Bacillus velezensis RMM03 and application of products thereof in prevention and treatment of rice false smut

Abstract

The application discloses Bacillus velezensis RMM03 and application of a product thereof in prevention and treatment of rice smut. Bacillus velezensis ) RMM03, which is preserved in the Guangdong Microbial Culture Collection Center on November 19, 2024, is located at No. 59 Building, 5th Floor, Guangzhou Xianlie Middle Road 100 Courtyard, and has a preservation number of GDMCC No. 65510. Indoor and field tests show that the bacterium has a significant inhibiting effect on the growth of a rice smut fungus, and can be prevented and treated 10-15 days before a break, 0-5 days before the break and 0-5 days after heading, so that a long prevention and treatment time span is achieved, and high yield and high quality of rice are effectively ensured.

Description

Technical Field

[0001] This invention belongs to the field of microbiology, and in particular relates to the application of Bacillus belye RMM03 and its products in the prevention and control of rice false smut. Background Technology

[0002] Rice false smut is a panicle disease caused by the fungus *Ustilaginoidea virens*. Its typical symptom is the formation of dark green or yellowish-green spherical sclerotia (rice false smut balls) on the grains during the grain-filling stage, containing numerous chlamydospores. These spores not only reduce rice yield and quality but also produce toxins (such as rice false smut toxin), threatening human and animal health.

[0003] Rice false smut fungus primarily invades rice during the flowering stage, manifests symptoms during the grain-filling stage, and becomes a major disease outbreak at maturity. Without control measures, the incidence of rice false smut can reach 80-100%. Currently, rice false smut control mainly relies on chemical methods. However, due to the short control window, the timing of application is crucial; pesticides must be applied 5-7 days before the rice heads emerge. Applying them earlier or later is ineffective. However, in southern regions like Zhejiang, the 5-7 days before heading coincide with the rainy season, and continuous rain can prevent pesticides from reaching their full effectiveness, resulting in poor control. To effectively control rice false smut, farmers repeatedly use chemical pesticides, increasing production costs and raising the risk of pesticide residue contamination in rice and the environment.

[0004] Active microbial biocontrol agents are an ideal alternative to chemical pesticides due to their advantages such as environmental safety, non-polluting nature, and lack of residue. To this end, Chinese invention patent application CN118773059A discloses a strain of Bacillus belyssus and its application. This strain, with accession number CCTCC M20241090, exhibits an inhibition zone of 33.56 mm against rice blast fungus, with an inhibition rate of 61.00%.

[0005] Chinese invention patent application CN114317329A discloses a Bacillus subtilis strain, its screening method and application. The specification of the application describes that the pot control efficacy of Bacillus belye is 57.69±0.13c and the field control efficacy is 56.42±0.60c.

[0006] Chinese invention patent application No. 118497045A discloses a straw-degrading bacterium with high efficiency in inhibiting crop pathogenic fungi and its application. This straw-degrading bacterium is *Bacillus velezensis* BD3, with accession number GDMCC No: 64386. This strain achieved an inhibition rate of 92.47 ± 0.35% against rice blast fungus.

[0007] However, current biocontrol agents all have very high time requirements for application; they must be applied 5-7 days before the rice heads emerge to achieve the desired control effect. Because existing biocontrol agents have a short treatment period of only three days, this presents a significant challenge to the control of rice false smut. Summary of the Invention

[0008] The purpose of this invention is to provide the application of Bacillus belye RMM03 and its products in the prevention and control of rice false smut. The strain of this invention has a significant inhibitory effect on the growth of rice false smut fungus and can control rice false smut in three time periods, with a long control period.

[0009] To address the problems existing in the prior art, the technical solution adopted in this invention is:

[0010] In a first aspect, the present invention provides a strain of Bacillus velezensis RMM03.

[0011] A strain of Bacillus velezensis RMM03 has been deposited at the Guangdong Provincial Center for Microbial Culture Collection (GDMCC) on November 19, 2024. The deposit address is: 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC No. 65510.

