Anammox Consortia Research Articles

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.

Insights into the carbon and nitrogen metabolism pathways in mixed-autotrophy/heterotrophy anammox consortia in response to temperature reduction

While the multi-coupled anammox system boasts a substantial research foundation, the specific characteristics of its synergistic metabolic response to decreased temperatures, particularly within the range of 13-15°C, remained elusive. In this study, we delve into the intricate carbon and nitrogen metabolism pathways of mixed-autotrophy/heterotrophy anammox consortia under conditions of temperature reduction. Our macrogenomic analyses reveal a compelling phenomenon: the stimulation of functional genes responsible for complete denitrification, suggesting an enhancement of this process during temperature reduction. This adaptation likely contributes to maintaining system performance amidst environmental challenges. Further metabolic functional recombination analyses highlight a dramatic shift in microbial community composition, with denitrifying MAGs (metagenome-assembled genomes) experiencing a substantial increase in abundance (up to 200 times) compared to autotrophic MAGs. This proliferation underscores the strong stimulatory effect of temperature reduction on denitrifying species. Notably, autotrophic MAGs play a pivotal role in supporting the glycolytic processes of denitrifying MAGs, underscoring the intricate interdependencies within the consortia. Moreover, metabolic variations in amino acid composition among core MAGs emerge as a crucial adaptation mechanism. These differences facilitate the preservation of enzyme activity and enhance the consortia's resilience to low temperatures. Together, these findings offer a comprehensive understanding of the microbial synergistic metabolism within mixed-autotrophy/heterotrophy anammox consortia under temperature reduction, shedding light on their metabolic flexibility and resilience in dynamic environments.

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.

Bacterial communication intelligently regulates their interactions in anammox consortia under decreasing temperatures

Bacterial communication could affect their interactions, but whether this regulation has “intelligence” is still unknown. Here, we operated an anammox reactor under temperature gradient from 35 °C to 15 °C. As results, expression abundance of bacterial communication genes increased by 12 % significantly after temperature declined. Division of labor among distinct signal molecules was evidenced by complementary roles of acyl-homoserine lactones (AHLs) and diffusible signal factor (DSF) in affecting bacterial interactions and niche differentiation respectively. DSF based inter-and intra-communication helped bacteria match their investments and rewards during cross-feedings. When temperature was below 25 °C, transcription regulator Clp governed by DSF inclined to promote folate and molybdenum cofactor biosynthesis, which coincidentally benefited one anammox species more than another. Meanwhile, for the anammox species with lower benefits, Clp also inclined to decrease biosynthesis of costly tryptophan and vitamin B1 rewarding others. Interestingly, bacterial communication inclined to influence the bacteria with many cooperators in the community or with high capacity to export cofactors for cross-feedings when temperature decreased. As results, these bacteria were enriched which could lead to closer interactions in whole community to adapt to low temperatures. The discovered intelligence of bacterial communication opened another window for understanding bacterial sociobiology.

High dosage of exogenous glycine in cross-feeding breaks the equilibrium of anammox system

Strategies to break the cross-feeding equilibrium by dosing essential metabolites and thus enriching and isolating anammox bacteria (AnAOB) have been proposed, but amino acids have not been investigated as important cross-feeding pathways in anammox consortia. In this study, long-term experiments were designed to verify the effect of exgenous glycine on anammox consortia and changes in the bacterial community and metabolic activity after the addition of glycine in a highly active anammox system were investigated. The results showed that the impact of glycine on the activity of anammox highly depended on glycine concentrations, which was manifested as a promotive effect at low concentrations (0–150 mg/L) and an inhibitory effect at high concentrations (300–600 mg/L). The addition of low concentrations of glycine not only enhanced the diversity and richness of bacterial communities but also promoted the enrichment of Candidatus Kuenenia. The qPCR results showed that the content of functional genes Hzs, NirS and anammox 16S rRNA increased while the expression of amoA and NrfA genes was suppressed when glycine acted positively on AnAOB; Hzs and amoA were suppressed when glycine acted negatively on AnAOB, while the expression of NirS and NrfA genes increased dramatically. Specific anammox activity (SAA) experimental results show that 50–150 mg/L glycine was the optimal amount for promoting the activity of AnAOB. Analysis of community metabolic functions indicated that the addition of exogenous glycine resulted in a imbalance of glycine cross-feeding and promoted AnAOB enrichment mainly by affecting pathways such as other glycan degradation and Lysosome. Overall, this study provided new insights into understanding the glycine cross-feeding, which is essential for improving anammox-based technologies.

