Bacillus vesiculosus fertilizer and its application to promote crop yield and / or disease resistance

Abstract

This invention discloses a Bacillus velezensis microbial fertilizer and its application for promoting crop yield and / or disease resistance, relating to the field of agricultural microbiology technology. The Bacillus velezensis microbial fertilizer comprises: Bacillus velezensis strain JBH BBLS3 microbial powder and a carrier, wherein the effective viable count of Bacillus velezensis strain JBH BBLS3 in the microbial fertilizer is ≥1.0 × 10⁻⁶. 11 CFU / g; wherein, the strain JBH BBLS3 is deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 28883. This invention's microbial fertilizer integrates growth promotion, disease resistance, and yield increase, demonstrating significant field application effects. The *Bacillus belyceae* strain JBH BBLS3 can be used to promote crop growth, control crop diseases, and increase crop yield. Furthermore, this invention effectively inhibits *Verticillium wilt*, increasing crop yield, while also effectively avoiding environmental pollution compared to chemical agents, demonstrating outstanding practical application value.

Description

Technical Field

[0001] This invention relates to the field of agricultural microbial technology, and in particular to a Bacillus berberis fertilizer and its application that promotes crop yield and / or disease resistance. Background Technology

[0002] With the increasing demand for green and sustainable development in modern agriculture, microbial fertilizers have received widespread attention due to their advantages such as improving soil microecology, promoting crop growth, enhancing crop resistance, and being environmentally friendly. Bacillus bacteria, due to their ability to produce highly resistant spores, are easy to process and store, making them an important microbial resource for microbial fertilizers.

[0003] Bacillus velezensis is a beneficial bacterium that has attracted much attention in recent years. Studies have shown that it has multiple functional potentials, including secreting antibacterial substances (such as lipopeptide antibiotics), promoting the production of plant growth hormones, and inducing systemic resistance. Known strains, such as Bacillus velezensis strain JBH BBLS3 (CGMCC No. 28883), have been reported for deodorization (see patent CN117701439A). However, to date, existing technologies have not revealed that this strain can increase crop yield, nor have there been any reports of its disease resistance effects on crops. Furthermore, existing production methods generally suffer from low viable bacteria concentration, insufficient field stability, and limited efficacy, restricting its large-scale application. Therefore, there is an urgent need for a low-cost, highly effective, disease-resistant, significantly growth-promoting and yield-increasing, high-density, and highly stable Bacillus velezensis inoculant fertilizer, and to clarify its comprehensive application effects on various important economic crops to solve the above problems. Summary of the Invention

[0004] To address the technical problems existing in the prior art, embodiments of the present invention provide a Bacillus baileyi bacterial fertilizer and its application that promotes crop yield increase and / or disease resistance. The technical solution is as follows:

[0005] A Bacillus velezensis microbial fertilizer that promotes crop yield and / or disease resistance, comprising: Bacillus velezensis strain JBH BBLS3 microbial powder and a carrier, wherein the effective viable count of Bacillus velezensis strain JBH BBLS3 in the microbial fertilizer is ≥1.0 × 10⁻⁶. 11 CFU / g;

[0006] The strain JBH BBLS3 is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 28883.

[0007] Optionally, the microbial fertilizer is in powder form;

[0008] And / or, the carrier includes: diatomaceous earth, starch, fermented soybean meal, sodium humate;

[0009] And / or, the starch is selected from corn starch;

[0010] And / or, the weight ratio of the Bacillus belyssus powder to the carrier is 1:5 to 1:20;

[0011] And / or, the weight ratio of the Bacillus belyssus powder to the carrier is 1:8 to 1:12.

[0012] The method for preparing the Bacillus vesiculosus fertilizer includes:

[0013] (1) The Bacillus velezensis strain JBH BBLS3 was fermented to obtain a fermentation broth;

[0014] (2) The fermentation broth is concentrated to obtain a concentrated solution;

[0015] (3) Add a preservative to the concentrate and dry it to obtain bacterial powder;

[0016] (4) The bacterial powder is mixed with the carrier to obtain the Bacillus vesiculosus fertilizer.

