Bacillus cereus L15 and application thereof

Abstract

The invention discloses bacillus cereus L15, and relates to the technical field of microorganisms, the bacillus cereus L15 is preserved in China General Microbiological Culture Collection Center (CGMCC) on January 2, 2025, and the preservation number is CGMCC No.33276. The bacillus cereus L15 has the advantages that the bacillus cereus L15 can be used for preparing the bacillus cereus L15; the new bacillus cereus L15 provided by the invention has obvious broad-spectrum antibacterial property, can have a good inhibition effect on rhizoctonia dinuclear, fusarium and phaseolus vulgaris, and can be used for preventing and treating plant root rot and stem rot caused by the fusarium, the rhizoctonia and the phaseolus vulgaris.

Description

Technical Field

[0001] The present invention relates to the field of microbial technology, and particularly relates to a Bacillus cereus L15 and its application. Background Art

[0002] Root rot or stem rot is a serious disease caused by pathogenic fungi, which can lead to root rot of plants, growth retardation and even death. Existing pathogenic fungi causing root rot or stem rot include Phytophthora, Fusarium, Rhizoctonia, Macrophomina, etc. At present, the control of root rot mainly relies on chemical agents, but the long-term use of chemical agents is likely to cause problems such as environmental pollution, enhanced pathogen resistance and imbalance of soil microbial communities. Therefore, it is of great significance to develop efficient and environmentally friendly biological control methods.

[0003] Bacillus is a kind of beneficial microorganism widely existing in the soil, with various functions such as antifungal, growth promotion and induction of plant disease resistance. In recent years, the application of Bacillus in the control of agricultural diseases has gradually matured. However, in the control of pathogenic fungi, most of the existing Bacillus strains have a single antibacterial spectrum and lack broad-spectrum property. In the actual application process, it is often necessary to use a combination of Bacillus and other strains to effectively control pathogenic fungi. Summary of the Invention

[0004] In view of the above deficiencies of the prior art, the present invention provides a Bacillus cereus L15 with broad-spectrum antibacterial property, which can simultaneously control plant root rot and stem rot caused by Rhizoctonia solani, Fusarium solani, Fusarium oxysporum and Macrophomina phaseolina, and its application.

[0005] In order to achieve the above invention object, the technical solution adopted by the present invention is as follows:

[0006] Provide a Bacillus cereus L15 or its variant or its offspring. Bacillus cereus L15 was deposited with the China General Microbiological Culture Collection Center on January 2, 2025, and the deposit number is CGMCC No. 33276.

[0007] Furthermore, Bacillus cereus L15 can inhibit the activities of Rhizoctonia solani, Fusarium solani, Fusarium oxysporum and Macrophomina phaseolina.

[0008] The present invention also provides the application of the above-mentioned Bacillus cereus L15 in the preparation of a functional product for controlling plant fungal diseases, wherein the plant fungal diseases include root rot and stem rot of plants caused by Rhizoctonia solani, Fusarium solani, Fusarium oxysporum and Macrophomina phaseolina.

[0009] Furthermore, the functional product for controlling plant fungal diseases includes biological pesticides, biocontrol agents or microbial fertilizers.

[0010] Furthermore, the functional product for preventing and controlling plant fungal diseases is a disease control product prepared by incorporating one or more of a bacterial suspension, spores, volatile metabolites, and fermentation products of Bacillus cereus L15.

[0011] Furthermore, the plants include osmanthus, ginkgo, cedar, rhododendron, peony, rice, sugarcane, pumpkin, cowpea, jute, alfalfa, strawberry, honeysuckle, apple, and loquat.

[0012] The present invention also provides the application of the above-mentioned Bacillus cereus L15 in inducing plants to enhance resistance to fungal diseases by spraying, root irrigation, or seed coating.

