Long-term multi-meta-omics resolves the ecophysiological controls of seasonal NO emissions during wastewater treatment.

Nitrous oxide (NO) is the third most important greenhouse gas and originates primarily from natural and engineered microbiomes. Effective emission mitigations are currently hindered by the largely unresolved ecophysiological controls of coexisting NO-converting metabolisms in complex communities. To address this, we used biological wastewater treatment as a model ecosystem and combined long-term metagenome-resolved metaproteomics with ex situ kinetic and full-scale operational characterization over nearly 2 years. By leveraging the evidence independently obtained at multiple ecophysiological levels, from individual genetic potential to actual metabolism and emergent community phenotype, the cascade of environmental and operational triggers driving seasonal NO emissions has ultimately been resolved. We identified nitrifier denitrification as the dominant NO-producing pathway and dissolved O as the prime operational parameter, paving the way to the design and fostering of robust emission control strategies. This work exemplifies the untapped potential of multi-meta-omics in the mechanistic understanding and ecological engineering of microbiomes towards reducing anthropogenic impacts and advancing sustainable biotechnological developments.