Compound reagent for preventing and treating tobacco bacterial wilt and application

Abstract

The invention discloses a compound reagent for preventing and treating tobacco bacterial wilt and application, and belongs to the technical field of tobacco disease prevention and control, the compound reagent is prepared by taking epsilon-polylysine and biocontrol bacterium fermentation liquor according to the mass ratio of (1-2): (3-5), then the compound reagent is sprayed on tobacco leaves, the cigar bacterial wilt can be effectively prevented and treated, and the yield of cigars is increased. By adopting the biological agent, the tobacco bacterial wilt is effectively prevented and treated, meanwhile, pathogenic bacteria are prevented from generating drug resistance, and meanwhile damage to the environment is relieved.

Description

Technical Field

[0001] The invention relates to the technical field of tobacco disease prevention and treatment, and in particular to a compound reagent for preventing and treating tobacco bacterial wilt and an application thereof. Background Art

[0002] Cigar is a tobacco product made from dried and fermented tobacco. Cigar tobacco is mainly produced in Brazil, Cameroon, Cuba and other countries. Some areas in my country also cultivate tobacco for making cigars.

[0003] In some cigar producing areas in my country, bacterial wilt is common. It is one of the main diseases of cigar tobacco. Some fields even have no harvest. The epidemic of bacterial wilt is becoming more and more serious. Therefore, the prevention and control of this disease should be paid great attention. In the early stage of the disease, 1 to 2 leaves can often be seen withering and drooping at noon on a sunny day, and they can recover at night. Observe the stem. There are often chlorotic stripes on the stem on one side of the wilting leaves. The root system on the side of the rotten stem rots into dark brown to black. If the wilting leaves are picked, the vascular bundles of the petioles can be seen to turn black. The infected leaves are still green and hanging on the stems at the beginning, so it is called "bacterial wilt". Because one side of the diseased leaves often wither in the early stage, it is commonly known as "half-side madness". When the weather is hot, the leaves of the diseased plants are irregularly burned, the leaves dry, and the edges fall off. The stems of the diseased plants are upright, with dead leaves hanging on the stems. When the soil humidity is high, the roots become soft and sticky, causing the whole plant to die. When tobacco stems with streaks are cut cross-section, the vascular tissue turns brown or brown. The pith begins to rot and becomes honeycomb-like, and a cavity forms at the base of the tobacco stem. There is sticky milky white to light brown glittering mucus at the vascular incision; sticky filaments can also be seen. To identify this disease, the diseased vascular tissue can be placed in a container filled with light salt water. After a few minutes, a milky white mist can be seen.

[0004] At present, most of the registered pesticides for bacterial wilt are chemical pesticides, such as thiophanate-methyl and trichloroisocyanuric acid. Long-term use of chemical pesticides to control diseases has been found to easily lead to drug resistance in pathogens, and long-term use also harms the environment. Therefore, it is urgent to find new and efficient control methods to improve this situation. Summary of the invention

[0005] The purpose of the present invention is to provide a composite agent for preventing and treating tobacco bacterial wilt and its application. ε-polylysine is mixed with a fermentation liquid of a biocontrol bacteria in a certain proportion and sprayed on tobacco leaves to effectively prevent and treat cigar tobacco bacterial wilt. The biological bacterial agent is used to effectively prevent and treat tobacco bacterial wilt, while preventing pathogens from developing drug resistance and reducing damage to the environment.

[0006] The present invention is achieved through the following technical solutions: A composite agent for preventing and treating tobacco bacterial wilt is prepared by taking epsilon-polylysine and biocontrol bacteria fermentation liquid in a mass ratio of 1-2:3-5.

[0007] Furthermore, the mass ratio of ε-polylysine to the biocontrol bacteria fermentation broth is 1:3.

[0008] Furthermore, the biocontrol bacteria fermentation liquid is prepared by mixing the fermentation liquid of Brevibacillus laterosporus and the fermentation liquid of Rheinheimerella aquae in a mass ratio of 0.8-1.1:1.9-2.3.

[0009] Furthermore, the mass ratio of the fermentation broth of Brevibacillus laterosporus to the fermentation broth of Rheinheimerella aquae is 1:2.

[0010] Furthermore, the fermentation broth of Brevibacillus laterosporus and the fermentation broth of Rheinheimerella aquatica are obtained by the following preparation method: (1) Inoculate Brevibacillus laterosporus or Rheinheimerella aquatica into LB solid medium for activation; (2) The activated Brevibacillus laterosporus and Rheinheimerella water were inoculated into LB liquid culture medium respectively, and cultured at 30°C and 160 r / min for 36 h to obtain seed solution; (3) The seed solution was transferred to new LB liquid culture medium, and cultured at 37°C and 200 r / min for 48 h to obtain the fermentation broth of Brevibacillus laterosporus and Rheinheimerella aquatica.

