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Wastewater pollution is one of the main causes of waterborne diseases (e.g., diarrheal diseases) because wastewater commonly contains a wide range of pathogenic microorganisms, notably human enteric viruses. Monitoring multiple pathogenic viruses in waters simultaneously is impractical and expensive, so monitoring through virus indicators is essential to ensure water safety and quality. Recently, pepper mild mottle virus (PMMoV) was found as one of the most prevalent viruses in the human gut microbiota and consistently present at high concentrations in human feces and domestic wastewater. This study reviewed the latest information on the presence of PMMoV and human enteric viruses in water environments and in wastewater (water) treatment systems to evaluate the suitability of PMMoV as an indicator virus for water safety and quality. PMMoV was present in all types of waters (e.g., wastewater, surface water, groundwater, coastal water and drinking water) in greater prevalence than human enteric viruses. PMMoV was also removed less or similar to human enteric viruses in various wastewater (water) treatment systems (including disinfection treatment). These results suggest that PMMoV can be used as a suitable indicator virus for 1) assessment of water quality polluted by domestic wastewater; 2) assessment of virus removal efficiency in drinking water treatment plants and 3) assessment of viral safety for drinking water.

Waterborne diseases caused by pathogenic human viruses are a major public health concern. To control the potential risk of viral infection through contaminated waters, a rapid, reliable tool to assess the infectivity of pathogenic viruses is required. Recently, an advanced approach (i.e., capsid integrity (RT-)qPCR) was developed to discriminate intact viruses (potentially infectious) from inactivated viruses. In this approach, samples were pretreated with capsid integrity reagents (e.g., monoazide dyes or metal compounds) before (RT -)qPCR. These reagents can only penetrate inactivated viruses with compromised capsids to bind to viral genomes and prevent their amplification, but they cannot enter viruses with intact capsids. Therefore, only viral genomes of intact viruses were amplified or detected by (RT-)qPCR after capsid integrity treatment. In this study, we reviewed recent progress in the development and application of capsid integrity (RT-)qPCR to assess the potential infectivity of viruses (including non-enveloped and enveloped viruses with different genome structures [RNA and DNA]) in water. The efficiency of capsid integrity (RT-)qPCR has been shown to depend on various factors, such as conditions of integrity reagent treatment, types of viruses, environmental matrices, and the capsid structure of viruses after disinfection treatments (e.g., UV, heat, and chlorine). For the application of capsid integrity (RT-)qPCR in real-world samples, the use of suitable virus concentration methods and process controls is important to control the efficiency of capsid integrity (RT-)qPCR. In addition, potential future applications of capsid integrity (RT-)qPCR for determining the mechanism of disinfection treatment on viral structure (e.g., capsid or genome) and a combination of capsid integrity treatment and next-generation sequencing (NGS) (capsid integrity NGS) for monitoring the community of intact pathogenic viruses in water are also discussed. This review provides essential information on the application of capsid integrity (RT-)qPCR as an efficient tool for monitoring the presence of pathogenic viruses with intact capsids in water.

Porous carbons are well-known efficient adsorbents for a variety of organic and inorganic pollutants, however, they have difficulty in virus removal. In this study, novel porous carbons (NPCs) (NPC-A, NPC-B, and NPC-C) derived from rice husks were compared with commercially available activated carbons (ACs) for their ability to remove MS2 bacteriophages (MS2) in a batch experiment. NPC-A was produced by the silica removal process. NPC-B was prepared with an additional steam activation applied to NPC-A. NPC-C was obtained with an additional acid rinse applied to NPC-B. The NPCs (particularly NPC-C) exhibited effective removal of up to 5.3 log10 of MS2, which was greater than that of less than 2.7 log10 obtained by other ACs under 10 g/L during the same contact time (60 min). The pore size distribution of the porous carbon adsorbents was found to influence their virus removal performance. The adsorbents with a larger proportion of pores ranging from 200–4500 nm in diameter were able to achieve higher virus removal rates. Thus, NPCs (particularly NPC-C), which had a larger volume of pores ranging from 200–4500 nm in size, demonstrated the potential for use as efficient adsorbents for removing viruses during water purification.

