Method for promoting rapid construction of acid soil aggregates by using green manure

Abstract

The application discloses a method for promoting rapid construction of acid soil aggregates by using green manure, which comprises the following steps: applying green manure and bacterial agent into acid soil, wherein the application amount of the green manure is 2g C kg ‑1 of soil, the bacterial agent is a bacterial agent of Penicillium ZC1 strain, and the use amount of the bacterial agent is greater than or equal to 5x10 9 cfu of ZC1 bacteria per kilogram of soil. The method can rapidly promote the formation of soil aggregates, improve the stability of the soil aggregates, and improve the pH value of the soil. The method can be combined with a calcium-containing conditioner, so that the pH value is better improved, the construction of the aggregates is not affected, and the improvement effect of the aggregates is better. The method can effectively utilize the green manure to promote the improvement of the soil, the construction of the soil aggregates is rapidly promoted, the content of soil organic carbon is not affected, the acid soil is greatly improved, and the method has a wide application prospect.

Description

Technical Field

[0001] This invention belongs to the field of soil acidification improvement technology, specifically relating to a method for promoting the rapid construction of aggregates in acidic soil using green manure. Background Technology

[0002] In soil improvement, methods such as returning straw to the field have traditionally been used to increase soil organic matter and nutrient content, improve soil physical properties, and promote crop root growth and development. However, straw decomposes slowly, and the presence of mixed weed seeds, insect eggs, and pathogens can easily induce pests and diseases. Compared with straw and other organic materials, green manure has the characteristics of a low carbon-to-nitrogen ratio and rapid microbial decomposition. After being applied to the soil, it decomposes and releases nutrients through mineralization, increasing soil organic matter, improving soil structure, and promoting nutrient cycling, thus ensuring stable and high crop yields.

[0003] Soil aggregates are structural units formed by the aggregation of soil particles through various physical, chemical, and biological processes. Soil aggregates are crucial for soil health and function, influencing its physical, chemical, and biological properties. The stability of aggregates directly affects the water-soil interface behavior at the soil surface, particularly its relationship with rainfall infiltration and soil erosion. Soil aggregates possess porosity and water stability, maintaining a harmonious balance between the solid, liquid, and gaseous phases of the soil; therefore, aggregates are often considered an indicator of soil fertility.

[0004] Acidic soils have poor site conditions, shallow soil layers, and weakened soil aggregates and stability. They are characterized by acidity, poor soil quality, and stickiness, further leading to insufficient soil nutrient supply, particularly a significant reduction in soil organic carbon, resulting in a severe decline in soil productivity and crop quality. Green manure is rich in nutrients and has a low carbon-to-nitrogen ratio, but its ability to improve acidic soils is limited. Therefore, promoting the improvement of acidic soils with green manure is particularly important, especially in promoting the formation of soil aggregates, which is one of the indicators of soil fertility. Summary of the Invention

[0005] Therefore, one objective of this invention is to provide a method for promoting the rapid construction of aggregates in acidic soil using green manure, comprising applying green manure in combination with a microbial agent into acidic soil, wherein the amount of green manure applied is 2g / kg. -1 The soil inoculant is a Penicillium ZC1 strain inoculant, and the dosage of the inoculant is greater than or equal to 5 × 10⁻⁶ per kilogram of soil. 9ZC1 strain of CFU. The ZC1 strain is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 40237, dated July 1, 2022, at the Institute of Microbiology, Chinese Academy of Sciences, No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing. The Penicillium fungicide can be prepared as follows: ZC1 strain is streaked onto potato dextrose agar (PDA) at 28°C for 7 days; then, single colonies are picked and cultured in PDA liquid medium on a shaker to prepare a bacterial suspension. The bacterial concentration is determined using a hemocytometer. After centrifugation, the original medium is discarded, and the bacteria are resuspended in 1 / 10 PDA medium, with the bacterial concentration adjusted to be greater than 10. 8 cfu / mL. The amount of green manure added is 2g C kg. -1 The amount of soil added is equivalent to an average of 30,000 kg / ha of green manure per year. -2 The amount of decomposition of fresh grass in the 0-20cm soil layer.

[0006] Preferably, the soil is purple soil or red soil with a pH value of 5-5.5.

