Wheat Plants Reduce N₂O Emissions from Upland Soil Subject to Transient and Permanent Waterlogging

Abstract

Climate change is expected to increase the frequency of extreme soil moisture events, such as winter waterlogging followed by spring drought, particularly in temperate regions of Europe, North America and Northeast China. While N₂O emissions from paddy soils under waterlogging and subsequent drainage have been widely studied, knowledge of upland arable soils under wheat cultivation remains limited. We hypothesized that: (1) in upland soils, combined waterlogging and drought reduces N₂O emissions compared to continuous waterlogging, and (2) plant presence mitigates soil nitrate accumulation and N₂O emissions across different moisture regimes. A greenhouse experiment was conducted using intact upland soil cores with and without wheat under four moisture treatments: control (60% water-holding capacity, WHC), drought (30% WHC), waterlogging, and waterlogging followed by drought. Daily and cumulative N₂O fluxes, soil mineral nitrogen (NH₄⁺-002DN and NO₃⁻-N), and total nitrogen uptake by wheat shoots were measured. Prolonged waterlogging resulted in the highest cumulative N₂O emissions, whereas the transition from waterlogging to drought triggered a sharp but transient N₂O peak, particularly in soils without plants. Wheat presence consistently reduced N₂O emissions, likely through nitrate uptake, which limited substrate availability for incomplete denitrification. Moisture regimes strongly affected nitrate dynamics, with drought promoting nitrate accumulation and waterlogging enhancing nitrate loss. These findings highlight the vulnerability of upland soils in regions prone to seasonal moisture extremes. Effective management of soil moisture and nitrogen, including the promotion of plant growth, is essential to mitigate N₂O emissions and improve nitrogen use efficiency under future climate scenarios.

1. Introduction

Greenhouse gases, particularly carbon dioxide (CO₂) and nitrous oxide (N₂O), are a major environmental concern due to their role in global warming and climate change. The global warming potential of N₂O is 273 times higher than that of CO₂, which makes its mitigation crucial in agricultural ecosystems [1]. In this context, N₂O accounts for 6.4% of the total pool of greenhouse gases, and agriculture contributes 74% of total anthropogenic N₂O emissions [2]. Soil moisture is one of the key factors that regulates N₂O emissions in soil as it controls the balance between nitrification and denitrification. Nitrification is an aerobic process where ammonium is oxidized to nitrate, releasing N₂O as an intermediate. On the contrary, denitrification occurs under anaerobic conditions, reducing nitrate to gaseous forms of N such as N₂O and dinitrogen (N₂). Therefore, soil moisture determines which pathway dominates the N₂O production, whereas nitrification prevails at moderate moisture levels, while denitrification is the dominant process under waterlogged soils, often leading to large N₂O pulses [3,4,5]. Climatic extremes such as heavy rainfall can significantly amplify N₂O emissions [6]. Climate change has brought unprecedented challenges for global agriculture, and regions with a temperate climate are no exception. The hydrological cycle is being altered by increasing temperatures, uneven rain patterns and extreme weather events, significantly impacting soil moisture regimes and crop productivity [7]. Extreme weather events such as heavy precipitation and droughts have become more frequent globally over the last 30 years [8]. Estimates indicate that Germany will face increased rainfall in winter, causing waterlogging, followed by hot and dry summers, resulting in droughts [9,10]. These events have been estimated to increase by 35% until 2050 in Europe [11]. This dual stress scenario of waterlogging followed by drought carries a unique challenge for arable soils under changing climate conditions. Similar sequences of flooding drying have been extensively studied in paddy soils, particularly through alternate drying and wetting practices (intermittent irrigation), where they have been shown to alter microbial processes including nitrification and denitrification, nitrogen dynamics and greenhouse gas fluxes [12,13,14]. In contrast, comparable studies that explicitly examine waterlogging followed by drought are scarce in upland arable soils, with most studies focusing on paddy soils. This highlights a knowledge gap in understanding how such dual stress scenarios driven by climate change affect upland soils, which are now more frequently exposed to such hydrological extremes [15,16,17].

