Single-stage nitritation-anammox combines the growth of aerobic ammonium-oxidizing bacteria (AOB) and anaerobic ammonium oxidizing bacteria (AnAOB) in one reactor. The necessary compromise of their milieu conditions often leads to the growth of nitrite-oxidizing bacteria (NOB). For this study, a sequencing batch reactor (SBR) for nitritation-anammox was operated for 180 days with sewage sludge reject water (removal capacity, 0.4 kg N m(-3) day(-1)). The growth of NOB was favored by enhanced oxygen supply rather than extended aerobic phases. Suspended-type biomass from this SBR was taken regularly and sieved into three size fractions (all of them <1,000 μm). Batch experiments as well as fluorescence in situ hybridization were performed to study the distribution and activity of AnAOB, AOB, and NOB within those size fractions. Both the measured conversion rates and detected abundances decreased with increasing size fraction. The highest anammox conversion rates (15 g NH4 (+)-N per kilogram VSS per hour) and the highest abundances of Brocadia fulgida were found in the medium size fraction (100-315 μm). The batch experiments proved to be accurate tools for the monitoring of multiple processes in the reactor. The results were representative for reactor performance during the 6 months of reactor operation.