Denitrifying Anaerobic Methane Oxidation Research Articles

The Effects of Model Insoluble Copper Compounds in a Sedimentary Environment on Denitrifying Anaerobic Methane Oxidation (DAMO) Enrichment.

The contribution of denitrifying anaerobic methane oxidation (DAMO) as a methane sink across different habitats, especially those affected by anthropogenic activities, remains unclear. Mining and industrial and domestic use of metals/metal-containing compounds can all cause metal contamination in freshwater ecosystems. Precipitation of metal ions often limits their toxicity to local microorganisms, yet microbial activity may also cause the redissolution of various precipitates. In contrast to most other studies that apply soluble metal compounds, this study investigated the responses of enriched DAMO culture to model insoluble copper compounds, malachite and covellite, in simulated sedimentary environments. Copper ≤ 0.22 µm from covellite appeared to cause immediate inhibition in 10 h. Long-term tests (54 days) showed that apparent methane consumption was less impacted by various levels of malachite and covellite than soluble copper. However, the medium-/high-level malachite and covellite caused a 46.6-77.4% decline in denitrification and also induced significant death of the representative DAMO microorganisms. Some enriched species, such as Methylobacter tundripaludum, may have conducted DAMO or they may have oxidized methane aerobically using oxygen released by DAMO bacteria. Quantitative polymerase chain reaction analysis suggests that Candidatus Methanoperedens spp. were less affected by covellite as compared to malachite while Candidatus Methylomirabilis spp. responded similarly to the two compounds. Under the stress induced by copper, DAMO archaea, Planctomycetes spp. or Phenylobacterium spp. synthesized PHA/PHB-like compounds, rendering incomplete methane oxidation. Overall, the findings suggest that while DAMO activity may persist in ecosystems previously exposed to copper pollution, long-term methane abatement capability may be impaired due to a shift of the microbial community or the inhibition of representative DAMO microorganisms.

Methane-driven microbial nitrous oxide production and reduction in denitrifying anaerobic methane oxidation cultures

Methane (CH4) and nitrous oxide (N2O) are two important greenhouse gases (GHGs), and it is very important to achieve the reduction of both gases in the case of increasingly serious climate change. The study confirms the simultaneous mitigation of potent GHGs N2O and CH4 in denitrifying anaerobic methane oxidation (DAMO) enrichment, supplementing the understanding that DAMO microorganisms do not produce and reduce N2O. We observed rapid accumulation and consumption of N2O in DAMO cultures subjected to high nitrate and nitrite loadings. Moreover, when N2O served as the sole electron acceptor, a significant correlation was found between the flux of N2O reduction and CH4 oxidation. Inhibition of the particulate methane monooxygenase (pMMO) led to a notable decrease of 95% in N2O reduction and 50% in CH4 oxidation. Metagenomic analysis revealed that Candidatus Methanoperedens and Candidatus Methylomirabilis were the dominant genera, with all functional genes related to denitrification and methane oxidation identified, including NO reductase (nor) of Ca. Methylomirabilis. Transcriptomic analysis further confirmed that the transcripts of nor were one order of magnitude higher than other functional genes. These results indicate that when the substrate NO2- surpasses the dismutation capacity of NO dismutase (nod), DAMO bacteria can alternatively induce nor to reduce NO to N2O. However, considering that N2O reductase (nosZ) is still missing in Ca. Methylomirabilis, the partner Methyloversatilis may be responsible for N2O reduction. Our results provide new insights into the link between GHG emissions in DAMO systems under anaerobic conditions.

