Increasing Topsoil Depth Improves Yield and Nitrogen Fertilizer Use Efficiency in Maize

Abstract

Topsoil degradation poses a significant threat to agricultural production worldwide. However, whether degraded topsoil is a net nitrogen source or sink depends on crop uptake and nutrient loss, and how it affects the sustainability of agricultural production remains unclear. To fill this gap in understanding, we conducted a three-year experiment with five topsoil depth treatments: 10 cm (D10), 20 cm (D20), 30 cm (D30), 40 cm (D40), and 50 cm (D50). Increasing topsoil depth significantly increased grain yield by a maximum of 49.4% (between D10 and D50). With increasing topsoil depth, the NFUE rises from 14.2% to 64.9% (between D10 and D50 treatments), while the G-NFUE climbs from 9.0% to 36.2% (between D10 and D40 treatments). Increasing topsoil depth reduced topsoil N depletion and the percentage of change in soil N stocks. In addition, N fertilizers applied during the season were generally enriched in soil at a depth of 30–40 cm. Therefore, increasing the depth of topsoil can effectively increase the source of nutrients absorbed by a crop by increasing access to additional resources stored in deeper soils, which ultimately increases maize grain yield and N fertilizer use efficiency. In this study, the threshold for maize to achieve high yields and efficiency was a topsoil depth of 30 cm. This study elucidated the differences in maize grain yield and resource utilization at different topsoil depths and established a link with soil N characteristics, and thus, it will provide a theoretical basis for the sustainable management of topsoil.

1. Introduction

Sustainability of agriculture is facing serious challenges due to climate change [1] and looming nutrient supply shortages [2]. In many agricultural regions, wind, water, and tillage erosion have accelerated the loss of fertile topsoil to levels exceeding those of soil formation [3]. In general, rates of soil loss on conventionally tilled farms far outpace natural rates of erosion [4,5]. Soils and agriculture are key factors for sustainable land management under future environmental change and require a multidisciplinary approach to address the challenges stemming from multiple socioeconomic and physical pressures [6]. When soil management is unsustainable, the goal of a food-secure future is severely threatened because of deteriorating soil health and accelerating topsoil loss [7].

Because soil nutrients are mainly concentrated in the topsoil [8,9], topsoil loss reduces nutrient availability to plants. In previous studies, the negative effects of topsoil loss on crop yields are attributed mainly to losses in soil organic carbon (C) and mineral nutrients, especially nitrogen (N), phosphorus (P), and potassium (K) [10,11]. Zöbisch et al. [12] showed that topsoil loss of 3–6 t ha⁻¹ resulted in reductions in P and K uptake by 24% and 28%, respectively, for maize and by 15% and 16%, respectively, for legumes. In China, soil erosion reduces soil ammonium concentration by 66% on the Loess Plateau [13] and soil organic C, total N, and total P by 33%, 28%, and 27%, respectively, in alpine grasslands [14]. In addition, reduced topsoil depth inhibits crop root proliferation and thus reduces root nutrient acquisition [15]. Soil erosion is leading to shallow soils across much of the agricultural Midwest in the United States [16]. Shallow soils may pose additional challenges to agricultural production as the climate changes. For example, water-holding capacity is reduced in shallow soils, and therefore, they cannot absorb rainfall from large precipitation events, leading to increased runoff and insufficient soil water available for plants during dry periods [17].

Farmland subsoil (i.e., the soil layer below the topsoil that is regularly tilled) may store nearly 50% of the total N reserves [18] and 25% to 70% of the total P reserves [19], which are difficult for crops to utilize. The organic C (OC) stored in global topsoil (0–30 cm) will be the most active participant in the C cycle under future climate change [20]. Because reductions in topsoil lead to reductions in soil productivity and function, overall soil health and the ability to support multifaceted soil and plant ecosystem services are jeopardized [21,22]. Depending on the amount and severity of topsoil loss, the key soil physical, chemical, including soil C, N, and other fertility pools, and biological properties are negatively altered [23,24]. As a result, soil conditions are unfavorable for proper crop growth throughout the growth stage, which limits crop development and ultimately affects crop yields [22]. Therefore, understanding how depth of farmland topsoil affects soil nutrient availability and plant utilization is important in assessing the vulnerability of agriculture and preventing soil erosion.

