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In this study, the effect of mixotrophic, heterotrophic, photoautotrophic (CO2of air source), and photoautotrophic (bicarbonate source) cultivation of Chlorella vulgariswas investigated on microalgae growth rate and nutrient removal under different kinds of nitrogen sources. The highest N–NH3+, N–NO3−, P–PO43−(in ammonium source), and P–PO43−(in nitrate source) removal efficiency (87.28 ± 0.89, 70.00 ± 1.20, 87.00 ± 3.10, and 78.10 ± 1.95%, respectively) was reached in the mixotrophic culture. In all cultivations, when nitrate was used as the nitrogen source, specific growth rate, biomass productivity, and specific oxygen production rate (SOPR) were higher than the one with ammonium source due to decreased pH. However, in all cultures except for photoautotrophic cultivation (in the CO2of the air), nitrogen removal rates improved in the ammonium source. In addition, in both ammonium and nitrate source, the respiration activity of mixotrophic microalgae was higher than the photosynthetic activity of the cells. When CO2was used as inorganic carbon, the specific growth rate of microalgae was lower than with the bicarbonate source. Based on the obtained results, mixotrophic conditions would be the most useful cultivation for application in N-rich wastewater treatment systems.