Denitrifying Anaerobic Methane Oxidation Bacteria Research Articles

Effects of Antibiotics on the DAMO Process and Microbes in Cattle Manure.

Denitrifying anaerobic methane oxidation (DAMO) can mitigate methane emissions; however, this process has not been studied in cattle manure, an important source of methane emissions in animal agriculture. The objective of this study was to investigate the occurrence of DAMO microbes in cattle manure and examine the impacts of veterinary antibiotics on the DAMO process in cattle manure. Results show that DAMO archaea and bacteria consistently occur at high concentrations in beef cattle manure. During the long-term operation of a sequencing batch reactor seeded with beef cattle manure, the DAMO activities intensified, and DAMO microbial biomass increased. Exposure to chlortetracycline at initial concentrations up to 5000 μg L-1 did not inhibit DAMO activities or affect the concentrations of the 16S rRNA gene and functional genes of DAMO microbes. In contrast, exposure to tylosin at initial concentrations of 50 and 500 μg L-1 increased the activities of the DAMO microbes. An initial concentration of 5000 μg L-1 TYL almost entirely halted DAMO activities and reduced the concentrations of DAMO microbes. These results show the occurrence of DAMO microbes in cattle manure and reveal that elevated concentrations of dissolved antibiotics could inhibit the DAMO process, potentially affecting net methane emissions from cattle manure.

Microbial stratification protects denitrifying anaerobic methane oxidation archaea and bacteria from external oxygen shock in membrane biofilm reactor

Different gradients of dissolved oxygen (DO) regulate the microbial community and nitrogen removal pathways of denitrifying anaerobic methane oxidation (DAMO) and anaerobic ammonium oxidation (Anammox) coupled process in a batch biofilm reactor. Under completely anaerobic condition, approximately 72 mg NO3–-N/L was removed at a daily rate of 6.55 mg N/L, whereas a peak accumulation of 95 mg NO3–-N/L was observed during DO reached 0.5 mg/L. There is a decrease in the abundance of Candidatus Methylomirabilis (24.1%), Candidatus Methanoperedens (23.3%), and Candidatus Kuenenia (22.6%) to below 5% when DO levels reached 0.2 mg/L. Moreover, key genes associated with the reverse methanogenesis (mcrA) and anaerobic ammonium oxidase (hzo) decreased. These findings indicate that during oxygen shock, methanotrophs and denitrifiers replace Anammox bacteria on the outer sphere of the biofilm, whereas DAMO bacteria and archaea are protected from external oxygen shock due to the microbial stratification of biofilm.

Long-term effect of acetate and biochar addition on enrichment and activity of denitrifying anaerobic methane oxidation microbes

The denitrifying anaerobic methane oxidation (DAMO) process plays a crucial role in the global carbon/nitrogen cycles and methane emission control, and also has application potential in biological wastewater treatment. However, given that DAMO microbes are susceptible to external conditions such as additional carbon source in the system, it is essential to evaluate the effect of alternative carbon substance on the enrichment efficiency and metabolic activity of DAMO microbes. To this end, this study investigated the effect of acetate (0.1 mmol/L-R2, 0.5 mmol/L-R3) and biochar addition (R4) on the enrichment and activity of DAMO microbes. The long-term operation showed that the NO2−-N and CH4 consumption rates in the reactors almost presented the sequence of R4>R2>R3>R1. However, the short-term activity test with isotope labelling showed the sequence of R2>R4>R1>R3. Furthermore, the addition of acetate and biochar improved the electrochemical activity and extracellular polymeric substance (EPS) secretion in the systems. In R4 reactor, the proportion of DAMO bacteria was the highest (7.20%), indicating that the addition of biochar could promote the enrichment of DAMO bacteria, and Thauera was co-enriched with the proportion increasing from 0.26% to 6.73%. While in R1, R2 and R3 reactors, DAMO bacteria were enriched with relatively low abundances (0.10%, 0.23%, 0.15%, respectively), together with methanogens and denitrifiers. This study showed that biochar and acetate with appropriate concentration could enhance the enrichment and activity of DAMO bacteria, the results can provide reference for the enrichment of DAMO microbes and its application in the biological nitrogen removal of wastewater.

