Anammox Sludge Research Articles

Sulfide addition accelerates anammox sludge granulation and promotes microbial cooperation

The granular anaerobic ammonium oxidation (anammox) system has attractive advantages in tolerance to environmental-stress and enhancement of nitrogen removal capacity. Sulfide addition can improve nitrogen removals in anammox systems via inducing sulfur denitrification, yet its function in the improvement of the property of anammox granular sludge remains unclear. Herein, we investigated the variations in the morphological and microbial properties of the anammox sludge response to different sulfide concentrations (Na2S: 10–100 mg/L) through a long-term experiment. By comparing the sludge diameter and heme c content, it comes that a relatively low sulfide (S/N [nitrate] molar ratio of 0.18–0.50) significantly promoted the average diameter and heme c concentration of sludge by 25–175 % and 75–95 %, respectively, compared to that of both without sulfide addition and a high sulfide addition (S/N > 0.85). This enhancement is primarily because a low amount of sulfide had stimulated the secretion of extracellular polymeric substance, induced slight biogenic sulfur accumulation as microbial nuclei, and facilitated the appropriate amount of filamentous bacteria proliferation. Microbial metabolism functions analyses revealed a robust granular anammox coupled with sulfur denitrification in the sulfide-mediated anammox reactor, and the assembled granules exhibited exceptional tolerance to environmental stress. Significantly, the anammox bacteria (Candidatus_Brocadia) dominating the granules displayed satisfactory anammox activity (21.8 ± 2.1 mg N/g VSS h), and their produced nitrate was efficiently removed by the sulfur-oxidizing bacteria (Thiobacillus) that predominantly occurred in the flocs. This collaboration ensured an efficient sulfide-mediated anammox granules system, achieving nitrogen removal efficiency exceeding 95 %. These results highlight the function of sulfide in improving the morphological property of anammox sludge as well as the creation of a favorable ecological niche for the functional microorganism, which is important to maintain the efficiency and robustness of the anammox process in treating wastewater.

Enhancement of the reactivation process of long-term starved anammox granular sludge with gravel balls: Microbial succession and metabolic impact

Anaerobic ammonium oxidation (Anammox) process is an economical and energy-efficient method of wastewater nitrogen removal. However, they are highly susceptible to starvation stress caused by sudden environmental changes. Rapid reactivation of starved anammox sludge is a crucial method to address seed sludge shortages and expand practical applications. This study investigated the impact of gravel balls on the reactivation of long-term starved anammox granular sludge (628 days). The results showed that gravel balls enhanced the recovery of nitrogen removal performance in starved anammox sludge, with nitrogen removal efficiency being 19.88% higher than the control group at the end of the recovery phase. The gravel balls also increased extracellular polymeric substance (EPS) secretion, contributing to the stability of the anammox system. Furthermore, the gravel balls promoted the proliferation of anammox bacteria, with the relative abundance of anammox bacteria reaching 38.25% on the 80th day. The analyses of microbial functions indicated that gravel balls facilitated cross-feeding and co-metabolism among microbes, while enhancing quorum sensing associated with anammox bacteria, forming a multifunctional community network centered on anammox bacteria. This indicates that gravel balls can effectively accelerate the reactivation process of long-term starved anammox sludge, aiding the reutilization of long-term starved anammox sludge.

Start-up of a full-scale two-stage partial nitritation/anammox (PN/A) process treating reject water from high solid anaerobic sludge digestion (HSAD)