[0012] Secondly, the present invention provides the application of Bacillus berberis described in the first aspect above in the preparation of products for the prevention and control of rice false smut.

[0013] Specifically, the product is a biocontrol agent.

[0014] Thirdly, the present invention provides a biocontrol agent comprising *Bacillus belye* as described in the first aspect, wherein the viable count in the biocontrol agent is 1 × 10⁻⁶. 5 cfu / mL -1×10 10 cfu / mL.

[0015] Furthermore, it also includes a protectant and sterile water, wherein the protectant is one or more of carboxymethyl cellulose, methyl cellulose, starch, cyclodextrin, chitosan, and lignin; the concentration of the protectant in the biocontrol agent is 1-10% (m / m).

[0016] Fourthly, the present invention provides a method for preparing the biocontrol agent described in the third aspect above, comprising the following steps:

[0017] ① Inoculate Bacillus belye RMM03 into LB liquid medium and place it in a constant temperature shaker at 26-35℃ for 24-60 hours to obtain fermentation broth;

[0018] ② Add a preservative and sterile water, and adjust the concentration of the cultured fermentation broth to a viable cell count of 1×10⁻⁶. 5 cfu / mL-

[0019] 1×10 10 cfu / mL.

[0020] Fifthly, the present invention provides a method for preventing rice false smut, comprising the following steps:

[0021] Spray the biocontrol agent described in the third aspect above, or the biocontrol agent prepared according to the method described in the fourth aspect above, 15 days before the rice heads break and 5 days after heading. The spraying concentration of the biocontrol agent is 1×10⁻⁶. 5 cfu / mL -1×10 10 If it rains within 4 hours after application, apply the same concentration again after the rain stops.

[0022] Furthermore, the period of 15 days before the rice seedlings break through the heading stage to 5 days after heading refers to 10-15 days before the seedlings break through the heading stage, 0-5 days before the seedlings break through the heading stage, or 0-5 days after heading, preferably 10-15 days before the seedlings break through the heading stage.

[0023] Furthermore, the biocontrol agent described in the third aspect or the biocontrol agent prepared according to the method described in the fourth aspect is sprayed 10-15 days before the rice heads emerge, with a spraying concentration of 1×10⁻⁶. 5 cfu / mL -1×10 7 cfu / mL.

[0024] Furthermore, when spraying the biocontrol agent described in the third aspect above or the biocontrol agent prepared according to the method described in the fourth aspect above 0-5 days before the rice heads break or 0-5 days after heading, a supplementary spray is required 3 days after the first spray, with a spray concentration of 1×10⁻⁶. 8 cfu / mL -1×10 10 cfu / mL.

[0025] Preservation instructions:

[0026] Strain name: Bacillus velezensis RMM03;

[0027] Accession number: GDMCC No. 65510;

[0028] Taxonomic name: Bacillus velezensis;

[0029] Deposit date: November 19, 2024;

[0030] Preservation institution: Guangdong Provincial Center for Microbial Culture Collection;

[0031] Address: 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou.

[0032] The advantages and beneficial effects of this invention are:

[0033] This invention relates to Bacillus belye RMM03, a strain isolated from soil in rice blast disease-affected areas and screened through indoor confrontation experiments. Indoor and field trials have shown that this bacterium has a significant inhibitory effect on the growth of rice blast fungus, and can control the disease 10-15 days before heading, 0-5 days before heading, and 0-5 days after heading, providing a long control period and effectively ensuring high and high yields of rice. Attached Figure Description

[0034] Figure 1 The figure shows the plate antagonistic effect of strain RMM03 of the present invention against Aspergillus oryzae (5 days after inoculation); Figure A shows the growth status of Aspergillus oryzae (Ustilaginoidea virens) on PDA medium; Figure B shows the growth status of Aspergillus oryzae on PDA medium containing RMM03.