[0007] Preferably, the green manure is *Vigna glabra*. *Vigna glabra* requires pretreatment, including collecting *Vigna glabra* plants, first blanching the fresh samples in an oven at 105℃ for 30 minutes, then cooling to 60-70℃ until dried, and finally crushing the plants to <0.25mm for subsequent experiments.

[0008] Preferably, the proportions of sand, silt, and clay particles in the purple soil are 20.6%, 38.1%, and 41.3%, respectively, and the proportions of sand, silt, and clay particles in the red soil are 25.8%, 45.4%, and 28.8%, respectively.

[0009] Preferably, the green manure applied to the soil is also combined with a calcium-containing conditioner.

[0010] Preferably, the calcium-containing conditioner is quicklime, and the amount used is 0.1% of the soil. The amount of quicklime used should not be too high, as this will affect the effectiveness of ZC1 bacteria.

[0011] Preferably, the aggregate is a water-stable aggregate.

[0012] A second objective of this invention is to provide the application of Penicillium ZC1 (accession number: CGMCC No. 40237) in promoting soil aggregate formation. Experiments based on this invention have shown that Penicillium ZC1 can improve soil and promote soil aggregate formation.

[0013] Preferably, the soil is acidic soil.

[0014] This invention improves acidic purple soil and red soil by combining green manure with Penicillium ZC1 inoculant to promote the decomposition of green manure. Extensive experiments have verified that this method, which utilizes green manure to rapidly construct soil aggregates in acidic soils, significantly improves the quality and stability of soil aggregates, and significantly increases the proportion of water-stable aggregates larger than 0.25 mm. The combination of calcium-containing conditioners and inoculants not only promotes the formation of large soil aggregates but also significantly increases soil pH. This invention effectively utilizes green manure to improve soil, rapidly promoting soil aggregate construction without affecting soil organic carbon content, and significantly increasing soil pH, thus greatly improving acidic soils and showing broad application prospects. Attached Figure Description

[0015] Figure 1 The effect of adding calcium-containing conditioners and microbial agents to promote the growth of green manure on the content of >0.25mm water-stable aggregates in acidic purple soil and red soil;

[0016] Figure 2 The effect of adding calcium-containing conditioners and microbial agents to promote the organic carbon content of green manure on acidic purple soil and red soil;

[0017] Figure 3 The effects of adding calcium-containing conditioners and microbial agents on the pH of acidic purple soil and red soil were investigated. Detailed Implementation

[0018] The present invention will be described in detail below with reference to embodiments. These embodiments are for illustrative purposes only and are not intended to limit the scope of application of the present invention. The present invention is not limited to the following embodiments or examples. Any modifications and variations made without departing from the spirit of the present invention should be included within the scope of the present invention.

[0019] I. Material Preparation

[0020] 1. Materials: Green manure, calcium-containing conditioner, microbial agent, and soil. The green manure was *Vigna glabra*, the calcium-containing conditioner was quicklime powder, the microbial agent was *Penicillium* ZC1 inoculum (ZC1 strain has been deposited at the China General Microbiological Culture Collection Center, accession number CGMCC No. 40237, deposit date July 1, 2022, deposit address: Institute of Microbiology, Chinese Academy of Sciences, No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing), and the soil consisted of acidic purple soil and acidic red soil.

[0021] 2. Material preparation:

[0022] Vetch (Vigna glabra): Physicochemical properties: Organic carbon content 341.9 g·kg⁻¹ -1 Total nitrogen content: 27.7 g·kg -1Before the experiment, Vigna glabra plants were collected. Fresh samples were first blanched in an oven at 105℃ for 30 minutes, then cooled to 60-70℃ until dried. The plants were then crushed to <0.25mm for subsequent experiments.

[0023] Penicillium ZC1 inoculum: Penicillium grows well at 25-35℃, but growth is limited above 50℃. Penicillium ZC1 was streaked onto potato dextrose agar (PDA) at 28℃ for 7 days. Single colonies were then picked and cultured in PDA liquid medium on a shaker to prepare a bacterial suspension. The bacterial concentration was determined using a hemocytometer. After centrifugation, the original medium was discarded, and the cells were resuspended in 1 / 10 PDA medium, with the bacterial concentration adjusted to be greater than 10. 8 cfu / mL, to obtain Penicillium ZC1 inoculum.