Long-term research demonstrates that implementation of measures such as reduced tillage, no-tillage, crop straw return, organic amendment inputs, cover crops, and crop rotations can improve degraded soil properties and thus improve functioning related to cropping system objectives [24,25,26]. However, there are few short-term studies on the relevant characteristics of degraded soils, and especially, the characteristics of soil fertilizer nutrient supply and plant utilization within a growing season are rarely reported. Therefore, to investigate how topsoil degradation affects the mechanisms related to high yields and efficient resource utilization of maize, we used ¹⁵N stable isotope labeling to analyze the differences in N fertilizer nutrient migration, enrichment, and utilization by maize plants within a growing season under different topsoil depths. The objective of this study was to elucidate the mechanisms determining differences in maize grain yield under different topsoil depths. We expect the results will provide the theoretical basis and technical guidance for using appropriate tillage practices to reduce soil degradation and improve sustainability of highly efficient, high-yielding maize crops.

2. Materials and Methods

2.1. Site Description

The study was conducted at the Gongzhuling Experimental Station of the Jilin Academy of Agricultural Sciences, Jilin Province, China (43°52′ N, 124°81′ E; 206 m a.s.l.) during 2021–2023. The region is in temperate and cold temperate zones with humid and semihumid climates. Winters are cold and dry, and summers are warm and short. Average annual duration of sunlight in the experimental region was 2624 h; total solar radiation was 5551 MJ m⁻²; the frost-free stage was 144 d; average annual temperature was 6.7 °C, and average annual precipitation was 572.7 mm during the past 15 years. The climate, combined with good soil quality, explains why the northern spring maize region is the main maize-producing area in China. Meteorological data of the experimental site during the study period growing seasons are represented in Figure S1. The soil is a typical Mollisols [27], which is known locally as “black soil”. Soil chemical properties in the 0–20-cm tillage layer were as follows: organic matter, 15.1 ± 1.0 g kg⁻¹; alkaline N, 192.9 ± 13.7 mg kg⁻¹; fast-acting P, 10.1 ± 0.8 mg kg⁻¹; fast-acting K, 292.3 ± 19.8 mg kg⁻¹; and pH, 7.9 ± 0.3. The main crop grown before initiation of the experiment was maize (Zea mays L.), with continuous maize also a dominant practice in the region.

To establish an independent system of experimental plots (each measuring 1.8 m × 4.0 m, with a total area of 7.2 m²), the plots were constructed using iron frames (50 cm in height) and plastic sheets (Figure S2). A 200-mesh (0.074 mm pore size) nylon net was placed at the bottom of each plot to simulate the restrictive effect of the soil plow pan on maize root growth (Figure S2B). Before beginning the experiment, the cultivated soil was excavated to five different depths: 10 cm, 20 cm, 30 cm, 40 cm, and 50 cm. Subsequently, the iron frames were inserted vertically into the excavated areas, and the root-limiting nylon nets were laid horizontally at the corresponding target soil depths. The soil layer beneath each nylon net interface was compacted to mimic the natural density of the plow pan. Following compaction, the excavated soil was backfilled into the regions above the nylon nets. The backfilled soil was then watered and left to settle naturally to ensure soil structure stability (Figure S2C).

2.2. Experimental Design and Agronomic Management

The experiment was set up with five topsoil depth treatments: 10 cm (D10), 20 cm (D20), 30 cm (D30), 40 cm (D40), and 50 cm (D50). Each treatment was replicated in three plots in a completely randomized block design. The maize cultivar was Fumin985, which is one of the main cultivars grown in the region. Plant density was 6 plants m⁻², and, in each plot, there were three planted rows within the width of 1.8 m with a row spacing of 60 cm. In addition, to minimize plot margin effects on maize growth and development, there were two extra rows of maize on either side of each plot, and the ends of the planted rows were extended. The distance between all neighboring plots was sufficient to ensure the independence of each plot.

In 2021 and 2022, all treatments received the same levels of fertilizers (N, 200 kg ha⁻¹; P₂O₅, 90 kg ha⁻¹; K₂O, 100 kg ha⁻¹), with N, P, and K applied once as a base fertilizer before sowing. The N fertilizer (resin-coated urea, 46% N) (Jilin Difu Fertilizer Technology Co., Changchun, China) was slow-release to supply N throughout the maize reproductive period.

