Activity and growth of anammox biomass on aerobically pre-treated municipal wastewater

Direct treatment of municipal wastewater (MWW) based on anaerobic ammonium oxidizing (anammox) bacteria holds promise to turn the energy balance of wastewater treatment neutral or even positive. Currently, anammox processes are successfully implemented at full scale for the treatment of high-strength wastewaters, whereas the possibility of their mainstream application still needs to be confirmed. In this study, the growth of anammox organisms on aerobically pre-treated municipal wastewater (MWW(pre-treated)), amended with nitrite, was proven in three parallel reactors. The reactors were operated at total N concentrations in the range 5-20 mg(N)∙L(-1), as expected for MWW. Anammox activities up to 465 mg(N)∙L(-1)∙d(-1) were reached at 29 °C, with minimum doubling times of 18 d. Lowering the temperature to 12.5 °C resulted in a marked decrease in activity to 46 mg(N)∙L(-1)∙d(-1) (79 days doubling time), still in a reasonable range for autotrophic nitrogen removal from MWW. During the experiment, the biomass evolved from a suspended growth inoculum to a hybrid system with suspended flocs and wall-attached biofilm. At the same time, MWW(pre-treated) had a direct impact on process performance. Changing the influent from synthetic medium to MWW(pre-treated) resulted in a two-month delay in net anammox growth and a two to three-fold increase in the estimated doubling times of the anammox organisms. Interestingly, anammox remained the primary nitrogen consumption route, and high-throughput 16S rRNA gene-targeted amplicon sequencing analyses revealed that the shift in performance was not associated with a shift in dominant anammox bacteria ("Candidatus Brocadia fulgida"). Furthermore, only limited heterotrophic denitrification was observed in the presence of easily biodegradable organics (acetate, glucose). The observed delays in net anammox growth were thus ascribed to the acclimatization of the initial anammox population or/and the development of a side population beneficial for them. Additionally, by combining microautoradiography and fluorescence in situ hybridization it was confirmed that the anammox organisms involved in the process did not directly incorporate or store the amended acetate and glucose. In conclusion, these investigations strongly support the feasibility of MWW treatment via anammox.