Anammox Process Research Articles

Arginine enhances activity of anammox consortia and process stability with increased nitrogen loading

Cross-feeding based on amino acids metabolism is an important strategy by which anammox bacteria and the co-existing heterotrophs facilitate their own growth and survival. Arginine is one of the necessary amino acids required for bacterial protein biosynthesis but whether adding arginine could benefit growth of anammox bacteria remains unknown. In this study, arginine was supplemented at dose of 5 mg·L−1 to promote the nitrogen removal performance of anammox bioreactors under varied loading rates. The results showed that nitrogen removal efficiency increased by 10.2 % under higher loading rates. Arginine addition substantially simulated the secretion of extracellular proteins and polysaccharides within anammox consortia as a strategy against unfavorable conditions. Canditatus Kuenenia dominated the anammox consortia and their 16S rRNA abundance and anammox-related functional genes were significantly increased by up to 0.42 times and 5.81 times, respectively. The findings of this study provided a feasible strategy to improve the performance of anammox reactors with arginine supplementation.

The green footprint of anammox processes under simulated actual operating conditions: Focusing on the nitrous oxide and methane production

The anammox process has attracted increasing attention due to its advantages of low-carbon and energy-saving, nevertheless, greenhouse gas was still generated during its engineering applications process. Hence, it is vital to comprehensively understand the production characteristics and mechanisms of N2O and CH4 in anammox processes by responding to practical conditions including dissolved oxygen, temperature, and salinity. Results showed that N2O production increased by 192 %–358 %, while nitrogen removal efficiency (NRE) increased by 64.2 %–86.8 % with increasing temperature. The increased salinity inhibits 40.60 %–65.33 % N2O production with a decrease NRE of 7.85 %–18.2 %. CH4 production was the highest at 18–27 °C, reaching 3.07 ± 0.11–4.06 ± 0.16 mg·L−1, which were 1.59–2 and 1.29–1.38 times higher than that at 8–17 °C and 28–37 °C, respectively. Denitratisoma, Thauera, and Nitrosomonas were the main functional microbes for greenhouse gas production in anammox consortia. Notably, H2O2-induced intracellular Fenton reaction may be critical for the CH4 production in anammox consortia. This work provides valuable insights into achieving efficient nitrogen removal and minimizing carbon footprint in anammox systems and provides a theoretical basis for implementing the net-zero emission idea in wastewater treatment plants.

Formate-facilitated recovery of wastewater anammox granular sludge from oxygen inhibition: Effects and mechanisms

Oxygen leakage due to poor reactor sealing or accidental aeration can have detrimental impacts on the performance of wastewater anammox processes. Strategies of restoring functionality of anammox consortia are scarce, and providing appropriate chemical reducing reagents may help reduce oxidative damage. This study characterized the responses of anammox granular sludge (AnGS) to oxygen inhibition, and identified formate as an effective promoter capable of facilitating activity recovery, while other organics (acetate, glucose, methanol) were less effective. Upon 12 h of air exposure (DO: 1.2 ± 0.1 mg/L), the total nitrogen removal efficiency of a lab-scale AnGS reactor decreased from 85.15 % to 45.46 %. Compared with natural recovery (without chemical addition), potassium formate at 10 mg COD/L for 3 days significantly increased the total nitrogen removal efficiency, both in the short-term (66.68 % vs 57.09 %, 3 days post inhibition) and long-term (82.34 % vs 72.88 %, 30 days post inhibition). By examining sludge morphology, metagenome and transcriptome, the following potential mechanisms were proposed: 1) rapidly decreasing intracellular ROS; 2) enhancing nitrite turnover; 3) maintaining granular sludge integrity; 4) facilitating interspecies metabolic interactions. The ability of formate to facilitate sludge recovery from oxygen inhibition also implies its potential in mitigating other oxidative stresses in wastewater anammox reactors.

Response of ammonium transformation in bioanodes to potential regulation: Performance, electromicrobiome and implications

Understanding how potential regulation affects ammonium transformation in bioanodes is crucial for promoting their application. This study explored the performance, electrochemical properties, electromicrobiome of bioanodes across potentials from 0.0 V to 0.4 V vs. standard hydrogen electrode (SHE). Higher anode potentials enhanced the performance of electroactive biofilms and ammonium removal but suppressed nitrite oxidation while favoring dissimilatory nitrate reduction (DNRA), leading to increased nitrite accumulation. A reduction in nitrite-oxidizing bacteria (NOB) and an increase in DNRA-related genes resulted in an optimal nitrite-to-ammonium ratio of 1.32 for the Anammox process. Higher anodic potentials (0.3 and 0.4 V) were less effective for TN removal than lower potentials (0, 0.1, and 0.2 V), likely due to increased NOB and denitrification genes at lower potentials enhancing nitrite oxidation and denitrification. These findings indicate that regulating anodic potential effectively directs ammonium transformation in bioanodes, optimizing its conversion to N2 or nitrite.

