Bacillus tequilensis ZW-35 and application thereof

Abstract

The invention provides bacillus tequilensis ZW-35 and application thereof, and relates to the technical field of microbial control. The bacillus tequilensis ZW-35 is preserved in the China General Microbiological Culture Collection Center (CGMCC), the classification name of the bacillus tequilensis ZW-35 is bacillus tequilensis, and the preservation number of the bacillus tequilensis ZW-35 is CGMCC No. 32878. Volatile substances in the metabolic process of the bacillus tequilensis ZW-35 can effectively inhibit the growth of colletotrichum gloeosporioides; and the volatile substances have efficient diffusion capability in air and soil gaps, so that the volatile substances can not be in direct contact with the fruits or leave residues on the surfaces of the fruits in the prevention and treatment of the fruit diseases, and potential risks caused by direct contact are avoided.

Description

Technical Field

[0001] The present invention relates to the technical field of microbial control, and specifically to a strain of Bacillus tequila ZW-35 and an application thereof. Background Art

[0002] Anthrax is a fungus belonging to the subphylum Ascomycota, class Coelosporales, order Melanosporales, family Melanosporaceae, and genus Anthrax. The morphology of Anthrax includes conidia, septate hyphae, conidiophores, setae, and other components. Its life cycle includes the conidia stage and hyphae stage. It enters the plant body by infecting plant wounds or natural openings, causing necrosis and decay of plant tissues.

[0003] When plants are infected with anthrax, yellow or white spots will appear on leaves, stems, and fruits. As the disease worsens, the spots gradually expand, tissue necrosis occurs, and the plant eventually dies. When plants are infected with anthrax, their photosynthesis capacity decreases and nutrient synthesis decreases, resulting in slow plant growth and reduced yields. In addition, plant anthrax may also reduce the plant's resistance to stress and make it more susceptible to infection by other pathogens.

[0004] At present, the prevention and control of anthrax mainly include chemical control, physical control, agricultural control and biological control. Chemical control mainly uses chemical agents to prevent and control diseases caused by plant anthrax. Commonly used chemical agents include carbendazim, thiophanate-methyl, difenoconazole, etc., but the use of chemical agents is easy to cause pollution to plants and the environment; physical control mainly uses ultraviolet light to kill conidia on the surface of plants to reduce the number of pathogens, or use high temperature treatment, but the timing of plant treatment is required to be high, and the final treatment may also affect the growth of plants, so the application scope is small; biological control uses microbial resources for prevention and control. Some microorganisms such as Bacillus and Trichoderma have the effect of inhibiting the growth of plant anthrax and can be used as candidate strains for biological control. Relatively speaking, biological control has the advantages of environmental protection and sustainability, but the effect of biological control requires more verification and determination of relevant microorganisms. Summary of the invention

[0005] In view of the shortcomings of the prior art, the present invention provides a strain of Bacillus tequila ZW-35 and its application. The volatile substances metabolized by Bacillus tequila ZW-35 can effectively inhibit the growth of anthrax and control the deterioration and development of fruit anthrax, providing effective biological control for the prevention and control of anthrax.

[0006] To achieve the above objectives, the present invention is implemented through the following technical solutions: A strain of Bacillus tequila ( Bacillus tequilensis) ZW-35, the tequila bacillus ZW-35 is deposited in the General Microbiological Center of China Microbiological Culture Collection Administration Committee and is classified and named as tequila bacillus Bacillus tequilensis , the deposit number is CGMCC No.32878.

[0007] Bacillus tequila ZW-35 is used to inhibit anthrax bacteria.

[0008] Preferably, the application method is to use the volatile substances of Bacillus tequila ZW-35 to inhibit anthrax.

[0009] Bacillus tequila ZW-35 is used to prevent and control fruit anthracnose.

[0010] Preferably, the application method is to use the volatile substances of Bacillus tequila ZW-35 to prevent and treat fruit anthracnose. The present invention provides a strain of Bacillus tergillosis ZW-35 and its application, which has the advantages of: The invention provides a strain of Bacillus tequila ZW-35, volatile substances in the metabolic process of which can effectively inhibit the growth of anthrax; and the volatile substances have high-efficiency diffusion ability in the air and soil gaps. In the prevention and control of fruit diseases, the volatile substances can avoid direct contact with the fruit and will not leave residues on the surface of the fruit, thereby avoiding potential risks caused by direct contact. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] Figure 1 Schematic diagram of the colony morphology of Bacillus tequila ZW-35 of the present invention; Figure 2 is a phylogenetic tree of Bacillus tequila ZW-35 of the present invention; Figure 3 Schematic diagram of the antibacterial effect of the plate in Example 2 of the present invention for 8 days, wherein the left side shows the antibacterial effect of the plate in the control group, and the right side shows the antibacterial effect of the plate in the group treated with volatile substances of Bacillus tequila; Figure 4 This is a schematic diagram of the structure of anthrax hyphae cells cultured for 8 days in the blank control group in Example 2 of the present invention; Figure 5 This is a schematic diagram of the structure of anthrax hyphae cells cultured for 8 days in the volatile substance treatment group of Bacillus tequila in Example 2 of the present invention; Figure 6 This is a schematic diagram of the growth of lesions on mangoes after 12 days in Example 3 of the present invention, wherein the left side is a sterile water treatment group, and the right side is a volatile substance treatment group of Bacillus tequila. DETAILED DESCRIPTION

