Defluviicoccus vanus-related glycogen accumulating organisms (GAO) regularly proliferate in industrial wastewater treatment plants handling high carbon but nitrogen deficient wastes. When GAO dominate, they are associated with poor performance, characterised by slow settling biomass and turbid effluents. Although their ecophysiology has been studied thoroughly in domestic waste treatment plants, little attention has been paid to them in aerobic industrial systems. In this study, the effect of nitrogen addition on GAO carbon metabolism was investigated during an 8 h cycle. Activated sludge dominated by GAO from a winery wastewater sequencing batch reactor was incubated under different carbon to nitrogen (COD:N) ratios (100:1, 60:1 and 20:1) using 13C — acetate and 15N — urea. GAO cell assimilation was quantified using FISH-NanoSIMS. The activated sludge community was assessed by 16S rRNA gene profiling, DNA and storage polymer production. Carbon and nitrogen quantification at the cellular level by NanoSIMS revealed that low (COD:N of 100:1) or null nitrogen concentrations enhanced GAO carbon uptake. COD:N ratios of 60:1 and 20:1 reduced GAO carbon uptake and promoted whole microbial community DNA production. Nitrogen dosing at COD:N ratios of 60:1 or higher was demonstrated as feasible strategy for controlling the excessive GAO growth in high COD waste treatment plants.