Jatuwat Sangsanont

Norovirus, a leading cause of acute gastroenteritis, significantly impacts global health, especially in developing countries where foodborne and environmental transmission remain challenging. This study examines the occurrence and seasonality of norovirus genogroups I (GI) and II (GII), their inter-annual variation and the prevalence of pepper mild mottle virus (PMMoV) as a potential viral indicator in oysters. Oysters were collected from one coastal sampling area on the Perfume River estuary, Hue City, Vietnam—a region with a tropical climate and limited wastewater treatment—over a 31-month period from March 2019 to October 2021 (34 sampling events, 240 individual oysters). Quantitative polymerase chain reaction (qPCR) was used to analyse viral genomes. Norovirus GI (45.4%, 109/240) was detected at a significantly higher rate than GII (35.4%, 85/240; χ²(1) = 4.576, p = 0.032). No significant difference in detection was found between dry and wet seasons for either GI (F(1,19) = 1.474, p = 0.240) or GII (Welch’s F(1,11.7) = 0.248, p = 0.627). However, significant inter-annual differences in dry season detection were observed for both GI (F(2,13) = 7.102, p = 0.008) and GII (Welch’s F(2,7.7) = 32.653, p < 0.001), with markedly lower detection in 2020 that may be associated with COVID-19 restrictions rather than climate variability, as rainfall data analysis did not support a climatic explanation. PMMoV was consistently detected across seasons (95.0%, 228/240) but showed no significant correlation with norovirus GI (r = 0.272, p = 0.120) or GII (r = 0.150, p = 0.396), suggesting it may not be a reliable quantitative proxy for norovirus contamination in oysters in this tropical setting. These findings highlight the importance of continuous oyster-based viral surveillance and improved wastewater management to reduce health risks associated with shellfish consumption in tropical coastal regions.

Flow diagram showing sampling in Thailand, nucleic acid extraction, PMMoV qPCR detection, and comparison with ToBRFV and crAssphage across wastewater and animal feces. Pepper mild mottle virus (PMMoV) is one of the most abundant ribonucleic acid viruses in human feces and is widely used in wastewater-based epidemiology (WBE) and microbial source tracking. Its performance has not been evaluated in Thailand. This study analyzed archived nucleic acid extracts from sewage (n = 16), wastewater treatment plant (WWTP) influents (n = 16), and pooled animal feces (n = 35). PMMoV was detected in all sewage and WWTP samples, with concentrations of 4.26–5.83 log10 copies/100 mL in sewage (median 5.49) and 4.51–5.67 log10 copies/100 mL in WWTP influents (median 5.25). Only one pooled pig sample was positive (3.02 log10 copies/100 mg). PMMoV concentrations were higher than Tomato brown rugose fruit virus ToBRFV in sewage (p < 0.001) and WWTP influents (p < 0.05), and greater than crAssphage in WWTP influents (p < 0.001). Correlation analysis showed positive associations with ToBRFV (Spearman′s rho = 0.57–0.62), crAssphage (rho = 0.29–0.59), and Escherichia coli (rho = 0.25–0.52). These findings demonstrate that PMMoV is abundant in Thai wastewater, rarely detected in pooled animal feces, and is suitable as a human-specific marker and WBE normalizer.

Financial feasibility is usually a concern in water reclamation projects. Aside from internal benefits, water reclamation in industrial parks delivers health and environmental benefits not normally considered in cost–benefit analyses (CBA). This study investigated the influence of environmental benefits on the feasibility of water reclamation projects with flow rate scenarios in accordance with industrial parks in Chonburi, Thailand. CBAs of water reclamation plants for industrial water supply, consisting of ultrafiltration (UF) and reverse osmosis (RO), with flow rates of 5200, 10,000, 15,000, and 25,000 m³/day and discount rates of 3%, 5%, 7%, 9% and 11% were conducted. Considering only the direct costs and benefits, none of the projects were financially feasible. However, when the environmental benefits were included, the projects became profitable in all cases except those with a flow rate of 5200 m³/day and discount rates of 5%, 7%, 9%, and 11% and those with flow rates of 10,000 and 25,000 m³/day and an 11% discount rate. Further, CBAs of water reclamation projects in industrial parks for irrigation were conducted with post-treatment processes consisting of sand filtration and chlorine disinfection for flow rates of 240, 480, 2400, 3600, and 4800 m³/day. The projects are profitable, regardless of environmental benefits.

Aims A continuous quench‐flow (CQF) reactor was developed to collect samples at the reaction times of less than one second. The reactor is applied to determine ozone disinfection kinetics of poliovirus and to study whether EMA‐qPCR can assess the viral infectivity after ozone disinfection. Methods Ozone disinfection of poliovirus was conducted in the developed CQF, and the disinfection kinetics were tested in the range of 0.7 – 5.0 seconds at ozone concentration of 0.08 and 0.25 mg/L. Inactivation, damage on viral genome and damage on capsid integrity were determined by plaque assay, quantitative reverse transcription polymerase chain reaction (RT‐qPCR), and ethidium monoazide treatment coupled with RT‐qPCR (EMA‐qPCR), respectively. Results By using CQF, 2.18 and 2.76 log₁₀ reductions were observed at the reaction time of 0.7s and ozone concentration of 0.08 and 0.25 mg/L, respectively, followed by tailing. Ozone disinfection kinetics of poliovirus 1 were better fit by the efficiency factor Hom (EFH) model than by the Chick‐Watson model, or the modified Chick‐Watson model. Kinetics observed were similar between RT‐qPCR and EMA‐qPCR assays at the reaction times of less than 2.0 s and ozone concentrations of 0.08 and 0.25 mg/L. At reaction times > 5 seconds, viral concentration evaluated by EMA‐qPCR was reduced in comparison to stable RT‐qPCR results. Both assays still underestimated the virus inactivation. Conclusion The simple developed reactor can be used to investigate viral ozone disinfection kinetics and to elucidate inactivation characteristics or mechanisms at very short exposure times. Significance and Impact of the Study The developed CQF reactor is beneficial for better understanding of virus inactivation by ozone, and the reactor can be used to better elucidate disinfection kinetics and mechanisms for future research. This work constitutes an important contribution to the existing knowledge of the application and limitation of the EMA/PMA‐qPCR to assess virus infectivity after ozone disinfection.