Preparation and application of paenibacillus polymyxa microbial inoculum for promoting growth of degraded grassland elymus dahuricus
By combining Bacillus polymyxa YF inoculant with chemical or organic fertilizers, the problems of inhibited growth of crested wheatgrass and soil degradation have been solved, resulting in improved yield and quality of crested wheatgrass and soil improvement. This method is suitable for forage planting and ecological restoration in cold and barren areas of northern China.
Patent Information
- Application Number
- CN202511420073.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-09-30
- Publication Date
- 2026-02-17
AI Technical Summary
In the arid and infertile environment of northern China, the growth of crested wheatgrass is inhibited. The long-term use of chemical fertilizers has led to the deterioration of soil physical and chemical properties and the imbalance of micro-ecology. The existing green fertilizers are insufficient and cannot meet the needs of high-quality development of the forage industry and ecological protection.
By combining Bacillus polymyxa YF inoculant with chemical or organic fertilizers, and then sowing the seeds after soaking them, the growth of Leymus chinensis can be promoted and the physical and chemical properties of the soil can be improved.
It significantly improves the yield and quality of crested wheatgrass, improves soil properties, reduces the risk of soil compaction, enhances stress resistance, maintains micro-ecological balance, is suitable for large-scale promotion, and is low-cost and environmentally friendly.
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Figure CN121533416A_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The application relates to the field of microbial agent preparation, in particular to application of Paenibacillus polymyxa YF in Qinghai degraded grassland soil remediation and growth promotion of elytrigia. BACKGROUND
[0002] Elytrigia is a high-quality forage with a long history of artificial cultivation in Qinghai region, and has been applied in local livestock production and ecological restoration for more than ten years. Elytrigia has high biomass and is rich in high-quality nutrients such as crude protein and crude fiber, which can not only provide high-cost-performance forage for livestock, but also effectively reduce soil erosion. It is not only an important forage guarantee for regional livestock development, but also a key vegetation for maintaining grassland ecological balance, and has very high production value and ecological value.
[0003] In recent years, with the expansion of livestock scale and the increasing demand for ecological restoration in Qinghai region, the planting area of elytrigia has been expanding year by year, and the planting area has also been extended to surrounding plots with more complex soil conditions. However, due to the environmental conditions of drought, poor soil and unbalanced nutrients in the north, combined with the long-term reliance on chemical fertilizers to increase yield, the soil physical and chemical properties have deteriorated, and the growth of elytrigia has been significantly restricted. At the same time, the risk of soil pathogenic bacteria breeding is increasing, which further affects the yield and nutritional quality of forage, seriously restricting the green development of local forage industry and the promotion of ecological restoration work.
[0004] Chemical fertilizer is a common means to improve soil fertility in elytrigia planting at present, but long-term single application not only leads to soil compaction and reduction of organic matter content, but also causes imbalance of nitrogen, phosphorus and potassium nutrients, and aggravates soil degradation. In addition, the excessive use of chemical inputs also destroys the micro-ecological balance of grassland soil, reduces the number of beneficial microorganisms, and forms a vicious cycle of "the more fertilizer used, the more barren". At the same time, there are few special green fertilizer improvement and stress regulation products for elytrigia planting, which is difficult to meet the dual needs of high-quality development and ecological protection of the current forage industry.
[0005] The Paenibacillus polymyxa YF agent described in the present application, combined with the base fertilizer of chemical fertilizer and organic fertilizer, can effectively promote the growth of elytrigia and significantly improve the soil physical and chemical properties after seed soaking and sowing. The combination of the agent and base fertilizer can not only be used as a green planting aid, but also provide technical support for forage ecological planting, and has broad market application prospects in the field of forage industry and ecological restoration in Qinghai and even the north. SUMMARY
[0006] In view of the deficiencies in the prior art, the purpose of the present application is to provide the application of Paenibacillus polymyxa YF or its fermentation liquor in promoting the growth of Elymus nutans, and the Paenibacillus polymyxa YF was preserved in the China General Microbiological Culture Collection Center on December 21, 2020, with a preservation number of CGMCC No. 21517.
[0007] Preferably, the Paenibacillus polymyxa YF is used in combination with a fertilizer.
[0008] Preferably, the fertilizer comprises one or more of chemical fertilizer, potassium fertilizer, and organic fertilizer.
[0009] The second purpose of the present application is to provide the application of Paenibacillus polymyxa YF or its fermentation liquor in increasing the yield of Elymus nutans, and the Paenibacillus polymyxa YF was preserved in the China General Microbiological Culture Collection Center on December 21, 2020, with a preservation number of CGMCC No. 21517.
