The coagulation process for enveloped and non-enveloped virus removal in turbid water: Removal efficiencies, mechanisms and its application to SARS-CoV-2 Omicron BA.2.

The coagulation process has a high potential as a treatment method that can handle pathogenic viruses including emerging enveloped viruses in drinking water treatment process which can lower infection risk through drinking water consumption. In this study, a surrogate enveloped virus, bacteriophage Փ6, and surrogate non-enveloped viruses, including bacteriophage MS-2, T4, ՓX174, were used to evaluate removal efficiencies and mechanisms by the conventional coagulation process with alum, poly­aluminum chloride, and ferric chloride at pH 5, 7, and 9 in turbid water. Also, treatability of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a recent virus of global concern by coagulation was evaluated as SARS-CoV-2 can presence in drinking water sources. It was observed that an increase in the coagulant dose enhanced the removal efficiency of turbidity and viruses, and the condition that provided the highest removal efficiency of enveloped and non-enveloped viruses was 50 mg/L of coagulants at pH 5. In addition, the coagulation process was more effective for enveloped virus removal than for the non-enveloped viruses, and it demonstrated reduction of SARS-CoV-2 Omicron BA.2 over 0.83-log with alum. According to culture- and molecular-based assays (qPCR and CDDP-qPCR), the virus removal mechanisms were floc adsorption and coagulant inactivation. Through inactivation with coagulants, coagulants caused capsid destruction, followed by genome damage in non-enveloped viruses; however, damage to a lipid envelope is suggested to contribute to a great extend for enveloped virus inactivation. We demonstrated that conventional coagulation is a promising method for controlling emerging and re-emerging viruses in drinking water.

Removal of antibiotics, bacterial toxicity, and occurrence of antibiotic resistance genes in secondary hospital effluents treated with granular activated carbon and the impact of preceding chlorination.

This comprehensive investigation highlighted the complex adsorption behaviors of antibiotics onto granular activated carbon (GAC), the effectiveness of this adsorption in reducing bacterial toxicity, and the reduction of antibiotic resistance genes (ARGs) and antibiotic resistant bacteria (ARB) in hospital wastewater (HWW) effluents. Six GACs were characterized for their physicochemical properties, and their ability to adsorb six antibiotics in the background matrix of actual HWW was evaluated. Coconut shell-derived GAC (Co-U), which had the highest hydrophobicity and lowest content of oxygen-containing acidic functional groups, demonstrated the highest adsorption capacities for the tested antibiotics. Bacterial toxicity tests revealed that GACs could eliminate the bacterial toxicity from antibiotic intermediates present in chlorinated HWW. By contrast, the bacterial toxicity could not be removed by GACs in non-chlorinated HWW due to the greater presence of intermediate components identified by LC-MS/MS. The intraparticle diffusion coefficient of antibiotics adsorbed onto Co-U could be calculated by adsorption kinetics derived from the linear driving force model and the homogenous intraparticle diffusion model associated with the linear adsorption isotherms (0-150 µg/L). Meropenem and sulfamethoxazole exhibited the highest adsorption capacities in a single-solute solution compared to penicillin G, ampicillin, cetazidime, and ciprofloxacin. However, the greater adsorption capacities of meropenem and sulfamethoxazole disappeared in mixed-solute solutions, indicating the lowest adsorption competition. GAC can eliminate most ARGs while also promoting the growth of some ARB. Chlorination (free chlorine residues at 0.5 mg Cl2/L) did not significantly affect the overall composition of ARGs and ARB in HWW. However, the accumulation of ARGs and ARB on GAC in fixed bed columns was lower in chlorinated HWW than in non-chlorinated HWW due to an increase in the adsorption of intermediates.

Detection of enteroviruses related to hand foot and mouth disease in wastewater of Asian communities.

