Heterotrophic Denitrifiers Research Articles

Effect of magnetic powder (Fe3O4) on heterotrophic-sulfur autotrophic denitrification efficiency and electron transport system activity for marine recirculating aquacultural wastewater treatment

As an efficient nitrogen removal process, heterotrophic-sulfur autotrophic denitrification (HSAD) has attracted extensive attention in wastewater treatment. However, the effects of magnetic powder (Fe3O4) on the electron transport activity in HSAD process remain unclear. Therefore, in this study, a heterotrophic-sulfur autotrophic denitrification system was established to remove nitrogen from marine recirculating aquacultural wastewater for evaluating the effects of Fe3O4. At the optimal Fe3O4 concentration of 50 mg/L, the nitrogen removal efficiency reached 100% with lower sulfate accumulation, and the start-up time was shortened. The assays of denitrifying enzymes and electron transport system activity showed that Fe3O4 improved the activities of nitrate and nitrite reductases, and increased the efficiency of electron transport. Microbial community analysis revealed that Fe3O4 enriched heterotrophic denitrifier Thauera and sulfur autotrophic denitrifier Canditatus Thiobios, and thus enhanced denitrification efficiencies. This study demonstrated that Fe3O4 is an efficient denitrification accelerator in HSAD for treating marine recirculating aquacultural wastewater.

Fimbriimonadales performed dissimilatory nitrate reduction to ammonium (DNRA) in an anammox reactor

Bacteria belonging to the order Fimbriimonadales are frequently detected in anammox reactors. However, the principal functions of these bacteria and their potential contribution to nitrogen removal remain unclear. In this study, we aimed to systematically validate the roles of Fimbriimonadales in an anammox reactor fed with synthetic wastewater. High-throughput 16S rRNA gene sequencing analysis revealed that heterotrophic denitrifying bacteria (HDB) were the most abundant bacterial group at the initial stage of reactor operation and the abundance of Fimbriimonadales members gradually increased to reach 38.8 % (day 196). At the end of reactor operation, Fimbriimonadales decreased to 0.9 % with an increase in anammox bacteria. Correlation analysis demonstrated nitrate competition between Fimbriimonadales and HDB during reactor operation. Based on the phylogenetic analysis, the Fimbriimonadales sequences acquired from the reactor were clustered into three distinct groups, which included the sequences obtained from other anammox reactors. Metagenome-assembled genome analysis of Fimbriimonadales allowed the identification of the genes narGHI and nrfAH, responsible for dissimilatory nitrate reduction to ammonium (DNRA), and nrt and nasA, responsible for nitrate and nitrite transport. In a simulation based on mass balance equations and quantified bacterial groups, the total nitrogen concentrations in the effluent were best predicted when Fimbriimonadales was assumed to perform DNRA (R2 = 0.70 and RMSE = 18.9). Moreover, mass balance analysis demonstrated the potential contribution of DNRA in enriching anammox bacteria and promoting nitrogen removal. These results prove that Fimbriimonadales compete with HDB for nitrate utilization through DNRA in the anammox reactor under non-exogenous carbon supply conditions. Overall, our findings suggest that the DNRA pathway in Fimbriimonadales could enhance anammox enrichment and nitrogen removal by providing substrates (nitrite and/or ammonium) for anammox bacteria.

Comammox rather than AOB dominated the efficient autotrophic nitrification-denitrification process in an extremely oxygen-limited environment

The discovery of complete ammonia oxidizer (comammox) has challenged the traditional understanding of the two-step nitrification process. However, their functions in the oxygen-limited autotrophic nitrification-denitrification (OLAND) process remain unclear. In this study, OLAND was achieved using comammox-dominated nitrifying bacteria in an extremely oxygen-limited environment with a dissolved oxygen concentrations of 0.05 mg/L. The ammonia removal efficiency exceeded 97 %, and the total nitrogen removal efficiency reached 71 % when sodium bicarbonate was used as the carbon source. The pseudo-first- and second-order models were found to best fit the ammonia removal processes under low and high loads, respectively, suggesting distinct ammonia removal pathways. Full-length 16S rRNA gene sequencing and metagenomic results revealed that comammox-dominated under different oxygen levels, in conjunction with anammox and heterotrophic denitrifiers. The abundance of enzymes involved in energy metabolism indicates the coexistence of anammox and autotrophic nitrification-heterotrophic denitrification pathways. The binning results showed that comammox bacteria engaged in horizontal gene transfer with nitrifiers, anammox bacteria, and denitrifiers to adapt to an obligate environments. Therefore, this study demonstrated that comammox, anammox, and heterotrophic denitrifiers play important roles in the OLAND process and provide a reference for further reducing aeration energy in the autotrophic nitrogen removal process.