The dynamics of saturation-induced N₂O fluxes are complex. Prolonged saturation increases N₂O emissions by stimulating denitrifying communities such as Pseudomonas and Paracoccus, while the re-oxygenation phase can favor nitrification. In addition, the conditions at the beginning of soil drainage are prone to extreme N₂O emissions peaks because of excessive soil moisture and increasing gas diffusion within soil and the soil–atmosphere interface [18]. The presence of plants can reduce these emissions throughout the flooding and drying cycle by reducing soil mineral N pools [19]. Several incubation studies and field reports suggest that the flooding–drying (FD) cycle triggers significantly amplified N₂O fluxes compared to prolonged flooding (PF) alone. This disturbance sequence effect was particularly relevant in grasslands, where FD increased N₂O emissions by over two orders of magnitude [20]. Short wet dry alterations further show that rewetting dry soils can produce rapid bursts of N₂O emissions driven by denitrifying microbes within minutes to hours, underscoring the importance of budgeting N₂O emissions from sequential stresses [21,22]. Landscape-scale observations show that temporary flooded patches can act as N₂O emissions hotspots, weeks after the water recedes [23]. In poorly drained waterlogged soils, the choice of crop or cover crop, specifically its rooting system, mediates the N₂O:N₂ ratio. This, in turn, determines the magnitude of the N₂O pulse that follows post-drying [24]. These findings indicate the importance of investigating sequential hydrological stresses in upland soils.

The cultivation of crops has been demonstrated to exert an influence on these dynamics. The presence of vegetation influences soil N transformations, including denitrification and associated N₂O emissions. Plant roots contribute to these changes by taking up mineral N, changing soil oxygen concentrations and providing labile carbon via rhizodeposition [25]. Certain plant species release root exudates that suppress nitrifying microorganisms, a process known as biological nitrogen inhibition (BNI), which can reduce nitrate losses and N₂O emissions [26]. Plants can also favor the complete reduction of N₂O by affecting soil carbon availability [27]. In general, vegetation reduces N₂O emissions relative to bare soil by taking up nitrate and thus limiting the substrate availability for denitrification [28]. However, the rhizosphere is a zone of high microbial activity where root exudates can stimulate denitrifiers and lead to increased N₂O emissions under favorable conditions regarding nitrate and water availability [29]. N₂O emissions increase during early plant growth and ripening stages due to low N requirements and root exudation of labile C [30]. Furthermore, foliar endophytes (ammonia-oxidizers) can add up to 1.21 Tg N₂O-N/year to global N₂O emissions [25]. Thus, the overall impact of plant presence is context-dependent, affected by factors such as plant growth stage, root biomass and soil moisture fluctuations.

In this context, it was hypothesized that (1) in upland soils, combined waterlogging and drought stress result in lower N₂O emissions compared to continuous waterlogging stress; and (2) wheat plants reduce soil nitrate concentrations and subsequent N₂O emissions irrespective of soil moisture conditions. The present study investigated the effect of the combined stress of waterlogging followed by drought compared to continuous waterlogging, drought, and optimum soil moisture on N₂O emissions from a cultivated upland soil. For that, intact soil cores were used in a greenhouse setting to replicate field conditions and also to investigate the influence of plants on soil N. This research aimed at providing new insights into how upland soils respond to hydrological stresses in terms of mineral N transformations and N₂O emissions, and how plants impact these processes.

2. Materials and Methods

2.1. Soil Characteristics

Intact soil cores were collected for the experiment from the Kiel University’s experimental farm Hohenschulen (54°18′ N, 9°58′ E), 24,239 Achterwehr, Germany. To do so, the plastic tubes were both inserted into and extracted from the soil using a tractor (John Deere, Deere & Company, Moline, IL, USA) equipped with a front loader. The tubes were 58 cm long with a diameter of 16 cm. The reason for using undisturbed soil was to replicate field conditions within a controlled environment. The soil had a sandy loam texture (clay 13%, silt 29%, and sand 58%) and was classified as Luvisol, having the following properties: a pH of 6.5, a bulk density of 1.37 g cm⁻³, a total organic C of 1.5% and total N of 0.1%, and a water holding capacity (WHC) of 37% [31,32].