[0017] Optionally, in step (2), the concentration is achieved using membrane concentration;

[0018] And / or, the membrane concentration uses a hollow fiber ultrafiltration membrane system;

[0019] And / or, in the hollow fiber ultrafiltration membrane system, the molecular weight cutoff of the ultrafiltration membrane is 10-50 kDa;

[0020] And / or, in the hollow fiber ultrafiltration membrane system, the operating pressure is 0.1-0.3 MPa and the temperature is 25-35℃;

[0021] And / or, in step (2), the fermentation broth is concentrated to 1 / 10 of its original volume.

[0022] Optionally, in step (3), the drying is spray drying;

[0023] And / or, in step (3), the protective agent comprises, by weight %: 10% skim milk powder, 6% trehalose, and 13% maltodextrin;

[0024] And / or, in step (4), the carrier includes: diatomaceous earth, starch, fermented soybean meal, and sodium humate;

[0025] And / or, the starch is selected from corn starch;

[0026] And / or, the weight ratio of the bacterial powder to the carrier is 1:5 to 1:20;

[0027] And / or, the weight ratio of the bacterial powder to the carrier is 1:8 to 1:12.

[0028] The application of the aforementioned Bacillus vesiculosus fertilizer in methods for promoting crop growth, preventing crop diseases, and / or increasing crop yield.

[0029] Optionally, the crop is at least one of cucumber, wheat, and cotton.

[0030] Optionally, the disease includes cotton verticillium wilt.

[0031] Optionally, the method includes applying the Bacillus berberis fertilizer to the crop via basal application.

[0032] Optionally, the application rate of the Bacillus vesiculosus fertilizer is 0.5-2.0 kg per mu or 1.0 kg per mu.

[0033] The beneficial effects of the technical solutions provided in the embodiments of the present invention include at least the following:

[0034] This invention discloses a Bacillus velezensis microbial fertilizer for disease resistance and crop yield promotion, as well as its preparation and application. The active ingredient of this microbial fertilizer is Bacillus velezensis strain JBH BBLS3, which is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 28883. The Bacillus velezensis strain exhibits rapid growth and is easy to cultivate, which is beneficial for industrial production and widespread application.

[0035] 1. This invention employs a specific membrane concentration technology, which can efficiently and gently concentrate and enrich Bacillus belye cells and beneficial metabolites in the fermentation broth, significantly increasing the effective viable cell concentration of the final product (≥1.0×10⁻⁶). 11 The CFU / g ratio reduced drying energy consumption and carrier usage, lowered production costs, and ensured the activity of the bacteria.

[0036] 2. The bacterial fertilizer product prepared by this invention contains strain JBH BBLS3, which has high activity and strong stress resistance.

[0037] 3. The microbial fertilizer of this invention integrates growth promotion, disease resistance, and yield increase, with significant field application effects. This invention has discovered that the *Bacillus vesiculosus* strain JBH BBLS3 can be used to promote crop growth, control crop diseases, and increase crop yield. Furthermore, this invention has also found that the microbial fertilizer prepared from the *Bacillus vesiculosus* strain JBH BBLS3 has a significant control effect on *Verticillium wilt*, effectively reducing the incidence of cotton *Verticillium wilt* by 57.14%. This invention effectively inhibits *Verticillium wilt*, increases crop yield, and, compared with chemical agents, effectively avoids environmental pollution and other problems, possessing outstanding practical application value.

[0038] Biological Preservation Information:

[0039] The Bacillus velezensis strain JBH BBLS3 of this invention is deposited at the China General Microbiological Culture Collection Center (CGMCC) on November 6, 2023, at the Institute of Microbiology, Chinese Academy of Sciences, No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 28883. Attached Figure Description

[0040] To more clearly illustrate the technical solutions in the embodiments of the present invention, the accompanying drawings used in the description of the embodiments will be briefly introduced below. Obviously, the accompanying drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0041] Figure 1 This is a plate colony morphology diagram of Bacillus berberis JBH BBLS3 provided in Embodiment 1 of the present invention;

[0042] Figure 2 This is an oil immersion image of Bacillus berberis JBH BBLS3 provided in Embodiment 1 of the present invention. Detailed Implementation

[0043] The technical solution of the present invention will now be described with reference to the accompanying drawings.