[0010] The third purpose of the present application is to provide the application of Paenibacillus polymyxa YF or its fermentation liquor in increasing the plant length and leaf width, and the Paenibacillus polymyxa YF was preserved in the China General Microbiological Culture Collection Center on December 21, 2020, with a preservation number of CGMCC No. 21517.
[0011] The fourth purpose of the present application is to provide the application of Paenibacillus polymyxa YF or its fermentation liquor in increasing the fresh weight content of aboveground and underground parts, and the Paenibacillus polymyxa YF was preserved in the China General Microbiological Culture Collection Center on December 21, 2020, with a preservation number of CGMCC No. 21517.
[0012] The fifth purpose of the present application is to provide the application of Paenibacillus polymyxa YF or its fermentation liquor in increasing the dry weight content of aboveground and underground parts, and the Paenibacillus polymyxa YF was preserved in the China General Microbiological Culture Collection Center on December 21, 2020, with a preservation number of CGMCC No. 21517.
[0013] The sixth objective of this invention is to provide the application of Paenibacillus polymyxa YF or its fermentation broth in the preparation of a growth promoter for Leymus chinensis. The Paenibacillus polymyxa YF was deposited on December 21, 2020, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 21517.
[0014] The seventh objective of this invention is to provide a method for promoting the restoration of degraded grasslands, characterized in that Paenibacillus polymyxa YF and other fertilizers are applied to the degraded grasslands. The Paenibacillus polymyxa YF was deposited at the China General Microbiological Culture Collection Center on December 21, 2020, with accession number CGMCC No. 21517. The fertilizers include organic fertilizers and / or inorganic fertilizers.
[0015] By adopting the above technical solution, the present invention has achieved the following beneficial effects:
[0016] The method described in this invention can significantly increase the yield of *Elymus chinensis*, increase plant height and leaf width, and increase fresh and dry weight. Compared with the control group that applied chemical fertilizer or organic fertilizer alone, the compound combination can increase the key growth indicators (plant height, leaf width, fresh weight, and dry weight) of *Elymus chinensis* by more than 17%, especially in cold and arid environments, it can effectively alleviate growth inhibition and ensure forage yield. Regarding the improvement of soil physical and chemical properties: when combined with Formula 1 (chemical fertilizer-type base fertilizer): soil pH increases by 5.55%, and organic matter... The first formula increased available potassium by 0.087%, available phosphorus by 18.66%, available phosphorus by 22.37%, and total nitrogen by 9.62%. When combined with formula two (organic-inorganic compound base fertilizer), soil pH increased by 8.55%, organic matter by 8.46%, available potassium by 42.84%, available phosphorus by 22.8%, available nitrogen by 9.65%, and total nitrogen by 12.57%. At the same time, it reduced excess total potassium (reduced by 12.86%) and total phosphorus (reduced by 9.33%) in the soil, thus alleviating nutrient imbalance. It solves the core problems of soil infertility and insufficient nutrient supply in northern pasture planting areas; at the same time, it adjusts the soil pH to a suitable range, improves soil aeration and water retention capacity, and avoids soil compaction caused by single chemical fertilizers; it replaces some chemical inputs with Bacillus polymyxa YF inoculant, eliminating the risk of environmental pollution and pesticide residues, while also increasing the abundance of beneficial soil microorganisms, maintaining the micro-ecological balance of grassland soil, and contributing to the ecological restoration of degraded grasslands and the sustainable development of the pasture industry; the application cost is low and the operation is simple: the base fertilizer (chemical fertilizer, organic fertilizer) and Bacillus polymyxa YF inoculant are readily available, the preparation process does not require complex equipment, and the seed soaking and base fertilizer application process is simple, making it suitable for large-scale pasture planting and promotion in areas such as Qinghai, saving manpower and material costs. Attached Figure Description
[0017] Figure 1 Differences in plant height of Leymus chinensis in different treatment plots
[0018] Figure 2 Differences in leaf width of Leymus chinensis in different treatment plots
[0019] Figure 3 Differences in fresh weight of Leymus chinensis in different treatment plots
[0020] Figure 4 Differences in dry weight of Leymus chinensis in different treatment plots
[0021] Figure 5 Differences in total nitrogen in soil from different treatment plots
[0022] Figure 6 Differences in total potassium in soil from different treatment plots
[0023] Figure 7 Differences in total phosphorus in soil from different treatment plots
[0024] Figure 8 Differences in available potassium in soil among different treatment plots
[0025] Figure 9 Differences in soil organic matter in different treatment plots
[0026] Figure 10 Differences in available nitrogen in soil among different treatment plots
[0027] Figure 11 Differences in available phosphorus in soil among different treatment plots
[0028] Figure 12 Differences in soil pH among different treatment plots Detailed Implementation
[0029] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention.
