Denitrification Research Articles

Magnetic biochar accelerates microbial succession and enhances assimilatory nitrate reduction during pig manure composting

Biochar promotes microbial metabolic activities and reduces N2O on aerobic composting. However, the effects of magnetic biochar (MBC) on the microbial succession and N2O emissions during pig manure composting remain unclear. Herein, a 42-day composting experiment was conducted with five treatment regimes: pig manure without biochar (CK), 5 % pig manure-based biochar (5 % PBC), 2 % MBC (2 % MBC), 5 % MBC (5 % MBC) and 7.5 % MBC (7.5 % MBC)), to clarify the variation in functional microorganisms and genes associated with nitrogen and direct interspecies electron transfer via metagenomics. Fourier-transform infrared spectroscopy showed that MBC possessed more stable aromatic structures than pig manure-based biochar (PBC), indicating its greater potential for nitrous oxide reduction. MBC treatments were more effective in composting organic matter and improving the carbon/nitrogen ratio than PBC. The microbial composition during composting varied significantly, with the dominant phyla shifting from Firmicutes to Proteobacteria, Actinobacteria, and Bacteroidota. Network and hierarchical clustering analyses showed that the MBC treatment enhanced the interactions of dominant microbes (Proteobacteria and Bacteroidota) and accelerated the composting process. The biochar addition accelerated assimilatory nitrate reduction and slowed dissimilatory nitrate reduction and denitrification. The Mantel test demonstrated that magnetic biochar potentially helped regulate composting nutrients and affected functional nitrogen genes. These findings shed light on the role of MBC in mitigating greenhouse gas emissions during aerobic composting.

Pilot study on the filling rate of parallel AA‐MBBR fillers in plateau areas

AbstractComparative analysis of the effects of different filling rates of polypropylene filler on the operational effectiveness, microbial abundance and diversity, and metabolism of the AA‐MBBR system for the treatment of domestic wastewater. The results showed that under the condition of the optimal filling rate of 40%, the average removal rates of BOD5, COD, NH3‐N, TN, and TP in the AA‐MBBR system were 93.63%, 90.02%, 81.25%, 80.07%, and 74.51%, respectively, and the effluent COD was 143.05 mg/L and the effluent NH3‐N was 37.03 mg/L; the number of operational taxonomic units (OTUs) in the AA‐MBBR system with the optimal fill rate was 595, which was higher than 10%, 20%, 30%, and 50%, and it was also found that the relative abundance of denitrifying bacteria (DNB) and polyphosphate bacteria (Paos) increased with the increase of biofilm fill rate, DNB increased by about 32.9% at TCL (fill rate) 40 compared to TCL10, Paos increased by about 21.6% at TCL30 compared to TCL10. the AA‐MBBR system with a fill rate of 40% showed better pollutant removal than the other systems with other fill rates The bacterial diversity was higher than that of the systems with other filling rates.

Long-term effect of microplastics on anammox systems: From macro efficiency to micro metabolic mechanisms and antibiotic resistance genes proliferation

Wastewater treatment plants (WWTPs) are sinks for various types of microplastics (MPs), which have an undeniable impact on anaerobic ammonia oxidation (anammox) systems, yet their differential effects remain elusive. In this study, the long-term effects of non-biodegradable polyvinyl chloride (PVC) MPs and biodegradable polylactic acid (PLA) MPs in anammox systems were investigated from macro efficiency, micro metabolic mechanism, and antibiotic resistance gene proliferation. Results showed that the introduced 0.5 mg L−1 PVC MPs caused a 20.64 % decrease in nitrogen removal efficiency (NRE) compared to the control one, while PLA MPs resulted in a 10.73 % increase in NRE. Accordingly, the relative abundance of Ca. Kuenenia with the introduced PLA MPs was 16.68 % higher than the control group, while it was 10.89 % lower with the introduced PVC MPs. Furthermore, MPs inhibit the reactor performance by inducing intracellular reactive oxygen species (ROS) production in anammox cells. However, PLA MPs, as a slow biodegradable organic, have a reversible negative effect on ROS induction, which further improves nitrogen removal performance and sludge proliferation through dissimilatory nitrate reduction to ammonium (DNRA) and microbial interactions. Nevertheless, this also poses an increased risk of antibiotic resistance genes (ARGs) spread and proliferation in anammox sludge. This study clarified the comprehensive effects of different MPs types in anammox processes, highlighting the feasibility and potential risks in the engineering applications of anammox processes for MPs-contained wastewater treatment.

