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Water quality in reclaimed distribution systems is shaped by multiple factors, particularly hydraulic stagnation in pipes and storage reservoirs. Stagnation can alter microbial community stability and facilitate persistence of pathogenic taxa. To investigate how prolonged stagnation affects microbial community structure and function, we integrated metagenomics and metaproteomics analyses of biofilms under flow and stagnant conditions over 3, 5 and 7\u2009months. Prolonged stagnation caused pronounced compositional shifts, including strong reductions in nitrogen\u2010removing taxa such as\n                    <jats:italic>Nitrospira<\/jats:italic>\n                    and\n                    <jats:italic>Nitrosomonas<\/jats:italic>\n                    . Correspondingly, key nitrification and denitrification proteins were depleted \u2265\u2009twofold under stagnation, indicating impaired nitrogen conversion processes. Stagnation also enriched motility\u2010 and transport\u2010related functions and promoted\n                    <jats:italic>Acidovorax<\/jats:italic>\n                    persistence, a genus including phytopathogenic species. In contrast, flow conditions sustained nitrogen\u2010cycling activity, contaminant\u2010degrading enzymes, and quorum\u2010quenching proteins, supporting greater biostability. Overall, our findings show that prolonged stagnation disrupts microbial community balance, suppresses essential nitrogen\u2010cycling and detoxification pathways, and reduces the functional robustness of treated wastewater. Maintaining hydraulic flow within reclaimed water systems is therefore critical for preserving microbial functionality and ensuring safe and reliable reuse in irrigation and other non\u2010potable applications.\n                  <\/jats:p>","DOI":"10.1111\/1462-2920.70372","type":"journal-article","created":{"date-parts":[[2026,7,14]],"date-time":"2026-07-14T14:32:49Z","timestamp":1784039569000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Prolonged Stagnation Reduces Treated Wastewater Biostability by Altering Microbial Community: Insights From Metaproteomics"],"prefix":"10.1111","volume":"28","author":[{"given":"Fatimah","family":"Almulhim","sequence":"first","affiliation":[{"name":"Bioscience Program, Biological and Environmental Science and Engineering Division King Abdullah University of Science and Technology (KAUST)  Thuwal Saudi 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