This study explores the influence of variable aeration strategies on microbial interactions within the partial nitritation/anammox (PN/A) process. To overcome the limitations of relying solely on microbial abundance, this research integrates high-throughput DNA sequencing, reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and batch tests to link microbial function and adaptation to variable aeration strategies. Four operational stages were investigated: continuous aeration with a dissolved oxygen (DO) concentration of 1.5 mg O₂/L; intermittent aeration at a ratio of 1:2 with the same DO; increased DO to 2.5 mg O₂/L while maintaining the intermittent regime; and equal duration of aerated and non-aerated phases (1:1) with DO at 2.5 mg O₂/L. Among these, the third stage intermittent aeration at 2.5 mg O₂/L proved to be the most favorable for the PN/A process, yielding the highest anammox activity and effective suppression of nitrite-oxidizing bacteria. Results show that the rate of nitrogen conversion is related to the particular structure of microbial communities, as revealed by network analysis. Importantly, even low-abundance functional groups contributed significantly to nitrogen removal under favorable environmental conditions. The combined methodological approach allowed for an effective correlation between microbial structure and activity, offering a valuable framework for interpreting microbial responses to operational changes.
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