[0012] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention is clearly and completely described below in combination with the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0013] The following strain of Bacillus tequila ZW-35 is deposited in the General Microbiology Center of China Microbiological Culture Collection Administration, the deposit address is: No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, and is classified as Bacillus tequila Bacillus tequilensis The deposit number is CGMCC No.32878, and the deposit time is December 2, 2024. Embodiment 1:

[0014] Isolation and Identification of Bacillus tequila ZW-35: Bacillus tequila ZW-35 was isolated from healthy tea leaves: Endogenous bacteria in tea leaves were isolated by the dilution plate method. The specific operation was as follows: healthy tea leaves were immersed in 75% ethanol solution for 2-3 seconds, then washed three times with sterile water to remove the residual ethanol solution, and finally the healthy tissue was cut with a sterile blade and transferred to LB solid medium. It was cultured in a constant temperature box at 28℃ for 48 hours, and the strain was purified by streak method ( Figure 1 ), and finally, the purified strain was identified using 16S rRNA. The results showed that the full length of the 16S rRNA sequence of Bacillus tekila ZW-35 was 1438 bp, which was similar to that of Bacillus tekila Bacillus tequilensis SJ33 and Bacillus tequila Bacillus tequilensis 10b have sequence similarities of 99.50% and 99.40%, respectively (Table 1), and are similar to Bacillus tequila Bacillus tequilensis The strains are in a phylogenetic branch ( Figure 2 ), so strain ZW-35 was identified as Bacillus tequila Bacillus tequilensis .

[0015] Table 1 Embodiment 2:

[0016] 1. Antibacterial effect of volatile substances of Bacillus tequila ZW-35 on anthrax: Use a cork borer to punch out a 5 mm anthrax cake and inoculate it in the center of the left medium of the two-divided medium. Apply 100 μL of 107 CFU / mL of Bacillus tergillosis suspension, with the blank medium without Bacillus on the right as the control, each treatment was repeated 3 times, cultured at 28℃ for 8 days, the colony diameter of the tested pathogens was measured, and the inhibition rate was calculated. The specific results are shown in Figure 3 As shown in Table 2 below: Table 2

[0017] It can be seen from the above table that the volatile substances of Bacillus tequila have a good inhibitory effect on pine needle anthracnose, and the inhibition rate after 8 days is 55.6%.

[0018] 2. Destruction of anthrax hyphae cells by volatile substances of Bacillus tequila ZW-35: The hyphae of anthrax pathogens were collected by the inverted method. The purified anthrax cake taken by the puncher was placed in the middle of the prepared PDA solid medium. Then, the LB solid medium coated with 200 μL of Bacillus tequila ZW-35 was inverted on the PDA solid medium with the anthrax cake. After sealing with a sealing film, it was cultured at 28°C for 5-8 days. The uninoculated LB medium was used as a blank control. After 8 days, the pine needle anthrax hyphae of the control group and the treatment group were cut with a scalpel, fixed with glutaraldehyde, and observed by transmission electron microscopy. Figure 4 (blank control group) and Figure 5 (Bacillus tequila volatile substances treatment group).

[0019] The results showed that the volatile substances of Bacillus tequila destroyed the structural integrity of anthrax hyphae cells, damaged the cell walls and cell membranes, and caused the cell contents to leak out. Embodiment 3:

[0020] Volatile substances of Bacillus tequila ZW-35 can prevent and control mango anthracnose: Mango fruits with no mechanical damage, uniform size, no pests and diseases, and uniform maturity were selected as experimental materials, disinfected with sodium hypochlorite solution, washed with sterile water and dried naturally for use. A hole was pierced at the equator of the fruit with a sterilized gun tip (4 mm deep and 4 mm in diameter), and 10 μL of anthrax spore suspension at a concentration of 10 was quantitatively injected into the hole with a micropipette. 4 CFU / mL, 3 fruits were treated in each treatment, repeated three times, and then the mangoes were placed in plastic boxes. 7The LB solid medium containing CFU / mL Tequila Bacillus ZW-35 suspension was opened and placed in the middle of the box containing mangoes inoculated with pathogens, and then the plastic box lid was closed. The culture dish without Bacillus inoculation was used as a control. The incidence and lesions of the fruit were observed every day. The growth of lesions on day 12 was as follows: Figure 6 The formula for calculating the incidence rate is as follows: Incidence rate (%) = (number of diseased wells / total number of wells) × 100%. Each treatment was repeated 3 times, and the specific incidence rates are shown in Table 3: Table 3

[0021] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the same. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some of the technical features may be replaced by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A strain of Bacillus tequila ( Bacillus tequilensis )ZW-35, characterized by: The Bacillus tequila ZW-35 is deposited in the General Microbiological Center of China Microbiological Culture Collection Administration and is classified and named as Bacillus tequila Bacillus tequilensis , the deposit number is CGMCC No.32878.

2. Use of the Bacillus tequila ZW-35 as claimed in claim 1 in inhibiting anthrax bacteria.

3. The use according to claim 2, characterized in that: The application method is to use the volatile substances of Bacillus tequila ZW-35 to inhibit anthrax.

4. The Bacillus tequila ZW-35 as claimed in claim 1 is used for preventing and controlling anthracnose of fruits.

5. The use according to claim 4, characterized in that: The application method is to use the volatile substances of Bacillus tequila ZW-35 to prevent and control fruit anthracnose.

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