[0044] In embodiments of the present invention, words such as "exemplarily," "for example," etc., are used to indicate that something is an example, illustration, or description. Any embodiment or design described as "exemplary" in the present invention should not be construed as being more preferred or advantageous than other embodiments or designs. Specifically, the use of the word "exemplary" is intended to present the concept in a concrete manner. Furthermore, in embodiments of the present invention, the meaning expressed by "and / or" can be both, or either one.

[0045] This invention specifically relates to a microbial fertilizer containing Bacillus velezensis, its preparation method, and its application in promoting crop growth, resisting pathogens, and increasing yield.

[0046] The purpose of this invention is to overcome the shortcomings of the prior art and provide a Bacillus vesiculosus fertilizer with high viable count, good stability, and both disease resistance and growth promotion functions, as well as its preparation method and application.

[0047] To achieve the above objectives, the present invention adopts the following technical solution:

[0048] In a first aspect, this invention provides a Bacillus velezensis microbial fertilizer that promotes disease resistance and crop yield increase. Its active ingredient is Bacillus velezensis strain JBH BBLS3, which has been deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 28883, on November 6, 2023. The microbial fertilizer contains ≥1.0 × 10⁻⁶ viable bacteria. 11 CFU / g.

[0049] Secondly, the present invention provides a method for preparing the Bacillus vesiculosus fertilizer.

[0050] In a preferred embodiment, the preparation method includes the following steps:

[0051] (1) Activation and fermentation of the strain: The preserved Bacillus berberis JBH BBLS3 was inoculated into liquid seed culture medium and cultured overnight at 35-42℃ and 150-200 rpm to obtain seed liquid; the seed liquid was inoculated into fermentation medium at a volume ratio of 10-15% and fermented at 35-42℃ and 150-200 rpm under aeration for 20-40 h until the spore formation rate reached more than 90% to obtain fermentation broth. The seed culture medium and fermentation medium were both ordinary meat broth culture medium, with the following composition: peptone 8-15 g / L, beef extract powder 4-6 g / L, sodium chloride 4-6 g / L, pH 7.0±0.2.

[0052] (2) Membrane Concentration: After pretreatment to remove large particulate impurities, the fermentation broth obtained in step (1) is concentrated using a hollow fiber ultrafiltration membrane system. The specific membrane concentration process parameters are as follows: using an ultrafiltration membrane with a molecular weight cutoff of 10-50 kDa, the process is carried out under the conditions of transmembrane pressure of 0.1-0.3 MPa, temperature of 25-35℃, and circulation flow rate, concentrating the fermentation broth to 1 / 10 of its original volume to obtain a high-concentration bacterial concentrate. This step can efficiently retain the target bacteria and some metabolites, while removing some water and small and medium molecular impurities, significantly increasing the number of viable bacteria per unit volume.

[0053] (3) Addition of protectant and formulation: Add freeze-dried protectant to the high concentration of bacterial concentrate obtained in step (2) and mix evenly. The protectant formula is 10% skim milk powder, 6% trehalose and 13% maltodextrin.

[0054] (4) Drying: The mixture from step (3) is spray-dried to obtain Bacillus vesiculosus raw powder with high viable bacterial count;

[0055] (5) Carrier adsorption and preparation: Mix the raw powder obtained in step (4) with the sterilized carrier evenly, add an appropriate amount of dispersant, and adjust the moisture content so that the effective live bacteria count of the final product meets the requirements, thus obtaining the microbial fertilizer product.

[0056] Thirdly, the present invention provides the application of the Bacillus vesiculosus fertilizer in promoting crop growth, preventing crop diseases, and increasing crop yield.

[0057] Preferably, the crops include, but are not limited to, cucumbers, wheat, and cotton.

[0058] Preferably, the application method includes: applying the microbial fertilizer as a base fertilizer. The recommended dosage is: 1.0 kg of powder per acre (effective viable bacteria count ≥ 1.0 × 10⁻⁶). 11 (CFU / g), which can be adjusted according to crop type and application method.