Denitrifying Anaerobic Methane Oxidation Activity and Microbial Mechanisms in Riparian Zone Soils of the Yulin River, a Tributary of the Three Gorges Reservoir

Riparian zones are recognized as major sources of greenhouse gas emissions, particularly methane (CH4). Denitrifying anaerobic methane oxidation (DAMO) has garnered growing attention due to its significant contribution to mitigating CH4 emissions in wetland environments. Nonetheless, the specific role and microbial mechanisms of DAMO in controlling CH4 release within riparian zones are still not well comprehended. This study employed isotopic labeling experiments to measure the nitrate-dependent anaerobic methane oxidation (NaDAMO) and nitrite-dependent anaerobic methane oxidation (NiDAMO) potential of soil samples from riparian zones that were collected during different hydrological cycles. Moreover, soil physicochemical properties, DAMO activity, and microbial abundance were integrated to analyze the key factors and mechanisms influencing DAMO in riparian zone soils. The isotope tracer results showed that NaDAMO activities (1.4–11.93 nmol 13CO2 g-1day-1) were significantly higher than NiDAMO activities (0.66–9.19 nmol 13CO2 g-1day-1) in the riparian zone (p<0.05). NiDAMO activities were more strongly influenced by hydrological variations compared to NaDAMO activities, exhibiting higher levels during the discharge period (2.78–9.19 nmol 13CO2 g-1day-1) compared to the impoundment period (0.66–4.10 nmol 13CO2 g-1day-1). The qPCR analysis showed that the gene copies of NaDAMO archaeal mcrA (107 copies g-1) were approximately ten times greater than those of NiDAMO bacterial pmoA (106 copies g-1) in the majority of the sampling sites. Correlation analyses revealed that NiDAMO activity was influenced by soil pH (p<0.05), while NaDAMO microbes were influenced by temperature, organic carbon, and ammonia nitrogen concentrations (p<0.05). In summary, this research explored how hydrological changes in the riparian zone influence DAMO activities and their underlying mechanisms, providing a theoretical basis for mitigating CH4 emissions in riparian zones of reservoir regions.

Biochar modulates intracellular electron transfer for nitrate reduction in denitrifying anaerobic methane oxidizing archaea

Denitrifying anaerobic methane oxidizing (DAMO) archaea plays a significant role in simultaneously nitrogen removal and methane mitigation, yet its limited metabolic activity hinders engineering applications. This study employed biochar to explore its potential for enhancing the metabolic activity and nitrate reduction capacity of DAMO microorganisms. Sawdust biochar (7 g/L) was found to increase the nitrate reduction rate by 2.85 times, although it did not affect the nitrite reduction rate individually. Scanning electron microscopy (SEM) and fluorescence excitation-emission matrix (EEM) analyses revealed that biochar promoted microbial aggregation, and stimulated the secretion of extracellular polymeric substances (EPS). Moreover, biochar bolstered the redox capacity and conductivity of the biofilm, notably enhancing the activity of the electron transfer system by 1.65 times. Key genes involved in intracellular electron transport (Hdr, MHC, Rnf) and membrane transport proteins (BBP, ABC, NDH) of archaea were significantly up-regulated. These findings suggest that biochar regulates electrons generated by reverse methanogenesis to the membrane for nitrate reduction.

Simultaneous methane mitigation and nitrogen removal by denitrifying anaerobic methane oxidation in lake sediments

Methane (CH4) is a potent greenhouse gas, with lake ecosystems significantly contributing to its global emissions. Denitrifying anaerobic methane oxidation (DAMO) process, mediated by NC10 bacteria and ANME-2d archaea, links global carbon and nitrogen cycles. However, their potential roles in mitigating methane emissions and removing nitrogen from lake ecosystems remain unclear. This study explored the spatial variations in activities of nitrite- and nitrate-DAMO and their functional microbes in Changdanghu Lake sediments (Jiangsu Province, China). The results showed that although the average abundance of ANME-2d archaea (5.0 × 106 copies g−1) was significantly higher than that of NC10 bacteria (2.1 × 106 copies g−1), the average potential rates of nitrite-DAMO (4.59 nmol 13CO2 g−1 d−1) and nitrate-DAMO (5.01 nmol 13CO2 g−1 d−1) showed no significant difference across all sampling sites. It is estimated that nitrite- and nitrate-DAMO consumed approximately 6.46 and 7.05 mg CH4 m−2 d−1, respectively, which accordingly achieved 15.07–24.95 mg m−2 d−1 nitrogen removal from the studied lake sediments. Statistical analyses found that nitrite- and nitrate-DAMO activities were both significantly related to sediment nitrate contents and ANME-2d archaeal abundance. In addition, NC10 bacterial and ANME-2d archaeal community compositions showed significant correlations with sediment organic carbon content and water depth. Overall, this study underscores the dual roles of nitrite- and nitrate-DAMO processes in CH4 mitigation and nitrogen elimination and their key environmental impact factors (sediment organic carbon and inorganic nitrogen contents, and water depth) in shallow lake, enhancing the understanding of carbon and nitrogen cycles in freshwater aquatic ecosystems.