[0013] The beneficial effects of the present invention are as follows:

[0014] The present invention provides a new Bacillus cereus L15, which has obvious broad-spectrum antibacterial properties and can have good inhibitory effects on Fusarium, Rhizoctonia, and Macrophomina phaseolina, and can be used to prevent and control root rot and stem rot of plants caused by Fusarium, Rhizoctonia, and Macrophomina phaseolina. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 It is a comparative diagram of the antibacterial effect of Bacillus cereus L15 on Rhizoctonia binucleata G1-3 on a plate;

[0016] Figure 2 It is a colony morphology diagram of Bacillus cereus L15;

[0017] Figure 3 It is a phylogenetic tree of Bacillus cereus L15;

[0018] Figure 4 It is an antibacterial effect diagram of Bacillus cereus L15 on 3 pathogenic fungi. DETAILED DESCRIPTION OF THE INVENTION

[0019] The following describes the specific embodiments of the present invention to facilitate the understanding of those skilled in the art of the present technology. However, it should be clear that the present invention is not limited to the scope of the specific embodiments. For those ordinary skilled in the art of the present technology, as long as various changes are within the spirit and scope of the present invention defined and determined by the appended claims, these changes are obvious, and all inventions made using the concept of the present invention are within the scope of protection.

[0020] Example 1 Isolation and Screening of Bacillus cereus L15

[0021] The test materials of the present invention were collected from the rhizosphere soil of osmanthus and ginkgo root rot diseased plants in Hong'en Temple Forest Park, Chongqing. The soil suspension was diluted by gradient concentration (10 -3 、10 -4 、10-5 and 10 -6 ) Take 0.1 mL of the dilution and spread it evenly on the LB solid medium, and incubate it upside down at a constant temperature of 28 °C. Pick single colonies with different colony sizes, colors, and edge morphologies, and use the streak plate method for purification. After purification three times, inoculate the strain into the LB liquid medium and incubate it with shaking at 200 rpm for 12 h at 28 °C. Add an equal volume of 50% glycerol and store it in a -80 °C ultra-low temperature refrigerator for later use.

[0022] Using the binucleate Rhizoctonia G1-3, the pathogen of osmanthus root rot, as the indicator bacterium, the antagonistic bacterial strains were screened by the plate confrontation culture method. First, use an inoculation loop to streak the bacterial strain to be screened in the center of the PDA solid plate, and then use a punch with a diameter of 5 mm to take a G1-3 agar disc and place it 3 cm away from the bacterial streak. At the same time, use only the inoculation of G1-3 as a control, and set 3 replicates for each strain. After incubating at a constant temperature of 28 °C for 5 d, measure the colony radius of the binucleate Rhizoctonia G1-3, and calculate the inhibition rate according to the formula:

[0023]

[0024] As Figure 1 shown, G1-3 is the control group inoculated only with the binucleate Rhizoctonia, and G1-3 + L15 is the treatment group;

[0025] The strain with the highest inhibition rate calculated by the above method was numbered L15, and its average inhibition rate was 51.95%.

[0026] Inoculate the L15 strain on the LB medium and activate it for 24 h, and observe the colony morphology and perform Gram staining. The colony morphology and Gram staining results are as Figure 2 shown, where Figure 2 A is the colony morphology diagram of L15 on the whole LB medium, Figure 2 B is the Gram staining diagram of the colony. From Figure 2 A, it can be seen that the L15 colony is nearly circular, light yellow, opaque, and has a neat edge; from Figure 2 B, it can be seen that L15 is stained blue-violet by Gram staining and is a Gram-positive bacterium, rod-shaped.

[0027] Perform phylogenetic analysis on the obtained L15 strain based on the 16s rRNA sequence. The phylogenetic tree of the L15 strain is as Figure 3As shown, strain L15 belongs to the genus Bacillus spp. and has a high genetic relationship with the sequences of B. cereus CCM 2010 and B. cereus NBRC15305. Combining the morphological characteristics of the strain and the 16S rRNA sequence information, it is determined that strain L15 belongs to Bacillus cereus and is named Bacillus cereus L15. It was deposited in the General Microbiological Center of the China Committee for Culture Collection of Microorganisms on January 2, 2025. The deposit address is No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, and the deposit number is CGMCC No. 33276.

