Improved methods for the detection and quantification of SARS-CoV-2 RNA in wastewater

Since the start of the COVID-19 pandemic, different methods have been used to detect the presence of genetic material of SARS-CoV-2 in wastewater. The use of wastewater for SARS-CoV-2 RNA detection and quantification showed different problems, associated to the complexity of the matrix and the lack of standard methods used to analyze the presence of an enveloped virus, such as coronavirus. Different strategies for the concentration process were selected to carry out the detection and quantification of SARS-CoV-2 RNA in wastewater: (a) aluminum hydroxide adsorption-precipitation, (b) pre-treatment with glycine buffer and precipitation with polyethylene-glycol (PEG) and (c) ultrafiltration (Centricon). Our results showed that the reduction of organic matter, using the pre-treatment with glycine buffer before the concentration with Centricon or aluminum hydroxide adsorption-precipitation, improved the recovery percentage of the control virus, Mengovirus (MgV) (8.37% +/- 5.88 n = 43; 6.97% +/- 6.51 n = 20, respectively), and the detection of SARS-CoV-2 in comparison with the same methodology without a pre-treatment. For the concentration with Centricon, the use of 100 mL of wastewater, instead of 200 mL, increased the MgV recovery, and allowed a positive detection of SARS-CoV-2 with N1 and N2 targets. The quantity of SARS-CoV-2 RNA detected in wastewater did not show a direct correlation with the number of confirmed cases, but the study of its upwards or downwards trend over time enabled the detection of an increase of epidemiological data produced in September 2020, January 2021 and April 2021.