Strain, bacterial agent for preventing and treating corn small spot and application thereof

Abstract

The present application belongs to the field of microorganisms, and relates to a strain for preventing and treating corn small spot, a bacterial agent and application thereof. The strain for preventing and treating corn small spot is bacillus velezensis A122, which can resist salt and alkali, and has a good inhibitory effect on dicladiodendron commixtum. The strain for preventing and treating corn small spot has a strong ability to produce cellulase, amylase, protease and indole-3-acetic acid (IAA), and has a good growth promoting effect on corn. The bacterial agent prepared by using the biocontrol bacteria does not pollute the ecological environment and cause pollution during use. The biocontrol bacteria can reduce the use amount of other chemical pesticides, reduce the prevention and control cost of corn diseases, promote the growth of corn, and has good popularization value.

Description

Technical Field

[0001] This invention belongs to the field of microbiology, and in particular relates to strains, inoculants and their applications for the prevention and control of corn leaf blight. Background Technology

[0002] Maize leaf spot is a globally significant disease of maize caused by *Bipolaris maydis*, primarily affecting leaves, leaf sheaths, husks, and ears. It can occur throughout the entire maize growth cycle, but is most severe during the tasseling to grain-filling stage. The spread of maize leaf spot depends on three key factors: large-scale planting of susceptible varieties, sufficient overwintering inoculum in the field, and suitable temperature and humidity. The pathogen overwinters as mycelium or conidia on diseased plant debris, surviving for 1-2 years. The following year, it spreads via air currents and rainwater, germinating and infecting within 4-8 hours. Lesions form within 3-4 days, producing new spores for repeated infections, leading to rapid and widespread outbreaks. Furthermore, excessive planting density, poor ventilation and light penetration, and improper fertilizer and water management further increase the risk of disease occurrence.

[0003] Current control of maize leaf spot relies primarily on integrated management measures, but these have significant limitations. For example, long-term and excessive use of chemical control leads to pathogen resistance, creating a vicious cycle that necessitates continuously increasing pesticide dosages. Furthermore, chemical agents easily cause environmental pollution and harm non-target organisms, contradicting the needs of green agriculture. While crop rotation, removal of diseased plant debris, and appropriate planting density can reduce inoculum sources or improve the field environment, these measures are limited by geographical location and planting patterns, making them insufficient to address large-scale disease outbreaks and resulting in limited control effectiveness.

[0004] Biological control has become an important development direction for the prevention and control of maize leaf spot due to its advantages such as environmental friendliness and low likelihood of developing resistance. However, existing biocontrol strains generally suffer from problems such as narrow antibacterial spectrum, weak environmental adaptability, and unstable control efficacy. Therefore, screening novel biocontrol strains with characteristics such as strong antibacterial activity, broad antibacterial spectrum, good environmental adaptability, and good growth-promoting effect is of great significance for improving the control level of maize leaf spot and ensuring the sustainable development of the maize industry. Summary of the Invention

[0005] The purpose of this invention is to provide a strain of Bacillus velezensis for controlling maize leaf spot, an inoculant, and its application. The strain for controlling maize leaf spot is Bacillus velezensis A122, isolated from the soil of a mango orchard in Damao Village, Yazhou District, Sanya City (109°12′5″E, 18°24′38″N). It is salt-tolerant and can inhibit various pathogens and maize diseases.

[0006] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0007] The strain for controlling maize leaf spot disease is Bacillus velezensis A122, deposited at the Guangdong Provincial Microbial Culture Collection Center, accession number GDMCC No:66572, deposited on June 23, 2025, classified as Bacillus velezensis, and located at the Institute of Microbiology, Guangdong Academy of Sciences, 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou.

[0008] The present invention also provides a microbial agent for controlling corn leaf spot disease, wherein the microbial agent comprises a bacterial suspension of the above-mentioned strain.

[0009] Furthermore, the bacterial concentration in the bacterial suspension is 1~9×10⁻⁶. 7 CFU / mL.