2.2. Experimental Design

The tubes were sealed at their bottom, except for a small opening (3.5 cm) covered with mesh for water drainage. The top of the tubes was kept open except for the time of gas sampling. The study comprised four different moisture levels, each with and without plants. The experiment was performed in a greenhouse at a temperature of 16° at night and 22° during the daytime with a photoperiod of 16/8 h (day/night). The relative humidity was kept at 85% and 65% at night and day, respectively. The treatments are described in

Table 1. Description of the treatments and their respective abbreviation and water holding capacities implemented in this study.

Treatments	Abbreviation	Water Holding Capacity (WHC, %)
Control	60% WHC	60
Drought	30% WHC	30
Waterlogging	WL	100
Waterlogging followed by drought	WL + D	100→30

Note: All the treatments were implemented in an upland soil with (+Plants) and without plants (−Plants).

.

The treatments were replicated four times, both in with and without plants. The wheat (Triticum aestivum L.) variety KWS Scirocco was sown. Wheat was selected as the test crop because it is one of the most widely cultivated upland cereals in temperate regions such as northern Europe and represents typical non-leguminous systems that rely entirely on soil and fertilizer-derived nitrogen. Unlike legumes that form symbiotic associations with nitrogen-fixing rhizobia, wheat does not fix atmospheric nitrogen, making it an ideal model crop to study soil-derived nitrogen transformations and N₂O emissions under varying moisture regimes. The use of wheat thus allows a clearer assessment of how hydrological stresses alone influence soil N dynamics without the confounding effects of biological nitrogen fixation.

The soil fertilizers were applied in all the pots (including treatments without plants) at the following rates, macronutrients (mg kg⁻¹): N = 476.7 (split dose half on 25th day, other half 45th day), K = 446, p = 407, Mg = 557.7, micronutrients (mg kg⁻¹): Fe = 19.2, Cu = 6.6, Zn = 11, Mn = 15.4, B = 9.5, and Mo = 0.61. The plants were grown for two months at 60% WHC prior to treatment application to ensure proper establishment and enhance their ability to withstand subsequent waterlogging and/or drought. The treatments were established on the 61st day, and the gas sampling was started the next day and continued until the end of the experiment. The plants were harvested on the 100th day, and the last soil sampling was performed at the 102nd day at the end of the experiment (Figure 1). The pots were monitored for soil moisture every day to maintain 60% WHC and 30% WHC. The waterlogging in the pots was implemented by placing the pots in large containers filled with water. The water entered the pots from the holes at their bottom by capillarity till waterlogging was reached. The waterlogging treatment was kept this way until the end of the experiment. The “waterlogging followed by drought” treatment was kept under waterlogging for one week after waterlogging establishment, and then the pots were removed from the containers filled with water on the 69th day, allowing natural drainage. The treatment achieved drought in 10 days.

2.3. Gas Sampling

A new syringe was used for gas sampling each time, and gas samples were taken at 0, 20, 40, and 60 min. The pots were left open the whole time and only closed at the time of gas sampling. The wheat plants were carefully folded inside the plastic tubes, and lids were placed on top of the pots to collect gas samples. The gas sampling was performed every third day during the experiment. The collected gas samples were analyzed using a gas chromatograph (Agilent 7890A GC, Agilent Technologies, Santa Clara, CA, USA) on the same day of gas sampling. N₂O concentrations were measured using an electron capture detector adjusted at 300 °C with N₂ as carrier gas. The gas chromatography was calibrated with certified gas standards for each measurement. The rate of N₂O emission was calculated based on the linear relation between N₂O concentration in the enclosed pot headspace and time. The following equation was employed to calculate N₂O-N emission rates:

EN₂O-N (µg h⁻¹ m⁻²) = (R × 60 × Vgas × AR)/(Area × Vm) × 2

where EN₂O is the emission rate of N₂O calculated in micrograms per hour per m²; R stands for the rate of N₂O emissions from each pot (ppm min⁻¹);

-

Vgas is the volume of gas in each pot (dm⁻³)

-

Area is the pot area (m²)

-

AR is the atomic mass of nitrogen (14 g mol⁻¹)

-

Vm is the molar volume of gas at 25 °C

Linear interpolation between adjacent sampling points was used to estimate emissions for the non-sampling days. Subsequently, total N₂O-N emissions were calculated by summing the estimated daily emissions over the entire incubation period.

