Average Total Nitrogen Removal Efficiency Research Articles

Nitrogen removal from urea effluent in an anammox sequencing batch biofilm reactor with valorised dishwashing scrubber as biocarrier

BackgroundThis study aimed for efficient nitrogen removal from urea wastewater in an anammox SBBR with valorised dishwashing scrubber as biocarrier. MethodsThe study (225 d) comprised three discrete phases: Phase I [1 – 26 d; Inf. NH4+-N (Simulated ww)], Phase II [27 – 131 d; Inf. NH4+-N (Real ww) + Inf. NH4+-N (Simulated ww)] and Phase III [132 – 225 d; Inf. NH4+-N (Real ww)] in which the reactor was supplemented with varied ratios (0 – 7) of influent NH4+-N reflecting varying combinations of real and simulated wastewater. Significant findingsThe reactor exhibited average total nitrogen removal efficiency of approximately 83 %, 81 %, and 80 % in the subsequent phases I, II, and III, respectively at varied nitrogen loading rates (100 – 232 g N m−3 d−1). The 16S rRNA gene sequencing revealed significant enrichment of Candidatus Kuenenia (from 41.69 % to 48.11 %) on the 225th day which was primarily attributed to significant decline in Nitrospira from 23.9 % to 1.24 % (due to nitrite oxidizing bacteria inhibition on account of high free ammonia concentrations in the reactor) and Nitrosomonas from 4.32 % to 0.01 %. Additionally, genera that were not initially present (on the 1st day) including Stenotrophobacter (17.34 %), Sphingobium (8.41 %), Rhodococcus (5.88%) and Ohtaekwangia (5.84 %) were observed to emerge by the end of the experiment (225 d).

Effect of linear anionic surfactant (sodium dodecyl sulfate) dosage on nitrogen removal performance of an anammox reactor

AbstractThe impact of sodium dodecyl sulfate (SDS, as model linear anionic surfactant) dosage on overall nitrogen removal performance of anammox reactor alongside its microbial population and sludge properties was investigated. In this study (day 136), an anmmox sequencing batch biofilm reactor was subjected to gradual dosage of SDS from 0 to 20 mg L−1. Intriguingly, results revealed that SDS at ≥7.5 mg L−1 prompted sludge disintegration, evidenced by increased protein and polysaccharide content in the effluent. Nevertheless, reactor's average total nitrogen removal efficiency slightly improved from 83.12% (0 mg L−1) to 86.3% (20 mg L−1). The 16S rRNA gene sequencing revealed that SDS dosing significantly suppressed the unwanted and unavoidable nitrite oxidizing bacteria (NOBs) in the reactor as the abundance of genus Nitrospira declined from 40.68% (day 1) to 19.15% (day 136). The abundance of anammox genus Candidatus Kuenenia significantly improved from 1.86% (day 1) to 40.02% (day 136) as a result of NOB suppression. This study revealed that low concentration of surfactants in wastewater does not affect the anammox bacteria in a biofilm reactor. Furthermore, adding low concentrations of SDS (≥7.5 to 20 mg L−1) to wastewater may effectively suppress notorious NOBs in biofilm‐based anammox systems.

Decrease in Oxygen Concentration for the Fast Start-Up of Partial Nitritation/Anammox without Inoculum Addition

Initiating the partial nitritation and anammox (PN/A) process without inoculation poses a significant challenge. Thus, there is a notable amount of interest in devising a straightforward strategy for the start-up of PN/A. This study demonstrates the feasibility of achieving the rapid start-up of a one-stage PN/A process within a moving-bed sequencing batch biofilm reactor (MBSBBR) by reducing the oxygen concentrations: 3.0 mg O2/L (Stage I), 2.0 mg O2/L (Stage II), and 1.0 mg O2/L (Stage III). The anammox activity was observed 15 days after a gradual decrease in the oxygen concentration and confirmed using a specific anammox activity test (5.9 mg N/gVSS∙h). During Stage III, the average total inorganic nitrogen (TIN) removal efficiency was 60.6%. The relative abundance of planctomycetes, a typical phylum representing anammox microorganisms, increased almost three times from Stage I (2.8%) to Stage III (7.1%). These results demonstrate the potential of a decrease in oxygen concentration for the fast start-up of the one-stage partial nitritation and anammox process without inoculation. The implementation of the studied DO strategy has practical implications for wastewater treatment plant operators, particularly in the start-up of the PN/A processes. Additionally, batch assays allow for the rapid assessment of treatment plant performance, providing real-time insights into its functionality and, thereby, optimizing wastewater treatment practices.