In 2023, maize was fertilized at the 3-leaf stage. Five representative maize plants per plot were selected for isotope labeling before fertilization. Plants were fertilized at a 1:9 ratio of ¹⁵N stable isotope-labeled urea (10.11% abundance, 46% N; Shanghai Research Institute of Chemical Industry Co., Shanghai, China) to resin-coated urea. The actual abundance of the ¹⁵N stable isotope in the applied N fertilizer was therefore 1.011%. The only type of N fertilizer applied to the unlabeled plants was resin-coated urea. All plants were fertilized with the same level of N (N, 200 kg ha⁻¹), P (P₂O₅, 90 kg ha⁻¹), and potash (K₂O, 100 kg ha⁻¹).

2.3. Sampling and Measurements

2.3.1. Grain Yield and Its Components

At the maturity stage, we chose 10 standard ears per sampling point for morphological analysis, recording the average number of rows per ear (rows) and kernels per row (kernels). Ten maize ears were randomly collected from the central three rows of each plot and only from the interior of a row (away from the ends of the plot). Kernel number per ear (θ) and 1000-kernel weight (W) were determined from the harvested ears. Maize grain yield was calculated using Equation (1), with grain water content adjusted to 14% [28,29].

Y = θ × W × 10⁻³/(1 − 0.14),

(1)

where Y is the grain yield (g plant⁻¹), θ is the number of kernels per ear, and W is the 1000-kernel weight (g). The minimum standard of moisture content suggested in the literature for maize in storage is 14%.

The coefficient of variation (CV) is a statistical measure used to represent the stability of maize yield over different years, with a lower CV indicating higher yield stability. The sustainable yield index (SYI) reflects the sustainable production capacity of an ecosystem, and the higher the value, the better the soil capacity to sustain crop yields over the long term [30].

SYI = (Y − σ)/Y_max,

(2)

where σ is the standard deviation, Y is the mean grain yield (2021–2023), and Y_max is the maximum grain yield in the experiment over the years (2021–2023) for each treatment.

2.3.2. Nitrogen Fertilizer Use Efficiency in Maize

Three ¹⁵N stable isotope-labeled maize plants were collected from each plot at maize physiological maturity. The labeled maize plants were divided into stems, leaves, spike, bracts, cob, and kernels, which were oven-dried at 80 °C to constant weight to determine dry weights. Plant tissues were crushed and ground to pass through a 0.35-mm sieve. Total N was determined using an elemental analyzer (Elementar Various MICRO cube, Elementar Analysensysteme Ltd., Wittenberg, Germany), and ¹⁵N abundance was determined using an isotope mass spectrometer (Loprime100, Elementar Analysensysteme Ltd., Wittenberg, Germany). The proportion of maize plant N accumulation from in-season N fertilization (P-N_season) was calculated according to the following equation [31]:

P-N_season = (δ¹⁵N_p − δ¹⁵N₀)/(δ¹⁵N_f − δ¹⁵N₀) × 100,

(3)

where P-N_season (%) represents the percentage of N accumulation in maize plants from seasonal N fertilizer; δ¹⁵N_p (%) represents the abundance of ¹⁵N stable isotopes in maize plants; δ¹⁵N_f (%) represents the abundance of ¹⁵N stable isotopes in applied N fertilizer; and δ¹⁵N₀ (%) represents the environmental ¹⁵N stable isotope natural abundance, which was 0.3663% in this study. Nitrogen fertilizer use efficiency was estimated from P-N_season, as follows [29,31]:

$$\mathrm{NFUE}=\left({\sum }_{\mathrm{i}}^{\mathrm{n}}=\mathrm{DM}\times \mathrm{TN}\times {P-N}_{\mathrm{season}}\right)/\mathrm{TNF},$$

(4)

G-NFUE = (GDM × GTN × P-N_season)/TNF,

(5)

where NFUE represents the N fertilizer use efficiency of maize plants (%); n represents the number of organs in a maize plant; DM represents the dry matter accumulation of each maize organ (kg ha⁻¹); TN represents the N content of each maize organ (%); and TNF represents the N application in a season, which was 200 kg ha⁻¹ in this study. In Equation (4), G-NFUE represents the N fertilizer use efficiency of maize grain (%); GDM represents the grain matter accumulation of maize (kg ha⁻¹); and GTN represents the N content of maize grain (%).

2.3.3. Soil Nitrogen Characteristics

Soil samples were collected between maize rows using a soil corer before planting (BS) and at anthesis (VT) and physiological maturity (R6) stages. Soil was sampled only above the nylon mesh interface and was stratified in 10-cm increments. Thus, soil samples were collected in one to five layers for treatments D10 to D50, respectively. Soil was collected at three points from each plot and then mixed. Soils were dried naturally in the shade for determination of soil total N and ¹⁵N stable isotope contents. Soil samples were ground and pulverized and passed through a 0.25 mm sieve; total N was determined using an elemental analyzer (Elementar vario MICRO cube), and ¹⁵N abundance was determined using an isotope mass spectrometer (Loprime100).

