Ammonia Oxidation Activity Research Articles

Autotrophic growth activity of complete ammonia oxidizers in an upflow biological contact filter for drinking water treatment.

Biological filters effectively remove ammonium from drinking water via nitrification. In a pilot-scale upflow biological contact filter (U-BCF), complete ammonia oxidizers (comammox), which are capable of oxidizing ammonia to nitrate in one cell, were more abundant than ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB). However, little information is available on the contribution of comammox to nitrification. In this study, we evaluated the autotrophic growth activity of comammox associated with biological activated carbon (BAC) in a U-BCF by DNA-stable isotope probing (DNA-SIP). BAC samples collected from the U-BCF were continuously fed mineral medium containing 0.14mg N L-1 ammonium and 12C- or 13C-labeled bicarbonate for 20 days. DNA-SIP analysis revealed that comammox (clades A & B) as well as AOA, assimilated bicarbonate after 10 days of incubation, proving that dominant comammox could contribute to nitrification. Contrarily, AOB remained inactive throughout the observation period. Amplicon sequencing of the 13C-labeled DNA fractions of comammox revealed that specific genotypes other than the most dominant genotype in the original sample were more enriched under the incubation condition for the DNA-SIP experiment. Thus, dominant genotypes of comammox in a U-BCF might utilize organic nitrogen to fuel nitrification in ammonia-limited environments.

Metabolic inhibitor-free assessment of the heterotrophic ammonia-oxidizing activity in activated sludge

When the contributions of three ammonia-oxidizing pathways (heterotrophic or autotrophic aerobic ammonia oxidization, and anammox) to wastewater biological nitrogen removal systems was compared by determining their ammonia-oxidizing activities, the key question is how to accurately determine the potential heterotrophic aerobic ammonia-oxidizing (PHAe) activity when the potential autotrophic aerobic ammonia-oxidizing (PAAe) activity (by ammonia-oxidizing bacteria (AOB) or archaea, or complete ammonia oxidization bacteria) also contributes to ammonia oxidization in PHAe activity assay medium. Using a AOB species and three heterotrophic AOB species as inocula, we demonstrated the feasibility of PHAe activity evaluation in the absence of a metabolic inhibitor, i.e., by subtracting the PAAe activity determined in PAAe activity assay medium from a combination of PAAe and PHAe activity determined in PHAe activity assay medium. Binary organic carbon sources (i.e., glucose and acetate) were included in the PHAe activity assay medium to fulfill the carbon requirements of most heterotrophic AOB genera. Higher ammonia-oxidizing activity in AOB biomass than heterotrophic AOB biomass (35.6 vs. 2.6–10.0 mg NH4+–N g−1 MLSS h−1) provides the remarkable advantages of autotrophic aerobic ammonia oxidization in biological nitrogen removal systems. Ammonia removal in three full-scale biological nitrogen removal systems for sewage treatment was predominantly mediated by PAAe activity (1.9–3.3 vs. 0.0–0.3 mg NH4+–N g1 MLSS h−1).

Salinity changes the nitrification activity and community composition of comammox Nitrospira in intertidal sediments of Yangtze River estuary.