Niche differentiation of denitrifying anaerobic methane oxidation microbes in Taihu Lake of China

Denitrifying anaerobic methane oxidation (DAMO) microbes distribute widely and consume methane/nitrogen in freshwater environments. The survival preference of DAMO microbes directly affects their habitat preference, enabling them playing different roles in different environments. There are few studies regarding the preference and adaptability of DAMO microbes in the environment. In this work, the survival characteristics of DAMO microbes in Taihu Lake were studied. The relative abundance of DAMO archaea was relatively high in samples D7 and D8 of the moderately eutrophic region. The abundance of DAMO bacteria was 1.82 × 10 4 –6.11 × 10 6 copies g − 1 sediment, which was the highest in D1 of the mildly eutrophic region. The microbial association network analysis showed a significant difference in the connector number between the Moderate-NA (54 connectors) and Mild-NA (27 connectors), and DAMO bacteria played the connector role in the Mild-NA. The OTU numbers showing a positive and negative correlation with DAMO bacteria were 7 and 8 in Mild-NA, respectively, while 7 and 10 in Moderate-NA, showing a stronger competition relationship. For DAMO archaea, the corresponding OTUs in Mild-NA were 8 and 5, respectively, while 11 and 2 in Moderate-NA, showing a stronger cooperation association. DAMO microbes were differently affected by the antagonism of Rhodocyclaceae and Anammox bacteria. These results showed that DAMO bacteria and archaea had different survival strategies and displayed niche differentiation in the Taihu Lake. The results expanded our understanding of the ecological functions of DAMO microbes in freshwater and even global environment. • Niche differentiation of DAMO bacteria and archaea in Taihu Lake is evidenced. • DAMO bacteria get suitable niche and become key species in mildly eutrophic region. • DAMO archaea survive more suitably in the moderately eutrophic region. • Rhodocyclaceae and Anammox bacteria cause strong antagonism on DAMO microbes.

Response of denitrifying anaerobic methane oxidation enrichment to salinity stress: Process and microbiology

Denitrifying anaerobic methane oxidation (DAMO) is a novel biological process which could decrease nitrogen pollution and methane emission simultaneously in wastewater treatment. Salinity as a key environmental factor has important effects on microbial community and activity, however, it remains unclear for DAMO microorganisms. In this study, response of the enrichment of DAMO archaea and bacteria to different salinity was investigated from the aspect of process and microbiology. The results showed that the increasing salinity from 0.14% to 25% evidently deteriorated DAMO process, with the average removal rate of nitrate and methane decreased from 1.91 mg N/(L·d) to 0.07 mg N/(L·d) and 3.22 μmol/d to 0.59 μmol/d, respectively. The observed IC50 value of salinity on the DAMO culture was 1.73%. Further microbial analyses at the gene level suggested that the relative abundance of DAMO archaea in the enrichment decreased to 46%, 39%, 38% and 33% of the initial value. However, DAMO bacteria suffered less impact with the relative abundance maintaining over 75% of the initial value (except 1% salinity). In functional genes of DAMO bacteria, pmoA, decreased gradually from 100% to 86%, 43%, 15% and 2%, while mcrA (DAMO archaea) maintained at 67%–97%. This difference probably indicated DAMO bacteria appeared functional inhibition prior to community inhibition, which was opposite for the DAMO archaea. Results above-mentioned concluded that, though the process of nitrate-dependent anaerobic methane oxidation was driven by the couple of DAMO archaea and bacteria, they individually featured different response to high salinity stress. These findings could be helpful for the application of DAMO-based process in high salinity wastewater treatment, and also the understanding to DAMO microorganisms.