[0010] The present invention also provides a method for preparing the above-mentioned fungal agent for controlling corn leaf blight, wherein the above-mentioned strain for controlling corn leaf blight is inoculated into a fermentation medium for fermentation culture to obtain the fungal agent for controlling corn leaf blight.

[0011] The present invention also provides a method for preparing the above-mentioned inoculant for controlling corn leaf blight, wherein the fermentation temperature is 30~40℃, the rotation speed is 180~220 rpm, and the time is 40~50 h.

[0012] The present invention also provides the application of the above-mentioned strains for preventing and controlling maize leaf spot in promoting maize growth.

[0013] Furthermore, promoting corn growth includes increasing corn plant height, root length, and stem diameter.

[0014] The present invention also provides the application of the above-mentioned strains for controlling maize leaf spot in inhibiting Bipolar Helicobacter spp.

[0015] The present invention also provides the application of the above-mentioned strains for controlling maize leaf spot in the production of extracellular hydrolases.

[0016] The present invention also provides the application of the above-mentioned strains for controlling maize leaf spot in the decomposition of cellulose, starch or protein.

[0017] Beneficial effects

[0018] The strain of *Bacillus berberis* A122 provided by this invention for controlling corn leaf blight is salt-tolerant and exhibits good inhibitory effects against *Bacillus diplostilbene*, effectively controlling corn leaf blight. *Bacillus berberis* A122 also possesses a strong ability to produce cellulase, protease, amylase, and indole-3-acetic acid (IAA), and has a good growth-promoting effect on corn. The biocontrol agent prepared using this biocontrol bacterium will not pollute the ecological environment during use and is harmless. Using the biocontrol bacterium of this invention can reduce the amount of other chemical pesticides used, lower the cost of corn disease control, and promote corn growth, thus having good promotional value. Attached Figure Description

[0019] Figure 1 Phylogenetic tree of Bacillus belyssus A122;

[0020] Figure 2 This refers to the whole-genome biosynthetic gene cluster of Bacillus belyssus A122;

[0021] Figure 3 The image shows the inhibitory effect of Bacillus belyssus A122 on Bipolar Helicobacter zearalensis.

[0022] Figure 4 The images show the results of salt stress tolerance determination for Bacillus belyss A122, and the salt tolerance plate phenotypes of Bacillus belyss A122 and the model strain FZB42, respectively.

[0023] Figure 5 Figure 1 shows the results of the acid and alkali stress resistance test of Bacillus belyssus A122, and the plate phenotype results of acid and alkali resistance of Bacillus belyssus A122 and FZB42. Figure 5 The pH of A is 2. Figure 5 The pH of B is 3. Figure 5 The pH of C is 11. Figure 5 The pH of D is 12;

[0024] Figure 6 HPLC analysis of secondary metabolites of Bacillus belyssus A122;

[0025] Figure 7 Figure showing the antibacterial effect of crude extract of secondary metabolites of Bacillus belyssus A122;

[0026] Figure 8 The results of detecting protease, cellulase, amylase, and ACC deaminase produced by Bacillus belyssus A122 are shown in the figure. Figure 8 A in the middle is a protease producer. Figure 8 B is cellulase, Figure 8 C in the middle represents amylase, Figure 8 D in the middle represents ACC deaminase;

[0027] Figure 9 Determination of the ability of Bacillus belyssus A122 to produce indole-3-acetic acid (IAA);

[0028] Figure 10 The diagram shows the disease control effect of Bacillus vesicles A122 on small spot disease on maize plants;

[0029] Figure 11 This image shows the effect of Bacillus belychnophorus A122 on the growth promotion of maize plants; among them, Figure 11 In the middle, A represents plant height. Figure 11 In the middle, B represents the root length. Figure 11 C represents the fresh weight of the above-ground portion. Figure 11 D represents the dry weight of the above-ground portion. Figure 11 E represents the fresh weight of the underground portion. Figure 11 F represents the dry weight of the underground portion. Figure 11 G represents the stem diameter.

