A type of Bacillus belye and its biological agents and applications

Abstract

This application discloses a type of *Bacillus belye*, its biological agents, and applications, belonging to the field of microbial technology. The *Bacillus belye* of this application is an endophytic *Bacillus belye* from rice, and is classified as follows: Bacillus velezensis HA-B13, deposited at the China Center for Type Culture Collection (CCTCC) with accession number CCTCC NO:M 20251098, exhibits excellent biological activity against *Urspora oryzae*, achieving a 94% inhibition rate. It also demonstrates good biological activity against various rice pathogens, including *Rhizoctonia solani*, *Acanthocephala oryzae*, *Bacillus oryzae*, and *Colocynella oryzae*. Derived from rice plants, it can colonize well within the plant, providing long-term control of diseases such as panicle rot. This significantly reduces the need for pesticides and is safer, demonstrating great potential for pest and disease control.

Description

Technical Field

[0001] This application relates to the field of microbial technology, specifically to a Bacillus belyssus and its biological agents and applications. Background Technology

[0002] Rice panicle rot is a fungal disease caused by pathogens such as *Ustilago maydis* and *Acanthocephala maydis*, and it occurs widely in all rice-growing areas of China. In the field of microbial control, most of the microorganisms found to have inhibitory activity against *Ustilago maydis* are fungi. Biocontrol bacteria have only been reported in a few cases, such as *Pseudomonas*, but *Pseudomonas* are mostly opportunistic pathogens, posing certain safety risks and limiting their widespread application in practice.

[0003] Bacillus belesiensis ( Bacillus velezensis It possesses advantages such as strong resistance, rapid growth, easy cultivation, safety, and no pollution, and performs excellently in plant disease control and plant growth promotion. Currently, most *Bacillus bellis* strains exhibiting antibacterial activity against rice pathogens are derived from soil, showing control effects against rice blast, bacterial leaf blight, and bacterial leaf streak. Compared to soil-derived *Bacillus bellis*, endophytic *Bacillus bellis* strains originate from plants, making them safer for humans. Furthermore, after spraying, they can colonize within the plant, forming a biological barrier and providing long-term control. However, currently, there are relatively few endophytic *Bacillus bellis* strains with biological activity against rice pathogens isolated, and most originate from other crops such as tomatoes, peppers, and sugarcane. Due to the complex symbiotic relationship formed through long-term co-evolution between endophytic bacteria and host plants, the application of biocontrol bacteria isolated from these crops to rice carries a relatively high risk of colonization. Endophytic biocontrol bacteria isolated from rice plants can colonize within the rice plant, thus exhibiting long-term control effects against rice diseases and pests, and have greater application potential. Currently, Bacillus belladonna has been isolated from rice plants and has been shown to be effective against bacterial basal rot and rice blast fungus, but no endophytic Bacillus belladonna of rice with highly effective antibacterial activity against rice black spore fungus has been found. Summary of the Invention

[0004] In view of this, this application provides a novel Bacillus belye, classified and named as follows: Bacillus velezensis HA-B13 ， The depositary institution is the China Center for Type Culture Collection (CCTCC), located at Wuhan University, Wuhan, China. The accession number is CCTCC NO: M 20251098, and the deposit date is May 19, 2025. *Bacillus bereaves* HA-B13 exhibits excellent biological activity against *Urspora oryzae*, and also shows good inhibitory activity against various rice pathogens such as *Rhizoctonia solani*, *Acanthocephala oryzae*, *Bacillus oryzae*, and *Colocynophytum oryzae*, making it effective for the control of rice diseases.

[0005] The embodiments of this application are implemented as follows:

[0006] Firstly, this application discloses a type of Bacillus belyes, specifically an endophytic Bacillus belyes from rice, classified and named as follows: Bacillus velezensis HA-B13 is deposited at the China Center for Type Culture Collection (CCTCC), accession number: CCTCC NO: M 20251098.

[0007] In some embodiments, the Bacillus belye is isolated from rice plants; and / or, the 16S rDNA base sequence of the Bacillus belye is shown in SEQ ID NO: 1.

[0008] In some embodiments, the Bacillus belye is obtained by fermentation and isolation, wherein the fermentation conditions are as follows: the Bacillus belye is inoculated into a liquid culture medium and cultured at a pH of 6.5-7.5 and a temperature of 28-37°C for 16-20 h.