The newly discovered complete ammonia-oxidizing (comammox) Nitrospira has been identified in different environments, including coastal environments, where salinity is one of the most important factors for the abundance and activity of nitrifiers. Here, we demonstrate the effect of salinity on comammox Nitrospira, canonical ammonia-oxidizing bacteria (AOB), and ammonia-oxidizing archaea (AOA) in the intertidal sediments of the Yangtze River estuary based on microcosm experiments, DNA stable-isotope probing (DNA-SIP), and potential ammonium-oxidation rate (PAR) tests for different groups of ammonia oxidizers with selective inhibitors. During microcosm incubations, the abundance of comammox Nitrospira was more sensitive to increased salinity than that of other ammonia oxidizers. The results obtained with DNA-SIP heavy fractions showed that the dominant phylotype in clade A.2 (containing genes involved in the adaptation to haloalkaline environments) had high proportions in comammox Nitrospira community under both freshwater (0.06% salinity) and highly saline water (3% salinity) conditions. In contrast, another phylotype of clade A.2 (which lacks these genes) was dominant only under freshwater conditions. The PARs confirmed that comammox Nitrospira presented greater contributions to nitrification under freshwater conditions with a PAR of 4.37 ± 0.53 mg N·day-1·kg soil-1 (54%) than under saline water conditions with a PAR of 0.60 ± 0.94 mg N·day-1·kg soil-1 (18%). Moreover, AOA were specific to saline water conditions, whereas AOB were common under both freshwater and saline water conditions (44% and 52%, respectively). The present study provided evidence that salinity markedly affects the activity of comammox Nitrospira, and that the salt sensitivity of different phylotypes varies. IMPORTANCE Complete ammonia oxidation (comammox) is a newly discovered type of nitrification through which ammonia is oxidized to nitrate in an organism. Comammox Nitrospira were abundantly found in coastal ecosystems and demonstrated high community diversity. Changes in salinity are considered one of the most important factors to comammox Nitrospira in coastal ecosystems; however, reports on the correlation between them remain inconsistent. Therefore, it is critical to experimentally determine the influence of salinity on comammox Nitrospira in the coastal ecosystem. This study demonstrated a clear effect of salinity on the abundance, activity, and relative contribution of different ammonia oxidizers, especially for comammox Nitrospira. To the best of our knowledge, this is the first study demonstrating comammox Nitrospira activity at seawater salinities, implying the existence of a salt-tolerant type comammox Nitrospira, despite its activity being much lower than in freshwater conditions. The indicated correlation between the activity of specific comammox Nitrospira and salinity is anticipated to provide insights into the distribution of comammox Nitrospira and their potential contributions in estuaries and coastal ecosystems.

Thickness of hydrogel for nitrifying biomass entrapment determines the free ammonia susceptibility differently in batch and continuous modes

Hydrogels immobilizing nitrifying bacteria with different thicknesses of 0.55 and 1.13 cm (HG-0.55 and HG-1.13, respectively) were produced. It was recognized that the thickness of media is a crucial parameter that affects both the stability and efficiency of wastewater treatment. Batch mode experiments were conducted to quantify specific oxygen uptake rate (SOUR) values at various total ammonium nitrogen (TAN) concentrations and pH levels. In the batch test, HG-0.55 exhibited 2.4 times higher nitrifying activity than HG-1.13, with corresponding SOUR values of 0.00768 and 0.00317 mg-O2/L mL-PVA min, respectively. However, HG-0.55 was more susceptible to free ammonia (FA) toxicity than HG-1.13, resulting in a reduction of 80% and 50% in SOUR values for HG-0.55 and -1.13, respectively, upon increasing the FA concentration from 15.73 to 118.12 mg-FA/L. Continuous mode experiments were conducted to assess the partial nitritation (PN) efficiency in practical applications, where continuous wastewater inflow maintains low FA toxicity through high ammonia-oxidizing rates. With step-wise TAN concentration increases, HG-0.55 experienced a gentler increase in FA concentration compared to HG-1.13. At a nitrogen loading rate of 0.78–0.95 kg-N/m3 day, the FA increase rate for HG-0.55 was 0.0179 kg-FA/m3 day, while that of HG-1.13 was 0.0516 kg-FA/m3 day. In the batch mode, where wastewater is introduced all at once, the high accumulation of FA posed a disadvantage for the FA-susceptible HG-0.55, which made it unsuitable for application. However, in the continuous mode, the thinner HG-0.55, with its larger surface area and high ammonia oxidation activity, proved to be suitable and demonstrated its effectiveness. This study provides valuable insights and a framework for the utilization strategy of immobilized gels in addressing the toxic effects of FA in practical processes.

Metagenomic and metaproteomic insights into the microbiome and the key geobiochemical potentials on the sandstone of rock-hewn Beishiku Temple in Northwest China

Metagenomics and metaproteomics analyses were used to determine the microbial diversity and taxon composition, as well as the biochemical potentials of the microbiome on the sandstone of Beishiku Temple located in Northwest China. Taxonomic annotation of the metagenomic dataset revealed the predominant taxa of the stone microbiome on this cave temple with characteristics of resistance to harsh environmental conditions. Meanwhile, there were also taxa in the microbiome that showed sensitivity to environmental factors. The taxa distribution and the metabolic functional distribution patterns by the metagenome and metaproteome, respectively, showed clear differences. The high abundance of energy metabolism represented in the metaproteome suggested that there were active geomicrobiological cycles of elements within the microbiome. The taxa responsible for reactions in the nitrogen cycle from both metagenome and metaproteome supported a metabolically active nitrogen cycle, and the high activity of Comammox bacteria indicated the strong metabolic activity of ammonia oxidation to nitrate in the outdoor site. The SOX-related taxa involved in the sulfur cycle showed higher activity outdoors than indoors, and on the outdoor ground than at the outdoor cliff, as detected through metaproteomic analysis. The development of petrochemical industry in the vicinity resulting in the deposition of sulfur/oxidized sulfur via atmosphere may stimulate the physiological activity of SOX. Our findings provide metagenomic and metaproteomic evidence for microbially driven geobiochemical cycles that result in the biodeterioration of stone monuments.