Electron transfer mechanism of intracellular carbon-dependent DNRA inside anammox bacteria

Generally, anaerobic ammonium oxidation (anammox) converts nitrite (NO2−) and ammonium (NH4+) to nitrogen gas (N2) but generates some nitrate (NO3−) (equivalent to 11% of inlet total nitrogen (TN)). Although it reported that anammox bacteria could degrade NO3− via dissimilatory nitrate reduction to ammonium (DNRA) pathway using the intracellular carbon as the electron donor, it is still unclear the specific electron transfer mechanism in this intracellular carbon-dependent DNRA inside anammox bacteria, and whether the sole anammox bacteria could achieve higher TN removal efficiency more than the theoretical maximum of 89%. In this study, transcriptome analysis and metabolic inhibitor experiments demonstrated that NADH generated from the decomposition of the intracellular carbon (glycogen) supplied electrons for the NO3−conversion; the electrons were transferred from NADH to nitrate reductase (Nar) and nitrite reductase forming ammonium (NrfA) from ubiquinone (UQ) and complex III, respectively. Combining the intracellular carbon-dependent DNRA with normal anammox process, an average TN removal efficiency of 95% was achieved by the sole anammox bacteria in a sequencing batch reactor. Fluorescent in situ hybridization (FISH) images and real-time fluorescence quantitative PCR (qPCR) results illustrated anammox bacteria could survive and proliferate in the SBR. Our work improved the understanding of the electron transfer mechanism inside anammox bacteria, and further exploit its potential in nitrogen pollutants removal.

Pyrite and sulfur-coupled autotrophic denitrification system for efficient nitrate and phosphate removal

The inefficiency of nitrogen removal in pyrite autotrophic denitrification (PAD) and the low efficiency of PO43−-P removal in sulfur autotrophic denitrification (SAD) limit their potential for engineering applications. This study examined the use of pyrite and sulfur coupled autotrophic denitrification (PSAD) in batch and column experiments to remove NO3−-N and PO43−-P from sewage. The effluent concentration of NO3−-N was 0.32 ± 0.11 mg/L, with an average Total nitrogen (TN) removal efficiency of 99.14%. The highest PO43−-P removal efficiency was 100% on day 18. There was a significant correlation between pH and the efficiency of PO43−-P removal. Thiobacillus, Thiomonas and Thermomonas were found to be dominant at the bacterial genus level in PSAD. Additionally, the abundance of Thermomonas in the PSAD was greater than that observed in the SAD reactor. This result indirectly indicates that the PSAD system has more advantages in reducing N2O.

Intensifying single-stage denitrogen by a dissolved oxygen-differentiated airlift internal circulation reactor under organic matter stress: Nitrogen removal pathways and microbial interactions

