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Climate change, unpredictable weather patterns, and droughts are depleting water resources in some parts of the globe, where recycling and reusing wastewater is a strategy for different purposes. To counteract this, the EU regulation for water reuse sets minimum requirements for the use of reclaimed water for agricultural irrigation, including a reduction in human enteric viruses. In the present study, the occurrence of several human enteric viruses, including the human norovirus genogroup I (HuNoV GI), HuNoV GII, and rotavirus (RV), along with viral fecal contamination indicator crAssphage was monitored by using (RT)-qPCR methods on influent wastewater and reclaimed water samples. Moreover, the level of somatic coliphages was also determined as a culturable viral indicator. To assess the potential viral infectivity, an optimization of a capsid integrity PMAxx-RT-qPCR method was performed on sewage samples. Somatic coliphages were present in 60% of the reclaimed water samples, indicating inefficient virus inactivation. Following PMAxx-RT-qPCR optimization, 66% of the samples tested positive for at least one of the analyzed enteric viruses, with concentrations ranging from 2.79 to 7.30 Log10 genome copies (gc)/L. Overall, most of the analyzed reclaimed water samples did not comply with current EU legislation and contained potential infectious viral particles. [ABSTRACT FROM AUTHOR]

The current popularity of minimally processed foods is an opportunity for natural antimicrobial agents to be combined with mild heat treatments to act synergistically in reducing viral foodborne pathogens. Viral inactivation by heat-treatments (at 25, 40, 50 and 63 °C for 30 min) combined with aged green tea extract (aged-GTE) was initially evaluated in phosphate buffered saline (PBS) against murine norovirus (MNV-1) and hepatitis A virus (HAV) by cell culture, and against human norovirus by in situ capture RT-qPCR. The combination of aged-GTE and heat treatment at 50 °C for 30 min exerted strong antiviral activity, reducing by more than 5 log MNV-1 infectivity in PBS. Heating at 40 °C for 30 min reduced the binding of norovirus to porcine gastric mucine (PGM) to 41.5% and the addition of aged-GTE further decreased the binding to 4.7%. Additionally, the reduction of MNV-1 and HAV infectivity was investigated in two different types of juices exposed to mild heat treatments alone, and combined with aged-GTE. The addition of aged-GTE increased to more than 4 log the inactivation of MNV-1 in juices exposed to 50 °C for 30 min. However, this synergistic effect of aged-GTE combined with heat treatments was not observed for HAV in any of the juices. Aged-GTE, then, could be considered as an additional control measure to improve the food safety of mild heat pasteurized juices. • Viral inactivation by combined heat treatments and aged-GTE was shown. • A synergistic effect of GTE and heat was observed in PBS for MNV and norovirus. • Aged-GTE enhanced the inactivation of MNV in juices exposed to mild heat treatments. [ABSTRACT FROM AUTHOR]

• Wastewater-based epidemiology (WBE) co-detects multiple viral pathogens. • Enteric viruses were present in wastewater samples, with RVs showing winter peaks. • Co-circulation of SARS-CoV-2, RSV, and IAV respiratory viruses was proved by WBE. • Correlations with clinical data and predictive models highlight WBE's reliability. • Physicochemical parameters outperformed viral indicators as normalising factors. Wastewater-based epidemiology (WBE) has lately arised as a promising tool for monitoring and tracking viral pathogens in communities. In this study, we analysed WBE's role as a multi-pathogen surveillance strategy to detect the presence of several viral illness causative agents. Thus, an epidemiological study was conducted from October 2021 to February 2023 to estimate the weekly levels of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), Respiratory Syncytial virus (RSV), and Influenza A virus (IAV) in influent wastewater samples (n = 69). In parallel, a one-year study (October 2021 to October 2022) was performed to assess the presence of pathogenic human enteric viruses. Besides, monitoring of proposed viral fecal contamination indicators crAssphage and Pepper mild mottle virus (PMMoV) was also assessed, along with plaque counting of somatic coliphages. Genetic material of rotavirus (RV), human astrovirus (HAStV), and norovirus genogroup I (GI) and GII was found in almost all samples, while hepatitis A and E viruses (HAV and HEV) only tested positive in 3.77 % and 22.64 % of the samples, respectively. No seasonal patterns were overall found for enteric viruses, although RVs had a peak prevalence in the winter months. All samples tested positive for SARS-CoV-2 RNA, with a mean concentration of 5.43 log genome copies per liter (log GC/L). The tracking of the circulating SARS-CoV-2 variants of concern (VOCs) was performed by both duplex RT-qPCR and next generation sequencing (NGS). Both techniques reliably showed how the dominant VOC transitioned from Delta to Omicron during two weeks in Spain in December 2021. RSV and IAV viruses peaked in winter months with mean concentrations 6.40 and 4.10 log GC/L, respectively. Moreover, the three selected respiratory viruses strongly correlated with reported clinical data when normalised by wastewater physico-chemical parameters and presented weaker correlations when normalising sewage concentration levels with crAssphage or somatic coliphages titers. Finally, predictive models were generated for each respiratory virus, confirming high reliability on WBE data as an early-warning system and communities illness monitoring system. Overall, this study presents WBE as an optimal tool for multi-pathogen tracking reflecting viral circulation and diseases trends within a selected area, its value as a multi-pathogen early-warning tool stands out due to its public health interest. [Display omitted] [ABSTRACT FROM AUTHOR]