Safely managed water supply and wastewater services have been playing key roles in maintaining good water environmental quality, stopping the spread of disease and protecting public health during infectious outbreaks, including of ongoing coronavirus outbreak 2019 (COVID-19) caused by SARS-CoV-2. Recently, there are a growing number of scientific research findings, and reports around the world, which revealed that SARS-CoV-2 is detected in untreated domestic and hospital wastewater, and it is spreading through human feces. In order to address this emerging challenge and minimize associated human health risks, it is essential to have a better understanding on possible routes of SARS-CoV-2 infections and contamination across water and wastewater service chain based on recent scientific findings. Consequently, it will assist local authorities in identifying and implementing appropriate preventive countermeasures to stop possible COVID-19 transmission. Roles of using regular viral surveillance in untreated wastewater in the affected areas have also been highlighted in the chapter, as an effective early-warning tool, for revealing true scale of the coronavirus outbreak, trends of the pandemic, as well as providing early warnings to the community. If SARS-CoV-2 can be effectively monitored in infected communities at an early stage through wastewater-based epidemiology, together with results from clinical diagnostic testing or even in the areas where clinical surveillance is poor, effective interventions and preparedness actions can be taken as early as possible to restrict the movements of infected population, as well as to minimize the pathogen spread and threat to public health.

The presence of pathogenic viruses in drinking water is a major public health concern. Although viability RT-qPCR methods were developed to quantify infectious viruses, they may not always reflect viral infectivity, therefore leading to false-positive results. In this study, sodium deoxycholate (SD) pre-treatment was used to improve the efficiency of viability RT-qPCR methods with respect to exclusive quantification of infectious viruses. The ability of SD pre-treatment to enhance the penetration of three viability markers, namely, ethidium monoazide (EMA, 100 µM), propidium monoazide (PMA, 100 µM), and cis-dichlorodiammineplatinum (CDDP, 1000 µM), into heat-treated (90 °C for 1 min) Aichi virus at various concentrations (0.01–0.5%) was evaluated. The optimal SD concentration was found to be 0.1% for all markers. EMA/PMA/CDDP-RT-qPCR with 0.1% SD pre-treatment was significantly more effective than without SD pre-treatment in determining AiV inactivation after heat (50, 60, 70, 80, or 90 °C for 1 min) or chlorine treatment (1 mgCl2/L for 1, 2, 5, or 10 min). Among the viability RT-qPCR methods tested, CDDP-RT-qPCR with SD pre-treatment (SD-CDDP-RT-qPCR) was the most effective in reflecting viral infectivity. Performance testing of SD-CDDP-RT-qPCR in concentrated drinking water samples did not reveal any significant effects of SD-CDDP treatment. Thus, SD-CDDP-RT-qPCR could be a useful tool for monitoring infectious virus presence in drinking water.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomes have been detected in wastewater worldwide. However, the assessment of SARS-CoV-2 infectivity in wastewater has been limited due to the stringent requirements of biosafety level 3. The main objective of this study is to investigate the applicability of capsid integrity RT-qPCR for the selective detection of intact SARS-CoV-2 in wastewater. Three capsid integrity reagents, namely ethidium monoazide (EMA, 0.1–100 μM), propidium monoazide (PMA, 0.1–100 μM), and cis-dichlorodiammineplatinum (CDDP, 0.1–1000 μM), were tested for their effects on different forms (including free genomes, intact and heat-inactivated) of murine hepatitis virus (MHV), which was used as a surrogate for SARS-CoV-2. CDDP at a concentration of 100 μM was identified as the most efficient reagent for the selective detection of infectious MHV by RT-qPCR (CDDP-RT-qPCR). Next, two common virus concentration methods including ultrafiltration (UF) and polyethylene glycol (PEG) precipitation were investigated for their compatibility with capsid integrity RT-qPCR. The UF method was more suitable than the PEG method since it recovered intact MHV (mean ± SD, 38% ± 29%) in wastewater much better than the PEG method did (0.013% ± 0.015%). Finally, CDDP-RT-qPCR was compared with RT-qPCR alone for the detection of SARS-CoV-2 in 16 raw wastewater samples collected in the Greater Tokyo Area. Five samples were positive for SARS-CoV-2 when evaluated by RT-qPCR alone. However, intact SARS-CoV-2 was detected in only three positive samples when determined by CDDP-RT-qPCR. Although CDDP-RT-qPCR was unable to determine the infectivity of SARS-CoV-2 in wastewater, this method could improve the interpretation of positive results of SARS-CoV-2 obtained by RT-qPCR., Graphical abstract Unlabelled Image