[0030] Biological Preservation Instructions

[0031] The Bacillus velezensis A122 provided by this invention is deposited at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC No:66572, deposit date June 23, 2025, classified as Bacillus velezensis, and deposited at the Institute of Microbiology, Guangdong Academy of Sciences, 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou. Detailed Implementation

[0032] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0033] Example 1

[0034] Taxonomic analysis of Bacillus belyssae A122 based on its core genome

[0035] Bacillus belyssus strain A122 was inoculated into liquid LB medium and incubated at 37°C for 12 hours at 200 rpm. The cells were collected by centrifugation, and genomic DNA was extracted using a bacterial DNA extraction kit (Hunan Aikerui Biotechnology Co., Ltd.). The 16S rRNA gene was then amplified by PCR using forward primer 27F: 5'-AGAGTTTGATCCTGGCTCAG-3' (SEQ ID No. 3) and reverse primer 1492R: 5'-GGTTACCTTGTTACGACTT-3' (SEQ ID No. 4). The PCR conditions were: 95°C, 5 min; 95°C, 15 s; 56°C, 1 min; 72°C, 1 min; 32 cycles; 72°C, 10 min; and stored at 4°C. The PCR product was sent to Sangon Biotech (Shanghai) Co., Ltd. for sequencing. The 16S rRNA gene sequence of Bacillus belyssus A122 is shown in SEQ ID No. 1.

[0036] The 16S rRNA gene sequence of Bacillus belyssus A122:

[0037]

[0038] PCR amplification and sequencing of the gyrB gene:

[0039] Forward primer gyrB-F:

[0040] 5'-GAAGTCATCATGACCGTTCTGCAYGCNGGNGGNAARTTYGA-3' (SEQ ID No. 5);

[0041] Reverse primer gyrB-R:

[0042] 5'-AGCAGGGTACGGATGTGCGAGCCRTCNACRTCNGCRTCNGTCAT-3' (SEQ ID No. 6); PCR reaction conditions for the gyrB gene are the same as above.

[0043] The partial gene sequence of gyrB is approximately 1200 bp in length, as shown in SEQ ID No. 2 of the sequence listing.

[0044] gyrB gene sequence:

[0045]

[0046] 16S rRNA and gyrB identification results showed that strain A122 belongs to *Bacillus belyescens*. Secondly, based on the genome sequence of A122 and 31 housekeeping genes from all samples, 19 strains most closely related at the species level were selected, and a phylogenetic tree was constructed using MEGA 6.0 software selection. The results showed that strain A122 is most closely related to *Bacillus belyescens* in terms of evolutionary distance. These results confirm that strain A122 belongs to *Bacillus belyescens*.

[0047] Whole-genome sequencing of Bacillus was performed using an Illumina Hiseq 4000 and PacBio SMRT sequencing system. The biosynthetic gene clusters (BGCs) of Bacillus A122 were predicted using the online tool antiSMASH (https: / / dl.secondarymetabolites.org / releases / 4.0.2 / ). Three lipopeptide compounds and three polyketide compounds were predicted in the BGCs of Bacillus A122 chromosome, responsible for the biosynthesis of Fengycin, Surfactin, Iturin, Bacillaene, Difficidin, and Macrolide H, respectively.

[0048] Example 2

[0049] Determination of the antibacterial activity of Bacillus belyssus A122

[0050] The antibacterial ability of Bacillus belysus was preliminarily determined, and the selected pathogenic fungi included Bipolaris maydis.

[0051] Determination of the antibacterial activity of Bacillus belyss against pathogenic fungi: A certain number of mycelial cakes were punched from the outermost edge of the original pathogenic fungal culture plate using a sterile punch with a diameter of approximately 0.6 cm. The mycelial cakes (mycelial side) were then transferred to the center of a new potato dextrose agar (PDA) culture plate. A Bacillus belyss A122 mycelial block was then placed at the same distance from the mycelial cakes. The plate was incubated at 28 ℃, and the antibacterial effect was observed. The results are as follows: Figure 3 As shown, this indicates that Bacillus belye A122 has a certain inhibitory effect on Bipolar Helicobacter zearalensis.

