Wastewater-based surveillance of COVID-19 and removal of SARS-CoV-2 RNA across a major wastewater treatment plant in San Antonio, Texas

Wastewater-based epidemiology (WBE) targeting SARS-CoV-2 RNA detection in municipal wastewater is considered a valuable tool for COVID-19 surveillance in a community. However, the persistence and removal of SARS-CoV-2 RNA in wastewater treatment plants (WWTPs) have not been well investigated. This study is aimed at detecting SARS-CoV-2 RNA in wastewater to correlate viral concentrations with clinical COVID-19 cases in the sewershed and determine whether the SARS-CoV-2 genetic material is detectable after treatment. Raw influent, primary effluent (after primary clarification), secondary effluent (after activated sludge treatment), and final effluent (after chlorination) samples were collected two times a week from the largest WWTP in San Antonio (Texas) during April to November 2021 and analyzed for SARS-CoV-2 RNA (N1 and N2 genes) concentrations using the reverse transcription droplet digital polymerase chain reaction (RT-ddPCR). SARS-CoV-2 RNA was detected in 98.5% (n = 34 weeks) of the raw influent samples and anticipated the trends of the COVID-19 outbreak. Furthermore, a higher correlation between viral concentrations and COVID-19 cases was observed for two days a week sampling frequency (ρ = 0.75, p <0.001) than one day per week (ρ = 0.60, p <0.001). Despite the high SARS-CoV-2 RNA concentrations in raw sewage, a significant amount of viral RNA was removed at primary and secondary clarifiers (removal efficiencies were 54% and 94%, respectively) and was undetectable in final effluents. These results demonstrate the performance of the WWTP in reducing the SARS-CoV-2 RNA concentration and further highlight the role of tertiary treatment and chlorination in eliminating SARS-CoV-2 RNA in receiving waters. © 2023 RSC.

Assessment of Concentration, Recovery, and Normalization of SARS-CoV-2 RNA from Two Wastewater Treatment Plants in Texas and Correlation with COVID-19 Cases in the Community

The purpose of this study was to conduct a correlative assessment of SARS-CoV-2 RNA concentrations in wastewater with COVID-19 cases and a systematic evaluation of the effect of using different virus concentration methods and recovery and normalization approaches. We measured SARS-CoV-2 RNA concentrations at two different wastewater treatment plants (WWTPs) in the Bexar County of Texas from October 2020 to May 2021 (32 weeks) using reverse transcription droplet digital PCR (RT-ddPCR). We evaluated three different adsorption-extraction (AE) based virus concentration methods (acidification, addition of MgCl2, or without any pretreatment) using bovine coronavirus (BCoV) as surrogate virus and observed that the direct AE method showed the highest mean recovery. COVID-19 cases were correlated significantly with SARS-CoV-2 N1 concentrations in Salitrillo (rho = 0.75, p < 0.001) and Martinez II (rho = 0.68, p < 0.001) WWTPs, but normalizing to a spiked recovery control (BCoV) or a fecal marker (HF183) reduced correlations for both treatment plants. The results generated in this 32-week monitoring study will enable researchers to prioritize the virus recovery method and subsequent correlation studies for wastewater surveillance.