It is a challenge to apply anaerobic ammonium oxidation (anammox) for nitrogen removal from wastewater at low temperatures. Maintenance of anammox- and aerobic ammonia oxidizing bacteria (AOB) and suppression of nitrite oxidizing bacteria (NOB) are key issues. In this work, a nitritation-anammox moving bed biofilm pilot reactor was operated at 19-10°C for 300d. Nitrogen removal was decreasing, but stable, at 19-13°C. At 10°C removal became unstable. Quantitative PCR, fluorescence in situ hybridization and gene sequencing showed that no major microbial community changes were observed with decreased temperature. Anammox bacteria dominated the biofilm (0.9-1.2×1014 16S rRNA copies m-2). Most anammox bacteria were similar to Brocadia sp. 40, but another smaller Brocadia population was present near the biofilm-water interface, where also the AOB community (Nitrosomonas) was concentrated in thin layers (1.8-5.3×1012 amoA copies m-2). NOB (Nitrobacter, Nitrospira) were always present at low concentrations (<1.3×1011 16S rRNA copies m-2).
Partial nitritation-anammox (PNA) permits energy effective nitrogen removal. Today PNA is used for treatment of concentrated and warm side streams at wastewater treatment plants, but not the more diluted and colder main stream. To implement PNA in the main stream, better knowledge about microbial communities at the typical environmental conditions is necessary. In order to investigate the response of PNA microbial communities to decreasing substrate availability, we have operated a moving bed biofilm reactor (MBBR) at decreasing reactor concentrations (311-27mg-N l(-1) of ammonium) and low temperature (13 degrees C) for 302days and investigated the biofilm community using high throughput amplicon sequencing; quantitative PCR; and fluorescence insitu hybridization. The anammox bacteria (Ca. Brocadia) constituted a large fraction of the biomass with fewer aerobic ammonia oxidizing bacteria (AOB) and even less nitrite oxidizing bacteria (NOB; Nitrotoga, Nitrospira and Nitrobacter). Still, NOB had considerable impact on the process performance. The anammox bacteria, AOB and NOB all harboured more than one population, indicating some diversity, and the heterotrophic bacterial community was diverse (seven phyla). Despite the downshifts in substrate availability, changes in the relative abundance and composition of anammox bacteria, AOB and NOB were small and also the heterotrophic community showed little changes in composition. This indicates stability of PNA MBBR communities towards decreasing substrate availability and suggests that even heterotrophic bacteria are integral components of these communities.
Bacteria capable of anaerobic ammonium oxidation (anammox) enable autotrophic nitrogen removal. Organic carbon in wastewater can instead be utilized for energy production. However, anammox-based processes are not yet used at any extent for treatment of the main stream at wastewater treatment plants. One of the reasons for this is the challenge for the slow growing autotrophic bacteria to work at low temperatures. Here we investigate one-stage nitritation-anammox at temperatures of 13-16°C in a pilot moving bed biofilm reactor (MBBR) receiving reject water from anaerobic sludge digestion. At a target nitrogen loading rate of 1 g NH4+-N m-2 d-1 the average nitrogen removal rate was 0.81 g NH4+-N m-2 d-1 and 0.55 g NH4+-N m-2 d-1 at 16°C and 13°C respectively. At low temperatures oxygen control is important to avoid oxygen penetration to the deeper parts of the biofilm, which causes inhibition of the anammox bacteria and as a result nitrite accumulation. Hence, the process was operated at conditions to limit the activity of the aerobic ammonium oxidizing bacteria (AOB) by oxygen availability. The biofilm biomass was dominated by anammox bacteria, with 1.0 × 1014 copies m-2 (16S rRNA), with considerably fewer AOB of 2.1 × 1012 copies m-2 (amoA), as measured by quantitative PCR. Cell specific conversion rates of anammox bacteria and AOB were estimated at 0.3-0.5 fmol N cell-1 d-1 and 7-9 fmol N cell-1 d-1, respectively. The study shows the applicability of one-stage nitritation-anammox in MBBRs at low temperatures and highlights the importance of quantification of AOB and anammox bacteria for understanding process performance.