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.

Partitioned granular sludge coupling with membrane-aerated biofilm reactor for efficient autotrophic nitrogen removal.

The partial nitritation-anammox process based on a membrane-aerated biofilm reactor (MABR) faces several challenges, such as difficulty in suppressing nitrite-oxidizing bacteria (NOB), excessive effluent nitrate, and ineffective synergy between denitrification and anammox bacteria. Therefore, a novel partitioned granular sludge coupling with MABR (G-MABR) was constructed. The chemical oxygen demand (COD) and nitrogen removal efficiency were 88.8 ±â€¯1.8 %-92.6 ±â€¯1.2 % and 88.8 ±â€¯1.5 %-93.6 ±â€¯0.7 %, respectively. The COD was mainly lowered in the lower granular sludge-zone, while nitrogen was removed in the upper MABR-zone. NOB was significantly suppressed in the MABR-zone due to competition for substrate with denitrifying bacteria and anammox bacteria. This partitioned configuration reduced the C/N ratio in the MABR-zone, thus facilitating autotrophic nitrogen removal. Both partial nitrification and denitrification provided nitrite for anammox bacteria in granular sludge, whereas partial nitrification mainly supplied nitrite to the anammox bacteria in membrane biofilms.

Achieving endogenous partial denitrification by cultivating denitrifying glycogen-accumulating organisms.

Although anaerobic ammonia oxidation (anammox) is considered a promising process due to its high efficiency and low energy in nitrogen removal, nitrite inadequacy was one of the bottlenecks for the application of anammox. However, endogenous partial denitrification (EPD) has been emerging as a stable pathway to provide nitrite for anammox. Furthermore, denitrifying glycogen-accumulating organisms (DGAOs) are believed to be associated with EPD. In this study, firstly, GAOs were gradually enriched in a sequencing batch reactor (SBR) with the dual strategy of influent phosphorus limitation and withdrawal after the anaerobic stage. DGAOs were successfully induced by adding sodium nitrate solution at the end of the anaerobic stage, resulting in NO3--N concentration increasing from 15 to 30 mg/L. During a typical SBR cycle, DGAOs contributed up to 96% of the conversion of intracellular carbon sources and up to around 95% of nitrate reduction during the anoxic stage. The maximum nitrate-to-nitrite transformation ratio (NTR) of the system reached 80%. Microbial community analysis demonstrated that the Ca. Compatibactors were the dominant functional bacteria for EPD, with a relative abundance of 31.12%. However, the relative abundance of phosphorous-accumulating organisms (PAOs) was only 1.02%. This study reveals the important role of DGAOs in the EPD process, which can provide a stable nitrite for anammox.

Metabolic evolution and bottleneck insights into simultaneous autotroph-heterotroph anammox system for real municipal wastewater nitrogen removal.

When biological nitrogen removal (BNR) systems shifted from treating simulated wastewater to real wastewater, a microbial succession occurred, often resulting in a decline in efficacy. Notably, despite their high nitrogen removal efficiency for real wastewater, anammox coupled systems operating without or with minimal carbon sources also exhibited a certain degree of performance reduction. The underlying reasons and metabolic shifts within these systems remained elusive. In this study, the simultaneous autotrophic/heterotrophic anammox system demonstrated remarkable metabolic resilience upon exposure to real municipal wastewater, achieving a nitrogen removal efficiency (NRE) of 82.83 ± 2.29 %. This resilience was attributed to the successful microbial succession and the complementary metabolic functions of heterotrophic microorganisms, which fostered a resilient microbial community. The system's ability to harness multiple electron sources, including NADH oxidation, the TCA cycle, and organics metabolism, allowed it to establish a stable and efficient electron transfer chain, ensuring effective nitrogen removal. Despite the denitrification channel's nitrite supply capability, the analysis of the interspecies correlation network revealed that the synergistic metabolism between AOB and AnAOB was not fully restored, resulting in selective functional bacterial and genetic interactions and the system's PN/A performance declined. Additionally, the enhanced electron affinity of PD increased interconversion of NO3--N and NO2--N, limiting the efficient utilization of electrons and thereby constraining nitrogen removal performance. This study elucidated the metabolic mechanism of nitrogen removal limitations in anammox-based systems treating real municipal wastewater, enhancing our understanding of the metabolic functions and electron transfer within the symbiotic bacterial community.