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.

The diversification and potential function of microbiome in sediment-water interface of methane seeps in South China Sea.

The sediment-water interfaces of cold seeps play important roles in nutrient transportation between seafloor and deep-water column. Microorganisms are the key actors of biogeochemical processes in this interface. However, the knowledge of the microbiome in this interface are limited. Here we studied the microbial diversity and potential metabolic functions by 16S rRNA gene amplicon sequencing at sediment-water interface of two active cold seeps in the northern slope of South China Sea, Lingshui and Site F cold seeps. The microbial diversity and potential functions in the two cold seeps are obviously different. The microbial diversity of Lingshui interface areas, is found to be relatively low. Microbes associated with methane consumption are enriched, possibly due to the large and continuous eruptions of methane fluids. Methane consumption is mainly mediated by aerobic oxidation and denitrifying anaerobic methane oxidation (DAMO). The microbial diversity in Site F is higher than Lingshui. Fluids from seepage of Site F are mitigated by methanotrophic bacteria at the cyclical oxic-hypoxic fluctuating interface where intense redox cycling of carbon, sulfur, and nitrogen compounds occurs. The primary modes of microbial methane consumption are aerobic methane oxidation, along with DAMO, sulfate-dependent anaerobic methane oxidation (SAMO). To sum up, anaerobic oxidation of methane (AOM) may be underestimated in cold seep interface microenvironments. Our findings highlight the significance of AOM and interdependence between microorganisms and their environments in the interface microenvironments, providing insights into the biogeochemical processes that govern these unique ecological systems.

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.

Simultaneous denitrification and electricity generation in a methane-powered bioelectrochemical system.

Bioelectrochemical system is a novel method for controlling down nitrate pollution, yet the feasibility of using methane as the electron donors for denitrification in this system remains unknown. In this study, using the effluent from mother BESs as inocula, a denitrifying anaerobic methane oxidation bioelectrochemical system was successfully started up in 92 days. When operated with 50 mmol/L phosphate buffer solution at pH7 and 30°C, the maximum methane consumption, nitrate, and total nitrogen removal load reached 0.23 ± 0.01 mmol/d, 551.0 ± 22.1mg N/m3 /d, and 64.0 ± 18.8mg N/m3 /d, respectively. Meanwhile, the peak voltage of 93 ± 4mV, the anodic coulombic efficiency of 6.99 ± 0.20%, and the maximum power density of 219.86 mW/m3 were obtained. The metagenomics profiles revealed that the dominant denitrifying bacteria in the cathodic chamber reduced most nitrate to nitrite through denitrification and assimilatory reduction. In the anodic chamber, various archaea including methanotrophs and methanogens converted methane via reverse methanogenesis to form formate (or H2 ), acetate, and methyl compounds, which were than utilized by electroactive bacteria to generate electricity. PRACTITIONER POINTS: A denitrifying anaerobic methane oxidation BES was successfully started up in 92 d. Simultaneous removal of methane and nitrate was achieved in the DAMO-BES. Functional genes related to AMO and denitrification were detected in the DAMO-BES. Methylocystis can mediate AMO in the anode and denitrification in the cathode.

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.