Pt電極触媒のアンモニア酸化活性に及ぼすMoの添加効果

In recently years, hydrogen production by water electrolysis using renewable energy is promoted from a carbon-neutral perspective. However, hydrogen is difficult to store and transport, and low volumetric energy density. Therefore, ammonia obtained by the Haber-Bosch process is attracting much attention as an energy carrier because of its easy liquefaction and handling. Ammonia is toxic, but it can be detected easily if ammonia leaks outside owing to its characteristic smell. Thus, a direct ammonia fuel cells (DAFCs) that use ammonia as fuel is promising candidate as a new power generation system. Generally, Pt is used as anode catalyst for ammonia oxidation reaction in DAFC. But it is desired to develop an alternative catalyst for DAFC anode due to the price escalation of Pt, limitation of Pt reserves and the problems of inactivation caused by Nads poisoning on Pt. Therefore, Pt-Mo alloy catalyst was prepared by RF-magnetron sputtering and the ammonia oxidation activity was investigated in this study. Pt-Mo alloy catalysts showed higher ammonia oxidation activity than Pt one. Especially, Pt-44.0at％Mo indicated 31.4 mA cm−2 and it is the highest activity among Pt-Mo alloys. And then, Pt-44.0at％Mo is promising candidate for anode catalyst of DAFC.

Enhanced photocatalytic ammonia oxidation activity and nitrogen selectivity over Ag/AgCl/N-TiO2 photocatalyst

The removal of ammonia (NH3) emitted from agricultural and industrial activities is of great significance to protect human health and ecological environment. Photocatalytic NH3 oxidation to N2 under mild conditions is a promising strategy. However, developing visible light photocatalysts for NH3 oxidation is still in its infancy. Here, we fabricate N-TiO2 and Ag/AgCl/N-TiO2 photocatalysts by sol-gel and photodeposition methods, respectively. The introduction of N not only endows TiO2 with visible light response (absorption edge at 460 nm) but also results in the formation of heterophase junction (anatase and rutile). Thus, N-TiO2 shows 2.0 and 1.8 times higher than those over anatase TiO2 and commercial TiO2 for NH3 oxidation under full spectrum irradiation. Meanwhile, surface modification of Ag can simultaneously enhance visible light absorption (generating localized surface plasmon resonance effect) and charge separation efficiency. Therefore, the photocatalytic activity of Ag/AgCl/N-TiO2 is further improved. Furthermore, the presence of N and Ag also enhances the selectivity of N2 product owing to the change of reaction pathway. This work simultaneously regulates photocatalytic conversion efficiency and product selectivity, providing some guidance for developing highly efficient photocatalysts for NH3 elimination.

Shifts in soil nitrogen availability and associated microbial drivers during stand development of Mongolian pine plantations

AbstractAfforestation on degraded lands is an effective measure to control desertification and soil erosion, but these functions are often constrained by soil nitrogen (N) availability. Moreover, soil N availability usually shifts along plantation development, yet we have a limited understanding of factors driving this shift. Here, we examined dynamics of soil N mineralization and nitrification rates along plantation development, and explored mechanisms behind these dynamics from the perspective of N‐cycling microbes. We measured needle litter quality, N‐hydrolyzing enzyme activity, biomass and community composition of soil microbes (phospholipid fatty acid analysis) and ammonia oxidizers (real‐time quantitative PCR and high‐throughput sequencing), and net N mineralization and nitrification rates along a chronosequence of Pinus sylvestris var. mongolica stands with six age classes ranging from 15‐ to 61‐year‐old in the Three‐North region of China. Results showed that stand development increased net N mineralization rate by 54%, nitrate‐N concentration by 106%, and nitrate‐N:ammonium‐N ratio from 1.09 to 2.63. Gram‐positive:gram‐negative bacterial ratio and β‐N‐acetylglucosaminidase activity initially increased and subsequently decreased along stand development with the highest values in 40‐year‐old stands, while fungal:bacterial ratio showed the opposite pattern with the lowest value in 40‐year‐old stands. These shifts in soil microbial properties were associated with age‐related changes in needle litter C:N ratio and soil pH. Net N mineralization rate was positively related to β‐N‐acetylglucosaminidase activities, but not to soil microbial biomass and community composition. Net nitrification rate was negatively correlated with ammonia‐oxidizing archaeal abundances and positively with ammonia‐oxidizing bacterial abundances. Collectively, our results indicate that soil N availability increases and N cycling accelerates with plantation growth, and suggest that microbial N recycling driven by β‐N‐acetylglucosaminidase is tightly related to soil N dynamics along stand development.