Quorum sensing mediated response mechanism of anammox consortia to anionic surfactant: Molecular simulation and molecular evidence

The widespread use of surfactants raise challenges to biological wastewater treatment. Anaerobic ammonium oxidation (anammox) process has the potential to treat wastewater containing anionic surfactants, but the response of anammox consortia at the molecular level under long-term exposure is unclear. Using high-throughput sequencing and gene quantification, combined with molecular docking, the effect of sodium dodecyl sulfonate (SDS) on anammox consortia were investigated. Levels of reactive oxygen species (ROS) might be lower than the threshold of oxidative damage, while the increase of lactate dehydrogenase (LDH) represented the cell membrane damage. Decreased abundance of functional genes (hdh, hzsA and nirS) indicated the decrease of the anammox bacterial abundance. Trace amounts of N-acyl homoserine lactone (AHL, C6-HSL, C8-HSL and C12-HSL) contained in influent could induce endogenous quorum sensing (QS), which could regulate the correlation between functional bacteria to optimize the microbial community and strengthen the resistance of anammox consortia to SDS. In addition, the proliferation of disinfectant resistance genes might increase the environmental pathogenicity of sewage discharge. This work highlights the potential response mechanism of anammox consortium to surfactants and provides a universal microbial-friendly bioenhancement strategy based on QS.

Deciphering behaviors of 6:2 chlorinated polyfluorinated ether sulfonate (alternative-PFOS) on anammox processes: Nitrogen removal efficiency and microbial adaptability

This study investigates the behaviors and effects of F-53B, an alternative to perfluorooctane sulfonate on anaerobic ammonium oxidation (anammox) processes. Results showed that the nitrogen removal efficiency (NRE) reached 83.8 % at a F-53B concentration of 0.5 mg·L−1, while NRE decreased to 66.9 % with 5 mg·L−1 of F-53B. The defluorination rates of 17.8 % (0.5 mg·L−1) and 9.3 % (5 mg·L−1) were observed, respectively, suggesting the occurrence of F-53B degradation. The relative abundance of Ca. Kuenenia decreased from 26.1 % to 16.2 % with the F-53B concentration increasing from 0.5 mg·L−1 to 5 mg·L−1. Meanwhile, Denitratisoma was selectively enriched with a relative abundance of 40.7 % at an F-53B concentration of 0.5 mg·L−1. Ca. Kuenenia could reduce reactive oxygen species induced by F-53B to maintain the balance of oxidative stress. This study gains insight into the behaviors and metabolic mechanisms of F-53B in anammox consortia, suggesting the feasibility of anammox processes for industrial wastewater.

Quest for Nitrous Oxide-reducing Bacteria Present in an Anammox Biofilm Fed with Nitrous Oxide.

N2O-reducing bacteria have been examined and harnessed to develop technologies that reduce the emission of N2O, a greenhouse gas produced by biological nitrogen removal. Recent investigations using omics and physiological activity approaches have revealed the ecophysiologies of these bacteria during nitrogen removal. Nevertheless, their involvement in‍ ‍anammox processes remain unclear. Therefore, the present study investigated the identity, genetic potential, and activity‍ ‍of N2O reducers in an anammox reactor. We hypothesized that N2O is limiting for N2O-reducing bacteria‍ ‍and an‍ ‍exogeneous N2O supply enriches as-yet-uncultured N2O-reducing bacteria. We conducted a 1200-day incubation of N2O-reducing bacteria in an anammox consortium using gas-permeable membrane biofilm reactors (MBfRs), which efficiently supply N2O in a bubbleless form directly to a biofilm grown on a gas-permeable membrane. A 15N tracer test indicated that the supply of N2O resulted in an enriched biomass with a higher N2O sink potential. Quantitative PCR and 16S rRNA amplicon sequencing revealed Clade II nosZ type-carrying N2O-reducing bacteria as protagonists of N2O sinks. Shotgun metagenomics showed the genetic potentials of the predominant Clade II nosZ-carrying bacteria, Anaerolineae and Ignavibacteria in MBfRs. Gemmatimonadota and non-anammox Planctomycetota increased their abundance in MBfRs despite their overall lower abundance. The implication of N2O as an inhibitory compound scavenging vitamin B12, which is essential for the synthesis of methionine, suggested its limited suppressive effect on the growth of B12-dependent bacteria, including N2O reducers. We identified Dehalococcoidia and Clostridia as predominant N2O sinks in an anammox consortium fed exogenous N2O because of the higher metabolic potential of vitamin B12-dependent biosynthesis.