[0059] To make the technical problems, technical solutions and advantages of the present invention clearer, a detailed description will be given below in conjunction with the accompanying drawings and specific embodiments, but the scope of protection of the present invention is not limited thereto.

[0060] Unless otherwise specified, the experimental methods described in the following embodiments are conventional experimental methods well known to those skilled in the art, and are performed according to the techniques or conditions described in the literature in this field or according to the product instructions. Where specific conditions are not specified in the experimental methods, they are generally operated under conventional conditions.

[0061] Unless otherwise specified, all materials and reagents described in the following examples are commercially available.

[0062] The culture medium formulation used in this invention is as follows:

[0063] The composition of broth agar medium is as follows: 1g peptone, 0.3g beef extract, 0.5g sodium chloride, 1.5g agar, 100mL purified water, pH 7.4. The above formula, without the addition of agar, yields broth liquid medium.

[0064] Example 1: Isolation and identification of Bacillus belyssus JBH BBLS3

[0065] One g of long-term stored pig manure sample was inoculated into 100 mL of broth containing 50 U / mL nystatin and cultured for 12 h. This was then transferred to the same medium and cultured for another 12 h, repeated five times to remove mold. The culture was then treated at 85°C for 15 min to kill non-spore-forming bacteria. The treated culture was diluted with sterile water at different ratios, and the diluted solutions were spread onto broth agar plates and incubated overnight at 37°C. Single colonies were picked and preserved on broth agar slants. The slants were purified by streak plating three times, and the resulting pure cultures were preserved on broth agar slants for later use. This process yielded strain JBH BBLS3.

[0066] Morphological identification of the strain was performed, and the results are as follows: Figure 1 and Figure 2 As shown.

[0067] Figure 1 This is a single colony image of Bacillus bellis JBH BBLS3. From... Figure 1 As can be seen, the colonies of this strain are milky white, opaque, round or oval, 2-4 mm in diameter, with an irregular surface, a concave center, and wrinkled appearance. They are viscous and easily picked up. The Gram staining result is blue-purple, indicating Gram-positive bacteria.

[0068] Figure 2 This is a microscopic morphological image of Bacillus belyssus JBH BBLS3. From Figure 2 As can be seen from the microscopic examination, the bacterial cells are rod-shaped, with a size of 0.5×(1.5~3.5)μm, and elliptical spores can be generated at both ends of the bacterial cells.

[0069] Further 16S rRNA gene sequencing and homology alignment between the target sequence and sequences in the GenBank nucleic acid sequence library using the BLAST system in the NCBI database confirmed that the strain was Bacillus velaeticus.

[0070] Therefore, JBH BBLS3 was identified as Bacillus velaeticus.

[0071] The sequencing results and phylogenetic tree of this strain have been reported in patent application CN117701439A, and this invention represents a novel application.

[0072] Example 2: Preparation of microbial fertilizer

[0073] 1. Fermentation: The activated JBH BBLS3 strain seed liquid was inoculated into a 50 L fermenter containing 30 L of fermentation medium at a 10% inoculation rate. The temperature was controlled at 37℃, the aeration rate VVM 1:1.2, the pressure at 0.03 MPa, the stirring speed at 200 rpm, and the fermentation time was 24 hours. The spore formation rate was >95% under microscopic examination.

[0074] 2. Membrane Concentration: After filtering the fermentation broth through a 200-mesh sieve, it was pumped into a hollow fiber ultrafiltration membrane system (molecular weight cutoff 30 kDa) and concentrated by circulation at 0.2 MPa pressure and 30°C until it was concentrated to 1 / 10 of its original volume, yielding a concentrated bacterial solution with a viable count of 3.30 × 10⁻⁶. 10 cfu / mL.

[0075] 3. Add preservatives: Add 10% skim milk powder, 6% trehalose, and 13% maltodextrin to the concentrated bacterial solution and stir well.

[0076] 4. Spray drying: The mixture is dispensed and sprayed at a temperature of 160℃, an outlet temperature of 86℃, an air volume of 45%, and a peristaltic pump volume of 25% to obtain bacterial powder. The viable count is 3.00 × 10⁻⁶. 11 CFU / g.