NH2-MIL-101(Fe)-mediated photo-Fenton reaction enhanced simultaneous removal of nitrogen and refractory organics in anammox process through interfacial electron transfer

In this study, H2O2 (0.1 ‰) and NH2-MIL-101(Fe)-driven (150 mg/L) photo-Fenton-coupled anammox were proposed to simultaneously improve the removal efficiency of nitrogen and humic acid. Long-term experiments showed that the total nitrogen removal efficiency was increased by the photo-Fenton reaction to 91.9 ± 1.5 % by altering the bioavailability of refractory organics. Correspondingly, the total organic carbon removal efficiency was significantly increased. Microbial community analyses indicated that Candidatus_Brocadia maintained high activity during photo-Fenton reaction and was the most abundant genus in the reactor. Dissimilatory nitrate reduction to ammonium process and denitrification process were enhanced, resulting in reduced NO3–-N production. The establishment of electron transfer between microorganisms and NH2-MIL-101 (Fe) improved the charge separation efficiency of the quantum dots and increased the intracellular adenosine triphosphate content of anammox bacteria. These results indicated that photo-Fenton-anammox process promoted the removal of nitrogen and refractory organics in one reactor which had good economic value and application prospects.

Functional similarity, despite taxonomical divergence in the millipede gut microbiota, points to a common trophic strategy

BackgroundMany arthropods rely on their gut microbiome to digest plant material, which is often low in nitrogen but high in complex polysaccharides. Detritivores, such as millipedes, live on a particularly poor diet, but the identity and nutritional contribution of their microbiome are largely unknown. In this study, the hindgut microbiota of the tropical millipede Epibolus pulchripes (large, methane emitting) and the temperate millipede Glomeris connexa (small, non-methane emitting), fed on an identical diet, were studied using comparative metagenomics and metatranscriptomics.ResultsThe results showed that the microbial load in E. pulchripes is much higher and more diverse than in G. connexa. The microbial communities of the two species differed significantly, with Bacteroidota dominating the hindguts of E. pulchripes and Proteobacteria (Pseudomonadota) in G. connexa. Despite equal sequencing effort, de novo assembly and binning recovered 282 metagenome-assembled genomes (MAGs) from E. pulchripes and 33 from G. connexa, including 90 novel bacterial taxa (81 in E. pulchripes and 9 in G. connexa). However, despite this taxonomic divergence, most of the functions, including carbohydrate hydrolysis, sulfate reduction, and nitrogen cycling, were common to the two species. Members of the Bacteroidota (Bacteroidetes) were the primary agents of complex carbon degradation in E. pulchripes, while members of Proteobacteria dominated in G. connexa. Members of Desulfobacterota were the potential sulfate-reducing bacteria in E. pulchripes. The capacity for dissimilatory nitrate reduction was found in Actinobacteriota (E. pulchripes) and Proteobacteria (both species), but only Proteobacteria possessed the capacity for denitrification (both species). In contrast, some functions were only found in E. pulchripes. These include reductive acetogenesis, found in members of Desulfobacterota and Firmicutes (Bacillota) in E. pulchripes. Also, diazotrophs were only found in E. pulchripes, with a few members of the Firmicutes and Proteobacteria expressing the nifH gene. Interestingly, fungal-cell-wall-degrading glycoside hydrolases (GHs) were among the most abundant carbohydrate-active enzymes (CAZymes) expressed in both millipede species, suggesting that fungal biomass plays an important role in the millipede diet.ConclusionsOverall, these results provide detailed insights into the genomic capabilities of the microbial community in the hindgut of millipedes and shed light on the ecophysiology of these essential detritivores.6T24awGbkoVb6oAp62-bVTVideo