Salinity alters the toxicity of copper nanoparticles on anammox consortia through modulating extracellular polymeric substances and membrane permeability

Salinity alters the toxicity of copper nanoparticles on anammox consortia through modulating extracellular polymeric substances and membrane permeability

Integration of double nitrite autotrophic shunt enhances anammox-based wastewater treatment for sustainable nitrogen removal and energy efficiency

Anammox holds significant promise as a next-generation technology for achieving energy-positive wastewater treatment. However, its application remains challenging due to unstable nitrite supply and excessive nitrate residue. To address this, a groundbreaking Anammox bioprocess incorporating a double nitrite autotrophic shunt was developed within single system to realize energy-efficient nitrogen removal from wastewater. Partial nitrification/Anammox converted a portion of ammonia in leachate to nitrogen gas in oxic stage. Significantly, sulfur-driven partial denitrification utilizing sulfide as electron donor, efficiently reduced generated nitrate to nitrite in subsequent anoxic stage, facilitating nitrite reutilization via Anammox and removing residual ammonia. Over 200-day monitoring, bioprocess demonstrated desirable effluent quality with 9.0 mg/L NH4+-N, 0.6 mg/L NO2−-N, and 6.2 mg/L NO3−-N. Delightedly, efficient nitrogen removal efficiency of 97.8 % was achieved even under fluctuation conditions. Priority position held by Anammox consortium (Candidatus Kuenenia 11.2 %) mainly derives from similar growth rate of autotrophic bacteria within their shared ecological niche and stable nitrite supply, enabling Anammox to contribute impressive 72.4 % of nitrogen removal. By combing electron flow preference kinetic, stability toward fluctuation, core microbial community, the ecological niche of Candidatus Kuenenia and self consistent logic governing functional microorganisms were revealed. The cost-effective system exhibited significant advantages over conventional bioprocess, reducing organic matter demand by 100 %, aeration energy consumption by 61.7 %, and biomass production by 82.9 %.

Simultaneous removal of NH4+ and SO42− in Sulfate-reducing anammox scale reactor using FDAARGOS_798 strain/Anammox integration

In this study, an exploratory experiment conducted to assess the efficiency of time reducing Bacillus cereus FDAARGOS_798 strain in the Sulfate-reducing anaerobic ammonium oxidation –Sulfammox- process at lab-scale reactor. The sulfammox process was examine in four different semi-continuous stirred (SCS) reactors receiving a treated synthetic wastewaters by 1 % of FDAARGOS_798 strain acclimated and carried with 2 % anaerobic granular sludge (sludge) in the presence and absence of the 1 % anammox consortia granular sludge (Anammox). Subsequently, the previous work outcomes of optimized anaerobic acclimation method isolating FDAARGOS_798 strain. The SCS-2 reactor with sulfammox integration (FDAARGOS_798/Anammox) has shown significant performance in anoxic simultaneous removal of NH4 + and SO42-, 77 ± 0.2 % and 50 ± 1.0 %, respectively, with the sole substrate (NH4)2SO4 0.1 g/L. The NH4+ and SO42- average decrease of 40 ± 0.2 % and 46 ± 0.17 % in 26 days, respectively, in SCS-4 reactor treated by sulfammox (FDAARGOS_798/Anammox) in (NH4)2SO4 0.5 g/L. Subsequently, the N/S ratio is calculated at around 1.8 ± 0.2 as evidence of reaching sulfammox process. The sulfammox process could not detect at SCS-1and SCS-3 reactors which are in the absence of sulfammox integration. Meanwhile the exact sulfammox process pathway was not detectable due to complicated scenarios. In conclusion, the addition of Bacillus ceruse FDAARGOS_798 carried by Anammox consortium is a well indicate to an effective and economical way that accelerate the start-up of the sulfate-reducing anammox process.

