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Anaerobic ammonia oxidation (Anammox) coupling sulfur autotrophic denitrification is an effective method for the advanced nitrogen removal from the wastewater with limited carbon source. The influence of S 2 O 3 -S addition on Anammox coupling sulfur autotrophic denitrification was investigated by adding different concentrations of S 2- -S addition on Anammox coupling sulfur autotrophic denitrification was investigated by adding different concentrations of S 2 -S (0, 39, 78, 156 and 312 mg/L) to the Anammox system. The contribution of sulfur autotrophic denitrification and Anammox to nitrogen removal at S 3 2- -S (0, 39, 78, 156 and 312 mg/L) to the Anammox system. The contribution of sulfur autotrophic denitrification and Anammox to nitrogen removal at S 2 O 3 -S concentrations up to 312 mg/L, and only sulfur autotrophic denitrification occurred. The isotopic effects of NO 2- -S concentrations of 156 mg/L was 75% ∼83% and 17%∼25%, respectively, and the mixed system achieved completely nitrogen removal. However, Anammox bioactivity was completely inhibited at S 2 O 3 2- -S concentrations up to 312 mg/L, and only sulfur autotrophic denitrification occurred. The isotopic effects of NO 2 - -N (δ 15 N NO2 and δ 18 O NO2 ) during Anammox coupling sulfur autotrophic denitrification showed a gradual decrease trend with the increase of S 3 - -N (δ 15 N NO3 and δ 18 O NO3 ) during Anammox coupling sulfur autotrophic denitrification showed a gradual decrease trend with the increase of S 2 O 3 2- -S addition. The ratios of δ 15 N NO2 :δ 18 O NO2 and δ 15 N NO3 :δ 18 O NO3 was maintained at 1.30-2.41 and 1.36-2.52, respectively, which revealed that Anammox was dominant nitrogen removal pathway at S 2 -driven autotrophic denitrification and weakened the Anammox, leading to a gradually decreasing trend of the proportion of Candidatus Brocadia as Anammox bacteria from the initial 27% to 4% (S 3 2- -S of 156 mg/L). Yet Norank-f-Hydrogenophilaceae (more than 50%) and Thiobacillus (54%) as functional bacteria of autotrophic denitrification obviously increased. The appropriate amount of S 2 O 3 -S addition promoted the performance of Anammox coupling sulfur autotrophic denitrification achieved completely nitrogen removal.2- -S. The S 2 O 3 2- -S addition enhanced the S 2 O 3 2- -driven autotrophic denitrification and weakened the Anammox, leading to a gradually decreasing trend of the proportion of Candidatus Brocadia as Anammox bacteria from the initial 27% to 4% (S 2 O 3 2- -S of 156 mg/L). Yet Norank-f-Hydrogenophilaceae (more than 50%) and Thiobacillus (54%) as functional bacteria of autotrophic denitrification obviously increased. The appropriate amount of S 2 O 3 2- -S addition promoted the performance of Anammox coupling sulfur autotrophic denitrification achieved completely nitrogen removal., (Copyright © 2022. Published by Elsevier Ltd.)

The dual isotope fractionation of nitrogen (N) and oxygen (O) is an effective way to track the transformation of NO 3 -N in biological denitrification process. The Sulfur autotrophic denitrification combined with the different concentrations of S - -N in biological denitrification process. The Sulfur autotrophic denitrification combined with the different concentrations of S 2 was investigated using the dual isotope fractionation of nitrogen (N) and oxygen (O) to reveal the nitrogen removal mechanism of the activated sludge. Based on successful autotrophic denitrification incubation, the modified Logistic model responded to the short-term effects of S 3 2- was investigated using the dual isotope fractionation of nitrogen (N) and oxygen (O) to reveal the nitrogen removal mechanism of the activated sludge. Based on successful autotrophic denitrification incubation, the modified Logistic model responded to the short-term effects of S 2 O 3 2- addition on NO 3 - -N removal and SO 4 2- generation. Under the S 2 O 3 2- addition of 0.5, 1, 2 and 4 times of the incubation stage (49.29 mg/L-394.32 mg/L), the fractionation effect of N in NO 3 - -N ( 15 ε NO3 ) decreased from 8.74 ± 1.81‰ to 2.08 ± 0.06‰, and the fractionation effect of O in NO 3 - -N ( 18 ε NO3 ) declined from 11.34 ± 0.46‰ to 5.48 ± 0.46‰. The 15 ε NO3 / 18 ε NO3 was maintained at 0.46-0.94, indicating a negative correlation between addition amount and isotope effect, and the addition of high concentrations of S 2 O 3 -N was explored in the autotrophic denitrification. The kinetic models coupled with isotope fractionation effectively revealed the nitrogen removal characteristics in the autotrophic denitrification systems, and verified the difference between the activated sludge-based wastewater treatment process and the natural ecosystem.2- was not suitable for system stabilization. Moreover, the 18 O-labeled H 2 O (δ 18 O H2O ) tests significantly proved the presence of O exchange between NO 2 - -N/NO 3 - -N and H 2 O (67%/97%) during the nitrogen removal process, while the reoxidation of NO 2 - -N was explored in the autotrophic denitrification. The kinetic models coupled with isotope fractionation effectively revealed the nitrogen removal characteristics in the autotrophic denitrification systems, and verified the difference between the activated sludge-based wastewater treatment process and the natural ecosystem., Competing Interests: Declaration of competing interest All authors have no conflict of interest to declare., (Copyright © 2021 Elsevier B.V. All rights reserved.)

