Department of Civil & Environmental Engineering & Earth Sciences, University of Notre Dame, 156 Fitzpatrick Hall, Notre Dame, IN 46556, United States.
Department of Civil & Environmental Engineering & Earth Sciences, University of Notre Dame, 156 Fitzpatrick Hall, Notre Dame, IN 46556, United States.
Sci Total Environ. 2023 Dec 20;905:167190. doi: 10.1016/j.scitotenv.2023.167190. Epub 2023 Sep 21.
Viral pathogens are typically dilute in environmental waters, necessitating a concentration step prior to subsequent quantification or analysis. Historically, studies on viral concentration efficiency have been done by spiking known viruses into the sample; however, spike-in controls may not have the same behavior as "native" viruses exposed to environmental conditions. In this study, four concentration methods, including polyethylene glycol precipitation (PEG), skimmed milk flocculation (SMF), pH drop followed by filtration through a 0.45 μm filter (pH), and centrifugation using an Amicon filter (Amicon), were evaluated to concentrate native viral targets in wastewater. Viral targets included both indicators (crAssphage and pepper mild mottle virus) and pathogens (adenovirus, norovirus GII, human polyomavirus, and SARS-CoV-2) in addition to a bacterial marker (HF183). A non-native spike-in control was also added to compare native and spike-in recoveries. Recovery varied widely across targets and methods, ranging from 0.1 to 39.3 %. The Amicon method was the most broadly effective concentration for recovery efficiency. For the lowest-titer target, the PEG method resulted in the lowest number of non-detections, with 96.7 % positive detections for SARS-CoV-2, compared to 66.7 %, 80 %, and 76.7 % positive detections for SMF, pH, and Amicon, respectively. The non-native spike-ins chosen were only representative of a few native recovery trends, varying by both target and concentration method, and consistently under or over-estimated recovery. Overall, this study suggests the utility of including native targets in viral concentration evaluation and determining the efficiency of concentration methods for a specific target of interest.
病毒病原体在环境水中通常是稀释的，因此在进行后续定量或分析之前需要进行浓缩步骤。历史上，对病毒浓缩效率的研究是通过向样品中添加已知病毒来完成的；然而，添加的对照物可能与暴露于环境条件下的“天然”病毒的行为不同。在这项研究中，评估了四种浓缩方法，包括聚乙二醇沉淀（PEG）、脱脂乳絮凝（SMF）、pH 值下降后通过 0.45μm 过滤器过滤（pH）和使用 Amicon 过滤器进行离心（Amicon），以浓缩废水中的天然病毒靶标。病毒靶标包括指示物（crAssphage 和胡椒温和斑点病毒）和病原体（腺病毒、诺如病毒 GII、人多瘤病毒和 SARS-CoV-2）以及细菌标记物（HF183）。还添加了非天然添加对照物来比较天然和添加对照物的回收率。回收率因目标物和方法而异，范围从 0.1 到 39.3%。Amicon 方法是最广泛有效的浓缩回收率方法。对于最低滴度的靶标，PEG 方法导致 SARS-CoV-2 的非检测数量最少，阳性检测率为 96.7%，而 SMF、pH 和 Amicon 的阳性检测率分别为 66.7%、80%和 76.7%。选择的非天然添加对照物仅代表少数天然回收趋势，因目标物和浓缩方法而异，并且始终低估或高估回收率。总体而言，这项研究表明在病毒浓缩评估中包含天然靶标并确定特定感兴趣目标的浓缩方法效率的实用性。