Calcium regulates nitrogen removal in constructed wetlands: The interactions between dissolved organic matter and microbial response

Limited carbon bioavailability in low C/N tailwater aggravated the difficulty of nitrogen (N) removal. Calcium (Ca) can regulate carbon transformation, but the effects of Ca on N removal in constructed wetlands (CWs) remain elusive. To fill the gap, this study constructed two types of Ca-based CWs, and the results revealed that the removal efficiencies of nitrate (NO3−-N) and total nitrogen (TN) in Ca-based CWs increased by 9.83–22.25 % and 8.59–18.72 %, respectively, compared to control group. The proportion of protein-like DOM and P/H index (represent bioavailability of DOC) increased in Ca-based CWs, supporting the growth of heterotrophic denitrifiers and facilitated NO3−-N and TN removal. In addition to directly enhancing complete denitrification, Ca-based CWs also optimize the N removal pathway. Moreover, Ca enhanced complexity and robust of the microbial network, increased microbial communication, thereby jointly facilitating N removal. This study highlighted Ca improved N removal by collaborating the interactions of DOM transformation and microbial activity, which provided an abundant and cheap substrate for the treatment of low C/N wastewater by CWs

Ferrous-driven efficient removal of nitrate and various heavy metals by Zoogloea sp. ZP7 under oligotrophic conditions: Kinetics and heavy metal stress responses

Denitrification in surface water is affected by its oligotrophic characteristics and heavy metal pollution. In this study, a heterotrophic denitrifying bacterium Zoogloea resiniphila ZP7, which can utilize ferrous (Fe2+) as an additional electron donor under oligotrophic conditions, was isolated. Strain ZP7 achieved a 98.5 % nitrate (NO3−-N) removal efficiency (NRE) within 12 h using sodium acetate as carbon sources, with a Fe2+ concentration of 10.0 mg L−1, pH of 7.0, carbon to nitrogen ratio of 2.0, and NO3−-N concentration of 5.01 mg L−1, and no nitrite residue was observed. The denitrification performance and the removing ability of heavy metal by strain ZP7 were studied when exposed to copper (Cu2+), zinc (Zn2+), and cadmium (Cd2+) alone or in combination. When 1.0 mg L−1 Cu2+, Zn2+, and Cd2+ were added simultaneously, the strain ZP7 could achieve 91.3 % NRE in 14 h, and the removal efficiencies of Cu2+, Zn2+, and Cd2+ were 68.1, 89.2, and 83.5 %, respectively. The results of fluorescence region integration showed that the response of strain ZP7 to different heavy metal system stresses was different. Moreover, the changes in the content of extracellular polymeric substances (EPS) and humic acids indicated that they contributed to the enhancement of the resistance of strain ZP7 to heavy metals. Various characterization results showed that EPS and bio‑iron oxides were beneficial to remove heavy metals. In future studies, strain ZP7 can be considered to be immobilized in biomaterials to treat contaminated oligotrophic water bodies to explore its application prospects in actual contaminated water bodies.

