Carbon and Nitrogen Mineralization of Common Organic Amendments in Olive Grove Soils

Abstract

Cover crops (CC), pruning residues (PR), and olive mill pomace (OP) are common amendments to enhance soil quality in olive groves; however, there is limited information on the dynamics of carbon (C) and nitrogen (N) during their decomposition under varying conditions. In this laboratory decomposition experiment, we assessed the C and N mineralization of CC, PR, and OP at application rates of 2 and 5 mg C g⁻¹ in soils with varying organic C levels, both with and without the addition of NO₃⁻. The results indicated that C and N mineralization patterns varied significantly between amendments, although the predicted remaining C after one year was similar for CC and PR (46.7–48.9%) and slightly lower for OP (40.0%). Soil organic carbon level did not affect remaining amendment-C. The addition of N accelerated the decomposition rate of labile C by 15% but slowed down that of the recalcitrant C another 13%, with no significant effect on remaining C. Conversely, increasing the C dose led to a 13% overall reduction in amendment-C mineralization across all combinations of factors. CC decomposition released between −8 and 31% of the amendment-N by day 130, while PR and OP showed net N immobilization. Therefore, CC residues contribute to both C sequestration and N availability, while PR and OP residues can improve N retention in the olive groves while also contributing to C sequestration.

1. Introduction

Decades of agricultural intensification, characterized by the extensive use of pesticides and inorganic fertilizers, along with practices like intensive tillage and herbicide application that leave soil bare, have exacerbated soil degradation of soils devoted to olive groves in Andalusia (Spain), the world’s largest olive oil producer [1]. In this region, olive cultivation covers over 30% of the arable land and is often situated on steep slopes [2,3]. However, olive groves hold significant potential for carbon (C) sequestration and soil fertility improvement [4,5,6]. Applying farm-produced biomass, such as temporary spontaneous cover crops, pruning residues, or olive mill pomace, is one of the most technically and economically viable and scalable practices for reversing soil degradation [7].

Temporary cover crops and light pruning residues applied on top of the soil as mulching have proven effective in reducing erosion while improving soil organic carbon (SOC) levels and other soil properties [8,9,10]. Furthermore, they play a crucial role in promoting internal nutrient recycling [10,11,12,13]. Olive mill pomace is the primary byproduct of the olive oil industry, and its application in olive groves is an effective strategy to recycle the C and nutrients removed during the harvest, such as nitrogen (N) [14,15]. Despite its phytotoxicity, some studies have reported no negative effects on crop yields or soil properties following the application of raw olive pomace in olive groves; in fact, soil properties conditions improved [16].

In line with CAP 2023–27 [17] and EU environmental policies, such as the 4per1000 or the Zero Pollution Action Plan, as well as the anticipated establishment of a voluntary CO₂ credit market, the widespread adoption of spontaneous cover crops, along with the use of tree pruning and olive mill pomace, is expected in olive orchards. The dynamics of C and N during the decomposition of these amendments have been studied individually in laboratory and field studies [10,18,19,20]. However, few studies have evaluated their decomposition under the same conditions, despite their potential significance on a regional scale. Additionally, there is no available information on how the decomposition of these amendments might be influenced by soil types, application doses, and nitrogen addition.

Carbon and nitrogen mineralization is generally considered to be primarily influenced by the biochemical composition of the amendment organic matter, assuming uniform climatic and environmental conditions [21]. Key indicators of quality include the content of lignin, cellulose, polyphenols, and the stoichiometric ratio of carbon to nutrients, such as the carbon-to-nitrogen ratio (C/N ratio) [22]. High-quality organic matter, characterized by a high N content and low-molecular-weight compounds, decomposes more quickly and contributes more effectively to stable soil organic matter [23], while releasing available nitrogen. We hypothesize that the mineralization of C and N of spontaneous cover crops, tree pruning, and olive mill pomace varies significantly due to differences in nitrogen content and the C/N ratio. The addition of available N to offset the stoichiometric mismatch between the low C/N ratio of soil microorganisms and the high C/N ratio and low N content of organic amendments such as the tree pruning and the olive mill pomace may accelerate the mineralization of the amendment’s C while reducing net N immobilization. This has been previously demonstrated for other organic amendments [24]; however, the extent to which the application of available N, along with tree pruning residues and olive mill pomace, affects the dynamics of amendment-C and N mineralization remains unknown. This has significant implications for olive groves, as the expected net N immobilization during the decomposition of C-rich and N-poor materials, such as tree pruning and olive mill pomace, which could temporarily deprive trees of essential nutrients, may be alleviated.

The rate of organic amendment addition can impact not only the decomposition of the added C but also that of native SOC, potentially through the priming effect [25]. There is no clear consensus on how different rates of organic C inputs affect C and N mineralization, as studies have reported both significant reductions [26] and proportional increases in C decomposition [27]. High doses of high C/N ratio of amendments, such as olive tree pruning and olive mill pomace, can limit microbial activity due to N scarcity, thereby reducing the decomposition of organic materials. Since olive farmers can control the annual input of organic C through cover crops, tree pruning, and olive mill pomace management, it is important to investigate how varying doses of these C sources influence C and nitrogen mineralization, and thus organic C accumulation and net N mineralization.

We hypothesize that (i) cover crop residues offer the best balance between C sequestration and N availability among the amendments studied; (ii) higher native SOC stimulates mineralization of newly added material, increasing the amount of N available in the soil but hindering SOC accumulation; (iii) higher amendment doses slow down the C mineralization process, leading to greater C storage; and (iv) nitrate addition accelerates C mineralization, particularly during the decomposition of tree pruning and olive mill pomace.

To test these hypotheses, we conducted a controlled environment experiment in which cover crop residues, tree pruning residues, and olive mill pomace were applied as amendments at two different C doses, both with and without nitrate, to two soils from commercial olive groves with contrasting levels of SOC. We assessed their effects on C and N mineralization through CO₂ production and inorganic N content.

2. Materials and Methods

2.1. Samples Collection and Chemical Analyses

Samples of soils with contrasting levels of SOC were collected in two adjacent 30–50-year-old olive groves located in Ubeda (Spain). The soil with higher levels of SOC (SHC, hereafter) has been organically managed for more than 10 years, without any external fertilizer input. In the SHC farm, a spontaneous cover crop grows annually on all the soil surfaces, and every year the soil of the inter-row area receives alternatively shredded pruning residues or raw olive pomace in the late winter. On the other hand, the soil with lower levels of SOC (SLC, hereafter) is conventionally managed. In this farm, the soil is kept bare by pre-emergence herbicides in autumn and tillage in winter. Inorganic fertilization is applied, but every two years the shredded pruning residues are applied in the inter-row area.

Enough amount of top 15 cm soil (approximately 10 kg) was collected from 3 random points in the SHC and SLC olive farms. Soil samples were air-dried for 10 days and passed through a 2 mm sieve to separate the fine soil fraction and coarse fragments, whose contribution to the total weight was noted. The fine fraction was subjected to multiple analyses. The pH (1:2 soil–water v/v) was measured by potentiometry (Hannah Edge; Woonsocket, RI, USA). The soil texture (clay, silt, and sand) was estimated using the Robinson pipette method. Soil organic carbon (SOC) was determined by wet combustion following the Walkley–Black method, and soil organic matter (SOM) was estimated as 1.72 × SOC. Total nitrogen (TN) was analyzed by dry combustion with an autoanalyzer CNHS (LecoTruSpect Micro; St Joseph, MI, USA). Water field capacity (% volume) was calculated from the matric potential at −33 kPa, using the Richards membrane method, and from bulk density. Cation exchange capacity (CEC) was determined by the ammonium acetate method (pH 7) using atomic absorption spectrometry (Perkin Elmer AAnalyst 800; Shelton, CT, USA). A detailed description of these methods can be found in Pansu and Gautheyrou [28]. Alkaline phosphatase, β-glucosidase, and arylsulfatase activities were using the methods described in Page et al. [29], and a dehydrogenase assay was carried out following the method described by Schinner et al. [30]. Soil potential nitrification (transformation of NH₄⁺ to NO₃⁻) was determined following the method described by Schinner et al. [30]. In addition to the samples for these analyses, three unaltered cylinders of soil in each site were collected to calculate bulk density. Both soils were Vertic Rigosols [31]. Values of SOM, SOC, and TN were 2.0, 2.0, and 1.6 times higher, respectively, in SHC, while enzyme activities were, on average, 2.1 times higher (Table 1).