[0035] Figure 2 The colony morphology of the strain RMM03 of this invention is shown.

[0036] Figure 3 The figure shows the field control effect of the biocontrol agent with RMM03 strain of the present invention as the main component on rice false smut. In the figure: a is the rice panicle before control, and b is the rice panicle after control. Detailed Implementation

[0037] The technical solutions of the embodiments of this application will be clearly described below with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of this application. All other embodiments obtained by those skilled in the art based on the embodiments of this application are within the scope of protection of this application.

[0038] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, performed according to the techniques or conditions described in the literature in this field or according to the product instructions. Unless otherwise specified, the materials and reagents used in the following examples are commercially available.

[0039] LB liquid medium: 10g tryptone, 5g yeast extract, 10g sodium chloride, distilled water to make up to 1000ml, pH adjusted to 7.0-7.2, sterilized at 121℃.

[0040] LB solid plate medium: Based on LB liquid medium, add agar powder with a concentration of 15 g / L.

[0041] PDA solid plate culture medium: 200g potato, 20g glucose, distilled water to make up to 1000ml, add agar powder at a concentration of 15g / L, sterilize at 121℃.

[0042] Example 1: Isolation, purification, and identification of antagonistic bacteria RMM03

[0043] 1. Bacterial isolation and initial screening

[0044] Soil samples were collected from rice (Yongyou 31) fields in Yuecheng District, Shaoxing City, Zhejiang Province, where rice false smut was present. The samples were sealed in sterile self-sealing bags and brought back to the laboratory, where they were stored at 4°C for the purpose of isolating single colonies.

[0045] The target pathogen used in the experiment was *Ustilaginoidea virens*, which was isolated by our team from rice plants in a diseased area of ​​rice variety Yongyou 31 in Yuecheng District, Shaoxing City, Zhejiang Province. The bacterial culture medium was LB liquid medium, and the fungal culture medium was PDA solid agar plate medium.

[0046] Bacteria were isolated using the soil dilution method and continuously streaked until purified. The antagonistic effect of the bacteria on *Aspergillus oryzae* was determined using growth rate as an indicator. *Aspergillus oryzae* mycelial cakes were prepared using a 5mm diameter punch and inoculated onto PDA plates (control group) and PDA plates containing pre-mixed antagonistic bacteria (antagonistic group), respectively, and incubated in the dark at 28℃ for 8 days. The growth diameter of *Aspergillus oryzae* in the antagonistic group was measured, and strain RMM03, which showed a significant antagonistic effect compared to the control group, was preserved. The results of the plate antagonistic effect of antagonistic bacteria RMM03 on *Aspergillus oryzae* are as follows: Figure 1 As shown, Figure 1 Figure A shows the growth status of Aspergillus oryzae in the control group on PDA medium. Figure 1 Figure B shows the growth status of *Aspergillus oryzae* on PDA medium containing the antagonistic bacterium RMM03. (The text abruptly ends here.) Figure 1 It can be seen that in the antagonistic group of RMM03 antagonistic bacteria, Aspergillus oryzae basically did not grow, and the effect was the most prominent.

[0047] 2. Strain identification

[0048] 2.1 Morphological identification:

[0049] like Figure 2 As shown, the colony morphology of the antagonistic bacterium RMM03 is as follows: the surface is smooth, the colonies are relatively loose, with forks and loose lines, and are pale yellow.

[0050] 2.2 The antagonistic strain RMM03 was identified using molecular biological methods.

[0051] PCR amplification was performed using universal primers (27F and 1492R) for bacterial identification of 16S rDNA.

[0052] PCR reaction system (50μL system): 2μL each of primers, 25μL of 2×Mix, and ddH2O added to make up to 50μL.