Rapid enrichment of denitrifying methanotrophs in a series hollow-fiber membrane biofilm reactor

Denitrifying anaerobic methane oxidation (DAMO) process uses methane as electron donor to reduce nitrate/nitrite to dinitrogen, which is a potentially efficient, low-cost and clean biological nitrogen removal technology. However, slow microbial growth rate severely limits the application of this promising process. In this study, a series hollow-fiber membrane biofilm reactor (HfMBR) was operated for 90 days to achieve rapid enrichment of these denitrifying methanotrophs. Finally, the highest relative abundance of denitrifying methanotrophic archaea and bacteria (DAMO archaea and bacteria) reached 47.5% and 11.3%, respectively. And the average abundance of DAMO archaea and bacteria increased 92.9 and 136.6 times respectively during the 90-day enrichment. High growth rate of DAMO archaea with a doubling time of 11.6 days was achieved in the second HfMBR according to quantitative PCR results. The results implied that dissolved oxygen would inhibit the growth of DAMO archaea, but the series HfMBR could effectively counteract this unfavorable factor. This work provided theoretical guidance for the rapid enrichment of denitrifying methanotrophs and contributed to the application of methane-dependent denitrification process.

Microbial dynamics and activity of denitrifying anaerobic methane oxidizers in China's estuarine and coastal wetlands

Estuarine and coastal wetlands, which act as large sources of methane (CH4) and undergo substantial loading of anthropogenic nitrogen (N), provide ideal conditions for denitrifying anaerobic methane oxidation (DAMO) to occur. Yet the microbial mechanisms governing DAMO and the main driving factors in estuarine and coastal ecosystems remain unclear. This study investigated the spatiotemporal distribution and associated activity of DAMO microorganisms along a wide swath of China's coastline (latitudinal range: 22–41°N) using molecular assays and isotope tracing techniques. We uncovered significant spatial and seasonal variation in DAMO bacterial community structure, whereas DAMO archaeal community structure exhibited no seasonal differences. The abundance of DAMO bacterial pmoA gene (2.2 × 105–1.0 × 107 copies g−1) was almost one order of magnitude higher than that of DAMO archaeal mcrA gene (8.7 × 104 –1.8 × 106 copies g−1). A significant positive correlation between pmoA and mcrA gene abundances (p < 0.01) was observed, indicating that DAMO bacteria and archaea may cooperate closely and thus complete nitrate elimination. Potential DAMO rates, in the range of 0.09–23.4 nmol 13CO2 g−1 day−1 for nitrite-DAMO and 0.03–43.7 nmol 13CO2 g−1 day−1 for nitrate-DAMO, tended to be greater in the relatively warmer low-latitudes. Potential DAMO rates were weakly positively correlated with gene abundances, suggesting that DAMO microbial activity could not be predicted directly by gene abundance alone. The heterogeneous variability of DAMO was shaped by interactions among key environmental characteristics (sediment texture, N availability, TOC, Fe3+, salinity of water, and temperature). On a broader continental scale, potential N removal rates of 0.1–11.2 g N m−2 yr−1 were estimated via nitrite-DAMO activity in China's coastal wetlands. Overall, our results highlight the widespread distribution of DAMO microbes and their potential role in eliminating excess N inputs and reducing CH4 emissions in estuarine and coastal ecosystems, which could help mitigate global warming.

Magnetite-mediated electrically connected community for shortening startup of methane-dependent denitrification in a membrane biofilm reactor

Denitrifying anaerobic methane oxidation (DAMO) not only mitigates methane emission from wastewater treatment, but compensates for the deficiency of current nitrogen removal process. However, the doubling time of DAMO microbes is usually more than two weeks, resulting in a long-period startup of a methane-dependent denitrification reactor especially with anaerobic sludge as inoculum. The study presented here found that, in the presence of magnetite, the nitrate removal rate reached 17.6 ± 0.4 mgNO3−-N/L/d after the 120-day experiments, which was approximately 2.4 folds higher than that in the absence of magnetite (7.3 ± 0.1 mgNO3−-N/L/d). The nitrate removal rate in the presence of magnetite reached 7–8 mgNO3−-N/L/d at day 80–85, which shortened the startup periods of approximately 35–40 days. Microbial morphology based on scanning electron microscope and transmission electron microscope showed that, the surface of cells was deposited by magnetite and they might be connected via magnetite. Further investigations via cyclic voltammetry and electrochemical impedance spectroscopy showed that magnetite mediated an electrically connected community with a higher capacitance and lower resistance than that in the absence of magnetite. Microbial community analysis showed that, in the presence of magnetite, the potential DAMO bacteria (Methylacidimicrobium and Methylotetracoccus species), methanogens (Methanothrix species) as well as nitrate-reducing bacteria (Bellilinea and Dechloromonas species) were enriched. They might participate in direct interspecies electron transfer (DIET) via magnetite substituting for c-type cytochromes as the electrical connection components, since methane oxidation in the presence of magnetite was insensitive to the external addition of potential intermediates, such as hydrogen.

