Assessing the viral enrichment methods and their roles in indicating wastewater-associated pollution in aquatic environments

With the increasing need for monitoring viral contamination in aquatic environments, research has increasingly focused on utilizing viruses as indicators for microbial contamination assessment. However, studies on enrichment techniques for waterborne viruses and their occurrence and risk characteristics in the environment remain relatively limited. In this study, samples were collected over one year from a wastewater treatment plant and its receiving stream to evaluate the optimal enrichment method and to assess the presence of four representative viral indicators-human adenovirus (HAdV), crAssPhage, enterovirus (EV), and the pepper mild mottle virus (PMMoV)-in the urban water environment. The results showed that skimmed milk flocculation (SMF) and silica attachment method (SAM) achieved better viral enrichment performance in both wastewater and surface water, demonstrating greater seasonal consistency compared to other methods. Seasonal variations in virus concentrations were observed, with HAdV and crAssphage peaking in winter, while EV and PMMoV peaked in summer. Virus concentrations in wastewater treatment plants were reduced by 8.61 log10 copies/L from influent to effluent. However, residual viruses discharged into receiving streams still pose a significant environmental exposure risk, as indicated by Quantitative Microbial Risk Assessment (QMRA) results, which exhibited a strong correlation with population density. This study highlights the importance of waterborne viral indicators in developing effective water quality management strategies to ensure the safe control of viruses in aquatic environments.