Current research focuses on efficient single-stage nitrogen removal from organic matter wastewater using the partial nitritation-anammox (PNA) process. In this study, we constructed a single-stage partial nitritation-anammox and denitrification (SPNAD) system using a dissolved oxygen-differentiated airlift internal circulation reactor. The system was operated continuously for 364 days at 250 mg/L NH4+–N. During the operation, the COD/NH4+–N ratio (C/N) was increased from 0.5 to 4 (0.5, 1, 2, 3, and 4), and the aeration rate (AR) gradually increased. The results showed that the SPNAD system maintained efficient and stable operation at C/N = 1–2 and AR = 1.4–1.6 L/min, with an average total nitrogen removal efficiency of 87.2%. The removal pathways of pollutants in the system and the interactions between microbes were revealed by analyzing the changes in sludge characteristics and microbial community structure at different phases. As the influent C/N increased, the relative abundance of Nitrosomonas and Candidatus Brocadia decreased, and that of denitrifying bacteria, such as Denitratisoma, increased to 44%. The nitrogen removal pathway of the system gradually changed from autotrophic nitrogen removal to nitrification-denitrification. At the optimum C/N, the SPNAD system synergistically removed nitrogen through PNA and nitrification-denitrification. Overall, the unique reactor configuration facilitated the formation of dissolved oxygen compartments, providing a suitable environment for different microbes. An appropriate organic matter concentration maintained the dynamic stability of microbial growth and interactions. These enhance microbial synergy and enable efficient single-stage nitrogen removal.

The rapid start-up of CANON process through adding partial nitration sludge to ANAMMOX system

This study aimed to start up the completely autotrophic nitrogen removal over nitrite (CANON) process after adding partial nitration (PN) sludge to the ANAMMOX reactor, so as to help the rapid start-up and stable operation of the CANON process in practical engineering applications. There were three steps in the research: cultivating the PN sludge, building a reliable ANAMMMOX system, and finally starting and running the CANON process. The PN sludge was successfully cultivated in less than 45 days with around 90% nitrite accumulation rate. The ANAMMOX reactor enriched a significant quantity of red granular sludge within 70 days, achieving the maximum nitrogen removal rate of 1.74 kg/(m3·d). Eventually, the CANON reactor was started up successfully, which achieved 95.08% of average ammonium removal efficiency and 84.51% of average total nitrogen removal efficiency in 60 days. The residual recalcitrant nitrite-oxidizing bacteria in the CANON process was successfully inhibited by intermittent aeration and 12 mg/L free ammonia in UASB reactor. Besides, Candidatus Kuenenia, Candidatus Brocadia and Nitrosomonas were the main functional microorganisms involved in the CANON process.

Influence of plants on anammox process in constructed Wetland: Irrelevance, inhibition or enhancement

Anaerobic ammonia oxidation (anammox) can become the dominant process of nitrogen removal in constructed wetland, yet whether plants that are signature composition of wetland play a significant role in this process is still unknown. Therefore, this study investigated the influence of plants (Phragmites) on the anammox-driven wetlands by comparing the differences between plant group and non-plant group. The results indicated that the proportion range of anammox process in nitrogen metabolism of both groups was estimated to be 45 %–63 % according to the detected abundance of functional genes, and there was a significant positive relationship between anammox proportion and total nitrogen removal efficiency (R = 0.81, P < 0.05), which confirmed anammox process dominating the nitrogen transformation in all systems. The average total nitrogen removal efficiencies in the systems with plants addition (84–85 %) were significantly superior than those in the corresponding control systems (74–82 %), respectively (P < 0.05). The presence of plants significantly contributed to the increase in the anammox proportion of nitrogen metabolism and the total absolute abundances of anammox bacteria (P < 0.05). Partial Least Squares Path Analysis Model (PLSPM) furtherly revealed that the estimated weight coefficients of plants to anammox process (R = 1.24) showed the larger level than denitrification process (R = 0.25). Overall, plants played an enhancing role in the anammox process of anammox-driven wetlands, rather than irrelevant or potential inhibiting role. The reason was mainly related to the plant role for improving biofilm formation by extracellular polymeric substances (EPS), and a significant positive relationship between EPS concentrations (7–224 μg·g−1) and anammox bacteria abundances (1.63–7.66 × 108 cells·g−1) was found (R = 0.44, P < 0.05). Besides, the subsidiary reasons also lie in the minimal role of plant roots under higher nitrogen loading and the appropriate nitrogen component of wastewater.