Soil samples collected during the VT stage of the 2021 to 2023 growing seasons were used to assess the vertical distribution of soil N content at that critical stage during maize growth. Soil samples collected at the BS and R6 stages of the 2021 and 2022 growing seasons were used to assess the stock of soil N during maize growth. In addition, soil bulk density (g cm⁻³) was measured at this stage according to a cutting-ring method [32]. Soil bulk density was calculated using the following equation:

Soil bulk density (g cm⁻³) = soil dry weight (g)/cutting-ring volume (cm³).

(6)

Soil N content and soil bulk density were used to assess the stock of N, calculated with the following equations [33]:

$$\mathrm{S}={\sum }_{\mathrm{i}}^{\mathrm{k}}=1{\mathrm{C}}_{\mathrm{i}}\times {\mathrm{B}}_{\mathrm{i}}\times {\mathrm{H}}_{\mathrm{i}}\times {10}^3,$$

(7)

ΔS = S_BS − S_R6,

(8)

δS = ΔS/S_BS,

(9)

where S is the stock of N (kg ha⁻¹); C is the concentration of N (g kg⁻¹); B is the soil bulk density (g cm⁻³); and H is the soil sampling depth (10 cm), with k representing the number of layers in the soil profile, which in this study, was 1 to 5 for treatments D10 to D50, respectively. ΔS is the change in soil N stock between the R6 stage and BS (kg ha⁻¹), with S_BS the stock of N at BS and S_R6 the stock of N at R6. δS is the percentage of the change in soil N stock (%).

The soil samples collected at the VT stage of the 2023 growing season were also used to assess the fate of N from in-season applications of N fertilizer at that critical stage during maize growth, and the proportion of soil N derived from in-season N fertilizer (S-N_season) was calculated according to the following equation [31]:

S-N_season = (δ¹⁵N_s − δ¹⁵N₀)/(δ¹⁵N_f − δ¹⁵N₀) × 100,

(10)

where S-N_season (%) represents the percentage of seasonal N fertilizer in soil N; δ¹⁵N_s (%) represents the abundance of ¹⁵N stable isotopes in the soil; δ¹⁵N_f (%) represents the abundance of ¹⁵N stable isotopes in applied N fertilizer; and δ¹⁵N₀ (%) represents the environmental ¹⁵N stable isotope natural abundance, which was 0.3663% in this study.

2.3.4. Activities of Key Soil Nitrogen Metabolism Enzymes

The soil samples collected at the VT of the 2023 growing season were also used to assess the activities of soil nitrate reductase, soil nitrite reductase, and soil hydroxylamine reductase at that critical stage. Activities of all enzymes were measured using commercial kits (Suzhou Michy Biomedical Technology Co., Ltd., Suzhou, China) and an enzyme labeler (Spectramax i3x, Molecular Devices, LLC, Salzburg, Austria). Kit instructions were downloaded from the company’s website (www.michybio.com), which briefly describe assay principles and define units of activity.

Soil nitrate reductase (S-NR): S-NR catalyzes the reduction of nitrate to nitrite, while simultaneously inhibiting the degradation of the produced nitrite by nitrite reductase. In the assay, the nitrite reacts with the corresponding colorant to produce a (pink) red azo compound, and soil nitrate reductase activity is determined by its maximum absorption peak at 540 nm. One unit of enzyme activity (μg d⁻¹ g⁻¹) was defined as the production of 1 μg NO²⁻ per day per unit mass of soil sample (dry sample).

Soil nitrite reductase (S-NiR): S-NiR reduces NO²⁻ to NO, resulting in a reduction of NO²⁻ that participates in a diazotization reaction to produce a purplish-red compound. Change in the absorbance value at 540 nm corresponds to the activity of S-NiR in soil. One unit of enzyme activity (μg d⁻¹ g⁻¹) was defined as the reduction of 1 μmol NO²⁻ per day per unit mass of soil sample (dry sample).