Meticulous integration of N and C active sites in Ni2P electrocatalyst for sustainable ammonia oxidation and efficient hydrogen production

Ammonia, as an efficient hydrogen carrier, is emerging as an alternative energy resource to replace fossil fuels in the carbon–neutral era. Hydrogen production by water electrolysis seeks a lower potential dependent anodic reaction to overcome its energy-inefficiency that originates from the high potential anodic oxygen evolution reaction (OER). In this work, nickel phosphide supported on nitrogen doped-carbon (Ni2P@N-C) was prepared by one-pot synthesis for the bifunctional activity of hydrogen evolution (HER) and ammonia oxidation (AOR) reactions. The Ni2P@N-C electrocatalyst exhibits about 78% decomposition of ammonia compared to the initial concentration. The amount of hydrogen generated in a 0.5 M ammonia environment for 30 min is about 2.144 mmol(H2)/mol(NH3) h cm2. The fabricated ammonia electrolysis cell demonstrates a comparatively low energy consumption rate of 8.611 KWHkg(H2)−1. Thus, engineering a bifunctional electrocatalyst for a low potential anode oxidation reaction and the cathodic HER is a promising strategy to fabricate an energy efficient electrolysis cell for hydrogen production with a low cell potential requirement.

Mechanisms Driving the Distribution and Activity of Mineralization and Nitrification in the Reservoir Riparian Zone.

The riparian zone ecosystems have greater energy flow and elemental cycling than adjacent terrestrial and aquatic ecosystems. Mineralization and nitrification are important initiating processes in the nitrogen cycle, but their distribution and activity under different environmental conditions in the riparian zone and the driving mechanisms are still not clear. We investigated the effects of environmental and microbial factors on mineralization and nitrification activities by analyzing the community of alkaline (apr) and neutral (npr) metallopeptidase, ammonia-oxidizing archaea (AOA), and bacteria (AOB) in soils and sediments under different land-use types in the riparian zone of Miyun Reservoir, as well as measuring potential nitrogen mineralization and ammonia oxidation rates (AOR). The results showed that the mineralization and nitrification activities of soils were greater than those of sediments. AOA and AOB dominate the ammonia oxidation activity of soil and sediment, respectively. NH4+ content was a key factor influencing the ecological niche differentiation between AOA and AOB. The high carbon and nitrogen content of the woodland significantly increased mineralization and nitrification activity. Microbial communities were significantly clustered in the woodland. The land-use type, not the flooding condition, determined the distribution of microbial community structure. The diversity of npr was significantly correlated with potential N mineralization rates, while the transcript abundance of AOA was significantly correlated with ammonia oxidation rates. Our study suggests that environmental changes regulate the distribution and activity of mineralization and nitrification processes in the reservoir riparian zone by affecting the transcript abundance, diversity and community structure of the microbial functional genes.

Nitrification kinetics, N2O emission, and energy use in intermittently aerated hybrid reactor under different organic loading rates

This study investigated the impact of intermittent aeration strategies and reduction in the reactor’s organic and nitrogen loading rates on the course of particular stages of the nitrification process, as well as energy consumption and N2O emissions in a hybrid reactor with nitrification/denitrification. Each of the analysed series revealed the greatest ammonia oxidation activity in activated sludge flocs. The highest activity of nitrite nitrogen oxidation was demonstrated in the case of biofilm. A reduction in the reactor’s organic and nitrogen loading rate value had a greater effect on changes in the activity of ammonia-oxidizing bacteria than nitrite-oxidizing bacteria. In a system where the operation of air pumps was controlled through switching them and off according to the adopted ratio between non-aerated and aerated sub-phase times and the assumed oxygen concentration, a reduction in the duration of aerated sub-phases caused no decrease in energy use for aeration. Lower N2O emission was recorded when the reactor operated with a longer duration of aerated sub-phases.

