Simultaneous detection of various pathogenic Escherichia coli in water by sequencing multiplex PCR amplicons
Simultaneous detection of various pathogenic Escherichia coli in water by sequencing multiplex PCR amplicons
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DOI：
10.1007/s10661-022-10863-6
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发表时间：
2023-02-01
影响因子：
3
通讯作者：
Ishii，Satoshi
中科院分区：
文献类型：
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作者：
Suzuki，Yoshihiro;Shimizu，Hiroki;Ishii，Satoshi
由于水中病原体污染而引起的水传播疾病是世界范围内的一个严重问题。准确和同步检测水中的病原体对保护公众健康至关重要。本研究建立了一种通过多重PCR扩增子测序同时检测多种致病性大肠杆菌的方法。我们设计的多重PCR扩增了5个致病基因。coli（uidA、stx1、stx2、STh基因和LT基因）。还设计了另外两种PCR测定法（foraggRandeae），并将其纳入扩增子测序分析中。数字PCR（dPCR）也使用相同的测定。对于大多数靶向基因，在序列读段计数和dPCR结果之间观察到强正相关，表明我们的多重PCR扩增子测序方法可以提供定量信息。该方法还成功地应用于病原菌的检测。大肠杆菌的河水和废水样品。这里展示的方法可以通过靶向其他病原体的基因来扩展。
Waterborne diseases due to pathogen contamination in water are a serious problem all over the world. Accurate and simultaneous detection of pathogens in water is important to protect public health. In this study, we developed a method to simultaneously detect various pathogenicEscherichia coliby sequencing the amplicons of multiplex PCR. Our newly designed multiplex PCR amplified five genes for pathogenicE. coli(uidA,stx1,stx2, STh gene, and LT gene). Additional two PCR assays (foraggRandeae) were also designed and included in the amplicon sequencing analysis. The same assays were also used for digital PCR (dPCR). Strong positive correlations were observed between the sequence read count and the dPCR results for most of the genes targeted, suggesting that our multiplex PCR-amplicon sequencing approach could provide quantitative information. The method was also successfully applied to monitor the level of pathogenicE. coliin river water and wastewater samples. The approach shown here could be expanded by targeting genes for other pathogens.