Hand, foot, and mouth disease (HFMD) is contagious and predominantly affects children below the age of five. HFMD-associated serotypes of Enterovirus A (EVA) family include EVA71, Coxsackievirus A type 6 (CVA6), 10 (CVA10), and 16 (CVA16). Although prevalent in numerous Asian countries, studies on HFMD-causing agents in wastewater are scarce. This study aimed to conduct wastewater surveillance in various Asian communities to detect and quantify serotypes of EVA associated with HFMD. In total, 77 wastewater samples were collected from Indonesia, the Philippines, Thailand, and Vietnam from March 2022 to February 2023. The detection ratio for CVA6 RNA in samples from Vietnam was 40 % (8/20). The detection ratio for CVA6 and EVA71 RNA each was 25 % (5/20) for the Indonesian samples, indicating the need for clinical surveillance of CVA6, as clinical reports have been limited. For the Philippines, 12 % (2/17) of the samples were positive for CVA6 and EVA71 RNA each, with only one quantifiable sample each. Samples from Thailand had a lower detection ratio (1/20) for CVA6 RNA, and the concentration was unquantifiable. Conversely, CVA10 and CVA16 RNAs were not detected in any of the samples. The minimum and maximum concentrations of CVA6 RNA were 2.7 and 3.9 log10 copies/L and those for EVA71 RNA were 2.5 and 4.9 log10 copies/L, respectively. This study underscores the importance of wastewater surveillance in understanding the epidemiology of HFMD-associated EVA serotypes in Asian communities. Long-term wastewater surveillance is recommended to monitor changes in dominant serotypes, understand seasonality, and develop effective prevention and control strategies for HFMD.

Wastewater monitoring in tourist cities as potential sentinel sites for near real-time dynamics of imported SARS-CoV-2 variants.

Wastewater-based epidemiology (WBE) complements the clinical surveillance of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its variants' distribution in populations. Many developed nations have established national and regional WBE systems; however, governance and budget constraints could be obstacles for low- and middle-income countries. An urgent need thus exists to identify hotspots to serve as sentinel sites for WBE. We hypothesized that representative wastewater treatment plants (WWTPs) in two international gateway cities, Bangkok and Phuket, Thailand, could be sentineled for SARS-CoV-2 and its variants to reflect the clinical distribution patterns at city level and serve as early indicators of new variants entering the country. Municipal wastewater samples (n = 132) were collected from eight representative municipal WWTPs in Bangkok and Phuket during 19 sampling events from October 2021 to March 2022, which were tested by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) using the US CDC N1 and N2 multiplex and variant (Alpha, Delta, and Omicron BA.1 and BA.2) singleplex assays. The variant detection ratios from Bangkok and Phuket followed similar trends to the national clinical testing data, and each variant's viral loads agreed with the daily new cases (3-d moving average). Omicron BA.1 was detected in Phuket wastewater prior to Bangkok, possibly due to Phuket's WWTPs serving tourist communities. We found that the Omicron BA.1 and BA.2 viral loads predominantly drove the SARS-CoV-2 resurgence. We also noted a shifting pattern in the Bangkok WBE from a 22-d early warning in early 2021 to a near real-time pattern in late 2021. The potential application of tourist hotspots for WBE to indicate the arrival of new variants and re-emerging or unprecedented infectious agents could support tourism-dependent economies by complementing the reduced clinical regulations while maintaining public health protection via wastewater surveillance.

UV inactivation of sewage isolated human adenovirus.

Adenoviruses are known to be one of the most resistant viruses to UV disinfection. This study determined the inactivation kinetics of adenovirus freshly isolated from sewage samples, and compared the results with reference adenovirus stocks grown in the laboratory. Human adenoviruses were isolated from sewage samples using the HEK 293 cell line. Inactivation kinetics for UV irradiation was determined for monochromatic low pressure (LP) mercury UV lamp (254 nm) and polychromatic medium pressure (MP) mercury UV lamp for each sewage isolate. Eleven (11) isolates were obtained from nine (9) different sewage samples with most isolates belonging to the enteric adenovirus group, specifically adenovirus 41. The average dose required for 4 log inactivation using LP UV lamps for sewage isolates (220 mJ/cm2) was not significantly different (p > 0.1) from the average dose reported for lab-grown enteric adenovirus (179.6 mJ/cm2). Interestingly, the average dose required for 4 log inactivation using MP UV lamps was significantly higher (p = 0.004) for sewage isolates (124 mJ/cm2) when compared to the average dose reported for laboratory stocks of adenovirus 40 and 41 (71 mJ/cm2). Viral capsid analysis using the propidium monoazide (PMA)-qPCR method showed that adenovirus isolates from group F were less affected by exposure to MP UV Lamps than adenoviruses from group D and C. Adenovirus isolates obtained from sewage samples showed greater resistance to UV irradiation compared to laboratory grown strains, although required doses for MP UV were still considerably lower than LP UV. These data suggest that the required fluence for inactivation of adenoviruses in real-world waters may be higher than previously understood.