[0052] Example 3

[0053] Determination of stress resistance of Bacillus belyssus A122

[0054] The stress resistance of Bacillus belye is closely related to its disease resistance and growth promotion ability. Therefore, the stress resistance ability of Bacillus belye A122 was determined.

[0055] 1. Determination of salt stress tolerance in Bacillus belyssus A122

[0056] The salt stress resistance of strain A122 and the model strain FZB42 was determined by simulating different salt stresses of 1%, 3%, 5%, 7%, 9%, 11%, 13%, 13.1%, and 13.2%. Single colonies of the tested strains were picked and placed in LB liquid medium and incubated overnight at 37 °C and 200 rpm for 24 h until the bacterial concentration reached OD200. 600 When the concentration of bacterial culture reached 2.0, 5 μL of bacterial culture was transferred to LB solid medium with different salt gradients. The medium was then placed in a 37 ℃ incubator. After 2 days, the colony growth of each strain under different salt gradients was observed. The results are as follows: Figure 4 As shown in the figure. The results indicate that Bacillus belyeis A122 can grow under salt stress conditions with a salt content of 13.1%, and its growth status is better than that of the model strain FZB42, indicating that it has a certain salt stress tolerance.

[0057] 2. Determination of the acid-base stress tolerance of Bacillus belyssus A122

[0058] Different acid-base stresses (pH gradients: 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12) were simulated to determine the resistance of *Bacillus belyssae* A122 and the model strain FZB42 to acid-base stress. Single colonies of the tested strains were picked and placed in LB liquid medium, incubated overnight at 37℃ and 200 rpm for 24 h on a shaker. 5 μL of the bacterial culture was then transferred to LB liquid medium at different pH gradients. The medium was then placed in a 37℃ shaking incubator. After 2 days, 5 μL of the bacterial culture was transferred to LB solid medium to observe the colony growth of each strain under different pH conditions. Each treatment was repeated three times. The results are shown below. Figure 5 As shown in the figure. The results indicate that Bacillus belye A122 is difficult to grow under acidic stress at pH 2 and alkaline stress at pH 12.

[0059] Example 4

[0060] Extraction, identification and antibacterial activity determination of secondary metabolites of Bacillus belyssus A122

[0061] 1. Extraction and identification of secondary metabolites of Bacillus belyssus A122

[0062] Bacillus belyssus A122 was activated on LB solid medium and incubated at 37 ℃ until single colonies grew. A single colony was picked and transferred to 20 mL of LB liquid medium and cultured at 37 ℃ with shaking at 200 rpm for 12 h. The Bacillus belyssus A122 culture was then transferred at a 1% ratio to 200 mL of LB liquid medium and cultured at 37 ℃ with shaking at 200 rpm for 48 h. 6 mL of resin (XAD-16N) was added to each A122 culture, and the culture was continued with shaking for 24 h. The culture was centrifuged at 8000 rpm for 10 min, and the bacterial cells were collected. The bacterial cells were dissolved in 30 mL of methanol, and the suspension was cultured at 37 ℃ with shaking at 200 rpm for 4 h. The suspension was centrifuged at 8000 rpm for 10 min, the supernatant was collected and filtered through filter paper. The supernatant was concentrated to 2 mL using a rotary evaporator, and then... The crude extract was filtered through a μm filter membrane, and the formation of lipopeptide compounds was identified and detected by high-performance liquid chromatography (HPLC). The results are as follows: Figure 6 As shown, this indicates that Bacillus belye A122 can produce Fengycin, Surfactin, and Iturin.