Enhancement of nitrogen removal in coupling Anammox and DAMO via Fe-modified granular activated carbon

Anaerobic ammonium oxidation (Anammox) coupled with Denitrifying anaerobic methane oxidation (DAMO) is an attractive technology to simultaneously remove nitrogen and mitigate methane emissions from wastewater. However, its nitrogen removal rate is usually limited due to the low methane mass transfer efficiency, low metabolic activity and slow growth rate of functional microorganisms. In this study, GAC and Fe-modified GAC (Fe-GAC) were added into Anammox-DAMO process to investigate their effects on nitrogen removal rates and then reveal the mechanism. The results showed that after 80-day experiments, the total nitrogen removal rate was slightly improved in the presence of GAC (3.94 mg L−1·d−1), while it reached high as 16.66 mg L−1·d−1 in the presence of Fe-GAC, which was ca.17 times that of non-amended control group (0.96 mg L−1·d−1). The addition of Fe-GAC stimulated the secretion of extracellular polymeric substance (EPS), improved the electron transfer capability and promoted the production of Cytochrome C. Besides, the key functional enzyme activities (HZS, HDH and NAR) were highest in the Fe-GAC group, which were approximately 1.06–1.56 times higher than those of GAC-amended and blank control groups. Microbial community analysis showed that the abundance of the DAMO archaea (Candidatus Methanoperedens) and Anammox bacteria (Candidatus Brocadia) were remarkably increased with the addition of Fe-GAC. Functional genes associated with nitrogen removal and methane oxidation in Fe-GAC system were up-regulated. This study provides a promising strategy for achieving high rate of nitrogen removal upon Anammox-DAMO process.

Electro-stimulated anaerobic oxidation of methane with synergistic denitrification by adding AQS: Electron transfer mode and mechanism

Denitrifying anaerobic methane-oxidizing (DAMO) processes, which link anaerobic methane oxidation (AMO) and denitrification, have a promising prospect in anaerobic wastewater treatment. In bioelectrochemical systems (BES), DAMO consortium presented potent metabolic activity. However, the extracellular electron transfer (EET) in BES was poorly understood. This study investigated the EET mechanisms and modes of electron transport in BES dominated by anaerobic methanotrophic bacteria. In the bioreactors with the auxiliary voltage of 0.5 and 1.1 V, named EMN-0.5 and EMN-1.1, respectively, biological voltages of 0.198 and 0.329 V were generated with power densities of 0.6 and 1.20 mW/m2, after removing the voltage. High throughput and metagenome analyses demonstrated that main methanotrophs were DAMO bacteria and Methylocystis sp. The electroactive bacteria detected were Pseudomonas sp., Hypomicrobium sp., Thiobacillus sp, and Rhodococcus sp. The pil, cytochrome c, hdr, and he/fp genes related to EET were present on the electrode surfaces. Carbon 13 isotope tracing and chemicals analysis by GC-MS exhibited that methanol was an intermediate product released to extracellular environment and acted as the electronic carrier to drive the EET in methane oxidation. Extracellular electron transfer was achieved through the collaboration of DAMO bacteria, Methylocystis sp., and Pseudomonas sp. Anthraquinone 2-sulfonic acid ester (AQS) could improve the rate of electron transfer to the extracellular space, especially in the EMN-0.5 reaction system. This study provides a new understanding of AMO consortium metabolism in BES and may provide a scientific basis for developing methane control technology.

Biogeographic distributions of microbial communities associated with anaerobic methane oxidation in the surface sediments of deep-sea cold seeps in the South China Sea.

