Nitrogen-cycling processes under long-term compound heavy metal(loids) pressure around a gold mine: Stimulation of nitrite reduction.

Mining and tailings deposition can cause serious heavy metal(loids) pollution to the surrounding soil environment. Soil microorganisms adapt their metabolism to such conditions, driving alterations in soil function. This study aims to elucidate the response patterns of nitrogen-cycling microorganisms under long-term heavy metal(loids) exposure. The results showed that the diversity and abundance of nitrogen-cycling microorganisms showed negative feedback to heavy metal(loids) concentrations. Denitrifying microorganisms were shown to be the dominant microorganisms with over 60% of relative abundance and a complex community structure including 27 phyla. Further, the key bacterial species in the denitrification process were calculated using a random forest model, where the top three key species (Pseudomonas stutzei, Sphingobium japonicum and Leifsonia rubra) were found to play a prominent role in nitrite reduction. Functional gene analysis and qPCR revealed that nirK, which is involved in nitrite reduction, significantly accumulated in the most metal-rich soil with the increase of absolute abundance of 63.86%. The experimental results confirmed that the activity of nitrite reductase (Nir) encoded by nirK in the soil was increased at high concentrations of heavy metal(loids). Partial least squares-path model identified three potential modes of nitrite reduction processes being stimulated by heavy metal(loids), the most prominent of which contributed to enhanced nirK abundance and soil Nir activity through positive stimulation of key species. The results provide new insights and preliminary evidence on the stimulation of nitrite reduction processes by heavy metal(loids).

Disentangling microbial coupled fillers mechanisms for the permeable layer optimization process in multi-soil-layering systems.

The multi-soil-layering (MSL) systems is an emerging solution for environmentally-friendly and cost-effective treatment of decentralized rural domestic wastewater. However, the role of the seemingly simple permeable layer has been overlooked, potentially holding the breakthroughs or directions to addressing suboptimal nitrogen removal performance in MSL systems. In this paper, the mechanism among diverse substrates (zeolite, green zeolite and biological ceramsite) coupled microorganisms in different systems (activated bacterial powder and activated sludge) for rural domestic wastewater purification was investigated. The removal efficiencies performed by zeolite coupled with microorganisms within 3 days were 93.8% for COD, 97.1% for TP, and 98.8% for NH4+-N. Notably, activated sludge showed better nitrification and comprehensive performance than specialized nitrifying bacteria powder. Zeolite attained an impressive 89.4% NH4+-N desorption efficiency, with a substantive fraction of NH4+-N manifesting as exchanged ammonium. High-throughput 16S rRNA gene sequencing revealed that aerobic and parthenogenetic anaerobic bacteria dominated the reactor, with anaerobic bacteria conspicuously absent. And the heterotrophic nitrification-aerobic denitrification (HN-AD) process was significant, with the presence of denitrifying phosphorus-accumulating organisms (DPAOs) for simultaneous nitrogen and phosphorus removal. This study not only raises awareness about the importance of the permeable layer and enhances comprehension of the HN-AD mechanism in MSL systems, but also provides valuable insights for optimizing MSL system construction, operation, and rural domestic wastewater treatment.

Transcriptome analysis of the degradation process of organic nitrogen by two heterotrophic nitrifying and aerobic denitrifying bacteria.

The heterotrophic nitrification aerobic denitrification bacteria (HNDS) can perform nitrification and denitrification at the same time. Two HNDS strains, Achromobacter sp. HNDS-1 and Enterobacter sp. HNDS-6 which exhibited an amazing ability to solution nitrogen (N) removal have been successfully isolated from paddy soil in our lab. When peptone or ammonium sulfate as sole N source, no significant difference in gene expression related to nitrification and denitrification of the strains was found according to the transcriptome analysis. The expression of phosphomethylpyrimidine synthase (thiC), ABC transporter substrate-binding protein, branched-chain amino acid ABC transporter substrate-binding protein, and RNA polymerase (rpoE) in HNDS-1 were significantly upregulated when used peptone as N source, while the expression of exopolysaccharide production protein (yjbE), RNA polymerase (rpoC), glutamate synthase (gltD) and ABC-type branched-chain amino acid transport systems in HNDS-6 were significantly upregulated. This indicated that these two strains are capable of using organic N and converting it into NH4+-N, then utilizing NH4+-N to synthesize amino acids and proteins for their own growth, and strain HNDS-6 can also remove NH4+-N through nitrification and denitrification.