Ditch level-dependent N removal capacity of denitrification and anammox in the drainage system of the Ningxia Yellow River irrigation district

Drainage networks, consisting of different levels of ditches, play a positive role in removing reactive nitrogen (N) via self-purification before drainage water returns to natural water bodies. However, relatively little is known about the N removal capacity of irrigation agricultural systems with different drainage ditch levels. In this study, we employed soil core incubation and soil slurry 15N paired tracer techniques to investigate the N removal rate (i.e., N2 flux), denitrification, anaerobic ammonium oxidation (anammox), and dissimilatory nitrate reduction to ammonium (DNRA) rates in the Ningxia Yellow River irrigation district at various ditch levels, including field ditches (FD), paddy field ditches (PFD), lateral ditches (LD1 and LD2), branch ditches (BD1, BD2, BD3), and trunk ditches (TD). The results indicated that the N removal rate ranged from 44.7 to 165.22 nmol N g−1 h−1 in the ditches, in the following decreasing order: trunk ditches > branch ditches > paddy field ditches > lateral ditches > field ditches. This result suggested that the N removal rate in drainage ditches is determined by the ditch level. In addition, denitrification and anammox were the primary pathways for N removal in the ditches, contributing 68.40–76.64 % and 21.55–30.29 %, respectively, to the total N removal. In contrast, DNRA contributed only 0.82–2.15 % to the total nitrate reduction. The N removal rates were negatively correlated with soil EC and pH and were also constrained by the abundances of denitrification functional genes. Overall, our findings suggest that the ditch level should be considered when evaluating the N removal capacity of agricultural ditch systems.

Dominant nitrogen metabolisms of a warm, seasonally anoxic freshwater ecosystem revealed using genome resolved metatranscriptomics.

Nitrogen (N) availability is one of the principal drivers of primary productivity across aquatic ecosystems. However, the microbial communities and emergent metabolisms that govern N cycling in tropical lakes are both distinct from and poorly understood relative to those found in temperate lakes. This latitudinal difference is largely due to the warm (>20°C) temperatures of tropical lake anoxic hypolimnions (deepest portion of a stratified water column), which result in unique anaerobic metabolisms operating without the temperature constraints found in lakes at temperate latitudes. As such, tropical hypolimnions provide a platform for exploring microbial membership and functional diversity. To better understand N metabolism in warm anoxic waters, we combined measurements of geochemistry and water column thermophysical structure with genome-resolved metatranscriptomic analyses of the water column microbiome in Lake Yojoa, Honduras. We sampled above and below the oxycline in June 2021, when the water column was stratified, and again at the same depths and locations in January 2022, when the water column was mixed. We identified 335 different lineages and significantly different microbiome membership between seasons and, when stratified, between depths. Notably, nrfA (indicative of dissimilatory nitrate reduction to ammonium) was upregulated relative to other N metabolism genes in the June hypolimnion. This work highlights the taxonomic and functional diversity of microbial communities in warm and anoxic inland waters, providing insight into the contemporary microbial ecology of tropical ecosystems as well as inland waters at higher latitudes as water columns continue to warm in the face of global change.IMPORTANCEIn aquatic ecosystems where primary productivity is limited by nitrogen (N), whether continuously, seasonally, or in concert with additional nutrient limitations, increased inorganic N availability can reshape ecosystem structure and function, potentially resulting in eutrophication and even harmful algal blooms. Whereas microbial metabolic processes such as mineralization and dissimilatory nitrate reduction to ammonium increase inorganic N availability, denitrification removes bioavailable N from the ecosystem. Therefore, understanding these key microbial mechanisms is critical to the sustainable management and environmental stewardship of inland freshwater resources. This study identifies and characterizes these crucial metabolisms in a warm, seasonally anoxic ecosystem. Results are contextualized by an ecological understanding of the study system derived from a multi-year continuous monitoring effort. This unique data set is the first of its kind in this largely understudied ecosystem (tropical lakes) and also provides insight into microbiome function and associated taxa in warm, anoxic freshwaters.