Characterization of EPS subfractions from a mixed culture predominated by partial-denitrification functional bacteria.

Extracellular polymeric substances (EPS) play a crucial role in the aggregation of partial denitrification (PD) consortia, as EPS is closely linked to bioreactor performance. However, the structural and compositional properties of EPS from PD consortia have not yet been investigated. In this study, photometric measurements indicated that PD consortia contained significantly more EPS (168.81 ± 2.10 mg/g VSS) compared to conventional activated sludge (79.79 mg/g VSS). The EPS of PD consortia exhibited a significant predominance of proteins over polysaccharides, with a protein/polysaccharide ratio of 1.43 ± 0.10. FTIR analysis revealed that the EPS of PD consortia contained fewer hydrophilic functional groups, particularly carboxyl and carbonyl groups, indicating a high aggregation potential. The content comparison of EPS and functional groups across three stratified EPS subfractions from PD consortia consistently followed the sequence: TB-EPS > LB-EPS > S-EPS. XPS results corroborated the FTIR findings and the protein/polysaccharide ratio determined by photometric measurements, all of which suggested that the EPS of PD consortia exhibited a higher abundance of hydrophobic functional groups. However, the higher α-helix/(ß-sheet + random coil) ratio (0.99) suggested that the proteins in PD consortia had a compact structure, making inner hydrophobic groups difficult to expose. This compact protein structure could limit aggregation among bacterial cells, indicating the need for process optimization to enhance sludge aggregation in PD-related processes. Overall, understanding the aggregation characteristics of PD consortia could improve the application of PD-based processes.

Advanced N removal from low C/N sewage via a plug-flow anaerobic/oxic/anoxic (AOA) process: Intensification through partial nitrification, endogenous denitrification, partial denitrification, and anammox (PNEnD/A).

Achieving low-cost advanced nitrogen (N) removal from municipal wastewater treatment plants (WWTPs) remains a challenge. A plug-flow anaerobic/oxic/anoxic (AOA) system with a mixtures bypass (MBP) integrating partial nitrification (PN), endogenous carbon denitrification (EnD), partial denitrification (PD), and anaerobic ammonium oxidation (Anammox), was constructed to treat actual sewage with a low C/N ratio. The effluent concentrations and removal efficiency of total inorganic nitrogen (TIN) during stable operation were 2.9 ± 0.9 mg/L and 93.1 ± 2.0 %, respectively. EnD was enhanced by the MBP through the efficient utilization of polyhydroxyalkanoates generated in the anaerobic zone. PD was promoted by the addition of carries and sodium acetate to the anoxic tank and the subsequent implantation of the Anammox biofilm successfully coupled PD/A. Stable PN was obtained with a satisfactory nitrite accumulation ratio of 92.6 %, facilitated by carriers and the introduction of hydroxylamine in the oxic zone. Mass balance analysis revealed that EnD and Anammox contributed 40.8 % and 48.2 % of TIN removal, respectively. The enrichment and synergistic effects of ammonia-oxidizing bacteria, denitrifying bacteria, glycogen-accumulating organisms, and anaerobic ammonia-oxidizing bacteria formed a diverses bacterial basis for the establishment of PN, EnD, PD, and Anammox (PNEnD/A) in the AOA system. The successful integration of PNEnD/A into the AOA process provides an innovative approach for low-cost advanced N removal in WWTPs.

Genomic synergistic efficient carbon fixation and nitrogen removal induced by excessive inorganic carbon in the anammox-centered coupling system.