[0009] Secondly, this application provides a biological agent comprising the aforementioned Bacillus belyssus, or comprising a fermentation product obtained by fermenting the aforementioned Bacillus belyssus.

[0010] In some embodiments, the fermentation product includes fermentation broth, active proteins and secondary metabolites of Bacillus belysium isolated after fermentation, and proteins and secondary metabolites screened and developed using Bacillus belysium as the source.

[0011] Thirdly, this application also provides the application of the aforementioned Bacillus berberis or the aforementioned biological agent in the prevention and control of rice diseases.

[0012] In some embodiments, this application provides the use of Bacillus vesicles or the biological agents in the control of rice panicle rot.

[0013] In some embodiments, the pathogen causing rice panicle rot is *Urspora riceensis* or *Acanthocephala riceensis*. Alternaria padwickii One or more of them.

[0014] This application provides the application of the above-mentioned Bacillus vesiculosus or the above-mentioned biological agents in the prevention and control of rice diseases, wherein the rice diseases include one or more of sheath blight, leaf sheath black spot, and rice blast.

[0015] In some embodiments, the rice disease includes *Urspora oryzae*. Nigrospora oryzae Rice Trichophyton Alternaria padwickii Rhizoctonia solani Rhizoctonia solani , kyphotic spiral cystidia Cochliobolus geniculatus and rice blast fungus Pyricularia oryzae It is caused by at least one pathogen.

[0016] This application provides information on the above-mentioned Bacillus vesiliflorus or the above-mentioned biological agents in *Urspora rice*. Nigrospora oryzae Rice Trichophyton Alternaria padwickii Rhizoctonia solani Rhizoctonia solani , kyphotic spiral cystidia Cochliobolus geniculatus Rice blast fungus Pyricularia oryzae Applications in the prevention and control of one or more plant diseases caused by these factors.

[0017] In some embodiments, the Bacillus berreatus or the biological agent is used for seed treatment and / or applied to plants.

[0018] The *Bacillus belye* strain described in this application is an endophytic *Bacillus belye* of rice, deposited at the China Center for Type Culture Collection (CCTCC) with accession number CCTCC NO: M 20251098. Derived from rice plants, this *Bacillus belye* strain can colonize rice plants effectively, thus providing long-term control against rice rot pathogens such as *U. belye* and *Acanthocephala indicum*, significantly reducing the need for pesticides and making it safer. This bacterium also exhibits excellent biological activity against various pathogens, including rice sheath blight, rice blast, and leaf sheath black spot, with an inhibition rate exceeding 90% against *U. belye*, *Rhizoctonia solani*, and *Colocynophytum geniculateum*, demonstrating significant potential for control applications. Attached Figure Description

[0019] To more clearly illustrate the technical solutions in the embodiments of this application, 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 this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0020] Figure 1 The colony morphology of Bacillus belyss HA-B13 on agar plates;

[0021] Figure 2 Gram staining image of Bacillus belyssus HA-B13;

[0022] Figure 3 Spore staining image of Bacillus belyss HA-B13;

[0023] Figure 4 The growth curve of Bacillus belyssus HA-B13;

[0024] Figure 5The image shows the inhibitory effect of Bacillus belye HA-B13 on Ulva rice.

[0025] Figure 6 The image shows the inhibitory effect of Bacillus belyss HA-B13 on Conophytum indica.

[0026] Figure 7 The image shows the inhibitory effect of Bacillus belyss HA-B13 on Rhizoctonia solani.

[0027] Figure 8 The image shows the inhibitory effect of Bacillus belyssus HA-B13 on Coelomyces pubescens.

[0028] Figure 9 The image shows the inhibitory effect of Bacillus belyi HA-B13 on rice blast fungus.

[0029] Figure 10 Comparison of the inhibitory effects of Bacillus vesicularis HA-B13 and two commercial rice biocontrol bacteria on Ulva rice.