On the other hand, five random samples of aboveground biomass of the cover crop (CC) were manually collected in the SHC farm and stored in bags. Biomass samples were dried at 60 °C for 4 days and milled to powder (<1 mm) with a hammer mill. An aliquot of each sample was sent for analysis, and then a composite sample of 200 g was prepared.

Three samples of 1 kg of light pruning residues (PR) (branches < 2 cm) were freshly collected in the SLC farm. These samples were divided into branches, twigs, and leaves and dried at 60 °C for 10 days to determine the proportion of each fraction. Then, fractions were milled to powder (<1 mm) and mixed, maintaining the original proportions. Again, an aliquot of each sample was sent for analysis, and a composite sample of 200 g was prepared.

Olive mill pomace (OP) was obtained from a two-phase extraction olive mill located in Jabalquinto (Spain). A single 20 L sample was collected from three spaced discharges from a tanker truck. Three subsamples were separated after homogenization and dried for 10 days at 60 °C to determine water content. The rest was kept at 4 °C in the refrigerator until the experiment was set up. Three aliquots of the 20 L sample were sent for analysis.

Total carbon (TC) and total nitrogen (TN) of the aliquots of the three organic amendments were analyzed by an autoanalyzer CNHS (Leco TruSpect Micro; St Joseph, MI, USA). The results of the analysis are shown in Table 2.

2.2. Experimental Design

C mineralization and net N mineralization of CC, PR, and OP were studied at 2 mg C g⁻¹ soil (C2) and 5 mg C g⁻¹ soil (C5) application rates with 100 µg NO₃⁻-N g⁻¹ soil (N100) or without (N0) external addition of NO₃⁻-N. These combinations of treatments were tested in two soils: one with a higher level of organic carbon (SHC) and the other with a lower organic carbon content (SLC). Additionally, C mineralization and net N mineralization from SHC and SLC soils at both levels of nitrate addition with no organic carbon amendment was evaluated (Scheme 1).

The experiment was conducted in 0.5 L jars. In each jar, 100 g of dry weight soil from SHC or SLC was pre-incubated for 10 days at 25 °C after the addition of 10 mL of water. After pre-incubation, organic amendments (CC, or PR, or OP) at the corresponding C dose (2 or 5 mg C g⁻¹) were added to jars. The soils were then hydrated to 60% of their water holding capacity and manually homogenized. In half of the jars, hydration was performed entirely with distilled water (i.e., 0 µg NO₃⁻-N g⁻¹ soil), while the other half received the same volume of liquid but of a water solution containing the necessary nitrate amount to apply 100 µg NO₃⁻-N g⁻¹ soil. Unamended control treatments of SHC and SLC, with and without nitrate addition, were also established. Each combination of treatments was repeated in triplicate. In addition, three extra repetitions of CON N0 treatments of both soils were preincubated to determine the initial N content. The amount of each amendment applied to the 100 g of soil to meet the C dose, and the N contained in it, is shown in Table 3. The experiment was conducted at 22 °C in the dark over 130 days after residue/nitrate application, and soil water holding capacity was maintained throughout on a weight basis.

2.3. CO₂ Production and C Mineralization

CO₂-C emissions were measured in each jar on days 4, 8, 13, 22, 33, 48, 63, 76, 90, and 130 using the alkaline trap method [32]. The cumulative CO₂-C production of control treatments (CON N0 and CON N100) in the SHC and SLH at each time was divided by the total amount of SOC contained in the jars to obtain the specific SOC mineralization or SOC min_day (i.e., the proportion of SOC released as CO₂). The cumulative CO₂-C productions of control treatments (CON N0 or CON N100) were subtracted from the cumulative CO₂-C production of the amended soils to calculate the cumulative CO₂-C production derived from amendment-C. The cumulative CO₂-C production derived from the amendments at each time was divided by the C dose applied (200 or 500 mg C per jar) to calculate both the specific C mineralization (AOC min_day) and the specific C remaining (AOC rem_day) of the amendments.

The C remaining of soil-C and amendment-C in all combinations of soil types, amendment types, carbon doses, and nitrogen doses was fitted to a four parameter double exponential decay function [22], following Equation (1).

(1)${\mathrm{C} \mathrm{rem}}_{\mathrm{day}}={\mathrm{Ae}}^{-\mathrm{kt}}+\left(1-\mathrm{A}\right){\mathrm{e}}^{-\mathrm{ht}}$

where C rem_day is the remaining C from soil-C or amendment-C (proportion) at time t, A is the pool size of easily decomposable C expressed as a proportion, k is the decomposition rate constant of the labile or easily decomposable fraction (day⁻¹), (1 − A) is the pool size of hardly decomposable C expressed as proportion, and h is the decomposition rate constant of the recalcitrant or hardly decomposable fraction (day ⁻¹).

2.4. Inorganic N Dynamics and Net N Mineralization

Inorganic N was extracted from subsamples (3 g) of the fresh soil from the jars at days 13, 33, 76, 90, and 130 using 2 M KCl in a 1-to-4 ratio of soil to extractant and filtered through Whatman No. 44 filter paper. In the same way, inorganic N was extracted at day 0 in pre-incubated control treatments without external N addition (CON N0) to calculate the initial inorganic N content, which was assumed to be 100 µg NO₃⁻-N g⁻¹ higher in the control treatments with N addition (CON N100). Ammonium (NH₄⁺-N) concentration was measured by the Koroleff method [33], whereas nitrate (NO₃⁻-N) was first reduced to nitrite, and nitrite concentration was measured following the Shinn method [34]. The sum of ammonium and nitrites plus nitrate concentrations was assumed to be the total inorganic N.

The inorganic N content of control soils (CON N0 and CON N100) in SHC and SLC at day 130 were divided by the total amount of soil organic nitrogen (SON) contained in the jars (200 and 116 mg N per jar in SHC and SLC, respectively) to obtain the specific SON mineralization at day 130 or SON min₁₃₀ (i.e., the proportion of SON released as inorganic N) (Equation (2)). Moreover, the inorganic N content at day 130 of control soils (CON N0 and CON N100) minus the added N (0 and 100 µg NO₃⁻-N g⁻¹ soil in CON N0 and CON N100, respectively) was divided by the number of days of decomposition to calculate the daily net N mineralization of SON in absolute terms (Equation (3)).

(2)${\mathrm{SON} \min }_{130}\left(\mathrm{proportion} \mathrm{of} \mathrm{SON}\right)={\displaystyle \frac{\mathrm{inorganic} \mathrm{N} \mathrm{control} \mathrm{soil} \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1} \right)-{\mathrm{added} \mathrm{N} \mathrm{as} \mathrm{NO}}_3^{-} \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1}\right)}{\mathrm{total} \mathrm{SON} \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1}\right)}}$

(3)$\mathrm{Daily} \mathrm{SON}\min \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1}{ \mathrm{soil} \mathrm{d}}^{-1}\right)={\displaystyle \frac{\mathrm{inorganic} \mathrm{N} \mathrm{control} \mathrm{soil} \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1} \right)-{\mathrm{added} \mathrm{N} \mathrm{as} \mathrm{NO}}_3^{-} \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1}\right)}{130 \mathrm{days}}}$

The inorganic N content of control soils was subtracted from that of amended soils to calculate the net N mineralization of amendments. The net N mineralization of amendments was divided by the total N of amendments to obtain the specific amendment net N mineralization at day 130 or AON min₁₃₀ (i.e., the proportion of amendment-N released or immobilized as inorganic N) (Equation (4)). Moreover, the net N mineralization of amendments at day 130 was divided by the number of days decomposing to calculate the daily net N mineralization of amendments in absolute terms (Equation (5)).