Long-term effect of zero-valent iron on one-stage partial nitritation and anammox

Partial nitritation and anammox (PN/A) is an energy- and carbon-saving process for nitrogen removal from organic deficient wastewater. However, the PN/A effluent often requires polishing to eliminate the nitrate produced by anammox. Iron as the essential element for living organisms has been tested for nitrate reduction and phosphorus recovery in wastewater treatment processes. Here, we proposed a novel process coupling PN/A with iron-based denitrification for efficiently removing nitrogen and phosphorus in a single reactor. Two laboratory-scale bioreactors were operated under oxygen-limited conditions in parallel. Fe0 with a dosage of ∼ 27 mg Fe/L-wastewater was continuously dosed in the experimental reactor, with the other serving as a control. Throughout 300 days of operation, the results showed that experimental system always had significantly high removal efficiencies of organic carbon, nitrogen and phosphorus compared with control system. The total nitrogen removal achieved 90.1 ± 1.2 % at the rate of 0.96 ± 0.03 kg N/(m3•d) due to the improved activity of ammonia oxidizers, anammox bacteria and denitrifiers and enriched nitrogen conversion pathway (i.e., iron-based autotrophic denitrification) with Fe0 supplement. The total phosphorus removal was synchronously improved to 80.2 ± 9.1 % mainly driven by the phosphate precipitation with iron. This study provides new insights into achieving high-level nitrogen and phosphorus removal in a single reactor, and deepens our understanding of the improved nutrients removal mechanism mediated by iron.

Creating metal-carbide interactions to boost ammonia oxidation activity for low-temperature direct ammonia fuel cells

Low-temperature direct ammonia fuel cell (DAFC) can easily transform the chemical energy into green in-commission power, yet remains great challenging due to kinetically sluggish ammonia oxidation reaction (AOR). The interface engineering is a promising strategy to take advantage of synergistic proficiencies and electronic properties, which was widely applied in heterogeneous catalysis but rarely studied in AOR. Herein, non-oxide-based metal–metal carbide interfaces were employed to construct excellent AOR catalysts. The as-built Pt/WC and PtIr/WC catalysts with the excellent peak current densities of 96.5 and 61.7 A gPGM−1, respectively, which are over 3.5-fold and 2.7-fold than those of carbon-supported catalysts. Both experimental and theoretical studies reveal that the charge transfer between PtIr NPs and WC regulates the work function and d-band center of catalysts and further alters adsorption energies of intermediates. The WC also plays an important role for water dissociation and hydroxyl group activation. Based on the understanding, maximizing active interfaces by dispersing PtIr–WC composites onto CNTs (PtIr–WC/CNT) achieves a compelling peak power density of 140.0 mW cm−2 at 80 °C in a DAFC test, which is at the top of previously reported catalysts at similar conditions. Our study provides a way to design highly-active catalysts for AOR and DAFC by interface engineering.

Heterogeneous Electrochemical Ammonia Oxidation with a Ru-bda Oligomer Anchored on Graphitic Electrodes via CH-π Interactions.

Molecular catalysts can promote ammonia oxidation, providing mechanistic insights into the electrochemical N2 cycle for a carbon-free fuel economy. We report the ammonia oxidation activity of carbon anodes functionalized with the oligomer {[RuII(bda-κ-N 2 O 2)(4,4'-bpy)]10(4,4'-bpy)}, Rubda-10, where bda is [2,2'-bipyridine]-6,6'-dicarboxylate and 4,4'-bpy is 4,4'-bipyridine. Electrocatalytic studies in propylene carbonate demonstrate that the Ru-based hybrid anode used in a 3-electrode configuration transforms NH3 to N2 and H2 in a 1:3 ratio with near-unity faradaic efficiency at an applied potential of 0.1 V vs Fc+/0, reaching turnover numbers of 7500. X-ray absorption spectroscopic analysis after bulk electrolysis confirms the molecular integrity of the catalyst. Based on computational studies together with electrochemical evidence, ammonia nucleophilic attack is proposed as the primary pathway that leads to critical N-N bond formation.