[0030] Figure 11 This study compares the inhibitory effects of Bacillus vesiculosus HA-B13 and two commercially available rice biocontrol bacteria on Rhizoctonia solani. Detailed Implementation

[0031] The technical solutions of the embodiments of this application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of this application, and not all embodiments. Based on the embodiments of this application, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this application. In addition, it should be understood that the specific embodiments described herein are only for illustration and explanation of this application and are not intended to limit this application. Furthermore, in the description of this application, the term "comprising" means "including but not limited to".

[0032] Various embodiments of this application may exist in the form of a range; it should be understood that the description in the form of a range is merely for convenience and brevity and should not be construed as a hard limitation on the scope of this application; therefore, it should be considered that the range description has specifically disclosed all possible sub-ranges and single numerical values ​​within that range. For example, it should be considered that the range description from 1 to 6 has specifically disclosed sub-ranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, etc., and single numbers within the range, such as 1, 2, 3, 4, 5, and 6, regardless of the range. Furthermore, whenever a numerical range is referred to herein, it means including any referenced number (fraction or integer) within the referred range.

[0033] In this application, "and / or" describes the relationship between related objects, indicating that three relationships can exist. For example, A and / or B can represent: A existing alone, A and B existing simultaneously, or B existing alone. A and B can be singular or plural.

[0034] In this application, "at least one" means one or more, and "more than one" means two or more. "At least one," "at least one of the following," or similar expressions refer to any combination of these items, including any combination of single or multiple items. For example, "at least one of a, b, or c," or "at least one of a, b, and c," can both mean: a, b, c, ab (i.e., a and b), ac, bc, or abc, where a, b, and c can be single or multiple.

[0035] This application provides a Bacillus belyceae HA-B13, classified and named as follows: Bacillus velezensis HA-B13, deposited at the China Center for Type Culture Collection (CCTCC) at Wuhan University, Wuhan, China, with accession number CCTCC NO: M 20251098, and deposited on May 19, 2025, exhibits excellent antibacterial activity against *Urspora oryzae*, as well as good antibacterial activity against various rice pathogens such as *Rhizoctonia solani*, *Acanthocephala oryzae*, *Bacillus oryzae*, and *Colocynophytum oryzae*. It can be used for the control of various rice diseases, including panicle rot, sheath blight, and leaf sheath black spot.

[0036] In one embodiment, the *Bacillus belye* HA-B13 is an endophytic *Bacillus belye* from rice. Specifically, *Bacillus belye* HA-B13 is isolated and purified from rice plants and originates from rice plants.

[0037] In this embodiment, the rice endophytic Bacillus not only has the advantages of strong resistance and environmental friendliness of Bacillus, but also, because it is isolated from rice plants, it is superior to strains from soil and other sources in terms of colonization and safety. It can coexist with rice for a long time, thereby playing a continuous role in prevention and control, and is safer for the consumption of rice and its processed products.

[0038] In some embodiments, when the Bacillus HA-B13 is cultured on LB plates at 37°C for 24 hours, the colonies are milky yellow, opaque, round, with neat, raised edges, smooth surface, and waxy appearance; after 48 hours, the colonies are milky white, opaque, round, with neat edges, adhere to the surface of the culture medium, and have 1-3 folds.

[0039] The *Bacillus belyssae* HA-B13 was identified using 16S rDNA. Its 16S rDNA was amplified using universal primers 8F and 1492R and sequenced. The result was then compared with the NCBI GenBank nucleic acid database to confirm that HA-B13 is indeed *Bacillus belyssae*. Bacillus velezensis .

[0040] This application also provides a biological agent, including the Bacillus berleis HA-B13 or a product prepared based on Bacillus berleis HA-B13.

[0041] In some embodiments, the biological agent can be prepared by liquid fermentation of Bacillus vesiculosus HA-B13 provided in this application.

[0042] Specifically, the fermentation conditions are as follows: Bacillus belye HA-B13 seed culture is transferred to a liquid culture medium and cultured for 16-20 hours at a pH of 6.5-7.5, a temperature of 28-37°C, and a shaking rate of 150-250 rpm. The liquid culture medium can be LB medium, TB medium, NA medium, etc.

[0043] In some embodiments, the biological agent may include fermentation products of Bacillus belyssus HA-B13. Specifically, the fermentation products may include fermentation broth, Bacillus belyssus isolated after fermentation, secondary metabolites, active proteins, or secondary metabolites and active proteins screened and developed using Bacillus belyssus HA-B13 as the source.

