In the Completely Autotrophic Nitrogen removal Over Nitrite (CANON) process, aerobic and anaerobic ammonia oxidizing bacteria cooperate to remove ammonia in one oxygen-limited reactor. Kinetic studies, microsensor analysis, and fluorescence in situ hybridization on CANON biomass showed a partial differentiation of processes and organisms within and among aggregates. Under normal oxygen-limited conditions (similar to5 muM O-2), aerobic ammonia oxidation (nitrification) was restricted to an outer shell (<100mum) while anaerobic ammonia oxidation (anammox) was found in the central anoxic parts. Larger type aggregates (>500 mum) accounted for 68% of the anammox potential whereas 65% of the nitrification potential was found in the smaller aggregates (<500 mum). Analysis with O-2 and NO2- microsensors showed that the thickness of the activity zones varied as a function of bulk O-2 and NO2- concentrations and flow rate. (C) 2004 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.