(4)${\mathrm{AON} \min }_{130}\left(\mathrm{proportion} \mathrm{of} \mathrm{N} \mathrm{added} \mathrm{by} \mathrm{amendment}\right)={\displaystyle \frac{\mathrm{inorganic} \mathrm{N} \mathrm{amended} \mathrm{soil} \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1} \right)-\mathrm{inorganic} \mathrm{N} \mathrm{control} \mathrm{soil} \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1}\right)}{\mathrm{total} \mathrm{N} \mathrm{added} \mathrm{through} \mathrm{amendment} \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1} \mathrm{soil}\right)}}$

(5)$\mathrm{Daily} \mathrm{AON}\min \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1}{ \mathrm{soil} \mathrm{d}}^{-1}\right)={\displaystyle \frac{\mathrm{inorganic} \mathrm{N} \mathrm{amended} \mathrm{soil} \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1} \right)-\mathrm{inorganic} \mathrm{N} \mathrm{control} \mathrm{soil} \left({\mu \mathrm{g} \mathrm{N} \mathrm{g}}^{-1}\right)}{130 \mathrm{days}}}$

2.5. Statistical Analyses

Statistical analyses were performed using R (version 4.3.0; R Foundation for Statistical Computing, Vienna, Austria). Differences in the specific C mineralization at day 130, the daily net N mineralization, and the specific net N mineralization of soils and amendments at day 130 were assessed using ANOVA and a Tukey post hoc test at a 0.05 level of significance. The independent variables considered were amendment type, soil type, C dose, and N dose. Additionally, differences between treatments in the parameters of the double exponential decay function were tested with ANOVA and Tukey post hoc tests at a 0.05 level of significance. All variables analyzed were tested for normality and homoscedasticity.

3. Results

3.1. CO₂ Production and C Mineralization

Figure 1 shows the cumulative production of CO₂-C throughout the experiment for each amendment type, soil type, C dose, and N dose, including control treatments. The cumulative production of CO₂-C followed the expected rank based on the level of C addition (CON < C2 < C5). Comparing CON treatments, soil SHC, which has twice the SOC of SLC, emitted only 1.6 times more CO₂-C after 130 days. In consequence, SOC min₁₃₀ was significantly lower in SHC (12%) than in SLC (15%), independently of the level of N addition (Figure S1, Tables S1 and S2). For soils amended with 2 mg C g⁻¹, CO₂-C production ranged from 2615 to 3033 and from 1821 to 2089 µg CO₂-C g⁻¹ in SHC and SLC, respectively. At the higher amendment-C dose of 5 mg g⁻¹, CO₂-C production varied between 3428 and 3781 µg CO₂-C g⁻¹ in SHC and between 2693 and 3176 µg CO₂-C g⁻¹ in SLC.

Values of AOC rem_day over the experiment were significantly fitted to a four-parameter double exponential model (Figure S2). For the whole set of treatments, the pool of easily decomposable fraction of C (A coefficient) ranged between 14.46% and 34.73% (Figure 2). This pool was significantly affected by amendment type, soil type, and C dose, as well as by multiple interactions (Tables S3 and S4). The A coefficient was, on average, 14% higher in SHC than in SLC, but this effect depended on the C dose. At the low C dose, the A coefficient was higher in SHC, whereas at the highest C dose, this parameter was higher in SLC (Figure 2). On the other hand, the effect of the C dose on the pool of easily decomposable C was larger and more distinct (Figure 2). The A coefficient increased by an average of 27% at the lowest C dose compared to the highest, regardless of other factors. Additionally, the amendment type affected the A coefficient, which was generally comparable or higher for PR than for CC, with few exceptions. In contrast, the OP amendment consistently produced the lowest A coefficient across all combinations of factors.

The decomposition rate of the labile fraction (k coefficient) ranged from 0.022 to 0.156 d⁻¹. The k coefficient was significantly affected by all main effects and multiple interactions (Tables S3 and S4). The effects of soil type and amendment type on the k coefficient were the most significant. Overall, decomposition rates of amendments-labile C were on average 58% higher in SLC than in SHC. On the other hand, mean values across all soils for C doses and N doses were more than 150% higher for CC and OP than for PR. On average, the addition of nitrate-N increased the k coefficient by 16%, but this depended on the soil type. The addition of nitrate-N had the effect of increasing the k coefficient by a mean of 25% in SLC but had a negligible effect on SHC. The mean k coefficient was 16% higher at the lower C dose, but this was highly dependent on the other factors. In SLC, the k coefficient was 25% to 59% higher at the high C dose compared to the low C dose, whereas in SHC, it was only 8% to 22% lower at the high C dose.

The decomposition rate of the refractory C (h coefficient) ranged between <0.0001 and 0.0035 d⁻¹ (Figure 2) and was affected by the main factors and interactions among them (Tables S3 and S4). On average, the h coefficient was 15% smaller in SHC compared to SLC, and it was 20% smaller under low compared to high C dose. The decomposition rate of refractory C with added nitrate-N was 13% lower compared to treatments without nitrate. The mean value for PR was, on average, 17% and 51% smaller than that of CC and OP, respectively. Moreover, we found a cross-interaction between amendment type, soil type, and C dose, given that decomposition rates of recalcitrant C were similar in both soils or slightly greater in SLC for all the amendments at both C doses except for OP, which had a notably greater h coefficient in SHC at the low C dose.

The coefficients of the model indicate that, although the pool of easily decomposable C was similar for CC and PR, it decomposed significantly slower in PR. In contrast, OP showed the smallest fraction of easily decomposable C, but it decomposed at the same decomposition rate as for CC. Moreover, OP showed the highest decomposition rate of the recalcitrant C under all conditions, except in SLC C2 N0. In contrast, PR showed the lowest decomposition rate of recalcitrant C when decomposing in SHC, while it was CC that showed the lowest decomposition rate of recalcitrant C in SLC, except for SLC C5 N0 treatment. As a result, decomposition processes of PR and OP were more continuous (see Figure 1), given the greater similarity between their k and h coefficients in comparison with CC across all soil types, C doses, and N doses (Table S3).

The observed AOC rem₁₃₀ was similar across all amendments for the same C dose, except in the SHC C2 N0 treatment, where it was significantly lower for OP. Notably, differences in AOC rem₁₃₀ between soils were minimal, except for C2 N0, where OP had a significantly lower value in SHC compared to SLC. As for the coefficients of the decay model, the AOM rem₁₃₀ was consistently lower at the high C dose. In contrast, the addition of nitrate did not reduce AOM rem₁₃₀ for any of the amendments. Overall, the modeled remaining C after 1 year of decomposition was, on average, 40.0% for OP, 46.7% for CC, and 48.9% for PR (Table S5).

3.2. Inorganic N Dynamics and N Mineralization

Figure 3 shows the changes in soil mineral N content across all treatments, including controls. Throughout the 130-day experiment, the concentration of available soil NH₄⁺ was very low (<3.9 μg NH4⁺-N g⁻¹ soil) or undetectable in all treatments, so inorganic N consisted mainly of nitrites and nitrates. Overall, soil mineral N in SHC was consistently higher compared to SLC, especially during the later stages of incubation. In soils amended with CC, soil mineral N levels were higher than or similar to those in the CON treatments. For PR and OP amendments, soil mineral N concentrations were typically lower than the CON, regardless of the C or N doses (Figure 3). With these two amendments, soil mineral N decreased relative to CON treatments during the first 30 days of incubation, particularly at higher doses of C and in treatments with added nitrate-N. However, during the latter half of the incubation, soil mineral N tended to increase, especially under the low C doses. Despite this increase, soil mineral N at day 130 in PR and OP treatments remained lower than in the respective controls.

Overall, SON min₁₃₀ in SHC control soil averaged 6.3% and 4.7% for N0 and N100 treatments, respectively, whereas mean values tended to be higher in SLC: 7.9% and 5.7% at N0 and N100, respectively (Tables S6 and S7).