2.4. Determination of Ammonium and Nitrate Concentrations in Soil

The soil samples for nitrate and ammonium analysis were taken from all pots, at the start of the experiment (Figure 1), in the middle of the experiment (day 65th), and at the end of the experiment (day 102nd). This was performed by inserting a soil auger at 15 cm soil depth. The soil mineral N analysis was performed by taking 10 g of fresh soil, mixing it with 1 mol dm⁻³ KCl solution (1:4) and shaking it for an hour. After filtering, the extracts were stored at 4 °C until analysis. A continuous flow analyzer of Skalar (San++ Automated Wet Chemistry Analyzer-Continuous Flow Analyzer (CFA), Breda, The Netherlands) was used to measure ammonium and nitrate concentrations. Additionally, a subsoil sample was taken and oven-dried for 24 h at 105 °C to calculate water content.

2.5. Plant Harvest and Total N

The plants’ above-ground biomass was harvested on the 100th day of the experiment. Then, the samples were oven-dried at 65 °C until constant weight, and the dry weights were recorded. The samples were ground and then milled into a fine powder in a ball mill (MM200, Retsch GmbH, Haan, Germany) for total N analysis, using an NC analyzer (Thermo Scientific, Flash EA 1112 series, Waltham, MA, USA).

2.6. Statistical Analysis

Data normality was verified using the Shapiro–Wilk test, and a two-way ANOVA was performed (p ≤ 0.05). In case of significant differences, the means were compared using Tukey’s HSD (Honestly Significant Difference) test (p ≤ 0.05). All the analyses were performed with the Statistics 10 software.

3. Results

3.1. N₂O Daily Emissions

The treatments without plants exhibited a higher rate of N₂O emissions as compared to the treatments with plants (Figure 2). Moreover, the N₂O emissions were higher at 60% WHC as compared to 30% WHC in both with plant and without plant treatments (Figure 2A). The largest daily N₂O emissions were found in WL treatments. A large peak of N₂O emissions was observed in the WL + D treatment when the waterlogging was suppressed, and natural drainage began. This peak was more prominent in WL + D treatment without plants than that observed for WL + D with plants (Figure 2B).

3.2. Total N₂O Emissions

The treatments without plants resulted in a substantial increase in N₂O emissions across all moisture treatments (Figure 3, p ≤ 0.05). In comparison to treatments with plants, emissions without plant treatments were 189% higher under 60% WHC, 48% higher under 30% WHC, 161% higher under WL and 218% higher under WL + D (p ≤ 0.05).

In treatments with plants, N₂O emissions decreased 90% under 30% WHC and increased 3732% under WL, and by 1984% under WL + D when compared with 60% WHC (p ≤ 0.05). The respective increases were even more pronounced under treatments without plants, with 3359% higher under WL, 2193% higher under WL + D, and 95% lower under 30% WHC compared to 60% WHC (p ≤ 0.05). The trend for emissions in treatments with plants from highest to lowest was observed as WL > WL + D > 60% WHC > 30% WHC (p ≤ 0.05). The same trend was observed in treatments without plants but with considerably higher N₂O emissions (Figure 3, p ≤ 0.05).

3.3. Ammonium and Nitrate in Soil

Ammonium concentrations were lower than nitrate concentrations in all the treatments (Figure 4). After N fertilizer application on 20th and 40th days, ammonium concentration increased until the 65th day of the experiment and subsequently declined. All the treatments followed this same trend for ammonium with slight variations in concentration among treatments (Figure 4A–D). Moreover, the treatments with and without plants showed similar ammonium concentrations during the whole experiment (Figure 4).

