Nmol CO2 Research Articles

Identification of Quality Characteristics of Autochthonous Karasüt Apple during Cold Storage

The main objective of the research was to assess weight loss, respiration rate, firmness, soluble solids content (SSC), titratable acidity and vitamin C content of the autochthonous Karasüt apple throughout cold storage. The ‘Granny Smith’ cultivar was used as positive control. The fruit was kept at a temperature of 0.0±0.5°C and relative humidity of 90±5%. Quality losses were observed in the apples during cold storage. The weight loss of Karasüt apple (6.70%) was higher than that of Granny Smith (2.20%) at the end of cold storage. A lower respiration rate was measured in the Karasüt apple (1.23 nmol CO2 kg-1 s-1), compared to the positive control (1.53 nmol CO2 kg-1 s-1) at harvest. On the contrary, the respiration rate was higher on days 30 and 60. The fruit firmness of Granny Smith (27.14 N) was higher than that of Karasüt (22.47 N) at the end of cold storage. During the cold storage, a higher SSC was obtained from Karasüt apple compared to the positive control. However, titratable acidity was lower in Karasüt apple. In the first two measurements of cold storage, the vitamin C of Karasüt apple was higher than that of the positive control. As a result, it was revealed that Karasüt apple had faster quality losses during cold storage than Granny Smith apple cultivar.

Vertical and temporal variations in activity, abundance, and composition of nitrite-driven anaerobic methanotrophs in a paddy field

Paddy soil is an important source of methane (CH4) emissions, and the nitrite-driven anaerobic oxidation of methane (AOM) mediated by Candidatus Methylomirabilis bacteria is a crucial process for CH4 mitigation. However, the vertical and temporal fluctuations in nitrite-driven AOM activity, as well as the abundance and composition of Methylomirabilis bacteria in paddy soil, remain poorly understood. We investigated the potential rate of nitrite-driven AOM and the composition of Methylomirabilis bacteria at varying depths (0–40 cm, at 10 cm intervals) within a paddy soil during different growth stages of rice, including tillering, jointing, flowering, and milky. The potential rate of nitrite-driven AOM ranged from 0.24 to 20.6 nmol CO2 g−1 (dry soil) d−1, and the abundance of Methylomirabilis bacterial 16S rRNA genes varied from 5.8 × 106 to 4.4 × 107 copies g−1 dry soil. The 10–20 cm soil depth showed higher potential activity and bacterial abundance compared to other depths, and the tillering and jointing stages exhibited greater activity and abundance. The composition of Methylomirabilis bacteria showed significant variation across different soil depths but remained stable throughout the growth stages of rice. Soil organic carbon content was positively correlated with potential nitrite-driven AOM rate, and nitrite content was negatively correlated with the abundance of Methylomirabilis bacteria. Soil organic carbon and ammonium content had a significant impact on the composition of Methylomirabilis bacteria. The results underscore the substantial influence of soil depth and the growth stage of rice on nitrite-driven AOM in paddy ecosystems.

Size matters: Aerobic methane oxidation in sediments of shallow thermokarst lakes

AbstractShallow thermokarst lakes are important sources of greenhouse gases (GHGs) such as methane (CH4) and carbon dioxide (CO2) resulting from continuous permafrost thawing due to global warming. Concentrations of GHGs dissolved in water typically increase with decreasing lake size due to coastal abrasion and organic matter delivery. We hypothesized that (i) CH4 oxidation depends on the natural oxygenation gradient in the lake water and sediments and increases with lake size because of stronger wind‐induced water mixing; (ii) CO2 production increases with decreasing lake size, following the dissolved organic matter gradient; and (iii) both processes are more intensive in the upper than deeper sediments due to the in situ gradients of oxygen (O2) and bioavailable carbon. We estimated aerobic CH4 oxidation potentials and CO2 production based on the injection of 13C‐labeled CH4 in the 0–10 cm and 10–20 cm sediment depths of small (~300 m2), medium (~3000 m2), and large (~106 m2) shallow thermokarst lakes in the West Siberian Lowland. The CO2 production was 1.4–3.5 times stronger in the upper sediments than in the 10–20 cm depth and increased from large (158 ± 18 nmol CO2 g−1 sediment d.w. h−1) to medium and small (192 ± 17 nmol CO2 g−1 h−1) lakes. Methane oxidation in the upper sediments was similar in all lakes, while at depth, large lakes had 14‐ and 74‐fold faster oxidation rates (5.1 ± 0.5 nmol CH4‐derived CO2 g−1 h−1) than small and medium lakes, respectively. This was attributed to the higher O2 concentration in large lakes due to the more intense wind‐induced water turbulence and mixing than in smaller lakes. From a global perspective, the CH4 oxidation potential confirms the key role of thermokarst lakes as an important hotspot for GHG emissions, which increase with the decreasing lake size.