Shifts of ammonia-oxidation process along salinity gradient in an estuarine wetland

Ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) play an important role in nitrogen (N) cycling of the estuarine sediments. However, the shifts of potential ammonia oxidation rates (PARs) of AOA and AOB along different salinity gradients are not well understood. Here, salinity-treated (0, 5, 15, 25 and 35 ‰) incubation experiments of sediment from the Pearl River Estuary were performed to explore the influence of salinity on ammonia oxidation process, including the PARs and gene abundances of AOA and AOB. Significant shifts of PARs and amoA gene abundances of ammonia oxidation process along salinity gradient were observed during the incubation (0–60 d). After a long-term incubation, both PARtotal (4.61 ± 0.93 μg N g−1 d−1) and PARaob (3.05 ± 1.04 μg N g−1 d−1) reached their maximum values at 25 ‰ salinity, and PARaoa (1.96 ± 0.10 μg N g−1 d−1) reached its maximum values at 35 ‰ salinity, which showed that PARs had higher values under high salinity conditions. AOB amoA gene was more abundant than AOA under moderate and high salinity, and only a significant linear correlation was observed between PARaob and AOB abundances. In addition, the ammonia oxidation activities showed a wide range of salinity tolerance, which is worthy of further study on community structure and transcript richness. Our study illustrates that salinity plays a key role in regulating the ammonia oxidation process in estuarine sediments, and has guiding significance for N fixation and N loss associated with saltwater incursion.

Removal of Mg2+ inhibition benefited the growth and isolation of ammonia-oxidizing bacteria: An inspiration from bacterial interaction.

Ammonia-oxidizing bacteria (AOB) play an important role in the global nitrogen cycle and have broad applications in the nitrogen removal from wastewater. However, the AOB species are sensitive to environmental factors and usually form tight relationships with other microbes, making the AOB isolation and maintenance are difficult and time-consuming. In this study, the relationship that occurred between AOB and their bacterial partners was found to be able to improve the ammonia oxidation; during the co-cultivation, the magnesium ions (Mg2+) with removal rate as high as 36.7% was removed from culture medium by the concomitant bacterial species, which was regarded as the main reason for improving ammonia oxidation. During the pure cultivation of AOB isolate, when the concentration of Mg2+ reduced to low levels, the ammonia-oxidizing activity was more than 5 times and the amoA gene expression was more than 12 times higher than that grown in the initial culture medium. Based on a newly designed culture medium, the ammonia oxidation of AOB isolate grown in liquid culture was significantly promoted and the visible AOB colonies with much more number and larger diameter were observed to form on agar plates. With the addition of high concentration of calcium carbonate (CaCO3), AOB colonies could be easily and specifically identified by following the hydrolytic zones that formed around AOB colonies. Another AOB isolates were successively obtained from different samples and within a short time, suggesting the feasibility and effectivity of this culture medium and strategy on the AOB isolation from environments.

Selective Enrichment of Comammox Nitrospira in a Moving Bed Biofilm Reactor with Sufficient Oxygen Supply.

The recent discovery of comammox (complete ammonia oxidation) Nitrospira has upended the long-held nitrification paradigm. Although comammox Nitrospira have been identified in wastewater treatment systems, the conditions for their dominance over canonical ammonia oxidizers remain unclear. Here, we report the dominance of comammox Nitrospira in a moving bed biofilm reactor (MBBR) fed with synthetic mainstream wastewater. Integrated 16S rRNA gene amplicon sequencing, fluorescence in situ hybridization (FISH), and metagenomic sequencing methods demonstrated the selective enrichment of comammox bacteria when the MBBR was operated at a dissolved oxygen (DO) concentration above 6 mg O2/L. The dominance of comammox Nitrospira over canonical ammonia oxidizers (i.e., Nitrosomonas) was attributed to the low residual ammonium concentration (0.02-0.52 mg N/L) formed in the high-DO MBBR. Two clade A comammox Nitrospira were identified, which are phylogenetically close to Candidatus Nitrospira nitrosa. Interestingly, cryosectioning-FISH showed these two comammox species spatially distributed on the surface of the biofilm. Moreover, the ammonia-oxidizing activity of comammox Nitrospira-dominated biofilms was susceptible to the oxygen supply, which dropped by half with the DO concentration decrease from 6 to 2 mg O2/L. These features collectively suggest a low apparent oxygen affinity for the comammox Nitrospira-dominated biofilms in the high-DO nitrifying MBBR.