The nitrate concentration in the 60% WHC treatment with plants increased until the middle of the experiment and then declined towards the end, while in the treatment without plants at 60% WHC, the nitrate concentration increased steadily from the start till the end. The drought treatment without plants showed the highest nitrate concentration (Figure 4F). The lowest nitrate concentration was observed in the treatment with plants under waterlogging (Figure 4G). In the waterlogging + drought treatment, soil nitrate concentrations were initially similar with and without plants but diverged towards the end, decreasing with plants, and increasing without plants (Figure 4H).

3.4. Nitrogen Uptake by Wheat Plants

The N uptake was significantly highest in the 60% WHC treatment. The 60% WHC values were 73% higher than those in 30% WHC, 65% higher than under WL, and 32% higher than under WL + D treatment. Furthermore, N uptake levels in the 30% WHC, WL, and WL + D treatments were statistically similar to each other (p ≤ 0.05) (Figure 5).

4. Discussion

The present study focused on the response of upland soils to the sequential stress of waterlogging followed by drought compared to continuous waterlogging, drought, and optimum soil moisture (60% WHC) on N cycling, especially N₂O emissions. These insights are particularly relevant with regard to climate change projections of intensified winter rains and extended summer droughts in temperate climate regions [33].

4.1. Effects of Permanent and Transient Waterlogging on N₂O Emissions

Continuous waterlogging (WL) consistently led to the highest cumulative N₂O emissions, whereas transient waterlogging followed by drainage (WL + D) induced a sharp but short-term N₂O peak (Figure 2 and Figure 3), which was generally lower in planted soils. This pattern aligns with earlier findings by Aulakh et al. [34], Kemmann et al. [24], and Ni et al. [31], who demonstrated that prolonged anoxic conditions favor denitrification-driven N₂O production, while subsequent re-aeration triggers transient emission peaks caused by rapid microbial processing of accumulated nitrate and labile carbon [35].

Globally, the combination of winter waterlogging and subsequent spring drought is becoming more frequent under climate change, not only in Europe but also in North America and Asia [7,12,36]. Extreme winter rainfall followed by spring drying has been reported to reduce cereal yields by 15–25% across several growing seasons [37]. In Asian rice systems, prolonged submergence can cause yield losses of up to 30%, particularly when followed by drought stress during reproductive stages [38]. Consistent with these broader patterns, our results showed that N uptake by wheat plants was markedly lower under WL, WL + D, and 30% WHC compared to 60% WHC (optimal moisture), demonstrating the negative effects of hydrological stress on plant nutritional status and growth.

For Europe, precipitation extremes are projected to increase by 30–35% by 2050 [39], likely amplifying both crop yield variability and N₂O fluxes. These observations illustrate that plant stress and soil biogeochemistry are interconnected across climatic regions, emphasizing the global relevance of this phenomenon.

At the physiological level, reactive oxygen species (ROS), such as superoxide (O₂•⁻) and hydrogen peroxide (H₂O₂), are generated in plant tissues under waterlogging due to impaired mitochondrial electron transport and altered antioxidative enzyme activity. Excessive ROS accumulation causes oxidative damage to lipids, proteins, and nucleic acids, reducing photosynthetic efficiency, root growth, and nutrient uptake [40,41]. These stress-induced impairments translate into yield losses reported for several crops under transient and prolonged waterlogging as indicated by Wollmer et al. [42,43] and Hussain et al. [44,45].

ROS-mediated plant stress directly influences soil nitrogen cycling and N₂O emissions. Oxidative root damage reduces root biomass and radial oxygen loss (ROL) into the rhizosphere, thereby decreasing micro-oxic zones that enable complete denitrification to N₂ [25,35]. As a result, partial denitrification pathways dominate, enhancing N₂O accumulation [20,35,36]. In addition, decomposition of damaged root tissue and changes in root exudate patterns increase the availability of labile carbon for heterotrophic denitrifiers, further stimulating N₂O production during post-waterlogging re-aeration [25,38].

Reduced oxygen availability also suppresses nitrifier activity, leading to nitrate accumulation in upper soil layers [35,36]. Upon drainage and re-oxygenation, rapid nitrification–denitrification cycles occur, generating distinct N₂O peaks that can contribute substantially to annual greenhouse-gas budgets [22,28,36]. This mechanistic linkage shows how plant physiological responses under waterlogging—particularly ROS formation—are closely tied to soil microbial dynamics and gaseous nitrogen fluxes.