[0053] The 16S rDNA primer sequences were 27F: (5'-AGAGTTT GATCCTGGCTCAG-3') (SEQ ID NO.1) and 1492R: (5'-GGTTACCTTGTTACGACTT-3') (SEQ ID NO.2). The reaction program was: 94℃ pre-denaturation for 3 min, 94℃ denaturation for 55 s, 50℃ annealing for 50 s, 72℃ extension for 1 min, 72℃ extension for 10 min, 35 cycles, followed by storage at 16℃. After PCR product detection on a 1.5% agarose gel electrophoresis, the product was sent to Beijing Qingke Biotechnology Co., Ltd. for 16S rDNA sequencing. The 16S rDNA gene sequence of the antagonistic bacterium RMM03 is shown in SEQ ID NO.3. The sequence was then BLAST-aligned using the Eziocloud database, identifying the antagonistic bacterium RMM03 as *Bacillus velezensis*. This strain was deposited at the Guangdong Provincial Microbial Culture Collection Center, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC No. 65510, and deposited on November 19, 2024. SEQ ID NO.3:

[0054] ACGCTGGCGGCGTGCCTAATACATGCAAGTCGAGCGGACAGATGGGAGCTTGCTCCCTGATGTTAGCGGCGGACGG

[0055] GTGAGTAACACGTGGGTAACCTGCCTGTAAGACTGGGATAACTCCGGGAAACCGGGGCTAATACCGGATGGTTGTC

[0056] TGAACCGCATGGTTCAGACATAAAAGGTGGCTTCGGCTACCACTTACAGATGGACCCGCGGCGCATTAGCTAGTTG

[0057] GTGAGGTAACGGCTCACCAAGGCGACGATGCGTAGCCGACCTGAGAGGGTGATCGGCCACACTGGGACTGAGACAC

[0058] GGCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCGCAATGGACGAAAGTCTGACGGAGCAACGCCGCGTG

[0059] AGTGATGAAGGTTTTCGGATCGTAAAGCTCTGTTGTTAGGGAAGAACAAGTGCCGTTCAAATAGGGCGGCACCTTG

[0060] ACGGTACCTAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCG

[0061] GAATTATTGGGCGTAAAGGGCTCGCAGGCGGTTTCTTAAGTCTGATGTGAAAGCCCCCGGCTCAACCGGGGAGGGT

[0062] CATTGGAAACTGGGGAACTTGAGTGCAGAAGAGGAGAGTGGAATTCCACGTGTAGCGGTGAAATGCGTAGAGATGT

[0063] GGAGGAACACCAGTGGCGAAGGCGACTCTCTGGTCTGTAACTGACGCTGAGGAGCGAAAGCGTGGGGAGCGAACAG

[0064] GATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAAGTGTTAGGGGGTTTCCGCCCCTTAGTGCTGCA

[0065] GCTAACGCATTAAGCACTCCGCCTGGGAGTACGGTCGCAAGACTGAAACTCAAAGGAATTGACGGGGGCCCGCAC

[0066] AAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCTCTGACAATCCTA

[0067] GAGATAGGACGTCCCCTTCGGGGGCAGAGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGG

[0068] GTTAAGTCCCGCAACGAGCGCAACCCTTGATCTTAGTTGCCAGCATTCAGTTGGGCACTCTAAGGTGACTGCCGGT

[0069] GACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATG

[0070] GACAGAACAAAGGGCAGCGAAACCGCGAGGTTAAGCCAATCCCACAAATCTGTTCTCAGTTCGGATCGCAGTCTGC

[0071] AACTCGACTGCGTGAAGCTGGAATCGCTAGTAATCGCGGATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGT

[0072] ACACACCGCCCGTCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTTATGGAGCCAGCCGCCGA

[0073] AGGTGGGACAGATGATTGGGGTGAAGT

[0074] Example 2

[0075] This embodiment provides a biocontrol agent, the preparation method of which includes the following steps:

[0076] ① Inoculate Bacillus belye RMM03 into LB liquid medium and culture it in a constant temperature shaker at 28℃ and 200r / min for 36h to obtain fermentation broth.