Performance characteristics and bacterial community analysis of a novel two-step-feed three-stage A/O-MBBR system for nitrogen removal in municipal wastewater

To obtain a high nitrogen removal efficiency and fully use the carbon sources in raw wastewater, a novel two-step-feed three-stage anoxic/oxic (A/O)-moving bed biofilm reactor (MBBR) system with ten chambers was established for municipal wastewater treatment. The designed ten chambers in the 2F-3S-A/O-MBBR system played different roles in nitrogen and organic removals, and they accomplished the task of achieving stable high nitrogen removal efficiency at different temperatures together. Concretely, the average total inorganic nitrogen (TIN) removal efficiency was improved to 72.88 % from 61.55 % without any external carbon sources by the second-stage A/O-MBBR subsystem due to the two-step-feed configuration, and it was further improved to 91.42 % by the third-stage A/O-MBBR subsystem due to the use of the external carbon sources. Specific functional biofilms with different bacterial communities (r = 0.85, P ≤ 0.001) and functional genes of denitrification and nitrification (r = 0.63, P ≤ 0.001) were formed in 4 groups of functional zones, including the first-stage anoxic, second-stage anoxic, third-stage anoxic and oxic chambers. This study developed an efficient 2F-3S-A/O-MBBR biological nitrogen removal process and investigated the performance characteristics and bacterial community to provide fundamental support for the application of this process.

Denitrification performance and microbial community analysis of sulfur autotrophic denitrification filter for low-temperature treatment of landfill leachate

This study investigated the denitrification performance of sulfur autotrophic denitrification filter (SADF) in five low-temperature ranges (14.0–15.0 ℃, 13.1–13.9 °C, 12.0–13.0 °C, 10.1–11.9 °C and 8.8–9.8 °C) environments and the nitrogen removal contribution along the filter cells. Results showed that the average total nitrogen removal efficiency (TNRE) of the bioreactor were 90.06%, 88.49%, 86.61%, 85.71% and 74.54%, respectively. However, when the temperature was lower than 9.8 °C, the process of sulfur autotrophic denitrification was obviously inhibited, but the inhibition was slowed down after low-temperature acclimation. The bottom-filter layer (0–200 mm) contributed 50.54% of the nitrogen removal, which was the greatest. The middle- filter layer (400–600 mm), which was at 35.86%, was next. And the upper- filter layer (800–1200 mm) made the least contribution, at 13.60%. It was demonstrated by high-throughput sequencing that in addition to Thiobacillus, the genus Sulfurimonas was also found and its average relative abundance was comparable to the former (24.03% for Thiobacillus and 23.10% for Sulfurimonas). The results of microbial function prediction showed that sulfur and nitrogen metabolism played a key role in nitrogen removal. The study of substrate environmental variables showed that the key bacterial groups of nitrogen removal along the process were negatively correlated with total nitrogen (TN) and positively correlated with pH value.

Enrichment of anammox sludge by using an Internal Circulation Reactor innoculated with anaerobic granular sludge and old landfill leachate

In recent years, anammox has not only economic benefits but also potential for wastewater treatment containing ammonium and low organic carbon. Along with breakthroughs ammonium treatment in the world, anammox process in internal circulation reactor (IC) is a new technology being researched and developed in high-load ammonium treatment. There In this study, an laboratory scale IC was used to enrich anammox sludge by seeding anaerobic granular sludge and feeding old leachate from the Go Cat municipal solid waste landfill. The seed sludge, which was anaerobic granular sludge taken from an UASB reactor of an industrial wastewater treatment plant was innoculated to IC at MLSS and MLVSS concentrations of 55.1 g.L-1 and 45 g.L-1, respectively, equivalent to the MLVSS:MLSS ratio of 0.82. Since 80 days of operation, the sludge in the IC was divided into two separate zones: a granular zone at the IC bottom part and the floc zone in the upper part. The average total nitrogen (TN) removal efficiency was 45% at nitrogen loading rate 0.4 – 0.6 kg N. m-3.day. The SAA of floc sludge and granular sludge on day 61, increased 6 and 8 times compared to thosed on day 27. The study illustrated that the enrichment of anammox sludge took a long time and granulation in the IC reactor was an important operating factor for anammox growth.