[0077] 5. Carrier Adsorption: The above-mentioned bacterial powder and sterilized corn starch (purchased from Henan Feitian Starch Co., Ltd., 25KG / bag) are thoroughly mixed in a mixer at a ratio of bacterial powder to corn starch of 1:10. The moisture content is adjusted to 10%, and the mixture is then packaged to obtain the powdered bacterial fertilizer. The viable count of the product is 1.30 × 10⁻⁶. 11 CFU / g.

[0078] Example 3: Preparation of Microbial Fertilizer

[0079] 1. Fermentation: The activated JBH BBLS3 strain seed liquid was inoculated into a 50 L fermenter containing 30 L of fermentation medium at a 10% inoculation rate. The temperature was controlled at 37℃, the aeration rate VVM 1:1.2, the pressure at 0.03 MPa, the stirring speed at 200 rpm, and the fermentation time was 24 hours. The spore formation rate was >95% under microscopic examination.

[0080] 2. Membrane Concentration: After filtering the fermentation broth through a 200-mesh sieve, it was pumped into a hollow fiber ultrafiltration membrane system (molecular weight cutoff 50 kDa) and concentrated by circulation at 0.3 MPa pressure and 35°C until it was concentrated to 1 / 10 of its original volume, yielding a concentrated bacterial solution with a viable count of 4.10 × 10⁻⁶. 10 cfu / mL.

[0081] 3. Add preservatives: Add 8% skim milk powder, 8% trehalose, and 15% maltodextrin to the concentrated bacterial solution and stir well.

[0082] 4. Spray drying: The mixture is dispensed and sprayed at a temperature of 160℃, an outlet temperature of 86℃, an air volume of 45%, and a peristaltic pump volume of 25% to obtain bacterial powder. The viable count is 3.50 × 10⁻⁶. 11 CFU / g.

[0083] 5. Carrier Adsorption: Thoroughly mix the above-mentioned bacterial powder with sterilized diatomaceous earth in a mixer at a ratio of 1:15 (bacterial powder:diatomaceous earth). Adjust the moisture content to 8%, and then package to obtain the powdered bacterial fertilizer. The viable count of the product is 1.20 × 10⁻⁶. 11 CFU / g.

[0084] Example 4: Application Experiment of Microbial Fertilizer on Cucumber

[0085] Experimental Location: Zhou Cunfeng's vegetable field in Taishan Village, Taishan Town, Huojia County, Henan Province (Location: 113.702366°E, 35.182092°N). The soil type is alluvial soil, with a texture of mixed soil and soil layers. The terrain is flat, the soil fertility is uniform, and irrigation and drainage are convenient. The topsoil nutrients in this field are: organic matter 24.1 g / kg, total nitrogen 1.25 g / kg, available phosphorus (P2O5) 38.7 mg / kg, available potassium (K2O) 169.2 mg / kg, and pH value 7.9. The previous crop was tomato, with a yield of 4500 kg / mu. During the tomato fruiting period, special water-soluble fertilizer was applied 4 times, 20 kg / mu each time. The test crop was cucumber, variety Bona No. 3, with a planting density of 2650 plants / mu. The tested microbial fertilizer was the microbial fertilizer prepared in Example 2, with an effective viable count ≥1.0×10⁻⁶. 11 CFU / g, strain is Bacillus belysinus, dosage form is powder.

[0086] Four treatments were set up: Treatment 1: conventional fertilization + 1 kg of microbial fertilizer preparation per acre before cucumber transplanting; Treatment 2: conventional fertilization + application of the same amount of the tested microbial fertilizer preparation substrate as Treatment 1 during the same period; Treatment 3: conventional fertilization + application of the same amount of sandy soil as Treatment 1 during the same period; Treatment 4: conventional fertilization. The plot area was 32 m². 2 The blocks were randomly arranged, and each treatment was repeated 3 times.

[0087] Experimental results:

[0088] The effects of basal application of microbial fertilizer on cucumber length, diameter and weight are shown in Table 1.

[0089] Table 1. Effects of basal application of microbial fertilizer on cucumber length, diameter, and weight.

[0090]

[0091] Note: All data in the table are the average of three repeated trials.