High ammonia loading rate and biofilm reattachment initiated partial nitrification and anammox in a membrane aerated biofilm reactor

Inhibition of nitrite oxidizing bacteria (NOB) is important for efficient wastewater treatment via anammox. Here, a novel strategy was demonstrated, which based on the detachment and reattachment of the biofilm to establish partial nitrification and anammox in a membrane aerated biofilm reactor (MABR). The results showed that using free ammonia as the sole inhibitor to start up partial nitrification took more time, and NOB acclimated to the free ammonia inhibition. Detaching the biofilm made it possible for free ammonia to contact NOB in deeper zones of the biofilm while at the same time washing NOB out of the reactor. Nitrite accumulation efficiency (NAE) increased to 70 % in 50 days when both high ammonia loading and biofilm reattachment were applied. With a short hydraulic retention time and high concentrations of ammonia and nitrite, anammox activity appeared in the reactor and resulted in a nitrogen removal efficiency of 60 %. Microbial community analysis indicated that Nitrosomonas, Nitrospira, and Candidatus Brocadia were the main functional taxa. Anaerobic ammonia oxidizing bacteria (AnAOB) proliferated within the biofilm, inhibiting NOB growth by competing for nitrite. Endogenous denitrifying bacteria also grew, coexisting with AnAOB. Predicted abundance of genes related to fermentation correlated with those involved in denitrification, indicating the possibility of heterotrophic denitrification. These findings help further our understanding of biofilm control-based strategies for NOB management and have implications for establishing autotrophic nitrogen removal using MABRs.

Diversity, composition, metabolic characteristics, and assembly process of the microbial community in sewer system at the early stage.

Sewer systems play vital roles in wastewater treatment facilities, and the microbial communities contribute significantly to the transformation of domestic wastewater. Therefore, this study conducted a 180-day experiment on a sewer system and utilized the high-throughput sequencing technology to characterize the microbial communities. Additionally, community assembly analysis was performed to understand the early-stage dynamics within the sewer system. The results demonstrated that the overall diversity of microbial communities exhibited fluctuations as the system progressed. The dominant phyla observed were Chloroflexi, Bacteroidetes, Firmicutes, and Proteobacteria, accounting for over 85.4% of the total relative abundances. At the genus level, bacteria associated with fermentation displayed a high relative abundance, particularly during days 75 to 180. A random-forest machine-learning model identified a group of microbes that confirmed the substantial contribution of fermentation. During the process of fermentation, microorganisms predominantly utilized propionate formation as the main pathway for acidogenesis, followed by acetate and butyrate formation. In terms of nitrogen and sulfur cycles, dissimilatory nitrate reduction and assimilatory sulfate reduction played significant roles. Furthermore, stochastic ecological processes had a dominant effect during the experiment. Dispersal limitation primarily governed the assembly process almost the entire experimental period, indicating the strong adaptability and metabolic plasticity of microorganisms in response to environmental variations. This experiment provides valuable insights into the metabolic mechanisms and microbial assembly associated with sewer systems.

Screening and diversity of culturable HNAD bacteria in the MBR sewage treatment system.

The activated sludge was collected from the Membrane BioReactor (MBR) pool of the sewage treatment system of Sanxing Town, Jintang County, Chengdu, to obtain a good population of heterotrophic nitrifying/aerobic denitrifying (HNAD) bacteria. After undergoing enrichment, isolation, and purification, the HNAD bacteria were selected using the pure culture method. The 16S rDNA molecular technology was used to determine the taxonomy of bacteria. The heterophic nitrifying ability and denitrification capacity of HNAD strains was ascertained through their growth characteristics in heterotrophic nitrification and denitrification media. The results showed that 53 HNAD strains selected from the MBR pool belonged to 2 phyla, 3 classes, 6 orders, 6 families, and 7 genera, with 26 species. Acinetobacter was the largest and dominant genus. Among these, strains numbered (bacterial strain) SW21HD14, SW21HD17, and SW21HD18 were potentially new species in the Acinetobacter genus. Each HNAD strain showed a significant heterotrophic nitrifying and aerobic denitrifying efficiency compared with the control strain (P < 0.05). Specifically, 10 strains demonstrated ammonia nitrogen degradation of greater than 70 mg·L-1 and 9 strains demonstrated nitrate nitrogen degradation above 150 mg·L-1. The HNAD bacteria, which were selected from the MBR pool of sewage treatment system of the Sanxing Town sewage treatment plant, exhibited rich diversity and strong nitrogen removal ability. These findings offered an effective strain source and theoretical basis for implementing biological denitrification technology that involves synchronous nitrification and denitrification.