[0028] Example 2 Verification of broad-spectrum antibacterial activity of Bacillus cereus L15

[0029] Agricultural and economic crops and garden plants are infected by fungi such as Fusarium, Rhizoctonia, and Macrophomina phaseolina, which cause diseases such as root rot and stem rot, resulting in reduced crop yields, withered and yellow leaves of garden plants, and in severe cases, large areas of plant death. In this example, the common pathogenic fungi of plants, Fusarium solani, Fusarium oxysporum, and Macrophomina phaseolina, were selected for the verification of the broad-spectrum antibacterial activity of Bacillus cereus L15. The above pathogenic fungi usually appear on agricultural and economic crops such as rice, sugarcane, pumpkin, cowpea, jute, alfalfa, strawberry, honeysuckle, apple, loquat, etc., and garden plants such as osmanthus, ginkgo, cedar, rhododendron, peony, etc.

[0030] Sterile punchers with a diameter of 5 mm were used to prepare pathogen fungal cakes, including Fusarium solani Y1-6, Fusarium oxysporum Y1-58, and Macrophomina phaseolina Y1-1, which were placed 30 mm away from the bacterial streaks. Using single inoculation of the pathogen as a control, each strain was set with 3 replicates. After culturing at a constant temperature of 28 °C for 7 days, the colony radius of Rhizoctonia solani G1-3 was measured, and the inhibition rate was calculated according to the formula. The results are as Figure 4 shown. Bacillus cereus L15 can inhibit the mycelial growth of Fusarium oxysporum Y1-58, Fusarium solani Y1-6, and Macrophomina phaseolina Y1-1, and the inhibition rates are 30.08%, 34.8%, and 44.77% respectively.

[0031] In summary, the newly isolated and screened Bacillus cereus L15 of this application can inhibit the mycelial growth of Rhizoctonia solani G1-3, Fusarium oxysporum Y1-58, Fusarium solani Y1-6, and Macrophomina phaseolina Y1-1, has broad-spectrum antibacterial properties, and can be used for the prevention and control of root rot and stem rot of plants caused by fungi.

Claims

1. A Bacillus cereus strain L15 or its variant or its progeny, wherein the Bacillus cereus L15 was deposited at the General Microbiological Center of the China Committee for Culture Collection of Microorganisms on January 2, 2025, with the deposit number CGMCC No. 33276.

2. The Bacillus cereus L15 according to claim 1, characterized in that, The Bacillus cereus L15 can inhibit the activities of Rhizoctonia solani, Fusarium solani, Fusarium oxysporum, and Macrophomina phaseolina.

3. Use of the Bacillus cereus L15 as claimed in claim 1 in the preparation of a functional product for preventing and controlling plant fungal diseases, characterized in that, The plant fungal diseases include root rot and stem rot of plants caused by Rhizoctonia solani, Fusarium solani, Fusarium oxysporum, and Macrophomina phaseolina.

4. The application according to claim 3, characterized in that, The functional products for preventing and controlling plant fungal diseases include biological pesticides, biocontrol agents, or microbial fertilizers.

5. The application according to claim 4, characterized in that The functional product for preventing and controlling plant fungal diseases is a disease control product prepared by incorporating one or more of the bacterial suspension, spores, volatile metabolites, and fermentation products of Bacillus cereus L15.

6. The application according to claim 3, characterized in that, The plants include osmanthus, ginkgo, cedar, rhododendron, peony, rice, sugarcane, pumpkin, cowpea, jute, alfalfa, strawberry, honeysuckle, apple, and loquat.

7. Use of the Bacillus cereus L15 according to claim 1 in inducing plants to enhance resistance to fungal diseases by spraying, root irrigation, or seed coating.

Images