[0092] Table 1 shows that compared with treatments 2, 3, and 4, treatment 1 resulted in cucumber length increases of 0.4 cm, 0.7 cm, and 0.9 cm, respectively; transverse diameter increases of 1.23 mm, 1.29 mm, and 1.44 mm, respectively; and single cucumber weight increases of 5.6 g, 7.5 g, and 7.9 g, respectively. These results indicate that, in addition to conventional fertilization, basal application of microbial fertilizer to cucumbers can increase cucumber length, transverse diameter, and weight, demonstrating that basal application of microbial fertilizer has a positive impact on the biological traits of cucumbers.

[0093] The effects of basal application of microbial fertilizer on cucumber yield are shown in Table 2.

[0094] Table 2. Effects of basal application of microbial fertilizer on cucumber yield

[0095]

[0096] As shown in Table 2, compared with treatment 2, treatment 1 yielded an average increase of 353.5 kg per mu, with an increase rate of 6.5%; compared with treatment 3, treatment 2 yielded an average increase of 123.6 kg per mu, with an increase rate of 2.3%; and compared with treatment 4, treatment 3 yielded an increase of 68.10 kg per mu, with an increase rate of 1.3%.

[0097] The results of the variance analysis of the yield for each treatment are shown in Table 3.

[0098] Table 3 Analysis of Variance Table

[0099]

[0100] As shown in Table 3, the yield differences between treatments reached a highly significant level.

[0101] The results of multiple comparisons using the PLSD method are shown in Table 4.

[0102] Table 4 Multiple Comparison Table

[0103]

[0104] Note: PLSD 0.05 =11.10, PLSD 0.01 =16.80.

[0105] As shown in Table 4, the yield differences between treatment 1 and treatments 2, 3, and 4 were extremely significant, while the yield differences between treatments 2, 3, and 4 were not significant.

[0106] The above results indicate that the application of basal microbial fertilizer formulations has a significant effect on increasing cucumber yield.

[0107] Example 5: Application test of microbial fertilizer on wheat

[0108] Experimental Location: Land leased to Gao Jiansong, Yanzhang Village, Batai Town, Wugang City, Henan Province (Location: N33°24'18'', E113°29'55''). The tested soil type was sandy black soil, with moderate fertility, heavy loam texture, uniform soil fertility, and convenient irrigation and drainage. The topsoil nutrients were: organic matter 12.7 g / kg, total nitrogen 0.79 g / kg, available phosphorus (P2O5) 18.2 mg / kg, available potassium (K2O) 92 mg / kg, and pH 6.1. Previous crop: corn, yield 520 kg / mu. 50 kg of special fertilizer (28-6-6) was applied as basal fertilizer per mu, and 10 kg of urea was applied as top dressing at the small trumpet stage. The tested crop was wheat, variety Zhengmai 605. The tested microbial fertilizer was the microbial fertilizer prepared in Example 2, with an effective viable count ≥1.0 × 10⁻⁶. 11 CFU / g, strain is Bacillus belysinus, dosage form is powder.

[0109] Four treatments were set up: Treatment 1: Conventional fertilization + topdressing with 1 kg of the tested microbial fertilizer preparation per mu before wheat greening; Treatment 2: Conventional fertilization + application of the same amount of the tested microbial fertilizer preparation substrate as Treatment 1 at the same time; Treatment 3: Conventional fertilization + application of the same amount of dry fine soil as Treatment 1 at the same time; Treatment 4: Conventional fertilization. The plot area was 40 m². 2 The blocks were randomly arranged, and each treatment was repeated 3 times.

[0110] The experiment was conducted based on local conventional fertilization practices. Conventional fertilization included: 200 kg of well-rotted manure + 50 kg of 45% compound fertilizer (24-15-5) per mu (667 square meters) as basal application before wheat sowing, and 10 kg of urea per mu as topdressing during the greening stage. Wheat in the experimental fields was sown on October 30, 2024, at a rate of 15 kg / mu. As per the experimental protocol, the tested microbial fertilizer preparation, substrate, and dry fine soil were applied to each plot on February 16, 2025 (before greening). Wheat growth was observed on April 4, 2025 (heading stage). On May 17, 10 plants were randomly selected from each treatment for biological trait surveys. Harvesting took place on May 23. At harvest, each plot was individually weighed and the actual yield was recorded. Except for the topdressing with the tested microbial fertilizer preparation, substrate, or dry fine soil before greening, as required by the protocol, other management practices were the same as for general wheat production.