Capsid integrity detection of pathogenic viruses in waters: Recent progress and potential future applications.

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.

SARS-CoV-2 RNA surveillance in large to small centralized wastewater treatment plants preceding the third COVID-19 resurgence in Bangkok, Thailand.

Wastewater surveillance for SARS-CoV-2 RNA has been a successful indicator of COVID-19 outbreaks in populations prior to clinical testing. However, this has been mostly conducted in high-income countries, which means there is a dearth of performance investigations in low- and middle-income countries with different socio-economic settings. This study evaluated the applicability of SARS-CoV-2 RNA monitoring in wastewater (n = 132) to inform COVID-19 infection in the city of Bangkok, Thailand using CDC N1 and N2 RT-qPCR assays. Wastewater influents (n = 112) and effluents (n = 20) were collected from 19 centralized wastewater treatment plants (WWTPs) comprising four large, four medium, and 11 small WWTPs during seven sampling events from January to April 2021 prior to the third COVID-19 resurgence that was officially declared in April 2021. The CDC N1 assay showed higher detection rates and mostly lower Ct values than the CDC N2. SARS-CoV-2 RNA was first detected at the first event when new reported cases were low. Increased positive detection rates preceded an increase in the number of newly reported cases and increased over time with the reported infection incidence. Wastewater surveillance (both positive rates and viral loads) showed strongest correlation with daily new COVID-19 cases at 22-24 days lag (Spearman's Rho = 0.85-1.00). Large WWTPs (serving 432,000-580,000 of the population) exhibited similar trends of viral loads and new cases to those from all 19 WWTPs, emphasizing that routine monitoring of the four large WWTPs could provide sufficient information for the city-scale dynamics. Higher sampling frequency at fewer sites, i.e., at the four representative WWTPs, is therefore suggested especially during the subsiding period of the outbreak to indicate the prevalence of COVID-19 infection, acting as an early warning of COVID-19 resurgence.

Detection of pepper mild mottle virus as an indicator for drinking water quality in Hanoi, Vietnam, in large volume of water after household treatment.

The aims of this study were to examine the removal of bacteria and viruses by household point-of-use (POU) treatments and to apply a previously developed large-volume virus concentration method (∼20 L). First, the removal of microbes by household POU treatment was investigated in the laboratory. Second, the prevalence of viruses in drinking water sources for households and the removal efficiency of microbes by POU treatments in two suburban communities in Hanoi, Vietnam, were investigated. Indigenous pepper mild mottle virus (PMMoV) was used as the main target together with adenovirus, Aichi virus, enterovirus, F-specific bacteriophage genogroup 1, and Escherichia coli to investigate the removal efficiency of household treatments. The results from laboratory and field survey were compared. From the laboratory study, ceramic membranes were not effective for removing viruses and bacteria from water; pathogen reduction was less than 1.5 log10. By contrast, reverse osmosis (RO) devices reduced microbes by 3 to > 5 log10. In a field study, PMMoV was found to be the most prevalent waterborne virus. Household sand filtration was ineffective for removing E. coli, total coliforms and PMMoV; the reduction was less than 1 order of magnitude. Boiling the water and then filtering it with a ceramic membrane reduced E. coli by 3 orders of magnitude, but this was not effective for removing PMMoV. RO filtration was one of the promising methods for removing E. coli, total coliforms and PMMoV to below their detection limits in most of the samples studied. The removal of E. coli, total coliforms and PMMoV was >2.3, >4 and >3 log10, respectively. The laboratory results of virus removal efficiency by POU devices agreed with the field study. Due to the prevalence and characteristics of PMMoV, it is a strong candidate for an indigenous indicator to investigate the viral removal efficiency of household POU treatments.