This research was conducted within Rasht County, Guilan Province, Iran, spanning an area between Limochah (49°48’36.54”E - 37°19’49.54”N) and Bala Mahalleh-ye Gafsheh (49°48’36.99”E - 37°20’7.56”N) (Fig. 1a,b ). The mean annual precipitation in this region totals approximately 1,359 mm. The soil in this area falls under the Aquept suborder, characterized predominantly by clay and silty clay loam textures.

Two specific sites (Fig. 1c) were selected for the study, representing land leveling activities that occurred 2 and 5 years prior to the study (L₂ and L₅, respectively). Additionally, adjacent unleveled (traditional) paddy fields were chosen as control sites (U₂ and U₅, respectively). In L₂, clover served as a cover crop (green manure) during the last growing season. Four leveled and four unleveled parcels were selected within the two aforementioned study sites considering soil type, rice varieties, soil management practices, microclimate conditions, etc.

Immediately following the rice harvest, five composite soil samples were systematically collected in August 2018 from the surface layer (0-20 cm) of each of the 16 parcels using a randomized sampling design. This process yielded a total of 80 soil samples, which were then transported to the laboratory. A portion of these samples underwent air-drying and sieving through 2 mm screens for the assessment of chemical properties, including pH, electrical conductivity (EC), available P, and potassium (K). Another subsample was ground through a 0.5 mm-mesh sieve for the determination of organic carbon (OC) and N. Furthermore, a separate set of samples was specifically preserved for subsequent biological analyses and stored refrigerated at 4°C.

Soil biological indexes, including metabolic (q_CO2) and microbial (q_mic) quotients, were calculated; q_CO2 is defined as the amount of C-_CO2 produced per unit of soil microbial biomass per unit of time (Anderson & Domsch, 1993); meanwhile, q_mic is the ratio between microbial biomass carbon and SOC (Powlson et al., 1997).

All data underwent descriptive statistical analysis and were assessed for normality using the non-parametric Kolmogorov-Smirnov test at a significance level of 5%. Paired t-tests were performed at probability levels of 0.1%, 1%, and 5% using SPSS version 8.2 to determine the overall impact of land leveling and the combined effects of land leveling and cover cropping on the datasets.

The relative change in the studied properties of leveled soils compared to unleveled ones (RCX_i) was calculated as a percentage using Eq. (2). Here, X_B and X_C represent the values of each studied parameter in the leveled and unleveled soils, respectively.

The findings of this study revealed notable differences in various soil chemical and biological parameters between soils that underwent leveling for five years (L₅) and the adjacent unleveled lands (U₅).

Following five years of land leveling, the soil pH in L₅ exhibited a substantial increase (p<0.01) compared to U₅, suggesting a shift towards a more alkaline environment. In contrast, soil EC showed a significant decrease (p<0.01) in L₅ when compared to U₅ (Table 1).

The levels of OC and N in the soil notably decreased (p<0.01) due to the five-year land leveling, indicating potential changes in SOC content and nutrient availability.

The available concentration of P was notably lower (p<0.01) in L₅ compared to U₅, suggesting a potential decline in the availability of this crucial nutrient. Conversely, the exchangeable K content exhibited a significant increase (p<0.01) in L₅ compared to U₅, implying potential enhancements in the availability of this nutrient (Table 1).

Biological parameter assessment indicated that L₅ displayed significantly lower microbial respiration and biomass carbon (p<0.01) in comparison to U₅ (Fig. 2). This suggests a potential reduction in soil microbial activity and carbon storage subsequent to the land leveling process. Moreover, the biological indexes of q_mic and q_CO2 were notably higher (p<0.05) in L₅ than in U₅ (Table 2).

Analyzing the changes in soil chemical characteristics brought about by the land leveling over a 5-year period (Fig. 3) showed a little rise in soil pH of 2.72%. On the other hand, there was a noticeable decrease in the soil’s EC, OC, N, and available P, with values ranging from 12.2% to 49.6%. Exchangeable K, in particular, showed a significant rise with a value of 47.62% (Fig. 3).

The results revealed significant alterations in various soil chemical and biological parameters in soils subjected to two years of land leveling and cover cropping (L₂) compared to adjacent unleveled lands (U₂).

After two years of land leveling and cover cropping, the soil pH in L₂ exhibited a notable decrease (p<0.01) compared to U₂, indicating a shift towards increased acidity. Conversely, soil EC displayed a significant increase (p<0.05) in L₂ compared to U₂. This rise in EC contrasted with the observed alteration in soil pH (Table 1).

The combination of two-year land leveling and cover cropping led to a substantial increase (p<0.01) in OC and N in the soil (Table 1). However, the available P concentration was notably lower (p<0.01) in L₂ compared to U₂, suggesting a potential decrease in the availability of this essential nutrient. On the other hand, the content of exchangeable K showed a significant increase (p<0.01) in L₂ compared to U₂, implying potential enhancements in its availability (Table 1).