Perturbation and mechanism of non-antibiotic drug in regulating resistome and metabolome of anammox consortia: An overlooked and underrated cause of multiresistance

Given that antibiotic resistance genes (ARGs) become emerging pollutants, their accumulation and transfer caused by extensive use of antibiotic and non-antibiotic drugs raise concerns. In contrast, the impacts of non-antibiotic drugs have been largely overlooked, and their regulation mechanism in anammox system remains unclear. In this study, the single and combined effects of typical antibiotic (doxycycline) and non-antibiotic drug (propranolol) on anammox consortia were evaluated and compared from process performance, community succession, metabolic and gene regulation perspectives. Significant reduction in the nitrogen removal efficiency (49.0 ± 0.5%) was only observed in the anammox process stressed by propranolol and doxycycline, but it was reversible. Metabolome results showed that anammox consortia upregulated amino acid metabolism under propranolol stress, while the abundance of pyrimidine and purine metabolic pathways significantly increased under doxycycline and multidrug stress. Microorganisms adapted to these stresses by changing the utilization preference for metabolites. Metagenomic analyses indicated that doxycycline and propranolol induced the high frequency of gene transfer through increasing oxidative stress level, membrane permeability, pili and efflux pumps. Significant correlations between ARGs and mobile genetic elements (MGEs), especially intI1, demonstrated the conjugative transfer of ARGs. Furthermore, structural equation models showed that microbial community, bacterial metabolism, MGEs, reactive oxygen species (ROS) and corresponding SOS response were key driving factors for ARGs transfer in the anammox systems. These findings fill the gap in the effect of non-antibiotic drugs on anammox system and their regulation mechanism, which further provides a theoretical guidance for anammox-based process to treat multidrug-containing wastewater.

The evolution of nitrogen transformation microorganism consortium under continued manganese domestication conditions

Manganese redox-stimulated bioremediation of nitrogen wastewater is receiving increasing attention. However, the nitrogen metabolic capacity and community evolution during manganese-mediated nitrogen transformation process under continued manganese domestication conditions are ambiguous. In this study, nitrogen- metabolizing microbial consortiums were incubated with synthesized Mn-humic acid complex (Mn-HA) for one month (M1), three months (M2) and six months (M3), respectively. During the Mn-HA incubation period, Bio-MnOx accompanying with bacterial consortiums (MnOB consortiums) with high TIN removal capacities were obtained. The TIN removal rates in M1, M2 and M3 were 0.220, 1.246 and 4.237 mg·L−1·h−1, respectively, which were 15.961, 90.006 and 1550.006 times higher than CK (Control Check group, no Mn-HA added group) (0.014 mg·L−1·h−1), respectively. Functional genes (amoA, AMX and narG) were most abundant in M3, which was associated with the highest nitrogen removal rate in M3. MnOB1 (bacterial consortium in M1), including Geobactor, Geothrix, Anaeromyxobacter and Bacillus, may be responsible for the Mnammox-NDMO (MnOx reduction coupled to ammonium oxidation – nitrate/nitrite-dependent low-valent Mn oxidation) process. MnOB3 (bacterial consortium in M2) enriched nitrifying bacteria Ellin6067, and denitrifying bacteria Denitratisoma, which dominated nitrogen transformation. MnOB6 (bacterial consortium in M3) enriched denitrifiers Denitratisoma, nitrifiers Ellin6067 and potential anammox bacteria SM1A02, Candidatus_Brocadia. Combined with the reduced abundance of Nitrospirae, a short-cut partial nitrification and denitrification (PND) or partial nitrification, denitrification and anammox (PNDA) could occurred in M2 and M3. It is suggested that community may have evolved into an energetically efficient short-cut nitrification, denitrification and anammox consortium to replace the full-range nitrification and denitrification community in M1 and CK under the continued manganese domestication conditions. Enhanced metabolic pathways of hydroxylamine oxidation and the nitric oxide reduction may confirm that PND or PNDA occurred in M2 and M3.

Enhancing anammox nitrogen removal by static magnetic field exposure: Performance, microbial community and symbiotic relationship analysis

Anammox batch reactors were operated with different static magnetic field (SMF) intensities (0 mT, 40 mT, and 100 mT). The impacts of SMF and the influent nitrogen concentration on nitrogen removal were analyzed. Sludge characteristics, microbial community structure, and metabolism pathways were explored. The results show that a high nitrogen load greatly decreased the nitrogen removal efficiency. Still, applying SMF with appropriate intensity enhanced the nitrogen removal performance, especially under high nitrogen loading conditions. Systems with 40 mT SMF at normal nitrogen loading conditions had the highest total nitrogen removal efficiency. In contrast, a higher SMF intensity was conducive to the anammox process with a high nitrogen load. Candidatus Kuenenia was fast enriched under high nitrogen loading conditions in the control systems. However, Candidatus Brocadia was more abundant than Ca. Kuenenia in the systems with SMF application. SMF was the main factor affecting the microbial metabolism, enhancing the metabolic pathways related to bacterial growth and resistance to environmental stress. The metabolic interactions between the anammox bacteria and other bacteria were enhanced by applying SMF to the anammox process, especially for the cross-feeding of cofactors and vitamins in the anammox consortia.

