ASSR-mediated sludge yield reduction couples deterministic enrichment of Nitrospira with metabolic resource partitioning

The anaerobic side-stream reactor (ASSR) process offers a microbiome-driven strategy for sustainable wastewater treatment, yet the ecological mechanisms governing its sludge yield reduction efficiency remain unresolved. Here, we demonstrate that a pilot-scale anaerobic-anoxic-oxic (AAO) system with integrated anaerobic side-stream reactor (ASSR) (designated AAO-ASSR/SR) reduced sludge production by 43.6 % compared to a conventional AAO system (designated AAO/CK), while maintaining effluent quality. Through integrated multi-omics and ecological modeling, we revealed the core microbiome-driven mechanism for ASSR-mediated sludge yield reduction. This mechanism is characterized by three key features: (1) enhanced microbial stability via cooperative networks, (2) deterministic assembly selecting slow-growing keystone taxa (e.g., Nitrospira, 18.6 % abundance in SR), and (3) metabolic resource partitioning from biomass synthesis to amino acid cross-feeding. Functional metagenomics revealed that Nitrospira (phylum Nitrospirota, comprising >99 % Nitrospira) and Novosphingobium (phylum Proteobacteria) mediated increased amino acid metabolism and reduced ATP biosynthesis in SR, contrasting with Bacteroidota-dominated biomass synthesis in CK through enhanced protein, nucleotide metabolism and ATP biosynthesis. By coupling deterministic microbial assembly with functional repartitioning, this work contributes to establish a design principle for targeted microbiome engineering in low-sludge systems, advancing sustainable wastewater management through ecological optimization of microbial resource allocation.