Given the significance of HCO3- for autotrophic anammox bacteria (AnAOB), excessive HCO3- was always provided in anammox-related systems and engineering applications. However, its impact mechanism on anammox process at genome-level remains unknown. This study firstly established an anammox-centered coupling system that entails heterotrophic partial denitrification (PD) and hydrolytic acidification (A-PDHA) fed mainly with inorganic carbon (high HCO3- concentration and low C/N ratio). Metagenomic binning and metatranscriptomics analyses indicated that high HCO3- concentration enhanced expression of natural most efficient phosphoenolpyruvate (PEP) carboxylase within AnAOB, by up to 30.59 folds. This further induced AnAOB to achieve high-speed carbon-fixing reaction through cross-feeding of phosphate and PEP precursors with heterotrophs. Additionally, the enhanced activity of transporters and catalytic enzymes (up to 4949-fold) induced by low C/N ratio enabled heterotrophs to eliminate extracellular accumulated energy precursors mainly derived from carbon fixation products of AnAOB. This maintained high-speed carbon-fixing reaction within AnAOB and supplemented heterotrophs with organics. Moreover, assimilated energy precursors stimulated nitrogen metabolism enzymes, especially NO2- reductase (968.14 times), in heterotrophs. This established an energy-saving PD-A process mediated by interspecies NO shuttle. These variation resulted in efficient nitrogen removal (>95 %) and reduced external organic carbon demand (67 %) in A-PDHA system. This study unveils the great potential of an anammox-centered autotrophic-heterotrophic coupling system for achieving cost-effective nitrogen removal and enhancing carbon fixation under excessive HCO3- doses.

Enhancing collaboration of anammox with heterotrophic microbes mediated selectively by iron of different valences: Activities balance, metabolic mechanism, and functional genes regulation.

The partial denitrification/anammox (PD/A) process is receiving increasing attention due to its cost-effectiveness advantages. However, effective strategies to alleviate organic matter inhibition and promote anammox activity have been proven to be a big challenge. This study investigated the effects of three types of iron (nano zero-valent iron (nZVI), Fe(II), and Fe(III)) on the PD/A process. It is worth noting that nZVI of 5-50 mg/L and Fe(III) of 5-120 mg/L promoted both PD and anammox activity. Long-term intermittent addition of nZVI (50 mg/L) resulted in a nitrogen removal efficiency of 98.2% in the mixotrophic PD/A system driven by iron and organic matter. The contribution of anammox for nitrogen removal reached as high as 93.8%. The organic carbon demand decreased due to the external electron donor provided by nZVI for PD. Multiple Fe-N metabolic pathways, primarily involving ammonia oxidation by Fe(III) and nitrate reduction by nZVI, play a crucial role in facilitating nitrogen transformation. Conversely, the direct addition of 30-120 mg/L Fe (II) resulted in a significant decrease in pH to below 5.0 and severe inhibition of PD and anammox activity. Following prolonged operation in the presence of nZVI, it was demonstrated that there is an enhancing effect on robust nitrite production for anammox. This was accompanied by a remarkable up-regulation of genes encoding nitrate reductase and iron-transporting proteins dominated by Thauera. Overall, this study has provided an efficient approach for advanced nitrogen removal through organic- and iron-driven anammox processes.

Enhanced and robust nitrogen removal using an integrated zeolite and partial denitrification anammox process.

Anammox has received increased attention due to its enhanced and cost-efficient approach to nitrogen removal. However, its practical application is complicated by strict influent NO2--N to NH4+-N ratio demands and an 11% nitrate production from the anammox process. This study was the first known research to propose and verify a system of zeolite integrated with partial denitrification and anammox (Z-PDA) in an up-flow anaerobic sludge bed (UASB) reactor. The enhanced and robust nitrogen removal resulted in an ultra-high nitrogen removal efficiency (NRE, 93.0 ± 2.0%). Zeolite adsorption and biological desorption of ammonium contributed to robust nitrogen removal with fluctuating influent NO2--N to NH4+-N ratios. Applying 16S rRNA gene sequencing found that Candidatus Brocadia and Thauera were the key bacteria responsible for anammox and partial denitrification (PD), respectively. Zeolite also acted as a biological carrier. This significantly enriched anammox bacteria with a higher relative abundance of Candidatus Brocadia, reaching 49.2%. Metagenomic analysis demonstrated that the multiple functional genes related to nitrogen removal (nrfA/H, narG/H/I) and the metabolic pathways (Biosynthesis of cofactors, the Two-component system, the Biosynthesis of nucleotide sugars, and Purine metabolism) ensured the resilience of the Z-PDA system despite influent fluctuations. Overall, this study provided novel insights into the impacts of zeolite in the PDA system. It described the fundamental mechanism of zeolite based on adsorption and biological desorption, and demonstrated a meaningful application of the anammox process in sewage treatment.