• Novel PDA-PFPN system was developed for real domestic wastewater and WAS treatment. • Nitrogen and phosphorus removal and sludge reduction were simultaneously achieved. • Effluent N and P reached 0 and 0.1 mg/L without addition of external carbon source. • Amino acids were important intracellular metabolites in biological P removal. • Tetrasphaera as simultaneous P removal and fermentation bacteria was dominant PAO. A novel process was developed for real domestic wastewater and waste activated sludge (WAS) treatment based on partial denitrification, anammox-biological phosphorus removal, fermentation and partial nitrification (PDA-PFPN). After 246 days of operation, the effluent concentrations of NH 4 +-N, NO 2 −-N and NO 3 −-N were below detection limits (0.1 mg/L), and the effluent concentration of PO 4 3−-P was 0.1 mg/L without the addition of external carbon source in PDA-PFPN system. Moreover, the sludge reduction efficiency reached 48.1% due to fermentation. The nitrite accumulation ratios by ammonia oxidation and nitrate reduction pathway were 60.6% and 87%, respectively. Intracellular metabolites measured by liquid chromatography mass spectrometer (LC-MS/MS) suggested that different intracellular amino acids were stored and consumed at different duration, and intracellular Valine, Glycine and Lysine were not utilized in oxic stage. Results of flow cytometry showed that the proportion of intact cells decreased from 94.7% to 82.9%, and necrotic cells increased from 5.3% to 17.1% with the increase of DNA content in sludge supernatant and cell decay rate, indicating the occurrence of cell death and lysis and leading to WAS reduction. Analysis of transcriptional community composition revealed that partial denitrification bacteria (Thauera), anammox bacteria (Candidatus Brocadia and Candidatus Kuenenia), simultaneous phosphorus removal and fermentation bacteria (Tetrasphaera) and partial nitrification bacteria (Nitrosomonas) coexisted and actually worked in PDA-PFPN system. The novel PDA-PFPN process simultaneously achieved highly efficient nitrogen and phosphorus removal and WAS reduction without the addition of external carbon source, which greatly reduced the operation cost of carbon source dosing and WAS treatment in wastewater treatment. [Display omitted] [ABSTRACT FROM AUTHOR]

• Synergy of endogenous denitrification and denitrifying P removal solved COD lack. • GAOs enriched at transcription level and Accumulibacter was the most active PAOs. • Transcriptional activity more closely related to operation status than gene level. • Denitrifying P removal was confirmed by transcriptional activity of Accumulibacter. The microbial community in endogenous denitrification and denitrifying phosphorus removal treatment at transcription level was unknown. This study first confirmed the expression of actually active bacteria in endogenous denitrification and denitrifying phosphorus removal system to treat low C/N municipal wastewater. No external carbon source was added to influent wastewater. The cDNA high throughput sequencing showed that Candidatus Accumulibacter was the most effective polyphosphate accumulating organisms (PAOs) that actually worked rather than Dechloromonas , which was different from the result at gene level. Reverse transcriptional PCR (RT-PCR) and analysis of Variance (ANOVA) suggested that the ratios of dead or dormant bacteria could monitor wastewater treatment process. Identification of active microbial community at transcription level demonstrated that the synergy of endogenous denitrification by glycogen accumulating organisms (GAOs) and denitrifying phosphorus removal by Candidatus Accumulibacter fully utilized the internal carbon source, and effectively solved the problem of carbon source deficiency in municipal wastewater treatment. [ABSTRACT FROM AUTHOR]

Autotrophic denitrification has become the main contributor to the nitrogen removal from wastewater due to its high efficiency, safety and economy. Effect of Fe on sulfur autotrophic denitrification coupled to heterotrophic denitrification was investigated to enhance the nitrogen removal. Under the substrate ratios of S 2 O 3 2-:Fe (II):CH 3 COO– of 1:1:3, 1:3:1 and 3:1:1, the microorganisms were wrapped by Fe minerals, and the normal metabolic pathways were blocked. By adjusting the substrate ratios of S 2 O 3 2-:CH 3 COO– of 1:3, 2:2 and 3:1 without Fe (II), the denitrification recovered with the removal efficiency of NO 3 –-N to 99%. Insufficient addition of S 2 O 3 2- and CH 3 COO– promoted the production of NO 2 –-N and N 2 O with high percentage of N 2 O of 5.70%. When S 2 O 3 2- and CH 3 COO– were sufficient, the nitrogen removal was effective and N 2 O production was low (＜1.35%). Modified Blotzman model and Logistic model were suitable to describe the removal of NO 3 –-N, modified Logistic model was suitable to reveal the generation of SO 4 2-. Dechloromonas acted as denitrifying bacteria in substrate ratios of S 2 O 3 2-:CH 3 COO– of 1:3 and 3:1 up to 58% and 56% at the genus level. Autotrophic Thiobacillus bacteria decreased from 19% (inoculated sludge) to 1% (1:3), 16% (2:2) and 17% (3:1), respectively. Thauera was corresponded to the heterotrophic denitrification which was successfully enriched from 0 (inoculated sludge) to 13% (1:3), 49% (2:2) and 2% (3:1). The different substrate ratios promoted the coexistence of autotrophic and heterotrophic bacteria and cooperated with Fe minerals to improve nitrogen removal from wastewater. [ABSTRACT FROM AUTHOR]