Succession of microbial communities reveals the inevitability of anammox core in the development of anammox processes

The lack of anammox seeds is regarded as the bottleneck of anammox-based processes. Although the interactions in anammox consortia have attracted increasing attention, little is known about the influence of inoculated sludge populations on the growth of anammox bacteria. In this study, four sludge of distinct communities mixed with anammox sludge (the relative abundance of Ca. Kuenenia was 1.96 %) were used as the seeds, respectively for the start-up of anammox processes. Notably, all these mixed microbial communities tend to form a similar microbial community, defined as the anammox core, containing anammox-bacteria (22.9 ± 5.9 %), ammonia-oxidizing-bacteria (0.8 ± 0.7 %), nitrite-oxidizing-bacteria (0.2 ± 0.2 %), Chloroflexi-bacteria (0.7 ± 0.4 %), and heterotrophic-denitrification-bacteria (0.3 ± 0.2 %). It also elucidated that the communities of Nitrosomonas-dominated sludge were the closest to the anammox core, and achieved the highest nitrogen-removal rate of 0.73 kg-N m−3 d−1. This study sheds light on the solution to the shortage of anammox seeds in the full-scale wastewater treatment application.

Metagenomic insights into the symbiotic relationship in anammox consortia at reduced temperature

Anammox as a promising biological nitrogen removal technology has attracted much attention. However, cold temperature would limit its wide application and little is known about the microbial interactions between anammox bacteria (AnAOB) and heterotrophic bacteria at cold temperature. Here, we observed reduced temperature (25–15 °C) promoted the secretion of EPS and thus stimulated bigger size of granular sludge in a laboratory-scale anammox reactor. We further combined co-occurrence network analysis and genome-centered metagenomics to explore the potential interactions between AnAOB and heterotrophic bacteria. Network analysis suggested 22 out of 25 positively related species were reported as definite heterotrophic bacteria in subnetwork of AnAOB. Genome-centered metagenomics analysis yielded 23 metagenomic assembly genomes (MAGs), and we found that Acidobacteriota-affiliated bacteria could biosynthesize most polysaccharides (PS) precursors and contain the most glycosyltransferases and transporters to facilitate exopolysaccharides biosynthesis, together with partial PS precursors produced by AnAOB. AMX1 as the only anammox genome could synthesize most amino acids and cross feed with some heterotrophs to affect the extracellular protein function. Additionally, Bacteroidota, Planctomycetota, Chloroflexota, and Proteobacteria could contribute folate and molybdopterin cofactor for AMX1 to benefit their activity and growth. Superphylum Patescibacteria could survive by cross-feeding with AnAOB and heterotrophic organisms about organic compounds (Glyceraldehyde-3P and lactate). These cross-feedings maintained the stability of anammox reactor performance and emphasize the importance of heterotrophs in anammox system at reduced temperature.

The organics-mediated microbial dynamics and mixotrophic metabolisms in anammox consortia under micro-aerobic conditions

The engineering applications of mainstream anaerobic ammonium oxidation (anammox) have raised increasing attention due to its energy-efficient, however, the organics-mediated microbial dynamics and mixotrophic metabolisms in anammox consortia under micro-aerobic conditions are still elusive. Here, the response of the anammox process to sodium acetate and glucose at a C/N ratio ranging from 0 to 0.5 was investigated under micro-aerobic conditions, respectively. Results showed that the additional glucose could promote the nitrogen removal efficiency (NRE) and nitrogen removal rate (NRR) of anammox processes at a low C/N ratio (0.3), representing 84.00% and 0.53 N kg·m−3·d−1. The introduced organics could regulate the diversity of the microbial community and simplify the microbial relationship in anammox consortia. Anammox could not benefit from the introduced sodium acetate, while glucose could effectively enhance the anammox activity and microbial interactions in anammox consortia. Glucose might also stimulate the mixotrophic mechanism of Ca. Kuenenia, further promotes the proliferation of anammox sludge under micro-aerobic conditions. This study reveals that glucose could positively mediate microbial interactions and mixotrophic metabolism in anammox consortia under micro-aerobic conditions, which raises a new horizon for the proliferation of anammox sludge for mainstream engineering applications.