Optimizing nitrogen removal in PD/A reactors: Effects of influent composition and temperature on system stability and microbial dynamics.

This study investigates the performance and microbial community dynamics in two partial denitrification/anammox (PD/A) reactors with different influent wastewater compositions (differ in the presence/absence of NO2-) subjected to a controlled temperature gradient reduction from mesophilic (30 °C) to room temperature (20.92 °C) over 76 days. Two lab-scale PD/A reactors (R1 and R2), both operated with a total inorganic nitrogen (TIN) concentrations of 70 mg N/L. R1 maintained a NH4+/NO2-/NO3- ratio of 3:3:1 and a COD/NO3- ratio of 2.0, while R2 had an NH4+/NO3- ratio of 3:4, and COD/NO3- ratios of 2.0 and 2.5. Our findings reveal distinct responses to the temperature transitions: the optimization of the NH4+/NO2-/NO3- ratio at 3:3:1 facilitated more stable nitrogen removal as temperatures decreased. This stability can be attributed to the enhanced synchronization between anammox bacteria and denitrifiers, promoting a balanced bioconversion process that is less susceptible to temperature-induced disruptions. Notably, the specific anammox activity (SAA) in both reactors declined linearly with the decrease in temperature, but the relative abundance of anammox bacteria (Ca. Brocadia) in R1 increased from 2.1 % to 9.7 %. Furthermore, the percentage of anammox-related key genes was higher in R1 than in R2, suggesting a microbial mechanism underlying the stable performance of R1. These results underscore the significant impact of influent nitrogen composition on PD/A performance amid temperature gradients and highlight the critical role of optimizing influent ratios for maintaining efficient nitrogen removal. This study offers valuable insights into enhancing the stability of PD/A systems under varying thermal conditions.

Effect of plant species on wastewater treatment performance of a subsurface vertical-flow constructed wetland with step-feeding at low temperature.

To improve the treatment performance of constructed wetlands under low-temperature conditions, this study investigated the effects of plant species on wastewater treatment performance at low temperature and the associated microbiological characteristics in a subsurface vertical-flow constructed wetland (VFCW) with step-feeding. The results showed that the redox microenvironment in the VFCW filter with step-feeding could be restored and optimized by planting appropriate species that can tolerate low temperature, ensuring a high nitrification performance for the system. Correspondingly, the abundance and activity of three functional microbes (namely nitrifiers, denitrifiers, and anammox bacteria) increased to different degrees in the system, eventually ensuring ideal nitrogen removal by the VFCW. Compared with the VFCW planted with Phragmites australis and Acorus gramineus, the operation performance of the VFCW planted with Iris wilsonii could be recovered at low temperature, and its chemical oxygen demand, total phosphorus, total nitrogen, and ammonium nitrate removal rates could respectively reach 95.7%, 99.2%, 93.0%, and 94.4%, respectively. Moreover, nitrogen removal in the system relied on the nitrification/denitrification and partial denitrification - anaerobic ammonium oxidation processes. Nitrosomonas, Nitrospira, Thauera, and Candidatus Brocadia were the four dominant bacterial genera in the filter layer.

Increasing biomass concentration facilitates simultaneous nitrogen removal and sludge reduction under low C/N conditions.

