Evaluation of Sars-Cov-2 Concentrations in Wastewater and River Water Samples

It is well-established that wastewater-based epidemiology (WBE) represents an important tool to monitor the occurrence of different viruses at population scale, including the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, there are few established methods for the concentration of encapsulated viruses, such as SARS-CoV-2, in water samples, which may limit the application of this approach. Thus, our study compared four methods used commonly to concentrate non-encapsulated gastrointestinal virus for the concentration of SARS-CoV-2 from wastewater and river water samples. The methods tested herein could be categorized in: vacuum filtration using electronegative membrane plus Mg2+ addiction; precipitation-based (aluminum hydroxide-based precipitation or polyethylene glycol (PEG) 8000 precipitation) and ultrafiltration (porosity of 10 and 50 kDa). Before the concentration step, filtration or centrifugation were performed to remove suspended particles from samples (pretreatment). To evaluate the efficiency (%), samples of SARS-CoV-2 from nasopharyngeal swabs obtained from RT-qPCR positive patients were used as spiked samples. The second part of the analysis consisted in quantifying SARS-CoV-2 viral copy number in the samples without SARS-CoV-2 spiked sample. Of all tested methods, centrifugation followed by ultrafiltration using a 50 kDa cut-off was the best concentration for wastewater samples with spiked sample added (efficiency of 54.3 or 113.01%). Considering the river samples with spiked sample added, filtration followed by PEG 8000 precipitation was the most efficient method (57.8 or 34.3%). However, analysis of the raw samples showed that ultracentrifugation using 10 or 50 kDa cut-off filters showed the best concentration efficiency based on copy number, regardless of pretreatment or sample type (values ranging from 1.3x105 to 9.8x103). Overall, the results found showed that the effectiveness of the concentration method can vary depending on the type of sample, pretreatment, and the concentration method approach. Thus, this study contribute to the  use of WBE for the control of SARS-CoV-2.