[0030] It should be noted that, unless otherwise specified, the methods described in the following embodiments are all conventional methods, and the reagents described are all commercially available.
[0031] It should be noted that in the following embodiments, the *Paenibacillus polymyxa* YF is deposited at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, with accession number CGMCC No. 21517 and contact number 010-64807355. In the following embodiments, *Paenibacillus polymyxa* YF will be abbreviated as *Paenibacillus polymyxa* YF.
[0032] In the following embodiments, the effective viable count of the Bacillus polymyxa YF liquid bacterial agent is ≥10. 9 CFU / mL.
[0033] In the following examples, the fermentation medium for Bacillus polymyxa YF in the composite combination is LB liquid medium, with a formulation of 1% peptone, 0.5% yeast extract, 1% sodium chloride, and the remainder being sterile water.
[0034] In the following examples, the specific fermentation conditions were: 30-32℃, 180-200rpm, and shaking culture for 24-36h.
[0035] In the following examples, the Bacillus megaterium de Bary was isolated by the inventor's research group and preserved in the laboratory.
[0036] In the following embodiments, the *Brevibacillus laterosporus* was isolated by the inventor's research group and preserved in the laboratory.
[0037] Example 1: A method for promoting the growth of crested wheatgrass and improving planting soil using a compound of Bacillus polymyxa YF-base fertilizer.
[0038] The method includes the following steps:
[0039] Step 1: Base fertilizer preparation: Formula 1: Pour the chemical fertilizer (agricultural grade, 10 kg / mu) and potassium fertilizer (agricultural grade, 10 kg / mu) into a mixing container, and stir at 180-200 rpm for 20 minutes at room temperature to ensure that the particles are evenly mixed and free of lumps; If formula 2 is selected, first pour the organic fertilizer (fully decomposed, crushed to a particle size ≤5 mm, 300 kg / mu) into the container, then add the chemical fertilizer and potassium fertilizer, and stir at 200-220 rpm for 30 minutes at room temperature to fully integrate the organic matrix and inorganic nutrients;
[0040] Step 2: Preparation of *Bacillus polymyxa* YF inoculum: Using LB liquid medium (1% peptone, 0.5% yeast extract, 1% sodium chloride, balance sterile water), inoculate *Bacillus polymyxa* YF into the medium. Control the fermentation conditions at 30-32℃, 180-200 rpm, and shake for 36-48 hours until the OD600 value of the inoculum reaches 1.5-1.8 and the viable cell concentration is ≥10⁻⁶. 9 CFU / mL, prepared as a liquid microbial inoculant;
[0041] Step 3: Seed treatment: Take the Bacillus polymyxa YF liquid inoculant prepared in Step 2, pour it into the seed soaking container, and completely immerse the Leymus chinensis seeds in the inoculant. Control the ambient temperature at 20-25℃ and soak for 24 hours, stirring gently once every 8 hours. After soaking, transfer the seeds to a cool and ventilated place to dry, avoiding direct sunlight to prevent germination or mold.
[0042] Step 4: Base fertilizer application and sowing preparation: The base fertilizer prepared in Step 1 is evenly spread on the surface of the experimental field soil. Mechanical tillage is used to mix the base fertilizer with the 0-20cm soil layer, ensuring even nutrient distribution and laying the foundation for subsequent sowing and microbial colonization. Comparative Example 1: A method for promoting the growth of *Leymus chinensis* and improving planting soil using a *Bacillus megaterium*-base fertilizer composite combination.
[0043] The method includes the following steps:
[0044] The other methods are the same as in Example 1, except that Bacillus polymyxa YF is replaced with Bacillus megaterium.
[0045] Comparative Example 2: A method for promoting the growth of Leymus chinensis and improving planting soil using a Bacillus lateralis-base fertilizer compound.
[0046] The method includes the following steps:
[0047] The other methods are the same as in Example 1, except that Bacillus polymyxa YF is replaced with Bacillus lateralis.
[0048] Example 2
[0049] The following experiments were conducted using the composite combinations prepared in Example 1, Comparative Example 1, and Comparative Example 2, respectively.