A novel simultaneous short-course nitrification, denitrification and fermentation process: bio-enhanced phenol degradation and denitrification in a single reactor.

The feasibility of a simultaneous nitrification, denitrification and fermentation process (SNDF) under electric stirrer agitation conditions was verified in a single reactor. Enhanced activated sludge for phenol degradation and denitrification in pharmaceutical phenol-containing wastewater under low dissolved oxygen conditions, additional inoculation with Comamonas sp. BGH and optimisation of co-metabolites were investigated. At a hydraulic residence time (HRT) of 28 h, 15 mg/L of substrate as strain BGH co-metabolised substrate degraded 650 ± 50 mg/L phenol almost completely and was accompanied by an incremental increase in the quantity of strain BGH. Strain BGH showed enhanced phenol degradation. Under trisodium citrate co-metabolism, strain BGH combined with activated sludge treated phenol wastewater and degraded NO2--N from 50 ± 5 to 0 mg/L in only 7 h. The removal efficiency of this group for phenol, chemical oxygen demand (COD) and TN was 99.67%, 90.25% and 98.71%, respectively, at an HRT of 32 h. The bioaugmentation effect not only promotes the degradation of pollutants, but also increases the abundance of dominant bacteria in activated sludge. Illumina MiSeq sequencing research showed that strain BGH promoted the growth of dominant genera (Acidaminobacter, Raineyella, Pseudarcobacter) and increased their relative abundance in the activated sludge system. These genera are resistant to toxicity and organic matter degradation. This paper provides some reference for the activated sludge to degrade high phenol pharmaceutical wastewater under the action of biological enhancement.

Improving denitrification estimation by joint inclusion of suspended particles and chlorophyll a in aquaculture ponds.

The denitrification process in aquaculture systems plays a crucial role in nitrogen (N) cycle and N budget estimation. Reliable models are needed to rapidly quantify denitrification rates and assess nitrogen losses. This study conducted a comparative analysis of denitrification rates in fish, crabs, and natural ponds in the Taihu region from March to November 2021, covering a complete aquaculture cycle. The results revealed that aquaculture ponds exhibited higher denitrification rates compared to natural ponds. Key variables influencing denitrification rates were Nitrate nitrogen (NO3--N), Suspended particles (SPS), and chlorophyll a (Chla). There was a significant positive correlation between SPS concentration and denitrification rates. However, we observed that the denitrification rate initially rose with increasing Chla concentration, followed by a subsequent decline. To develop parsimonious models for denitrification rates in aquaculture ponds, we constructed five different statistical models to predict denitrification rates, among which the improved quadratic polynomial regression model (SQPR) that incorporated the three key parameters accounted for 80.7% of the variability in denitrification rates. Additionally, a remote sensing model (RSM) utilizing SPS and Chla explained 43.8% of the variability. The RSM model is particularly valuable for rapid estimation in large regions where remote sensing data are the only available source. This study enhances the understanding of denitrification processes in aquaculture systems, introduces a new model for estimating denitrification in aquaculture ponds, and offers valuable insights for environmental management.

Pyric herbivory decreases soil denitrification despite increased nitrate availability in a temperate grassland.