Soil hydroxylamine reductase (S-HR): S-HR catalyzes the second step in the conversion of ammonia to nitrite via the intermediate hydroxylamine (NH₂OH). The Fe³⁺ in ferric ammonium sulfate oxidizes hydroxylamine to nitrogen gas and is reduced to Fe²⁺. The Fe²⁺ forms an orange-red complex with o-phenanthroline under weak acidic conditions, which has an absorption peak at 510 nm. The S-HR acts on hydroxylamine to reduce the amount of the complex produced, and the decrease in absorbance value at 510 nm is used to reflect the activity of S-HR. One unit of enzyme activity (μg d⁻¹ g⁻¹) was defined as the conversion of 1 μg hydroxylamine per day per unit mass of soil sample (dry sample).

2.4. Statistical Analyses

Statistical analyses were conducted using Microsoft Excel 2019 (Microsoft Inc., Redmond, WA, USA) and IBM SPSS Statistics v22.0 (SPSS Inc., Chicago, IL, USA). A mixed effects model with block as a random variable was used to assess the significance of different topsoil depths on grain yield, kernels per ear, 1000-kernel weight, NFUE, G-NFUE, soil N content (10 cm), ΔS, δS, S-NR, S-NiR, and S-HR, with the Tukey test used for multiple comparisons at a significance level of 0.05. Linear regression was performed to assess relations between grain yield and ΔS and between grain yield and δS.

3. Results

3.1. Grain Yield and Yield Components

Both year and topsoil depth had significant effects on grain yield and yield components, but their combined effect was not significant (

Table 1. Multivariate analysis of variance on grain yield and yield components. df1 and df2 represent the degrees of freedom of the numerator and the denominator, respectively.

Indicator	Statistics	Year (Y)	Topsoil Depth (D)	Y × D
Yield	df1	2	4	8
	df2	132	130	120
	F-value	7.1	70.1	1.4
	p-value	<0.001	0.003	0.247
Kernels per ear	df1	2	4	8
	df2	132	130	120
	F-value	9.6	49.5	3.2
	p-value	0.001	0.000	0.009
1000-kernel weight	df1	2	4	8
	df2	132	130	120
	F-value	32.3	26.8	1.1
	p-value	<0.001	<0.001	0.392

). Increasing topsoil depth significantly increased kernels per ear, 1000-kernel weight, and maize grain yields (

Table 2. The grain yield components, grain yield, sustainable yield index (SYI), and coefficient of variation of grain yield (CV) under different topsoil depth treatments (2021–2023) (Mean ± SD, n = 3). Topsoil depth treatments: D10, 10 cm; D20, 20 cm; D30, 30 cm; D40, 40 cm; D50, 50 cm. Different lowercase letters indicate significant differences (Tukey) between treatments in each year at p < 0.05.

Topsoil Treatment	Kernels per Ear	1000-Kernel Weight (g)	Grain Yield (g plant−1)	CV	SYI
D10	523.6 ± 41.8 c	276.0 ± 23.6 b	167.6 ± 15.4 b	0.92	0.78
D20	558.3 ± 34.3 b	284.4 ± 18.0 b	184.5 ± 15.6 b	0.85	0.79
D30	636.3 ± 27.5 a	316.3 ± 26.6 a	233.5 ± 14.8 a	0.63	0.86
D40	649.3 ± 19.6 a	328.1 ± 22.2 a	247.5 ± 14.5 a	0.59	0.86
D50	651.9 ± 43.0 a	331.0 ± 26.8 a	250.3 ± 19.4 a	0.78	0.81

). According to the average of three years (2021−2023), the number of kernels per ear increased by up to 128.3 kernels with increasing topsoil depth, and the maximum increases in 1000-kernel weight and maize grain yields with increasing topsoil depth were 19.9% and 49.4%, respectively, with values representing the differences between D10 and D50 treatments. The differences in grain yield under D30, D40, and D50 treatments were not significant, but yields in D30, D40, and D50 were all significantly higher than those in D10 and D20 treatments. Meanwhile, the coefficient of variation of grain yield (CV) of D30, D40, and D50 decreased by 0.29, 0.33, and 0.14, and the sustainable yield index (SYI) increased by 0.08, 0.08, and 0.03, respectively, compared to D10. Therefore, in this study, the threshold topsoil depth to ensure high yields of maize was 30 cm.

3.2. Nitrogen Fertilizer Use Efficiency of Maize Plants and Grain

Increasing topsoil depth effectively increased maize N fertilizer use efficiency (NFUE) and grain N fertilizer use efficiency (G-NFUE) (Figure 1). With increasing topsoil depth, the NFUE rises from 14.2% to 64.9% (between D10 and D50 treatments), while the G-NFUE climbs from 9.0% to 36.2% (between D10 and D40 treatments). The NFUE was not significantly different between D40 and D50 treatments, but it was significantly higher in D40 and D50 than in D10, D20, and D30 treatments. The G-NFUE was not significantly different among D30, D40, and D50 treatments, but it was significantly higher in D30, D40, and D50 than in D10 and D20 treatments. Therefore, in this study, NFUE and G-NFUE were maximized at topsoil depths greater than 30 cm.