Cold seeps are oasis for the microbes in the deep-sea ecosystems, and various cold seeps are located along the northern slope of the South China Sea (SCS). However, by far most microbial ecological studies were limited to specific cold seep in the SCS, and lack of comparison between different regions. Here, the surface sediments (0-4 cm) from the Site F/Haima cold seeps and the Xisha trough in the SCS were used to elucidate the biogeography of microbial communities, with particular interest in the typical functional groups involved in the anaerobic oxidation of methane (AOM) process. Distinct microbial clusters corresponding to the three sampling regions were formed, and significantly higher gene abundance of functional groups were present in the cold seeps than the trough. This biogeographical distribution could be explained by the geochemical characteristics of sediments, such as total nitrogen (TN), total phosphorus (TP), nitrate (NO3 -), total sulfur (TS) and carbon to nitrogen ratios (C/N). Phylogenetic analysis demonstrated that mcrA and pmoA genotypes were closely affiliated with those from wetland and mangroves, where denitrifying anaerobic methane oxidation (DAMO) process frequently occurred; and highly diversified dsrB genotypes were revealed as well. In addition, significantly higher relative abundance of NC10 group was found in the Xisha trough, suggesting that nitrite-dependent DAMO (N-DAMO) process was more important in the hydrate-bearing trough, although its potential ecological contribution to AOM deserves further investigation. Our study also further demonstrated the necessity of combining functional genes and 16S rRNA gene to obtain a comprehensive picture of the population shifts of natural microbial communities among different oceanic regions.

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.

Combining biological denitrification and electricity generation in methane-powered microbial fuel cells

Methane has been demonstrated to be a feasible substrate for electricity generation in microbial fuel cells (MFCs) and denitrifying anaerobic methane oxidation (DAMO). However, these two processes were evaluated separately in previous studies and it has remained unknown whether methane is able to simultaneously drive these processes. Here we investigated the co-occurrence and performance of these two processes in the anodic chamber of MFCs. The results showed that methane successfully fueled both electrogenesis and denitrification. Importantly, the maximum nitrate removal rate was significantly enhanced from (1.4 ± 0.8) to (18.4 ± 1.2) mg N/(L·day) by an electrogenic process. In the presence of DAMO, the MFCs achieved a maximum voltage of 610 mV and a maximum power density of 143 ± 12 mW/m2. Electrochemical analyses demonstrated that some redox substances (e.g. riboflavin) were likely involved in electrogenesis and also in the denitrification process. High-throughput sequencing indicated that the methanogen Methanobacterium, a close relative of Methanobacterium espanolae, catalyzed methane oxidation and cooperated with both exoelectrogens and denitrifiers (e.g., Azoarcus). This work provides an effective strategy for improving DAMO in methane-powered MFCs, and suggests that methanogens and denitrifiers may jointly be able to provide an alternative to archaeal DAMO for methane-dependent denitrification.

The denitrifying anaerobic methane oxidation process and microorganisms in the environments: A review

Methane (CH4) is an important greenhouse gas with a global warming potential 28 – 34 times that of CO2 over the 100-year horizon. Denitrifying anaerobic methane oxidation (DAMO) is a recently discovered process that potentially represents an important CH4 sink globally. This process involves two possible pathways: the nitrite-dependent DAMO mediated by NC10 bacteria and the nitrate-dependent DAMO by ANME-2d archaea. Both are widely detected in freshwater and coastal habitats using molecular tools. However, the distributions of these two processes and the functional microorganisms and their interactions with other N cycling pathways are far from clear. In this review, we conducted a scientometric analysis on a co-citation network consisting of 835 references derived from 354 citing articles closely related to the distribution of DAMO in the environment. Through this analysis, we found that current studies focus more on freshwater systems than coastal systems, and ANME-2d archaea are generally under-studied compared to NC10 bacteria. The emerging research topics in this area include AMO processes coupled to alternative electron acceptors and their role as CH4 sinks. We further reviewed papers focusing on DAMO distribution in freshwater and coastal environments guided by the result of the scientometric analysis. Finally, we identified several areas that require further research and proposed future research including comparisons of DAMO with other N cycling pathways and environmental conditions in the context of the river-estuary-sea continuum.

Enhancement of denitrifying anaerobic methane oxidation via applied electric potential.