Mean daily AON min was influenced by soil type, amendment type, C dose, and multiple interactions among the factors (Figure 4, Tables S8 and S9). For CC, daily AON min ranged from slightly negative (SLC soil at the lower C dose) to largely positive (SHC at the higher C dose). This pattern differs from PR and OP amendment treatments, whose mean values ranged from slightly negative (lower C dose in the high SOC soil) to largely negative (higher C doses in the low SOC soil). In the CC treatments, higher C doses resulted in increased net N mineralization. Conversely, in PR and OP treatments, higher C doses resulted in more negative daily net N mineralization rates. Therefore, increasing the C dose amplified the magnitude of the daily net N mineralization, either positively or negatively. Overall, the daily net N mineralization of amendments in SHC, averaging 0.015 µg N g⁻¹ d⁻¹, was superior to that of SLC, averaging −0.259 µg N g⁻¹ d⁻¹ (Figure 4), which indicates that N release from amendments was boosted by the soil with higher SOC. AON min₁₃₀ was also influenced by soil type, amendment type, C dose, and multiple interactions among the factors. The specific net N mineralization relative to the amendment-N ranged between −8.1 and 30.6% for CC, between −22.0 and −108.7% for PR and between −2.4 and −73.4% for OP (Tables S8 and S9). Specific net N immobilization was consistently greater for PR than for OP in any combination of factors.

4. Discussion

4.1. C and N Mineralization of the Different Amendments

We anticipated that the varying quality of amendments would lead to different decomposition patterns, as the effect of substrate quality in the decomposition process is well established [22]. Indeed, CC decomposed rapidly at first due to its high N content and C quality but slowed after 30 days as the remaining material became more resistant to decomposition. In contrast, the initial daily CO₂-C emissions from PR and OP were lower than those from CC but remained steady throughout the incubation experiment. This steady rate was likely due to the medium-high C/N ratios and high lignin content in both PR and OP [22,23]. Despite the different decomposition patterns, the average AOC rem₁₃₀ was similar across all three amendments (around 63%). This finding aligns with that of Marzi et al. [35], who found higher remaining C in organic amendments with lower C/N ratios after 120 days of decomposition under laboratory conditions, and with that of García-Ruiz et al. [36], who reported similar remaining C of wheat straw across a C/N ratio of 89–202 after 84 days of incubation. Nevertheless, the projected AOC rem₃₆₅ indicates that a larger fraction of CC and PR will remain in the soil compared with OP, suggesting a greater potential for C sequestration. This outcome is consistent with the growing body of evidence indicating that amendments with higher labile C content tend to retain more residue in the soil over the long term [23]. Moreover, the remaining OP-C after 130 days, consistent with previous studies of olive pomace decomposition [19], contradicts the general consensus on the toxicity of olive mill pomace over microbes [15].

The three amendments also demonstrated distinct patterns of N mineralization. CC was the only amendment showing neutral or positive net N mineralization from the beginning of the experiment. This is attributable to its high N content and low C/N ratio, both within the thresholds required for net N mineralization [21,37]. In our 130-day incubation experiment, between 14% (SLC) and 30% (SHC) of the CC-N was released as soil mineral N. Recently, Torrús-Castillo et al. [12] found that an average of 19.5 kg N ha⁻¹ was accumulated annually in the aboveground biomass of spontaneous cover crops in 48 commercial olive orchards of Andalusia. Our findings further indicate that this N retained by cover crops during the rainy season is gradually released during the subsequent growing period, ensuring a steady supply of N for olive groves. Conversely, the application of PR and OP in olive grove soils leads to significant N immobilization, with rates ranging from 22–109% for PR-N and 2–73% for OP-N, depending on the soil type (Table S8). Similar findings were reported by Repullo et al. [4] and Gómez-Muñoz et al. [18], who observed net N immobilization during the decomposition of olive tree pruning residues in both field and laboratory experiments. Although PR and OP will finally release the nutrients extracted during pruning and harvesting to the soil, their application in spring means that temporary N immobilization must be expected during the following months. This short-term immobilization could limit the availability of N for olive trees at crucial stages, but it serves as a viable strategy for promoting N retention in olive farming. Typically, fertilization in olive orchards is applied beneath the tree canopy to maximize nutrient uptake, leaving a portion of the nutrients in inter-row soils underutilized by the trees. Combining N fertilization beneath the canopy with the application of PR or OP in inter-row areas can help meet the N requirements of trees while minimizing nitrate leaching. Still, further research is needed to establish optimal application rates to prevent excessive nutrient competition between the amendments and the crop.

Based on these findings, cover crops appear to be the most effective amendment for promoting both C sequestration and N retention during periods of high leaching risk, while also releasing this N when the tree requires it. PR and OP amendments, while also valuable for storing C and replenishing nutrients lost through pruning and harvest, tend to immobilize N during critical growth phases. To maximize their benefits and mitigate their drawbacks, careful attention should be paid to application rates and placement. Additionally, alternative management practices like composting these residues could further enhance their effectiveness.

4.2. SOC Level Effects on Amendment-C and N Mineralization

Some studies indicate that soils subjected to conservative management practices such as no-tillage or the addition of organic amendments tend to show higher SOC content and lower specific SOC mineralization [38,39]. This reduced mineralization is related to mechanisms that protect SOM from microbial activities, such as the formation of microaggregates, the chemical interaction with silt and clay minerals, and the inherent resistance of complex organic molecules [40]. For example, Pareja-Sánchez et al. [41] reported that in 13 olive groves practicing conservative management, the physically and chemically protected SOC levels were 1.3 to 2.1 times and 1.4 to 1.8 times higher, respectively, compared to similar olive groves without such management practices. Consistently, the soil SHC, which achieved a higher SOC and humic compounds content through conservative management practices (Table 1), exhibited a lower SOC min₁₃₀ and SON min₁₃₀ compared to the soil SLC.

Conversely, the state of the soil (pH, porosity, nutrient availability, and microbial community) also plays a crucial role in the decomposition of newly added organic C [22]. Based on these features, we hypothesized that SHC soils, which exhibited a higher SOC content and elevated soil microbial activities (Table 1), would facilitate faster decomposition of added amendments—indicated by higher values of coefficients k and h and lower AOC rem₁₃₀—compared to SLC soils.

The results, however, were not uniformly consistent with this hypothesis and varied depending on the type of amendment applied. Notably, in the case of CC and PR amendments, SLC increased the k and h coefficients but decreased the A coefficient. This led to similar remaining C levels for these amendments in both soils. This finding is in agreement with previous studies, such as Nett et al. [42] and Murúa and Gaxiola [43], which indicated that substrate-specific amendment history had minimal or no effect on the decomposition of recently added amendments. In contrast, a notable exception was observed with OP. The decomposition rate of OP-C in SHC soil was significantly higher than in SLC soil when applied at a low dose and without the addition of nitrate. This difference is likely attributable to the long-term regular addition of OP to SHC soils over the past decade, as sustained management practices can alter microbial community structure [44]. Such ongoing treatments in SHC may have enhanced its capacity to decompose OP more efficiently.

On the other hand, SHC caused a greater amendment-N mineralization than SLC with a similar amendment-C mineralization. This observation suggests that microbial adaptation strategies to stoichiometric imbalances are different in the two soils [45].

The results indicate that while the general history of organic matter input influences soil microbial activity and SOC levels, the specific history of individual amendments does not play a crucial role in determining decomposition rates. Our results suggest the potential for C sequestration after the addition of spontaneous cover crop residues or tree pruning in olive groves seems to be independent on the SOC level, at least in the short term. However, N is cycled faster in soils with higher SOC levels, so more N available is expected after the addition of the organic amendments in SOC-rich soils.