High solid anaerobic digestion (HSAD) achieves the benefits of high volumetric loading rates and lower reject water production, which, however, results in much more concentrated reject water with a remarkable increase in organics and nitrogen compared with that from conventional AD with low solid content. The high concentrations of ammonium (2000–3500 mg/L) and COD (3000–4000 mg/L) were reported to exert inhibition on anammox bacteria (AnAOB), posing challenges to the application of the partial nitritation/anammox (PN/A). To date, no cases of PN/A process start-up for sludge HSAD reject water were reported. This study demonstrated the start-up process of a 480 m3/d PN/A project without anammox sludge inoculation and treating HSAD reject water from a centralized dewatered sludge treatment plant. The project did not construct new infrastructures but utilized previously constructed tanks to upgrade the process from existing short-cut nitrification-denitrification to a two-stage PN/A process. Although no external anammox sludge inoculation was performed to save seeding sludge cost, the start-up was successfully achieved in about 9 months (273 days) based on a three-step method of “AnAOB enrichment - sludge acclimation - capacity doubling”. During start-up, the relative abundance of AnAOB (Candidatus_Kuenenia) increased from near zero to 12.0%. After start-up, the total inorganic nitrogen (TIN) removal load reached 0.74 kgN/(m3•d), with a total nitrogen removal efficiency of over 90%. Compared to the traditional nitrification-denitrification process, the PN/A process remarkably reduces the addition of organic chemicals and aeration energy consumption, saving approximately 4.2 million yuan (RMB) in operational costs annually. In summary, this research provides a full-scale reference for the start-up of the PN/A process treating sludge HSAD reject water.

Efficient partial denitrification-anammox process enabled by a novel denitrifier with truncated nitrite reduction pathway

Partial denitrification coupled with anaerobic ammonium oxidation (PD-anammox) is a promising technology for cost-effective nitrogen removal from wastewater. Nitrite availability is crucial to anammox performance but often limited by the slow partial denitrification process. Here we report an efficient PD-anammox system driven by the novel denitrifier Bacillus velezensis C1–3 with truncated nitrite reduction pathway. Whole-genome sequencing analysis revealed that the lack of nitrite reductase genes nirS/nirK and norBC in strain C1–3 enabled nitrite accumulation without the need for precise control of carbon dosage. By coupling it with anammox sludge, over 79 % total nitrogen (TN) removal was stably achieved, under a TN loading rate of 660 mg/L/d and a carbon/nitrogen ratio below 1.0. Mechanism explorations indicate that the niche differentiation of C1–3 and anammox bacteria facilitated their mutualism while avoiding nitrite competition. This study demonstrates a novel strategy for establishing efficient PD-anammox process by harnessing the unique metabolic deficiency of denitrifiers, shedding light on the development of stable and sustainable biological nitrogen removal technologies with minimal carbon footprint.

Enhancing the nitrogen removal of anammox sludge by setting up novel redox mediators-mediated anammox process

Anaerobic ammonium oxidizing (anammox) bacteria have been proven weak-electroactive. However, the impact of exogenous anthraquinone-2,6-disulfonate (AQDS) on the anammox activity, although it usually plays essential roles in the life activities of many other electroactive microorganisms, is still unknown. Therefore, this study further explored the influences of AQDS on the anammox activity and the interaction mechanism with anammox bacteria, as well as the behaviors of NH4+, NO2−, and NO3−. The results showed that exogenous AQDS increased the ammonium and total nitrogen removal rates by 12.8% and 10.7%, respectively. Interestingly, the conversion from NO2− to NO3− was significantly reduced after adding AQDS, resulting in a 40.1% reduction in NO3− production of anammox process. In this study, we found for the first time that anammox bacteria could not only carry out the conventional anammox process but also perform a weak redox mediator-mediated anammox process, which could achieve the 1:1 consumption of NH4+ and NO2−. The redox mediator-mediated anammox process was related to an endogenous redox mediator (ERM) synthesized and secreted by anammox bacteria, whose redox midpoint potential was around −0.26 V (vs. Ag/AgCl). After adding AQDS, not only the ERM-mediated anammox process was enhanced, but also two novel redox mediator-mediated anammox processes were introduced, including the AQDS-mediated anammox process and ERM-AQDS-mediated anammox process. These three redox mediator-mediated anammox processes significantly improved the nitrogen removal performance of anammox bacteria and reduced energy consumption. These findings will help reduce the dependence of anammox technology on NO2−, reduce the cost of subsequent treatment of NO3−, and provide new visions for optimizing and applying anammox technology.