Enhanced nitrogen removal from secondary effluent of municipal wastewater using denitrification filter: Feasibility of refractory organics as a carbon source

Conventional advanced nitrogen removal in municipal wastewater is hindered by the limited availability of carbon sources in the secondary effluent. However, refractory organics present in it had the potential to serve as intrinsic carbon sources after hydrolysis for nitrogen removal via simultaneous denitrification and partial-denitrification anammox (PDA) processes. To assess this potential, a denitrification filter was set up in this study to evaluate its feasibility of concurrent processes. Results showed that increasing influent ammonium (NH4+-N) from 1.0 to 7.0 mg/L increased total nitrogen (TN) removal from 52.4 % to 89.9 %. Simultaneous occurrence of PDA and denitrification process were confirmed by the actual chemical oxygen demand (COD) consumption (0.8–1.2 mg/mg TN removal) from non-fluorescent organics. The presence of the anammox, hydrolytic and denitrifying bacteria further supported the achievement of nitrogen removal through PDA and denitrification processes by utilizing hydrolytic products biodegraded from refractory organics.

Monitoring the Biochemical Activity of Single Anammox Granules with Microbarometers.

Granule-based anaerobic ammonium oxidation (Anammox) is a promising biotechnology for wastewater treatments with extraordinary performance in nitrogen removal. However, traditional analytical methods often delivered an average activity of a bulk sample consisting of millions and even billions of Anammox granules with distinct sizes and components. Here, we developed a novel technique to monitor the biochemical activity of individual Anammox granules in real-time by recording the production rate of nitrogen gas with a microbarometer in a sealed chamber containing only one granule. It was found that the specific activity of a single Anammox granule not only varied by tens of folds among different individuals with similar sizes (activity heterogeneity) but also revealed significant breath-like dynamics over time (temporal fluctuation). Statistical analysis on tens of individuals further revealed two subpopulations with distinct color and specific activity, which were subsequently attributed to the different expression levels of heme c content and hydrazine dehydrogenase activity. This study not only provides a general methodology for various kinds of gas-producing microbial processes but also establishes a bottom-up strategy for exploring the structural-activity relationship at a single sludge granule level, with implications for developing a better Anammox process.

Surface-Adhered Microbubbles Enhance the Resistance of ANAMMOX Granular Sludge to Sulfamethoxazole Stress.

The granule-based anammox process has been reported to be more resistant to the stress of antibiotics; however, the underlying resistance mechanism is still not fully understood. In this study, we found that more microbubbles stably adhered to the surface layer of anammox granular sludge (AnGS, Gs) operating under long-term sulfamethoxazole (SMZ) stress of 2 mg/L, compared to that in the control reactor (Gc). The difference in covering content can be up to over three times (1.0 ± 0.1% vs 0.3 ± 0.0%). Batch tests showed that the coverage ratio of microbubbles on Gs reached approximately 32.5%, which significantly reduced SMZ transfer into AnGS due to the adsorption of SMZ by bubbles, thus alleviating the inhibition of anammox bacterial activity by 36.5%. The adhesion force between the microbubbles and the surface layer of Gs was found to be largely enhanced by 75.0% compared to that of Gc. The increased hydrophobicity of surface layer due to the increased extracellular polymer substance (EPS, mainly proteins) content, and the larger capillary force of surface layer, owing to the unique micronano structure, were identified as key factors responsible for the stable adhesion of microbubbles on the Gs. Consequently, this study demonstrated the vital roles of the surface-adhered microbubbles in resisting the stress of SMZ and shed light on the regulation and development of robust granule-based anammox processes.

Low Strength Wastewater Treatment Using a Combined Biological Aerated Filter/Anammox Process

