Sequential treatment using a hydrophobic resin and gel filtration to improve viral gene quantification from highly complex environmental concentrates
Sequential treatment using a hydrophobic resin and gel filtration to improve viral gene quantification from highly complex environmental concentrates
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DOI：
10.1016/j.watres.2020.115652
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发表时间：
2020-05-01
期刊：
影响因子：
12.8
通讯作者：
Katayama, Hiroyuki
中科院分区：
文献类型：
--
作者：
Hata, Akihiko;Furumai, Hiroaki;Katayama, Hiroyuki
基于聚合酶链反应（PCR）的检测方法被广泛应用于定量水生环境中的肠道病毒，以研究它们的命运和潜在的感染风险。然而，通过病毒浓缩过程富集的抑制物质可能导致不准确的定量。本研究的目的是找到一种方法，通过减轻抑制性环境浓缩物的影响，使用以前的知识的抑制性物质的属性，提高病毒定量。使用脊髓灰质炎病毒及其提取的RNA掺入腐殖酸溶液中，对阴离子交换树脂、凝胶过滤和疏水树脂（DAX-8）的性能进行了比较评估。这些溶液作为抑制性环境浓缩物的良好代表。使用DAX-8树脂和凝胶过滤的顺序处理产生了最有利的结果，即，稳定的低病毒损失和降低的抑制作用。此外，将顺序处理应用于另一组15种环境浓缩物。在没有顺序处理的情况下，在8份样品中测量到分子过程对照（鼠诺如病毒）的严重低估（>4.0 log（10）至1.1 log（10））。在处理后，对照组检测到
Assays based on the polymerase chain reaction (PCR) are widely applied to quantify enteric viruses in aquatic environments to study their fates and potential infection risks. However, inhibitory substances enriched by virus concentration processes can result in inaccurate quantification. This study aimed to find a method for improving virus quantification by mitigating the effects of inhibitory environmental concentrates, using previous knowledge of the properties of the inhibitory substances. Performances of anion exchange resins, gel filtration, and a hydrophobic resin (DAX-8) were comparatively evaluated using poliovirus and its extracted RNA spiked into humic acid solutions. These solutions served as good representatives of the inhibitory environmental concentrates. A sequential treatment using DAX-8 resin and gel filtration produced the most favorable results, i.e., low virus losses that were stable and a reduced inhibitory effect. Furthermore, the sequential treatment was applied to another set of 15 environmental concentrates. Without the sequential treatment, serious underestimation (>4.0 log(10) to 1.1 log(10)) of a molecular process control (murine norovirus) was measured for eight samples. With the treatment, the control was detected with