Immobilized denitrifying bacteria on modified oyster shell as biofilter carriers enhance nitrogen removal

Biocarriers play a vital role in water treatment in recirculating aquaculture systems. In this study, modified oyster shell (OS)-immobilizing denitrifying bacteria were applied as biocarriers in biofilters. The results revealed that CaCO3 was the main component of the OS calcined at 200 °C (OS200), 400 °C (OS400), and 600 °C (OS600). The surface area and microbe-immobilizing ability of OS200, OS400, and OS600 were enhanced. After immobilizing Pseudomonas putida DS2 and Pseudomonas chengduensis BF6, the average total nitrogen (TN) removal efficiencies of OS400-P (Pseudomonas sp. DS2 and BF6 immobilized onto OS400) and OS600-P (Pseudomonas sp. DS2 and BF6 immobilized onto OS600) reached 50.25% and 61.50%, respectively. The microbial and functional gene richness in the biofilters filled with biocarriers made of OS-immobilizing denitrifying bacteria changed. The genus Pseudomonas became the dominant microbe, which was positively correlated with NO3--N + NO2--N (p < 0.001), and TN removal (p < 0.01). Overall, these findings indicate that applying OS immobilizing denitrifying bacteria as biocarriers can be a novel strategy to enhance nitrogen removal in biofilters.

Stable year-round nutrients removal and recovery from wastewater by technical-scale algal Turf Scrubber (ATS)

In this study, three technical-scale Algal Turf Scrubber (ATS) systems were continuously operated in a greenhouse over one year to evaluate the nutrient removal and recovery ability. The independent and interdependent effects of water temperature and light intensity on dependent variables, such as nutrient removal and recovery, were identified via mathematical modelling. Over 7 days, the average total nitrogen and phosphorous removal rates and efficiencies were 30.93 and 8.95 mg/L and 51.2 and 89.5 %, respectively. Based on a statistical regression matrix, the quadratic models were chosen to analyze data on nutrient removal and recovery. In this study, light intensity was the dominant variable for ATS performance. Significant positive correlations were found between nutrient removal and recovery, and biomass productivity. The annual production costs for dried algal biomass, N and P were 2.27, 40 and 156 € kg−1, respectively. Our results indicated that the ATS technology can be efficient and cost-effective in nutrients removal and recovery from wastewaters.

Sulfammox forwarding thiosulfate-driven denitrification and anammox process for nitrogen removal

The coupled process of thiosulfate-driven denitrification (NO3−→NO2−) and Anammox (TDDA) was a promising process for the treatment of wastewater containing NH4+-N and NO3−-N. However, the high concentration of SO42− production limited its application, which needs to be alleviated by an economical and effective way to promote the application of TDDA process. In this study, TDDA process was started in a relatively short time by stepwise replacing nitrite with nitrate and operated continuously for 146 days. Results presented that the average total nitrogen removal efficiency of 82.18% can be acquired at a high loading rate of 1.98 kg N/(m3·d) with maximum nitrogen removal efficiency up to 87.04%. It was observed that the increase of S/N ratio improved the denitrification efficiency and slightly inhibit the Anammox process. Batch tests showed that Sulfammox process appeared in TDDA process under certain conditions, further contributing 2.59% nitrogen removal and 10.46% sulfur removal (14.42 mg/L NH4+-N and 37.68 mg/L SO42--S were removed). This finding was mainly attributed to the reduction of sulfate in TDDA system to elemental S0 or HS−, which subsequently was used as an electron donor to realize the recycling of sulfate (SO42--S) pollutants and promote the sulfur-nitrogen (S–N) cycle. High-throughput analysis displayed that Anammox bacteria (Candidatus_Kuenenia), Sulfur-oxidizing bacteria (Thiobacillus) with relatively high abundance of 5.37%, 7.74%, respectively, guaranteeing the excellent nitrogen and sulfate removal performance in the reactor. The enrichment of phyla Chloroflexi (31.79%), Proteobacteria (31.82%), class Ignavibacteriales (10.55%), genus Planctomycetes (13.57%) further verified the exitence of Sulfammox process in the TDDA reactor. This study provides a new perspective for the practical application of TDDA in terms of reducing the production of high concentration SO42− and saving operational cost and strengthening deeply nitrogen removal.

