New SARS-CoV-2 mutations are constantly emerging, raising concerns of increased transmissibility, virulence or escape from host immune response.

Hepatitis E virus (HEV) is an emerging causative agent of acute hepatitis worldwide. To provide insights into the epidemiology of HEV in Italy, a large-scale investigation was conducted into urban sewage over nine years (2011–2019), collecting 1374 sewage samples from 48 wastewater treatment plants located in all the 20 regions of Italy. Broadly reactive primers targeting the ORF1 and ORF2 regions were used for the detection and typing of HEV, followed by Sanger and next generation sequencing (NGS). Real-time RT-qPCR was also used to attempt quantification of positive samples. HEV RNA detection occurred in 74 urban sewage samples (5.4%), with a statistically significant higher frequency (7.1%) in central Italy. Fifty-six samples were characterized as G3 strains and 18 as G1. While the detection of G3 strains occurred in all the surveillance period, G1 strains were mainly detected in 2011–2012, and never in 2017–2019. Typing was achieved in 2 samples (3f subtype). Viral concentrations in quantifiable samples ranged from 1.2 × 10³ g.c./L to 2.8 × 10⁴ g.c./L. Our results suggest the considerable circulation of the virus in the Italian population, despite a relatively small number of notified cases, a higher occurrence in central Italy, and a noteworthy predominance of G3 strains.

Hepatitis A is a common form of viral hepatitis. It is usually transmitted through the ingestion of contaminated food and water. This systematic review was carried out to summarise the overall prevalence of Hepatitis A virus (HAV) in different water matrices: untreated and treated wastewater, surface water, groundwater, drinking water, and others (e.g., irrigation water and floodwater). The literature search was performed in four databases: PubMed, Web of Science, Global Index Medicus, and Excerpta Medica Database. Heterogeneity (I²) was assessed using the χ² test on the Cochran Q statistic and H parameters. A total of 200 prevalence data from 144 articles were included in this meta-analysis. The overall prevalence of HAV in water matrices was 16.7% (95% CI: 13.4–20.3). The prevalence for individual matrix was as follows: 31.4% (95% CI: 23.0–40.4) untreated wastewater, 18.0% (95% CI: 9.5–28.2) treated wastewater, 15.0% (95% CI: 10.1–20.5) surface water, 2.3% (95% CI: 0.1–6.0) in groundwater, 0.3% (95% CI: 0.0–1.7) in drinking water, and 8.5% (95% CI: 3.1–15.6) in other matrices. The prevalence was higher in low-income economies (29.0%). Africa and Eastern Mediterranean were the regions with higher HAV prevalence values. This study showed a high heterogeneity (I² > 75%) with a significant publication bias (p value Egger test < 0.001). The results of this review suggest that water matrices could be an important route of HAV transmission even in industrialized countries, despite the lower prevalence compared to less industrialized countries, and the availability of advanced water management systems. More effective water/wastewater treatment strategies are needed in developing countries to limit the environmental circulation of HAV.

The emergence of SARS-CoV-2 variants of concern (VOCs) and variants of interest (VOIs) poses an increased risk to global public health and underlines the need to prioritise monitoring and research to better respond to the COVID-19 pandemic. Wastewater monitoring can be used to monitor SARS-CoV-2 spread and to track SARS-CoV-2 variants. A long read amplicon sequencing approach based on the Oxford Nanopore technology, targeting the spike protein, was applied to detect SARS-CoV-2 variants in sewage samples collected in central Italy on April 2021. Next-generation sequencing was performed on three pooled samples. For variant identification, two approaches–clustering (unsupervised) and classification (supervised)–were implemented, resulting in the detection of two VOCs and one VOI. Key mutations of the Alpha variant (B.1.1.7) were detected in all of the pools, accounting for the vast majority of NGS reads. In two different pools, mutations of the Gamma (P.1) and Eta (B.1.525) variants were also detected, accounting for 22.4%, and 1.3% of total NGS reads of the sample, respectively. Results were in agreement with data on variant circulation in Italy at the time of wastewater sample collection. For each variant, in addition to the signature key spike mutations, other less common mutations were detected, including the amino acid substitutions S98F and E484K in the Alpha cluster (alone and combined), and S151I in the Eta cluster. Results of the present study show that the long-read sequencing nanopore technology can be successfully used to explore SARS-CoV-2 diversity in sewage samples, where multiple variants can be present, and that the approach is sensitive enough to detect variants present at low abundance in wastewater samples. In conclusion, wastewater monitoring can help one discover the spread of variants in a community and early detect the emerging of clinically relevant mutations or variants.