To achieve the in situ capacity expansion of the post-denitrification biological aerated filter (BAF-DN), the integration of BAF with the anammox process (BAF/AX) was proposed. With the objective of maximizing retaining ammonia nitrogen, the operational optimization of BAF was achieved by two distinct strategies. The treatment performance of BAF demonstrated that the removal efficiencies of chemical oxygen demand (COD) and ammonia nitrogen (NH4+-N) was 66.3~67.3% and 4~12%, respectively, under conditions of low aeration intensity (0.4 m3·m−2·h−1) or a shortened empty bed residence time (EBRT) of 30 min. Residual NH4+-N in the BAF effluent served as the ammonia substrate for the subsequent anammox process, which was successfully launched by using ceramic particles and sponges as carriers. Notably, the sponge carrier facilitated a shorter start-up period of 41 to 44 days. Furthermore, the sponge-based anammox reactor exhibited a superior NH4+-N removal capacity (≥85.7%), under operations of a shorter EBRT of 40 min, low influent NH4+-N concentrations (≤30 mg/L), and COD levels of ≤67 mg/L. In addition, a comprehensive evaluation of the BAF/AX process was conducted, which considered performance, cost-effectiveness, and engineering feasibility. The performance results illustrated that the effluent quality met the standard well (with a COD level of ≤ 50 mg/L, and a TN of ≤3.1~10.5 mg/L). Following a comparison against the low aeration intensity operation, it was recommended to operate BAF at a low EBRT within the BAF/AX process. Consequently, the treated volume was double the volume of the standalone BAF-DN, synchronously achieving low costs (0.413 yuan/m3).

Synchronous and efficient removal of carbon, nitrogen, and phosphorus from actual rural sewage by composite wetlands enhanced with functional fillers

A composite wetland (CECW) was constructed by introducing P-adsorption filler (EPAF) and activated sludge into traditional wetlands for treating actual sewage. The results showed that EPAF improved P removal through physico-chemical adsorption, and it could be stably regenerated after adsorption saturation without potential risks. Meanwhile, zeolite promoted NH4+-N reduction in sewage by cation exchange. In addition, simultaneous biological removal of carbon, nitrogen, and phosphorus was achieved through nitrification, denitrification, anammox, and aerobic P-accumulation processes induced by Nitrobacter, Proteus Hauser, Candidatus Paracaedibacter, and Brevundimonas. Under the coupling of filler interception/adsorption, microbial assimilation/transformation, flocculation, and plant uptake, CECW obtained the removal rates of 93.22 %, 85.75 %, 91.80 %, 95.38 %, 97.07 %, and 78.05 % for turbidity, TN, NH4+-N, TP, PO43−-P, and TCOD, which met the Class 1A standard (GB18918-2002). Therefore, the experiment systematically investigated the effects and mechanism of CECW in treating actual sewage, which could provide reference for rural sewage treatment and sludge utilization.

Meta-analysis: Global patterns and drivers of denitrification, anammox and DNRA rates in wetland and marine ecosystems

Nitrogen cycling is one of the most important biogeochemical processes on Earth, and denitrification, anammox and DNRA processes are important nitrogen cycling processes in estuarine ecosystems. However, due to the large input of anthropogenic nitrogen sources, a large number of environmental problems have now occurred in the estuary. But the global patterns and controlling factors of denitrification, anammox and DNRA rates in wetland marine ecosystems are not yet known. We reached our conclusions through a global synthesis of 546 observation sites from 78 peer-reviewed papers: The three rates were generally higher in areas near wetlands than in coastal areas. The rate of denitrification was highest in the subtropical region the seasonal variability was not significant; and TOC was the main factor controlling denitrification. The rate of anammox was significantly higher in the subtropical region than in the tropical and boreal zones, and the seasonal variability was significant; and at the same time, TN was the main driver of the anammox rate of the wetland ocean. DNRA rates were significantly higher in the tropics than in the subtropics and temperate zones; and the main driver of DNRA rates was temperature. Nitrogen cycle functional genes also had an indirect effect on their rates. With NH4 + -N significantly affecting nirK abundance and TN significantly affecting the gene abundance of nirS; TOC and TN had a greater effect on hzo abundance, which indirectly affected anammox rates; for DNRA, C/N significantly affects the gene abundance of nrfA, which indirectly affects the DNRA rate. Therefore, the findings of this study indicate that physicochemical indicators about N and climatic characteristics have a profound effect on the nitrogen cycling process, which provides a good feedback for studying the role of denitrification and provides a positive impact on global climate and environmental governance.

Achieving advanced nitrogen removal with anammox and endogenous partial denitrification driven by efficient hydrolytic fermentation of slowly-biodegradable organic matter

Anammox-based processes are pivotal for elevating nitrogen removal efficiency in municipal wastewater treatment. This study established a novel HF-EPDA system combined in-situ hydrolytic fermentation (HF) with endogenous partial denitrification (EPD) and anammox. Slowly-biodegradable organic matter (SBOM) was degraded and transformed into endogenous polymers for driving production of sufficient nitrite by EPD, further promoted the nitrogen removal via anammox process. Processes above formed positive feedback, guaranteeing the robustness and recoverability of system. After a 92-day suspension during operation, advanced nitrogen removal was still achieved with excellent nitrogen removal efficiency of 95.84 ± 1.73 %, treating with actual domestic wastewater and synthetic nitrate wastewater. Candidatus Brocadia and Candidatus Competibacter were dominant bacteria on biofilms responsible for the anammox and EPD process respectively, while the main hydrolytic fermentation organisms norank_o SBR1031 was enriched in floc sludge. This study highlights the reliable potential for expanding anammox application with simultaneous improvement of SBOM utilization.