[0011] Furthermore, in step (3), the number of viable bacteria in the fermentation broth of Brevibacillus laterosporus is greater than 1×10 8 cfu / mL; the number of viable bacteria in water Rheinheimerella fermentation broth is greater than 3×10 7 cfu / mL.

[0012] Use of any of the aforementioned composite reagents for preventing and controlling tobacco bacterial wilt in preventing and controlling tobacco bacterial wilt in cigar tobacco.

[0013] Furthermore, the biological agent is applied on the 9th to 12th day after the cigar tobacco is transplanted, and is applied again after an interval of 14 to 16 days.

[0014] Furthermore, the application amount each time is 80 ml / mu.

[0015] Compared with the prior art, the present invention has the following advantages and beneficial effects: 1. The present invention proposes a compound reagent prepared by using ε-polylysine and biocontrol bacteria fermentation liquid, which can be used in the prevention and treatment of tobacco bacterial wilt, especially for cigar tobacco bacterial wilt, and has a good prevention and treatment effect. At the same time, the compound reagent is a biological bacterial agent. During the application process, compared with the application of chemical agents, pathogens are not easy to develop drug resistance, and biological bacterial agents are more environmentally friendly.

[0016] 2. The present invention proposes a compound agent for preventing and treating tobacco bacterial wilt, which is a biological bacterial agent and provides a technical idea for preventing and treating diseases in the tobacco planting process.

[0017] 3. In the present invention, a composite agent for preventing and controlling tobacco bacterial wilt is proposed, which can be used alone in the disease prevention and control technology in the tobacco planting process, and can also be used in combination with chemical agents or used at intervals. It can reduce the use of chemical pesticides and the risk of drug resistance, protect the ecological environment, meet the requirements of green plant protection and ecological civilization construction, is safe and harmless to the environment, and has good development prospects and application potential.

[0018] 4. In the present invention, a composite agent is proposed for the optimal prevention and treatment of tobacco bacterial wilt. The mass ratio of ε-polylysine to the biocontrol bacteria fermentation liquid is 1:3, and the mass ratio of the fermentation liquid of Brevibacillus laterosporus to the fermentation liquid of Rheinheimerella aquatica is 1:2. The composite agent has an excellent control effect on the bacterial wilt of cigar tobacco, which can be as high as 83.48%, which is higher than the control effect of conventional chemical agents. DETAILED DESCRIPTION

[0019] The present invention is further described in detail below in conjunction with examples, but the embodiments of the present invention are not limited thereto.

[0020] In the following examples, the strains / reagents were derived from: The Brevibacillus laterosporus is Brevibacillus laterosporus SNB10, which is deposited in the General Microbiological Center of the China National Microbiological Culture Collection Administration with a deposit number of 14171; such as the Brevibacillus laterosporus disclosed in patent CN201710536055.8.

[0021] The water Rheinheimerella is the water Rheinheimerella disclosed in patent application CN202111612846, and the deposit number is CCTCC NO:M2015245.

[0022] When the compound agent for preventing and controlling tobacco bacterial wilt of the present invention is used for preventing and controlling tobacco bacterial wilt of cigar tobacco, the biological agent is applied 10 days after transplanting and applied again after an interval of 15 days, and the application amount each time is 80 ml / mu.

[0023] Example 1 1. Test materials Drugs: ε-polylysine, Brevibacillus laterosporus, Rheinheimerella water, and sterile water as blank control.

[0024] The preparation of LB liquid medium includes the following steps: 950 ml of deionized water, 10 g of tryptone, 5 g of yeast extract, and 10 g of NaCl. Use 1 M NaOH to adjust the pH value to 7.0 and adjust the volume to 1 L. After the above steps, add 15 g of agar powder to the LB solid medium, sterilize it by high pressure at 121°C for 20 min, and pour the solid medium into a plate when it cools to 55°C; the liquid medium is divided for use.