Electrochemical system for anaerobic oxidation of methane by DAMO microbes with nitrite as an electron acceptor

Denitrifying anaerobic methane oxidation (DAMO) is an important microbial metabolic process that simultaneously converts of methane and nitrite. In this study, electrochemical systems were investigated for DAMO with nitrite as an electron acceptor. The results showed that the auxiliary voltage enhanced anaerobic methane oxidation and nitrite reduction. The greatest methane conversion (26.61 mg L−1 d−1) was obtained at an auxiliary voltage of 1.6 V (EMN-1.6). Isotope tracing indicated that carbon dioxide was the oxidation product of methane, and methanol was the intermediate. The power density reached 0.60 (for EMN-0.5, the bioreactor with a voltage of 0.5 V) and 3.77 mW m−2 (for EMN-1.6). DAMO microbes, Methylocystis sp., and Methylomonas sp. were identified as methanotrophs. Rhodococcus sp., Hyphomicrobium sp., and Thiobacillus sp. were the dominant denitrifying bacteria. The conversion pathway was speculated to be as follows: methane was oxidized to carbon dioxide and nitrite was reduced to nitrogen. The two processes were independently completed by DAMO bacteria and oxygen was simultaneously generated. For the electron transfer pathway, methanotrophs utilized the oxygen released by DAMO bacteria to convert methane into organic matter (e.g. methanol). These organic compounds were utilized by Pseudoxanthomonas sp. and Pseudomonas sp., and the generated electrons were then released to the outside of the cells and transferred to the anode. Denitrifying bacteria received electrons at the cathode, transferred them to the interior of the cell, and then converted nitrite into nitrogen. This research explored an effective consortium and a method for methane and nitrogen removal.

Niche differentiation of denitrifying anaerobic methane oxidizing bacteria and archaea leads to effective methane filtration in a Tibetan alpine wetland

Denitrifying anaerobic methane oxidation (DAMO) is a vital methane sink in wetlands. However, the interactions and niche partitioning of DAMO bacteria and archaea in freshwater wetland soils, in addition to the interactions among microorganisms that couple methane and nitrogen cycling is still unclear, despite that these factors may govern the fate of methane and nitrogen in wetlands. Here, we evaluated the vertical distribution of DAMO bacteria and archaea in soil layers along with the potential interactions among populations in the methane-coupled nitrogen cycling microbial community of Tibetan freshwater wetlands. A combination of molecular biology, stable isotope tracer technology, and microbial bioinformatics was used to evaluate these interrelated dynamics. The abundances and potential methane oxidation rates indicated that DAMO bacteria and archaea differentially occupy surface and subsurface soil layers, respectively. The inferred interactions between DAMO bacteria and nitrogen cycling microorganisms within their communities are complex, DAMO bacteria apparently achieve an advantage in the highly competitive environment of surface soils layers and occupy a specific niche in those environments. Conversely, the apparent relationships between DAMO archaea and nitrogen cycling microorganisms are relatively simple, wherein high levels of cooperation are inferred between DAMO archaea and nitrate-producing organisms in subsurface soils layers. These results suggest that the vertical distribution patterns of DAMO bacteria and archaea enable them to play significant roles in the methane oxidation activity of different soil layers and collectively form an effective methane filtration consortium.