Pyric herbivory, the combination of controlled burning and targeted grazing, is an effective strategy for restoring abandoned, shrub-encroached rangelands to open ecosystems. This practice may impact soil nitrogen pools by altering soil nitrification and denitrification rates, and may lead to an increase of nitrogen losses through nitrate leaching and N-gas emissions. This research, located in the south-western Pyrenees, investigated the effects of pyric herbivory on soil nitrification and denitrification potentials and mineral nitrogen content in a gorse-encroached temperate rangeland six months after the burning was implemented. The study included three treatments: high-severity burning plus grazing, low-severity burning plus grazing, and unburned and ungrazed areas (control). We measured soil nitrification and denitrification potentials (net and gross), the limitation of denitrifiers by nitrogen or organic carbon, and the abundance of nitrite- and nitrous oxide-reducing bacteria. Additional soil and vegetation data complemented these measurements. Results showed that pyric herbivory did not significantly affect nitrification potential, which was low and highly variable. However, it decreased gross denitrification potential and nitrous oxide reduction to dinitrogen in high-severely burned areas compared to the control. Denitrification rates directly correlated with microbial biomass nitrogen, soil organic carbon, soil water content and abundance of nirS-harbouring bacteria. Contrary to the expected, soil nitrate availability did not directly influence denitrification despite being highest in burned areas. Overall, the study suggests that pyric herbivory does not significantly affect mid-term nitrification rates in temperate open ecosystems, but may decrease denitrification rates in intensely burned areas. These findings highlight the importance of assessing the potential impacts of land management practices, such as pyric herbivory, on soil nutrient cycling and ecosystem functioning.

Development of an integrated fixed-film activated sludge (IFAS) reactor treating landfill leachate for the biological nitrogen removal through nitritation-denitritation.

The current work investigated the performance of an Integrated Fixed-Film Activated Sludge Sequencing Batch Reactor (IFAS-SBR) for Biological Nitrogen Removal (BNR) from mature landfill leachate through the nitritation-denitritation process. During the experimental period two IFAS-SBR configurations were examined using two different biocarrier types with the same filling ratio (50%). The dissolved oxygen (DO) concentration ranged between 2 and 3 mg/L and 4-6 mg/L in the first (baseline-IFAS) and the second (S8-IFAS) setup, respectively. Baseline-IFAS operated for 542 days and demonstrated a high and stable BNR performance maintaining a removal efficiency above 90% under a Nitrogen Loading Rate (NLR) up to 0.45 kg N/m3-d, while S8-IFAS, which operated for 230 days, was characterized by a limited and unstable BNR performance being unable to operate sufficiently under an NLR higher than 0.20 kg N/m3-d. It also experienced a severe inhibition period, when the BNR process was fully deteriorated. Moreover, S8-IFAS suffered from extensive biocarrier stagnant zones and a particularly poor sludge settleability. The attached biomass cultivated in both IFAS configurations had a negligible content of nitrifying bacteria, probably attributed to the insufficient DO diffusion through the biofilm, caused by the low DO concentration in the liquid in the baseline case and the extensive stagnant zones in the S8-IFAS case. As a result of the high biocarrier filling ratio, the S8-IFAS was unstable and low. This was probably attributed to the mass transfer limitations caused by the biocarrier stagnant zones, which hinder substrate and oxygen diffusion, thus reducing the biomass activity and increasing its vulnerability to inhibitory and toxic factors. Hence, the biocarrier filling fraction is a crucial parameter for the efficient operation of the IFAS-SBR and should be carefully selected taking into consideration both the media type and the overall reactor configuration.

Enhanced nitrogen removal of the anaerobic ammonia oxidation process by coupling with an efficient nitrate reducing bacterium (Bacillus velezensis M3-1).