4.3. Amendment Dose Effects on Amendment-C and N Mineralization

To evaluate our third hypothesis, we examined the effect of two different amendment doses on C and N mineralization. Our results confirm that a higher C dose consistently reduced C mineralization when expressed per unit of amendment-C applied. For all amendments, across both soil types and nitrogen availabilities, C mineralization by day 130 was lower at the higher C dose, with reductions observed in all three parameters of the decay function. These findings contradict previous studies, which reported a proportional increase in C mineralization with higher C inputs [4,25,46], or even an enhancement due to the increased effectiveness of exoenzymes and microbial stimulation driven by the abundance of labile C and nutrients [27,47]. However, they align with Wei et al. [26], who found that higher organic C doses (three times the lower dose) emitted 7% less CO₂-C per unit of litter-C. Similarly, Saviozzi et al. [19] observed lower specific C decomposition of olive mill pomace at higher application doses in an incubation experiment.

N limitation cannot account for the observed reduction in C mineralization, as this decline also affected CC, which showed net N mineralization at both C doses. A more plausible explanation involves mechanisms related to soil microbial activity and C stabilization. Wei et al. [26] suggested that higher doses might cause aggregation of the applied material, preventing microbial access, whereas lower doses facilitate better contact between soil particles and microorganisms, enhancing decomposition. However, the extent to which undecomposable C is truly stabilized remains uncertain. For instance, Wei et al. [26] found that the higher C dose exhibited lower SOC formation efficiency after 80 days of decomposition. The studies of Mendoza et al. [25] and Don et al. [47] provide another explanation, namely a negative priming effect on SOC decomposition after the addition of increasing amendment doses. As we measured CO₂-C emissions, a lower SOC mineralization may have masked similar amendment decomposition across doses.

Our results suggest that applying higher doses of organic C to olive groves, regardless of SOC levels, may enhance short-term SOC accumulation efficiency (i.e., the proportion of amendment-C that remain in the soil in the short term). However, limited microbial transformation at higher doses may reduce the contribution of amendments to long-term SOC stabilization, warranting further investigation. These findings have important implications for managing C inputs in olive groves. For instance, while the cover crop is typically mowed and left on the soil surface or grazed and dispersed randomly, the management pruning residues and olive pomace can be more precisely controlled by farmers. When managing pruning material, it can be strategically collected either in all alleys or in alternate alleys. In the former approach, each alley receives the pruning residues from a single row of trees, while in the latter, half of the alleys receive the pruning material from two rows of trees, leaving the other half uncovered. Similarly, olive pomace is often applied using a spreader trailer or tank truck, allowing for flexible distribution. Such targeted management could optimize C sequestration and potentially improve soil health while reducing costs, although further studies are required to confirm whether this C becomes stabilized over time.

In addition, amendment doses should be carefully considered in relation to N management. We observed that net N immobilization increases proportionally to the amendment dose for PR or OP, despite reduced AOC min₁₃₀. Therefore, the greater potential of N immobilization in concentrated applications should also be considered in field applications. Particularly in SOC-poor soils, the concentrated application of great amounts of pruning residues may cause a localized N deficit that could affect nearby olive trees. In this case, smaller doses are recommended.

4.4. Nitrate Addition Effects on Amendment-C and N Mineralization

Overall, available nitrate promoted the decomposition of labile C while slowing the decomposition of refractory C, as predicted from previous evidence [24]. However, it has no consistent effect on AOC rem₁₃₀. This was expected for CC amendment, as it showed a positive net N mineralization that indicates sufficient nutrient availability, even when external N was not added. However, the lack of significant effects of nitrate availability on the PR-C and OP-C remaining at day 130 was unexpected, as these residues have a high C/N ratio and showed net N immobilization during the 130-day incubation experiment (Figure 3). In fact, Saviozzi et al. [19] found an increase in CO₂ emissions from olive mill pomace when nitrogen was added, at least during the first stage of decomposition. The lack of a consistent effect of nitrate availability in PR and OP-C mineralization could be due to the fact that the immobilization of available NO₃⁻-N by soil microorganisms to fulfill their N requirements under amendments with a high C/N ratio (Figure 3) leads to a higher microbial carbon-use efficiency, i.e., greater C being transformed in microbial biomass rather than respired. This hypothesis is consistent with Parton et al. [48], who showed that when N is added to soils with high C/N ratio organic matter, the added N is often immobilized by microbes rather than contributing to increased rates of C decomposition. This immobilization can lead to a temporary reduction in soil nitrate levels but does not necessarily promote the breakdown of the C-rich amendment.

We also expected that N addition would promote greater N immobilization at the greater application dose of PR and OP, as they exhausted the available soil native N. In fact, during the first 75 days of decomposition, N immobilization by PR and OP was greater with the external N addition, but this difference practically disappeared by the end of the experiment. These results suggest that the added N primarily serves to meet the N requirements of the crop, counteracting N immobilization, rather than promoting the mineralization of amendment-C. Therefore, in N-poor soils where large amounts of PR or OP are applied, fertilization can provide an excess of N available to olive trees during the growing season.

5. Conclusions

Our study provides insights into the dynamics of C and N mineralization of the main organic amendments applied to olive grove soils, highlighting key findings and implications for soil management.

The specific SOC and SON mineralization rates were lower in SOC-rich soils compared to soils with lower SOM content. This reduced mineralization is attributed to the higher level of protected C in the SOC-rich soils.

Differences in C decomposition among amendments with varying C/N ratios and C quality were most pronounced during the initial stages of decomposition. However, as the 130-day study progressed, these differences diminished, with the remaining C content after one year becoming relatively similar across all amendments (between 40 and 49%). These findings underscore the significant potential for short-term soil carbon accumulation following the application of organic amendments produced in olive orchards. Such practices are highly relevant for carbon farming strategies, emphasizing the importance of integrating organic amendments into soil management practices to enhance soil health and contribute to climate mitigation efforts.

The residues from cover crop residue were the only amendment that consistently demonstrated positive N mineralization throughout its decomposition. This highlights its potential capacity to provide N to olive trees during critical growth periods while also contributing to C sequestration.

The addition of external nitrogen had only a minor effect on C and N dynamics of cover crops, tree pruning, and olive mill pomace residues. Therefore, applying available N in conjunction with tree pruning and olive mill pomace residues may help alleviate soil N scarcity without negatively impacting SOC accumulation in the short term.

The quantity of amendment applied significantly reduced the amendment derived-C. By strategically applying organic amendments, farmers can enhance short-term SOC accumulation. However, substantial nitrogen immobilization is anticipated, particularly with tree pruning residues and olive mill pomace amendments.

Overall, our results suggest that the choice and application of organic amendments in olive groves should take into account the specific decomposition dynamics and nutrient interactions to optimize soil health and C sequestration. Future research should focus on calibrating amendment doses to enhance C sequestration and N retention while maintaining crop performance.

Author Contributions

Conceptualization, R.G.-R. and P.D.; methodology, R.G.-R. and P.D.; software, P.D.; validation, E.P.-S. and J.C.; formal analysis, P.D.; investigation, P.D. and R.G.-R.; resources, R.G.-R.; data curation, P.D.; writing—original draft preparation, P.D.; writing—review and editing, R.G.-R., E.P.-S. and J.C.; visualization, P.D.; supervision, R.G.-R.; project administration, R.G.-R.; funding acquisition, R.G.-R. All authors have read and agreed to the published version of the manuscript.

Data Availability Statement

The data are not publicly available because we only want to make them available under request.

Acknowledgments

Authors would like to thank Gustavo Ruiz-Cátedra and Laura P. Martín Coco for their collaboration during data acquisition and experimental measurements.

Conflicts of Interest

The authors declare no conflicts of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

Footnotes

Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Figures, Scheme and Tables

Scheme 1. Schematic representation of treatments.

Figure 1. Cumulative CO2 production (µg CO2−C g−1) in SHC and SLC soils receiving 0 (CON), 2 (C2), or 5 (C5) mg C g−1 of cover crops (CC) or tree pruning (PR) residues or olive mill pomace (OP) and 0 (N0) and 100 (N100) µg NO3−-N g−1. Data are the mean of three replicates and error bars stand for standard deviation.