A slight decrease in soil pH and available P was found when the 2-year land leveling was analyzed to examine the changes in soil chemical characteristics. Conversely, there was a slight rise in SOC, N, and exchangeable K, ranging from 2.56% to 3.97%. The value of 18.8% for soil EC in particular showed a notable rise (Fig. 3).

According to the biological parameter analysis, L₂ significantly outperformed U₂ in terms of biomass carbon and microbial respiration (p<0.01) (Fig. 2). This suggests that the combined effects of cover crops and land leveling have increased soil microbial activity and carbon storage. On the other hand, soils that were leveled and covered with cover crops for a period of two years did not exhibit a significant effect on q_mic and q_CO2 from L₂ (Table 2).

A key area of research in soil science is soil chemistry, which has a big impact on soil productivity, capacity for self-purification, and nutrient balance (Renkou et al., 2020). Our study clarifies the impact of cover crops and land leveling on the chemical characteristics of soil.

Soil pH, a critical measure of soil acidity or alkalinity, plays a pivotal role in determining overall soil health and productivity. Our findings demonstrate subtle changes in soil pH at L₂ and L₅, potentially affecting nutrient availability and crop uptake. We observed an increase in soil pH and a decrease in EC after five years of leveling (L₅), consistent with observations made by Oztekin et al. (2013). In contrast, Sharifi et al. (2014) observed a decrease in soil pH concurrent with an increase in EC after a four-year period of land leveling, which aligns with the trends found in our study concerning soil pH following a two-year period of leveling combined with cover cropping (L₂). Interestingly, Parfitt et al. (2013) reported no changes in soil pH after a three-month land leveling process. The exposure of subsoil to the surface subsequent to land leveling may induce significant pH variations (Robbins et al., 1997). Consistent with our findings from the combination of land leveling and cover cropping, Lin et al. (2022) observed the highest average soil pH in non-agricultural land consolidation areas. Additionally, in consolidated areas where activities such as constructing ditches, merging plots, employing organic fertilizers, and implementing comprehensive enhancements were carried out, soil pH tended to approach neutrality. This implies that the use of organic fertilizers can diminish reliance on inorganic alternatives, aiding in effective regulation of soil pH levels.

The alteration in soil pH observed in both the leveled (L₅) and the leveled and cover cropped soils (L₂) in this study was relatively modest, showing a marginal decrease of 0.04 units in L₂ and an increase of 0.19 units in L₅ (Table 1 and Fig. 3). These slight fluctuations may have limited implications on crop yield, especially when considering the dynamic nature of soil systems and the inherent buffering capacity of agricultural soil.

The impact of five-year land leveling resulted in a significant reduction in SOC content, whereas the combination of two-year land leveling with cover cropping showed an increase (Table 1). Specifically, the prolonged five-year land leveling practice led to a noticeable depletion of SOC, whereas the combined approach of land leveling and cover cropping indicated a tendency to enhance SOC storage. This transformation is evident in the alteration of SOC content, with L₅ experiencing a decline of approximately 45.9%, while L₂ showcased a modest increase of 2.6% (Fig. 3).

The documented decrease in SOC attributed to land leveling aligns with earlier research findings (e.g. Brye et al., 2004; Parfitt et al., 2013; Sharifi et al., 2014; Li et al., 2018). Notably, SOC plays a crucial role in influencing crop growth. Consequently, the decline of SOC in L₅ indicates a significant reduction in the soil’s capacity for crop production and exacerbates the issue of global warming (Li et al., 2018). The disturbance and removal of the soil surface during intensive land leveling practices significantly affect SOC dynamics. The installation of drainage systems and subsequent improvement of soil aeration can accelerate the decomposition of OC. Shepherd et al. (2001) reported an increase in microbial decomposition of soil organic matter due to land leveling. Moreover, the disruption of soil aggregates can amplify the decomposition and mineralization of OC, as emphasized by Sharifi et al. (2014). On the contrary, the combined practices of land leveling and cover cropping (L₂) resulted in an increase in SOC content, indicating a potential improvement in both SOC levels and nutrient availability. However, the overall change in SOC was relatively modest at +2.6%. This aligns with previous research by Nascente & Crusciol (2015), which highlighted the positive impact of cover crops on enhancing SOC content. The integration and widespread adoption of cover crops in agricultural systems enhance soil characteristics through various mechanisms. These include the establishment of diversified root systems that alleviate soil compaction, the absorption and cycling of nutrients from deeper soil layers, and the promotion of rhizosphere interactions that enhance nutrient allocation within the organic fraction of the soil (Arai et al., 2018).

The alterations in N in L₅ and L₂ sites closely mirrored the observed pattern in SOC content. It is important to note that approximately 95% of soil N consists of organic N. Hence, the fluctuations in N within this study can be readily understood through changes in SOC content. A significant decrease in N was evident at L₅, while an increase was observed at L₂.

Utilizing cover crops as green manure presents an effective method to augment nutrients within a cropping system. Furthermore, leguminous cover crops offer an added advantage by enhancing soil fertility through biological N fixation. This process involves the conversion of atmospheric dinitrogen gas (N₂) into plant-accessible ammonium (NH) within legume root nodules, facilitated by symbiotic interactions with nitrogen-fixing rhizobia bacteria (Parr et al., 2011).