Effect of gradual increase of atmospheric CO2 concentration on nitrate-dependent anaerobic methane oxidation in paddy soils

Nitrate-dependent anaerobic oxidation of methane (AOM) is a new pathway to reduce methane emissions from paddy ecosystems. The elevated atmospheric CO2 concentration can affect methane emissions from paddy ecosystems, but its impact on the process of nitrate-dependent AOM is poorly known. Based on the automatic CO2 control platform with open top chambers and the 13CH4 stable isotope experiments, the responses of the activity of nitrate-dependent AOM, abundance and community composition of Candidatus Methanoperedens nitroreducens (M. nitroreducens)-like archaea to the gradual increase of CO2 concentration were investigated in paddy fields. We set up two CO2 concentration treatments, including an ambient CO2 and a gradual increase of CO2(increase of 40 μL·L-1 per year above ambient CO2 concentration until 160 μL·L-1). The results showed the nitrate-dependent AOM rate of 0.7-11.3 nmol CO2·g-1·d-1 in the studied paddy fields, and quantitative PCR showed the abundance of M. nitroreducens-like archaeal mcrA genes of 2.2×106-8.5×106 copies·g-1. Compared to the ambient CO2 treatment, the slow elevated CO2 treatment enhanced the nitrate-dependent AOM rate and stimulated the abundance of M. nitroreducens-like archaea, particularly in 5-10 cm soil layer. The gradual increased CO2 concentration treatment did not change the community composition of M. nitroreducens-like archaea, but significantly decreased their diversity. The soil organic carbon content was an important factor influencing the nitrate-dependent AOM process. Overall, our results showed that the gradual increase of CO2 concentration could promote the nitrate-dependent AOM, suggesting its positive role in mitigating methane emissions from paddy ecosystems under future climate change.

Spatial variations of activity and community structure of nitrite-dependent anaerobic methanotrophs in river sediment

Rivers are an important site for methane emissions and reactive nitrogen removal. The process of nitrite-dependent anaerobic methane oxidation (n-damo) links the global carbon cycle and the nitrogen cycle, but its role in methane mitigation and nitrogen removal in rivers is poorly known. In the present study, we investigated the activity, abundance, and community composition of n-damo bacteria in sediment of the upper, middle, and lower reaches of Wuxijiang River (Zhejiang Province, China). The 13CH4 stable isotope experiments showed that the methane oxidation activity of n-damo was 0.11–1.88 nmol CO2 g−1 (dry sediment) d−1, and the activity measured from the middle reaches was significantly higher than that from the remaining regions. It was estimated that 3.27 g CH4 m−2 year−1 and 8.72 g N m−2 year−1 could be consumed via n-damo. Quantitative PCR confirmed the presence of n-damo bacteria, and their 16S rRNA gene abundance varied between 5.45 × 105 and 5.86 × 106 copies g−1 dry sediment. Similarly, the abundance of n-damo bacteria was significantly higher in the middle reaches. High-throughput sequencing showed a high n-damo bacterial diversity, with totally 152 operational taxonomic units being detected at 97 % sequence similarity cut-off. In addition, the n-damo bacterial community composition also varied spatially. The inorganic nitrogen (NH4+, NO2−, NO3−) level was found to be the key environmental factor controlling the n-damo activity and bacterial community composition. Overall, our results showed the spatial variations and environmental regulation of the activity and community structure of n-damo bacteria in river sediment, which expanded our understanding of the quantitative importance of n-damo in both methane oxidation and reactive nitrogen removal in riverine systems.

Subnanomolar Sensitive Stable Isotopic Determination in CO2 by Tunable Infrared Laser Absorption Spectroscopy.