Achieving rapid start-up and efficient nitrogen removal of partial-denitrification/anammox process using organic matter in brewery wastewater as carbon source

ABSTRACT To find a cost-efficient carbon source for the partial denitrification/anaerobic ammonium oxidation (anammox) (PD/A) process, the practicability of using the organic matter contained in brewery wastewater as carbon source was investigated. Quick self-enrichment of denitrifying bacteria was achieved by supplying brewery wastewater as organic carbon source and using the mature anammox sludge as the seeding sludge. The PD/A process was successfully established after 33-day operation and then the average total nitrogen removal efficiency reached 92.29% when the influent CODCr: NO3 −-N: NH4 +-N ratio was around 2.5: 1.0: 0.67. The relative abundance of Thauera increased from 0.03% in the seeding sludge to 54.29% on day 110, whereas Candidatus brocadia decreased from 30.66% to 2.08%. The metagenomic analysis indicated that the sludge on day 110 contained more nar and napA (total of 41.24%) than nirK and nirS (total of 11.93%). Thus NO2 −-N was accumulated efficiently in the process of denitrification and sufficient NO2 −-N was supplied for anammox bacteria in the PD/A process. Using brewery wastewater as carbon source not only saved the cost of nitrogen removal but also converted waste into resource and reduced the treatment expense of brewery wastewater.

Crucial role of extracellular polymeric substances from anammox sludge in wastewater phosphorus recovery via magnesium phosphate crystallization

The magnesium phosphate (Mg-P) crystallization based on anaerobic ammonia oxidation is of interest for simultaneous nitrogen and phosphorus removal in wastewater treatment due to cost-effectiveness. Understanding the role of extracellular polymeric substances (EPS) in crystal growth is pivotal for bio-induced Mg-P crystallization. In this work, the effects of EPS in Mg-P crystallization were thoroughly investigated. Results indicated that EPS decreased the initial crystallization rate but slightly improved the final crystallization yield. The maximum removal efficiencies of Mg and P ions increased by 2.8 % and 3.1 %, respectively, when EPS concentration was 80 mg/L. Additionally, the presence of EPS enhanced Mg3(PO4)2·10 H2O production and facilitated the formation of larger rhombic plate-like structures crystals (maximum particle size increased by 215 %). Analysis of EPS composition changes during the crystallization process and characterization of the final crystals suggested that tryptophan-like proteins preferentially involved in the Mg-P crystallization process, while β-sheet proteins potentially influencing crystal morphology. Furthermore, the formation of phosphate ester groups, hydrogen bonding, and COOH-Mg2+ complexes were considered triggering factors for the interaction between EPS and Mg-P crystals. This study elucidated the underlying mechanism of EPS on Mg-P crystallization, further enhancing the understanding of the interaction between inorganic mineralization processes and microorganisms.

Enhanced-nitrogen removal through Fe(III)-triggered partial dissimilatory nitrate reduction to ammonium coupling with anammox in anammox bioreactor

Anammox is recognized as a prospective alternative for future biological nitrogen removal technologies. However, the nitrate by-products produced by anammox bacteria limit its overall nitrogen removal efficiency below 88 %. This study introduced Fe(III) into the anammox bioreactor to enhance the nitrogen removal efficiency to approximately 95 %, surpassing the biochemical limit of 88 % imposed by anammox stoichiometry. Anammox sludge was demonstrated to utilize extracellular polymeric substances to reduce Fe(III) into Fe(II), and this process promoted the dominance of Ca. Brocadia. The iron addition improved the abundance of narGHI genes and facilitated the partial dissimilatory nitrate reduction to ammonium, with nitrite as the end product. The accumulated nitrite was then eliminated through the anammox pathway, along with the excess ammonium (30 mg/L) in the influent. Overall, this study deepens our understanding of the enhanced nitrogen removal triggered by Fe(III) in anammox sludge and offers an effective approach to boost anammox process.