It is important to evaluate the removal of enteric viruses by drinking water treatment processes so that viral infection risk can be assessed and managed. However, evaluating the removal of enteric viruses by full-scale treatment processes can be challenging due to the low numbers of viruses and the presence of substances in the water samples that inhibit detection. In this study, we evaluated the removal of pepper mild mottle virus (PMMoV) by microfiltration (MF) and slow sand filtration (SSF) at two full-scale drinking water treatment plants in Japan, quantifying virus concentrations with real-time polymerase chain reaction (qPCR). The removal of PMMoV by MF ranged from 0.0 to >0.9 log10, although concentrations were below the detection limit for half of the treated water samples. SSF removed PMMoV by up to 2.8 log10; however, the removal efficiency decreased to 0.0–1.0 log10 under cold water temperatures. Process control showed that nucleic acid extraction and qPCR efficiency were inhibited in nearly 40% of water samples. Dilution, DAX-8, and ferrihydrite treatments for purification were effective in mitigating these inhibitory effects.

Capsid integrity (RT-)qPCR has recently been developed to discriminate between intact forms from inactivated forms of viruses, but its applicability to identifying integrity of viruses in drinking water has remained limited. In this study, we investigated the application of capsid integrity (RT-)qPCR using cis-dichlorodiammineplatinum (CDDP) with sodium deoxycholate (SD) pretreatment (SD-CDDP-(RT-)qPCR) to detect intact viruses in surface water and tap water. A total of 63 water samples (surface water, n = 20; tap water, n = 43) were collected in the Kanto region in Japan and quantified by conventional (RT)-qPCR and SD-CDDP-(RT-)qPCR for pepper mild mottle virus (PMMoV) and seven other viruses pathogenic to humans (Aichivirus (AiV), noroviruses of genotypes I and II, enterovirus, adenovirus type 40 and 41, and JC and BK polyomaviruses). In surface water, PMMoV (100%) was more frequently detected than other human pathogenic viruses (30%–60%), as determined by conventional (RT-)qPCR. SD-CDDP-(RT-)qPCR also revealed that intact PMMoV (95%) was more common than intact human pathogenic viruses (20%–45%). In the tap water samples, most of the target viruses were not detected by conventional (RT-)qPCR, except for PMMoV (9%) and AiV (5%). PMMoV remained positive (5%), whereas no AiV was detected when tested by SD-CDDP-(RT-)qPCR, indicating that some PMMoV had an intact capsid, whereas AiV had damaged capsids. The presence of AiV in the absence of PMMoV in tap water produced from groundwater may demonstrate the limitation of PMMoV as a viral indicator in groundwater. In addition to being abundant in surface water, PMMoV was detected in tap water, including PMMoV with intact capsids. Thus, the absence of intact PMMoV may be used to guarantee the viral safety of tap water produced from surface water.

Groundwater pollution by arsenic is a major health threat in suburban areas of Hanoi, Vietnam. The present study evaluates the effect of the sedimentary environments of the Pleistocene and Holocene deposits, and the recharge systems, on the groundwater arsenic pollution in Hanoi suburbs distant from the Red River. At two study sites (Linh Dam and Tai Mo communes), undisturbed soil cores identified a Pleistocene confined aquifer (PCA) and Holocene unconfined aquifer (HUA) as major aquifers, and Holocene estuarine and deltaic sediments as an aquitard layer between the two aquifers. The Holocene estuarine sediments (approximately 25–40 m depth, 9.6–4.8 cal ka BP) contained notably high concentrations of arsenic and organic matter, both likely to have been accumulated by mangroves during the Holocene sea-level highstand. The pore waters in these particular sediments exhibited elevated levels of arsenic and dissolved organic carbon. Arsenic in groundwater was higher in the PCA (25–94 μg/L) than in the HUA (5.2–42 μg/L), in both the monitoring wells and neighboring household tubewells. Elevated arsenic concentration in the PCA groundwater was likely due to vertical infiltration through the arsenic-rich and organic-matter-rich overlying Holocene estuarine sediments, caused by massive groundwater abstraction from the PCA. Countermeasures to prevent arsenic pollution of the PCA groundwater may include seeking alternative water resources, reducing water consumption, and/or appropriate choice of aquifers for groundwater supply.