Oxygen and carbon stable isotope ratios (18O/16O, 13C/12C, and 17O/16O) of CO2 have been crucial in helping us understand Earth and planetary systems. These ratios have also been used in medicine for the noninvasive diagnosis of diseases from exhaled breath and for quantifying biochemical or metabolic reactions and in determining the production area of agricultural products. The current method for measuring the stable isotope ratios of CO2 is primarily gas-source isotope ratio mass spectroscopy (IRMS). Due to the recent demand for isotopic microanalysis of carbonates and organic compounds, the sample size required for isotopic measurements has been reduced to approximately 2 nmol CO2 (equivalent to 0.2 μg CaCO3 and 24 ng carbon) by using high-precision IRMS. We report a novel method using tunable mid-infrared laser direct absorption spectroscopy (TILDAS) for sensitive measurements of 18O/16O and 13C/12C in subnanomolar CO2. This method can accurately measure 18O/16O and 13C/12C in CO2 with a repeatability of less than 0.03‰ (n = 28) in a range of 0.3 nmol (equivalent to 0.03 μg CaCO3 and 3.8 ng carbon) to 30 nmol. This is a sample size 1 order of magnitude smaller than currently available sensitive analytical techniques. In addition, the TILDAS system measures 17O/16O simultaneously with a repeatability of less than 0.06‰ (n = 28). Our method is a major advance in supersensitive CO2 stable isotopic analyses for various fields.

Finding the pieces for the anaerobic methane oxidation jigsaw puzzle in mangrove wetlands

Estuarine mangrove wetlands are generally considered a major source of methane (CH4) emissions. Anaerobic methane oxidation (AMO) is an important pathway for reducing CH4 emissions and is known to be tightly linked to carbon, nitrogen, sulfur and metal cycling processes largely driven by microorganisms. However, these coupled processes and their contributions to mitigating global warming remain unclear in mangrove wetlands. In this study, the depth-specific potential methane oxidation rates of nitrate-, nitrite- and nitrous oxide-dependent anaerobic methane oxidation (nitrate-, nitrite- and N2O-AMO) and sulfate-dependent anaerobic methane oxidation (sulfate-AMO) processes were investigated using stable isotope tracing along with an assessment of the vertical nutrient and CH4 content profiles in mangrove sediments. The results indicated that nitrite-AMO, nitrate-AMO, N2O-AMO and sulfate-AMO were active and exhibited methane oxidation rates varying from 24.1 to 1077.4, 14.1 to 66.1, 14.3 to 40.9 and 4.2–36.7 nmol CO2 g−1 dry soil d−1, respectively. Furthermore, nitrite-AMO and N2O-AMO tended to be more active in the upper sediment layers, while nitrate-AMO and sulfate-AMO displayed higher rates in the deeper layers. The contribution rate of nitrite-AMO to the total of the 4 AMO processes examined in the mangrove sediments reached 28.90–95.80%, followed by nitrate-AMO (2.73–46.46%), sulfate-AMO (1.46–24.65%) and N2O-AMO (1.70–4.47%), suggesting that the nitrite-AMO process is a significant methane sink in mangrove wetland ecosystems. The temporal and spatial variations in the nitrite-AMO bacterial depth and genus-specific distribution, abundance, activity and sensitivity to environmental factors were then investigated. The absolute abundances of the nitrite-AMO bacterial 16S rRNA and pmoA genes were 0.51–6.55 × 107 and 0.79–8.43 × 106 copies g−1 ds, respectively, and these values decreased with sediment depth; most of the samples were characterized as belonging to Group B of NC10. In addition, NO2− is the limiting factor for nitrite-AMO bacteria in mangrove wetlands.

Response of nitrite-dependent anaerobic methanotrophs to elevated atmospheric CO2 concentration in paddy fields

Nitrite-dependent anaerobic methane oxidation (n-damo) catalyzed by Candidatus Methylomirabilis oxyfera (M. oxyfera)-like bacteria is a new pathway for the regulation of methane emissions from paddy fields. Elevated atmospheric CO2 concentrations (e[CO2]) can indirectly affect the structure and function of microbial communities. However, the response of M. oxyfera-like bacteria to e[CO2] is currently unknown. Here, we investigated the effect of e[CO2] (ambient CO2 + 200 ppm) on community composition, abundance, and activity of M. oxyfera-like bacteria at different depths (0–5, 5–10, and 10–20 cm) in paddy fields across multiple rice growth stages (tillering, jointing, and flowering). High-throughput sequencing showed that e[CO2] had no significant effect on the community composition of M. oxyfera-like bacteria. However, quantitative PCR suggested that the 16S rRNA gene abundance of M. oxyfera-like bacteria increased significantly in soil under e[CO2], particularly at the tillering stage. Furthermore, 13CH4 tracer experiments showed potential n-damo activity of 0.31–8.91 nmol CO2 g−1 (dry soil) d−1. E[CO2] significantly stimulated n-damo activity, especially at the jointing and flowering stages. The n-damo activity and abundance of M. oxyfera-like bacteria increased by an average of 90.9% and 50.0%, respectively, under e[CO2]. Correlation analysis showed that the increase in soil dissolved organic carbon content caused by e[CO2] had significant effects on the activity and abundance of M. oxyfera-like bacteria. Overall, this study provides the first evidence for a positive response of M. oxyfera-like bacteria to e[CO2], which may help reduce methane emissions from paddy fields under future climate change conditions.