Feasibility of simultaneous optimization of Anammox start-up and nitrogen removal performance by intermittent dosing of nanoscale zero-valent iron

The long acclimation period and sensitivity to environmental conditions of Anammox are the bottlenecks for its promotion and application. An innovative strategy was adopted to accelerate functional microbial enhancement and improve nitrogen removal performance by inoculating cryopreserved Anammox sludge and activated sludge with intermittent dosing of nanoscale zero-valent iron (nZVI). The acclimation time was shortened by 76 days with nitrogen removal efficiency (NRE) reaching up to 91.07 %. Anammox, NDFO (nitrate/nitrite-dependent Fe(II) oxidation), Feammox (Fe(III) reduction coupled with anaerobic ammonium oxidation) and abiotic reactions were coupled in the system with nZVI, contributing to 69.79 %, 15.14 %, 9.84 % and 0.25 % of nitrogen removal, respectively. Further microbial analysis demonstrated significant enrichment of functional microorganisms, such as Candidatus Jettenia, Acidovorax and Comamonas. High-efficient nitrogen removal was attribute to the increase of functional genes involved in Anammox, electronic transfer, heme C synthesis and iron metabolism. This work provides an inspiring idea for the mainstream Anammox application.

Deciphering the anammox microbial community succession with humic acid exposure to optimize large anammox granules for robust nitrogen removal

The robustness of the anaerobic ammonia oxidation (anammox) process in treating wastewater with high concentrations of humic acids (HAs), including landfill leachate and sludge anaerobic digestion liquid, has been paid great attention. This study revealed that the anammox sludge granule size of 1.0–2.0 mm could be robust under the HA exposure with high concentrations. The total nitrogen removal efficiency (NRE) was 96.2% at the HA concentration of 20–100 mg/L, while the NRE was 88.5% at the HA concentration of 500 mg/L, with reduced by 7.7%. The increased extracellular polymeric substances (EPS) content which was stimulated by the HA exposure favored the formation of large granules (1.0–2.0 mm) by enveloping medium and micro granules (0.2–1.0 mm). The abundance of anammox bacteria Candidatus Brocadia was found to be higher (14.2%) in large anammox granules sized 1.0–2.0 mm, suggesting a potentially high anammox activity. However, the abundance of denitrifiers Denitratisoma increased by 4.3% in ultra-large anammox granules sized >2.0 mm, which could be attributed to the high EPS content for heterotrophic denitrifiers metabolism as organic matter. The feedback mechanism of the anammox community for maintaining the ecological function under the HA exposure resulted in a closely related microbial community, with positive and negative correlations in the ecological network increased by 64.3%. This study revealed that the HA exposure of the anammox system resulted in the anammox granules of 1.0–2.0 mm size being the dominant granules with robust nitrogen removal, providing significant guidance for the optimization of anammox granules for an efficient treatment of HA-containing wastewater in anammox applications.

Nitrogen removal performance and the biocenosis with microalgae consortium Nitrosifying and anammox bacteria in an upflow reactor

This study introduced an innovative pathway utilizing an algal anaerobic ammonium oxidation (ALGAMMOX) system to treat ammonium wastewater. Lake bottom sludge and anammox sludge were used to cultivate functional microorganisms and microalgae for nitrogen removal in an upflow reactor made of transparent materials. The results showed that the ALGAMMOX system achieved 87.40 % nitrogen removal when the influent NH4+-N concentration was 100 mg-N/L. Further analysis showed that anammox bacteria Candidatus Brocadia (8.87 %) and nitrosobacteria Nitrosomonas (3.74 %) were crucial contributors, playing essential roles in nitrogen removal. The 16S rRNA gene showed that the anammox bacteria in the sludge transitioned from Candidatus Kuenenia to Candidatus Brocadia. The 18S rRNA gene revealed that Chlamydomonas, Bacillariaceae and Pinnularia were the dominant microalgae in the system at a relative abundance of 7.99 %, 3.64 % and 3.14 %, respectively. This novel approach provides a theoretical foundation for ammonium wastewater treatment.