[0111] Experimental results:

[0112] The effects of topdressing microbial fertilizer preparations on the biological traits of wheat are shown in Table 5.

[0113] Table 5 Effects of Topdressing with Microbial Fertilizer on Biological Traits of Wheat

[0114]

[0115] Note: All data in the table are the average of three repeated trials.

[0116] As shown in Table 5, compared with treatments 2, 3, and 4, treatment 1 resulted in increases in plant height of 0.6 cm, 0.8 cm, and 0.8 cm; spike length of 0.2 cm, 0.3 cm, and 0.3 cm; number of spikes per mu (667 square meters) of wheat of 0.9, 1.2, and 1.2, respectively; number of grains per spike of 0.9, 1.0, and 1.2, respectively; and thousand-grain weight of 0.6 g, 0.6 g, and 0.7 g, respectively. These results indicate that, in addition to conventional fertilization, topdressing with microbial fertilizer preparations can increase wheat plant height, spike length, number of spikes per mu, number of grains per spike, and thousand-grain weight.

[0117] The effects of topdressing microbial fertilizer formulations on wheat yield are shown in Table 6.

[0118] Table 6. Effects of Topdressing with Microbial Fertilizer on Wheat Yield

[0119]

[0120] As shown in Table 6, compared with treatment 2, treatment 1 yielded an average increase of 32.8 kg per mu, with an increase rate of 6.8%; compared with treatment 3, the average increase was 38.3 kg per mu, with an increase rate of 8.1%; and compared with treatment 4, the average increase was 40.6 kg per mu, with an increase rate of 8.6%.

[0121] Analysis of variance was performed on the experimental results, and the results are shown in Table 7.

[0122] Table 7 Analysis of Variance Table

[0123]

[0124] As shown in Table 7, the yield differences between treatments reached a significant level.

[0125] Multiple comparisons were performed using the PLSD method, and the results are shown in Table 8.

[0126] Table 8 Multiple Comparison Table

[0127]

[0128] Note: PLSD 0.05 =1.33, PLSD 0.01 =2.01.

[0129] As shown in Table 8, the differences between treatment 1 and treatments 2, 3, and 4 were statistically significant, while the differences between treatments 2, 3, and 4 were not statistically significant.

[0130] The above results demonstrate that the application of basal microbial fertilizer formulations has a very positive effect on increasing wheat yield.

[0131] Example 6: Disease Resistance Effect and Application Test of Microbial Fertilizer on Cotton

[0132] Experimental site: Xiaozhai Village, Xujiagou Town, Yindu District, Henan Province (114.081867 E, 36.147831 N). A plot of land with relatively uniform fertility, flat terrain, and a history of high incidence of Verticillium wilt was selected as the experimental field. The experiment included four treatments, arranged in a randomized block design, with three replicates. The plot area was 40 m². 2 The planting density was 5000 plants per mu (approximately 0.16 acres), and field management followed high-quality cotton cultivation techniques. The test crop was cotton, specifically the Yu Mian 7 variety. The tested microbial fertilizer was the one prepared in Example 2, with an effective viable bacteria count ≥1.0 × 10⁻⁶. 11 CFU / g, strain is Bacillus belysinus, dosage form is powder.

[0133] Treatment 1: Conventional fertilization + applying a mixture of microbial fertilizer and inoculant at a ratio of 1:50 (inoculant:water) to the cotton roots during the early budding stage, at a rate of 0.75 kg / mu (approximately 0.16 acres), once every 10 days, for a total of 2 applications; Treatment 2: Conventional fertilization + applying an equal amount of commercially available microbial fertilizer (Junliqing product purchased from Shandong Zhonggu Nongkang Fertilizer Co., Ltd.) to the cotton roots via drip irrigation. The plot area is 40m². 2 The blocks were randomly arranged, and each treatment was repeated 3 times.