Bacillus velezensis M3-1 strain isolated from the sediment of Myriophyllum aquatium constructed wetlands was found to efficiently convert NO3--N to NO2--N, and the requirements for carbon source addition were not very rigorous. This work demonstrates, for the first time, the feasibility of using the synergy of anammox and Bacillus velezensis M3-1 microorganisms for nitrogen removal. In this study, the possibility of M3-1 that converted NO3--N produced by anammox to NO2--N was verified in an anaerobic reactor. The NO3--N reduction ability of M3-1 and denitrifying bacteria in coupling system was investigated under different C/N conditions, and it was found that M3-1 used carbon sources preferentially over denitrifying bacteria. By adjusting the ratio of NH4+-N to NO2--N, it was found that the NO2--N converted from NO3--N by M3-1 participated in the original anammox.The nitrogen removal efficacy (NRE) of the coupled system was increased by 12.1%, compared to the control group anammox system at C/N = 2:1. Functional gene indicated that it might be a nitrate reducing bacterium.This study shows that the nitrate reduction rate achieved by the Bacillus velezensis M3-1 can be high enough for removing nitrate produced by anammox process, which would enable improve nitrogen removal from wastewater.

Synergistic mechanisms of denitrification in FeS2-based constructed wetlands: Effects of organic carbon availability under day-night alterations.

In constructed wetlands (CWs), carbon source availability profoundly affected microbial metabolic activities engaged in both iron cycle and nitrogen metabolism. However, research gaps existed in understanding the biotransformation of nitrogen and iron in response to fluctuations in organic carbon content under day-night alterations. Results demonstrated increased removal efficiency of NO3--N (95.7 %) and NH4+-N (75.70 %) under light conditions, attributed to increased total organic carbon (TOC). This enhancement promoted the relative abundance of bacteria involved in nitrogen and iron processes, establishing a more stable microbial network. Elevated TOC content also upregulated genes for iron metabolism and glycolysis, facilitating denitrification. Spearman correlation analysis supported the synergistic mechanisms between FeS2-based autotrophic denitrification and TOC-mediated heterotrophic denitrification under light conditions. The significant impact of carbon sources on microbial activities underscores the critical role of organic carbon availability in enhancing nitrogen removal efficiency, providing valuable insights for optimizing FeS2-based CWs design and operation strategies.

Coordination of elemental sulfur and organic carbon source stimulates simultaneous nitrification and denitrification toward low C/N ratio wastewater.

The feasibility of inducing simultaneous nitrification and denitrification (SND) by S0 for low carbon to nitrogen (C/N) ratio wastewater remediation was investigated. Compared with S0 and/or organics absent systems (-3.4 %∼5.0 %), the higher nitrogen removal performance (18.2 %∼59.8 %) was achieved with C/N ratios and S0 dosages increasing when S0 and organics added simultaneously. The synergistic effect of S0 and organics stimulated extracellular polymeric substances secretion and weakened intermolecular binding force of S0, facilitating S0 bio-utilization and reducing the external organics requirement. It also promoted microbial metabolism (0.16 âˆ¼ 0.24 µg O2/(g VSS·h)) and ammonia assimilation (5.9 %∼20.5 %), thereby enhancing the capture of organics and providing more electron donors for SND. Furthermore, aerobic denitrifiers (15.91 %∼27.45 %) and aerobic denitrifying (napA and nirS) and ammonia assimilating genes were accumulated by this synergistic effect. This study revealed the mechanism of SND induced by coordination of S0 and organics and provided an innovative strategy for triggering efficient and stable SND.

Nitrate and nitrite utilization during denitrifying phosphorus removal: Electron acceptor preference and feasible process combinations.

Denitrifying phosphorus removal (DPR), which is dominated by denitrifying polyphosphate-accumulating organisms (DPAOs), is a promising process for nitrogen and phosphorus removal. Denitrifying glycogen-accumulating organisms (DGAOs) and DPAOs typically coexist in the DPR sludge, complicating the study of DPAOs' denitrification capacity. In this study, two reactors were fed with nitrate and nitrite during the anoxic phase to cultivate nitrate-DPR and nitrite-DPR sludge. Both reactors yielded high and low DGAO abundance sludges, enabling the evaluation of the denitrification capacity of DPAOs. For the nitrate-DPR sludge, the nitrite reduction rate was 1.63 times higher than the nitrate reduction rate when DPAOs were the primary denitrifiers. For the nitrite-DPR sludge, the reduction rate of nitrite was more than three times that of nitrate, irrespective of DGAO abundance. These findings indicated that DPAOs preferred nitrite to nitrate and were well suited to reduce nitrite rather than reduce nitrate to supply nitrite.