[0025] 2. Preparation of compound reagents for controlling tobacco bacterial wilt According to each experimental group in Table 1, a certain mass ratio of ε-polylysine and biocontrol bacteria fermentation liquid was taken to prepare a composite agent for preventing and controlling tobacco bacterial wilt. Among them, the biocontrol bacteria fermentation liquid in groups 1 to 5 was a 1:2 uniform mixture of Brevibacillus laterosporus fermentation liquid and water Rheinheimerella fermentation liquid; the biocontrol bacteria fermentation liquid in groups 6 and 7 was a 0.8:1.1 uniform mixture of Brevibacillus laterosporus fermentation liquid and water Rheinheimerella fermentation liquid; the biocontrol bacteria fermentation liquid in groups 8 and 9 was a 1.1:1.8 uniform mixture of Brevibacillus laterosporus fermentation liquid and water Rheinheimerella fermentation liquid.

[0026] The Brevibacillus laterosporus fermentation liquid and the Rheinheimerella water fermentation liquid are obtained by the following preparation method: (1) Inoculate Brevibacillus laterosporus or Rheinheimerella aquatica into LB solid medium for activation; (2) The activated Brevibacillus laterosporus and Rheinheimerella water were inoculated into LB liquid culture medium respectively, and cultured at 30°C and 160 r / min for 36 h to obtain seed solution; (3) The seed solution was transferred to new LB liquid culture medium, and cultured at 37°C and 200 r / min for 48 h to obtain the fermentation broth of Brevibacillus laterosporus and Rheinheimerella aquatica.

[0027] In the obtained composite reagent, the number of viable bacteria in the fermentation liquid of Brevibacillus laterosporus is greater than 1×10 8 cfu / mL; the number of viable bacteria in water Rheinheimerella fermentation broth is greater than 3×10 7 cfu / mL.

[0028] 3. Application method Ten days after tobacco transplanting, the leaves were sprayed with the test agent, ε-polylysine, Brevibacillus laterosporus, Rheinheimerella water, and sterile water, and sprayed again 15 days later. It should be noted that, unless otherwise specified, tobacco planting, fertilization, and field management were carried out according to conventional cultivation methods.

[0029] After the disease begins, investigate the disease situation, use five-point sampling survey, survey 8 plants at each point, and survey 40 plants in each plot. Record the number of diseased plants, the number of diseased leaves per plant, the total number of leaves surveyed, and the disease index of each leaf. The grading standard of cigar tobacco bacterial wilt disease index is as follows (grading survey is conducted on leaves, and 5 leaves are surveyed per plant): Level 0: The whole plant is disease-free; Level 1: There are occasional chlorotic spots on the stem, or less than half of the leaves on the diseased side wilt; Level 3: There are black streaks on the stem, but not exceeding half of the stem height, or one-half to two-thirds of the leaves on the diseased side wilt; Level 5: The black streaks on the stem exceed half of the stem height but do not reach the top of the stem, or more than two-thirds of the leaves on the diseased side wilt; Level 7: Black streaks on the stem reach the top of the stem, or all leaves of the diseased plant wilt; Level 9: The diseased plants are basically dead.

[0030] Then, the tobacco morbidity, disease index and relative prevention effect were examined to comprehensively evaluate the prevention and control effect of the reagent. The results are shown in Table 1. The calculation method of the indicators involved is as follows: Incidence rate / % = number of diseased plants / total number of plants investigated × 100; Disease index = Σ (number of diseased leaves at each level × representative value at each level) / (total number of leaves surveyed × highest representative value) × 100; Relative prevention effect / %=(control disease index - treatment disease index) / control disease index×100.

[0031] Table 1: Investigation on the effectiveness of prevention of cigar tobacco bacterial wilt As can be seen from Table 1, the use of the compound reagent in this scheme, or the use of ε-polylysine, Brevibacillus laterosporus, and Rheinheimerella water alone, all have a certain control effect on cigar tobacco bacterial wilt, and the incidence rate is significantly reduced (the incidence rate of the blank group is 74%). From Groups 1 to 9, it can be seen that the incidence rate of cigar tobacco bacterial wilt is significantly lower than that of the group using ε-polylysine, Brevibacillus laterosporus, and Rheinheimerella water alone, and the relative prevention efficiency is also significantly improved. In this experimental group, the reagent in Group 8 has the best control effect on cigar tobacco bacterial wilt, with a relative prevention efficiency of 83.48%.

[0032] Comparative Example 1 In this comparative example, the control effect of some commercially available chemical agents on cigar tobacco bacterial wilt in the field was investigated.

[0033] 1. Test materials Drugs: 80% Bordeaux mixture wettable powder (commercially available), 77% copper calcium sulfate wettable powder (commercially available), and sterile water as blank controls.

