This　study　constructed　an　integrated　algae/partial　nitrification/anammox　biofilm　system　and　operated　it　for　240　days.　The　total　nitrogen　removal　efficiency　exceeded　90　%.　The　structure,　compositions,　and　function　of　this　symbiotic　biofilm,　which　played　a　pivotal　role　in　the　system,　were　analyzed　in　detail.　Microscope　photos　and　fluorescence　in　situ　hybridization　both　showed　that　bacteria　and　algae　were　well　integrated.　The　dissolved　oxygen　gradient　further　confirmed　that　different　functional　microorganisms　grew　at　varying　depths　within　biofilm.　Algae　formed　an　oxygen-producing　zone　(0–0.48　mm),　followed　by　ammonia　oxidizing　bacteria　(AOB)　consuming　oxygen　to　form　an　oxygen-consuming　zone　(0.48–0.86　mm),　and　anaerobic　ammonia　oxidizing　bacteria　(AnAOB)　removed　nitrogen　in　anaerobic　zone　(＞0.86　mm).　Chlorella,　Nitrosomonas　and　Candidatus_Kuenenia　were　identified　as　the　dominant　algae,　AOB　and　AnAOB,　with　relative　abundances　of　11.80　%,　19.77　%　and　3.07　%,　respectively.　This　layered　biofilm　benefitted　providing　a　suitable　environment　for　various　microorganisms　to　survive　within　a　complex　biofilm.　©　2024　Elsevier　Ltd