Microbial Coadaptation Drives the Dynamic Stability of Microecology in Mainstream and Sidestream Anammox Systems Under Exposure of Progesterone

Microbial cooperation determines the efficacy of wastewater biological treatment, and the adaptability of microorganisms to environmental stresses varies. Recently, extensive use of hormones results in their inevitable discharge into aquatic environment. Therefore, mainstream and sidestream anammox reactors were constructed in this study to evaluate their removal performance of progesterone and nitrogen simultaneously, the adaptability of anammox consortia to progesterone stress and the corresponding regulation mechanism. Both anammox processes had the resilience to progesterone stress, with the average nitrogen removal efficiency exceeding 90%. At the same time, progesterone removal efficiency also exceeded 70%. In contrast, microbial community in the mainstream reactors was more susceptible to progesterone interference. The adaptation of anammox consortia mainly depended on microbial cooperation and molecular regulation. Initially, bacteria secreted more extracellular polymeric substances to detain progesterone. Biodegradation also contributed to mitigating the side effect of progesterone, which was demonstrated by the proliferation of potential degrading bacteria such as Bacillus salacetis, Bacillus wiedmannii and Rhodococcus erythropolis. In addition, the enhancement of microbial interaction intensity drove their cooperation to enhance adaptability and maintain stable performance. Combined with metagenomic and metatranscriptomic analyses, such microbial adaptability was enhanced through molecular regulations, including the energy redistribution for amino acid synthesis and alteration of key metabolic pathways. Related functional gene expressions and microbial interactions were, in turn, regulated by quorum sensing. This work verifies the feasibility of anammox process in hormone-containing wastewater treatment and provides a holistic understanding of molecular mechanism of microbial interaction and coadaptation to stress.

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.

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.

Linking ecological niche and metabolic hierarchy in low-oxygen-driven anammox systems

Dissolved oxygen (DO) is regarded as a key regulatory factor for the practical engineering applications of anaerobic ammonium oxidation (anammox) processes, however, the adaptability and metabolic mechanisms of floc anammox consortia with low-oxygen as potential substrates involved in the holistic microbial metabolic system are still elusive. This study showed that the nitrogen removal efficiency (NRE) of floc anammox consortia was 16.45% and 35.71% higher at DO of 0.2–0.5 mg·L−1 than at DO <0.2 mg·L−1 and DO of 0.5–0.8 mg·L−1, respectively. Meanwhile, the activity of anammox bacteria (AnAOB) and ammonia-oxidizing bacteria (AOB) was the highest, representing 22.64 ± 3.20 mg N·g−1 V SS·d−1 and 103.22 ± 6.47 mg N·g−1 VSS·d−1, respectively. AnAOB applied the antioxidant enzymes (CCP, T-SOD) and Cbb3-type cytochrome c oxidase (Cbb3-type CcO) for oxygen detoxification and utilization. Moreover, AOB, aerobic/anoxic heterotrophic bacteria, denitrifying bacteria, etc. defined as the core members in anammox consortia could also consume DO and adjust the NH4+-N/NO2−-N ratio for AnAOB, establishing potent microbial interactions to relieve the pressure of DO on AnAOB and improve NRE. This study reveals that anammox consortia can utilize limited DO as a substrate to promote anammox processes, which advances the practical applications of anammox processes.

Enrichment and characterization of thermophilic anaerobic ammonium oxidizing bacteria from hot spring

Anaerobic ammonium oxidization (Anammox) process plays a crucial role in the global nitrogen cycle and sustainable biological nitrogen removal from wastewater. Although Anammox bacteria have been detected across mesophilic and thermophilic conditions, the direct cultivation of Anammox bacteria from thermal environments has remained elusive. This impedes limiting our understanding of their physiology and ecology in high-temperature habitats. Here, we successfully enriched Anammox bacteria from hot spring sediments at 45 °C, achieving an ammonium oxidation rate of 158.0 mg NH4+-N l-1d-1, with the genus ‘Candidatus Brocadia’ presenting 22.9 % of the total microbial community after about 500 days of operation. Metagenomic analysis recovered two high-quality genomes of novel Anammox bacteria, which we designed as ‘Candidatus Brocadia thermophilus’ and ‘Candidatus Brocadia thermoanammoxidans’. Both of them encoded and actively expressed key metabolic genes involved in Anammox process and several genes associated with thermotolerance, demonstrating their remarkable ability to perform Anammox reaction in thermophilic environments. Notably, phylotypes related to ‘Candidatus Brocadia thermoanammoxidans’ have frequently been retrieved from geographically distinct natural habitats. These findings expand our understanding of thermophilic Anammox bacteria and underscore their potential in the nitrogen cycle of thermal natural and engineering ecosystems.