[0050] The seeds of *Leymus chinensis* were derived from a high-quality variety selected locally in Haiyan County, Qinghai Province. These were healthy seeds harvested from the same batch, with a germination rate ≥85%. Experimental location: Forage planting base in the No. 7 Factory Area of the Atomic City Scenic Area, Haiyan County, Qinghai Province (36°57′27″N, 100°51′33″E). The soil type in this area is shale granular soil, with a basic pH of 6.5-7.0, organic matter content of 1.5-2.0%, and available potassium content of 50-80 mg / kg. Experimental area: 10 mu (approximately 1.65 acres).
[0051] Experimental groups: Empty shell treatment group (CK): Only formula one base fertilizer (10 kg / mu of chemical fertilizer + 10 kg / mu of potassium fertilizer) was applied, and the seeds were soaked in sterile water for 24 hours (20-25℃) before sowing;
[0052] Chemical fertilizer group (CF): Only one base fertilizer of formula 1 is applied, and seed microbial agent is used for treatment;
[0053] Organic fertilizer group (OF): Apply base fertilizer of formula two (300 kg / mu of organic fertilizer + 10 kg / mu of chemical fertilizer + 10 kg / mu of potassium fertilizer), and soak seeds in sterile water for 24 hours (20-25℃) before sowing.
[0054] Fertilizer group (SF): Apply base fertilizer of formula two (300 kg / mu of organic fertilizer + 10 kg / mu of chemical fertilizer + 10 kg / mu of potassium fertilizer), and sow seeds after treating them with microbial agents;
[0055] Experimental implementation: Base fertilizer application and tillage were completed on May 10, 2025. On May 15, the seeds of *Leymus chinensis*, which had been soaked and dried in *Bacillus polymyxa* YF inoculant, were evenly sown on the soil surface. After sowing, the seeds were lightly rake and covered with a 1-2 cm layer of soil. The sowing density was 2.5 kg / mu. No other fertilizers or inoculants were applied during the growth period.
[0056] (1) Monitoring experiment on the growth of Leymus chinensis
[0057] On July 22, 2025 (2 months after sowing) and August 15, 2025 (3 months after sowing, harvest period), the growth indicators of *Leymus chinensis* in each treatment group were measured: plant height (from the ground to the top of the plant) was measured with a ruler, and leaf width (maximum width of the third pair of fully expanded leaves at the top) was measured with a vernier caliper. After harvest, the fresh weight of each plant was weighed, and the dry weight was measured after drying at 65℃ to constant weight. Three quadrats (1m×1m) were randomly selected from each group, and 10 plants were measured in each quadrat, and the average value was taken.
[0058] Test results of different strains, such as Figures 1-4As shown in Figure 1 and Table 2, compared with Bacillus laterosporus and Bacillus megaterium, the combined application of organic fertilizer and Bacillus polymyxa inoculant during forage planting increased the height, leaf width, fresh weight, and dry weight of Leymus chinensis by 19.76%, 20.33%, 17.85%, and 17.75%, respectively. The growth data indicate that the Bacillus polymyxa YF-base fertilizer compound combination has a significant promoting effect on the growth of Leymus chinensis.
[0059] Table 1. Effects of microbial inoculant addition on plants under inorganic fertilizer treatment.
[0060]
[0061] Table 2. Effects of adding microbial agents on plants under organic fertilizer treatment.
[0062]
[0063]
[0064] (2) Soil physicochemical index monitoring test
[0065] On July 22, 2025, three sampling points were randomly selected from each treatment group containing Bacillus polymyxa to collect rhizosphere soil from 0-20 cm (top leaves removed, five-point sampling method was used). The soil was mixed and passed through a 2 mm sieve, and the eight major physicochemical properties of the soil were determined using conventional methods.
[0066] The test results are as follows Figures 5-12 As shown in Tables 3 and 4, compared to the control group, the combined application of inorganic fertilizer and Bacillus polymyxa resulted in a 5.55% increase in soil pH, a 0.087% increase in organic matter, an 18.66% increase in available potassium, a 22.37% increase in available phosphorus, and a 1.3% increase in available nitrogen in *Elymus sibiricum*, while total potassium decreased by 10.52%, total phosphorus decreased by 4.86%, and total nitrogen increased by 9.62%. Conversely, compared to the control group, the combined application of organic fertilizer and Bacillus polymyxa resulted in an 8.55% increase in pH, an 8.46% increase in organic matter, a 42.84% increase in available potassium, a 22.8% increase in available phosphorus, and a 9.65% increase in available nitrogen, while total potassium decreased by 12.86%, total phosphorus decreased by 9.33%, and total nitrogen increased by 12.57%. Organic fertilizer achieved superior results compared to inorganic fertilizer.