Figure 2. (a) Proportion of easily decomposable carbon (A coefficient); (b) decomposition coefficient for the easily decomposable carbon (k coefficient); and (c) decomposition coefficient for the refractory carbon (h coefficient). Points represent means and bands stands for the estimated 95% confidence intervals. The red lines are for comparison purposes: non-overlapping lines between different treatments indicate significant differences. (p [less than] 0.05, Tukey test).

Figure 3. Soil mineral N (µg inorganic N g−1) in SHC and SLC soils receiving 0 (CON), 2 (C2), or 5 (C5) mg C g−1 of cover crops (CC) or tree pruning (PR) residues or olive mill pomace (OP) and 0 (N0) or 100 (N100) µg NO3−-N g−1. Data are the mean of three replicates and error bars stand for standard deviation.

Figure 4. Daily net N mineralization (mg N g−1 soil d−1) of amendments in SHC and SLC soils receiving 0 (CON), 2 (C2), or 5 (C5) mg C g−1 of cover crops (CC) or pruning residues (PR) or olive mill pomace (OP) and 0 (N0) or 100 (N100) µg NO3−-N g−1. Points represent means and bands stands for the estimated 95% confidence intervals. Values above the dashed line represent N release and values below represent immobilization. The red lines are for comparison purposes: non-overlapping lines between different treatments indicate significant differences. (p [less than] 0.05, Tukey test).

Table 1

Main properties of the studied soils. Data are the mean of three replicates ± the standard deviation. Different superscript letters indicate differences between soils (p < 0.05, Tukey test).

	High Organic Carbon Soil (SHC)	Low Organic Carbon Soil (SLC)
Coordinates (latitude, longitude)	37.983012, −3.316353	37.983297, −3.315570
Soil type	Vertic regosol	Vertic regosol
pH	7.50 ± 0.09 a	7.48 ± 0.01 a
Sand (%)	27.3 ± 9.3 a	19.3 ± 1.2 a
Silt (%)	25.0 ± 4.6 a	23.7 ± 1.2 a
Clay (%)	47.7 ± 13.6 a	57.0 ± 0.0 a
Gravel content (%)	15.7 ± 10.3 a	13.2 ± 9.5 a
Top 30 cm bulk density (g cm−3)	1.36 ± 0.04 a	1.44 ± 0.07 a
Water field capacity (% volume)	40.5 ± 6.7 a	45.4 ± 0.2 a
SOM (%)	2.76 ± 0.14 a	1.42 ± 0.16 b
SOC (%)	1.61 ± 0.08 a	0.82 ± 0.09 b
TN (%)	0.20 ± 0.02 a	0.12 ± 0.02 b
CEC (cmol (+) kg−1)	29.7 ± 7.5 a	32.0 ± 1.0 a
Humic and fulvic acids (mg g−1)	6.54 ± 2.16 a	3.75 ± 0.08 a
Potential nitrification (µg N g−1 5 h−1)	10.4 ± 9.0 a	14.1 ± 1.3 a
Arisulfatase activity (µg p-NP g−1 h−1)	39.9 ± 5.2 a	16.0 ± 0.5 b
β-glucosidase activity (µg p-NP g−1 h−1)	262.9 ± 83.7 a	146.8 ± 28.1 a
Dehydrogenase activity (µg TPF g−1 h−1)	46.3 ± 41.5 a	14.8 ± 22.8 a
Phosphomonoesterase (µg p-NP g−1 h−1)	103.92 ± 39.77 a	83.40 ± 8.86 a

Table 2

Total carbon (TC), total nitrogen (TN), and C/N ratio on dry matter basis of spontaneous cover crop, pruning residues and raw olive mill pomace. Different superscript letters indicate differences (p < 0.05) between amendments.

	Spontaneous Cover Crop (CC)	Light Pruning Residues (PR)	Raw Olive Mill Pomace (OP)
TC (%)	40.5 ± 0.1 b	47.7 ± 0.2 a	47.6 ± 1.0 a
TN (%)	2.41 ± 0.07 a	0.90 ± 0.03 c	1.24 ± 0.17 b
C/N ratio	16.8 ± 0.5 c	52.7 ± 2.0 a	38.8 ± 4.9 b

Table 3

Total biomass, carbon, and nitrogen (on dry matter basis) added in the jars through amendments at 2 (C2) or 5 mg C g⁻¹ (C5) application rate for the residues of cover crop (CC), tree pruning (PR), and olive pomace (OP). The equivalent biomass applied per hectare considering 0.08 m depth in a hypothetical field application is also shown.

	Spontaneous Cover Crop (CC)		Light Pruning Residues (PR)		Raw Olive Mill Pomace (OP)
	C2	C5	C2	C5	C2	C5
Lab conditions
Biomass (mg jar−1)	493.8	1234.6	419.3	1048.2	420.2	1050.4
Carbon (mg jar−1)	200.0	500.0	200.0	500.0	200.0	500.0
Nitrogen (mg jar−1)	11.9	29.7	3.8	9.4	5.2	13.0
Field conditions
Biomass (kg ha−1)	4748.6	11,885.9	4035.8	10,104.0	4045.4	10,113.6
Carbon (kg ha−1)	1926.4	4816.0	1926.4	4816.0	1926.4	4816.0
Nitrogen (kg ha−1)	114.4	286.5	36.3	90.9	50.2	125.4

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/agriculture14111923/s1, Figure S1: Four parameter double exponential decay model for SOC remaining of SHC and SLC soils receiving 0 and 100 µg N g⁻¹; Figure S2: Four parametric double exponential decay model for AOC remaining for all the combinations of independent variables; Table S1: Parameters of the double exponential decay model for SOC remaining and observed SOC remaining at day 130 in the two types of soils; Table S2: ANOVA p-value results of the effect of independent variables on the parameters of the double exponential decay model for SOC decay and on the observed SOC remaining at day 130; Table S3: Coefficients of the double exponential decay model for AOC remaining, and observed AOC remaining at 130; Table S4: ANOVA p-value results of the effect of independent variables on the parameters of the double exponential decay model for AOC decay and on the AOC remaining at day 130; Table S5: Modeled values for AOC remaining at days 130 and 365 for all the combinations of independent variables; Table S6: Daily soil net N mineralization, absolute soil net N mineralization at day 130 and specific soil net N mineralization; Table S7: ANOVA p-value results of the effect of independent variables on daily soil net N mineralization, absolute soil net N mineralization at day 130 and specific soil net N mineralization; Table S8: Daily amendments net N mineralization, absolute amendment net N mineralization at day 130 and specific amendment net N mineralization; Table S9: ANOVA p-value results of the effect of independent variables on the net N mineralization of the amendments.

References

1. Infante-Amate, J.; Villa, I.; Aguilera, E.; Torremocha, E.; Guzmán, G.; Cid, A.; González de Molina, M. The Making of Olive Landscapes in the South of Spain. A History of Continuous Expansion and Intensification. Biocultural Diversity in Europe; Agnoletti, M.; Emanueli, F. Environmental History Springer International Publishing: Cham, Switzerland, 2016; Volume 5, pp. 157-179. ISBN 978-3-319-26313-7

2. Morugán-Coronado, A.; Linares, C.; Gómez-López, M.D.; Faz, Á.; Zornoza, R. The Impact of Intercropping, Tillage and Fertilizer Type on Soil and Crop Yield in Fruit Orchards under Mediterranean Conditions: A Meta-Analysis of Field Studies. Agric. Syst.; 2020; 178, 102736. [DOI: https://dx.doi.org/10.1016/j.agsy.2019.102736]

3. Rodríguez-Entrena, M.; Arriaza, M. Adoption of Conservation Agriculture in Olive Groves: Evidences from Southern Spain. Land Use Policy; 2013; 34, pp. 294-300. [DOI: https://dx.doi.org/10.1016/j.landusepol.2013.04.002]

4. Repullo, M.A.; Carbonell, R.; Hidalgo, J.; Rodríguez-Lizana, A.; Ordóñez, R. Using Olive Pruning Residues to Cover Soil and Improve Fertility. Soil Tillage Res.; 2012; 124, pp. 36-46. [DOI: https://dx.doi.org/10.1016/j.still.2012.04.003]