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.

Responses of suspended sludge and biofilm in a SNAD system under C/N elevation: Microbial activity, nitrogen conversion flux and molecular ecological network

The simultaneous partial nitrification, anammox and denitrification (SNAD) process had received widespread attention as an advanced wastewater treatment process. In this study, the SNAD mainstream nitrogen removal process with the incorporation of polyurethane sponge packing under different C/N conditions was investigated. Results showed that the highest nitrogen removal efficiency of the system was achieved at the C/N of 2.0, while the high C/N (3.5) significantly deteriorate the nitrogen removal efficiency. Meanwhile, high C/N (3.5) significantly inhibited the activity and abundance of anammox bacteria (mainly Candidatus_Kuenenia), resulting in the decreased contribution of anammox (from 63.14 % to 48.09 %). The significant divergence of microbial interactions in the suspended sludge and biofilm was observed with increasing C/N. Compared with suspended sludge, biofilm facilitated higher abundance and activity of anammox bacteria, and the molecular ecological network of biofilm displayed better stability and more efficient mass transfer efficiency between microorganisms. The C/N of 3.5 simplified the subnetworks of Chloroflexi and Proteobacteria but increased the positive interactions between Planctomycetota and other microbes. Anammox bacteria were found as keystone species only in biofilm system. This study provided a theoretical basis and technical guidance for the application of SNAD process in municipal wastewater treatment.

Polystyrene nanoparticles regulate microbial stress response and cold adaptation in mainstream anammox process at low temperature

Nanoplastic pollution has become one of the most pressing environmental issues, and its bioaccumulation in aquatic environment also causes a great difficulty in treatment. Therefore, this work investigated the microbial dynamics of mainstream anaerobic ammonia oxidizing (anammox) process to treat the wastewater containing typical nanoplastics, as well as the fate and regulation mechanism of polystyrene nanoparticles (PS-NPs) with different concentrations. The results showed that 0.1–0.5 mg L-1 of PS-NPs had no significant effect on the nitrogen removal efficiency (NRE). When the concentration of PS-NPs increased from 0.5 mg L-1 to 2 mg L-1, the NRE of R1 with PS-NPs decreased from 94.9 ± 2.3 % to 77.0 ± 1.6 %, while the control reactor R0 maintained a stable NRE. Notably, the relative abundance of Ca. Kuenenia decreased from 17.4 % to 14.8 %, and that of Ca. Brocadia slightly decreased from 5.9 % to 5.0 % in R1. In addition, PS-NPs induced oxidative stress in anammox consortia, leading to the significant increase in reactive oxygen species (ROS) and lactate dehydrogenase (LDH) as well as cell membrane damage. PS-NPs also downregulated the content of heme c and further inhibited anammox activity. Based on the molecular docking simulation and western blotting, cold shock proteins (CSPs) could bind to PS-NPs and reduce the performance of anammox processes at low temperatures.

Autotrophic biological nitrogen removal in anammox system at elevated temperature: EPS regulation and metagenomic analysis

Carbon reduction in wastewater treatment industry has been attracted global attention. Anaerobic ammonium oxidation (anammox) process is known as a promising and energy-efficient technology. In this study, the effects of elevated temperature on anammox system were thoroughly investigated. Analyses of effluent and microbial community indicated that long-term exposure to 45℃ led to system collapse and a huge shift in community structure. Network analysis revealed that the sudden drop in abundance of AnAOB at 45℃ (from 36.19 % to 10.71 %) made them uncompetitive with heterotrophic bacteria. The decrease of EPS concentration (a minimum of 7.22 mg·g−1-VSS at 45℃) combined with the enhancement of EPS metabolism at higher temperature suggested a shift in the role of EPS from protective function at 35℃ to utilization as a carbon source for heterotrophic bacteria at higher temperatures. Through constructing the cross-feeding relationships associated with amino acids, polysaccharides, and cofactors, we found the dependence of heterotrophs on AnAOB became weaker at high temperature, and the disturbances in interspecific relationships might be responsible for the collapse of the anammox system. This work presents significant insights into the engineering of anammox process under high temperature.