Microbial Involvement in Carbon Transformation via CH4 and CO2 in Saline Sedimentary Pool.

Simple SummaryMethane and carbon dioxide are commonly found in the environment and are considered the most important greenhouse gases. Transformation of these gases is in large carried by microorganisms, which occur even in extreme environments. This study presents methane-related biological processes in saline sediments of the Miocene Wieliczka Formation, Poland. Biological activity (carbon dioxide and methane production or methane oxidation), confirmed by stable isotope indices, occurred in all of the studied Wieliczka rocks. CH4-utilizing microbes constituted 0.7–3.6% while methanogens (represented by Methanobacterium) only 0.01–0.5% of taxa present in the Wieliczka Salt Mine rocks. Water activity was the key factor regulating microbial activity in saline subsurface sediments. Generally, CO2 respiration was higher in anaerobic conditions while methanogenic and methanotrophic activities were dependent on the type of rock.Methane and carbon dioxide are one of the most important greenhouse gases and significant components of the carbon cycle. Biogeochemical methane transformation may occur even in the extreme conditions of deep subsurface ecosystems. This study presents methane-related biological processes in saline sediments of the Miocene Wieliczka Formation, Poland. Rock samples (W2, W3, and W4) differed in lithology (clayey salt with veins of fibrous salt and lenses of gypsum and anhydrite; siltstone and sandstone; siltstone with veins of fibrous salt and lenses of anhydrite) and the accompanying salt type (spiza salts or green salt). Microbial communities present in the Miocene strata were studied using activity measurements and high throughput sequencing. Biological activity (i.e., carbon dioxide and methane production or methane oxidation) occurred in all of the studied clayey salt and siltstone samples but mainly under water-saturated conditions. Microcosm studies performed at elevated moisture created more convenient conditions for the activity of both methanogenic and methanotrophic microorganisms than the intact sediments. This points to the fact that water activity is an important factor regulating microbial activity in saline subsurface sediments. Generally, respiration was higher in anaerobic conditions and ranged from 36 ± 2 (W2200%t.w.c) to 48 ± 4 (W3200%t.w.c) nmol CO2 gdw−1 day−1. Methanogenic activity was the highest in siltstone and sandstone (W3, 0.025 ± 0.018 nmol CH4 gdw−1 day−1), while aerobic methanotrophic activity was the highest in siltstone with salt and anhydrite (W4, 220 ± 66 nmol CH4 gdw−1 day−1). The relative abundance of CH4-utilizing microorganisms (Methylomicrobium, Methylomonas, Methylocystis) constituted 0.7–3.6% of all taxa. Methanogens were represented by Methanobacterium (0.01–0.5%). The methane-related microbes were accompanied by a significant number of unclassified microorganisms (3–64%) and those of the Bacillus genus (4.5–91%). The stable isotope composition of the CO2 and CH4 trapped in the sediments suggests that methane oxidation could have influenced δ13CCH4, especially in W3 and W4.