[0063] 2. Determination of the antibacterial effect of secondary metabolites of Bacillus belyssus A122

[0064] *Bipolaris zebufo* was selected as the pathogenic fungus. A certain number of mycelial cakes were taken from the outermost edge of the original pathogenic fungal culture plate using a sterile punch with a diameter of approximately 0.5 cm. The mycelial cakes (with the mycelial side) were then transferred to the center of a new potato dextrose agar (PDA) culture plate and incubated at a suitable temperature until new mycelia grew around the cakes. Subsequently, holes were punched at the same distance from the mycelial cakes using a 0.5 cm diameter sterile punch. 50 µL of secondary metabolites of *Bacillus belyssae* A122 was added to the center of each hole, with methanol solution used as a control. After drying, the plates were returned to the incubator, and the antibacterial effect was observed. The results are as follows: Figure 7 As shown in the figure. The plate confrontation experiment results indicate that the secondary metabolites of Bacillus belye A122 have a good antagonistic effect on maize leaf spot pathogen.

[0065] 3. Determination of the ability of Bacillus belyssus A122 to produce extracellular hydrolases

[0066] Qualitative detection of protease production by *Bacillus belyssae* A122 was performed on skim milk agar plates: The ability of *Bacillus belyssae* to produce protease was determined in agar medium containing skim milk powder (20 g skim milk powder, 20 g agar, pH 7.0, sterilized at 115°C for 10 min). Single colonies of *Bacillus belyssae* were picked and incubated in LB broth at 37 °C, 200 r / min overnight for 12 h; 5 μL (OD) of the culture medium was added dropwise to the medium. 600 A bacterial suspension with a concentration of 1.0 g / cm³ was incubated at 37 °C for 24-36 h. Afterward, the presence of a clear zone around the colonies was observed. Results were as follows: Figure 8 As shown in Figure A.

[0067] The cellulase production capacity of *Bacillus belyssiensis* A122 was determined using sodium carboxymethyl cellulose (CMC-Na) medium: Single colonies of *Bacillus belyssiensis* were picked and cultured overnight at 37 °C and 200 rpm for 12 h. 5 μL of the bacterial suspension (OD) was then aspirated onto a CMC-Na agar plate. 600 =1.0), seal the plate and incubate it in a 37 °C incubator; after 2 days, remove the petri dish, pour Gram's iodine stain solution until it submerges the surface of the plate, let it stand for 4 minutes, then pour off the stain solution and observe whether a clear zone forms around the colonies. If a clear hydrolysis zone appears around the bacterial cells, it indicates that the strain can produce the corresponding enzyme; if no clear hydrolysis zone appears, it indicates that the strain cannot produce this type of enzyme. The results are as follows. Figure 8 As shown in B.

[0068] Detection of amylase activity in Bacillus belyssus A122 on starch agar medium: Colonies of Bacillus belyssus were picked and placed in LB broth, incubated at 37 °C and 200 rpm for 12 h, and then 5 μL of bacterial suspension (OD200) was added. 600 =1.0) was added to starch-containing culture medium and incubated in a 37°C incubator; after 2 days, the petri dishes were removed, Gram's iodine stain was poured in to submerge the surface of the plates, and after standing for 4 minutes, the stain was poured off. The experimental results were observed and recorded as follows. Figure 8 As shown in C.

[0069] Colonies of Bacillus belye were picked and placed in LB broth, incubated overnight at 37 °C and 200 rpm for 12 h, and then 5 μL of bacterial suspension (OD) was added. 600=1.0) was added to DF medium, DF (containing ammonium sulfate) medium, and ADF medium. The growth of the strain in the three different media was observed. When the strain grew well in ADF medium but poorly in DF medium, it indicated that the strain could grow with 1-aminocyclopropane-1-carboxylic acid (ACC) as the sole nitrogen source, which also indicated that the strain had the ability to produce 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase. The results are as follows. Figure 8 As shown in D.

[0070] The results showed that strain A122 could produce large transparent hydrolysis zones on skim milk powder medium, sodium carboxymethyl cellulose (CMC-Na) medium, and amylase detection medium, and could grow on ADF medium, indicating that it could produce cellulase, protease, amylase, and 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase.