3.3. Nitrogen Distribution in the Soil Profile

In this study, soil nutrient content was only analyzed above the nylon mesh interface, and therefore, the number of soil layers varied under the different topsoil depth treatments. Both year and topsoil depth had significant effects on soil (0–10 cm) N content (

Table 3. Multivariate analysis of variance on topsoil nitrogen of 0–10 cm. df1 and df2 represent the degrees of freedom of the numerator and the denominator, respectively.

Statistics	Year (Y)	Topsoil Depth (D)	Y × D
df1	2	4	8
df2	132	130	120
F-value	204.4	17.5	2.4
p-value	<0.001	<0.001	0.039

). Soil N content decreased each year of the three-year study (Figure 2). Nitrogen content of the effective soil (above the nylon mesh interface) was reduced by 32.3% in D10, 38.9% in D20, 43.9% in D30, 45.6% in D40, and 41.0% in D50 from 2021 to 2023, indicating increasing soil N depletion with deeper topsoil depth treatments. In addition, based only on the 0–10 cm depth, soil N content in D20, D30, D40, and D50 treatments was significantly higher than that in the D10 treatment. Based on the three-year averages, compared with D10, soil N content increased by 23.3% in D20, 26.9% in D30, 22.9% in D40, and 30.7% in D50 treatments. Thus, the decrease in topsoil depth increased N depletion in the topmost soil layer.

3.4. Soil Nitrogen Stock

In 2021 and 2022, increasing topsoil depth increased the loss of soil N stocks (ΔS) during the growth stage of maize but reduced the percentage of the loss in soil N stock (δS) (Figure 3). The mean value of ΔS of the D30, D40, and D50 treatments increased by 239.9 kg ha⁻¹ and 244.5 kg ha⁻¹ compared with ΔS values in D10 and D20 treatments, respectively, in 2021 (Figure 3A). The increases were 316.8 kg ha⁻¹ and 307.1 kg ha⁻¹, respectively, in 2022 (Figure 3B). The δS of D10 was significantly higher than that in the other treatments, with values 18.3, 10.3, 21.0, and 30.3 percentage points higher than those in D20, D30, D40, and D50 treatments, respectively, in 2021 (Figure 3C). The δS values of D10 were 18.1, 10.9, 19.9, and 10.7 percentage points higher, respectively, in 2022 (Figure 3D).

3.5. Relations Between Grain Yield and Soil Nitrogen Stock

Linear relations between yield and ΔS were significantly positive, whereas those between yield and δS were significantly negative (Figure 4). According to the regression equation, for every 1 kg N ha⁻¹ lost in the effective soil, maize grain yield increased by 15 kg ha⁻¹ in 2021 (Figure 4A) and by 8.6 kg ha⁻¹ in 2022 (Figure 4B), which indicated that the contribution from depletion of the soil N stock to the increase in maize grain yield decreased from year to year. However, for every 1% loss in N stock in the effective soil, maize grain yield decreased by 1184.2 kg ha⁻¹ in 2021 (Figure 4C) and by 2185.8 kg ha⁻¹ in 2022 (Figure 4D), which indicated that the relative reduction in the effective soil N stock limited the yield increase from year to year. Combined with the results of soil N stocks, the deeper topsoil depth treatments exhibited higher ΔS and lower δS than those of the shallow treatments, and thus, the deeper topsoil depth treatments were more favorable for increasing maize grain yields.

3.6. Distribution of Nitrogen Fertilizer in the Soil

The percentage of N fertilizer in soil N (S-N_season) indicated N from in-season fertilizer application migrated to deeper soils (Figure 5). The S-N_season of D20 and D30 treatments increased with soil depth and peaked at 20 cm and 30 cm depths, respectively; whereas the S-N_season of D40 and D50 treatments first increased and then decreased with soil depth, with S-N_season peaking in both treatments at 30 cm. The S-N_season at the 20-cm depth in the D20 treatment increased by 60% compared with that at the 10 cm depth, and at the 30-cm depth in D30, D40, and D50 treatments, it increased by 97.9%, 87.7%, and 95.9%, respectively, compared with that at the 10 cm depth. Therefore, in this study, a topsoil depth greater than 30 cm most benefited N fertilizer uptake by maize plants.