Mitigating nitrous oxide emissions from low carbon to nitrogen ratio wastewater treatment: Utilizing sugarcane bagasse fermentation liquid for constructed wetlands.

Sugarcane bagasse was recycled to produce fermentation liquid (FL) as a supplementary carbon source that was added to constructed wetlands (CWs) for regulating influent carbon to nitrogen ratio (C/N), and then being applied to investigate nitrogen transformations and greenhouse gas emissions. Results showed that this FL achieved faster NO3--N removal and lower N2O fluxes than sucrose did, and the lowest N2O flux (67.6 µg m-2h-1) was achieved when FL was added to CWs in a C/N of 3. In contrast, CH4 emissions were higher by the FL addition than by the sucrose addition, although the fluxes under both additions were in a lower range of 0.06-0.17 mg m-2h-1. The utilization of FL also induced significant variations in microbial communities and increased the abundance of denitrification genes. Results showed the application of FL from sugarcane bagasse can be an effective strategy for improving nitrogen removal and mitigating N2O emissions in CWs.

Optimization of a Novel Engineered Ecosystem Integrating Carbon, Nitrogen, Phosphorus, and Sulfur Biotransformation for Saline Wastewater Treatment Using an Interpretable Machine Learning Approach.

The denitrifying sulfur (S) conversion-associated enhanced biological phosphorus removal (DS-EBPR) process for treating saline wastewater is characterized by its unique microbial ecology that integrates carbon (C), nitrogen (N), phosphorus (P), and S biotransformation. However, operational instability arises due to the numerous parameters and intricates bacterial interactions. This study introduces a two-stage interpretable machine learning approach to predict S conversion-driven P removal efficiency and optimize DS-EBPR process. Stage one utilized the XGBoost regression model, achieving an R2 value of 0.948 for predicting sulfate reduction (SR) intensity from anaerobic parameters with feature engineering. Stage two involved the CatBoost classification and regression model integrating anoxic parameters with the predicted SR values for predicting P removal, reaching an accuracy of 94% and an R2 value of 0.93, respectively. This study identified key environmental factors, including SR intensity (20-45 mg S/L), influent P concentration (<9.0 mg P/L), mixed liquor volatile suspended solids (MLVSS)/mixed liquor suspended solids (MLSS) ratio (0.55-0.72), influent C/S ratio (0.5-1.0), anoxic reaction time (5-6 h), and MLSS concentration (>6.50 g/L). A user-friendly graphic interface was developed to facilitate easier optimization and control. This approach streamlines the determination of optimal conditions for enhancing P removal in the DS-EBPR process.

Evidence of nitrogen inputs affecting soil nitrogen purification by mediating root exudates of salt marsh plants.

Salt marsh has an important 'purification' role in coastal ecosystems by removing excess nitrogen that could otherwise harm aquatic life and reduce water quality. Recent studies suggest that salt marsh root exudates might be the 'control centre' for nitrogen transformation, but empirical evidence is lacking. Here we sought to estimate the direction and magnitude of nitrogen purification by salt marsh root exudates and gain a mechanistic understanding of the biogeochemical transformation pathway(s). To achieve this, we used a laboratory incubation to quantify both the root exudates and soil nitrogen purification rates, in addition to the enzyme activities and functional genes under Phragmites australis populations with different nitrogen forms addition (NO3-, NH4+ and urea). We found that NO3- and urea addition significantly stimulate P. australis root exudation of total acids, amino acids, total sugars and total organic carbon, while NH4+ addition only significantly increased total acids, amino acids and total phenol exudation. High total sugars, amino acids and total organic carbon concentrations enlarged nitrogen purification potential by stimulating the nitrogen purifying bacterial activities (including enzyme activities and related genes expression). Potential denitrification rates were not significantly elevated under NH4+ addition in comparison to NO3- and urea addition, which should be ascribed to total phenol self-toxicity and selective inhibition. Further, urea addition stimulated urease and protease activities with providing more NH4+ and NO2- substrates for elevated anaerobic ammonium oxidation rates among the nitrogen addition treatments. Overall, this study revealed that exogenous nitrogen could increase the nitrogen purification-associated bacterial activity through accelerating the root exudate release, which could stimulate the activity of nitrogen transformation, and then improve the nitrogen removal capacity in salt marsh.