[0077] ② Add a preservative and sterile water to the fermentation broth, and adjust the concentration of the cultured fermentation broth to a viable cell count of 1×10⁻⁶. 9 The biocontrol agent was obtained by dispersing cfu / ml. The protectant was starch and lignin in a mass ratio of 1:1, and the concentration of the protectant in the biocontrol agent was 6% (m / m).

[0078] Example 3

[0079] This embodiment provides a biocontrol agent. The only difference between this embodiment and Embodiment 2 is that the protective agent used is methylcellulose, and all other aspects are the same as in Embodiment 2.

[0080] Example 4

[0081] This embodiment provides a biocontrol agent. The only difference between this embodiment and Embodiment 2 is that the protective agent used is lignin, and all other aspects are the same as in Embodiment 2.

[0082] Example 5

[0083] This embodiment provides a biocontrol agent. The only difference between this embodiment and Embodiment 2 is that the protective agent used is carboxymethyl cellulose, and all other aspects are the same as in Embodiment 2.

[0084] Example 6

[0085] This embodiment provides a biocontrol agent. The only difference between this embodiment and Embodiment 2 is that the protective agent used is chitosan, and all other aspects are the same as in Embodiment 2.

[0086] Example 7

[0087] This embodiment provides a biocontrol agent. The only difference between this embodiment and Embodiment 2 is that the concentration of the protectant in the biocontrol agent is 1% (m / m), and all other aspects are the same as in Embodiment 2.

[0088] Example 8

[0089] This embodiment provides a biocontrol agent. The only difference between this embodiment and Embodiment 2 is that the concentration of the protectant in the biocontrol agent is 10% (m / m), and all other aspects are the same as in Embodiment 2.

[0090] Comparative Example 1

[0091] This comparative example provides a biocontrol agent, namely Bacillus vesiculosus C17271 (produced by Zhejiang Serike Cell Technology Co., Ltd.).

[0092] Comparative Example 2

[0093] This comparative example provides a biocontrol agent. The only difference between this comparative example and Example 2 is that sodium lignosulfonate is used instead of lignin in step ② of this comparative example. All other aspects are the same as in Example 2.

[0094] Comparative Example 3

[0095] This comparative example provides a biocontrol agent. The only difference between this example and Example 2 is that this example does not contain a protectant. In step ② of this example, only sterile water is added to the fermentation broth. All other steps are the same as in Example 2.

[0096] Example 9

[0097] This embodiment provides a method for preventing and controlling rice false smut, including the following steps:

[0098] 10-15 days before the rice heading stage, spray the biocontrol agent described in Example 2 at a concentration of 1×10⁻⁶. 5 cfu / mL -1×10 7 CFU / mL. If it rains within 4 hours after application, apply again with the same concentration after the rain stops. Application time: evening, when the temperature does not exceed 30℃.

[0099] Example 10

[0100] This embodiment provides a method for preventing and controlling rice false smut, including the following steps:

[0101] Spray the biocontrol agent described in Example 2 0-5 days before the rice heading stage, at a concentration of 1×10⁻⁶. 8 cfu / mL -1×10 10 CFU / mL. If it rains within 4 hours after application, apply a second application at the same concentration after the rain stops. Application time: evening, when the temperature does not exceed 30℃. Apply a second application at the same concentration 3 days after the first application.

[0102] Example 11

[0103] This embodiment provides a method for preventing and controlling rice false smut, including the following steps:

[0104] The biocontrol agent described in Example 2 was sprayed 0-5 days after the rice heading stage, with a spraying concentration of 1×10⁻⁶. 8 cfu / mL -1×10 10 CFU / mL. If it rains within 4 hours after application, apply a second application at the same concentration after the rain stops. Application time: evening, when the temperature does not exceed 30℃. Apply a second application at the same concentration 3 days after the first application.

