Chlorine Disinfection of Aggregated Adenovirus 2 in Source Water

Most viral inactivation studies have been conducted using purified, dispersed viral particles. However, it is widely believed that viral particles are less likely to be present in a dispersed state in source water than in viral aggregates and attached to other particles. More information is needed in order to understand the phenomenon of viral aggregation and to determine whether CT values (disinfectant concentration × exposure in min) generated from dispersed viral disinfection studies are sufficient for aggregated viruses. In this study, the extent of viral aggregation was measured at different pH and specific conductance levels to determine the impact of these water quality parameters on aggregation state and chlorine disinfection CT values. Disinfection experiments with human adenovirus 2 (HAdV2) in source water were conducted using 0.2 mg/L free chlorine at 5°C. The pH and ionic strength were adjusted to either pH 7 or 8 and 300 or 1,000 μS specific conductance. Experimental flasks were seeded with semi-purified aggregated or monodispersed HAdV2 preparations. Viral titers before and after disinfection were determined by cell culture plaque assays. The efficiency factor Hom (EFH) model was used to calculate CT values required to achieve 2-, 3-, and 4-log10 reductions in viral titers. Electron microscopic analysis of the aggregated virus stock and selected experimental samples was performed. The passive Sharp/Miller kinetic sedimentation particle counting procedure was used to make quantitative assessments of the character and number of particles. When diluted into source water, percent aggregation of HAdV2 (≥3 aggregated particles) was greater at higher specific conductance and pH levels. However, aggregation percentage was highly variable and ranged from 43-71%. Upon addition into source water at the start of disinfection experiments, the aggregation percentage dropped dramatically, but did not usually drop below 11% aggregation. On average, CT values for 3-log10 inactivation of aggregated HAdV2 were 2 times higher than those for dispersed HAdV2, indicating that aggregation substantially impacted disinfection rate. This information can be used by water utilities and regulators to guide decision making regarding disinfection of aggregated viruses in source water.