Metabolomic pathway regulation to achieve optimal control of inorganic carbon in anammox process

The significance of inorganic carbon (IC) for anaerobic ammonium oxidation (anammox) bacteria has been verified. However, the regulation of metabolic pathways under IC stress is not clear, limiting the optimization of IC supply. In this study, the regulatory pathways at IC concentration of 5–150 mg/L were explored to achieve optimal control of IC. The results show that the changes of metabolic pathway under IC stress determined anammox characteristics. At IC concentration of 5 mg/L, the anammox activity distinctly decreased due to the guanosine tetraphosphate (ppGpp) -mediated regulation under IC limitation. With less than 15 mg/L of IC, the decrease of carbon fixation limited the biosynthesis of gluconeogenesis and amino acids, causing the decline of extracellular polymeric substance synthesis. With more than 50 mg/L of IC, the improvement of purine and pyrimidine metabolism enhanced the electron transport capacity and growth potential of anammox bacteria. This study provides metabolic insights into IC influence on anammox consortia and a novel method of IC concentration optimization using metabolomics analysis.

Microbial dynamics reveal the adaptation strategies of ecological niche in distinct anammox consortia under mainstream conditions

The feasibility of anammox-based processes for nitrogen-contained wastewater treatment has been verified with different anammox bacteria, however, the ecological niche of anammox bacteria under mainstream conditions is still elusive. In this study, six sludge samples collected from different habitats were utilized to culture anammox bacteria under mainstream conditions, and two distinct anammox genera (Ca. Kuenenia and Ca. Brocadia) with a relative abundance of 6.31% (C1) and 3.09% (C3), respectively, were identified. Notably, the microbial dynamics revealed that anammox bacteria (AMX), ammonia-oxidizing bacteria (AOB), nitrite-oxidizing bacteria (NOB), Chloroflexi bacteria (CFX), and heterotrophic denitrification bacteria (HDB) were the core members in anammox consortia. However, Ca. Kuenenia and Ca. Brocadia occupied different ecological niches in anammox consortia. The dissolved oxygen and microbial structures of the anammox-continuous stirred tank reactor systems were the main factors to affect their niche differentiation. Meanwhile, comammox might exist in the systems and occupy the ecological niche of AOB in nitrogen cycling. The network analysis suggested that Ignavibacterium could be the associated bacteria in Ca. Kuenenia-dominated consortia, while Ca. Nitrotoga was that in the Ca. Brocadia-dominated consortia. Our findings reveal a valuable reference for the observation of distinct anammox genera under mainstream conditions, which provides theoretical guidance for the engineering application of mainstream anammox-based processes.

Genome-centric metagenomics insights into functional divergence and horizontal gene transfer of denitrifying bacteria in anammox consortia

Denitrifying bacteria with high abundances in anammox communities play crucial roles in achieving stable anammox-based systems. Despite the relative constant composition of denitrifying bacteria, their functional diversity remains to be explored in anammox communities. Herein, a total of 77 high-quality metagenome-assembled genomes (MAGs) of denitrifying bacteria were recovered from the anammox community in a full-scale swine wastewater treatment plant. Among these microbes, a total of 26 MAGs were affiliated with the seven dominant denitrifying genera that have total abundances higher than 1%. A meta-analysis of these species suggested that external organics reduced the abundances of genus Ignavibacterium and species MAG.305 of UTPRO2 in anammox communities. Comparative genome analysis revealed functional divergence across different denitrifying bacteria, largely owing to their distinct capabilities for carbohydrate (including endogenous and exogenous) utilization and vitamin (e.g., pantothenate and thiamine) biosynthesis. Serval microbes in this system contained fewer genes encoding biotin, pantothenate and methionine biosynthesis compared with their related species from other habitats. In addition, the genes encoding energy production and conversion (73 genes) and inorganic ion transport (53 genes) putatively transferred from other species to denitrifying bacteria, while these denitrifying bacteria (especially genera UTPRO2 and SCN-69–89) likely donated the genes encoding nutrients (e.g., inorganic ion and amino acid) transporter (64 genes) for other members to utilize new metabolites. Collectively, these findings highlighted the functional divergence of these denitrifying bacteria and speculated that the genetic interactions within anammox communities through horizontal gene transfer may be one of the reasons for their functional divergence.