Quantitative CrAssphage PCR Assays for Human Fecal Pollution Measurement
Quantitative CrAssphage PCR Assays for Human Fecal Pollution Measurement
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DOI：
10.1021/acs.est.7b02703
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发表时间：
2017-08-15
影响因子：
11.4
通讯作者：
Shanks, Orin C.
中科院分区：
文献类型：
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作者：
Stachler, Elyse;Kelty, Catherine;Shanks, Orin C.
监测环境沃茨的粪便污染，以保护公众健康和水资源。传统上，使用一般的粪便指示细菌;然而，它们不能区分人类粪便废物和其他动物污染源。最近，一种新的噬菌体，crAsphage，被发现的宏基因组数据挖掘，并报道了丰富的和密切相关的人类粪便废物。为了证实生物信息学预测，沿crAspherage基因组的长度设计了沿着384个引物组。基于初始筛选，设计了两种新型crAsAsbilage qPCR检测试剂盒（CPQ_056和CPQ_064），并在参考粪便样本和水基质中进行了评价。当对动物粪便参考库进行测试时，该测定显示出高特异性（98.6%），并且crAsbridage遗传标记在原污水和污水影响的水样中非常丰富。此外，在配对实验中比较了CPQ_056和CPQ_064与HF 183/BacR 287和HumM 2检测试剂的性能。研究结果证实，病毒crAsA扩增qPCR检测与成熟的细菌人类相关粪便来源鉴定方法的水平相似。这些新的基于病毒的检测方法可能成为重要的水质管理和研究工具。
Environmental waters are monitored for fecal pollution to protect public health and water resources. Traditionally, general fecal-indicator bacteria are used; however, they cannot distinguish human fecal waste from other animal pollution sources. Recently, a novel bacteriophage, crAssphage, was discovered by metagenomic data mining and reported to be abundant in and closely associated with human fecal waste. To confirm bioinformatic predictions, 384 primer sets were designed along the length of the crAssphage genome. Based on initial screening, two novel crAssphage qPCR assays (CPQ_056 and CPQ_064) were designed and evaluated in reference fecal samples and water matrices. The assays exhibited high specificities (98.6%) when tested against an animal fecal reference library, and crAssphage genetic markers were highly abundant in raw sewage and sewage impacted water samples. In addition, CPQ_056 and CPQ_064 performance was compared to HF183/BacR287 and HumM2 assays in paired experiments. Findings confirm that viral crAssphage qPCR assays perform at a similar level to well-established bacterial human-associated fecal-source-identification approaches. These new viral-based assays could become important water quality management and research tools.