The　discovery　of　complete　ammonium　oxidation　(comammox)　has　subverted　the　traditional　perception　of　two-step　nitrification,　which　plays　a　key　role　in　achieving　biological　nitrogen　removal　from　wastewater.　Floccular　sludge-based　treatment　technologies　are　being　applied　at　the　majority　of　wastewater　treatment　plants　in　service　where　detection　of　various　abundances　and　activities　of　comammox　bacteria　have　been　reported.　However,　limited　efforts　have　been　made　to　enrich　and　subsequently　characterize　comammox　bacteria　in　floccular　sludge.　To　this　end,　a　lab-scale　sequencing　batch　reactor　(SBR)　in　the　step-feeding　mode　was　applied　in　this　work　to　enrich　comammox　bacteria　through　controlling　appropriate　operational　conditions　(dissolved　oxygen　of　0.5　±　0.1　g-O2/m3,　influent　ammonium　of　40　g-N/m3　and　uncontrolled　longer　sludge　retention　time).　After　215-d　operation,　comammox　bacteria　gradually　gained　competitive　advantages　over　counterparts　in　the　SBR　with　a　stable　nitrification　efficiency　of　92.2　±　2.2　%:　the　relative　abundance　of　Nitrospira　reached　42.9　±　1.3　%,　which　was　13　times　higher　than　that　of　Nitrosomonas,　and　the　amoA　gene　level　of　comammox　bacteria　increased　to　7.7　±　2.1　×　106　copies/g-biomass,　nearly　50　times　higher　than　that　of　conventional　ammonium-oxidizing　bacteria.　The　enrichment　of　comammox　bacteria,　especially　Clade　A　Candidatus　Nitrospira　nitrosa,　in　the　floccular　sludge　led　to　(i)　apparent　affinity　constants　for　ammonium　and　oxygen　of　3.296　±　0.989　g-N/m3　and　0.110　±　0.004　g-O2/m3,　respectively,　and　(ii)　significantly　low　N2O　and　NO　production,　with　emission　factors　being　0.136　±　0.026　%　and　0.023　±　0.013　%,　respectively.　©　2024　Elsevier　Ltd