Pre-anoxic electro-stimulation enhanced simultaneous nitrification–denitrification in single-stage electrolysis-integrated sequencing batch biofilm reactor

Simultaneous nitrification–denitrification (SND) is a promising nitrogen removal process. However, total nitrogen (TN) removal is limited due to unsatisfactory denitrification. This study demonstrated that short-time (1 h) pre-anoxic electro-stimulation significantly enhanced SND efficiency in the aerobic phase by promoting the proliferation of mixotrophic and heterotrophic denitrifiers. SND and TN removal efficiencies at the optimal electric current (EC) (0.02 A) were 85.6 % and 93.9 %, which were 39.1 % and 17.2 % higher than control. Microbial community analysis indicated that the abundance of mixotrophic and heterotrophic denitrifiers significantly increased. H2 generated in the electro-stimulation process induced the proliferation of mixotrophic denitrifiers. The weak EC (0.02 A) promoted the activity and growth of heterotrophic denitrifiers by accelerating electron transfer. They concurrently mediated heterotrophic denitrification to enhance SND efficiency. PICRUSt2 analysis revealed that the abundance of denitrifying genes dramatically surged. This study provides new insights into applying electrolysis to achieve advanced SND while minimizing electricity consumption.

Functional and Microbiological Responses of Iron–Carbon Galvanic Cell-Supported Autotrophic Denitrification to Organic Carbon Variation and Dissolved Oxygen Shaking

Iron–carbon galvanic-cell-supported autotrophic denitrification (IC-ADN) is a burgeoning efficient and cost-effective process for low-carbon wastewater treatment. This study revealed the influence of organic carbon (OC) and dissolved oxygen (DO) on IC-ADN in terms of functional and microbiological characteristics. The nitrogen removal efficiency increased to 91.6% and 94.7% with partial organic carbon source addition to COD/TN of 1 and 3, respectively. The results of 16S rRNA high-throughput sequencing with nirS and cbbL clone libraries showed that Thiobacillus was the predominant autotrophic denitrifying bacteria (ADB) in the micro-electrolysis-based autotrophic denitrification, which obtained nitrogen removal efficiency of 80.9% after 96 h. The ADBs shifted gradually to heterotrophic denitrifying bacteria Thauera with increasing COD/TN ratio. DO concentration of 0.8 rarely affected the denitrification efficiency and the denitrifying communities. When the DO concentration increased to 2.8 mg/L, the nitrogen removal efficiency decreased to 69.1%. These results demonstrated that autotrophic denitrification was notably affected by COD/TN and high DO concentration, which could be used to acquire optimum conditions for nitrogen removal. These results provided an in-depth understanding of the influential factors for galvanic-cell-based denitrification and helped us construct a stable and highly efficient treatment process for insufficient carbon source wastewater.

A self-sustaining effect induced by iron sulfide generation and reuse in pyrite-woodchip mixotrophic bioretention systems: An experimental and modeling study

Dual electron donor bioretention systems have emerged as a popular strategy to enhance dissolved nitrogen removal from stormwater runoff. Pyrite-woodchip mixotrophic bioretention systems showed a promoted and stabilized removal of dissolved nutrients under complex rainfall conditions, but the sulfate reduction process that can induce iron sulfide generation and reuse was overlooked. In this study, experiments and models were applied to investigate the effects of filler configuration and dissolved organic carbon (DOC) dissolution rate on treatment performance and iron sulfide generation in pyrite-woodchip bioretention systems. Key parameters govern that DOC dissolution and microbe-mediated processes were obtained by experiments. The water quality models that integrate one-dimensional constant flow, sorption and microbial transformation kinetics were used to predict the performance of bioretention systems. Results showed that the mixotrophic bioretention system with woodchip mixed in the vadose zone and pyrite in the saturated zone achieves a better performance in both nitrogen removal efficiency and by-product control. Comparably, woodchip and pyrite mixed in the saturated zone could encounter a high secondary pollution risk. The sensitivity coefficients of oxic/anoxic DOC dissolution rates to total nitrogen removal are 0.36 and -2.43 respectively. Iron sulfide generation was affected by DOC distribution and the competition between heterotrophic denitrifiers, autotrophic denitrifiers, and sulfate-reducing bacteria (SRB). DOC accumulation has an antagonistic effect on iron production and sulfate reduction. Extra DOC accumulation favors sulfate reduction while high DOC concentration inhibits pyrite-based denitrification and reduces Fe(III) production. The recycling of iron sulfide can improve the robustness and sustainability of bioretention systems.