Characterization and Comparison of Anammox Immobilization in Polyvinyl Alcohol, Polyethylene Glycol and Water-Borne Polyurethane

Anammox bacteria were embedded with different mass fractions of polyvinyl alcohol (PVA), polyethylene glycol (PEG) and water-based polyurethane (WPU) materials. The embedded immobilized pellets with different particle sizes of about 2.8–3.2 mm were prepared. The effects of the mass fraction of the embedding material (PVA 6–12%, PEG 6–9%, WPU 10%) and the concentration of activated carbon added in the embedding process (0–4%) on the pellet was investigated. The performance of pellet formation, sedimentation rate, mechanical strength, expansion coefficient, and elasticity were compared and analyzed under different immobilization conditions, and the parameters of each embedding step were optimized. Anammox immobilized pellets prepared with 10% polyvinyl alcohol (PVA), 2% sodium alginate (SA), and 2% powdered activated carbon were proposed. The effects of salinity on anammox were investigated through a batch test, and the optimal reaction conditions were selected to carry out the operation test. The functional groups of embedded and unembedded anammox sludge were detected using the infrared spectrum. A continuous flow sequencing batch reactor (SBR) demonstrated stable operation with immobilized anammox. Scanning electron microscopy revealed that the immobilized anammox pellets appeared as irregular particles, with each micro-unit predominantly being spherical. Additionally, a minor presence of rod-shaped bacteria was also noted. After 30 days of stable operation of the reactor, the ammonia nitrogen removal rate reached 84.7%.

Enhanced sludge granulation and stable performance of an anammox expanded granular sludge bed (EGSB) reactor through the utilization of hydroxyapatite (HAP) particles

The reuse of hydroxyapatite particles (HAPs) as a granulation activator for anammox sludge was explored to address the remaining issues of time-consuming and unstable granular structure in anammox granulation. During the granulation, nitrogen removal capacity from 2.8 to 13.7 gN/L/d was obtained within 193 days, accompanied by an enhancement in bio-activity from 0.23 to 0.52 gN/gVSS/d. HAPs and anammox microorganisms coupled well to aggregate into granules for denser biomass, higher settleability, and stronger mechanical properties, which effectively improved the biomass retention capacity and structural strength of the sludge system. A skeleton structure formed by the HAPs was characterized during the transformation of the granules, playing a crucial role in strengthening the stability of the sludge. The intermediate processes of granulation were thus clarified to propose an evolutionary pathway for anammox-HAP granules. The pre-addition of HAPs is conducive to achieving faster anammox granulation and rapid process start-up for high-strength wastewater treatment.

External redox couple enhanced anammox sludge activity at low temperature: Insight into intracellular resource synthesis

Anaerobic ammonium oxidation (anammox), an energy-efficient deamination biotechnology, faces operational challenges in low-temperature environments. Enhancing the metabolic activity of anammox bacteria (AnAOB) is pivotal for advancing its application in mainstream municipal wastewater treatment. Inspired by the metabolic adaptability of AnAOB and based on our previous findings, this work investigated the enhancement of intracellular ATP and NADH synthesis through the exogenous supply of reduced humic acid (HAred) and H2O2 redox couple, aiming to augment AnAOB activity under low-temperature conditions. Our experimental setup involved continuous dosing of 0.0067 μmol g-1 volatile suspended solid of H2O2 and 10 mg g-1 volatile suspended solid of HAred into a mainstream anammox reactor operated at 15 °C with an influent TN content of 60 mg/L. The results showed that HAred / H2O2 couple succeeded in maintaining the effluent TN at 10.72 ± 0.91 mg l-1. The specific anammox activity, ATP and NADH synthesis levels of sludge increased by 1.34, 2.33 and 6.50 folds, respectively, over the control setup devoid of the redox couple. High-throughput sequencing analysis revealed that the relative abundance of Candidatus Kuenenia after adding HAred / H2O2 couple reached 3.65 % at the end of operation, which was 5.14 folds higher than that of the control group. Further metabolomics analysis underscored an activation in the metabolism of amino acids, nucleotides, and phospholipids, which collectively enhanced the availability of ATP and NADH for the respiratory processes. These findings may provide guidance on strategy development for improving the electron transfer efficiency of AnAOB and underscore the potential of using redox couples to promote the mainstream application of anammox technology.