[0044] Specifically, the secondary metabolites isolated after fermentation may include one or more of the following: lipopeptides (surfactin, fengycin, iturin), polyketides (difficidin, bacillaene), and growth promoters (IAAs). The active proteins isolated after fermentation may include one or more of the following: antimicrobial proteins (such as PrI / PrII, subtilisin), enzymes (such as α-amylase), and heat-stable proteins.

[0045] Understandably, in some embodiments, the biological agent may include, in addition to Bacillus berleis HA-B13 or products prepared based on Bacillus berleis HA-B13, other additives or other biocontrol bacteria. In some specific embodiments, the biological agent is a compound biological agent formed by Bacillus berleis HA-B13 and other biocontrol bacteria, such as a compound with other Bacillus berleis, Bacillus moghaves, Bacillus pleuropsis, Bacillus amyloliquefaciens, Bacillus megaterium, Bacillus polymyxa, Bacillus subtilis, and Bacillus fanniir, etc., that have biocontrol activity.

[0046] The Bacillus vesiculus HA-B13 and biological agents provided in this application can be used for the prevention and control of rice diseases.

[0047] The *Bacillus berreatus* HA-B13 and biological agents provided in this application can be used for the control of ear rot. The main pathogens of ear rot include *Urspora oryzae*. Nigrospora oryzae、 Rice hairy beetle Alternaria padwickii The *Bacillus belye* HA-B13 and its biological agents exhibit good biological activity against *U. oryzae* and *Acanthocephala oryzae*, with an inhibition rate of over 90% against *U. oryzae*. Therefore, *Bacillus belye* HA-B13 and its biological agents have good control effects on diseases of rice and other plants caused by *U. oryzae* and *Acanthocephala oryzae*.

[0048] The Bacillus berreatus HA-B13 and biological agents provided in this application are not limited to the inhibition and control of the aforementioned pathogens, but also applicable to Rhizoctonia solani. Rhizoctonia solani , kyphotic spiral cystidia Cochliobolus geniculatus Rice blast fungus Pyricularia oryzae Suppression and control of pathogens. Effective suppression of these pathogens enables effective control of plant diseases such as sheath blight, leaf sheath black spot, and rice blast.

[0049] The *Bacillus berleis* HA-B13 and its biological agents described in this application can be used not only for the control of rice diseases, but also for the effective control of diseases caused by one or more of *U. erinaceus*, *Acanthocephala*, *Rhizoctonia solani*, *Cyclocarya oryzae*, and *Bacillus blastus* in other plants (such as crops). Therefore, *Bacillus berleis* HA-B13 and its biological agents have good control effects on plant (rice or other plant) diseases caused by one or more of *U. erinaceus*, *Acanthocephala*, *Rhizoctonia solani*, *Cyclocarya oryzae*, and *Bacillus blastus*.

[0050] In some embodiments, when Bacillus berleis HA-B13 and its biological agents are applied to the prevention and control of plant diseases such as rice, Bacillus berleis HA-B13 and its biological agents can be used for seed treatment or sprayed onto the plant surface.

[0051] The technical solutions and effects of this application are described in detail below through specific embodiments and comparative examples. These embodiments are merely some examples of this application and are not intended to limit the scope of this application. It is understood that the reagents and materials used in the embodiments and comparative examples are commercially available products unless otherwise specified. The *Urspora oryzae*, *Acanthocephala oryzae*, *Rhizoctonia solani*, *Bacillus oryzae*, and *Colocynoptera oryzae* strains used in the following embodiments are all strains isolated and preserved from rice panicle rot plants and have undergone ITS1 sequence alignment.

[0052] Example 1: Isolation and purification of Bacillus belyssus HA-B13

[0053] Rice plants were collected from well-grown paddy fields in Huai'an. After rinsing them thoroughly with tap water, the rice stems were removed and cut into small sections. The surfaces were then disinfected with a solution of 5% sodium hypochlorite and 70% ethanol, followed by rinsing with sterile water at least four times. The water was blotted dry with sterile filter paper, and the mixture was transferred to a mortar and ground thoroughly. The grinding liquid was collected and diluted 10 times. 3 10 4 10 5 After double-distillation, 100 μL of the solution was spread onto a plate (LB plate preparation: weigh 10 g tryptone (BBI), 5 g yeast extract (BBI), 10 g sodium chloride, and 15 g agar, add water to make up to 1000 mL, stir well, sterilize at 121℃ for 20 min, pour the plate when it cools to 50-60℃, seal with sealing film after cooling and solidification, and store upside down in a 4℃ refrigerator for later use), and incubate overnight at 37℃.