Immobilized consortium of heterotrophic nitrifying-aerobic denitrifying bacteria on various matrices for nitrogen removal from synthetic wastewater

Environmental contamination by nitrogen compounds such as ammonium and nitrate has increased extensively in the recent past, which necessitates the development of eco-friendly remediation technologies. In this study, three matrix types including pumice, aquarium ceramic filter, and calcium alginate beads were used to facilitate nitrogen removal with an immobilized heterotrophic nitrifying-aerobic denitrifying (HNAD) bacterial consortium. The HNAD bacterial consortium was made of Pseudomonas monteilii Nht, Pseudomonas mendocina AquaN, Rhodococcus erythropolis R1, and Acinetobacter calcoaceticus SCC2. The quality parameters for immobilization, such as the number of immobilized cells and their viability, were assessed. The highest number of bacterial cells (3.4 × 10 9) was immobilized on the aquarium ceramic filter, with 53% cell viability at 30°Ⅽ for two months. Pumice, aquarium ceramic filter, and calcium alginate achieved NH4+-N removal efficiencies of 85.3 ± 1.7%, 87.3 ± 2.2%, and 77.5 ± 3.99% within 24 h, respectively, and removed NO3−-N by 88.23 ± 0.36%, 93.95 ± 0.00%, and 71.29 ± 6.49% over 60 h. Additionally, immobilized cells on pumice and ceramic filter retained up to 84% of NH4+-N removal efficiency after 14 reuse cycles. These findings indicate that the immobilized HNAD bacterial consortium on the aquarium ceramic filter can be used as a suitable biofilter for treatment of high nitrogen wastewater.

Integration of mixotrophic denitrification and anammox in Thiothrix-hydroxyapatite coupled particles treating municipal wastewater against low temperature and organic load shocks

Sulfur-driven autotrophic denitrification coupled with anammox (SPDA) is a promising process for autotrophic nitrogen removal, but autotrophic microorganisms are challenging to retain in wastewater treatment units. In this study, a single-stage mixotrophic denitrification coupled with anammox (MDA) process was established in an up-flow anaerobic sludge bed reactor with Thiothrix-hydroxyapatite coupled particles. The advancement of the sludge granulation process effectively strengthened the resistance of the MDA system to environmental stresses, and the SPDA-dominated nitrogen removal process achieved a total nitrogen removal efficiency of 88.6 %, a nitrogen removal rate of 0.432 KgN/m3/d at temperatures as low as 12.8 °C and an organic loading rate of 0.63 KgCOD/m3/d. Interestingly, the ecological niche integration of sulfur oxidizing bacteria (SOB), anammox bacteria and heterotrophic denitrifying bacteria in the granular sludge was observed during long-term operation. The FISH-CLSM results revealed the spatial distribution of functional microorganisms in the granular sludge and the potential pollutant transformation mechanism. The aggressive growth of SOB dominated by Thiothrix constructed a biological wall at the periphery of the granules, which ensured their priority for nitrate acquisition, supplied nitrite to the internal anammox bacteria, and acted as an important barrier against the shock of organic loads. Overall, this study presents a novel granulation approach for the biological treatment of sulfur-containing wastewater, highlighting the efficacy of functional integration within Thiothrix-hydroxyapatite coupled particles as an effective strategy to resist environmental stress.

Increasing fish production in recirculating aquaculture system by integrating a biofloc-worm reactor for protein recovery

Aquaculture, producing half of global fish production, offers a high-quality protein source for humans. Improving nitrogen use efficiency (NUE) through microbial protein recovery is crucial for increasing fish production and reducing environmental footprint. However, the poor palatability and high moisture content of microbial protein make its utilization challenging. Here, a biofloc-worm reactor was integrated into a recirculating aquaculture system (BW_RAS) for the first time to convert microbial protein into Tubificidae (Oligochaeta) biomass, which was used as direct feed for culturing fish. Batch experiments indicated that an aeration rate of 0.132 m3L−1h−1 and a worm density of 0.3 g cm−2 on the carrier were optimal for microbial biomass growth and worm predation, respectively. Compared to the biofloc reactor-based recirculating aquaculture system (B_RAS), the BW_RAS improved water quality, NUE, and fish production by 17.1 % during a 120-day aquaculture period. The abundance of heterotrophic aerobic denitrifier Deinococcus in BW_RAS was one order of magnitude higher than in B_RAS, while heterotrophic bacteria Mycobacterium was more abundant in B_RAS. Denitrifiers cooperated with organic matter degraders and nitrogen assimilation bacteria for protein recovery and gaseous nitrogen loss while competing with predatory bacteria. Function prediction and qPCR indicated greater aerobic respiration, nitrate assimilation, nitrification (AOB-amoA), and denitrification (napA, nirK, nirS, nosZI), but lower fermentation in BWR compared to BR. This study demonstrated that BW_RAS increased microbial protein production and aerobic nitrogen cycling through ongoing worm predation, further enhancing fish production to a commercially viable level.

