This study describes the role of microorganisms in a full-scale step-feed sequencing batch reactor (SBR) system for urban wastewater treatment. Chemical profiles for three different cycle times were measured under low aeration conditions with a high carbon-to-nitrogen ratio. The applied organic load was above 1.0 g chemical oxygen demand (COD)/L x d. The removal efficiencies were higher than 81%, 93%, and 76% for soluble COD, N-NH4+, and total Kjeldahl nitrogen, respectively. The ratio of volatile suspended solids (VSS) to total suspended solids was 78%, and the food-to-microorganism ratio was an average of 1.41 g COD/g VSS x d. The active biomass was comprised of 87.8% heterotrophic and 12.2% autotrophic organisms. Nitrifying organisms were found with a low amount of ammonia-oxidizing bacteria (5%) and a much higher amount of nitrite-oxidizing bacteria. Polyphosphate-accumulating organisms (PAOs) were found at high amounts (25%) compared to glycogen-accumulating organisms, even in a system with a high carbon to phosphorus ratio. The activity of denitrifying PAOs was 72%.