To overcome the issues of limited carbon source and high sludge production in partial denitrification/anammox (PD/A) process, the effects of mixed liquor suspended solids (MLSS) and carbon/nitrogen ratio (C/N) on PD/A were investigated through parallel experiments. Nitrogen removal efficiencies decreased significantly when C/N was reduced (1.5 â†’ 0.75). When MLSS was doubled, the nitrogen removal efficiencies in the two parallel reactors increased from 75.3 %, 72.9 % to 86.9 %, 89.7 %, respectively, and sludge yields decreased obviously. Combining with in-situ test, it was speculated when MLSS increased, fermentation was enhanced, providing substrate for partial denitrification. Thauera, involved in partial denitrification, decreased obviously with reduced C/N, but increased from 9.93 % to 38.16 % when MLSS doubled, which could promote the PD/A process. Terrimonas and Ignavibacterium (fermentative bacteria) increased from 1.26 %, 5.22 % to 6.62 %, 6.30 %, respectively. These results proved that increasing MLSS under low C/N ratios promoted fermentation in PD/A system, facilitating efficient nitrogen removal and sludge reduction.

Molecular evidence of internal carbon-driven partial denitrification in a mainstream pilot A-B system coupled with side-stream EBPR treating municipal wastewater.

Achieving mainstream short-cut nitrogen removal via nitrite has become a carbon and energy efficient way, but still remains challenging for low-strength municipal wastewaters. This study integrated sidestream enhanced biological phosphorus removal system in a pilot-scale adsorption/bio-oxidation (A-B) process (named A-B-S2EBPR system) and nitrite accumulation was successfully achieved for treating the municipal wastewater. Nitrite could accumulate to 5.5 ± 0.3 mg N/L in the intermittently aerated tanks of B-stage with the nitrite accumulation ratio (NAR) of 79.1 ± 6.5 %. The final effluent concentration and removal efficiency of total inorganic nitrogen (TIN) were 4.6 ± 1.8 mg N/L and 84.9 ± 5.6 %, respectively. In-situ process performance of nitrogen conversions, routine batch nitrification/denitrification activity tests and functional gene abundance of nitrifiers collectively suggested that the nitrite accumulation was mainly caused by partial denitrification rather than out-selection of nitrite oxidizing bacteria (NOB). Moreover, the single-cell Raman spectroscopy analysis first demonstrated that there was a specific microbial population that could utilize polyhydroxyalkanoates (PHA) as the potential internal carbon source during the partial denitrification process. The integration of S2EBPR brings unique features to the conventional A-B process, such as extended anaerobic retention time, lower oxidation-reduction potential (ORP), much higher and complex volatile fatty acids (VFAs) etc., which can largely reshape the microbial communities. The dominant genera were Acinetobacter and Comamonadaceae, which accounted for (17.8 ± 15.5)% and (6.7 ± 3.4)%, respectively, while the relative abundance of conventional nitrifiers was less than 0.2%. This study provides insights into phylogenetic and phenotypic shifts of microbial communities when incorporating S2EBPR into the sustainable A-B process to achieve mainstream short-cut nitrogen removal.

Rapid start-up of mainstream partial denitrification /anammox and enhanced nitrogen removal through inoculation of precultured biofilm for treating low-strength municipal sewage.

This study investigated the rapid start-up of mainstream partial denitrification coupled with anammox (PD/A) and nitrogen removal performance by inoculating precultured PD/A biofilm. The results showed mainstream PD/A in the anaerobic-anoxic-aerobic (A2O) process was rapidly established within 30 days. Nitrogen removal efficiency (NRE) improved by 23.8 % contrasted to the traditional A2O process. The mass balance showed that anammox contribution to total nitrogen (TN) removal were maintained at 37.9 %∼55.7 %, and reducing hydraulic retention time (HRT) strengthened simultaneously denitrification and anammox activity. The microbial community showed that the dominant bacteria such as denitrifying bacteria (DNBs) and glycogen accumulating organisms (GAOs) both in biofilm and flocculent sludge (floc), integrating with anammox bacteria (AnAOB) in biofilm might lead to enhanced nitrogen removal. Overall, this study offered a fast start-up strategy of mainstream PD/A with enhanced nitrogen removal, which are valuable for upgradation and renovation of existed municipal wastewater treatment plants (WWTPs).

Sulfamethoxazole exposure shifts partial denitrification to complete denitrification: Reactor performance and microbial community.