[0054] The disinfection effect of the tissue surface was tested using the rinsing solution test. 100 μL of the rinsing solution from the last rinsing of the disinfected material was spread on an LB agar plate and incubated simultaneously with the isolation plate. After incubation at 37°C overnight, the presence or absence of colonies was observed. If no colonies appeared, it indicated that the material surface was thoroughly disinfected and the colonies grown on the isolation plate could be used for the next step of the experiment. Otherwise, the plate was discarded and the isolation was repeated.

[0055] Colony characteristics on plates were observed using a stereomicroscope. Colonies with different morphologies were picked for liquid culture, plated again, and purified until the colony morphology on the plates was uniform. The colony morphology of HA-B13 grown on LB plates for 48 hours under a stereomicroscope is shown below. Figure 1 As shown. Colonies are milky white, opaque, round, with neat edges, adhere to the surface of the culture medium, and have 3 folds.

[0056] Example 2: Identification of Bacillus belyssus HA-B13

[0057] The preserved HA-B13 was removed and activated, and then stained using Gram staining solution and the Scharffer-Fulton spore staining kit. The staining results are shown in the figure. Figure 2 and Figure 3 , Figure 2 Gram staining diagram, Figure 3 This is a staining image of spores.

[0058] Simultaneously, 16S rDNA PCR amplification was performed using 8F / 1492R. Amplification was performed using Tks Gflex DNA polymerase (Takara, Beijing), with the following system: HA-B13 bacterial culture, 1 μL; 2×Gflex PCR Buffer (mg / mL). 2+ 25 μL of dNTP plus; 1 μL of TKS Gflex DNA polymerase; 2 μL of 8F (10 mM); 2 μL of 1492R (10 mM); and 19 μL of ddH2O. Amplification was performed using a PCR instrument with the following program: 94℃ pre-denaturation for 1 min, 98℃ denaturation for 10 s, 55℃ annealing for 15 s, and 68℃ extension for 2 min, for a total of 30 cycles, followed by a 72℃ extension for 10 min. After PCR, agarose gel electrophoresis was performed, and a single band of approximately 1500 bp was purified and sequenced. Sequence alignment with the NCBI GenBank nucleic acid database, combined with the morphological characteristics and relevant physiological and biochemical characteristics of the bacterium, confirmed that strain HA-B13 was *Bacillus belesiensis*. Bacillus velezensis It was named Bacillus belyssus HA-B13.

[0059] The 16S rDNA sequence of Bacillus belyssus HA-B13 is shown in SEQ ID NO: 1, and is as follows:

[0060]

[0061] Example 3: Growth curve of Bacillus belyssus HA-B13

[0062] The preserved HA-B13 was diluted and plated. Single colonies were picked and transferred to 5 mL of LB liquid medium and cultured with shaking at 200 rpm and 37°C until the OD600 reached 1.0, which served as seed culture. Fresh medium was then added at a ratio of 1:40, and 100 μL was collected every hour to measure the OD600 value. The growth curve is shown below. Figure 4 The x-axis represents incubation time (in hours), and the y-axis represents OD600.

[0063] Depend on Figure 4 It can be seen that Bacillus belye HA-B13 grows extremely fast, and a large number of cells can be obtained in a very short time. Its OD600 value reaches above 1.0 after 3 hours of transfer and above 2.0 after 5 hours.

[0064] Example 4: Inhibitory effect of Bacillus belye HA-B13 on rice panicle rot

[0065] Fold a 9cm diameter piece of paper in half, then fold it in half again to center the petri dish. Place a 0.5cm diameter *Ulva oryzae* fungus inside. Nigrospora oryzae The mycelium was inoculated onto the center point, and a seed of *Bacillus belye* HA-B13 was applied 1.5 cm from the center point using an inoculation loop. A plate inoculated only with *U. oryzae* was used as a control. The inhibitory effect of *Bacillus belye* HA-B13 on *U. oryzae* was observed after 7 days. (*Candida albicans*) Alternaria padwickii The inoculation method for the inhibition test was the same as that for *Ichthyophthirius multifiliis*. After 7 days, the inhibitory effect of *Bacillus belye* HA-B13 on *Conophytum glutenosa* was observed.