[0024] The tested soils were acidic purple soil and acidic red soil: the purple soil was collected from Jiangjin District, Chongqing (29°28′N, 105°49′E), and the red soil was collected from Chuxiong Prefecture, Yunnan Province (24°30′N, 100°35′E). Soil samples from the 0-40cm layer were collected, and after removing visible plant and animal remains, they were air-dried at room temperature, passed through a 2mm sieve, and mixed thoroughly. A portion was used for determining the basic physicochemical properties of the soils, and the remainder was used for the experiments. The basic physicochemical properties of the tested soils are shown in Table 1. In the acidic purple soil, the proportions of sand, silt, and clay particles were 20.6%, 38.1%, and 41.3%, respectively; in the acidic red soil, the proportions were 25.8%, 45.4%, and 28.8%, respectively.

[0025] Table 1 Physicochemical properties of the tested soils

[0026]

[0027]

[0028] II. Experimental Methods

[0029] Application: Green manure, calcium-containing conditioner, and microbial agent were applied to the tested soil. The amount of green manure applied was 2g / kg. -1 For the soil, the calcium conditioner is added at 0.1% of the soil volume, and the microbial agent is added at 5 × 10⁻⁶ per kilogram of soil. 9 CFU; The control group consisted of no green manure, a calcium-containing conditioner, and a microbial agent. Five treatment groups were specifically set up:

[0030] (1) CK: No material is applied;

[0031] (2) G: Apply green manure;

[0032] (3)GL: Apply green manure with quicklime;

[0033] (4) GP: Apply green manure in combination with Penicillium ZC1 inoculant;

[0034] (5) GLP: Apply green manure in combination with quicklime and Penicillium ZC1 inoculant.

[0035] Each treatment was repeated three times. Green manure was added at a rate of 2g C / kg. -1 For soil addition, the added material is mixed evenly in the soil, minimizing the breakage of aggregates. To maintain uniform conditions, the same procedure is performed on the soil in the control group. The soil-material mixture is then gently poured into culture flasks (250 mL), maintaining a bulk density of 1.2 g·cm³. -3 The soil was then moistened to 60% of field capacity. All culture bottles were stored in a 4°C incubator for 3 days to allow for water equilibration under conditions of minimal soil microbial activity, and then transferred to a 25±2°C incubator for 56 days. Throughout the experiment, the culture bottles were weighed periodically, and deionized water was added using a dropper to control soil moisture content. On days 0, 7, 14, 28, and 56, destructive sampling was performed to determine the dynamic changes in soil aggregates. The determination method is described in Part III, “Detection Methods,” below.

[0036] III. Detection Methods

[0037] 1. Determination and calculation of soil aggregates

[0038] The collected soil samples were placed on the top layer of a high-frequency vibrating sieve (from top to bottom, 2mm, 0.25mm, and 0.053mm) for dry sieving for 5 minutes to separate air-dried aggregates of different particle sizes (>2mm, 0.25-2mm, 0.053-0.25mm, and <0.053mm). The weights of each aggregate were recorded and stored for later use. The aggregates obtained from dry sieving were then mixed into 50g samples for wet sieving analysis. For the determination of water-stable aggregates, a wet sieving method was used. The prepared soil sample (50g) was placed on the top layer of the sieve (from top to bottom, 2mm, 0.25mm, and 0.053mm) to separate soil aggregates of four particle sizes: >2mm, 0.25-2mm, 0.053-0.25mm, and <0.053mm. The agglomerates at each level on the sieve were washed into a pre-weighed aluminum box. Then, each component was dried in an oven at 40°C until constant weight, cooled to room temperature, weighed, and stored in a self-sealing bag. Finally, four parts were obtained: viscous powder particles (<0.053mm), micro-agglomerates (0.25-0.053mm), fine and large agglomerates (2-0.25mm), and coarse and large agglomerates (>2mm).

[0039] Soil aggregate stability is described by two indicators: average weight diameter of aggregates (MWD) and the proportion of water-stable aggregates >0.25 mm (WSA). The calculation method is as follows:

[0040] In the formula Let be the average diameter (mm) of the i-th size agglomerate, and wi be the mass fraction (%) of the i-th size agglomerate.

