The development of methodology to differentiate mixed populations of Escherichia coli in the secondary habitat might improve monitoring of fecal pollution indicators and facilitate the development of strategies to mitigate bacterial pollution. The objective of this study was to determine the ability of denaturing gradient gel electrophoresis (DGGE) to differentiate mixed assemblages of E. coli in the natural environment. After confirming the identity of 184 environmental bacterial isolates as E. coli, each was subjected to polymerase chain reaction (PCR) of the beta-glucuronidase gene (uidA) followed by DGGE fingerprinting. The ability of DGGE to discriminate individual isolates at the strain level was determined by comparing fingerprints to those resulting from a standard, library-dependent fingerprinting method, BOX-PCR. Computerized analysis of fingerprints indicated that DGGE and BOX-PCR identified 15 and 21 unique phylotypes respectively. Rank-abundance plots comparing the numerical distribution of unique E. coli phylotypes detected by both methods revealed no difference in resolution at the population level. In water and sediment samples from two beaches, DGGE effectively distinguished indigenous E. coli populations with an average rate of correct classification (site-based) of 83%. Denaturing gradient gel electrophoresis of uidA genes isolated and PCR-amplified from environmental samples appears to be an effective tool to differentiate unique E. coli populations and should be useful to characterize E. coli dynamics in the secondary environment.

译文

:发展区分次级生境中大肠杆菌混合种群的方法学,可能会改善对粪便污染指标的监测,并有助于制定减轻细菌污染的策略。这项研究的目的是确定变性梯度凝胶电泳(DGGE)在自然环境中区分大肠杆菌混合组合的能力。在确认了184种环境细菌分离株的身份为大肠埃希菌后,对每种细菌进​​行β-葡萄糖醛酸苷酶基因(uidA)的聚合酶链反应(PCR),然后进行DGGE指纹分析。通过比较指纹与标准,依赖库的指纹分析方法BOX-PCR产生的指纹,可以确定DGGE在菌株水平上区分各个分离株的能力。指纹的计算机分析表明,DGGE和BOX-PCR分别鉴定了15种和21种独特的系统型。比较两种方法检测到的独特大肠杆菌系统型的数值分布的秩-丰度图显示,在种群水平上分辨率没有差异。在来自两个海滩的水和沉积物样本中,DGGE有效地区分了本土大肠杆菌种群,正确分类(基于现场)的平均比率为83%。从环境样品中分离并通过PCR扩增的uidA基因的变性梯度凝胶电泳似乎是区分独特的大肠杆菌种群的有效工具,并且对于表征次级环境中的大肠杆菌动态也应是有用的。

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