The presence of actively growing plants mitigated N₂O emissions via several mechanisms: (1) plant nitrate uptake reduced available substrate for denitrifiers, and (2) radial oxygen loss from roots promoted more complete denitrification to N₂ rather than N₂O [25,35,36]. The extent of this mitigation depended on plant species, root architecture, and stress tolerance, demonstrating the tight coupling between plant physiology and soil microbial processes.

Overall, these results confirm that N₂O emissions under waterlogging are not solely governed by soil properties but are strongly modulated by plant stress physiology, ROS dynamics, and their interaction with microbial nitrification and denitrification pathways. The observed differences between permanent and transient waterlogging (Figure 2 and Figure 3) therefore reflect both the temporal dynamics of soil redox potential and plant-mediated modification of the rhizosphere environment. Integrating plant physiological stress responses with microbial N dynamics is essential for accurate prediction of N₂O fluxes under climate-change-driven waterlogging events.

4.2. Ammonium and Nitrate Dynamics

The ammonium and nitrate concentration trends in this study highlight the differential effects of moisture regimes and wheat plant on N dynamics. The increase in ammonium concentration followed by fertilizer application and subsequent decline towards the end of the experiment reflects an active nitrification process (Figure 4A–D). The highest ammonium levels were observed under drought, suggesting lower nitrification rates due to low moisture, which restricts microbial activity (Figure 4B) [35].

Nitrate concentrations, however, showed more sensitivity towards different treatments and plant presence. In the control 60% WHC, nitrate concentration increased steadily without plants but decreased with plants towards the end of the experiment, likely due to plant uptake (Figure 4E) [25,36]. The nitrate accumulation was highest under drought conditions in the absence of plants (Figure 4F). This can be explained by the combined effects of reduced microbial activity and the limited availability of electron donors, which together suppress denitrification under dry soil conditions. As described by Firestone and Davidson [46], low soil water content maintains predominantly aerobic microsites, thereby restricting the activity of denitrifying microorganisms and limiting gaseous nitrogen losses. Similarly, Yi et al. [47] demonstrated that during the drying phase following flooding in a paddy soil, nitrate accumulated due to the shortage of available electron donors and reduced microbial activity, whereas rewetting reactivated denitrification and decreased nitrate concentrations. These findings collectively support the interpretation that restricted anaerobic zone formation under drought promotes nitrate preservation rather than reduction.

In contrast, nitrate concentrations remained low under waterlogged conditions, especially in soil with plants, pointing to enhanced denitrification and root-mediated N uptake (Figure 4G) [24]. Under the waterlogging followed by drought treatment, nitrate levels initially increased for with and without plants treatments due to fertilization and post-waterlogging nitrification. However, towards the end of the experiment, as drought was established, nitrate decreased in soil with plants likely due to root uptake, while it continued to increase in soil without plants, where uptake was absent and microbial activity was limited (Figure 4H) [24,36]. This divergence shows the critical role that plants play in regulating nitrate dynamics during the transition from wet to dry conditions.

Overall, the observed dynamics demonstrate that the balance between ammonium and nitrate in soil depends on the interaction of moisture regime, microbial activity, and plant presence. Drought favors nitrate accumulation through suppressed microbial turnover, whereas continuous waterlogging leads to nitrate depletion due to inhibited nitrification and enhanced denitrification [22,28,36]. These results emphasize the importance of considering moisture-driven redox changes and plant–microbe interactions when assessing nitrogen cycling and associated N₂O emissions under variable climatic conditions.

4.3. Implications for Climate Change Adaptation

The current findings emphasize that not only the duration but also the timing of soil saturation greatly influences N₂O emissions as well as nitrogen fluxes in the soil of upland cropping systems. This is consistent with current research findings, indicating that the timing of waterlogging events is a key factor in the severity of stress effects on the yield of various crop species. For example, Wollmer et al. [42,43,48] and Hussain et al. [44] have demonstrated that waterlogging impairs the vegetative and generative phases, thereby significantly reducing yield, depending on the plant species (e.g., wheat, rapeseed). The reason for this is that hypoxia/anoxia during the most sensitive stages of development leads to irreversible losses in productivity and nutrient uptake. Moreover, these findings highlight that the issue of timing and severity of waterlogging events should always be considered when defining experimental and management-related scenarios.

