Genome-Resolved Metatranscriptomics Provide Insights on Immigration Influence in Structuring Microbial Community Assembly of a Full-Scale Aerobic Granular Sludge Plant

Understanding the relative influence of immigration and species sorting in wastewater treatment systems is essential, as bacteria in influent wastewater can significantly impact treatment system functionality. This study investigated the contribution of immigration to the community assembly of different-sized microbial aggregates in a full-scale aerobic granular sludge (AGS) system using genome-resolved metatranscriptomics. Our novel analysis revealed that negative-net-growth-rate populations, which persist due to immigration, can exhibit substantial activity and potentially contribute to the AGS system's functionality. The results also highlighted that sulfate-reducing and fermenting bacteria, along with some nitrifiers and glycogen-accumulating organisms (GAOs), were more active in the influent wastewater, serving as a continuous source of both beneficial and competing immigrants to the AGS system. Granular sludge (size >0.2 mm) demonstrated a robust capacity to resist immigration effects from competing immigrants, whereas flocculent sludge (size <0.2 mm) was more susceptible. Importantly, flocculent sludge harbored functional microbial groups such as active nitrifiers and fermentative polyphosphate-accumulating organisms (PAOs) belonging to <i>Ca. Phosphoribacter</i>, while granular sludge enriched for active conventional PAOs such as <i>Ca. Accumulibacter</i>. These findings provide valuable insights for engineers to design and operate AGS systems by optimizing microbial aggregate sizes and emphasizing the importance of influent microbial characterization in the design of wastewater treatment plants to enhance the functionality and activity of AGS systems.