5. Repullo-Ruibérriz de Torres, M.A.; Carbonell-Bojollo, R.M.; Moreno-García, M.; Ordóñez-Fernández, R.; Rodríguez-Lizana, A. Soil Organic Matter and Nutrient Improvement through Cover Crops in a Mediterranean Olive Orchard. Soil Tillage Res.; 2021; 210, 104977. [DOI: https://dx.doi.org/10.1016/j.still.2021.104977]

6. Torrús-Castillo, M.; Calero, J.; García-Ruiz, R. Does Olive Cultivation Sequester Carbon?: Carbon Balance along a C Input Gradient. Agric. Ecosyst. Environ.; 2023; 358, 108707. [DOI: https://dx.doi.org/10.1016/j.agee.2023.108707]

7. Zipori, I.; Erel, R.; Yermiyahu, U.; Ben-Gal, A.; Dag, A. Sustainable Management of Olive Orchard Nutrition: A Review. Agriculture; 2020; 10, 11. [DOI: https://dx.doi.org/10.3390/agriculture10010011]

8. Arias-Giraldo, L.F.; Guzmán, G.; Montes-Borrego, M.; Gramaje, D.; Gómez, J.A.; Landa, B.B. Going Beyond Soil Conservation with the Use of Cover Crops in Mediterranean Sloping Olive Orchards. Agronomy; 2021; 11, 1387. [DOI: https://dx.doi.org/10.3390/agronomy11071387]

9. Rodríguez-Lizana, A.; Espejo-Pérez, A.J.; González-Fernández, P.; Ordóñez-Fernández, R. Pruning Residues as an Alternative to Traditional Tillage to Reduce Erosion and Pollutant Dispersion in Olive Groves. Water Air Soil Pollut.; 2008; 193, pp. 165-173. [DOI: https://dx.doi.org/10.1007/s11270-008-9680-5]

10. Ordóñez-Fernández, R.; Repullo-Ruibérriz De Torres, M.A.; Román-Vázquez, J.; González-Fernández, P.; Carbonell-Bojollo, R. Macronutrients Released during the Decomposition of Pruning Residues Used as Plant Cover and Their Effect on Soil Fertility. J. Agric. Sci.; 2015; 153, pp. 615-630. [DOI: https://dx.doi.org/10.1017/S0021859614000458]

11. Marañón-Jiménez, S.; Serrano-Ortíz, P.; Peñuelas, J.; Meijide, A.; Chamizo, S.; López-Ballesteros, A.; Vicente-Vicente, J.L.; Fernández-Ondoño, E. Effects of Herbaceous Covers and Mineral Fertilizers on the Nutrient Stocks and Fluxes in a Mediterranean Olive Grove. Eur. J. Agron.; 2022; 140, 126597. [DOI: https://dx.doi.org/10.1016/j.eja.2022.126597]

12. Torrús-Castillo, M.; Domouso, P.; Herrera-Rodríguez, J.M.; Calero, J.; García-Ruiz, R. Aboveground Carbon Fixation and Nutrient Retention in Temporary Spontaneous Cover Crops in Olive Groves of Andalusia. Front. Environ. Sci.; 2022; 10, 868410. [DOI: https://dx.doi.org/10.3389/fenvs.2022.868410]

13. Domouso, P.; Calero, J.; Ruiz-Cátedra, G.; García-Ruiz, R. Nitrogen Recycling across a Spectrum of Fertilization Strategies: An Assessment in Olive Groves. Agric. Ecosyst. Environ.; 2024; 372, 109096. [DOI: https://dx.doi.org/10.1016/j.agee.2024.109096]

14. Gómez-Muñoz, B.; Hatch, D.J.; Bol, R.; Garca-Ruiz, R. The Compost of Olive Mill Pomace: From a Waste to a Resource—Environmental Benefits of Its Application in Olive Oil Groves. Sustainable Development—Authoritative and Leading Edge Content for Environmental Management; Curkovic, S. InTech: Rijeka, Croatia, 2012; ISBN 978-953-51-0682-1

15. Ntougias, S.; Bourtzis, K.; Tsiamis, G. The Microbiology of Olive Mill Wastes. BioMed Res. Int.; 2013; 2013, 784591. [DOI: https://dx.doi.org/10.1155/2013/784591] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/24199199]

16. López-Piñeiro, A.; Albarrán, A.; Nunes, J.M.R.; Barreto, C. Short and Medium-Term Effects of Two-Phase Olive Mill Waste Application on Olive Grove Production and Soil Properties under Semiarid Mediterranean Conditions. Bioresour. Technol.; 2008; 99, pp. 7982-7987. [DOI: https://dx.doi.org/10.1016/j.biortech.2008.03.051] [PubMed: https://www.ncbi.nlm.nih.gov/pubmed/18462936]

17. Ministerio de Agricultura, Pesca y Alimentación (MAPA) 2023-2027 CAP Strategic Plan of Spain Version 3.3. 2022; Available online: https://www.mapa.gob.es/es/pac/pac-2023-2027/pepac-version-33_tcm30-692566.pdf (accessed on 22 October 2024).

18. Gómez-Muñoz, B.; Valero-Valenzuela, J.D.; Hinojosa, M.B.; García-Ruiz, R. Management of Tree Pruning Residues to Improve Soil Organic Carbon in Olive Groves. Eur. J. Soil Biol.; 2016; 74, pp. 104-113. [DOI: https://dx.doi.org/10.1016/j.ejsobi.2016.03.010]

19. Saviozzi, A.; Levi-Minzi, R.; Cardelli, R.; Biasci, A.; Riffaldi, R. Suitability of Moist Olive Pomace as Soil Amendment. Water Air Soil Pollut.; 2001; 128, pp. 13-22. [DOI: https://dx.doi.org/10.1023/A:1010361807181]

20. Rodríguez-Lizana, A.; Repullo-Ruibérriz de Torres, M.Á.; Carbonell-Bojollo, R.; Moreno-García, M.; Ordóñez-Fernández, R. Study of C, N, P and K Release from Residues of Newly Proposed Cover Crops in a Spanish Olive Grove. Agronomy; 2020; 10, 1041. [DOI: https://dx.doi.org/10.3390/agronomy10071041]

21. Bonanomi, G.; Sarker, T.C.; Zotti, M.; Cesarano, G.; Allevato, E.; Mazzoleni, S. Predicting Nitrogen Mineralization from Organic Amendments: Beyond C/N Ratio by 13C-CPMAS NMR Approach. Plant Soil; 2019; 441, pp. 129-146. [DOI: https://dx.doi.org/10.1007/s11104-019-04099-6]

22. Berg, B.; McClaugherty, C. Plant Litter: Decomposition, Humus Formation, Carbon Sequestration; Springer International Publishing: Cham, Switzerland, 2020; ISBN 978-3-030-59630-9

23. Cotrufo, M.F.; Wallenstein, M.D.; Boot, C.M.; Denef, K.; Paul, E. The Microbial Efficiency-Matrix Stabilization (MEMS) Framework Integrates Plant Litter Decomposition with Soil Organic Matter Stabilization: Do Labile Plant Inputs Form Stable Soil Organic Matter?. Glob. Change Biol.; 2013; 19, pp. 988-995. [DOI: https://dx.doi.org/10.1111/gcb.12113]

24. Gill, A.L.; Schilling, J.; Hobbie, S.E. Experimental Nitrogen Fertilisation Globally Accelerates, Then Slows Decomposition of Leaf Litter. Ecol. Lett.; 2021; 24, pp. 802-811. [DOI: https://dx.doi.org/10.1111/ele.13700]

25. Mendoza, O.; De Neve, S.; Deroo, H.; Li, H.; Sleutel, S. Do Interactions between Application Rate and Native Soil Organic Matter Content Determine the Degradation of Exogenous Organic Carbon?. Soil Biol. Biochem.; 2022; 164, 108473. [DOI: https://dx.doi.org/10.1016/j.soilbio.2021.108473]

