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

Abstract

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 (ρ = 0.75, <i>p</i> < 0.001) and Martinez II (ρ = 0.68, <i>p</i> < 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.