A survey of greenhouse gases production in central European lignites

Due to changes in the energy market, it is projected that lignite excavation will be reduced in the near future. Cessation of exploitation is associated with restitution of natural water conditions and flooding of the resources left in the mines. Flooded lignite mines are a potential source of greenhouse gases (GHG) (CH4, CO2 and N2O), which should be monitored due to growing environmental concerns. Here, we aim to recognize GHG release from the lignites collected from the main deposits of Poland, Slovenia and Serbia. GHG production was studied along with a range of physical and chemical parameters that are crucial for microbial growth and activity.The microcosm experiments showed that the main gas emitted from the lignites was carbon dioxide. Daily CO2 production was highly variable. The highest values were recorded for detroxylitic lignite collected from the Konin deposit (402.05 nmol CO2 g-1 day-1) while the lowest were for the Kolubara xylitic lignite (19.64 nmol CO2 g-1 day-1). Methane production was much lower and ranged from nearly zero to 66.75 nmol g dry mass-1 d-1. Nitrous oxide production was not detected. It was found that CO2 production, being a general measure of microbial activity, was positively affected by NO3- concentration and redox potential. With respect to methane formation, the lower atmospheric oxygen exposure of the sample from the Velenje underground mine compared to the samples from the opencast mines has been identified as a possible cause of the high methane production. The overall global warming potential (GWP) of the gases released by xylitic lignite was lowest among the samples. Preferential extraction of the detritic lignites is suggested as a means to reduce GHG emissions from the abandoned lignite mines.

Activity, abundance and community composition of nitrite-dependent methanotrophs in response to fertilization in paddy soils

Nitrite-dependent anaerobic oxidation of methane (AOM), which is mediated by Candidatus Methylomirabilis oxyfera (M. oxyfera)-like bacteria, has the great potential to reduce methane emissions from paddy ecosystems. Fertilization, a necessary practice for increasing crop yields, could greatly affect nitrite-dependent AOM process. Here, we investigated the abundance, community composition and activity of M. oxyfera-like bacteria at different layers (0–10, 20–30 and 40–50 cm) of paddy soils under three fertilization treatments, including unfertilized control (CK), chemical fertilization (CF) and organic fertilization (OF). The 16S rRNA gene abundance of M. oxyfera-like bacteria in soils under CF (6.1 × 106–2.2 × 107 copies g−1 dry soil) and OF (8.9 × 106–1.8 × 107 copies g−1 dry soil) treatments was higher than that under CK treatment (5.0 × 106–1.5 × 107 copies g−1 dry soil). An obvious separation of M. oxyfera-like bacterial communities was observed between different fertilization treatments, and a higher diversity of these bacteria was detected in CF and OF soils. The nitrite-dependent AOM activity was increased in soils under CF (3.6–11.5 nmol CO2 g−1 (dry soil) d−1) and OF (3.4–7.2 nmol CO2 g−1 (dry soil) d−1) treatments as compared to CK (1.6–4.4 nmol CO2 g−1 (dry soil) d−1) treatment, particularly at the layer of 20–30 cm. Overall, our results demonstrated that the communities and activity of M. oxyfera-like bacteria responded positively to both chemical and organic fertilization, which expanded our knowledge of the role of nitrite-dependent AOM in regulating methane emissions from paddy ecosystems.

Anaerobic oxidation of methane with denitrification in sediments of a subtropical estuary: Rates, controlling factors and environmental implications

Anaerobic oxidation of methane with denitrification (DAMO), as an important microbial process regulating methane emission, has been widely reported in freshwater ecosystems. However, the DAMO process and associated biogeochemical controls in estuaries remain poorly understood. Here, we used 13C- and 15N-labelling experiments to quantify the potential rates of DAMO and determined the crucial factors controlling the DAMO rates in the sediment of Yangtze Estuary. Potential rates of DAMO varied greatly across the estuary, ranging from 0.07 to 0.28 nmol CO2 g−1 d−1. Salinity negatively affected the DAMO and also showed an indirectly negative influence on DAMO process by high salinity inhibition on NO3− availability and denitrification. Nitrate concentrations were significantly correlated with the DAMO rates. Denitrification rates showed positive correlation with DAMO rates, implying that nitrate reduction drives the DAMO process. Sediment total organic carbon and NH4+ had important effects on DAMO rates. These results together indicate that DAMO process can occur and the DAMO rates were mainly controlled by sediment NO3− and denitrification in estuary. We further conclude that increasing NO3− load can drive the DAMO process with more important implications on methane sink in estuarine ecosystems.

