Department of Physiology, Wayne State University, Detroit, MI 48201, United States of America.
Department of Biochemistry, Microbiology, & Immunology, Wayne State University, Detroit, MI 48201, United States of America.
Sci Total Environ. 2024 Nov 10;950:175044. doi: 10.1016/j.scitotenv.2024.175044. Epub 2024 Jul 27.
Wastewater-based epidemiology (WBE) can be used as a part of a long-term strategy for detecting and responding rapidly to new outbreaks of infectious disease in the community. However, wastewater collected by grab samples may miss marker presence, and composite auto-sampling throughout a day is technically challenging and costly. Tampon swabs can be used as passive collectors of wastewater markers over hours, but recovery of the captured markers is a challenge. Our goal was to improve tampon elution methods for virus detection and variant analysis to increase the likelihood of detection near the Limit of Detection (LOD) and to potentially detect new or rare variants in a new outbreak. Counts of SARS-CoV-2 N1 and N2 markers in grab samples were compared to markers eluted from tampons that had been immersed in 3 sewersheds for 4-6 h during June to December 2023. We compared tampon elution methods that used different elution volumes, pressure, and amounts of Tween 20, evaluated after automated magnetic bead purification and RT-ddPCR of SARS-CoV-2 markers. Overall, method "SwabM2" in which tampons were eluted by high pressure squeeze in a 50 mL syringe after adding 2 mL of 0.5 X TE + 0.075 % Tween-20 yielded a median four-fold higher concentration of final purified SARS-CoV-2 markers than paired grab samples and significantly more than other tested tampon elution methods (p < 0.0001). Method SwabM2 was more likely to yield enough extracted nucleic acids for sequencing and also gave higher quality variant sequences than two other tampon elution methods. Variant analysis captured the Fall 2023 transition of variants from XBB to JN and "H" lineages. In summary, we demonstrated a tampon-based wastewater collecting and elution method that yielded higher counts, more detections near the LOD, and higher quality variant sequences compared to both grab samples and other tampon-based passive-collecting wastewater methods.
基于污水的流行病学（WBE）可以作为一种长期策略的一部分，用于检测和快速应对社区中新的传染病爆发。然而，通过 grab 采样收集的污水可能会错过标志物的存在，而一整天的复合自动采样在技术上具有挑战性且成本高昂。棉拭子可以作为污水标志物的被动收集器，持续数小时，但捕获标志物的回收是一个挑战。我们的目标是改进棉拭子洗脱方法，以用于病毒检测和变异分析，从而增加在接近检测限（LOD）附近检测的可能性，并有可能在新的爆发中检测到新的或罕见的变异。2023 年 6 月至 12 月期间，我们比较了 grab 样本中的 SARS-CoV-2 N1 和 N2 标志物计数与在 3 个污水流域中浸泡 4-6 小时的棉拭子洗脱标志物计数。我们比较了使用不同洗脱体积、压力和吐温 20 量的棉拭子洗脱方法，对 SARS-CoV-2 标志物进行自动磁珠纯化和 RT-ddPCR 后进行了评估。总体而言，在加入 2 mL 0.5 X TE+0.075%吐温-20 后，通过高压挤压在 50 mL 注射器中洗脱棉拭子的方法“SwabM2”产生的最终纯化 SARS-CoV-2 标志物浓度中位数比配对的 grab 样本高四倍，显著高于其他测试的棉拭子洗脱方法（p<0.0001）。方法 SwabM2 更有可能产生足够用于测序的提取核酸，并且比其他两种棉拭子洗脱方法产生更高质量的变异序列。变异分析捕获了 2023 年秋季从 XBB 到 JN 和“H”谱系的变异过渡。总之，我们展示了一种基于棉拭子的污水收集和洗脱方法，与 grab 样本和其他基于棉拭子的被动收集污水方法相比，该方法产生的计数更高，在接近 LOD 处的检测更多，并且变异序列质量更高。