[0105] Experimental Example 1: Field Trial of the Biocontrol Agent of the Present Invention against Rice False smut

[0106] 1.1 Overview of the experimental site: From August to September 2024, a control experiment was conducted in rice fields where rice false smut was present. The experimental site was located in Shanghuizao Village, Lanting Street, Keqiao District, Shaoxing City, Zhejiang Province. The experimental rice fields had good fertility management and covered a total area of ​​10 mu.

[0107] 1.2 Experimental Grouping: The experimental field was divided into four large areas: a water control area and three experimental areas. A 2-meter rice paddy spacing was maintained between each area. The water control area had an area of ​​10 m². 2 Each experimental area included 10 treatments: the biocontrol agent group prepared in Examples 2-8 and the control agent group prepared in Comparative Examples 1-3. Each treatment was repeated three times, resulting in 30 plots in each experimental area, with each plot measuring 10 m².2 All experimental areas followed the same planting pattern, and fertilizer and water management were carried out normally in each area during the experiment. Cultivation, water and fertilizer management, and weed control were consistent with local rice production. The rice variety used in the experiment was Yongyou 31.

[0108] Test method:

[0109] Water control area: No pesticides were applied throughout the growing season; only water was sprayed during the experiment.

[0110] Experimental Area 1: Control measures were applied 10-15 days before rice heading. The specific application time for this experiment was the evening of August 20, 2024, when the temperature did not exceed 30℃. The spraying concentration was 1×10⁻⁶. 7 cfu / mL.

[0111] Experimental Area 2: Control measures were applied 0-5 days before rice heading, with two applications. Specifically, the applications were made on September 2nd and 5th, 2024, in the evening when the temperature did not exceed 30℃. The spraying concentration was 1×10⁻⁶. 9 cfu / mL.

[0112] Experimental Area 3: Control measures were applied 0-5 days after rice heading, with two applications. Specifically, the applications were made on September 8th and 11th, 2024, in the evening when the temperature did not exceed 30℃. The spray concentration was 1×10⁻⁶. 9 cfu / mL.

[0113] 3. Survey on rice false smut control: A survey was conducted on September 20th in each plot. A random 5-point sampling method was used in each plot, with 10 rice panicles sampled at each point. The number of diseased grains was recorded, and the control rate was calculated. The statistical results are shown in Tables 1-4.

[0114] Particle control efficacy (%) = (Number of diseased particles in the control area - Number of diseased particles in the treatment area) / (Number of diseased particles in the control area) × 100%.

[0115] Figure 3 To assess the control effect of the biological agent prepared in Example 2 in experimental area 3. Figure 3 As shown, before treatment, the rice panicles had already suffered from relatively severe rice false smut, and the rice false smut balls had already developed into their full shape. Figure 3 a) Seven days after the second application of the biological agent prepared according to this invention, the further development of rice blast fungus was successfully inhibited, and the rice blast balls had shrunk and fallen off. Figure 3 b).

[0116] Table 1. Control efficacy (%) of one seed in the experimental area

[0117]

[0118] Table 2. Control efficacy (%) of 2 seeds in the experimental area

[0119]

[0120]

[0121] Table 3. Control efficacy (%) of 3 seeds in the experimental area

[0122]

[0123] Table 4. Grain control efficacy (%) in the water control group

[0124]

[0125] As shown in Tables 1-4, the biological agent with Bacillus vesiculosus RMM03 as the main active ingredient, when sprayed 15 days before rice heading to 5 days after heading, has a good control effect on rice false smut. The comparison between Examples 2-8 and Comparative Example 3 shows that the addition of a protectant to the antagonistic bacterium RMM03 significantly improves the control effect.

[0126] As shown in Tables 1-4, spraying the biocontrol agent described in Comparative Example 1 10-15 days before rice heading has a certain preventive effect against rice false smut, but the preventive effect drops significantly when sprayed 0-5 days before heading and 0-5 days after heading.