Ammonium nitrogen and phosphorus removal by bacterial-algal symbiotic dynamic sponge bioremediation system in micropolluted water: Operational mechanism and transformation pathways.

Construct a bacteria-algae symbiotic dynamic sponge bioremediation system to simultaneously remove multiple pollutants under micro-pollution conditions. The average removal efficiencies of NH4+-N, PO43--P, total nitrogen (TN), and Ca2+ were 98.35, 78.74, 95.64, and 84.92 %, respectively. Comparative studies with Auxenochlorella sp. sponge and bacterial sponge bioremediation system confirmed that NH4+-N and TN were mainly removed by bacterial heterotrophic nitrification - aerobic denitrification (HN-AD). PO43--P was removed by algal assimilation and the generation of Ca3(PO4)2 and Ca5(PO4)3OH, and Ca2+ was removed by algal electron transfer formation of precipitates and microbially induced calcium precipitation (MICP) by bacteria. Algae provided an aerobic environment for the bacterial HN-AD process through photosynthesis, while respiration produced CO2 and adsorbed Ca2+ to promote the formation of calcium precipitates. Immobilization of Ca2+ with microalgae via bacterial MICP helped to lift microalgal photoinhibition. The bioremediation system provides theoretical support for research on micropolluted water treatment while increasing phosphorus recovery pathways.

Elucidating biogeochemical characterization of nitrogen in the vadose zone integrating geochemistry, microorganism, and numerical simulation.

A thorough comprehension of nitrogen biogeochemical processes in the vadose zone is crucial for the effective prevention and remediation of soil-groundwater system contamination. Despite the growing research on this subject, the full scope of nitrogen biogeochemical characterization in different geological environments remains poorly understood. This study addresses this knowledge gap by integrating geochemical, microbiological and numerical simulation approaches to gain a deeper insight into nitrogen biogeochemistry in agriculture. Our findings indicate the biogeochemical behavior of nitrogen in the vadose zone is mediated by microorganisms, driven by hydraulics, influenced by geological conditions and environmental factors. Along the groundwater flow, NH4+-N was found to be heavily accumulated in the topsoil of 0-40 cm, while NO3--N was transported and driven by hydrodynamics from both vertical and horizontal directions. Microbial diversity, species composition and functional microorganisms were significantly influenced by soil depth, rather than geomorphological types. Oxidation-reduction potential (ORP), total organic carbon (TOC), soil moisture (MOI), bicarbonate (HCO3-), and ferrous (Fe2+) were identified as the principal environmental factors that regulate nitrogen metabolism and the dominant biochemical processes, encompassing nitrogen fixation, nitrification, and denitrification. Driven by hydrodynamics, NH4+-N, NO2--N and NO3--N tend to form distinct biochemical reaction zones in the vertical vadose zone. These areas are dynamic and subject to geomorphologies. It should be noted that NO3--N can migrate towards groundwater from the clayey sand in the Alluvial Plain, which presents a potential risk of groundwater contamination. The fissure structure of loess may serve as the major transport pathway for groundwater nitrogen contamination in the Loess Tableland. This finding highlights the importance of integrating microbiology, geochemistry and hydraulics to elucidate the biogeochemical processes of nitrogen in the vadose zone with a dynamic mindset.