Denitrifying Anaerobic Methane Oxidation and Anammox Process in a Membrane Aerated Membrane Bioreactor: Kinetic Evaluation and Optimization

Denitrifying anaerobic methane oxidation (DAMO) coupled to anaerobic ammonium oxidation (anammox) is a promising technology for complete nitrogen removal with economic and environmental benefit. In this work, a model framework integrating DAMO and anammox process was constructed based on suspended-growth systems. The proposed model was calibrated and validated using experimental data from a sequencing batch reactor and a membrane aerated membrane bioreactor (MAMBR). The model managed to describe removal rates of ammonium (NH4+), nitrite (NO2-), and total nitrogen, as well as biomass changes of DAMO archaea, DAMO bacteria, and anaerobic ammonium oxidizing bacteria (AnAOB) in both reactors. The estimated parameter values revealed that DAMO archaea possessed properties of faster growth and higher biomass yield in suspended-growth systems compared to those in attached-growth systems (e.g., biofilm). Model simulation demonstrated that solid retention time (SRT) was effective in washing out DAMO bacteria, but retaining DAMO archaea and AnAOB in the MAMBR. The optimal SRT and nitritation efficiency (the ratio of the NO2- to the sum of NH4+ and NO2- in the MAMBR influent) were simulated so that 99% of total nitrogen was removed to meet the discharge standard. MAMBR further suggested to be operated with SRT between 15 and 30 days so that the optimal nitritation efficiency could be minimized to 49% for cost savings.

Denitrifying anaerobic methane oxidation in marsh sediments of Chongming eastern intertidal flat.

Denitrifying anaerobic methane oxidation (DAMO) and associated microbial diversity and abundance in the marsh sediments of Chongming eastern intertidal flat, the Yangtze Estuary, were investigated using carbon-isotope tracing and molecular techniques. Co-existence of nitrate-DAMO archaea and nitrite-DAMO bacteria was evidenced, with higher biodiversity of DAMO archaea than DAMO bacteria. Abundance of DAMO archaeal mcrA gene and DAMO bacterial pmoA gene ranged from 4.2 × 103 to 3.9 × 1010 copies g-1 and from 4.5 × 105 to 6.4 × 106 copies g-1, respectively. High DAMO potential was detected, ranging from 0.6 to 46.7 nmol 13CO2 g-1 day-1 for nitrate-DAMO and from 1.3 to 39.9 nmol 13CO2 g-1 day-1 for nitrite-DAMO. In addition to playing an important role as a CH4 sink, DAMO bacteria also removed a substantial amount of reactive nitrogen (29.4 nmol N g-1 day-1) from the intertidal sediments. Overall, these results indicate the importance of DAMO bioprocess as methane and nitrate sinks in intertidal marshes.

Denitrifying anaerobic methane oxidation in intertidal marsh soils: Occurrence and environmental significance

Denitrifying anaerobic methane oxidation (DAMO), consisting of nitrite-dependent DAMO (nitrite-DAMO) and nitrate-dependent DAMO (nitrate-DAMO), has recently been discovered and considered an important link between carbon and nitrogen cycles. In this work, we investigated biodiversity, abundance, and potential methane (CH4) oxidation activity of nitrite-DAMO bacteria and nitrate-DAMO archaea in intertidal marsh soil cores, based on molecular and stable isotope tracing methods. Results evidenced the co-occurrence and vertical stratification of DAMO bacteria and archaea in intertidal marsh soils, with higher biodiversity of DAMO archaea compared with DAMO bacteria. The abundance of DAMO bacterial pmoA gene (8.2 × 105–3.0 × 107 copies g−1 dry soil) was approximate to that of DAMO archaeal mcrA gene (3.0 × 105–3.9 × 107 copies g−1 dry soil) in the intertidal marsh soil cores. Stable isotope experiments showed that DAMO bacteria and archaea were both active in the intertidal marshes, with CH4 oxidation potential of 0.1–3.8 nmol 13CO2 g−1 dry soil day−1 and 0.1–4.1 nmol 13CO2 g−1 dry soil day−1, respectively. The relative importance (including the abundance and CH4 oxidation activity) of DAMO bacteria and archaea showed significant vertical variations, with more contribution by DAMO bacteria at the soil-tidal water interface and soil-groundwater interface layers of the soil cores, which are largely due to the influence of tidal dynamics. In addition to playing an important role as CH4 sink, DAMO process was also a non-negligible pathway of nitrogen removal in intertidal marsh soils, with an estimated nitrogen removal rate of 0.4–10.1 nmol N g−1 dry soil day−1. Overall, these results illustrated the occurrence and environmental significance of DAMO bioprocess in intertidal marshes.

