Detection of SARS-CoV-2 in wastewater is influenced by sampling time, concentration method, and target analyzed

Abstract

The detection of SARS-CoV in wastewater has been proposed as a tool for monitoring COVID-19 at the community level. Although many reports have been published about detecting viral RNA in wastewater and its presence has been linked to infected people, appropriate analytical methodologies to use this approach have not yet been established. In this study, we compared ultrafiltration, polyethylene glycol precipitation, flocculation using AlCl3, and flocculation with skim milk for the recovery of SARS-CoV-2, using RNA from patients with positive diagnoses for COVID-19 and Pseudomonas phage φ6 as the control. We also evaluated the primers for detecting the E, RdRp, and N genes of the virus, as well as different storage times. Differences in the recovery efficiencies were evident with the different concentration methods, the best being ultrafiltration and precipitation with aluminum, which had recovery rates of 42.0% and 30.0%, respectively, when virus was present at high levels. Significant differences were found between the recoveries using wastewater and deionized water and between different storage times, with better recoveries for 6 and 12 h samplings. The E gene was the only one detected in all the samples analyzed. The results show that although this approach can provide important data for studying the pandemic, clear protocols are necessary for investigations to be comparable. HIGHTLIGHTS RNA of SARS-CoV-2 was used to evaluate the recovery of different methods in wastewater.; The utility of phage φ6 of Pseudomonas as a control for evaluation of recovery methods of SARS-CoV-2 from wastewater was demonstrated.; Differences due to the target selected for the detection of SARS were demonstrated.; The results show that it is possible to detect SARS RNA in wastewater, however its epidemiological usefulness will depend on the correct validation of the technique used for detection.;

Keywords

• covid-19
• epidemiology
• method validation
• rt-pcr
• sars-cov-2
• wastewater