。这些数据从实验上证实了 ycjM 的肠道特异性。我们的研究还表明，与常用的 tuf- 或 uidA 为基础的 PCR 方法相比，ycjPCR 检测方法在区分肠道大肠杆菌和ß-D-葡糖醛酸酶阳性环境细菌方面具有优越性。此外，对从水样中分离的 190 株大肠杆菌进行研究，使用 EPA 方法 1603 并结合 Biolog MicroStation™细菌鉴定和 ycjPCR 检测方法，表明 ycjM 在大肠杆菌水样分离株中的流行率存在显著的（p < 0.05，优势比）空间变化，从 69.6%到 93.8%不等。这些数据表明，使用 EPA 方法 1603 或其他ß-D-葡糖醛酸酶活性检测方法进行的大肠杆菌分析可能需要进一步使用 ycjM 分析来准确确定水中的粪便污染。