[0034] 2. Application method 10 days after tobacco transplanting, the test agent was sprayed on the tobacco leaves, and the control group was sprayed with the same amount of sterile water. After 15 days, it was sprayed again. Each treatment was repeated 3 times, and the planting, fertilization, field management and cultivation methods of cigar tobacco in the embodiment were also used. The amount of reagent used was the recommended amount in the reagent manual.

[0035] After the onset of disease, the incidence was investigated according to the grading standards and relative preventive efficacy in Example 1. The investigation results are shown in Table 2.

[0036] Table 2 Investigation of the efficacy of different pesticides As shown in Table 2, the commercially available 80% Bordeaux mixture wettable powder and 77% copper calcium sulfate wettable powder have similar relative control effects on cigar tobacco bacterial wilt, and the 77% copper calcium sulfate wettable powder is slightly better than the 80% Bordeaux mixture wettable powder, but their relative control effects are all lower than 70%. The composite reagents used in this scheme for preventing and controlling tobacco bacterial wilt, referring to Groups 1 to 9 in Example 1, have relative control effects of 77% copper calcium sulfate wettable powder and 77% Bordeaux mixture wettable powder. Higher than 76.43% , up to 83.48%, which is significantly higher than the anti-bacterial wilt effect of chemical reagents sold on the market, that is, the composite reagent of the present invention has a better effect of preventing and controlling cigar tobacco bacterial wilt.

[0037] At the same time, the compound reagent of this scheme is a biological agent, which improves the stress resistance of tobacco compared with traditional chemical pesticides, and is safe, non-toxic, pollution-free, and environmentally friendly. It can be put into research and production as a biological pesticide for preventing and controlling cigar tobacco wilt disease, and can partially replace the use of traditional chemical pesticides.

[0038] Although the preferred embodiments of the present invention have been described, those skilled in the art may make other changes and modifications to these embodiments once they have learned the basic creative concept. Therefore, the appended claims are intended to be interpreted as including the preferred embodiments and all changes and modifications that fall within the scope of the present invention.

[0039] Obviously, those skilled in the art can make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include these modifications and variations.

Claims

1. A composite agent for preventing and treating tobacco bacterial wilt, characterized in that: The product is prepared by taking ε-polylysine and biocontrol bacteria fermentation liquid in a mass ratio of 1-2:3-5.

2. The composite agent for preventing and treating tobacco bacterial wilt according to claim 1, characterized in that: The mass ratio of ε-polylysine to the biocontrol bacteria fermentation liquid is 1:

3.

3. The composite agent for preventing and treating tobacco bacterial wilt according to claim 1, characterized in that: The biocontrol bacteria fermentation liquid is prepared by mixing the fermentation liquid of Brevibacillus laterosporus and the fermentation liquid of Rheinheimerella aquatica in a mass ratio of 0.8-1.1:1.9-2.

3.

4. The composite agent for preventing and treating tobacco bacterial wilt according to claim 3, characterized in that: The mass ratio of the fermentation broth of Brevibacillus laterosporus to the fermentation broth of Rheinheimerella aquatica is 1:

2.

5. The composite agent for preventing and treating tobacco bacterial wilt according to claim 3, characterized in that: The Brevibacillus laterosporus fermentation liquid and the Rheinheimerella water fermentation liquid are obtained by the following preparation method: (1) Inoculate Brevibacillus laterosporus or Rheinheimerella aquatica into LB solid medium for activation; (2) The activated Brevibacillus laterosporus and Rheinheimerella water were inoculated into LB liquid culture medium respectively, and cultured at 30°C and 160 r / min for 36 h to obtain seed solution; (3) The seed solution was transferred to new LB liquid culture medium, and cultured at 37°C and 200 r / min for 48 h to obtain the fermentation broth of Brevibacillus laterosporus and Rheinheimerella aquatica.

6. The composite agent for preventing and treating tobacco bacterial wilt according to claim 5, characterized in that: In step (3), the number of viable bacteria in the fermentation broth of Brevibacillus laterosporus is greater than 1×10 8 cfu / mL; the number of viable bacteria in water Rheinheimerella fermentation broth is greater than 3×10 7 cfu / mL.

7. Use of the composite agent for preventing and treating tobacco bacterial wilt according to any one of claims 1 to 6 in preventing and treating tobacco bacterial wilt of cigar tobacco.

8. The use according to claim 7, characterized in that: The biological agent was applied on the 9th to 12th day after transplanting cigar tobacco, and then applied again after an interval of 14 to 16 days.

9. The use according to claim 8, characterized in that: The application amount is 80ml / mu each time.

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