Calcium nitrate as a bio-stimulant for anaerobic ammonium oxidation process

This study explored the role of calcium nitrate as a bio-stimulant for anaerobic ammonium oxidation (anammox) process. The anaerobic sequencing batch reactor was firstly inoculated with malodorous river sediment and only fed with calcium nitrate until no marked endogenous release of ammonium in effluent (Phase 1). Subsequently, nitrite and ammonium were supplied to test the performance of anammox process (Phase 2). During the operation of Phase 1, the effluent ammonium increased firstly and then decreased. Additionally, continuous nitrite (about 1.54 mgN/L) was observed in the effluent. The microbial analysis showed the simultaneous increase of the relative abundance of heterotrophic denitrifier Denitratisoma and sulfur autotrophic denitrifier Thiobacillus from 0.15% to 5.37% and 0.21% to 4.19%, respectively. Besides, ¹⁵N isotopes trace and qPCR results showed that the contribution of anammox to total nitrogen (TN) removal increased from 3.07% to 27.6%, and that the anammox functional gene hzsB increased from 1.37 × 10⁵ to 2.90 × 10⁶ copies/g. These results indicated that calcium nitrate may induce partial mixotrophic denitrification (heterotrophic and sulfur autotrophic denitrification) to provide nitrite as electron acceptor for anammox, thus promoting the occurrence of anammox. In Phase 2, rapid ammonium and TN removal were accomplished in the initial operation with the reduction efficiency of 80.1% and 90.0%, respectively. The relative abundance of anammox bacteria Candidatus_Brocadia significantly increased from 0.01% to 7.15% during the operation of Phase 2. These findings further confirmed the above deduction. Taken together, calcium nitrate can be a promising bio-stimulant for anammox process by promoting the coupling of mixotrophic denitrification with anammox.