[0066] In this embodiment, *Ustilago mays* Nigrospora oryzae and rice hairy beetle Alternaria padwickii These are strains isolated and preserved from rice panicle rot plants in our laboratory, and their ITS1 sequences have been compared. Among them, *Urspora oryzae*... Nigrospora oryzae and rice hairy beetle Alternaria padwickii It is the main pathogen of rice panicle rot.

[0067] See results Figure 5 and Figure 6 , Figure 5 The graph shows the inhibitory effect of Bacillus belysin HA-B13 on Ustilago oryzae. The left side is the Ustilago oryzae blank control, and the right side is the inhibitory effect of Bacillus belysin HA-B13 on Ustilago oryzae. Figure 6 The image shows the inhibitory effect of Bacillus belysin HA-B13 on Conophytum indica. The left side is the blank control of Conophytum indica, and the right side is the inhibitory effect of Bacillus belysin HA-B13 on Conophytum indica.

[0068] Depend on Figure 5 and Figure 6 It can be seen that Bacillus belye HA-B13 has a good inhibitory effect on the growth of *Ulva oryzae* and *Acanthocephala oryzae*, with an inhibition rate of 94% against *Ulva oryzae* and 81% against *Acanthocephala oryzae*. Bacillus belye HA-B13 has a good inhibitory and control effect on rice panicle rot.

[0069] Example 5: Inhibitory effect of Bacillus belyssus HA-B13 on Rhizoctonia solani.

[0070] Rhizoctonia solani Rhizoctonia solani The inoculation method for the inhibition test was the same as that for *Urspora oryzae*. Four days after inoculation, the inhibitory effect of *Bacillus belye* HA-B13 on *Rhizoctonia solani* was observed. In this example, *Rhizoctonia solani* was a strain preserved in the laboratory. Results are shown below. Figure 7 The left side shows the *Rhizoctonia solani* blank control, and the right side shows the inhibitory effect of *Bacillus belysiniana* HA-B13 on *Rhizoctonia solani*. Figure 7 It can be seen that Bacillus belye HA-B13 has an excellent inhibitory effect on the growth of Rhizoctonia solani, with an inhibition rate of approximately 92%.

[0071] Example 6: Inhibitory effect of Bacillus belyssus HA-B13 on Coelophysis davidii.

[0072] Knee-shaped spiral cystidia Cochliobolus geniculatus The inoculation method for the inhibition test was the same as that for *Urspora oryzae*. Seven days after inoculation, the inhibitory effect of *Bacillus belye* HA-B13 on *Colocasia oryzae* was observed. In this example, *Colocasia oryzae* was a strain isolated and preserved from rice plants infected with leaf sheath black spot disease in our laboratory, and its ITS1 sequence was verified. Results are shown below. Figure 8 The left side shows the blank control of *Coelophysis geniculateus*, and the right side shows the inhibitory effect of *Bacillus belysinus* HA-B13 on *Coelophysis geniculateus*. Figure 8 It can be seen that Bacillus belye HA-B13 has an excellent inhibitory effect on the growth of Coelomyces geniculateus, with an inhibition rate of approximately 91%.

[0073] Example 7: Inhibitory effect of Bacillus belye HA-B13 on rice blast fungus.

[0074] Rice blast fungus Pyricularia oryzae The inoculation method for the inhibition test was the same as that for *Urspora oryzae*. Seven days after inoculation, the inhibitory effect of *Bacillus belye* HA-B13 on *Bacillus oryzae* was observed. In this example, the *Bacillus oryzae* strain was a laboratory-preserved strain. Results are shown below. Figure 9 The left side shows the blank control of rice blast fungus, and the right side shows the inhibitory effect of Bacillus belye HA-B13 on rice blast fungus. Figure 9It can be seen that Bacillus belyi HA-B13 has a good inhibitory effect on the growth of rice blast fungus, with an inhibition rate of about 83%.

