Isotope analysis of nitrogen removal pathways and N2O production potential in the SDAD-anammox system under different N/S ratios.

This study explored the impact of varying nitrate to sulfide (N/S) ratios on nitrogen removal efficiency (NRE) in the sulfide-driven autotrophic denitrification and anammox (SDAD-anammox) system. Optimal nitrogen removal was observed at N/S ratios between 1.5 and 2.0. Isotope tracing results showed that the contribution of anammox to nitrogen removal was enhanced with increasing N/S ratios, reaching up to 37 % at the N/S ratio of 2.5. Additionally, complex nitrogen pathways were identified, including dissimilatory nitrate reduction to ammonium (DNRA). Furthermore, isotope tracing was innovatively applied to investigate N2O emissions, demonstrating that higher N/S ratios significantly reduced N2O emissions, with the lowest emissions at N/S ratio of 2.5. Gene expression analysis indicated that nitrogen and sulfide transformation genes decreased with increasing N/S ratios, while anammox-related genes first increased and then decreased, reflecting the system's microbial dynamics. These findings offer insights into nitrogen transformation pathways and N2O production mechanisms in the SDAD-anammox process.

Ultra-stable mixotrophic denitrification coupled with anammox under organic stress for mainstream municipal wastewater treatment.

Sulfur-based autotrophic denitrification (SAD) coupled with anammox is a promising process for autotrophic nitrogen removal in view of the stable nitrite accumulation during SAD. In this study, a mixotrophic nitrogen removal system integrating SAD, anammox and heterotrophic denitrification was established in a single-stage reactor. The long-term nitrogen removal performance was investigated under the intervention of organic carbon sources in real municipal wastewater. With the shortening of hydraulic retention time, the nitrogen removal rate of the mixotrophic system dominated by the autotrophic subsystem reached 0.46 Kg N/m³/d at an organic loading rate of 0.57 Kg COD/m³/d, with COD and total nitrogen removal efficiencies of 82.5 % and 94 %, respectively, realizing an ideal combination of autotrophic and heterotrophic systems. The 15NO3--N isotope labeling experiments indicated that thiosulfate-driven autotrophic denitrification was the main pathway for nitrite supply accounting for 80.6 %, while anammox exhibited strong competitiveness for nitrite under the dual electron supply of sulfur and organic carbon sources and contributed to 65.1 % of nitrogen removal. Sludge granulation created differential functional distributions in different forms of sludge, with SAD showing faster reaction rate as well as higher nitrite accumulation rate in floc sludge, while anammox was more active in granular sludge. Real-time quantitative PCR, RT-PCR and high-throughput sequencing results revealed a dynamically changing community composition at the gene and transcription levels. The decrease in heterotrophic denitrification bacteria abundance indicated the effectiveness of the operational strategy for introduction of thiosulfate and maintaining the dominance of SAD in denitrification process in suppressing the excessive growth of heterotrophic bacteria in the mixotrophic system. The high transcriptional expression of sulfur-oxidizing bacteria (SOB) (Thiobacillus and Sulfurimonas) and anammox bacteria (Candaditus_Brocadia and Candidatus_Kuenenia) played a crucial role in the stable nitrogen removal.

Rapid cultivation and enrichment of anammox bacteria solely using traditional activated sludge as inoculum and biocarrier in low-strength real sewage treatment.

In low-ammonia sewage anammox process, cultivation and enrichment of anammox bacteria (AnAOB) is a challenge especially from traditional activated sludge. To this end, a novel strategy solely using activated sludge as inoculum and biocarrier in a dynamic fixed-bed reactor was proposed in this study. During 115-day operation, excellent performance was achieved with influent total inorganic nitrogen (TIN) and effluent TIN of 55.3 mg·L-1 and 4.1 mg·L-1, respectively. Rapid enrichment of AnAOB (doubling time: 8.5 days) was demonstrated by augmented specific anammox activity (trace value to 1.85 mg N·g VSS-1·h-1) and increased hzsB gene number (106 to 109 copies·g-1 dry sludge), with predominance of Candidatus_Brocadia. Large-flocs aggregate was the primary habitat for AnAOB with highest abundance and capacity. The distinctive sludge properties, symbiotic microbial interactions and dynamic operation scheme facilitated AnAOB growth and retention. This study provides a simple, economic and workable approach for the start-up of mainstream anammox process.

