Virus Inactivation during Water Treatment

Virus removal and inactivation is still a major challenge for water treatment facilities in both industrialised nations and developing countries. This may seem surprising as chlorine disinfection started to spread broadly over a century ago. However, many viruses are more resistant to disinfection by chlorine and other oxidants than other pathogens. Additionally, some viruses are known to be particularly difficult to disinfect by UV. Finally, viruses are extremely small (18-120 nm diameter), which makes sedimentation impossible and filtration difficult. As no real-time methods exist to enumerate viruses, disinfectant doses are based on lab experiments typically conducted with dispersed viruses. However, viruses in wastewater and natural environments can be present as aggregates. Previous studies have shown that aggregates protect viruses from disinfection, but it remains unclear what renders these aggregates more resistant compared to dispersed viruses. In order to elucidate this observation, aggregates of bacteriophage MS2 of well-defined sizes up to 1 μm diameter were produced by lowering the solution pH, and aggregates were inactivated by peracetic acid (PAA). Aggregates were re-dispersed before enumeration to obtain the residual number of individual infectious viruses. In contrast to enumerating whole aggregates, this approach allowed an assessment of disinfection efficiency which remains applicable even if the aggregates disperse in post-treatment environments. Aggregation reduced the apparent inactivation rate constants 2-6 fold, depending on the aggregate size. The larger the aggregate and the higher the PAA concentration, the more pronounced was the inhibitory effect of aggregation on disinfection. A reaction diffusion model, developed to simulate aggregate disinfection, showed that the inhibitory effect of aggregation arises from consumption of the disinfectant within the aggregate, but that diffusion of the disinfectant into the aggregates is not a rate-limiting factor. Aggregation therefore has a large inhibitory effect if highly reactive disinfectants are used, whereas inactivation by mild disinfectants is less affected. This finding leads to the counterintuitive notion that mild disinfectants, rather than aggressive ones, should be used when virus aggregates are present. During UV disinfection, viruses disinfection curves frequently exhibit a tailing after an initial exponential decay. Aggregation, light shielding, genome recombination or resistant virus sub-populations were proposed as explanations. However, none of these options has conclusively been demonstrated. We investigated how aggregation affects virus inactivation by UV254 in general, and the tailing phenomena in particular. A similar experimental set-up was used as described above with the difference that UV254 disinfection was applied instead of PAA addition. Results showed that initial inactivation kinetics were similar for viruses incorporated in aggregates and dispersed viruses. However, aggregated viruses started to tail more readily than dispersed ones. Neither light shielding, nor the presence of resistant sub-populations could account for the tailing. Instead, tailing was consistent with genome recombination arising as a result of the simultaneous infection of the host by several impaired viruses. We argue that UV254 treatment of aggregates permanently fuses a fraction of viruses, which increased the likelihood of multiple infection of a host cell and ultimately enables the production of infective viruses via recombination. Our results suggest that UV disinfection followed by the addition of a mild disinfectant should yield efficient disinfection for waters containing viral aggregates. Outside Europe and North America wastewater is rarely efficiently treated as treatment costs are often too high and additionally require highly-skilled personnel. To mitigate this issue, waste stabilisation ponds (WPSs) are a viable option because both construction and maintenance are inexpensive and easy to be executed. Pathogen inactivation in these pond systems is mainly attributed to sunlight. Although viruses are among the most resistant pathogens, little is known about sunlight-mediated inactivation mechanisms for these pathogens. Viruses can either be inactivated directly by UV light absorption by the viral genomes, or indirectly via light absorption by sensitizers. The excited sensitizers lead to the formation of reactive species, typically oxidants, which can further react with the viruses and lead to inactivation. Two bacteriophages were chosen as model viruses to investigate these inactivation mechanisms; phiX174, which is resistant to oxidants, and MS2, which is relatively resistant to direct UV inactivation. The efficiency of direct inactivation by solar UVB light was determined, and the rate constants associated with the inactivation by four potentially important reactive species present in sunlit surface waters (singlet oxygen, triplet state organic matter, hydroxyl and carbonate radicals) were quantified. A model was developed that computes the contribution of each of these inactivation mechanisms for ponds with different depths and solution parameters. Direct inactivation was found to be the major inactivation mechanism for phiX174 and also X contributed importantly to inactivation of MS2. Singlet oxygen was the most important reactive species for both virus below 0.5 m. Nevertheless, it did not contribute significantly to the overall inactivation of phiX174. These results suggest that maturation ponds should either be shallow or continuously-mixed to achieve good disinfection results. They furthermore demonstrate that virus inactivation can be reasonably approximated based on easy to determine solution parameters, along with information regarding the sensitivity to viruses toward direct inactivation and few selected reactive species.