Department of Earth, Marine, and Environmental Sciences, Institute of Marine Science, University of North Carolina at Chapel Hill, Morehead City, North Carolina, United States of America.
PLoS One. 2022 Jun 24;17(6):e0270659. doi: 10.1371/journal.pone.0270659. eCollection 2022.
Wastewater based epidemiology (WBE) is useful for tracking and monitoring the level of disease prevalence in a community and has been used extensively to complement clinical testing during the current COVID-19 pandemic. Despite the numerous benefits, sources of variability in sample storage, handling, and processing methods can make WBE data difficult to generalize. We performed an experiment to determine sources of variability in WBE data including the impact of storage time, handling, and processing techniques on the concentration of SARS-CoV-2 in wastewater influent from three wastewater treatment plants (WWTP) in North Carolina over 19 days. The SARS-CoV-2 concentration in influent samples held at 4°C did not degrade significantly over the 19-day experiment. Heat pasteurization did not significantly impact the concentration of SARS-CoV-2 at two of the three WWTP but did reduce viral recovery at the WWTP with the smallest population size served. On each processing date, one filter from each sample was processed immediately while a replicate filter was frozen at -80°C. Once processed, filters previously frozen were found to contain slightly higher concentrations (<0.2 log copies/L) than their immediately processed counterparts, indicating freezing filters is a viable method for delayed quantification and may even improve recovery at WWTP with low viral concentrations. Investigation of factors contributing to variability during sample processing indicated that analyst experience level contributed significantly (p<0.001) to accepted droplet generation while extraction efficiency and reverse transcription efficiency contributed significantly (p<0.05) to day-to-day SARS-CoV-2 variability. This study provides valuable practical information for minimizing decay and/or loss of SARS CoV-2 in wastewater influent while adhering to safety procedures, promoting efficient laboratory workflows, and accounting for sources of variability.
基于污水的流行病学（WBE）可用于追踪和监测社区疾病流行水平，并且在当前 COVID-19 大流行期间已被广泛用于补充临床检测。尽管有许多好处，但由于样本储存、处理和处理方法的变异性，WBE 数据难以推广。我们进行了一项实验，以确定 WBE 数据中的变异性来源，包括储存时间、处理和处理技术对北卡罗来纳州三个污水处理厂（WWTP）污水进水口 SARS-CoV-2 浓度的影响超过 19 天。在 19 天的实验中，4°C 下储存的污水进水样本中的 SARS-CoV-2 浓度没有明显降解。在两个 WWTP 中，热巴氏消毒法对 SARS-CoV-2 浓度没有显著影响，但在服务人口最少的 WWTP 中，病毒回收率降低。在每个处理日期，从每个样本中取一个过滤器立即处理，而另一个过滤器在-80°C 下冷冻。一旦处理，以前冷冻的过滤器发现含有略高的浓度（<0.2 log 拷贝/L）比其立即处理的对应物，表明冷冻过滤器是一种可行的延迟定量方法，甚至可能提高病毒浓度低的 WWTP 的回收率。对样品处理过程中变异性的因素的调查表明，分析师的经验水平对接受液滴生成有显著影响（p<0.001），而提取效率和逆转录效率对 SARS-CoV-2 的日常变异性有显著影响（p<0.05）。本研究为在遵守安全程序的同时，最大限度地减少污水进水口 SARS-CoV-2 的衰减和/或损失，促进高效的实验室工作流程，并考虑变异性来源提供了有价值的实践信息。