Response mechanism of denitrifying anaerobic methane oxidation microorganisms to ammonia

Environmental contextDenitrifying anaerobic methane oxidation (DAMO) is a new process in wastewater treatment with the potential to provide cheap and sustainable development. To better apply this technology to the large scale, we studied the response mechanism of DAMO microorganisms to ammonia, the main form of nitrogen in the nitrogenous wastewater. The results can provide a theoretical basis for the stable and efficient operation of DAMO processes. AbstractThe dominant microorganisms in the denitrifying anaerobic methane oxidation (DAMO) process are primarily DAMO bacteria and DAMO archaea, which can simultaneously realise methane oxidation and denitrification. Ammonia is the primary form of nitrogen found in wastewater. This study focuses on a coexistence system that contains both DAMO bacteria and DAMO archaea (DAMO co-system). The short- and long-term effects of NH4+-N on the DAMO co-system were investigated at both the macro level (such as denitrification performance) and the micro level (such as microbial structure and community). Short-term experimental studies demonstrated that the safe concentration of ammonia for this system was 250mgNL−1. When the ammonia concentration was 500mgNL−1, the nitrogen removal efficiency was significantly inhibited. With an increase in concentration and an extension of time, the inhibitory effect of ammonia was enhanced. Long-term experimental studies showed that the nitrogen removal performance of DAMO was completely inhibited when the ammonia concentration reached 1000mgNL−1 and that ammonia had a toxic accumulation effect on the DAMO co-system. The results of the pH experimental study demonstrated that free ammonia (FA) was the limiting factor in the alkaline condition, while ionised NH4+ was the limiting factor in neutral and acidic conditions. Scanning electron microscopy (SEM) demonstrated that the microbes in the DAMO co-system shrank after short-term exposure and that the microorganisms shrank in the shape of polygons. High-throughput sequencing analysis demonstrated that the community structure of the DAMO co-system changed substantially, and the species diversity and abundance decreased distinctly after long-term inhibition. A genus analysis indicated that the reduction in Nitrospirae may be an internal reason for the decrease in the denitrification performance of the DAMO co-system.

The short- and long-term effects of nitrite on denitrifying anaerobic methane oxidation (DAMO) organisms.

The denitrifying anaerobic methane oxidation (DAMO) process can achieve methane oxidation and denitrification at the same time by using nitrate or nitrite as an electron acceptor. The short- and long-term effects of nitrite on DAMO organisms were studied from macro (such as denitrification) to micro (such as microbial structure and community) based on two types of DAMO microbial systems. The results showed that the inhibitory effects of nitrite on the two DAMO microbial systems increased with rising concentration and prolonged time. In the short-term inhibitory phase, nitrite with concentrations below 100mgNL-1 did not inhibit the two distinct DAMO enrichments. When nitrite concentration was increased to 950mgNL-1, the nitrogen removal performance was completely inhibited. However, in the long-term inhibition experiment, when nitrite concentration was increased to 650mgNL-1, the nitrogen removal performance was completely inhibited. In addition, in acidic conditions, the real inhibitor of nitrite is FNA (free nitrous acid), while in alkaline conditions, the real inhibitor is the ionized form of nitrite. By using high-throughput sequencing technology, the species abundance and diversity of the two DAMO microbial systems showed an apparent decrease after long-term inhibition, and the community structure also changed significantly. For the enrichment culture dominated by DAMO bacteria, the substantial drop of Methylomonas may be the internal cause of the decreased nitrogen removal rate, and for the coexistence system that hosted both DAMO bacteria and archaea, the reduction of Nitrospirae may be an internal reason for the decline of the denitrification rate.