26. Wei, Y.; Xiong, X.; Ryo, M.; Badgery, W.B.; Bi, Y.; Yang, G.; Zhang, Y.; Liu, N. Repeated Litter Inputs Promoted Stable Soil Organic Carbon Formation by Increasing Fungal Dominance and Carbon Use Efficiency. Biol. Fertil. Soils; 2022; 58, pp. 619-631. [DOI: https://dx.doi.org/10.1007/s00374-022-01647-8]

27. Fang, X.; Zhao, L.; Zhou, G.; Huang, W.; Liu, J. Increased Litter Input Increases Litter Decomposition and Soil Respiration but Has Minor Effects on Soil Organic Carbon in Subtropical Forests. Plant Soil; 2015; 392, pp. 139-153. [DOI: https://dx.doi.org/10.1007/s11104-015-2450-4]

28. Pansu, M.; Gautheyrou, J. Handbook of Soil Analysis: Mineralogical, Organic and Inorganic Methods; Springer: Berlin/Heidelberg, Germany, 2006; ISBN 978-3-540-31210-9

29. Methods of Soil Analysis. Part 2. Chemical and Microbiological Properties; 2nd ed. Page, A.L. Agronomy American Society of Agronomy: Madison, WI, USA, 1982; Volume 2.

30. Schinner, F.; Öhlinger, R.; Kandeler, E.; Margesin, R. Methods in Soil Biology; Springer: Berlin/Heidelberg, Germany, 1996; ISBN 978-3-642-64633-1

31. IUSS Working Group WRB World Reference Base for Soil Resources. International Soil Classification System for Naming Soils and Creating Legends for Soil Maps; 4th ed. International Union of Soil Sciences: Vienna, Austria, 2022; ISBN 979-8-9862451-1-9

32. Methods of Soil Analysis. Part 2: Microbiological and Biochemical Properties; Weaver, R.W.; Angle, S.; Bottomley, P.; Bezdicek, D.; Smith, S.; Tabatabai, A.; Wollum, A. Soil Science Society of America book series; Soil Science Society of America: Madison, WI, USA, 1994; ISBN 978-0-89118-810-0

33. Koroleff, F. Direct Spectrophotometric Determination of Ammonia in Precipitation. Tellus; 1966; 18, pp. 562-565. [DOI: https://dx.doi.org/10.3402/tellusa.v18i2-3.9343]

34. Shinn, M.B. Colorimetric Method for Determination of Nitrite. Ind. Eng. Chem. Anal. Ed.; 1941; 13, pp. 33-35. [DOI: https://dx.doi.org/10.1021/i560089a010]

35. Marzi, M.; Shahbazi, K.; Kharazi, N.; Rezaei, M. The Influence of Organic Amendment Source on Carbon and Nitrogen Mineralization in Different Soils. J. Soil Sci. Plant Nutr.; 2020; 20, pp. 177-191. [DOI: https://dx.doi.org/10.1007/s42729-019-00116-w]

36. García-Ruiz, R.; Carranza-Gallego, G.; Aguilera, E.; González De Molina, M.; Guzmán, G.I. C and N Mineralisation of Straw of Traditional and Modern Wheat Varieties in Soils of Contrasting Fertility. Nutr. Cycl. Agroecosystems; 2019; 113, pp. 167-179. [DOI: https://dx.doi.org/10.1007/s10705-019-09973-4]

37. Anand, R.; Germon, J.-C.; Groffman, P.M.; Norton, J.M.; Philippot, L.; Prosser, J.I.; Schimel, J.P. Nitrogen Transformations. Handbook of Soil Science: Properties and Processes; Huang, P.M.; Li, Y.; Sumner, M.E. Taylor and Francis Group: Boca Raton, FL, USA, 2012.

38. Huang, X.; Jia, Z.; Guo, J.; Li, T.; Sun, D.; Meng, H.; Yu, G.; He, X.; Ran, W.; Zhang, S. et al. Ten-Year Long-Term Organic Fertilization Enhances Carbon Sequestration and Calcium-Mediated Stabilization of Aggregate-Associated Organic Carbon in a Reclaimed Cambisol. Geoderma; 2019; 355, 113880. [DOI: https://dx.doi.org/10.1016/j.geoderma.2019.113880]

39. Yang, S.; Jansen, B.; Absalah, S.; Kalbitz, K.; Chunga Castro, F.O.; Cammeraat, E.L.H. Soil Organic Carbon Content and Mineralization Controlled by the Composition, Origin and Molecular Diversity of Organic Matter: A Study in Tropical Alpine Grasslands. Soil Tillage Res.; 2022; 215, 105203. [DOI: https://dx.doi.org/10.1016/j.still.2021.105203]

40. Six, J.; Conant, R.T.; Paul, E.A.; Paustian, K. Stabilization Mechanisms of Soil Organic Matter: Implications for C-Saturation of Soils. Plant Soil; 2002; 241, pp. 155-176. [DOI: https://dx.doi.org/10.1023/A:1016125726789]

41. Pareja-Sánchez, E.; Domouso, P.; Gómez-Muñoz, B.; Heras-Linares, M.T.; García-Ruíz, R. Conservation Practices Boost Soil-Protected Organic Carbon Stocks in Olive Orchards. Agriculture; 2024; 14, 1354. [DOI: https://dx.doi.org/10.3390/agriculture14081354]

42. Nett, L.; Ruppel, S.; Ruehlmann, J.; George, E.; Fink, M. Influence of Soil Amendment History on Decomposition of Recently Applied Organic Amendments. Soil Sci. Soc. Am. J.; 2012; 76, pp. 1290-1300. [DOI: https://dx.doi.org/10.2136/sssaj2011.0190]

43. Murúa, J.M.; Gaxiola, A. Variability in Terrestrial Litter Decomposition Can Be Explained by Nutrient Allocation Strategies among Soil Decomposer Communities. Funct. Ecol.; 2023; 37, pp. 1642-1652. [DOI: https://dx.doi.org/10.1111/1365-2435.14321]

44. Stark, C.H.; Condron, L.M.; O’Callaghan, M.; Stewart, A.; Di, H.J. Differences in Soil Enzyme Activities, Microbial Community Structure and Short-Term Nitrogen Mineralisation Resulting from Farm Management History and Organic Matter Amendments. Soil Biol. Biochem.; 2008; 40, pp. 1352-1363. [DOI: https://dx.doi.org/10.1016/j.soilbio.2007.09.025]

45. Mooshammer, M.; Wanek, W.; Hämmerle, I.; Fuchslueger, L.; Hofhansl, F.; Knoltsch, A.; Schnecker, J.; Takriti, M.; Watzka, M.; Wild, B. et al. Adjustment of Microbial Nitrogen Use Efficiency to Carbon: Nitrogen Imbalances Regulates Soil Nitrogen Cycling. Nat. Commun.; 2014; 5, 3694. [DOI: https://dx.doi.org/10.1038/ncomms4694]

46. Mitchell, E.; Scheer, C.; Rowlings, D.; Conant, R.T.; Cotrufo, M.F.; Grace, P. Amount and Incorporation of Plant Residue Inputs Modify Residue Stabilisation Dynamics in Soil Organic Matter Fractions. Agric. Ecosyst. Environ.; 2018; 256, pp. 82-91. [DOI: https://dx.doi.org/10.1016/j.agee.2017.12.006]

47. Don, A.; Rödenbeck, C.; Gleixner, G. Unexpected Control of Soil Carbon Turnover by Soil Carbon Concentration. Environ. Chem. Lett.; 2013; 11, pp. 407-413. [DOI: https://dx.doi.org/10.1007/s10311-013-0433-3]

48. Parton, W.; Silver, W.L.; Burke, I.C.; Grassens, L.; Harmon, M.E.; Currie, W.S.; King, J.Y.; Adair, E.C.; Brandt, L.A.; Hart, S.C. et al. Global-Scale Similarities in Nitrogen Release Patterns During Long-Term Decomposition. Science; 2007; 315, pp. 361-364. [DOI: https://dx.doi.org/10.1126/science.1134853]