Over-exploited groundwater is expected to remain the predominant source of domestic water in suburban areas of Hanoi, Vietnam. In order to evaluate the effect on groundwater recharge, of decreasing surface-water bodies and land-use change caused by urbanization, the relevant groundwater systems and recharge pathways must be characterized in detail. To this end, water levels and water quality were monitored for 3 years regarding groundwater and adjacent surface-water bodies, at two typical suburban sites in Hanoi. Stable isotope (δ18O, δD of water) analysis and hydrochemical analysis showed that the water from both aquifers and aquitards, including the groundwater obtained from both the monitoring wells and the neighboring household tubewells, was largely derived from evaporation-affected surface-water bodies (e.g., ponds, irrigated farmlands) rather than from rivers. The water-level monitoring results suggested distinct local-scale flow systems for both a Holocene unconfined aquifer (HUA) and Pleistocene confined aquifer (PCA). That is, in the case of the HUA, lateral recharge through the aquifer from neighboring ponds and/or irrigated farmlands appeared to be dominant, rather than recharge by vertical rainwater infiltration. In the case of the PCA, recharge by the above-lying HUA, through areas where the aquitard separating the two aquifers was relatively thin or nonexistent, was suggested. As the decrease in the local surface-water bodies will likely reduce the groundwater recharge, maintaining and enhancing this recharge (through preservation of the surface-water bodies) is considered as essential for the sustainable use of groundwater in the area.

Molecular assays are currently the most widely used method to quantify pathogenic viruses in water; however, their performance is often disrupted by matrices present in environmental samples. The present study used ferrihydrite (Fh) treatment to mitigate inhibition of RT-qPCR virus detection from environmental water concentrates. Fh treatment was performed to improve the detection of spiked Aichi virus 1 (AiV) and Qβ bacteriophage (Qβ) in commercial humic acids and seawater concentrates. The optimal Fh doses were found to be 200, 500, and 1000 mgFe/L for humic acid concentrates at UV254 of 1.5, 3.0, and 6.0 cm-1 respectively, whereas this value was 1000 mgFe/L for the seawater concentrates at UV254 of 1.5 cm-1. At these optimal doses, the recoveries of spiked AiV and Qβ ranged from 2% to 12% in the humic acid and seawater concentrates, respectively. In addition, high levels of indigenous viruses (including AiV ≥ 2.81 log10 copies/mL and pepper mild mottle virus ≥ 2.77 log10 copies/mL) were detected from seawater concentrates after Fh treatment, while none were detected without Fh treatment. Fh treatment effectively mitigated the inhibitive effects from environmental water samples.

Multi-drug resistant microbes, pathogenic viruses, metals, and pharmaceuticals and personal care products (PPCPs) in water has become the crux of urban sustainability issues. However, vulnerability due to pollutant concurrences, source apportionment, and identification of better faecal indicators needs to be better understood. The present study focuses on the vulnerability of urban Guwahati, the largest city in Northeastern India, through analyzing the concurrence of PPCPs, enteric viruses, antibiotic resistant bacteria, metal, and faecal contamination in water. The study strives to identify a relevant marker of anthropogenic pollution for the Indian scenario. Samples from the Brahmaputra River (n = 4), tributary Bharalu River (an unlined urban drain; n = 3), and Ramsar recognized Lake (Dipor Bil; n = 1) indicate caffeine > acetaminophen > theophylline > carbamazepine > crotamiton for PPCPs and pepper mild mottle virus (PMMoV) > aichi > hepatitis A > norovirus GII > norovirus GI for enteric viruses. PMMoV was the better indicator of faecal pollution due to its prevalence, specificity and ease of detection. Antibiotic resistance was neither correlated with the prevalence of PPCPs nor E. coli. As, Co and Mn appear to be inducing antibiotic resistance in E. coli. While the risk quotient of the urban drain (Bharalu River) indicates one order higher magnitude than reported for other Indian rivers, the Lake exhibited the least pollution and better resilience. The concurrence of pollutants and multi-drug resistant E. coli, owing to the complete absence of wastewater treatment, puts the city in a highly vulnerable state. Pollution is being regulated only by the dilution capability of the Brahmaputra River, which needs to be further researched for seasonal variation.