Characteristics of Greenhouse Gas Emissions from Constructed Wetlands Vegetated with Myriophyllum aquatic: The Effects of Influent C/N Ratio and Microbial Responses

This study designed surface flow constructed wetlands (SFCWs) with Myriophyllum aquaticum (M. aquaticum) to evaluate how different influent C/N ratios (0:1 (C0N), 5:1 (C5N), 10:1 (C10N), and 15:1 (C15N)) affect pollutant removal, greenhouse gas (GHG) emissions, and microbial communities. The results showed that effluent ammonia nitrogen (NH4+-N), nitrate nitrogen (NO3−-N), and total nitrogen (TN) concentrations decreased, but effluent chemical oxygen demand (COD) concentration increased with increasing influent C/N ratios. The highest removal rates of TN (73.17%) and COD (74.56%) were observed with C5N. Regarding GHG emissions, a few changes in CO2 fluxes were caused by the influent C/N ratio, whereas CH4 fluxes obviously increased with the increasing influent C/N ratio. The highest N2O emission occurred with C0N (211.03 ± 44.38 mg-N·m−2·h−1), decreasing significantly with higher C/N ratios. High-throughput sequencing revealed that different influent C/N ratios directly influenced the microbial distribution and composition related to CH4 and N2O metabolism in SFCWs. The highest abundance (46.24%) of denitrifying bacteria (DNB) was observed with C5N, which helped to achieve efficient nitrogen removal with a simultaneous reduction in N2O emissions. Methanogen abundance rose with higher C/N ratios, whereas methanotrophs peaked under C5N and C10N conditions. Additionally, the random forest model identified influent C/N ratio and Rhodopseudomonas as primary factors influencing CH4 and N2O emissions, respectively. This highlights the importance of the influent C/N ratio in regulating both pollutant removal and GHG emissions in constructed wetlands.

Enhanced adaptability of pyrite-based constructed wetlands for low carbon to nitrogen ratio wastewater treatments: Modulation of nitrogen removal mechanisms and reduction of carbon emissions

Pyrite-based constructed wetlands (CWs) stimulated nitrate removal performance at low carbon to nitrogen (C/N) ratio has been gaining widely attention. However, the combined effects of pyrite and C/N on the nitrate removal mechanisms and greenhouse gases (GHGs) reduction were ignored. This study found that pyrite-based CWs significantly enhanced nitrate removal in C/N of 0, 1.5 and 3 by effectively driving autotrophic denitrification with high abundance of autotrophs denitrifiers (Rhodanobacter) and nitrate reductase (EC 1.7.7.2), while the enhancement was weakened in C/N of 6 by combined effect of mixotrophic denitrification and dissimilatory nitrate reduction to ammonium (DNRA) with high abundance of organic carbon-degrading bacteria (Stenotrophobacter) and DNRA-related nitrite reductase genes (nrf). Moreover, pyrite addition significantly reduced GHGs emissions from CWs in all stages with the occurrence of iron-coupled autotrophic denitrification. The study shed light on the potential mechanism for pyrite-based CWs for treating low C/N ratio wastewater.

