Department of Engineering, University of Yamanashi, 4-3-11 Takeda, Kofu, Yamanashi 400-8511, Japan.
Interdisciplinary Center for River Basin Environment, University of Yamanashi, 4-3-11 Takeda, Kofu, Yamanashi 400-8511, Japan.
Sci Total Environ. 2022 Nov 20;848:157613. doi: 10.1016/j.scitotenv.2022.157613. Epub 2022 Jul 25.
Several virus concentration methods have been developed to increase the detection sensitivity of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in wastewater, as part of applying wastewater-based epidemiology. Polyethylene glycol (PEG) precipitation method, a method widely used for concentrating viruses in wastewater, has some limitations, such as long processing time. In this study, Pegcision, a PEG-based method using magnetic nanoparticles (MNPs), was applied to detect SARS-CoV-2 in wastewater, with several modifications to increase its sensitivity and throughput. An enveloped virus surrogate, Pseudomonas phage φ6, and a non-enveloped virus surrogate, coliphage MS2, were seeded into wastewater samples and quantified using reverse transcription-quantitative polymerase chain reaction to assess the recovery performance of the Pegcision. Neither increasing MNP concentration nor reducing the reaction time to 10 min affected the recovery, while adding polyacrylic acid as a polyanion improved the detection sensitivity. The performance of the Pegcision was further compared to that of the PEG precipitation method based on the detection of SARS-CoV-2 and surrogate viruses, including indigenous pepper mild mottle virus (PMMoV), in wastewater samples (n = 27). The Pegcision showed recovery of 14.1 ± 6.3 % and 1.4 ± 1.0 % for φ6 and MS2, respectively, while the PEG precipitation method showed recovery of 20.4 ± 20.2 % and 18.4 ± 21.9 % (n = 27 each). Additionally, comparable PMMoV concentrations were observed between the Pegcision (7.9 ± 0.3 log copies/L) and PEG precipitation methods (8.0 ± 0.2 log copies/L) (P > 0.05) (n = 27). SARS-CoV-2 RNA was successfully detected in 11 (41 %) each of 27 wastewater samples using the Pegcision and PEG precipitation methods. The Pegcision showed comparable performance with the PEG precipitation method for SARS-CoV-2 RNA concentration, suggesting its applicability as a virus concentration method.
几种病毒浓缩方法已被开发出来，以提高废水中严重急性呼吸综合征冠状病毒 2（SARS-CoV-2）的检测灵敏度，这是应用基于废水的流行病学的一部分。聚乙二醇（PEG）沉淀法是一种广泛用于浓缩废水中病毒的方法，但存在处理时间长等局限性。在这项研究中，Pegcision 是一种使用磁性纳米颗粒（MNP）的 PEG 方法，已被应用于检测废水中的 SARS-CoV-2，并进行了一些修改以提高其灵敏度和通量。包膜病毒假型物假单胞菌噬菌体 φ6 和非包膜病毒假型物噬菌体 MS2 被接种到废水样品中，并使用逆转录定量聚合酶链反应进行定量，以评估 Pegcision 的回收性能。增加 MNP 浓度或将反应时间缩短至 10 分钟均不会影响回收效果，而添加聚丙烯酸作为多阴离子可提高检测灵敏度。进一步将 Pegcision 的性能与基于 SARS-CoV-2 和假型病毒（包括本地辣椒轻斑驳病毒（PMMoV））检测的 PEG 沉淀法进行比较，包括在废水样品中（n = 27）。Pegcision 对 φ6 和 MS2 的回收率分别为 14.1 ± 6.3％和 1.4 ± 1.0％，而 PEG 沉淀法的回收率分别为 20.4 ± 20.2％和 18.4 ± 21.9％（n = 27）。此外，Pegcision（7.9 ± 0.3 log 拷贝/L）和 PEG 沉淀法（8.0 ± 0.2 log 拷贝/L）之间观察到可比的 PMMoV 浓度（n = 27）（P > 0.05）。使用 Pegcision 和 PEG 沉淀法分别成功检测到 27 个废水样品中的 11 个（41％）SARS-CoV-2 RNA。Pegcision 对 SARS-CoV-2 RNA 浓度的浓缩性能与 PEG 沉淀法相当，表明其作为病毒浓缩方法的适用性。