Effective remediation of agricultural drainage at three influent strengths by bioaugmented constructed wetlands filled with mixture of iron­carbon and organic solid substrates: Performance and mechanisms.

Agricultural drainage containing a large quantity of nutrients can cause quality deterioration and algal blooming of receiving water bodies, thus needs to be effectively remediated. In this study, iron­carbon (FeC) composite-filled constructed wetlands (Fe-C-CWs) were employed to treat farmland drainage at three pollution levels, and organic solid substrates (walnut shells) and phosphate-accumulating denitrifying bacteria (Pseudomonas sp. DWP1) were supplemented to enhance the treatment performance. The results showed that the Fe-C-CWs exhibited notably superior removal efficiency for total nitrogen (TN, 52.0-58.2 %), total phosphorus (TP, 67.8-70.2 %) and chemical oxygen demand (COD, 56.7-70.4 %) than the control systems filled solely with gravel (28.5-32.5 % for TN, 33.2-40.5 % for TP and 30.2-55.0 % for COD) at all influent strengths, through driving autotrophic denitrification, Fe-based dephosphorization, and organic degradation processes. The addition of organic substrates and functional bacteria markedly enhanced pollutant removal in the Fe-C-CWs. Furthermore, use of FeC and organic substrates and denitrifier inoculation decreased CO2 and CH4 emissions from the CWs, and reduced global warming potential of the CWs at low influent strength. Pollutant removal efficiencies in the CWs were only marginally impacted by the increasing influent loads except for NO3--N, and pollutant removal mass was largely increased with the increase of influent strengths. The microbial community in the FeC composite-filled CWs exhibited distinct distribution patterns compared to the gravel-filled CWs regardless of the influent strengths, with obviously higher proportions of dominant genera Trichococcus, Geobacter and Ferritrophicum. Keystone taxa associated with pollutant removal in the Fe-C-filled CWs were identified to be Pseudomonas, Geobacter, Ferritrophicum, Denitratisoma and Sediminibacterium. The developed augmented Fe-C-filled CWs show great promises for remediating agricultural drainage with varied pollutant loads.

[Start-up and Recovery Performance of ANAMMOX Reactors Under Long-term Starvation].

The application of ANAMMOX technology is constrained by sluggish growth and difficulty in enriching ANAMMOX bacteria. Long-term starvation of functioning bacteria due to limited substrate supply makes the steady operation of ANAMMOX reactors more difficult. Re-examining the start-up and recovery performance of the ANAMMOX reactor and identifying its resistance mechanism are important from the standpoint of long-term starvation. By inoculating nitrifying and denitrifying sludge under various operating circumstances, the ANAMMOX reactors were successfully started. Under various start-up procedures, the tolerance mechanism and recovery performance were examined. The outcomes demonstrated that the denitrifying sludge-inoculated reactor operated steadily with a high substrate concentration and low flow rate. After 85 days of operation, the removal efficiencies of NH4+-N, NO2--N, and total nitrogen reached 98.7%, 99.3%, and 89.3%, respectively. After 144 days of starvation and 30 days of recovery, the better nitrogen removal performance was achieved at a low substrate concentration and high flow rate, and the removal efficiencies were 99.8% （NH4+-N）, 99.8% （NO2--N）, and 93.6% （total nitrogen）. During the starvation, extracellular polymeric substances wrapped the ANAMMOX bacteria and kept them intact to resist long-term starvation stress. The expression of nirS, hzsA, and hdh genes ensured the synthesis of nitrite/nitric oxide oxidoreductase, hydrazine synthase, and hydrazine dehydrogenase to maintain ANAMMOX activity. There was no significant difference in the relative abundance of ANAMMOX bacteria before and after starvation recovery. Candidatus Kuenenia had better anti-hunger ability, and the relative abundance increased by more than 86% after 30 days of recovery, confirming its tolerance to long-term starvation.