Factors affecting the enrichment of heterotrophic nitrifying aerobic denitrifying bacteria in municipal wastewater treatment systems and analysis of differences in nitrogen metabolism

In this study, a four-stage MPSR process was employed for the treatment of urban wastewater. Three different conditions were established by controlling dissolved oxygen (DO) concentration, organic load rate (OLR), and influent pH value to examine pollutant removal effects and the changes of the heterotrophic nitrification-aerobic denitrification (HN-AD) functional bacteria communities. The factors driving changes in the HN-AD bacterial community and differences in nitrogen metabolism were revealed in the urban wastewater treatment system. The results showed that by forming anoxic/aerobic alternate oxygen environments and increasing influent OLR and pH, various HN-AD bacteria (Pseudomonas, Aeromonas, Flavobacterium) were enriched in the system, leading to improved removal capacity for NH4+-N and TN. Under optimal condition, the removal rates of NH4+-N and TN were 94.0 % and 78.7 % respectively. The dominant species of HN-AD bacteria were influenced by OLR and pH. The HN-AD bacteria exhibited the capacity for heterotrophic nitrification and aerobic denitrification within the system. This study provides a technological basis for the application of HN-AD bacteria for wastewater treatment.

Response of sulfide autotrophic denitrification process and microbial community to oxytetracycline stress

The coexistence of antibiotics with sulfide and nitrate is common in sewage. Thus, this study explored the removal performance of nitrate and sulfide, and the response of extracellular polymer substances (EPS) and the microbial community to the sulfide autotrophic denitrification (SAD) process under oxytetracycline (OTC) stress. In Phase Ⅰ, the SAD system showed favouranle performance (nitrate removal rate ＞ 92.57%, sulfide removal rate ＞ 97.75%). However, in Phase Ⅳ, at OTC concentrations of 10, 15, and 20 mg/L, the NRE decreased to 76.13%, 40.71%, 11.37%, respectively, and the SRE decreased to 97.58%, 97.09%, 92.84%, respectively. At OTC concentrations of 0, 10, 15, and 20 mg/L, the EPS content were 1.62, 1.75, 2.03, and 1.42 mg/gVSS, respectively. The results showed that SAD performance gradually deteriorated under OTC stress. In particular, when the OTC concentration was 20 mg/L, the EPS content was lower than that of the control test, which could be attributed to the occurrence of microbial death. Finally, high-throughput sequencing results showed that OTC exposure led to gradual domination by heterotrophic denitrifying bacteria.

Enhanced denitrification driven by a novel iron-carbon coupled primary cell: chemical and mixotrophic denitrification

Iron-carbon micro-electrolysis system is a promising method for promoting electron transfer in nitrate removal. However, many traditional approaches involving simple physical mixing inevitably suffered from the confined iron-carbon contact area and short validity period, leading to the overuse of iron. Here, a ceramsite-loaded microscale zero-valent iron (mZVI) and acidified carbon (AC) coupled-galvanic cell (CMC) was designed to support chemical, autotrophic and heterotrophic denitrification. Long-term experiments were conducted to monitor the nitrogen removal performance of denitrification reactors filled with CMC and thus optimized the denitrification performance by improving fabrication parameters and various operating conditions. The denitrification contributions test showed that the chemical denitrification pathway contributed most to nitrate removal (57.3%), followed by autotrophic (24.6%) and heterotrophic denitrification pathways (18.1%). The microbial analysis confirmed the significant aggregation of related denitrifying bacteria in the reactors, while AC promoted the expression of relevant nitrogen metabolism genes because of accelerated uptake and utilization of iron complexes. Meanwhile, the electrochemical analysis revealed a significantly improved electron transfer capacity of AC compared to pristine carbon. Overall, our study demonstrated the application of a novel mZVI-AC coupled material for effective nitrate removal and revealed the potential impact of CMC in the multipathway denitrification process.Graphical

"Candidatus Siderophilus nitratireducens": a putative nap-dependent nitrate-reducing iron oxidizer within the new order Siderophiliales.