[0041] WSA = (Mr>0.25) / MT = 1 - (Mr<0.25) / MT, where Mr<0.25 is the weight (g) of agglomerates with a particle size less than 0.25 mm, Mr>0.25 is the weight (g) of agglomerates with a particle size greater than 0.25 mm, and MT is the total weight (g) of agglomerates.

[0042] 2. Measurement of soil organic carbon and pH value

[0043] Determination of soil organic carbon content: Naturally air-dried soil was ground and passed through a 100-mesh sieve. The soil organic carbon content was determined using the potassium dichromate external heating method. Soil pH was determined using the potentiometric method.

[0044] IV. Experimental Results

[0045] 1. Soil water-stable aggregates

[0046] The average mass diameter of soil aggregates and the content of water-stable aggregates >0.25 mm are important parameters characterizing the size distribution of soil aggregates. The higher the value, the higher the degree of soil aggregation, and the stronger the structural stability and erosion resistance.

[0047] The differences in aggregate size distribution among different treatments in purple soil and red soil are shown in Tables 2 and 3. Compared with the control (CK), all treatments increased the mass fraction of soil aggregates with particle sizes >2 mm and 0.25-2 mm. In purple soil, the content of macroaggregates (>0.25 mm) in the G, GL, GP, and GLP treatments increased in the first two weeks and then stabilized. Among them, the content of macroaggregates (>0.25 mm) in the GP and GLP treatments increased more rapidly. The average content of macroaggregates (>0.25 mm) in each treatment group at 7, 14, 28, and 56 days was obtained by summing the contents of macroaggregates (>0.25 mm) in each treatment group. Compared with the control (CK), the average content of macroaggregates (>0.25 mm) in G, GL, GP, and GLP increased significantly by 74.3%, 92.0%, 98.4%, and 109.9%, respectively. Figure 1 (Left). In red soil, the content of macroaggregates (>0.25 mm) in the G, GL, GP, and GLP treatments increased in the first two weeks and then stabilized. Similarly, the content of macroaggregates (>0.25 mm) in the GP and GLP treatments increased more rapidly. Compared with the control (CK), the average content of macroaggregates (>0.25 mm) increased significantly by 27.6%, 33.1%, 49.0%, and 56.2%, respectively. Figure 1(Right). In both soil types, all treatments containing green manure showed a significant increase in large aggregate content. Among them, the treatments GP and GLP, which were combined with microbial agents, showed a faster and greater increase in large aggregates (>0.25 mm), with GLP showing better results.

[0048] Table 2. Effects of different treatments on the content of aggregates of different particle sizes in purple soil.

[0049]

[0050]

[0051] Note: At four different particle size levels, different letters in a column indicate significant differences in the percentage of aggregate content between different treatments, P<0.05.

[0052] Table 3. Effects of different treatments on the content of aggregates of different particle sizes in red soil.

[0053]

[0054] Note: At four different particle size levels, different letters in a column indicate significant differences in the percentage of aggregate content between different treatments, P<0.05.

[0055] Table 4 shows the differences in mean weight diameter of aggregates in purple soil and red soil among different treatments. In both purple soil and red soil, all treatments increased the mean weight diameter of aggregates to varying degrees compared to the control (CK). The mean weight diameter peaked on day 7 and remained relatively stable for the remainder of the incubation period. Different treatments with added green manure significantly increased the mean weight diameter, showing a pattern of GLP>GP>GL>G>CK. In purple soil, the overall mean weight diameter of aggregates in the G, GL, GP, and GLP treatments (the average of the mean weight diameters at 7, 14, 28, and 56 days) was significantly increased by 48.3%, 61.4%, 66.9%, and 77.2% compared to the CK, respectively. In red soil, the overall mean weight diameter of aggregates in the G, GL, GP, and GLP treatments was significantly increased by 21.0%, 24.2%, 39.3%, and 43.8% compared to the CK, respectively.

[0056] In summary, soil aggregation can occur rapidly within a short period of time after the addition of organic materials. The addition of microbial agents has a significant promoting effect on the formation of soil aggregates. Furthermore, the effect of aggregate formation is even more pronounced when combined with calcium-containing conditioners on the basis of microbial agents.