Different responses of nitrite- and nitrate-dependent anaerobic methanotrophs to increasing nitrogen loading in a freshwater reservoir

Nitrite (NO2−)- and nitrate (NO3−)-dependent anaerobic oxidation of methane (AOM) are two new additions in microbial methane cycle, which potentially act as important methane sinks in freshwater aquatic systems. Here, we investigated spatial variations of community composition, abundance and potential activity of NO2−- and NO3−-dependent anaerobic methanotrophs in the sediment of Jiulonghu Reservoir (Zhejiang Province, China), a freshwater reservoir having a gradient of increasing nitrogen loading from upstream to downstream regions. High-throughput sequencing of total bacterial and archaeal 16S rRNA genes showed the cooccurrence of Candidatus Methylomirabilis oxyfera (M. oxyfera)-like and Candidatus Methanoperedens nitroreducens (M. nitroreducens)-like anaerobic methanotrophs in the examined reservoir sediments. The community structures of these methanotrophs differed substantially between the sediments of upstream and downstream regions. Quantitative PCR suggested higher M. oxyfera-like bacterial abundance in the downstream (8.6 × 107 to 2.8 × 108 copies g−1 dry sediment) than upstream sediments (2.4 × 107 to 3.5 × 107 copies g−1 dry sediment), but there was no obvious difference in M. nitroreducens-like archaeal abundance between these sediments (3.7 × 105 to 4.8 × 105 copies g−1 dry sediment). The 13CH4 tracer experiments suggested the occurrence of NO2−- and NO3−-dependent AOM activities, and their rates were 4.7–14.1 and 0.8–2.6 nmol CO2 g−1 (dry sediment) d−1, respectively. Further, the rates of NO2−-dependent AOM in downstream sediment were significantly higher than those in upstream sediment. The NO3− concentration was the key factor affecting the spatial variations of abundance and activity of NO2−-dependent anaerobic methanotrophs. Overall, our results showed different responses of NO2−- and NO3−-dependent anaerobic methanotrophs to increasing nitrogen loading in a freshwater reservoir.

Depth-specific distribution and significance of nitrite-dependent anaerobic methane oxidation process in tidal flow constructed wetlands used for treating river water.

Tidal flow constructed wetlands (TF CWs) have been considered an effective approach to treat contaminated river water, as well as a significant role in global matter cycles, especially for carbon and nitrogen. Notably, it has been thought that methane oxidation was completely catalyzed by the aerobic process, ignoring the anaerobic methane oxidation, such as the nitrite-dependent anaerobic methane oxidation (n-damo) process. In our current work, therefore, we used molecular and stable isotopes to investigate the biodiversity, quantity and potential rate of n-damo bacteria in the TF CWs located in the Xisha Wetland Park in the Yangtze River estuary, China. The results revealed that n-damo process was active in the collected soil cores, with a decreasing trend along water depths and rates ranging from 8.48 to 23.45nmol CO2 g-1 dry soil d-1. The n-damo bacterial contributions to CH4 oxidation and N2 production in TF CWs reached 9.49-26.26% and 20.73-47.11%, respectively, suggesting that n-damo bacteria was an important nitrogen and methane sink in the TF CWs, but had been previously overlooked. The copy numbers of total bacterial 16S rRNA and pmoA genes were 1.84-11.21×109 and 0.59-2.72×106 copies g-1 ds, respectively, as the higher abundance was found in the soil at lower water levels during tidal submergence. Diverse n-damo bacterial 16S rRNA gene sequences belonged to group B, C and D were measured, and it was found that group B and C were the most frequently measured n-damo clusters in the TF CWs. In addition, nitrite was the key factor regulating the n-damo bacterial distribution in the TF CWs. This study would broaden our horizons and help us better understand the nitrogen and methane cycles in tidal ecosystems.

Coupled methane and nitrous oxide biotransformation in freshwater wetland sediment microcosms

Anaerobic oxidation of methane (AOM) coupled to denitrification is becoming the focus of scientific inquiry due to its potential contribution to global carbon and nitrogen cycles. AOM has been previously reported to proceed with nitrate (NO3−) or nitrite (NO2−). However, little research has been conducted on the simultaneous use of methane (CH4) and nitrous oxide (N2O). Here, coupled CH4 and N2O biotransformation in a freshwater wetland sediment was obtained in a 7-day anaerobic sediment incubation assay. The significant CO2 accumulation and decrease of CH4 emission in sediment microcosms was attributed to two mechanisms: inhibition of methanogenesis and N2O-dependent AOM. To further confirm the coupled CH4 and N2O transformation, a 13C-labelled stable isotope tracer assay after anaerobic incubation was conducted with N2O and/or CH4 amendments. The N2O-dependent AOM rate was 3.41 ± 0.13 nmol CO2 g−1 dry sediment·day−1. According to metagenomic analysis, addition of N2O stimulated AOM by increasing the activity and abundance of methanotrophic bacteria and by increasing enzymatic activities in the electron transport chain. Based on these results, we propose coupled CH4 and N2O biotransformation in the sediment microcosms for the first time, carried out by unidentified methanotroph(s) via intra‑oxygen produced in the presence of N2O. Such a process has the potential to reduce the emission of two highly potent greenhouse gases and makes a significant contribution to the link of global carbon and nitrogen cycles in anoxic environments.

