Protozoan Communities and Their Contribution to Predation on E. coli in Aerobic Granular Sludge

Protozoa contribute to water purification through predation in wastewater treatment systems. Full-scale aerobic granular sludge (AGS) reactors treating municipal wastewater contain AGS of varying sizes, with those larger than 2 mm dominating. These size fractions exhibit different sludge morphologies and microbial communities. To date, little is known about protozoan communities and their role in the removal of human-associated bacteria (like pathogens) in AGS plants, particularly across different size fractions. This study conducted uptake experiments with fluorescent Escherichia coli, as a model for human-associated bacteria, followed by microscopic observation to investigate protozoan communities and their predatory behavior in six AGS size fractions and activated sludge collected from full-scale municipal wastewater treatment plants. Sessile ciliates, particularly Epistylis and Vorticella, dominated protozoan populations across six AGS size fractions, with Epistylis being more abundant in larger AGS fractions (>1 mm) and Vorticella in smaller fractions (<1 mm). Additionally, microcosm experiments under aerobic (including predation) and anoxic conditions (excluding predation) revealed that predation was likely to be the main E. coli removal pathway, contributing an additional 0.5 to 2.5 log₁₀ CFU mL^-1 reduction over a combination of non-predatory biological and abiotic processes. Larger AGS fractions showed greater predation capacity, linked to higher Epistylis abundance, while activated sludge, dominated by Vorticella, resembled smaller AGS fractions with lower predation capacity. These findings advance the understanding of the distribution of protozoan communities and their contribution to E. coli removal by predation in AGS wastewater treatment.