[0127] As shown in Tables 1-4, Comparative Example 2 showed a significant decrease in the control effect against rice false smut due to the use of sodium lignosulfonate instead of lignin.

[0128] As shown in Tables 1-4, in Comparative Example 3, due to the lack of a protectant, Bacillus belye RMM03 died in large numbers because it could not adapt to environmental conditions and thus could not exert a control effect.

[0129] In summary, the biological agent of this invention not only has a very good control effect on rice false smut, but also extends the control time of rice false smut from the traditional 5-7 days before rice heading to 15 days before rice heading to 5 days after heading, which greatly reduces the difficulty and intensity of rice false smut control and improves the control effect of rice false smut.

[0130] The embodiments of this application have been described above with reference to the accompanying drawings. However, this application is not limited to the specific embodiments described above. The specific embodiments described above are merely illustrative and not restrictive. Those skilled in the art can make many other forms under the guidance of this application without departing from the spirit and scope of the claims, and all of these forms are within the protection scope of this application.

Claims

1. A strain of Bacillus belye ( Bacillus velezensis RMM03, characterized in that: It is deposited at the Guangdong Provincial Center for Microbial Culture Collection, on November 19, 2024, at the address of Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC No: 65510.

2. The use of Bacillus belye as described in claim 1 in the preparation of products for the prevention and control of rice false smut.

3. The product according to claim 2, characterized in that: The product is a biocontrol agent.

4. A biocontrol agent, characterized in that: The biocontrol agent contains *Bacillus belye* as described in claim 1, and the viable count in the biocontrol agent is 1 × 10⁻⁶. 5 cfu / mL -1×10 10 cfu / mL.

5. The biocontrol agent according to claim 4, characterized in that: It also includes a protectant and sterile water, wherein the protectant is one or more of carboxymethyl cellulose, methyl cellulose, starch, cyclodextrin, chitosan, and lignin; the concentration of the protectant in the biocontrol agent is 1-10% (m / m).

6. The method for preparing the biocontrol agent according to any one of claims 4-5, characterized in that: Includes the following steps: ① Inoculate Bacillus belye RMM03 into LB liquid medium and place it in a constant temperature shaker at 26-35 ℃ for 24-60 h to obtain fermentation broth; ② Add a preservative and sterile water, and adjust the concentration of the cultured fermentation broth to a viable cell count of 1×10⁻⁶. 5 cfu / mL -1×10 10 cfu / mL.

7. A method for preventing and controlling rice false smut, characterized in that: Includes the following steps: Spray the biocontrol agent according to any one of claims 4-5 or the biocontrol agent prepared according to the method of claim 6 15 days before heading and 5 days after heading, wherein the spraying concentration of the biocontrol agent is 1×10 5 cfu / mL -1×10 10 cfu / mL.

8. The method for preventing and controlling rice false smut according to claim 7, characterized in that: The period from 15 days before the rice seedlings to 5 days after heading refers to 10-15 days before the seedlings break, 0-5 days before the seedlings break, or 0-5 days after heading.

9. The method for preventing and controlling rice false smut according to claim 8, characterized in that: Spray the biocontrol agent according to any one of claims 4-5 or the biocontrol agent prepared according to the method of claim 6 10-15 days before the rice heads break.

10. The method for preventing and controlling rice false smut according to claim 9, characterized in that: The biocontrol agent according to any one of claims 4-5 or the biocontrol agent prepared according to the method of claim 6 is sprayed 10-15 days before the rice heading stage, with a spraying concentration of 1×10⁻⁶. 5 cfu / mL -1×10 7 cfu / mL.

11. The method for controlling rice false smut according to claim 8, characterized in that: Spray the biocontrol agent according to any one of claims 4-5 or the biocontrol agent prepared according to the method of claim 6 0-5 days before the rice heads break or 0-5 days after heading. A supplementary spray should be applied 3 days after the first spray, with a concentration of 1×10⁻⁶. 8 cfu / mL -1×10 10 cfu / mL.

Images