The aim was to verify effect of sulphides on the course and effectiveness of autotrophic and heterotrophic denitrification by non-adapted activated sludge. Joint application of organic substrate and sulphides indicates heterotrophic and autotrophic denitrification does not run simultaneously, but the organic substrate is used preferably and only after exhaustion of organic substrate autotrophic denitrification with sulphide proceeds. Results offer a possible solution for wastewater with a lack of organic carbon as well as for some sources of dissolved sulphides available especially from external desulphurization of biogas.