Our investigation revealed a significant decrease in the availability of P and a corresponding increase in exchangeable K within L₅ and L₂ when compared to their unleveled counterparts (Table 1). The reduction in available P was recorded at 2.6% for L₂ and a more notable 12.12% for L₅ (Fig. 3). Particularly, the decline in available P was more pronounced in L₅ compared to L₂ (Fig. 3), a trend that could potentially be attributed to the comparatively lower OC content found in leveled soils in L₅. Mohebbi (2014) previously reported a positive correlation between P and organic matter. Unlike K, which can be leached from the surface and accumulate in lower soil profiles, the mineralization of organic components contributes partially to fulfilling plant P requirements. The decrease in available P in leveled soils can also be linked to the lower P content observed in the exposed subsoil (Sharifi et al., 2014).

Our findings of reduced available P after land leveling are consistent with prior research (e.g., Brye et al., 2004; Parfitt et al., 2013; Sharifi et al., 2014). Conversely, the slight decrease in available P within L₂ can also be attributed to the introduction of cover crops. These crops can act as “catch crops,” retrieving less available nutrients from deeper soil horizons and enhancing fertility in the rhizosphere. Specifically, the uptake of P by cover crops can result in its release into shallower soil horizons following the decomposition of plant residues and mineralization within the microbial immobilized pool (Alamgir et al., 2012).

In both L₅ and L₂, there was a notable increase in exchangeable K compared to adjacent unleveled soils (Table 1). The alteration in the percentage of exchangeable K was more prominent in L₅ than in L₂ (Fig. 3). The detected rise in exchangeable K after land leveling is consistent with findings from studies by Brye et al. (2004) and Parfitt et al. (2013).

In the study conducted by Parfitt et al. (2013), it was noted that the average K content deviated from anticipated levels, showing an increase subsequent to land leveling. This rise is attributed to the higher presence of K in the soil resulting from the application of inorganic K fertilizers. As K is a mobile cation in soil, it was expected to leach from the surface and accumulate in the subsoil (Parfitt et al., 2013). The presence of clay minerals, particularly illite, in the subsurface horizon of argillic soils, may also contribute to enhancing the exchangeable K in leveled soils (Brye et al., 2004). However, Sharifi et al. (2014) reported a notable decrease in exchangeable K subsequent to land leveling, without identifying the specific cause for this decline.

Our findings emphasize that the five-year land leveling (L₅) resulted in a significant decrease in both microbial respiration and biomass carbon (C) compared to unleveled soils. This decline is likely due to the decrease in OC and N content. Previous research has shown that microbial biomass C decreases when soil C and N are reduced following land leveling activities (Sharifi et al., 2014). The observed drop in microbial respiration could also be linked to inadequate leveling practices, excavation procedures in leveled areas, and the absence of proper soil management protocols. Land leveling interventions have the potential to disrupt the nutrient cycle balance, resulting in reduced native bacterial populations, increased ecological risks, altered biodiversity, and shifts in microbial populations (Yu et al., 2010). In contrast, the combination of land leveling with cover cropping (L₂) led to increased microbial respiration and biomass C. This elevation might be attributed to the release of essential plant nutrients from organic matter, which acts as a primary driver for enhanced respiration. Studies have indicated that the addition of plant residues to soil elevates microbial biomass C (Zhang et al., 2010). There is a recognized positive correlation between SOC and microbial biomass C, indicating their close relationship (Xue & Huang, 2013). Parfitt et al. (2013) also reported an increase in microbial C biomass following land leveling, attributing it to the severe disturbance of the soil due to the breakdown of larger aggregates during the leveling operation. The introduction of cover cropping in the two-year leveled lands likely contributed to enhanced soil organic matter content and provided favorable aeration conditions conducive to microbial growth and activity. Despite the adverse impacts of land leveling on soil biological properties evident in the five-year land leveling areas, the introduction of cover cropping emerges as a crucial factor for the slight yet significant increase in microbial respiration and biomass C observed in L₂.

The microbial quotient (q_mic), which measures the amount of C-CO₂ produced per unit of soil microbial biomass over a specific time, provides insights into microbial activity and its potential impact on organic matter breakdown in the soil (Feketeova et al., 2021). Our findings indicate that the five-year land leveling intervention led to a 45% reduction in SOC (Fig. 3) and a 23% decrease in microbial C biomass (Fig. 2). As a result, a more substantial decrease in OC compared to microbial C biomass significantly raised the q_mic in the five-year leveled soils. The metabolic quotient (q_CO2), a ratio between microbial biomass C and SOC, serves as an indicator of microbial stress; higher values suggest unfavorable conditions for microbial growth (Xue & Huang, 2013). An elevation in q_CO2 was noted following a five-year land leveling process, contrasting with the absence of significant changes observed in areas where a two-year land leveling procedure was combined with cover cropping. The observed increase in q_CO2 within the L₅ context could potentially be linked to ecological disturbances resulting from land leveling practices, coinciding with reductions in SOC and fertility parameters such as N and available P.