This study elucidated the influence on a partial denitrification (PD) system under 0-1 mg/L sulfamethoxazole (SMX) stress in a sequencing batch reactor. The results showed that the nitrite accumulation rate (NAR) significantly (P ≤ 0.01) decreased from 68.68 ± 9.00% to 49.05 ± 11.68%, while the total nitrogen removal efficiency significantly (P ≤ 0.001) increased from 23.19 ± 4.42% to 31.36 ± 2.73% in presence of SMX. The results indicated that SMX exposure switched the PD process to complete denitrification through the deterioration of the nitrite accumulation and the promotion of further nitrite reduction. The SMX removal loading rate increased from 0.21 ± 0.04 to 5.03 ± 0.77 mg-SMX/(g-MLVSS·d) with the extended reactor operation under SMX stress. Low SMX concentration exposure increased extracellular polymeric substances (EPS) content from 107.69 ± 20.78 mg/g-MLVSS (0.05 mg-SMX/L) to 123.64 ± 9.66 mg/g-MLVSS (0.5 mg-SMX/L), while EPS secretion was inhibited under high SMX concentration exposure (i.e., 1 mg-SMX/L). Moreover, SMX exposure promoted the synthesis of aromatic protein-like compounds and changed the functional groups as revealed by EEM and FTIR analysis. Additionally, SMX exposure significantly shifted the microbial community structures at both phylum and genus levels. Particularly, the abundance of Thauera, i.e., functional bacteria related to PD, considerably decreased from 41.69% to 11.62% after SMX exposure, whereas the abundances of Denitratisoma and SM1A02 significantly rose under different SMX concentrations. These outcomes hinted that the addition of SMX resulted in the shifting of partial denitrification to complete denitrification.

Starvation resilience in anammox-based bioreactors: A stable nitrogen removal route on partial denitrification/anammox (PD/A).

This study investigates the performance, resilience and microbial community dynamics of two anaerobic processes, i.e. pure anammox (R1) and partial denitrification/anammox (PD/A) (R2), following a 30-day starvation period. The tolerance to starvation was assessed by comparing nitrogen removal efficiency and microbial activity across both reactors. Results show that the PD/A process recovery to pre-starvation performance levels within just one day, as compared to the pure anammox process. Notably, although the activity of anammox bacteria decreased in both processes during starvation, the decay rate in R1 was 69.59 % higher than in R2, potentially explaining the quicker recovery of R2. Furthermore, enhanced secretion of extracellular polymeric substance (EPS) during starvation served as a protective mechanism. The potential functions and genes in microorganisms, as well as the pathway of nitrogen cycling, were demonstrated through analyses using the KEGG database. This research reveals essential mechanistic insights and strategic guidance for the effective implementation of anammox-based biological nitrogen removal processes.

Novel Anoxic-Anaerobic-Oxic process successfully enriched anammox bacteria under actual municipal wastewater.

Anaerobic ammonia oxidation (Anammox) exhibits promise for wastewater treatment,but the enrichment of anammox bacteria (AnAOB) in municipal wastewater treatment plants is a significant challenge. This study constructed a novel Anoxic-Anaerobic-Oxic (AAnO) process with a pure biofilm anoxic zone fed with actual fluctuating municipal wastewater and operated for six months to enrich AnAOB at ambient temperature. High-throughput sequencing (HTS), qPCR, and fluorescence in situ hybridization showed that AnAOB were successfully enriched in the anoxic biofilms, reaching 1.56 % relative abundance on day 75 detected by HTS. During the period from day 130 to day 186, the anammox process contributed to 55.8 ± 19.2 % of the nitrogen removal in the anoxic zone. Phylogenetic analysis revealed this AnAOB species was closely related to Candidatus Brocadia fulgida. This study provides technical support for the application of anammox in mainstream wastewater.

Metagenomic insights into microbial metabolic mechanisms of a combined solid-phase denitrification and anammox process for nitrogen removal in mainstream wastewater treatment.