Morphological, kinetic, and microbial community characterization of anammox bacteria with different inoculations and biofilm types for low-ammonium wastewater treatment

Granular anammox sludge treating high-ammonium wastewater and biofilm anammox sludge treating low-ammonium wastewater were inoculated in two identical reactors to treat low-ammonium wastewater. The morphological and microbial community characterizations were investigated. Kinetic models were applied to understand kinetic characterizations and predict reactor performances. Both types of sludge adapted well in 10 and 5 days with the average total inorganic nitrogen removal efficiencies >85%. The average particle size of the granular sludge decreased from 843 μm to 515 μm. The biomass and thickness of the biofilm sludge increased by 17% and 66% when NH4+-N, NO2−-N and temperature increased from 10 mg/L to 33 mg/L, <0.5 mg/L to 41 mg/L, and circa 13 °C to 35 °C, respectively. The granular sludge had higher maximum utilization rate constant and saturation value constant (16.393 and 12.934 mg/(L·h)) than biofilm sludge (1.017 and 0.080 mg/(L·h)), indicating the inherent difference of inoculations. The granular sludge inoculated from high nitrogen loading rate (NLR) (0.36 g-N/(g-VSS·d)) condition had the potential to withstand a higher NLR than that in present reactor (0.14 g-N/(g-VSS·d)). The results of Grau second-order model indicated that both reactors performed good nitrogen removal efficiency despite inoculations and types differences, and can predict the nitrogen removal efficiency well. Microbial community diversities deceased, but granular bacteria richness increased due to the lower free ammonium and free nitrous acid. The dominant anammox bacteria in the granular sludge shifted from Candidatus Kuenenia to Candidatus Brocadia. Candidatus Kuenenia was always the dominant anammox bacteria in the biofilm sludge.

Food waste hydrolysate as a carbon source to improve nitrogen removal performance of high ammonium and high salt wastewater in a sequencing batch reactor

The high ammonium and high salt (HAHS) wastewater generated from the anaerobic digestate of food waste is usually difficult to be treated by biological process because of its low C/N ratio. Herein, food waste hydrolysate (FWH) is rich in readily biodegradable organic matter, was utilized as carbon source to enhance the nitrogen removal of HAHS in the activated-sludge system. Results showed that compared with the control average total nitrogen removal efficiency increased from 73.4% to 94.9% and effluent declined from 281.4 mg/L to 53.9 mg/L by adding FWH at the C/N ratio of 6, satisfying the sewage discharge standard regulated by China. Besides, FWH utilization led to higher selectivity of the species responsible for nitrogen removal in related to glucose-adding group, which were dominated by Flavobacteriaceae, Melioribacteraceae, PHOS-HE36, and Rhodobacteraceae after a long-term operation. In general, FWH is an alternative carbon source to enhance nitrogen removal in HAHS wastewater treatment.

Realization of partial nitrification and in-situ anammox in continuous-flow anaerobic/aerobic/anoxic process with side-stream sludge fermentation for real sewage

A continuous-flow anaerobic/aerobic/anoxic reactor with complete suspended activated sludge using sludge alkaline fermentation products as carbon source was utilized to strengthen nitrogen removal performance for low C/N ratio (<4) wastewater. Long-time performance indicated that the nitrite accumulation rate reached 60.40%, which strengthened the contribution of anammox. The average total inorganic nitrogen removal efficiency improved 19.40%. The abundance of ammonia oxidizing bacteria has not changed, but the abundance of nitrite oxidizing bacteria reduced from 5.79% to 0.69%. Quantitative PCR results demonstrated that the abundance of anammox bacteria has raised by 80.5 times. These results indicated that side-stream sludge alkaline fermentation promoted the mainstream partial nitrification, consequently accelerating the in-situ enrichment of anammox bacteria. No external carbon source dosing and short oxic hydraulic retention time (5.3 h) save energy and reduce consumption significantly in this system.