Unveiling long-term combined effect of salinity and Lead(II) on anammox activity and microbial community dynamics in saline wastewater treatment

The study assessed the effect of salinity and lead (Pb(II)) on the anammox sludge for nitrogen removal from saline wastewater. Results showed decreased nitrogen removal and specific anammox activity (SAA) with elevated salinity and Pb(II). SAA reduced from 541.3 ± 4.3 mg N g−1 VSS d−1 at 0.5 mg/L Pb(II) to 436.0 ± 0.2 mg N g−1 VSS d−1 at 30 g/L NaCl, further to 303.6 ± 7.1 mg N g−1 VSS d−1 under 30 g/L NaCl + 0.5 mg/L Pb(II). Notably, the combined inhibition at salinity (15–20 g/L NaCl) and Pb(II) (0.3–0.4 mg/L) exhibited synergistic effect, while higher salinity and Pb(II) aligned with independent inhibition models. Combined inhibition decreased protein/polysaccharides ratio, indicating more severe negative effect on anammox aggregation capacity. Metagenomics confirmed decreased Candidatus Kuenenia, and enhanced denitrification under elevated salinity and Pb(II) conditions. This study offers insights into anammox operation for treating saline wastewater with heavy metals.

A new substrate equalization method for optimizing the influent conditions and fluid flow patterns of a multifed upflow anaerobic sludge blanket reactor with mature anammox granules

To improve nitrogen removal efficiency (NRE) and achieve homogenous distribution of anammox sludge and substrate, a new substrate equalization theory and a cumulative overload index was proposed for multifed upflow anaerobic sludge bed (MUASB) reactors with mature anammox granules. The performance and flow patterns of MUASB reactors were investigated under various influent conditions. The results showed that the nitrogen removal performance and stability of MUASB reactors could be optimized by minimizing the cumulative load. The NRE gradually increased from 83.3 ± 2.2 %, 86.8 ± 4.2 % to 89.3 ± 4.1 % and 89.7 ± 1.6 % in feeding flow tests and feeding port tests, respectively. Furthermore, the flow patterns were compared based on residence time distribution and computational fluid dynamics, indicating that a better equilibrium distribution of microorganisms and substrates could be achieved in the MUASB reactors under the lowest cumulative load. Therefore, substrate equalization theory can be used to optimize the nitrogen removal performance of MUASB reactors with low-carbon footprints.

Metabolic Profiles and Microbial Synergy Mechanism of Anammox Biomass Enrichment and Membrane Fouling Alleviation in the Anammox Dynamic Membrane Bioreactor.

The anammox dynamic membrane bioreactor (DMBR) is promising in applications with enhanced anammox biomass enrichment and fouling alleviation. However, the metabolic mechanism underlying the functional features of anammox sludge and the biofilm membrane is still obscure. We investigated the metabolic networks of anammox sludge and membrane biofilm in the DMBR. The cooperation between anammox and dissimilatory nitrate reduction to ammonium processes favored the robust anammox process in the DMBR. The rapid bacterial growth occurred in the DMBR sludge with 1.33 times higher biomass yield compared to the MBR sludge, linked to the higher activities of lipid metabolism, nucleotide metabolism, and B vitamin-related metabolism of the DMBR sludge. The metabolism of the DMBR biofilm microbial community benefited the fouling alleviation that the abundant fermentative bacteria and their cooperation with the anammox sludge microbial community promoted organics degradation. The intensified degradation of foulants by the DMBR biofilm community was further evidenced by the active carbohydrate metabolism and the upregulated vitamin B intermediates in the biofilms of the DMBR. Our findings provide insights into key metabolic mechanisms for enhanced biomass enrichment and fouling control of the anammox DMBR, guiding manipulations and applications for overcoming anammox biomass loss in the treatment of wastewater under detrimental environmental conditions.