[0075] Example 8: Comparison of the inhibitory effect of Bacillus belyssus HA-B13 on rice pathogens with commercially available biocontrol bacteria.

[0076] Currently, there are no registered Bacillus vesiculosus microbial pesticides for the control of rice diseases and pests. Therefore, commercially available formulations containing Bacillus subtilis and Bacillus cereus were purchased from the market, diluted, plated, and single colonies were collected and preserved by shaking. Then, the inhibition rates of Bacillus subtilis and Bacillus cereus against *U. oryzae* and *Rhizoctonia solani* were determined using the same method as the HA-B13 inhibition test. The inhibition rates of HA-B13 against *U. oryzae* were compared with those of the two commercially available biocontrol bacteria. Figure 10 The inhibition rates against Rhizoctonia solani were compared. Figure 11 .Depend on Figure 10 and Figure 11 It can be seen that Bacillus belye HA-B13 has a significantly better inhibitory effect on *Urspora oryzae* than commercially available biocontrol agents *Bacillus subtilis* and *Bacillus cereus*, with inhibition rates 11% and 33% higher, respectively; its inhibition rate on *Rhizoctonia solani* is comparable to that of commercially available biocontrol agents *Bacillus subtilis* and *Bacillus cereus*.

[0077] As can be seen from the above embodiments, the Bacillus vesiculosus HA-B13 provided in this application has good inhibitory activity against a variety of rice pathogens. Among them, the inhibition rate against *Ustilago maydis*, *Rhizoctonia solani*, and *Colocynophytum genicum* reaches more than 90%, and the inhibition rate against *Acanthocephala indicum* and *Bacillus oryzae* reaches 80% or more. It can be used for the prevention and control of various rice diseases such as panicle rot, sheath blight, and leaf sheath black spot, and can also play a good role in the prevention and control of diseases of other plants caused by the above pathogens.

[0078] In addition, Bacillus belye HA-B13 was isolated from rice plants, and it faces fewer obstacles to colonization or application on rice compared to biocontrol bacteria from other sources, making it a promising candidate for application.

[0079] The above provides a detailed description of Bacillus belyssus and its biological agents and applications provided in the embodiments of this application. Specific examples have been used to illustrate the principles and implementation methods of this application. The descriptions of the above embodiments are only for the purpose of helping to understand the methods and core ideas of this application. At the same time, for those skilled in the art, there will be changes in the specific implementation methods and application scope based on the ideas of this application. Therefore, the content of this specification should not be construed as a limitation of this application.

Claims

1. A type of Bacillus belesii, characterized in that, The *Bacillus belye* mentioned is an endophytic *Bacillus belye* from rice, classified and named as follows: Bacillus velezensis HA-B13, deposited at the China Center for Type Culture Collection (CCTCC) with accession number M 20251098, is a Bacillus belye, isolated and purified from rice plants and used for the control of rice diseases.

2. The Bacillus belye according to claim 1, characterized in that, The *Bacillus belyssus* was obtained through fermentation. The fermentation conditions were as follows: the *Bacillus belyssus* was inoculated into a liquid culture medium and cultured at a pH of 6.5-7.5 and a temperature of 28-37°C for 16-20 h.

3. A biological agent, characterized in that, The biological agent includes Bacillus belyssus as described in claim 1 or 2.

4. The application of *Bacillus belye* according to claim 1 or 2, or the biological agent according to claim 3, in the control of rice panicle rot; wherein the pathogen of rice panicle rot is *Urspora oryzae*. Nigrospora oryzae or rice hyacinth Alternaria padwickii One or more of them.

5. The Bacillus belye according to claim 1 or 2, or the biological agent according to claim 3, in Rhizoctonia solani Rhizoctonia solani Application in the prevention and control of plant diseases caused by rice; the plant in question is rice.

6. The Bacillus berberis according to claim 1 or 2, or the biological agent according to claim 3, in rice blast fungus. Pyricularia oryzae Application in the prevention and control of plant diseases caused by rice; the plant in question is rice.

7. The application according to any one of claims 4-6, characterized in that, The Bacillus berreatus or the biological agent is used for seed treatment and / or applied to plants.

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