A novel strategy for enhancing the partial denitrification to treat domestic wastewater by feeding sludge fermentation liquid.

Partial-denitrification (PD; NO3-→NO2-) has recently been proposed to be a feasible choice of NO2--N supply for anammox bacteria. In this study, an aerobic/anoxic process for treating domestic wastewater was operated for 176 days to evaluate the feasibility of using sludge fermentation liquid for partial denitrification of the wastewater. Results show that, with the ratio of C/N (COD/ NO3--N) increased at anoxic stage, the average NO2--N concentration in the effluent and nitrate-to-nitrite transformation ratio (NTR) at anoxic stage showed relative growth. High-throughput sequencing analysis demonstrated that the enhancement of PD can be explained by the increases of Thauera, Paracoccus and Enterobacteriaceae. Moreover, Candidatus_Brocadia (0.13%) was detected as the predominant anammox bacteria. Ex-situ isotopic tracing technique analysis assessed that the ratio of anammox role (ra%) was 7.29%. This study has a great potential for being coupled with the anammox bacteria for advanced nitrogen removal.

Performance of the anammox process treating low-strength municipal wastewater under low temperatures: Effect of undulating seasonal temperature variation.

In the anammox process treating low-strength municipal wastewater, the effect of common seasonal temperature variation (15.1 °C-22.2 °C) on performance was studied. In autumn and winter, the nitrogen removal rate (NRR) decrement of 0.038kgN/(m3·d) (17.9 °C â†’ 15.1 °C) was nearly threefold higher than 0.014kgN/(m3·d) (22.2 °C â†’ 17.9 °C), which showed that lower temperature laid more negative impact on nitrogen removal. 15N isotope tracing tests confirmed that the contribution of denitrification to nitrogen removal was far less than anammox, and anammox contributed more at 15.1 °C (91.7%) than 21.9 °C (78.9%). Anammox bacteria could adapt to lower temperature after short-term acclimatization, especially the dominant genus Ca. Brocadia increased from 1.8% to 2.5% and its abundance was significantly correlated with nitrogen consumption (p < 0.05). Above findings suggest that the adaptability of Ca. Brocadia could provide the possibility to maintain nitrogen removal performance at lower temperature. In spring, the improved maximum anammox activity from 2.85 to 3.23mgNH4+-N/(gVSS·h) indicated the recovered removal capacity.

Recovering partial nitritation in a PN/A system during mainstream wastewater treatment by reviving AOB activity after thoroughly inhibiting AOB and NOB with free nitrous acid.

Starting up or recovering partial nitritation is a major challenge for achieving or maintaining stable partial nitritation/anammox (PN/A) during mainstream wastewater treatment. This study presents a novel strategy for recovering the nitrite pathway by selectively reviving ammonium oxidizing bacteria (AOB) after thoroughly inhibiting AOB and nitrite oxidizing bacteria (NOB) using free nitrous acid (FNA). A sequencing batch reactor was operated for PN/A to treat real domestic wastewater for 423 days, during which twice FNA treatment was temporarily implemented. Results showed that with a single 0.45 mg/L FNA treatment on flocculent sludge, the NO3--N concentration during the aerobic period showed an uptrend again and the partial nitritation performance was deteriorated. In contrast, 1.35 mg/L FNA treatment induced the inhibition of both AOB and NOB leading to regressive ammonium oxidation, but a subsequently higher DO (1.5 mg/L) and longer aeration duration recovered partial nitritation. For the relative abundances of the acquired biomass related to nitrogen conversion, Nitrosomonas, Nitrospira and Nitrolancea increased to 9.65%, 10.27% and 4.35%, respectively, at the beginning of the 1.35 mg/L FNA treatment, and Nitrospira and Nitrolancea decreased to 2.80% and 0.03% whereas Nitrosomonas declined to 8.71% after 76 days. Ca. Brocadia showed less resilience after the 1.35 mg/L FNA treatment, with the relative abundance decreasing from 13.38% to 0.62% due to insufficient nitrite. Molecular ecological network analysis indicates that among anammox taxa, Ca. Kuenenia and Ca. Brocadia formed important links with other N cycle processes. Moreover, the proposed strategy shows operational flexibility because it can be easily used to control NOB in mainstream PN/A applications offered by flocculent sludge systems.