Chlorate as a comammox Nitrospira specific inhibitor reveals nitrification and N2O production activity in coastal wetland

Specific nitrification inhibitors (NIs) have been widely used to disentangle the contribution of ammonia-oxidizing archaea (AOA), ammonia-oxidizing bacteria (AOB) as well as nitrite oxidizers to the nitrification process in specific environments. However, if these previously reported NIs can also be used to evaluate the activity of the newly discovered complete ammonia oxidizers (comammox) Nitrospira, remains understudied. Here we evaluated various NIs for their impact and specificity regarding inhibition of comammox Nitrospira in batch cultures of pure and mixed strains of AOA, AOB and comammox. Using these cultures, we observed that chlorate could specifically inhibit the ammonia oxidation and nitrite oxidation activity of comammox Nitrospira, while it had no effect on the tested AOA and AOB strains. This inhibitory effect of chlorate on comammox Nitrospira was subsequently confirmed based on 13CO2-DNA-stable isotope probing (13C-DNA-SIP) analysis. Furthermore, by applying a set of specific NIs, the nitrification and nitrous oxide (N2O) production rates of comammox Nitrospira in coastal wetlands were estimated as 17.45 ng N g−1 h−1 (26.9% of the total rate) and 0.0083 μmol−1 L h−1 (28.5%). Altogether, we identified and applied an effective and specific inhibitor of comammox Nitrospira, which allowed quantifying comammox activity in wetlands of the Yangtze Estuary, shedding new light on the ecological roles of comammox bacteria in coastal wetland environments.

Promotion of nitrogen removal in a zero-valent iron-mediated nitrogen removal system operated in co-substrate mode

In this study, the performance and mechanism of nitrogen removal were investigated in a zero-valent iron-mediated nitrogen removal system operated in co-substrate mode with sodium acetate as the organic carbon source. The results showed that the additional organic matter had the capacity to promote NH4+-N and total inorganic nitrogen (TIN) removal with efficiencies of 91.09% and 84.10%, and increases of 60.06% and 75.32% compared with the control group, respectively. The organic matter also stimulated the production of extracellular polymer substances that reduced the passivation and toxicity of iron to microorganisms. The ammonia oxidation activity was 2.5 times higher than that in the control group, and the anammonia oxidation activity and denitrification activity were substantially higher than in the control group with TIN removal efficiencies of 1.02 and 1.19 mgN/(gVSS·d), respectively. In addition, the organic matter increased the enrichment of the heterotrophic denitrification bacterium Diaphorobacter and facultative iron salt-based bacterium Dechloromonas.

Unraveling Nitrogen, Sulfur, and Carbon Metabolic Pathways and Microbial Community Transcriptional Responses to Substrate Deprivation and Toxicity Stresses in a Bioreactor Mimicking Anoxic Brackish Coastal Sediment Conditions

Microbial communities are key drivers of carbon, sulfur, and nitrogen cycling in coastal ecosystems, where they are subjected to dynamic shifts in substrate availability and exposure to toxic compounds. However, how these shifts affect microbial interactions and function is poorly understood. Unraveling such microbial community responses is key to understand their environmental distribution and resilience under current and future disturbances. Here, we used metagenomics and metatranscriptomics to investigate microbial community structure and transcriptional responses to prolonged ammonium deprivation, and sulfide and nitric oxide toxicity stresses in a controlled bioreactor system mimicking coastal sediment conditions. Ca. Nitrobium versatile, identified in this study as a sulfide-oxidizing denitrifier, became a rare community member upon ammonium removal. The ANaerobic Methanotroph (ANME) Ca. Methanoperedens nitroreducens showed remarkable resilience to both experimental conditions, dominating transcriptional activity of dissimilatory nitrate reduction to ammonium (DNRA). During the ammonium removal experiment, increased DNRA was unable to sustain anaerobic ammonium oxidation (anammox) activity. After ammonium was reintroduced, a novel anaerobic bacterial methanotroph species that we have named Ca. Methylomirabilis tolerans outcompeted Ca. Methylomirabilis lanthanidiphila, while the anammox Ca. Kuenenia stuttgartiensis outcompeted Ca. Scalindua rubra. At the end of the sulfide and nitric oxide experiment, a gammaproteobacterium affiliated to the family Thiohalobacteraceae was enriched and dominated transcriptional activity of sulfide:quinone oxidoreductases. Our results indicate that some community members could be more resilient to the tested experimental conditions than others, and that some community functions such as methane and sulfur oxidation coupled to denitrification can remain stable despite large shifts in microbial community structure. Further studies on complex bioreactor enrichments are required to elucidate coastal ecosystem responses to future disturbances.