Denitrifying Anaerobic Methane Oxidation: A Previously Overlooked Methane Sink in Intertidal Zone.

The intertidal zone is an open ecosystem rich in organic matter and plays an important role in global biogeochemical cycles. It was previously considered that methane was mainly removed by sulfate-dependent anaerobic methane oxidation (sulfate-AOM) process in marine ecosystems while other anaerobic methane oxidation processes were ignored. Recent researches have demonstrated that denitrifying anaerobic methane oxidation (DAMO), consisting of nitrite-dependent anaerobic methane oxidation (nitrite-AOM) and nitrate-dependent anaerobic methane oxidation (nitrate-AOM), can also oxidize methane. In this work, the community structure, quantity and potential methane oxidizing rate of DAMO archaea and bacteria in the intertidal zone were studied by high-throughput sequencing, qPCR and stable isotope tracing method. The results showed that nitrate-AOM and nitrite-AOM were both active in the intertidal zone and showed approximate methane oxidation rates. The copy number of 16S rRNA gene of DAMO archaea and DAMO bacteria were 104 ∼ 105 copies g-1 (dry sediment), whereas NC10 bacteria were slightly higher. The contribution rate of DAMO process to total anaerobic methane removal in the intertidal zone reached 65.6% ∼ 100%, which indicates that DAMO process is an important methane sink in intertidal ecosystem. Laboratory incubations also indicated that DAMO archaea were more sensitive to oxygen and preferred a more anoxic environment. These results help us draw a more complete picture of methane and nitrogen cycles in natural habitats.

Effect of methane partial pressure on the performance of a membrane biofilm reactor coupling methane-dependent denitrification and anammox

Complete nitrogen removal has recently been demonstrated by integrating anaerobic ammonium oxidation (anammox) and denitrifying anaerobic methane oxidation (DAMO) processes. In this work, the effect of methane partial pressure on the performance of a membrane biofilm reactor (MBfR) consisting of DAMO and anammox microorganisms was evaluated. The activities of DAMO archaea and DAMO bacteria in the biofilm increased significantly with increased methane partial pressure, from 367 ± 9 and 58 ± 22 mg-N L−1d−1 to 580 ± 12 and 222 ± 22 mg-N L−1d−1, respectively, while the activity of anammox bacteria only increased slightly, when the methane partial pressure was elevated from 0.24 to 1.39 atm in the short-term batch tests. The results were supported by a long-term (seven weeks) continuous test, when the methane partial pressure was dropped from 1.39 to 0.78 atm. The methane utilization efficiency was always above 96% during both short-term and long-term tests. Taken together, nitrogen removal rate (especially the nitrate reduction rate by DAMO archaea) and methane utilization efficiency could be maintained at high levels in a broad range of methane partial pressure (0.24–1.39 atm in this study). In addition, a previously established DAMO/anammox biofilm model was used to analyze the experimental data. The observed impacts of methane partial pressure on biofilm activity were well explained by the modeling results. These results suggest that methane partial pressure can potentially be used as a manipulated variable to control reaction rates, ultimately to maintain high nitrogen removal efficiency, according to nitrogen loading rate.

The content of trace element iron is a key factor for competition between anaerobic ammonium oxidation and methane-dependent denitrification processes