[0067] Table 3. Effects of microbial inoculant addition on soil physicochemical properties under inorganic fertilizer treatment.
[0068]
[0069] Table 4. Effects of adding microbial agents on soil physicochemical properties under organic fertilizer treatment.
[0070]
[0071] In summary, the present invention (1) significantly increases soil pH by 5.55% compared to applying chemical fertilizer alone and by 8.55% compared to applying organic fertilizer alone; (2) increases soil organic matter content by 8.46% compared to applying organic fertilizer alone; (3) increases soil available potassium content by 42.84% compared to applying organic fertilizer alone; (4) increases soil available phosphorus content by ≥22.37%; and (5) increases soil total nitrogen content by 12.57% compared to applying organic fertilizer alone. By constructing a soil microenvironment buffer layer for Bacillus polymyxa YF as basal fertilizer, on the one hand, the rapid loss of bacterial agent activity under the cold and arid conditions of Qinghai is reduced, and its colonization cycle and functional duration in the soil are extended; on the other hand, the organic / inorganic nutrients in the basal fertilizer can complement the active substances secreted by the bacterial agent, promote the transformation of insoluble nitrogen, phosphorus and potassium in the soil, and improve the nutrient absorption efficiency of Leymus chinensis. Furthermore, this compound combination can exert a highly efficient synergistic effect in water conservation and moisture retention, improving soil physical and chemical properties, and enhancing the stress resistance of forage grasses. It can reduce the risk of soil compaction caused by single chemical fertilizers, and enhance the cold resistance and drought tolerance of crested wheatgrass through microbial agents. It is an optimized combination that balances high-quality and high-yield crested wheatgrass in Qinghai Province with grassland ecological restoration. The compound combination and its application of this invention have no environmental pollution or ecological side effects, significantly promote the accumulation of crested wheatgrass biomass, effectively improve soil infertility, and significantly enhance the feed nutritional value and soil water retention capacity of forage grasses. The Bacillus polymyxa YF-base fertilizer compound combination and its preparation and application methods of this invention can specifically solve the dual problems of crested wheatgrass growth and soil improvement in the cold and barren areas of northern China. It can ensure forage grass yield and quality while maintaining the ecological environment, and has extremely high practical application value and promotion prospects.
Claims
1. Application of Paenibacillus polymyxa YF or its fermentation broth in promoting the growth of Leymus chinensis, wherein the Paenibacillus polymyxa YF was deposited at the China General Microbiological Culture Collection Center on December 21, 2020, with accession number CGMCC No. 21517.
2. The application as described in claim 1, characterized in that, The aforementioned polymyxa Bacillus YF is used in combination with fertilizer.
3. The application as described in claim 1, characterized in that, The fertilizers mentioned include one or more of chemical fertilizers, potash fertilizers, and organic fertilizers.
4. Application of Paenibacillus polymyxa YF or its fermentation broth in increasing the yield of crested wheatgrass. The Paenibacillus polymyxa YF was deposited at the China General Microbiological Culture Collection Center on December 21, 2020, with the accession number CGMCC No. 21517.
5. Application of Paenibacillus polymyxa YF or its fermentation broth in increasing plant length and leaf width, wherein the Paenibacillus polymyxa YF was deposited at the China General Microbiological Culture Collection Center on December 21, 2020, with accession number CGMCC No. 21517.
6. Application of Paenibacillus polymyxa YF or its fermentation broth in increasing the fresh weight content of aboveground and underground parts, wherein the Paenibacillus polymyxa YF was deposited at the China General Microbiological Culture Collection Center on December 21, 2020, with accession number CGMCC No. 21517.
7. Application of Paenibacillus polymyxa YF or its fermentation broth in increasing the dry weight content of the aboveground and underground parts, wherein the Paenibacillus polymyxa YF was deposited at the China General Microbiological Culture Collection Center on December 21, 2020, with accession number CGMCC No. 21517.
8. Application of Paenibacillus polymyxa YF or its fermentation broth in the preparation of crested wheatgrass growth promoter, wherein the Paenibacillus polymyxa YF was deposited at the China General Microbiological Culture Collection Center on December 21, 2020, with accession number CGMCC No. 21517.
9. A method for promoting the restoration of degraded grasslands, characterized in that, The degraded grassland was treated with Paenibacillus polymyxa YF and other fertilizers. Paenibacillus polymyxa YF was deposited at the China General Microbiological Culture Collection Center on December 21, 2020, with accession number CGMCC No. 21517. The fertilizers included organic fertilizers and / or inorganic fertilizers.