[0071] 4. Determination of the ability of Bacillus belyssus A122 to produce indole-3-acetic acid (IAA)

[0072] Preparation of Salkowski colorimetric solution: Weigh 0.81 g of FeCl3 and dissolve it in 10 mL of ddH2O. Ensure complete dissolution and mix thoroughly to obtain a 0.5 mol / L stock solution. Prepare a 35% perchloric acid solution. Then, take 50 mL of the perchloric acid solution and mix it thoroughly with 1 mL of the 0.5 mol / L FeCl3 stock solution.

[0073] First, a stock solution of the bacterial strain was obtained using LB medium at 37 ℃ and 200 rpm. Then, the stock solution was transferred to YMB liquid medium at a ratio of 1% and incubated at 37 ℃ and 200 rpm for 48 h. After incubation, 2 mL of the bacterial culture was centrifuged at 12000 rpm to collect the supernatant. An equal volume of Salkowski colorimetric solution was added to the supernatant, and the mixture was thoroughly mixed. The mixture was then reacted in the dark for 30 min. If the solution turned red, it indicated that the bacterium could produce indole-3-acetic acid (IAA). Using an equal volume of YMB liquid medium as a blank control, standard solutions of indole-3-acetic acid (IAA) at concentrations of 2.5 μg / mL, 5.0 μg / mL, 7.5 μg / mL, 10.0 μg / mL, and 12.5 μg / mL were prepared. The OD values ​​of the standard and sample solutions were measured using a spectrophotometer. 530 The value was used to obtain the regression equation = 0.0428x - 0.0257. The calculated indole-3-acetic acid (IAA) production capacity of *Bacillus belyssus* A122 was 7.70 μg / mL, and that of the model strain FZB42 was 7.59 μg / mL. The results are as follows: Figure 9As shown in the figure. The results indicate that Bacillus belyeis A122 can produce indole-3-acetic acid (IAA), and the ability of strain A122 to produce indole-3-acetic acid (IAA) is higher than that of the model strain FZB42, indicating that it has great potential in promoting plant growth and development and has the potential to be developed into a biocontrol agent.

[0074] Example 5

[0075] Test of the ability of Bacillus vesiculosus A122 to control maize leaf spot disease

[0076] Select whole and plump corn seeds, soak them in a 30% sodium hypochlorite solution for 120 seconds, then soak them in 75% ethanol for 30 seconds, and then rinse them repeatedly with ddH2O 5 times until there is no irritating odor remaining on the surface of the corn seeds. Mix the nutrient soil and vermiculite evenly in a 2:1 ratio, sterilize at 121 ℃ for 20 minutes under high temperature and high pressure, and set aside. Fill the flower pot with substrate soil, use tweezers to pick up the sterilized corn seeds, plant them in the soil, cover them with another layer of soil, place the flower pot in a tray with an appropriate amount of water, and incubate at 28 ℃ in a greenhouse until the corn plants enter the large trumpet stage (4-6 leaf stage). Select corn plants to carry out disease prevention experiments. Activate Bacillus belye A122 and the model bacterium FZB42 on LB solid medium. Pick a single colony with a sterile toothpick and place it in 20 mL of LB liquid medium, and incubate overnight at 37 ℃ and 200 rpm with shaking. The following day, the culture was transferred at 1%, and after 48 hours of incubation, the OD of the bacterial culture was measured. 600 Adjust to 0.5 (10) 7 (CFU / mL) was prepared for use. Wounds were created on corn leaves by rubbing with quartz sand. The wounds were then inoculated with fungal discs of *Bacillus berberis*, the causal agent of corn leaf blight, while simultaneously sprayed with a diluted suspension of *Bacillus belyssus*. The control group was sprayed with water. All treatments were incubated under a film-supported moist environment for 3 days, after which the film was removed. Disease occurrence was recorded 1-2 weeks later.