[0057] Table 4. Effects of different treatments on the average weight diameter (mm) of purple soil and red soil.

[0058]

[0059] Note: At four different particle size levels, different letters in a column indicate significant differences in the average weight diameter of aggregates between different treatments, P<0.05.

[0060] 2. Soil organic carbon content

[0061] Figure 2 To determine the organic carbon content of purple soil and red soil under different treatments, the soil was cultured for 7-56 days. Compared with the control (CK), all treatments increased the soil organic carbon content to varying degrees, but the differences between treatment groups were not significant. This indicates that calcium-containing conditioners and microbial agents had no significant effect on the organic carbon content of soils with added green manure. In fact, the organic carbon content of GL and GLP in red soil was slightly lower than that of other treatments, suggesting that the promotion of soil aggregates was not promoted by organic matter.

[0062] 3. Soil pH value

[0063] Figure 3 The pH values ​​of purple soil and red soil under different treatments are shown. Soil pH can directly characterize the intensity of soil active acidity. Without green manure application, the pH of purple soil was 5.04 (data is the average over 7-56 days, the same below), and the pH of red soil was 5.31. Green manure significantly increased soil pH. In purple soil, the GLP and GL treatments had the highest pH, significantly increasing by 21.6% and 18.7% compared to the control (CK), followed by the GP and G treatments, increasing by 8.9% and 6.0%, respectively. In red soil, the GL and GLP treatments had the highest pH, significantly increasing by 13.7% and 13.0% compared to the control (CK), followed by the GP and G treatments, increasing by 6.2% and 5.1%, respectively. This indicates that green manure can increase the pH of acidic purple soil and red soil. Combining it with calcium-containing conditioners and microbial agents can better reduce soil acidity, especially with calcium-containing conditioners, which effectively neutralize soil acidity and significantly increase soil pH.

[0064] In summary, the results show that green manure vetch can promote the formation of soil aggregates, increase soil organic carbon content, and improve soil pH. Penicillium ZC1 inoculant can promote better and faster formation of soil aggregates in soil containing green manure vetch, but since it has no significant effect on organic carbon content, it indicates that Penicillium ZC1 primarily promotes soil aggregate formation and thus improves soil structure. Furthermore, the addition of calcium conditioner, along with Penicillium ZC1, has a better effect on promoting soil aggregate formation and simultaneously increasing soil pH.

[0065] The conventional techniques and solutions not described in detail in the above embodiments are all well known in the art, and therefore will not be elaborated upon here. The above experimental examples describe the preferred embodiments of the present invention in detail. However, the present invention is not limited to the specific details of the above embodiments. Within the scope of the technical concept of the present invention, various simple modifications can be made to the technical solutions of the present invention, and these simple modifications all fall within the protection scope of the present invention.

Claims

1. A method for promoting the rapid construction of aggregates in acidic soil using green manure, characterized in that, This includes applying green manure mixed with microbial agents to acidic soil, wherein the amount of green manure applied is 2 g / kg. -1 The soil inoculant is a Penicillium ZC1 strain inoculant, and the application rate of the inoculant is greater than or equal to 5 × 10⁻⁶ per kilogram of soil. 9 The cfu ZC1 strain; the preservation number of the Penicillium ZC1 strain is CGMCC No. 40237; the green manure is Vigna glabra.

2. The method as described in claim 1, characterized in that, The soil is purple soil or red soil with a pH value of 5-5.

5.

3. The method as described in claim 2, characterized in that, The proportions of sand, silt, and clay in the purple soil are 20.6%, 38.1%, and 41.3%, respectively, while the proportions of sand, silt, and clay in the red soil are 25.8%, 45.4%, and 28.8%, respectively.

4. The method as described in claim 1, characterized in that, The green manure applied to the soil is also combined with a calcium-containing conditioner.

5. The method as described in claim 4, characterized in that, The calcium-containing conditioner is quicklime, and the amount used is 0.1% of the soil composition.

6. The method according to any one of claims 1-5, characterized in that, The aggregates are water-stable aggregates.

7. The application of Penicillium ZC1 strain in promoting soil aggregate formation, characterized in that, The preservation number of the Penicillium ZC1 strain is CGMCC No. 40237; the soil is acidic soil.

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