Enhancing total nitrogen removal in constructed wetlands: A Comparative study of iron ore and biochar amendments

Efficient nitrogen removal in constructed wetlands (CWs) remains challenging when treating agricultural runoff with a low carbon-to-nitrogen ratio (C/N). However, using biochar, iron ore, and FeCl3-modified biochar (Fe-BC) as amendments could potentially improve total nitrogen (TN) removal efficiency in CWs, but the underlying mechanisms associated with adding these substrates are unclear. In this study, five CWs: quartz sand constructed wetland (Control), biochar constructed wetland, Fe-BC constructed wetland, iron ore constructed wetland, and iron ore + biochar constructed wetland, were built to compare their treatment performance. The rhizosphere microbial community compositions and their co-occurrence networks were analyzed to reveal the underlying mechanisms driving their treatment performance. The results showed that iron ore was the most efficient amendment, although all treatments increased TN removal efficiency in the CWs. Ammonia-oxidizing, heterotrophic denitrifying, nitrate-dependent anaerobic ferrous oxidizing (NAFO), and Feammox bacteria abundance was higher in the iron ore system and led to the simultaneous removal of NH4+-N, NO3−-N, and NO2−-N. Visual representations of the co-occurrence networks further revealed that there was an increase in cooperative mutualism (the high proportion of positive links) and more complex interactions among genera related to the nitrogen and iron cycle (especially ammonia-oxidizing bacteria, heterotrophic denitrifying bacteria, NAFO bacteria, and Feammox bacteria) in the iron ore system, which ultimately contributed to the highest TN removal efficiency. This study provides critical insights into how different iron ore or biochar substrates could be used to treat agricultural runoff in CWs.

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

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

Exposure to zinc and dialkyldimethyl ammonium compound alters bacterial community structure and resistance gene levels in partial sulfur autotrophic denitrification coupled with the Anammox process

Dialkyldimethyl ammonium compound (DADMAC) is widely used in daily life as a typical disinfectant and often co-exists with the heavy metal zinc in sewage environments. This study investigated the effects of co-exposure to zinc (1 mg/L) and DADMAC (0.2–5 mg/L) on the performance, bacterial community, and resistance genes (RGs) in a partial sulfur autotrophic denitrification coupled with anaerobic ammonium oxidation (PSAD-Anammox) system in a sequencing batch moving bed biofilm reactor for 150 days. Co-exposure to zinc and low concentration (0.2 mg/L) DADMAC did not affect the nitrogen removal ability of the PASD-Anammox system, but increased the abundance and transmission risk of free RGs in water. Co-exposure to zinc and medium-to-high (2–5 mg/L) DADMAC led to fluctuations in and inhibition of nitrogen removal, which might be related to the enrichment of heterotrophic denitrifying bacteria dominated by Denitratisoma. Co-exposure to zinc and high concentration DADMAC (5 mg/L) stimulated the secretion of extracellular polymeric substances and increased the proliferation risk of intracellular RGs in sludge. This study provided insights into the application of PSAD-Anammox system and the ecological risks of wastewater containing zinc and DADMAC.