Start-up a novel coupling process with partial nitritation, partial denitrification and anaerobic ammonia oxidation in a single sequencing batch reactor

A novel process, named single-stage PNDA, that integrates the partial nitritation, partial denitrification and anaerobic ammonia oxidation in a single lab-scale sequencing batch reactor was developed to treat moderate-strength ammonia wastewater (220 mg N/L). Firstly, a single stage partial nitritation-anammox process was started up with the total nitrogen removal efficiencies of approx. 76.22%, and the remaining free ammonia concentration could produce a more sustained inhibition of nitrite oxidizing bacteria(NOB) than low DO condition. Then, the proposed one-stage PNDA process achieved an average total nitrogen removal efficiency of 85.29% by adding sodium acetate in the influent at a concentration of 50 mg COD/L. Microbial community characterization revealed that ammonia-oxidizing bacteria, anammox bacteria, NOB and partial denitrification bacteria coexist in the suspended sludge. The significant enrichment of Thauera and the dominance of Ca. Anammoxoglobus in the Anammox biomass indicate their important role in the improved nitrogen removal performance. This single-stage PNAD process can potentially be applied to further improve the total nitrogen removal efficiency of PN/A process.

The coupling of mixotrophic denitrification, dissimilatory nitrate reduction to ammonium (DNRA) and anaerobic ammonium oxidation (anammox) promoting the start-up of anammox by addition of calcium nitrate

This study discovered one nitrate-calcium-based anammox start-up pathway. Compared with control, the start-up time of anammox was saved by 33.3%, and the average total nitrogen removal efficiency increased from 29.6% to 53.7% during the start-up. Besides, the continuous nitrite accumulation (1.18 mg/L) and a marked increase in the relative abundance of denitrifying and anammox bacteria were observed in the only Ca(NO3)2-added group. These results suggested that calcium nitrate induced partial denitrification to provide nitrite for anammox. Additionally, the role of dissimilatory nitrate reduction to ammonium (DNRA) in the Ca(NO3)2-added systems also deserved attention, for the contribution of DNRA to nitrate removal as well as the relative abundance of DNRA bacteria were both increased for the Ca(NO3)2-added groups. These results suggested that a mutualistic symbiosis among denitrification, DNRA and anammox exists in the calcium nitrate-added systems, which may explain the reason for acceleration of anammox start-up by adding calcium nitrate.

Unravelling the importance of Ca2+ and Mg2+ as essential in anammox culture medium

The mechanism and nitrogen removal performance of anammox process under different concentrations of Ca2+ and Mg2+ were explored from the perspective of molecular biology analysis based on the metabolic changes of the second messenger cyclic diguanylate (c-di-GMP). After 100-day operation, reactor with 98 mg/L Ca2+ and 30 mg/L Mg2+ achieved a higher anammox performance with an average total nitrogen (TN) removal efficiency of 85.8%. Under the Mg2+concentration of 30 mg/L, a higher Ca2+ could accelerate anammox process by promoting the amplification of Candidatus Brocadia (0.62%) and production of Diguanylate cyclase (DGC-s: 6.54 × 108 copies/μL DNA) which function was to synthesize c-di-GMP. While under the Ca2+concentration of 49 mg/L, Mg2+ concentration at appropriate rang could promote the degradation process of c-di-GMP. Since Ca2+ had positive linear relationship with TN removal (R2 = 0.96), a higher Ca2+ concentration is recommended in the culture medium. This study provided a potential method for optimization of anammox process.