Coupling of anaerobic ammonium oxidation (Anammox) with denitrifying anaerobic methane oxidation (DAMO) is a sustainable pathway for nitrogen removal and reducing methane emissions from wastewater treatment processes. However, studies on the competitive relation between Anammox bacteria and DAMO bacteria are limited. Here, we investigated the effects of variations in the contents of trace element iron on Anammox and DAMO microorganisms. The short-term results indicated that optimal concentrations of iron, which obviously stimulated the activity of Amammox bacteria, DAMO bacteria and DAMO archaea, were 80, 20, and 80 μM, respectively. The activity of Amammox bacteria increased more significant than DAMO bacteria with increasing contents of trace element iron. After long-term incubation with high content of trace element iron of 160 μM in the medium, Candidatus Brocadia (Amammox bacteria) outcompeted Candidatus Methylomirabilis oxyfera (DAMO bacteria), and ANME-2d (DAMO archaea) remarkably increased in number and dominated the co-culture systems (64.5%). Meanwhile, with further addition of iron, the removal rate of ammonium and nitrate increased by 13.6 and 9.2 times, respectively, when compared with that noted in the control. As far as we know, this study is the first to explore the important role of trace element iron contents in the competition between Anammox bacteria and DAMO bacteria and further enrichment of DAMO archaea by regulating the contents of trace element iron.

Achieving high-level nitrogen removal in mainstream by coupling anammox with denitrifying anaerobic methane oxidation in a membrane biofilm reactor.

To achieve energy neutral wastewater treatment, mainstream anaerobic ammonium oxidation (anammox) has attracted extensive attention in the past decade. However, the relatively high effluent nitrogen concentration (>10 mg N L-1) remains a significant barrier hindering its practical implementation. A novel technology integrating the anammox and denitrifying anaerobic methane oxidation (DAMO) reactions in a membrane biofilm reactor (MBfR) was developed in this study to enhance the mainstream anammox process. With the hydraulic retention time (HRT) progressively decreased from 12 to 4 h, the total nitrogen (TN) removal rate increased stepwise from 0.09 to 0.28 kg N m-3 d-1, with an effluent TN concentration below 3.0 mg N L-1 achieved. Mass balance analysis showed that 30-60% of the nitrate produced by the anammox reaction was reduced back to nitrite by DAMO archaea, and the anammox and DAMO bacteria were jointly responsible for nitrite removal with contributions of >90% and <10%, respectively. Additionally, the established MBfR was robust and achieved consistently high effluent quality with >90% TN removal when the influent nitrite to ammonium molar ratio varied in the range of 1.17-1.55. Fluorescence in situ hybridization (FISH) and 16S rRNA gene sequencing indicated that anammox bacteria, DAMO bacteria and DAMO archaea jointly dominated the biofilm, and were likely the key contributors to nitrogen removal. This is the first study that a high nitrogen removal rate (>0.2 kg N m-3 d-1) and satisfactory effluent quality (∼3 mg TN L-1) were achieved simultaneously by integrating anammox and DAMO reactions in mainstream wastewater treatment.

Co-existence of Anaerobic Ammonium Oxidation Bacteria and Denitrifying Anaerobic Methane Oxidation Bacteria in Sewage Sludge: Community Diversity and Seasonal Dynamics.

Anaerobic ammonium oxidation (ANAMMOX) and denitrifying anaerobic methane oxidation (DAMO) have been recently discovered as relevant processes in the carbon and nitrogen cycles of wastewater treatment plants. In this study, the seasonal dynamics of ANAMMOX and DAMO bacterial community structures and their abundance in sewage sludge collected from wastewater treatment plants were analysed. Results indicated that ANAMMOX and DAMO bacteria co-existed in sewage sludge in different seasons and their abundance was positively correlated (P<0.05). The high abundance of ANAMMOX and DAMO bacteria in autumn and winter indicated that these seasons were the preferred time to favour the growth of ANAMMOX and DAMO bacteria. The community structure of ANNAMOX and DAMO bacteria could also shift with seasonal changes. The "Candidatus Brocadia" genus of ANAMMOX bacteria was mainly recovered in spring and summer, and an unknown cluster was primarily detected in autumn and winter. Similar patterns of seasonal variation in the community structure of DAMO bacteria were also observed. Group B was the dominant in spring and summer, whereas in autumn and winter, group A and group B presented almost the same proportion. The redundancy analysis revealed that pH and nitrate were the most significant factors affecting community structures of these two groups (P<0.01). This study reported the diversity of ANAMMOX and DAMO in wastewater treatment plants that may be the basis for new nitrogen removal technologies.