Microbial community dynamics in a two-stage treatment wetland: Insights from treating seasonal ski resort wastewater

Microbial community studies in treatment wetlands (TW) have not investigated cold-climate TW and have focused primarily on laboratory mesocosms. This study determined microbial community composition and dynamics using 16s rRNA sequencing in a partially-saturated two-stage, vertical-flow pilot TW treating ski resort wastewater from December–April. Indicator species analysis found organisms capable of the primary nitrogen transformations observed in each stage at 3 °C, with nitrifiers in the unsaturated second stage and heterotrophic denitrifiers in the first stage where recycled nitrate is removed. During seasonal TW operation, microbial communities developed significant differences between TW stages and α-diversity decreased. Microbial communities were compared to batch mesocosm TW communities, which were significantly different from the pilot. The study is the first to discern community dynamics and key microorganisms during seasonal loading of a cold-climate, field-scale TW. Results raise questions regarding comparability of mesocosms to pilot systems and indicate more field-scale TW investigations are needed.

The fate of various organic compounds including pharmaceuticals in the treatment of landfill leachate and anaerobic digestate using deammonification

Studies focusing on the fate of organic compounds specifically recalcitrant organics and pharmaceuticals in a deammonification plant are scarce. This study downscaled a full-scale deammonification plant, which was being operated with the same wastewater streams of this study and evaluated for the first time the fate of different fractions of organic compounds in deammonification process along with the removal of nitrogenous compounds. Organic compounds in the feed and effluent were analyzed and their changes in the deammonification process were characterized. The results demonstrate that deammonification achieved over 90% of ammonium removal. COD monitoring revealed that around 11% of COD was reduced during the process mainly due to aerobic COD degradation and heterotrophic denitrifying bacteria. Analysis of the organic fractions revealed that high molecular weight components (around 11 kDa) are susceptible to removal and their concentrations tend to decrease in the reactor. Concentrations of humic substances, specifically fulvic acid-like substances, were reduced. Tracking nine different pharmaceutical residues revealed that morphine and cotinine were completely removed, while pregabalin and oxazepam which are known to be highly recalcitrant were more than 60% removed. The research finding can help in optimizing treatment processes by revealing the fate of different organic compounds, especially pharmaceuticals, within the deammonification process, thereby contributing to better effluent quality, regulatory compliance, and environmental protection.

A pilot study of Mn redox driven nitrogen removal in municipal tailwater: Nitrogen metabolic pathway and electron transport mechanism

Microbial-mediated manganese (Mn) cycle coupled with nitrogen removal in municipal tailwater has received increasing attention. Herein, this study successfully extended the application of this technique to an 800 L biofilter, achieving a daily capacity of 2.4 m3 for treating municipal tailwater with a low C/N ratio. The operation results over 180 days indicated that the MnOx biofilter performed more efficiently than gravel control biofilter, achieving ∼ 40.2 % for nitrate and ∼ 35.9 % for total nitrogen (TN) removal efficiencies, respectively, under the conditions of hydraulic retention time (HRT) of 4 ∼ 5 h and low temperature (6.3–10.5 °C). To better elucidate the potential mechanisms, various functional bacteria involved in nitrogen and Mn transformation were identified, including heterotrophic denitrifying bacteria, such as Denitratisoma, Thauera, and Anaerolineae (cumulative relative abundance: 16.94–23.14 % in MnOx system vs. 9.86–11.90 % in control system) and Mn(II) oxidizing denitrifying bacteria, such as Pseudomonas and Acinetobacter (12.90–18.13 % vs. 3.59–4.64 %). Furthermore, metagenomic analysis indicated that the Mn cycle enhanced the expression of genes responsible for electron generation in organics/Mn metabolic pathways (e.g. TCA cycle and Mn oxidation), promoted electron transfer by enriching the Mn(II)-oxidizing denitrifying bacteria, and improved the abundance of genes associated with electron utilization (e.g. nitrate and nitrite reduction). The enhancement of the electron generation-transport-utilization due to Mn circulation contributed to efficient nitrate removal. This study provides further guidance of large-scale application and expands the understanding of the nitrogen removal mechanism in the Mn cycle-mediated process in municipal tailwater.