In this study, single-chamber three-electrode electrochemical sequencing batch reactor (ESBR) was set up to investigate the impact of applying potential on denitrifying anaerobic methane oxidation (DAMO) process. When the applied potential was +0.8V, the conversion rate of nitrite to nitrogen was superior to those of other potentials. With the optimal potential of +0.8V for 60 days, the nitrite removal rate of ESBR could reach 3.34±0.28mgN/L/d, which was 4.5 times more than that of the non-current control (0.74±0.16mgN/L/d). The DAMO functional bacteria Candidatus Methylomirabilis exhibited noticeable enrichment under applying potential, and its functional gene of pmoA was significantly expressed. Through untargeted LC-MS metabolomics analysis, applied potential was shown to affect the regulation of prior metabolites including spermidine, spermine and glycerophosphocholine that were related to the metabolic pathways of glycerophospholipid metabolism and arginine and proline metabolism, which had positive effects on DAMO process. These results show that applying electric potential could be a useful strategy in DAMO process used for methane and nitrogen removal.

Enhancement of nitrate-dependent anaerobic methane oxidation via granular activated carbon

Denitrifying anaerobic methane oxidation (DAMO) is a bioprocess utilizing methane as the electron source to remove nitrate or nitrite, but denitrification rate especially for nitrate-dependent DAMO is usually limited due to the low methane mass transfer efficiency. In this research, granular active carbon (GAC) was added to enhance the nitrate-dependent DAMO process. The results showed that the maximum nitrate removal rate of GAC assisted DAMO system reached as high as 61.17 mg L−1 d−1, 8 times higher than that of non-amended control SBR. The porous structure of GAC can not only adsorb methane, but also keep the internal DAMO archaea from being washed out, and thus benefits for DAMO archaea enrichment. The relative abundance of DAMO archaea accounted for 96.3% in GAC-SBR, which was significantly higher than that of non-amended control SBR system (29.9%). Furthermore, GAC amendment up-regulated metabolic status of denitrification and methane oxidation based on gene sequence composition. The absolute abundances of function genes (NC10 pmoA and ANME mcrA) in GAC-SBR were almost 20 times higher than that of non-amended control SBR. This study provides a novel technique to stimulate the nitrate-dependent DAMO process.

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.

Influences of longitudinal gradients on methane-driven membrane biofilm reactor for complete nitrogen removal: A model-based investigation

Integrating anammox with denitrifying anaerobic methane oxidation (DAMO) in the membrane biofilm reactor (MBfR) is a promising technology capable of achieving complete nitrogen removal from wastewater. However, it remains unknown whether reactor configurations featuring longitudinal gradients parallel to the membrane surface would affect the performance of the CH4-driven MBfR. To this end, this work aims to study the impacts of longitudinal heterogeneity potentially present in the gas and liquid phases on a representative CH4-driven MBfR performing anammox/DAMO by applying the reported modified compartmental modeling approach. Through comparing the modeling results of different reactor configurations, this work not only offered important guidance for better design, operation and monitoring of the CH4-driven MBfR, but also revealed important implications for prospective related modeling research. The total nitrogen removal efficiency of the MBfR at non-excessive CH4 supply (e.g., surface loading of ≤0.064 g-COD m−2 d−1 in this work) was found to be insensitive to both longitudinal gradients in the liquid and gas phases. Comparatively, the longitudinal gradient in the liquid phase led to distinct longitudinal biomass stratification and therefore played an influential role in the effective CH4 utilization efficiency, which was also related to the extent of reactor compartmentation considered in modeling. When supplied with non-excessive CH4, the MBfR is recommended to be designed/operated with both the biofilm reactor and the membrane lumen as plug flow reactors (PFRs) with co-current flow of wastewater and CH4, which could mitigate dissolved CH4 discharge in the effluent. For the reactor configurations with the biofilm reactor designed/operated as a PFR, multi-spot sampling in the longitudinal direction is needed to obtain a correct representation of the microbial composition of the MBfR.