Anoxic and aerobic batch reactor assays using a two-by-two factorial design were developed to determine the potential activity of autotrophic denitrification and heterotrophic nitrification in biological nutrient removal (BNR) activated sludge systems, especially those accomplishing simultaneous BNR (SBNR). Results from the application of these assays to three full scale closed loop bioreactors previously documented to be accomplishing SBNR demonstrated that these activities were minimal in comparison with the conventionally recognized activities of heterotrophic denitrification and autotrophic nitrification. Activity within the mixed liquor consistent with current theories for phosphorus accumulating organisms (PAOs) was also observed. Along with other observations, this suggests the presence of PAOs in the facilities studied.