Molecular and stable isotopic evidence for the occurrence of nitrite-dependent anaerobic methane-oxidizing bacteria in the mangrove sediment of Zhangjiang Estuary, China.

Nitrite-dependent anaerobic methane oxidation (n-damo), which is mediated by "Candidatus Methylomirabilis oxyfera-like" bacteria, is unique in linking the carbon and nitrogen cycles. However, the niche and activity of n-damo bacteria in the mangrove ecosystem have not been confirmed. Here, we report the occurrence of the n-damo process in the mangrove wetland of the Zhangjiang Estuary, China. The widespread occurrence of n-damo bacteria in mangrove wetland was confirmed using real-time quantitative polymerase chain reaction (qPCR) assay, which showed that the abundance of Methylomirabilis oxyfera-like bacterial 16S rRNA and pmoA genes ranged from 2.43 × 106 to 2.09 × 107 and 2.07 × 106 to 3.38 × 107copies per gram of dry soil in the examined sediment cores. The highest amount of targeting genes was all detected in the upper layer (0-20cm). Phylogenetic analyses of n-damo bacterial 16S rRNA and pmoA genes illustrated the depth-specific distribution and high diversity of n-damo bacteria in the mangrove wetland. Stable isotope experiments further confirmed the occurrence of n-damo in the examined mangrove sediments, and the potential n-damo rates ranged from 25.93 to 704.08nmol CO2 per gram of dry soil per day at different depths of the sediment cores, with the n-damo being more active in the upper layer of the mangrove sediments. These results illustrate the existence of active M. oxyfera-like bacteria and indicate that the n-damo process is a previously overlooked microbial methane sink in the mangrove wetlands.

Continuous-flow IRMS technique for determining the <sup>17</sup>O excess of CO<sub>2</sub> using complete oxygen isotope exchange with cerium oxide

Abstract. This paper presents an analytical system for analysis of all single substituted isotopologues (12C16O17O, 12C16O18O, 13C16O16O) in nanomolar quantities of CO2 extracted from stratospheric air samples. CO2 is separated from bulk air by gas chromatography and CO2 isotope ratio measurements (ion masses 45 / 44 and 46 / 44) are performed using isotope ratio mass spectrometry (IRMS). The 17O excess (Δ17O) is derived from isotope measurements on two different CO2 aliquots: unmodified CO2 and CO2 after complete oxygen isotope exchange with cerium oxide (CeO2) at 700 °C. Thus, a single measurement of Δ17O requires two injections of 1 mL of air with a CO2 mole fraction of 390 μmol mol−1 at 293 K and 1 bar pressure (corresponding to 16 nmol CO2 each). The required sample size (including flushing) is 2.7 mL of air. A single analysis (one pair of injections) takes 15 minutes. The analytical system is fully automated for unattended measurements over several days. The standard deviation of the 17O excess analysis is 1.7‰. Multiple measurements on an air sample reduce the measurement uncertainty, as expected for the statistical standard error. Thus, the uncertainty for a group of 10 measurements is 0.58‰ for Δ 17O in 2.5 h of analysis. 100 repeat analyses of one air sample decrease the standard error to 0.20‰. The instrument performance was demonstrated by measuring CO2 on stratospheric air samples obtained during the EU project RECONCILE with the high-altitude aircraft Geophysica. The precision for RECONCILE data is 0.03‰ (1σ) for δ13C, 0.07‰ (1σ) for δ18O and 0.55‰ (1σ) for δ17O for a sample of 10 measurements. This is sufficient to examine stratospheric enrichments, which at altitude 33 km go up to 12‰ for δ17O and up to 8‰ for δ18O with respect to tropospheric CO2 : δ17O ~ 21‰ Vienna Standard Mean Ocean Water (VSMOW), δ18O ~ 41‰ VSMOW (Lämmerzahl et al., 2002). The samples measured with our analytical technique agree with available data for stratospheric CO2.