Maximizing pollutant removal and greenhouse gas emission reduction in vertical flow constructed wetlands: an orthogonal experimental approach.

Owing to the impact of the effluent C/N from the secondary structures of urban domestic wastewater treatment plants, the denitrification efficiency in constructed wetlands (CWs) is not satisfactory, limiting their widespread application in the deep treatment of urban domestic wastewater. To address this issue, we constructed enhanced CWs and conducted orthogonal experiments to investigate the effects of different factors (C/N, fillers, and plants) on the removal of conventional pollutants and the reduction of greenhouse gas (GHG) emission. The experimental results indicated that a C/N of 8, manganese sand, and calamus achieved the best denitrification efficiencies with removal efficiencies of 85.7%, 95.9%, and 88.6% for TN, NH4+-N, and COD, respectively. In terms of GHG emission reduction, this combination resulted in the lowest global warming potential (176.8 mg/m2·day), with N2O and CH4 emissions of 0.53 and 1.25 mg/m2·day, respectively. Characterization of the fillers revealed the formation of small spherical clusters of phosphates on the surfaces of manganese sand and pyrite and iron oxide crystals on the surface of pyrite. Additionally, the surface Mn (II) content of the manganese sand increased by 8.8%, and the Fe (III)/Fe (II) and SO42-/S2- on pyrite increased by 2.05 and 0.26, respectively, compared to pre-experiment levels. High-throughput sequencing indicated the presence of abundant autotrophic denitrifying bacteria (Sulfuriferula, Sulfuritalea, and Thiobacillus) in the CWs, which explains denitrification performance of the enhanced CWs. This study aimed to explore the mechanism of efficient denitrification and GHG emission reduction in the enhanced CWs, providing theoretical guidance for the deep treatment of urban domestic wastewater.

Gemmobacter denitrificans sp. nov., a denitrifying bacterium, isolated from pond water for Litopenaeus vannamei.

A novel Gram-stain-negative strain, designated JM10B15T, was isolated from pond water for Litopenaeus vannamei collected from Jiangmen City, Guangdong province, south PR China. Cells of the strain were aerobic, rod-shaped, and motile by lateral flagella. JM10B15T could grow at 15-40 °C, pH 6.0-9.5, and in 0-3.0 % NaCl, with optimal growth at 25-35 °C, pH 7.5-8.5, and in 0 % NaCl, respectively. Furthermore, this strain grew well on Reasoner's 2A agar but not on nutrient broth agar or Luria-Bertani agar. JM10B15T was a denitrifying bacterium capable of removing nitrites and nitrates, and three key functional genes, nasA, nirS, and nosZ, were identified in its genome. The results of phylogenetic analyses based on the 16S rRNA gene and genome sequences indicated that JM10B15T belonged to the genus Gemmobacter. JM10B15T showed the highest 16S rRNA sequence similarity to Gemmobacter lutimaris YJ-T1-11T (98.8 %), followed by Gemmobacter aquatilis IFAM 1031T (98.6 %) and Gemmobacter serpentinus HB-1T (98.1 %). The average nucleotide identity and digital DNA-DNA hybridization values between JM10B15T and the other type strains of genus Gemmobacter were 78.1-82.1 % and 18.4-22.1 %, respectively. The major fatty acids of strain JM10B15T were summed feature 8 (C18 : 1 ω6c and/or C18 : 1 ω7c) and C18 : 1 ω7c 11-methyl. In addition, the major respiratory quinone of this novel strain was Q-10, and the predominant polar lipids were phosphatidylglycerol, phosphatidylethanolamine, four unidentified phospholipids, three unidentified lipids, and an unidentified aminophospholipid. Results of analyses of the phylogenetic, genomic, physiological, and biochemical characteristics indicated that JM10B15T represents a novel species of the genus Gemmobacter, for which the name Gemmobacter denitrificans sp. nov. is proposed. The type strain is JM10B15T (=GDMCC 1.4148T=KCTC 8140T).