Simultaneous removal of ammonia nitrogen and manganese by anoxic/oxic (A/O) process from electrolytic manganese residue leachate

Ammonia nitrogen and manganese contained in the leachate of electrolytic manganese residue are causing serious damage to the ecological environment. In this study, anaerobic/oxic (A/O) process as a biological method to achieve simultaneous removal of nitrogen and manganese produced during the actual production of manganese. Results indicated that after 137 d of operation, the system achieved 96.73 ± 1.25%, 98.42 ± 0.60%, 96.97 ± 0.19%, and 100% removal of Mn(II), NH4+-N, TN, and chemical oxygen demand (COD), respectively. Extracellular polymeric substances (EPS) secreted by microorganisms was converted to carbon source to promote denitrification. Scanning electron microscopy (SEM) showed that Mn(II) formed manganese crystals in the process of complete reflection with sludge, acting as electron donor to achieve simultaneous removal of nitrogen and manganese, and removal of Mn(II) was found by X-ray photoelectron spectroscopy (XPS) through redox and valence changes. 16S rRNA indicated that Proteobacteria, Deinococcus-Thermus, Bacteroidete, and Firmicutes were involved in the process of removal nitrogen and manganese. The presence of Limnobacter allowed a more stable accumulation of NO2−-N in reactor, thus achieving shortcut nitrification-denitrification, and Flavihumibacter promoted the secretion of EPS. Collectively, the study reveals that the interaction of autotrophic denitrifying bacteria (Chryseolinea), heterotrophic denitrifying bacteria (Fusibacter, Thauera), and manganese-oxidizing bacteria (Pseudaminobacter, Bacillus, Pedomicrobium, Aeromonasr) achieved simultaneously removal of nitrogen and manganese from the reactor shortcut nitrification-denitrification. This study provides a new insight for the treatment of electrolytic manganese residue leachate produced in the process of manganese production.

Optimization of nitrogen removal by immobilized heterotrophic nitrification aerobic denitrifying bacterium using response surface methodology

An innovative bacterial carrier complex embedded with polyvinyl alcohol and sodium alginate complex (P.S.) was developed, using the heterotrophic nitrification-aerobic denitrification (HN-AD) strain Paracoccus sp. QD-21, which was carefully selected from activated sludge. The performance of the immobilized bacterium on ammonia nitrogen, nitrate nitrogen, and nitrite nitrogen was thoroughly analyzed, aiming to investigate the synergistic effects of the P.S.-encapsulated bacterial carrier on the HN-AD strain QD-21. To assess the adaptability of the immobilized bacterium, various single-factor analysis experiments were conducted, including the assessment of carbon source, C/N, temperature, pH, rotational speed, and salinity. Subsequently, response surface methodology was employed to conduct further optimization, leading to the identification of optimum conditions (147.31 rpm, 36°C, C/N 8.72, pH 8). Under these conditions, an impressive ammonia nitrogen removal rate of 94.82 ± 0.82% was achieved. Furthermore, practical wastewater treatment experiments were carried out, demonstrating a remarkable 63.31% increase in ammonia nitrogen removal when the carrier with HN-AD bacterium attached was employed. Notably, the addition of a specific carbon source remarkably enhanced the treatment efficiency, resulting in a complete removal rate of 100%. These significant findings lay a solid foundation for the engineering application of the HN-AD bacterium and highlight the promising potential of the P.S.-encapsulated bacterial carrier complex in various applications.

Microbial interactions facilitating efficient methane driven denitrification via in-situ utilization of short chain fatty acids

High nitrate pollution in agriculture and industry poses a challenge to emerging methane oxidation coupled denitrification. In this study, an efficient nitrate removal efficiency of 100 % was achieved at an influent loading rate of 400 mg-N/L·d, accompanied by the production of short chain fatty acids (SCFAs) with a maximum value of 80.9 mg/L. Batch tests confirmed that methane was initially converted to acetate, which then served as a carbon source for denitrification. Microbial community characterization revealed the dominance of heterotrophic denitrifiers, including Simplicispira (22.8 %), Stappia (4.9 %), and the high‑nitrogen-tolerant heterotrophic denitrifier Diaphorobacter (19.0 %), at the nitrate removal rate of 400 mg-N/L·d. Notably, the low abundance of methanotrophs ranging from 0.24 % to 3.75 % across all operational stages does not fully align with the abundance of pmoA genes, suggesting the presence of other functional microorganisms capable of methane oxidation and SCFAs production. These findings could facilitate highly efficient denitrification driven by methane and contributed to the development of denitrification using methane as an electron donor.