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Nitrous oxide (N2O), a major greenhouse gas and ozone depleter, is emitted from drained organic soils typically developed in floodplains. We investigated the effect of the water table depth and soil oxygen (O2) content on N2O fluxes and their nitrogen isotope composition in a drained floodplain fen in Estonia. Measurements were done at natural water table depth, and we created a temporary anoxic environment by experimental flooding. From the suboxic peat (0.5–6 mg O2/L) N2O emissions peaked at 6 mg O2/L and afterwards decreased with decreasing O2. From the anoxic and oxic peat (0 and >6 mg O2/L, respectively) N2O emissions were low. Under anoxic conditions the δ15N/δ14N ratio of the top 10 cm peat layer was low, gradually decreasing to 30 cm. In the suboxic peat, δ15N/δ14N ratios increased with depth. In samples of peat fluctuating between suboxic and anoxic, the elevated 15N/14N ratios (δ15N = 7–9‰ ambient N2) indicated intensive microbial processing of nitrogen. Low values of site preference (SP; difference between the central and peripheral 15N atoms) and δ18O-N2O in the captured gas samples indicate nitrifier denitrification in the floodplain fen.

<p>Freezing and thawing are common phenomena and potential sources of N2O emissions in ecosystems at high latitudes. Earlier it was hypothesized that the frozen soil layer might trap the underlying production of N2O and release this as the top layer is thawed. However, newer research has found other factors playing role in the de novo emissions such as fluctuating availability of organic matter, nitrates, and ammonia, microbial activity, and changing oxic conditions of the soil. But, the variation in the abundance of genes involved in the nitrogen cycle during these events is rarely explained thus, a generally accepted theory of the impact of freeze-thaw on N2O fluxes is still missing.<br />To further investigate the relationships between physical, chemical, and microbial parameters with N2O emissions, we conducted a two-week experiment of three thaw-freeze events in March 2022 using artificial heating with electrical cables installed in collars of greenhouse gas sampling chambers conducted in a drained Downy birch peatland forest. Gas and soil samples were obtained on three non-consecutive days from these collars. Soil temperature, soil water content (SWC), NH4-N, and NO3-N were measured in the soil. Also, the abundances of functional genes involved in the nitrification (bacterial, archaeal, and comammox (complete ammonia oxidation) amoA) and denitrification (nirS, nirK, nosZI, and nosZII) were known using qPCR.<br />Our results show that artificial heating induced the thawing of the frozen top layer of soil during our experiment. The increase in soil surface temperature positively correlated with the soil water content in the top layer (R=0.58, p<0.01). N2O emissions also increased with heating and correlated with SWC (R=0.38, p<0.01). Ammonia in soil decreased and was negatively associated with N2O emissions (R=−0.28, p<0.05), suggesting active nitrification as the amount of nitrates also increased during heating. The abundance of all functional genes significantly increased during the heating except for those responsible for the consumption of N2O (nosZ genes) during<br />denitrification. Although we found evidence of both active nitrification and denitrification, the multiple regressions between N2O emissions and the proportion of different functional genes suggest that the nirK-type denitrifiers dominated in the denitrification as well as in the overall production of the N2O (p<0.001). Meanwhile, the inactivation of N2O consumers (nosZ) at thawing temperatures resulted in the emission of N2O during the thawing events in the drained peatland’s nitrogen-rich soil.</p>