[0077] Table 1. Control efficacy of Bacillus belye A122 against maize leaf spot.

[0078]

[0079] Experimental results showed that spraying maize leaves with Bacillus vesicles A122 reduced the severity of maize leaf spot disease. The maize plants treated with Bacillus vesicles A122 had the smallest lesion area, measuring 8.19 mm. 2 The lesion area was reduced by 16.2 mm compared to the blank control group. 2 The Bacillus berberis A122 treatment showed good control of maize leaf spot disease, and its control effect was similar to that of the FZB42 treatment group.

[0080] Example 6

[0081] Study on the growth-promoting effect of Bacillus vesiculosus A122 on maize plants

[0082] Disinfection and germination of corn seeds: Select whole and plump seeds, soak them in 30% sodium hypochlorite solution for 120 seconds, then soak them in 75% ethanol for 30 seconds, and then rinse them repeatedly with ddH2O 5 times until there is no irritating odor remaining on the surface of the corn seeds; place sterile filter paper in a 9 cm sterilized glass petri dish, add an appropriate amount of sterile water with a pipette so that the filter paper in the dish is completely wetted with water; use sterile tweezers to pick up corn seeds and place them on the filter paper, making sure that there is a certain distance between the seeds so that they can germinate;

[0083] Planting corn seedlings: Mix nutrient soil and vermiculite in a 2:1 ratio, sterilize at 121 ℃ for 20 minutes under high temperature and pressure, and set aside. Use a hole punch with a diameter of about 2-3 cm to punch a hole in the center of a disposable plastic cup, then fill it with substrate soil. Use tweezers to pick up corn seeds that have grown radicles, plant them in the soil with the roots facing down, cover them with another layer of soil, place the disposable plastic cup in a tray, add an appropriate amount of water to the tray, and place it in a greenhouse at 28 ℃ for about 20 days.

[0084] Root irrigation treatment with Bacillus vesicles A122 fermentation broth: After the corn seedlings have grown to the stage of two leaves and one bud, select plants with uniform growth and dilute the seed culture of Bacillus vesicles A122 and the model strain FZB42 with sterile water to a concentration of 10. 7 CFU / mL, inject 50 mL into each corn root to a final concentration of 10. 7 A bacterial suspension of CFU / mL was used, with sterile water as a blank control. A second treatment was performed after 7 days, and growth promotion data were collected after 15 days of culture. Results are as follows: Figure 11 As shown.

[0085] The results showed that strain A122 had a good promoting effect on maize plant growth. Among the strains, strain A122 showed the best promoting effect on the dry and fresh weight of the aboveground parts of maize, with promotion rates as high as 115.60% and 89.06%, respectively. Strain A122 also showed good promoting effects on the dry and fresh weight of the underground parts of maize, with promotion rates of 59.99% and 59.52%, respectively. The growth promotion rate of this strain on maize was higher than that of the model strain FZB42, indicating that Bacillus belyeis A122 has a good growth-promoting effect and certain development potential.

[0086] As can be seen from the above embodiments, the Bacillus berberis A122 provided by the present invention has the ability to resist salt and alkali and can effectively prevent and control small leaf spot disease. It has a good growth-promoting effect on corn and has great application potential in the field of biological pesticides.

[0087] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. Bacillus belye ( Bacillus velezensis The application of A122 in simultaneously preventing and controlling maize leaf spot and promoting maize growth is characterized by, The described *Bacillus belyssus* A122 is deposited at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC No:66572, deposit date June 23, 2025, and classified as... Bacillus velezensis The depositary address is the Institute of Microbiology, Guangdong Academy of Sciences, 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou. The promotion of maize growth includes increasing maize plant height and stem diameter; the prevention and control of maize leaf spot disease refers to preventing maize leaf spot disease during the large trumpet stage.

2. The application according to claim 1, characterized in that, The application includes spraying or drenching corn plants with a suspension of Bacillus berreatus A122.

Images