Monitoring the water quality is a requisite to prevent outbreaks related to waterborne diseases, predominantly caused by enteric pathogens, such as bacteria and viruses. Enteric RNA viruses are transmitted to humans usually via the fecal-oral route and water is recognized as an important transmission vehicle. This project aimed to monitor Enterovirus, Norovirus from two genogroups (NoV GI and NoV GII), hepatitis A virus (HAV), and hepatitis E virus (HEV), in water matrices in Portugal: natural water from two surface sources; drinking water of two Water Treatment Plants (WTPs), and at the water distribution network; untreated and treated wastewater of three Wastewater Treatment Plants (WWTPs). Samples of surface and drinking water were collected in May 2018 and in a sampling campaign from January to December 2019. Wastewater samples were collected between November 2019 and March 2020. The detection and quantification of viral RNA were performed by a reverse transcription quantitative PCR (RT-qPCR) approach and the infectivity of RT-qPCR positive samples was evaluated in cultured Vero E6 cells. Surface water and drinking water samples were also taken for the analysis of fecal indicator bacteria (FIB) by cultural methods. Viral RNAs from Enterovirus, HAV, HEV, NoV I and NoV were detected and quantified in the two sources of surface water, river and dam reservoir (23 in 27 samples), as well as in drinking water sampled at the outlet of WTP_R, WTP_D and the distribution network (27 in 36 samples). HEV and HAV RNAs were the most and the least frequently detected, respectively. RNAs of Enterovirus and NoV I detected in surface waters, appeared to be eliminated with the treatment at both WTPs. However, HEV, NoV II, and HAV RNAs (0–109,687.5 gc/L), found in surface water samples, were still detected in drinking water, although usually at lower concentrations (0–5,617.1 gc/L). Our results also evidenced the existence of samples positive for infectious HEV (8 in 19 samples analyzed), in both types of water matrix. On the other hand, FIB were only detected in surface water, having been eliminated by WTPs to non-detectable values in the drinking water. Only a positive correlation was found between enteric viruses and FIB; this was in surface water, between NoV I and intestinal enterococci. In untreated wastewater, viral RNA was detected in all samples collected, being Enterovirus and NoV II RNAs the most frequently detected (14 in 14 samples). Norovirus was the viral genus whose representatives evidenced the highest RNA concentrations (5,448,041.8 gc/L). In treated wastewater, all collected samples were positive for RNA from at least one of the viruses under study, NoV II and NoV I having been the most frequently detected (14 in 14 samples and 13 in 14, respectively). Viral RNA was differently eliminated in the three WWTPs, depending on the type of treatment used, having been confirmed that the highest rates of reduction were achieved with tertiary treatment (31.4–100%). In this context, it is very important to consider the insertion of enteric viruses in the mandatory/regulatory analysis of water quality and to maintain their monitoring over time with the aim to prevent risks of transmission in the community.

Changes in the occurrence, distribution, and seasonal variation of waterborne pathogens due to global climate change may increase the risk of human exposure to these microorganisms, thus heightening the need for more reliable surveillance systems. Routine monitoring of drinking water supplies and recreational waters is performed using fecal indicator microorganisms, such as Escherichia coli, Enterococcus spp., and coliphages. However, the presence and numbers of these indicators, especially E. coli and Enterococcus spp., do not correlate well with those of other pathogens, especially enteric viruses, which are a major cause of waterborne outbreaks associated with contaminated water and food, and recreational use of lakes, ponds, rivers, and estuarine waters. For that reason, there is a growing need for a surveillance system that can detect and quantify viral pathogens directly in water sources to reduce transmission of pathogens associated with fecal transmission. In this review, we present an updated overview of relevant waterborne enteric viruses that we believe should be more commonly screened to better evaluate water quality and to determine the safety of water use and reuse and of epidemiological data on viral outbreaks. We also discuss current methodologies that are available to detect and quantify these viruses in water resources. Finally, we highlight challenges associated with virus monitoring. The information presented in this review is intended to aid in the assessment of human health risks due to contact with water sources, especially since current environmental and adaptive changes may be creating the need for a paradigm shift for indicators of fecal contamination.