<p>Due to the complexity and diversity of nitrogen cycle processes, different methods, e.g., microbiological and isotope analysis, are used to study them. Their combined application helps make the most accurate estimates of the processes occurring, which is essential for the future management of drained peatlands to mitigate soil degradation and negative atmospheric impact. Nitrification and denitrification processes in soil are the main processes behind the harmful greenhouse gas nitrous oxide (N<sub>2</sub>O) emission.</p><p>This study aimed to investigate the effect of drainage and rewetting on nitrification and denitrification processes and N<sub>2</sub>O emissions using real-time PCR and isotope methods. In the summer of 2020, the 1 m<sup>2</sup> triangle-shaped mesocosms were established to achieve varying oxygen conditions for flooding and drainage experiment in Estonia's <em>Oxalis</em> site-type drained peatland forest. In the experiments, heavy nitrogen tracers of potassium nitrate <sup>15</sup>N 98% atom (Sigma Aldrich) and ammonium chloride <sup>15</sup>N 98% atom (Sigma Aldrich) were applied to soil to amplify and get an insight into N<sub>2</sub>O production mechanisms and on its soil moisture dependence. N<sub>2</sub>O concentration was measured, and soil samples were collected six times from the study sites between October 2020 and January 2021. Besides different physical and chemical parameters measured of soil samples, quantitative real-time PCR was used to measure the abundance of bacterial and archaeal specific 16S rRNA, nitrification (bacterial and archaeal <em>amoA</em> genes) and denitrification (<em>nirK</em>, <em>nirS</em>, <em>nosZI</em> and <em>nosZII</em> genes) marker genes from the samples. Isotope composition of soil and gas samples were also measured.</p><p>This study indicates that different hydrological regimes influence nitrification and denitrification processes. Regarding control of N<sub>2</sub>O fluxes, nitrification played a major role on drained sites, and denitrification was the main process in rewetted sites, which is easily related to the oxygen content in the soils. This is supported by a higher proportion of <sup>15</sup>N-N<sub>2</sub>O in <sup>15</sup>N-NO<sub>3</sub> treatment in rewetted mesocosms. In the case of <sup>15</sup>N-NH<sub>4</sub> treatment, the highest proportion of heavy N was found in the drained mesocosms. Overall, heavy nitrogen proportion in both alpha and beta positions was higher in the N<sub>2</sub>O produced by denitrification, whereas N<sub>2</sub>O contained only one <sup>15</sup>N atom per N<sub>2</sub>O molecule. Abundances of <em>nosZI</em> and <em>nosZII</em> genes behaved differently in drained and rewetted mesocosms. Both microbiological and isotope methods showed similar results and backed each other very well, which makes either of them a perfect tool for predicting N<sub>2</sub>O emissions.</p>

<p>Nitrous oxide (N2O) is a major greenhouse gas whose presence in atmosphere is continuously increasing. Hence it’s important to understand its production and consumption mechanisms. During the summer of 2020, we conducted lab experiments using heavy nitrogen tracers of Potassium Nitrate 15N 98% atom (Sigma Aldrich) and Ammonium Chloride 15N 98% atom (Sigma Aldrich) under different moisture conditions to get an insight into N2O production mechanisms and on their dependence on soil moisture. We applied the tracer to peat samples (Kärevere, Estonia) placed in 36 (12 control, 12 nitrate treatment & 12 ammonia treatment) plastic buckets (radius-10cm, height-20cm) with soil height of 10 cm and a 10 cm head space left for gas collection. We installed oxygen sensors, water table indicators and temperature sensors on all buckets. We focused on studying physical conditions (soil oxygen, temperature, water table and soil moisture), gas (N2O) emission data, soil chemistry, gas isotope 15N, soil isotope and soil microbiology to get a complete picture of the processes involved in production of N2O gas. Under the ammonia treatment, emissions increased more than ten-fold which could be due to multiple processes of the nitrogen cycle in play. N2O emissions increased as the oxygen conditions shifted from anoxic (Omg/L=0) to sub-oxic (Omg/L=0.5–6) and then decreased as oxygen conditions reached the oxic (Omg/L>6) state. Furthermore, we witnessed negative site preference and 18O values during the nitrate treatment indicating nitrifier-denitrification. Under the ammonia treatment, we recorded both negative as well as high positive site preference values indicating presence of multiple production mechanisms. This was expected as ammonia triggers multiple processes in the nitrogen cycle. In some samples, we observed N2O consumption with little change in site preference as compared to the N2O producing samples. This indicates some bacterial-denitrification along with the prevailing nitrifier-denitrification. We also observed that under both treatments, heavy oxygen increased with increasing site preference. This indicates reduction of N2O (Ostrom et al, 2007) as redox supports 15N and 18O enrichments. After these lab experiments, we conducted the same experiment at a large scale in a drained peatland forest in Agali, Estonia. In this experiment, we established 1m2 triangle-shape mesocosms using experimental draining and flooding to achieve varying oxygen conditions. Preliminary results of qPCR analysis of N-cycle control genes support the domination of ammonia oxidation and denitrification as sources of N2O.</p>