To overcome the significant challenges associated with nitrite supply and nitrate residues in mainstream anaerobic ammonium oxidation (anammox)-based processes, this study developed a combined solid-phase denitrification (SPD) and anammox process for low-strength nitrogen removal without the addition of nitrite. The SPD step was performed in a packed-bed reactor containing poly-3-hydroxybutyrate-co-3-hyroxyvelate (PHBV) prior to employing the anammox granular sludge reactor in the continuous-flow mode. The removal efficiency of total inorganic nitrogen reached 95.7 ± 1.2% under a nitrogen loading rate of 0.18 ± 0.01 kg N·m3·d-1, and it required 1.02 mol of nitrate to remove 1 mol of ammonium nitrogen. The PHBV particles not only served as biofilm carriers for the symbiosis of hydrolytic bacteria (HB) and denitrifying bacteria (DB), but also carbon sources that facilitated the coupling of partial denitrification and anammox in the granules. Metagenomic sequencing analysis indicated that Burkholderiales was the most abundant HB genus in SPD. The metabolic correlations between DB (Betaproteobacteria, Rhodocyclaceae, and Anaerolineae) and anammox bacteria (Candidatus Brocadiac and Kuenenia) in the granules were confirmed through microbial co-occurrence networks analysis and functional gene annotations. Additionally, the genes encoding nitrate reductase (Nap) and nitrite reductase (Nir) in DB primarily facilitated nitrate reduction, thereby supplying nitric oxide to anammox bacteria for subsequent nitrogen removal with hydrazine synthase (Hzs) and hydrazine dehydrogenase (Hdh). The findings provide insights into microbial metabolism within combined SPD and anammox processes, thus advancing the development of mainstream anammox-based processes in engineering applications.

Enhance PD/A biofilm formation via a novel biochar/tourmaline modified-biocarriers to treat low-strength contaminated surface water: Initial adhesion and high-substrate microenvironment.

In this work, a novel polyurethane carrier modified with biochar and tourmaline/zeolite powder at ratio of 1:1 and 1:2 was developed to promote the formation of biofilms and the synergy of overall bacterial activity for Partial Denitrification/Anammox to treat low-nitrogen contaminated surface water. Based on the batch experiment, the modified biocarrier, BTP2 (biochar: tourmaline = 2: 1), exhibited the highest total nitrogen removal efficiency (83.63%) under influent total nitrogen of 15 mg/L and COD/NO3- of 3. The dense biofilm was formed in inner side of biocarrier owing to the increased surface roughness and various functional groups suggested by scanning electron microscopy and Fourier-transform infrared analysis. The EPS content increased from 200.15 to 220.26 mg/g VSS in BTP2 system. Besides, the rapid NH4+ capture and organics release of the modified carrier fueled the growth of anammox and denitrification bacteria, with the activity of 2.13 ± 0.52 mg N/gVSS/h and 6.70 ± 0.52 mg N/gVSS/h (BTP2). High-throughput sequencing unraveled the increased abundances of Candidatus_Competibacter (0.82%), Thauera (0.60%) and Candidatus_Brocadia (0.55%) which was responsible for the synergy of incomplete reduction of NO3- to NO2- and NH4+ oxidation. Overall, this study provided a valid and simple-control guide for biofilm formation towards rapid enrichment and great collaboration of Anammox and denitrification bacteria.

Metagenomic unveils the promotion of mainstream PD-anammox process at lower nZVI concentration and inhibition at higher dosage.

The partial-denitrification-anammox (PdNA) process exhibits great potential in enabling the simultaneous removal of NO3--N and NH4+-N. This study delved into the impact of exogenous nano zero-valent iron (nZVI) on the PdNA process. Adding 10 mg L-1 of nZVI increased nitrogen removal efficiency up to 83.12 % and maintained higher relative abundances of certain beneficial bacteria. The maximum relative abundance of Candidatus Brocadia (1.6 %), Candidatus Kuenenia (1.5 %), Ignavibacterium (1.3 %), and Azospira (1.2 %) was observed at 10 mg L-1 of nZVI. However, the greatest relative abundance of Thauera (1.3 %) was recorded under 50 mg L-1. Moreover, applying nZVI selectively enhanced the abundance of NO3--N reductase genes. So, keeping the nZVI concentration at 10 mg L-1 or below is advisable to ensure a stable PdNA process in mainstream conditions. Considering nitrogen removal efficiency, using nZVI in the PD-anammox process could be more cost-effective in enhancing its adoption in industrial and mainstream settings.