Nitrate leaching from agricultural soils is increasingly found in groundwater, a primary source of drinking water worldwide. This nitrate influx can potentially stimulate the biological oxidation of iron in anoxic groundwater reservoirs. Nitrate-dependent iron-oxidizing (NDFO) bacteria have been extensively studied in laboratory settings, yet their ecophysiology in natural environments remains largely unknown. To this end, we established a pilot-scale filter on nitrate-rich groundwater to elucidate the structure and metabolism of nitrate-reducing iron-oxidizing microbiomes under oligotrophic conditions mimicking natural groundwaters. The enriched community stoichiometrically removed iron and nitrate consistently with the NDFO metabolism. Genome-resolved metagenomics revealed the underlying metabolic network between the dominant iron-dependent denitrifying autotrophs and the less abundant organoheterotrophs. The most abundant genome belonged to a new Candidate order, named Siderophiliales. This new species, "Candidatus Siderophilus nitratireducens," carries genes central genes to iron oxidation (cytochrome c cyc2), carbon fixation (rbc), and for the sole periplasmic nitrate reductase (nap). Using thermodynamics, we demonstrate that iron oxidation coupled to nap based dissimilatory reduction of nitrate to nitrite is energetically favorable under realistic Fe3+/Fe2+ and NO3-/NO2- concentration ratios. Ultimately, by bridging the gap between laboratory investigations and nitrate real-world conditions, this study provides insights into the intricate interplay between nitrate and iron in groundwater ecosystems, and expands our understanding of NDFOs taxonomic diversity and ecological role.

Anaerobic oxidation of ammonium and short-chain gaseous alkanes coupled to nitrate reduction by a bacterial consortium.

The bacterial species "Candidatus Alkanivorans nitratireducens" was recently demonstrated to mediate nitrate-dependent anaerobic oxidation of short-chain gaseous alkanes (SCGAs). In previous bioreactor enrichment studies, the species appeared to reduce nitrate in two phases, switching from denitrification to dissimilatory nitrate reduction to ammonium (DNRA) in response to nitrite accumulation. The regulation of this switch or the nature of potential syntrophic partnerships with other microorganisms remains unclear. Here, we describe anaerobic multispecies cultures of bacteria that couple the oxidation of propane and butane to nitrate reduction and the oxidation of ammonium (anammox). Batch tests with 15N-isotope labelling and multi-omic analyses collectively supported a syntrophic partnership between "Ca. A. nitratireducens" and anammox bacteria, with the former species mediating nitrate-driven oxidation of SCGAs, supplying the latter with nitrite for the oxidation of ammonium. The elimination of nitrite accumulation by the anammox substantially increased SCGA and nitrate consumption rates, whereas it suppressed DNRA. Removing ammonium supply led to its eventual production, the accumulation of nitrite, and the upregulation of DNRA gene expression for the abundant "Ca. A. nitratireducens". Increasing the supply of SCGA had a similar effect in promoting DNRA. Our results suggest that "Ca. A. nitratireducens" switches to DNRA to alleviate oxidative stress caused by nitrite accumulation, giving further insight into adaptability and ecology of this microorganism. Our findings also have important implications for the understanding of the fate of nitrogen and SCGAs in anaerobic environments.

Implications for nitrogen and sulphur cycles: phylogeny and niche-range of Nitrospirota in terrestrial aquifers.

