The bacterium Escherichia coli is commonly associated with the presence of faecal contamination in environmental samples, and is therefore subject to statutory surveillance. This is normally done using a culture-based methodology, which can be slow and laborious. Nucleic acid amplification for the detection of E. coli DNA sequences is a significantly more rapid approach, suited for applications in the field such as a point of sample analysis, and to provide an early warning of contamination. An existing, high integrity qPCR method to detect the E. coli ybbW gene, which requires almost an hour to detect low quantities of the target, was compared with a novel, isothermal RPA method, targeting the same sequence but achieving the result within a few minutes. The RPA technique demonstrated equivalent inclusivity and selectivity, and was able to detect DNA extracted from 100% of 99 E. coli strains, and exclude 100% of 30 non-target bacterial species. The limit of detection of the RPA assay was at least 100 target sequence copies. The high speed, and simple, isothermal amplification chemistry may indicate that RPA is a more suitable methodology for on-site E. coli monitoring than an existing qPCR technique.

中文翻译：

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重组酶聚合酶扩增用于快速、选择性、基于 DNA 的粪便指示性大肠杆菌检测

大肠杆菌通常与环境样本中的粪便污染有关，因此受到法定监督。这通常是使用基于文化的方法来完成的，这可能是缓慢而费力的。用于检测大肠杆菌 DNA 序列的核酸扩增是一种明显更快的方法，适用于现场应用，例如样品分析点，并提供污染的早期预警。将现有的高完整性 qPCR 检测大肠杆菌 ybbW 基因的方法与一种新型等温 RPA 方法进行了比较，该方法需要近一个小时才能检测到少量的目标，该方法针对相同的序列，但在几个小时内即可获得结果。分钟。RPA 技术展示了等效的包容性和选择性，并且能够检测 100% 从 99 种大肠杆菌菌株中提取的 DNA，并 100% 排除 30 种非目标细菌物种。RPA 检测的检测限为至少 100 个目标序列拷贝。高速、简单、等温扩增化学可能表明 RPA 是一种比现有 qPCR 技术更适合现场大肠杆菌监测的方法。