In a recent study, Hussain et al. [44] investigated the impact of waterlogging stress on the performance of hybrid and inbred winter wheat varieties. The experimental design involved exposing the plants to waterlogging stress at two stages of development: the vegetative stage (BBCH 30/31) and the generative stage (BBCH 51). The results of the study indicated significant yield losses, ranging from 17–18% for the early stage to 30–31% for the late stage, in response to the waterlogging stress. These losses were accompanied by a substantial decline in sulfur uptake and an impact on grain quality. These results indicate that wheat serves as a suitable model plant for studying physiological and biogeochemical responses to temporary and permanent waterlogging. This is due to its relevance to agronomic practices and its mechanistic responsiveness to fluctuating soil oxygen conditions.

Furthermore, the reduction in stress responses through nutrient- and signal-based interventions has recently been investigated. Hussain et al. [45] reported that the combined application of melatonin and sulfur under waterlogging conditions significantly reduced markers of oxidative damage (H₂O₂, MDA), improved photosynthetic performance, and increased biomass production in Brassica napus. These results underscore the notion that biochemical strategies can complement physical and agronomic approaches by stabilizing redox homeostasis and nitrogen transformations under conditions of fluctuating soil moisture.

The implementation of these findings into climate-friendly management can be translated into several practical strategies. Improved drainage or precise irrigation can prevent waterlogging during critical growth phases and minimize the extent of N₂O pulses, similar to the phase-specific sensitivity observed by Wollmer et al. [43]. Breeding or selecting genotypes with improved waterlogging tolerance, as identified by Husain et al. [44], has the potential to reduce both yield and gaseous nitrogen losses.

The application of redox-modulating compounds or sulfur additives [45] has also been demonstrated to improve the antioxidant capacity of plants and reduce denitrification-induced N₂O formation.

These adaptation strategies are consistent with the objectives of the European Green Deal “Farm to Fork” initiative and the EU Nitrates Directive, which aim to reduce reactive nitrogen losses by 50% by 2030.

Subsequent research endeavors should integrate these findings from greenhouse and controlled environmental conditions with long-term field trials that encompass hydrological monitoring, plant diversity, and the analysis of microbial communities. The integration of these data with process-based models and sensor technologies will facilitate more precise estimation of annual N₂O fluxes and the identification of sustainable management thresholds. The integration of physiological, biochemical, and agronomic approaches provides a holistic framework for developing resilient cropping systems that maintain productivity and minimize greenhouse gas emissions under increasingly variable climatic conditions.

5. Conclusions

The present study offers clear evidence that soil moisture dynamics and the presence of plants have a significant impact on N₂O emissions and nitrogen cycling. Firstly, the findings of this study demonstrate that the waterlogging–drought process results in reduced overall N₂O emissions in comparison to continuous waterlogging. While the transition from anaerobic to aerobic conditions triggered a sharp, transient N₂O pulse, the cumulative emissions remained lower than under permanent water saturation. This finding underscores the notion that continuous waterlogging sustains conditions conducive to denitrification, whereas drought reduces gaseous losses by constraining both denitrification and nitrification processes. Secondly, the presence of plants consistently reduced soil nitrate concentrations and N₂O emissions across all moisture treatments. This finding highlights the pivotal role of plant nitrate uptake in limiting substrate availability for microbial nitrogen transformations, even under conditions of extreme moisture fluctuations.

These insights have practical implications for the management of upland agricultural systems in the face of increasingly frequent climatic extremes. The optimization of drainage, the timing of fertilizer applications, and the management of soil cover in consideration of soil moisture variability have been demonstrated to mitigate greenhouse gas emissions and nutrient losses. The findings of this study underscore the necessity of incorporating plant–soil interactions and dynamic moisture regimes into adaptive agricultural strategies that are aimed at sustainable nitrogen management.