Depth-specific distribution and importance of nitrite-dependent anaerobic ammonium and methane-oxidising bacteria in an urban wetland

Anaerobic ammonium oxidation (anammox) and nitrite-dependent anaerobic methane oxidation (n-damo) are two recently discovered processes in the nitrogen cycle that are catalysed by anammox bacteria and n-damo bacteria, respectively. Here, the depth-specific distribution and importance of anammox bacteria and n-damo bacteria were studied in an urban wetland, Xixi Wetland, Zhejiang Province (China). Anammox bacteria related to Candidatus Brocadia, Candidatus Kuenenia and Candidatus Anammoxoglobus, and n-damo bacteria related to “Candidatus Methylomirabilis oxyfera” were present in the collected soil samples. The abundance of anammox bacteria (2.6–8.6 × 106 copies g−1 dry soil) in the shallow soils (0–10 cm and 20–30 cm) was higher than that (2.5–9.8 × 105 copies g−1 dry soil) in the deep soils, whereas the abundance of n-damo bacteria (0.6–1.3 × 107 copies g−1 dry soil) in the deep soils (50–60 cm and 90–100 cm) was higher than that (3.4–4.5 × 106 copies g−1 dry soil) in the shallow soils. Anammox activity was detected at all depths, and higher potential rates (12.1–21.4 nmolN2g−1 dry soil d−1) were observed at depths of 0–10 cm and 20–30 cm compared with the rates (3.5–8.7 nmolN2g−1 dry soil d−1) measured at depths of 50–60 and 90–100 cm. In contrast, n-damo was mainly occurred at depths of 50–60 cm and 90–100 cm with potential rates of 0.7–5.0 nmol CO2 g−1 dry soil d−1. This study suggested the niche segregation of the anammox bacteria and n-damo bacteria in wetland soils, with anammox bacteria being active primarily in deep soils and n-damo bacteria being active primarily in shallow soils.

Vertical distribution of nitrite-dependent anaerobic methane-oxidising bacteria in natural freshwater wetland soils.

Nitrite-dependent anaerobic methane oxidation (n-damo) is a recently discovered process that is catalysed by "Candidatus Methylomirabilis oxyfera". In the present study, the vertical distribution (0-10, 20-30, 50-60 and 90-100 cm) of M. oxyfera-like bacteria was investigated in Xiazhuhu wetland, the largest natural wetland on the southern Yangtze River (China). Phylogenetic analyses showed that group A of M. oxyfera-like bacteria and pmoA genes occurred primarily at depths of 50-60 and 90-100 cm. Quantitative PCR further confirmed the presence of M. oxyfera-like bacteria in soil cores from different depths, with the highest abundance of 5.1 × 10(7) copies g(-1) dry soil at depth of 50-60 cm. Stable isotope experiments demonstrated that the n-damo process occurred primarily at depths of 50-60 and 90-100 cm, with the potential rates ranging from 0.2 to 14.5 nmol CO2 g(-1) dry soil d(-1). It was estimated that the methane flux may increase by approximately 2.7-4.3% in the examined wetland in the absence of n-damo. This study shows that the deep wetland soils (50-60 and 90-100 cm) are the preferred habitats for M. oxyfera-like bacteria. The study also highlights the potential importance of these bacteria in the methane and nitrogen cycles in deep wetland soils.

Evaluation of an optimal extraction method for measuring d-ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) in agricultural soils and its association with soil microbial CO2 assimilation

Abstract Assimilating atmospheric carbon (C) into terrestrial ecosystems is recognized as a primary measure to mitigate global warming. Ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) is the dominant enzyme by which terrestrial autotrophic bacteria and plants fix CO 2 . To investigate the possibility of using RubisCO activity as an indicator of microbial CO 2 fixation potential, a valid and efficient method for extracting soil proteins is needed. We examined three methods commonly used for total soil protein extraction. A simple sonication method for extracting soil protein was more efficient than bead beating or freeze–thaw methods. Total soil protein, RubisCO activity, and microbial fixation of CO 2 in different agricultural soils were quantified in an incubation experiment using 14 C-CO 2 as a tracer. The soil samples showed significant differences in protein content and RubisCO activity, defined as nmol CO 2 fixed g −1 soil min −1 . RubisCO activities ranged from 10.68 to 68.07 nmol CO 2 kg −1 soil min −1 , which were closely related to the abundance of cbbL genes ( r = 0.900, P = 0.0140) and the rates of microbial CO 2 assimilation ( r = 0.949, P = 0.0038). This suggests that RubisCO activity can be used as an indicator of soil microbial assimilation of atmospheric CO 2 .