Mechanistic insight into the aggregation ability of anammox microorganisms: Roles of polarity, composition and molecular structure of extracellular polymeric substances

The chemical characteristics of extracellular polymeric substances (EPS) of anammox bacteria (AnAOB) play a crucial role in the rapid enrichment of AnAOB and the stable operation of wastewater anammox processes. To clarify the influential mechanisms of sludge EPS on AnAOB aggregation, multiple parameters, including the polarity distribution, composition, and molecular structure of EPS, were selected, and their quantitative relationship with AnAOB aggregation was analyzed. Compared to typical anaerobic sludge (anaerobic floc and granular sludge), the anammox sludge EPS exhibited higher levels of tryptophan-like substances (44.82–56.52 % vs. 2.57–39.81 %), polysaccharides (40.02–53.49 mg/g VSS vs. 30.22–41.69 mg/g VSS), and protein structural units including α-helices (20.70–23.98 % vs. 16.48–19.32 %), β-sheets (37.43–42.98 % vs. 25.78–36.72 %), and protonated nitrogen (Npr) (0.065–0.122 vs. 0.017–0.061). In contrast, it had lower contents of β-turns (20.95–27.39 % vs. 28.17–39.04 %). These biopolymers were found to originate from different genera of AnAOB. Specifically, the α-helix-rich proteins were mainly derived from Candidatus Kuenenia, whereas the extracellular proteins related to tryptophan and Npr were closely associated with Candidatus Brocadia. Critically, these EPS components could drive anammox aggregation through interactions. Substantial amounts of tryptophan-like substances facilitated the formation of β-sheet structures and the exposure of internal hydrophobic clusters, which benefited the anammox aggregation. Meanwhile, extracellular proteins with high Npr content played a pivotal role in the formation of mixed protein-polysaccharide gel networks with the electronegative regions of polysaccharides, which could be regarded as the key component in the maintenance of anammox sludge stability. These findings provide a comprehensive understanding of the multifaceted roles of EPS in driving anammox aggregation and offer valuable insights into the development of EPS regulation strategies aimed at optimizing the anammox process.

Granulation characteristics of anammox sludge in response to different signal-molecule-stimulants; mediated through programmed cell death

During the anammox process, mitigation of biomass washout to increase sludge retention is an important parameter of process efficiency. Signal molecular stimulants (SMS) initiate the sludge granulations controlled by programmed cell death (PCD) of microorganisms. In this study, the aerobic granular sludge (AGS), cell fragments, extracellular polymeric substances (EPS), and AGS process effluent were tested as SMS to identify their effect on anammox granulation. The results showed that the addition of SMS increased the nitrogen removal efficiency to varying degrees, whereas the addition of AGS process supernatant, as SMS, increased the ammonia removal efficiency up to 96%. The addition of SMS was also found to increase EPS production and contributed to sludge granulation. In this process, the proportion of PCD increased and both Gaiella and Denitratisoma abundance increased from 3.54% to 5.59%, and from 1.8% to 3.42%, respectively. In conclusion, PCD was found important to increase anaerobic ammonia oxidation performance through the granulation mechanism.

Roles of red mud-based biochar carriers in the recovery of anammox activity: characteristics and mechanisms.

Anaerobic ammonium oxidation (anammox) sludge is easily deactivated in the process of treating ammonia-laden wastewater. To investigate an effective recovery method, red mud-based biochar carriers (RMBC) were prepared and added to a deactivated anammox reactor; the operation of this reactor had been interrupted for 6months with starvation and low temperature. The deactivated sludge with added RMBC was recovered rapidly after 31days, with the specific anammox activity rapidly increasing to 0.84g N/(g VSS∙day), and the recovery efficiency of nitrogen removal rate increased by four times compared to the unadded control. The granulation degree and extracellular polymeric substances secretion of the anammox sludge with the added RMBC were significantly higher than that of the control group. In addition, a large number of spherical anammox bacteria were observed moored at the porous channels of RMBC, and the copy numbers of functional genes of anammox bacteria were approximately twice that of the control group. Hence, RMBC is a potential sludge activator, and it can provide a "house" to protect anammox bacteria, enhance the metabolic activity and the agglomerative growth of anammox bacteria, and synergistically achieve rapid recovery of deactivated anammox sludge.