Hepatitis E virus (HEV) is a non-enveloped single-stranded positive-sense RNA virus, belonging to the Hepeviridae family, resistant to environmental conditions, and transmitted by the consumption of contaminated water. This virus is responsible for both sporadic and epidemic outbreaks, leading to thousands of infections per year in several countries, and is thus considered an emerging disease in Europe and Asia. This study refers to a survey in Portugal during 2019, targeting the detection and eventual quantification of enteric viruses in samples from surface and drinking water. Samples positive for HEV RNA were recurrently found by reverse transcription quantitative PCR (RT-qPCR), in both types of matrix. The infectivity of these samples was evaluated in cultured Vero E6 cells and RNA from putative viruses produced in cultures evidencing cytopathic effects and was subjected to RT-qPCR targeting HEV genomic RNA. Our results evidenced the existence of samples positive either for HEV RNA (77.8% in surface water and 66.7% in drinking water) or for infectious HEV (23.0% in surface water and 27.7% in drinking water). These results highlight the need for effective virological control of water for human consumption and activities.

Monitoring the quality of water is a requisite to prevent outbreaks related to waterborne diseases, predominantly caused by pathogens like enteric viruses, usually transmitted via the fecal-oral route. This study aimed to survey a group of enteric viruses (Enterovirus, Norovirus genogroups I and II, and hepatitis A virus) in two surface water sources of drinking water, also intending to evaluate the extent of their elimination in the two water treatment plants (WTPs) involved in drinking water production. Correlations between these viruses and fecal indicator bacteria (FIB) were also evaluated. Positive samples for viral RNA were recurrently found by reverse transcription quantitative PCR (RT-qPCR) and quantified, in genomic copies per liter (gc/L) of sampled water. Viral RNAs were detected in 14 out of 27 samples of surface water, and 21 out of 36 samples of drinking water, NoV II having been the most frequently detected in both (0–78.6 gc/L and 0–12.5 gc/L, respectively). Both WTPs showed variable efficacies in the elimination of viral RNA. Only one correlation was found with FIB, between NoV II and intestinal enterococci. These results recommend the monitoring of enteric viruses over time and their inclusion in the mandatory analysis of water quality.

Water-related infectious diseases are important causes of morbidity and mortality. Enterovirus and Hepatitis A and E viruses are, among others, representative of the enteric viruses, which are disease-causing agents mainly transmitted by the oral-fecal route, through water. The objective of this study is to search for enteric viruses by reverse transcription followed by Real-time PCR (RT-qPCR), in samples of natural surface water and in drinking water. It is also intended to evaluate the eventual association of these viruses with other fecal contamination indicators, and the efficacy of the water treatment plants in their elimination. It was confirmed the adequacy of the methodology implemented since, in the 15 samples analyzed so far, Hepatitis A virus and Enterovirus RNAs were detected in natural surface water samples, in two and three samples, respectively. Both viruses RNAs were detected in one of these samples. No viral nucleic acids were detected in drinking water samples. Fecal coliforms (microbial indicators of fecal contamination) were detected in natural water samples, but not in drinking water samples. Viral RNA and coliform detection only partially co-occurred.

Water quality monitoring is a fundamental tool in the management of freshwater resources. The purpose of monitoring is to provide meaningful quality data for local action planning and catchment-wide decision making. The assessment of water quality is crucial to guarantee the efficient operation of the Water Treatment Plants (WTPs), promoting health conditions and contributing for a more sustainable urban water cycle. In accordance, the objective of this study was to evaluate key target chemical and microbiological water quality parameters, some of them already monitored within Portuguese/EU legal framework and others still not regulated, but with environmental and human heath relevance. A local monitoring database model, using a 6-year period (from 2014 to 2019) of water quality data, regarding water samples collected on representative sampling locations covering the freshwater abstraction sites, conventional WTPs and distribution network was assessed. This work provides new knowledge regarding occurrence and seasonal behaviour for both microbiological and chemical water quality parameters, essential to understand/manage the water supply system. Additionally, relationships between the target variables were also assessed. Particularly, strong correlations were identified between TOC and THMs formation at distribution network (r = 0.69; p ≤ 0.001); nitrates were the water quality parameter that revealed the best correlation between surface water source and treated water (r = 0.81; p ≤ 0.001), suggesting that treatment yield/performance is dependent on surface water load. The local and continuous monitoring of water systems are crucial to implement new approaches to guarantee the best quality of drinking water throughout the supply system.