[0134] The number of plants infected with Verticillium wilt in treatment 1 and treatment 2 was investigated and recorded daily. When the incidence of Verticillium wilt was fully developed, the control effect and yield were calculated.

[0135] Experimental results:

[0136] The statistical results are shown in Table 9.

[0137] Table 9. Results of efficacy tests of Bacillus vesicular inoculants against Verticillium wilt in cotton.

[0138]

[0139] Note: All data in the table are the average of three repeated trials.

[0140] As shown in Table 9, treatment with Bacillus vesicularis microbial inoculant significantly reduced the damage caused by Verticillium wilt in cotton. Compared with treatment 2, treatment 1 increased the control rate of Verticillium wilt by 17.67%. The yield per mu (unit of land area) increased by 33.84 kg / mu, indicating that Bacillus vesicularis microbial fertilizer preparation has a good control and yield-promoting effect on Verticillium wilt in cotton.

[0141] The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present invention should be included within the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be determined by the scope of the claims.

Claims

1. A Bacillus vesiculosus fertilizer that promotes crop yield and / or disease resistance, characterized in that, The Bacillus velezensis microbial fertilizer comprises: Bacillus velezensis strain JBH BBLS3 microbial powder and a carrier, wherein the effective viable count of Bacillus velezensis strain JBH BBLS3 in the microbial fertilizer is ≥1.0 × 10⁻⁶. 11 CFU / g; The strain JBH BBLS3 is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 28883.

2. The Bacillus vesiculosus fertilizer according to claim 1, characterized in that, The microbial fertilizer is in powder form; And / or, the carrier includes: diatomaceous earth, starch, fermented soybean meal, sodium humate; And / or, the starch is selected from corn starch; And / or, the weight ratio of the Bacillus belyssus powder to the carrier is 1:5 to 1:20; And / or, the weight ratio of the Bacillus belyssus powder to the carrier is 1:8 to 1:

12.

3. The method for preparing Bacillus vesiculosus fertilizer according to any one of claims 1-2, characterized in that, The method includes: (1) The Bacillus velezensis strain JBH BBLS3 was fermented to obtain a fermentation broth; (2) The fermentation broth is concentrated to obtain a concentrated solution; (3) Add a preservative to the concentrate and dry it to obtain bacterial powder; (4) The bacterial powder is mixed with the carrier to obtain the Bacillus vesiculosus fertilizer.

4. The method according to claim 3, characterized in that, In step (2), the concentration is achieved using membrane concentration; And / or, the membrane concentration uses a hollow fiber ultrafiltration membrane system; And / or, in the hollow fiber ultrafiltration membrane system, the molecular weight cutoff of the ultrafiltration membrane is 10-50 kDa; And / or, in the hollow fiber ultrafiltration membrane system, the operating pressure is 0.1-0.3 MPa and the temperature is 25-35℃; And / or, in step (2), the fermentation broth is concentrated to 1 / 10 of its original volume.

5. The method according to claim 3, characterized in that, In step (3), the drying is spray drying; And / or, in step (3), the protective agent comprises, by weight %: 10% skim milk powder, 6% trehalose, and 13% maltodextrin; And / or, in step (4), the carrier includes: diatomaceous earth, starch, fermented soybean meal, and sodium humate; And / or, the starch is selected from corn starch; And / or, the weight ratio of the bacterial powder to the carrier is 1:5 to 1:20; And / or, the weight ratio of the bacterial powder to the carrier is 1:8 to 1:

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6. The use of Bacillus vesiculosus fertilizer according to any one of claims 1-2 in methods for promoting crop growth, preventing crop diseases and / or increasing crop yield.

7. The application according to claim 6, characterized in that, The crop is at least one of cucumber, wheat, and cotton.

8. The application according to claim 6 or 7, characterized in that, The disease mentioned includes cotton Verticillium wilt.

9. The application according to claim 6, characterized in that, The method includes applying the Bacillus vesiculosus fertilizer to crops via basal application.

10. The application according to claim 9, characterized in that, The application rate of the Bacillus vesiculosus fertilizer is 0.5-2.0 kg per mu or 1.0 kg per mu.

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