3.7. Key Enzymes of Soil Nitrogen Metabolism

Increasing topsoil depth significantly increased soil nitrate reductase (S-NR) and nitrite reductase (S-NiR) activities but did not significantly affect hydroxylamine reductase (S-HR) activity (Figure 6). Activities of S-NR and S-NiR increased significantly between treatments D10 and D50, with the increases reaching 135.1% and 48.2%, respectively (Figure 6A,B). Although S-NR activity was not significantly different among D30, D40, and D50 treatments, activities were all significantly higher than those in D10 and D20 treatments. The S-NiR activity was not significantly different between D40 and D50 treatments, but activities in both treatments were significantly higher than those in D10, D20, and D30 treatments. Therefore, in this study, the N metabolism in maize farmland soils was most stimulated by topsoil depth treatments of 30 to 40 cm.

4. Discussion

4.1. Increasing Topsoil Depth Improves Maize Grain Yield and Nitrogen Fertilizer Use Characteristics

Reduced topsoil depth directly limits crop yield increases [34]. In dryland crops such as wheat, soybean, and maize, removal of 5–10 cm of topsoil reduces grain yield by 5% to 13%, whereas removal of 20–30 cm of topsoil reduces grain yields by 37% to 73% [35,36]. In this study, the maximum increase in maize grain yields with increasing topsoil depth was 49.4%, and kernels per ear and 1000-kernel weight were mainly responsible for the increases in yield with increasing topsoil depth (Table 2). Differences in grain yield response to topsoil depth among various studies might be due to the crop species or environmental conditions. Previous studies indicate that insufficient soil and fertilizer N limit rice yield mainly by affecting tiller production, leaf area development, canopy light interception, canopy photosynthesis, and biomass production [37,38]. Thus, it is reasonable to expect that the limitation of topsoil depth on N uptake and utilization in maize is closely related to yield.

Reducing topsoil depth reduces crop N utilization, which reduces crop yield and quality [39]. Nitrogen, as one of the important mineral elements affecting maize yield and quality, contributes approximately 45% to maize yield [40]. In this study, increasing the topsoil depth significantly increased the NFUE and G-NFUE of maize plants (Figure 1). The results are consistent with those of previous studies that found increasing topsoil depth effectively improves the soil environment, which promotes crop root growth into deeper soils and increases the contact area between biological organs and the soil and thus improves nutrient absorption ability [41,42].

4.2. Increasing Topsoil Depth Optimizes Soil Nitrogen Content and Stock Characteristics

Nutrients in topsoil are important factors affecting soil fertility and crop yields in agricultural production systems, and topsoil degradation may increase the mineralization of organic nutrients, thereby increasing the processes of leaching and volatilization of nutrients and resulting in decreases in nutrient contents. According to previous research, deep tillage improves subsoil nutrient availability, thereby increasing crop yields in the presence of nutrient deficiencies in the topsoil [43]. Increasing topsoil depth increases the index of ecosystem multifunctionality in the soil layer at 20–40 cm depths and is significantly correlated with nutrient content [44]. The results of this study are consistent with that observation, because increasing topsoil depth significantly increased soil N content in the 0–10 cm topsoil layer (Figure 2). This result might be due to reduced root competition for soil nutrients with increasing topsoil depth, with roots exploiting nutrients from deeper soil layers (20–50 cm) and thereby reducing the extent of nutrient depletion in the 0–10 cm topsoil layer [43]. However, soil nutrient content generally decreases with soil depth, and the roots of deep-rooted crops generally do not take up enough nutrients from deeper soils to compensate for nutrient competition in the topsoil; thus, with adequate nutrient content, topsoil remains the most important source for crop uptake [45]. According to Artacho et al. [46], realizing high yield on high-fertility soil requires less N fertilizer than on low- and medium-fertility soils, with yields on low- and medium-fertility soils not reaching the yield levels on high-fertility soil even with high fertilizer N input. Therefore, reduced N content in the topsoil (0–10 cm) could be one of the mechanisms by which the reductions in topsoil depth limited maize grain yield in this study.

