Wastewater Surveillance for SARS-CoV-2 in Northern Italy: An Evaluation of Three Different Gene Targets

Wastewater-based epidemiology has emerged as a complementary tool for the monitoring of COVID-19 pandemic waves and for the circulation of viral variants. The selection, standardization, and dynamics of different SARS-CoV-2 RNA targets in wastewater requires further investigation. In the present study, 106 wastewater samples were collected over a 24-month period from the wastewater treatment plant of Sondrio, north Italy, and were analyzed for the presence of SARS-CoV-2 RNA through the quantification of ORF1b, N1, and N3 gene targets via one-step real-time qPCR. In general, the three RNA targets demonstrated different performances and dynamics over the studied time period, underlying the usefulness of multiple viral targets in the surveillance of SARS-CoV-2 in wastewater. During the first 12 months, the quantification of the selected SARS-CoV-2 viral targets also correlated with the reported clinical cases in the same geographical area; however, from the overall data analysis this did not appear to significantly anticipate the epidemic waves. In conclusion, this study further supports the use of wastewater surveillance as a real time indicator of the human circulation of SARS-CoV-2. Moreover, the use of multiple viral gene targets has been shown to improve the reliability of SARS-CoV-2 surveillance in wastewater over time.