Quantification of Particle-Associated Viruses in Secondary Treated Wastewater Effluent

Viruses can interact with a broad range of inorganic and organic particles in water and wastewater. These associations can protect viruses from inactivation by quenching chemical disinfectants or blocking ultraviolet light transmission, and a much higher dosage of disinfectants is required to inactivate particle-associated viruses than free viruses. There have been only few studies of the association of viruses with particles in wastewater, particularly in secondary treated effluent. As secondary effluent is the source water to the reclaimed water treatment system, this study quantified indigenous enteric viruses, and viral indicators associated with particles in secondary effluents collected from five full-scale water reclamation facilities in the United States. Particle-associated viruses were enumerated using a sequential filtration followed by microfluidic digital PCR. This study showed that enteric viruses and viral indicators (crAssphage and pepper mild mottle virus, PMMoV) were attached to particles of different sizes in secondary effluent. Significantly higher concentrations of RNA viruses including PMMoV, norovirus, and enterovirus were detected in filtrate of the sequential filtration, which contained particles < 0.45 µm. DNA viruses including adenovirus and crAssphage were found to be more associated with larger particles in secondary effluent. Overall, high correlations were observed between viral indicators and enteric viruses, supporting the use of crAssphage and PMMoV to evaluate virus removal efficiency in water and wastewater treatment processes. The association of viruses with particles in wastewater has significant implications on wastewater treatment and disinfection processes as well as virus enumeration in wastewater.