Estimates of soil N stocks are important for assessing potential nitrogen oxide emissions due to land-use changes and agricultural practices [47]. Soil N stocks are an important component of N cycles [48] and are a regulating factor in terrestrial C sequestration, with N also one of the most limiting nutrients in world agricultural production [49]. In general, a positive balance in the soil N pool can be guaranteed by fertilization in agricultural production systems [33]. In this study, the N pool in the effective soil in different topsoil depth treatments was a net N source, and soil N stock was depleted during the maize reproductive period (Figure 3). The lost N seemed to support the increase in maize yield in this study, because ΔS was significantly positively related to maize grain yield. However, the decrease in δS limited the increase in maize grain yield, and the degree of limitation increased from year to year (Figure 4). In this study, the deeper topsoil depth treatments exhibited higher ΔS and lower δS than those of the other treatments, and thus, the deeper topsoil depth treatments favored increases in maize grain yield.

4.3. Increasing Topsoil Depth Optimizes the Distribution Characteristics of Nitrogen Fertilizer in Topsoil

Generally, not all the N input into farmland is used by crops. The unused N input is typically discharged into the environment in runoff and leaching, which cause water pollution in rivers and groundwater [50]. Increasing topsoil depth can effectively increase N uptake by crops and reduce nitrate leaching, which are important for the protection of environmental resources such as soil and water [51]. An article discussing agricultural practices and topsoil decline highlights that topsoil erosion can indirectly increase leaching rates by reducing the soil’s capacity to retain nutrients [52]. In this study, increasing topsoil depth significantly increased the S-N_season at the 30-cm or 40-cm depth in the D30, D40, and D50 treatments (Figure 5). Although the fate of S-N_season from deeper soils under the shallow topsoil treatments (D10 and D20) was unclear in this study, the N in deeper soils under D10 and D20 treatments was clearly not available. Therefore, an important result of increasing the depth of topsoil is an increase in the space available to the crop to absorb fertilizer nutrients. When fertilizer nutrients were enriched at a depth of 30–40 cm, it matched the area of nutrient uptake by maize root systems [53]. Therefore, in D30, D40, and D50 treatments, the ability of maize root systems to take up N increased, thereby reducing fertilizer N losses. When nutrients in the topsoil are a limiting factor and affect crop growth and development, a deeper root system can disrupt the mechanical structure of soil and provide the crop with resources from the deeper soils, thus alleviating yield suppression.

4.4. Increasing Topsoil Depth Improves Soil Nitrification

Soil enzymes are involved in soil biochemical processes that affect nutrient mineralization and subsequent nutrient utilization by plants [54]. Variations in topsoil depth may contribute to general differences in soil microbial communities, as well as to differences in maize root biology, root secretions, and plant nutrient utilization strategies, ultimately leading to differences in crop regulation of soil enzyme activities [55,56,57]. In previous studies, soil N and P deficiencies stimulate enzyme catalysis and thus increase activities of urease, phosphatase, and invertase in degraded soils; however, nutrients mineralized by enzymes are not sufficient to alleviate nutrient limitations in maize [15,58]. In this study, increasing topsoil depth effectively increased soil nitrate reductase and nitrite reductase activities (Figure 6). The results are consistent with previous findings that deep plowing can effectively increase enzyme activity in deep soils, which in turn accelerates nutrient cycling in the subsoil [59]. In addition, maize plants with normal growth and development in deeper topsoil treatments may have higher nutrient requirements, which stimulates enzyme activity. However, changes in topsoil depth had little effect on hydroxylamine reductase activity, suggesting that topsoil depth mainly affected the process of soil nitrification, with nitrate N readily transported in soil [60].

5. Conclusions

Reduced topsoil depth is a substantial barrier to acquisition of soil resources by maize plants and leads to severe reductions in the efficiency of plant uptake and utilization of N. This mainly includes crop uptake and N migration to deeper soils, which lead to depletion of topsoil nutrient resources, ultimately limiting crop uptake. Increased depletion of N stocks in the effective soil reduces the ability of soil to supply N and limits the activity of key enzymes involved in soil N metabolism, which reduces the process of soil N mineralization. Therefore, increasing the depth of topsoil can effectively increase the source of nutrients absorbed by a crop by increasing access to additional resources stored in deeper soils, which ultimately increases maize grain yield and N fertilizer use efficiency (Figure 7). This study elucidates the relationship between soil depth and grain yield as well as nutrient uptake under specific conditions by examining different soil depths. In this study, the threshold to achieve the objective of high maize yield and efficiency was a topsoil depth of 30 cm under specific soil and climatic conditions. Further studies across various pedoclimatic conditions are required to elucidate the specific relationship between topsoil depth and grain yield.