No abstract

Detection of SARS-CoV-2 in wastewater in Italy is described for the first time.• Use of the PEG/dextran concentration method for SARS-CoV-2 is reported. • A novel nested PCR assay specific for SARS-CoV-2 was designed. • Wastewater-based epidemiology can be applied for COVID-19 surveillance.

Several studies have demonstrated the advantages of environmental surveillance through the monitoring of sewer systems for the assessment of viruses circulating in a given community (wastewater-based epidemiology, WBE). During the COVID-19 public health emergency, many reports have described the presence of SARS-CoV-2 RNA in stools from COVID-19 patients, and a few studies reported the occurrence of SARS-CoV-2 in wastewaters worldwide. Italy is among the world's worst-affected countries in the COVID-19 pandemic, but so far there are no studies assessing the presence of SARS-CoV-2 in Italian wastewaters. To this aim, twelve influent sewage samples, collected between February and April 2020 from Wastewater Treatment Plants in Milan and Rome, were tested adapting, for concentration, the standard WHO procedure for Poliovirus surveillance. Molecular analysis was undertaken with three nested protocols, including a newly designed SARS-CoV-2 specific primer set. SARS-CoV-2 RNA detection occurred in volumes of 250 mL of wastewaters collected in both areas at high (Milan) and low (Rome) epidemic circulation, according to clinical data. Overall, 6 out of 12 samples were positive. One of the positive results was obtained in a Milan wastewater sample collected a few days after the first notified Italian case of autochthonous SARS-CoV-2. The study shows that WBE has the potential to be applied to SARS-CoV-2 as a sensitive tool to study spatial and temporal trends of virus circulation in the population.

The qualified presumption of safety (QPS) was developed to provide a safety pre‐assessment within EFSA for microorganisms. Strains belonging to QPS taxonomic units (TUs) still require an assessment based on a specific data package, but QPS status facilitates fast track evaluation. QPS TUs are unambiguously defined biological agents assessed for the body of knowledge, their safety and their end use. Safety concerns are, where possible, to be confirmed at strain or product level, and reflected as ‘qualifications’. Qualifications need to be evaluated at strain level by the respective EFSA units. The lowest QPS TU is the species level for bacteria, yeasts and protists/algae, and the family for viruses. The QPS concept is also applicable to genetically modified microorganisms used for production purposes if the recipient strain qualifies for the QPS status, and if the genetic modification does not indicate a concern. Based on the actual body of knowledge and/or an ambiguous taxonomic position, the following TUs were excluded from the QPS assessment: filamentous fungi, oomycetes, streptomycetes, Enterococcus faecium, Escherichia coli and bacteriophages. The list of QPS‐recommended biological agents was reviewed and updated in the current opinion and therefore now becomes the valid list. For this update, reports on the safety of previously assessed microorganisms, including bacteria, yeasts and viruses (the latter only when used for plant protection purposes) were reviewed, following an Extensive Literature Search strategy. All TUs previously recommended for 2016 QPS list had their status reconfirmed as well as their qualifications. The TUs related to the new notifications received since the 2016 QPS opinion was periodically evaluated for QPS status in the Statements of the BIOHAZ Panel, and the QPS list was also periodically updated. In total, 14 new TUs received a QPS status between 2017 and 2019: three yeasts, eight bacteria and three algae/protists.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is responsible for the coronavirus disease COVID-19, a public health emergency worldwide, and Italy is among the most severely affected countries. The first autochthonous Italian case of COVID-19 was documented on February 21, 2020. We investigated the possibility that SARS-CoV-2 emerged in Italy earlier than that date, by analysing 40 composite influent wastewater samples collected - in the framework of other wastewater-based epidemiology projects - between October 2019 and February 2020 from five wastewater treatment plants (WWTPs) in three cities and regions in northern Italy (Milan/Lombardy, Turin/Piedmont and Bologna/Emilia Romagna). Twenty-four additional samples collected in the same WWTPs between September 2018 and June 2019 (i.e. long before the onset of the epidemic) were included as ‘blank’ samples. Viral concentration was performed according to the standard World Health Organization procedure for poliovirus sewage surveillance, with modifications. Molecular analysis was undertaken with both nested RT-PCR and real-rime RT-PCR assays. A total of 15 positive samples were confirmed by both methods. The earliest dates back to 18 December 2019 in Milan and Turin and 29 January 2020 in Bologna. Virus concentration in the samples ranged from below the limit of detection (LOD) to 5.6 × 10 4 genome copies (g.c.)/L, and most of the samples (23 out of 26) were below the limit of quantification of PCR. Our results demonstrate that SARS-CoV-2 was already circulating in northern Italy at the end of 2019. Moreover, it was circulating in different geographic regions simultaneously, which changes our previous understanding of the geographical circulation of the virus in Italy. Our study highlights the importance of environmental surveillance as an early warning system, to monitor the levels of virus circulating in the population and identify outbreaks even before cases are notified to the healthcare system.