The　construction　of　aerobic　denitrification　(AD)　systems　in　an　antibiotic-stressed　environment　is　a　serious　challenge.　This　study　investigated　strategy　of　cyclic　stress　with　concentration　gradient　(5–30　mg/L)　of　sulfamethoxazole　(SMX)　in　a　sequencing　batch　reactor　(SBR),　to　achieve　operation　of　AD.　Total　nitrogen　removal　efficiency　of　system　increased　from　about　10　%　to　95　%.　Original　response　of　abundant-rare　genera　to　antibiotics　was　changed　by　SMX　stress,　particularly　conditionally　rare　or　abundant　taxa　(CRAT).　AD　process　depends　on　synergistic　effect　of　heterotrophic　nitrifying　aerobic　denitrification　bacteria　(Paracoccus,　Thauera,　Hypomicrobium,　etc).　AmoABC,　napA,　and　nirK　were　functionally　co-expressed　with　multiple　antibiotic　resistance　genes　(ARGs)　(acrR,　ereAB,　and　mdtO),　facilitating　AD　process.　ARGs　and　TCA　cycling　synergistically　enhance　the　antioxidant　and　electron　transport　capacities　of　AD　process.　Antibiotic　efflux　pump　mechanism　played　an　important　role　in　operation　of　AD.　The　study　provides　strong　support　for　regulating　activated　sludge　to　achieve　in　situ　AD　function.　©　2024　Elsevier　Ltd