Increasing evidence suggests Nitrospirota are important contributors to aquatic and subsurface nitrogen and sulphur cycles. We determined the phylogenetic and ecological niche associations of Nitrospirota colonizing terrestrial aquifers. Nitrospirota compositions were determined across 59 groundwater wells. Distributions were strongly influenced by oxygen availability in groundwater, marked by a trade-off between aerobic (Nitrospira, Leptospirillum) and anaerobic (Thermodesulfovibrionia, unclassified) lineages. Seven Nitrospirota metagenome-assembled genomes (MAGs), or populations, were recovered from a subset of wells, including three from the recently designated class 9FT-COMBO-42-15. Most were relatively more abundant and transcriptionally active in dysoxic groundwater. These MAGs were analysed with 743 other Nitrospirota genomes. Results illustrate the predominance of certain lineages in aquifers (e.g. non-nitrifying Nitrospiria, classes 9FT-COMBO-42-15 and UBA9217, and Thermodesulfovibrionales family UBA1546). These lineages are characterized by mechanisms for nitrate reduction and sulphur cycling, and, excluding Nitrospiria, the Wood-Ljungdahl pathway, consistent with carbon-limited, low-oxygen, and sulphur-rich aquifer conditions. Class 9FT-COMBO-42-15 is a sister clade of Nitrospiria and comprises two families spanning a transition in carbon fixation approaches: f_HDB-SIOIB13 encodes rTCA (like Nitrospiria) and f_9FT-COMBO-42-15 encodes Wood-Ljungdahl CO dehydrogenase (like Thermodesulfovibrionia and UBA9217). The 9FT-COMBO-42-15 family is further differentiated by its capacity for sulphur oxidation (via DsrABEFH and SoxXAYZB) and dissimilatory nitrate reduction to ammonium, and gene transcription indicated active coupling of nitrogen and sulphur cycles by f_9FT-COMBO-42-15 in dysoxic groundwater. Overall, results indicate that Nitrospirota are widely distributed in groundwater and that oxygen availability drives the spatial differentiation of lineages with ecologically distinct roles related to nitrogen and sulphur metabolism.

Enhanced nitrogen removal through aerobic denitrifying bacteria in horizontal subsurface flow constructed wetlands: Influencing factors and microbial community structure

Variation in the demand for dissolved oxygen during nitrification and denitrification processes is one of the main constraints for nitrogen removal performance in conventional constructed wetlands. This study aimed to enhance the performance of horizontal submerged flow constructed wetland (HSCW) systems through aerobic denitrification (AD). The effects of different operating conditions on the performance and microbial community structure of the system were investigated using three experimental systems including a non-aerated HSCW without bacterial inoculation (CWC), a non-aerated HSCW with bacterial inoculation (CWM) and an intermittently aerated HSCW with bacterial inoculation (CWMA). Results showed that C/N ratio of 10 and aeration rate of 0.2 L/(min·L) were the optimum conditions for CWMA, which showed 29.01 % and 12.17 % higher COD and TN removals than CWC, even at an average temperature of 8.80℃. Microbial community analysis revealed that the addition of Pseudomonas stutzeri significantly increased the microbial diversity and abundance within the HSCWs. This study provides a new idea for the treatment of nitrogenous wastewater and the AD-CW system has a promising application in low-temperature areas.

Nitrate reduction capacity of the oral microbiota is impaired in periodontitis: potential implications for systemic nitric oxide availability

The reduction of nitrate to nitrite by the oral microbiota has been proposed to be important for oral health and results in nitric oxide formation that can improve cardiometabolic conditions. Studies of bacterial composition in subgingival plaque suggest that nitrate-reducing bacteria are associated with periodontal health, but the impact of periodontitis on nitrate-reducing capacity (NRC) and, therefore, nitric oxide availability has not been evaluated. The current study aimed to evaluate how periodontitis affects the NRC of the oral microbiota. First, 16S rRNA sequencing data from five different countries were analyzed, revealing that nitrate-reducing bacteria were significantly lower in subgingival plaque of periodontitis patients compared with healthy individuals (P < 0.05 in all five datasets with n = 20–82 samples per dataset). Secondly, subgingival plaque, saliva, and plasma samples were obtained from 42 periodontitis patients before and after periodontal treatment. The oral NRC was determined in vitro by incubating saliva with 8 mmol/L nitrate (a concentration found in saliva after nitrate-rich vegetable intake) and compared with the NRC of 15 healthy individuals. Salivary NRC was found to be diminished in periodontal patients before treatment (P < 0.05) but recovered to healthy levels 90 days post-treatment. Additionally, the subgingival levels of nitrate-reducing bacteria increased after treatment and correlated negatively with periodontitis-associated bacteria (P < 0.01). No significant effect of periodontal treatment on the baseline saliva and plasma nitrate and nitrite levels was found, indicating that differences in the NRC may only be revealed after nitrate intake. Our results suggest that an impaired NRC in periodontitis could limit dietary nitrate-derived nitric oxide levels, and the effect on systemic health should be explored in future studies.