Bacterial fertilizer containing Bacillus velez and application thereof

Abstract

The present invention provides a bacterial fertilizer containing Bacillus velezensis, and the Bacillus velezensis is named Bacillus velezensis QN3NO‑3, registered and preserved in the China Center for Type Culture Collection, and the preservation number is CCTCC NO: M 2021304. The Bacillus velezensis of the present invention has a broad-spectrum antibacterial activity, has a good antagonistic effect on tomato wilt pathogens, etc., and the strain has the ability of efficient nitrogen fixation and IAA production, promotes plant growth, and also has the activity of producing cellulase, chitinase and β‑1,3‑glucanase. The present invention prepares the Bacillus velezensis into a bacterial fertilizer, which can give full play to the antibacterial and nitrogen fixation abilities and other biological characteristics of the Bacillus velezensis, can reduce the dependence on pesticides and fertilizers, improve the yield and quality of crops, and reduce the negative impact on the environment at the same time, and has a wide range of application potentials in the fields of agriculture and the environment.

Description

Technical Field

[0001] The invention relates to the field of fertilizers, and in particular to a bacterial fertilizer containing Bacillus velezinus and application thereof. Background Art

[0002] Nitrogen (N) in NPK fertilizer is one of the most important nutrients in agricultural production. It is a basic chemical element necessary for the life activities of plants and animals. However, there is a widespread problem of nitrogen deficiency in farmland soil. Increasing the nitrogen content in organic fertilizers and chemical fertilizers to improve the productivity of the ecosystem is an effective way to achieve increased production benefits. However, excessive nitrogen application will lead to environmental pollution, which will not only reduce the utilization efficiency of nitrogen, but also have a negative impact on the quality of water and atmospheric environment, and pose a threat to the balance of animal and plant ecosystems and human health. China is one of the countries with the largest consumption of fertilizer nitrogen in the world. Although the large-scale use of nitrogen fertilizers has promoted the development of my country's agriculture, it has also caused problems such as eutrophication of water bodies. With the growth of population and the increase in demand for agricultural products, the consumption of fertilizer nitrogen will continue to increase. Therefore, improving the utilization efficiency of nitrogen, avoiding excessive use of fertilizer nitrogen, preventing adverse effects on environmental quality (such as soil compaction, etc.), and taking into account economic and environmental benefits have become the basic principles and important tasks of agricultural nitrogen management. While ensuring crop yields, we should actively explore ways to cultivate soil nitrogen fertility to achieve sustainable agricultural development.

[0003] Microorganisms are the oldest organisms on Earth, with a history of at least 4.1 billion years. They exist in soil, water, air, and in plants and animals in different forms, physiological characteristics, and metabolic capabilities. The diversity of Earth's microorganisms, such as bacteria, fungi, and viruses, has enriched the microbial resource library and provided valuable resources for scientific research and application in the fields of agriculture, medicine, and the environment. Nitrogen-fixing bacteria, as a special type of bacteria, play an important role in Earth's microorganisms. They can convert nitrogen (N2) in the atmosphere into nitrogen compounds that can be used by plants, such as ammonia (NH3) and nitrates (NO3 -), which is crucial to the nitrogen cycle of the earth's ecosystem. Nitrogen is an important element for organisms to synthesize essential substances such as proteins and nucleic acids, but most organisms cannot directly use nitrogen in the atmosphere. Nitrogen-fixing bacteria convert nitrogen into a form that can be used by other organisms through nitrogen fixation, thereby promoting the healthy development of biodiversity and ecosystems. Nitrogen-fixing bacteria are widely present in environments such as soil, water bodies and plant rhizosphere, and form a symbiotic relationship with plants. For example, rhizobia of legumes combine with the roots of legumes to promote nitrogen fixation and plant growth. In addition, nitrogen-fixing bacteria can also live together with other microorganisms to form complex microbial communities and participate in key ecological processes such as soil organic matter decomposition and nutrient cycling. Research on the classification, physiological characteristics, nitrogen fixation mechanism and interaction with plants and other microorganisms of nitrogen-fixing bacteria will help reveal the nitrogen cycle mechanism of the earth's ecosystem, improve agricultural production efficiency and promote sustainable development. However, the research on nitrogen-fixing bacteria still faces some challenges, such as the exploration and rational use of nitrogen-fixing bacteria resources, the development and utilization of nitrogen-fixing bacteria, the improvement of nitrogen fixation efficiency and the influence of environmental factors on nitrogen fixation. Therefore, further research still requires continuous efforts to give full play to the important role of nitrogen-fixing bacteria in the earth's microorganisms.

[0004] Endophytes are considered to be microorganisms that live in healthy plants at least at a certain stage or even throughout their life cycle without causing obvious diseases in the plants. Since K1eopper et al. first mentioned them in 1992, they have attracted people's attention. Their diversity determines their wide application in the fields of biological control, medicine, forestry, etc. Under the dual impact of the strengthening of the concept of sustainable agricultural development and the difficulty of chemical control, microorganisms have gained more and more research and development space due to their advantages such as wide sources, large quantities, and low costs, including improving soil fertility, repairing soil, increasing crop yield and quality, and improving crop stress resistance. After entering the soil, microorganisms with antagonistic functions establish a competition mechanism with plant pathogens, seize ecological sites, compete for nutrients and oxygen, etc., or inhibit pathogens by parasitism and dissolution, induce crop resistance, and reduce the incidence rate. Previous reports have shown that the metabolites produced by antagonistic strains during metabolism can improve the disease resistance of plants, or destroy the cell structure of pathogens, causing the hyphae to fission and twist, and lose the ability to infect. Zeng Yibo studied the growth-promoting and disease-resistant functions of photosynthetic bacteria PSB06 and found that the extracellular proteins produced by the strain metabolism can effectively inhibit rice blast. Anming et al. isolated a biological control strain Y-1 resistant to the genus Pseudomonas from the stems of Dendrobium nobile, and determined through proteomics that its polypeptide metabolites inhibited the energy production and amino acid biosynthesis in the respiratory chain of pathogens. As a biological pesticide, Bacillus velez has certain advantages in controlling crop diseases. It can produce a series of antibacterial substances, such as antibiotics, enzymes and antimicrobial peptides, which have inhibitory effects on a variety of plant pathogens. Exploring and developing Bacillus velez biological pesticide products can effectively reduce the use of chemical pesticides, reduce environmental pollution and harm to human health, and improve crop yield and quality. However, for specific applications, it is necessary to select appropriate Bacillus velez strains and application methods according to different crops and diseases. Summary of the invention

[0005] The purpose of the present invention is to overcome the deficiencies in the prior art and provide a bacterial fertilizer containing Bacillus velezensis and application thereof.

[0006] The first aspect of the present invention is to provide an application of Bacillus velezensis or its fermentation liquid in promoting plant growth; the Bacillus velezensis is named Bacillus velezensis QN3NO-3, registered and preserved in the China Center for Type Culture Collection, with a preservation number of CCTCC NO: M 2021304.

[0007] Preferably, the Bacillus Velezii promotes plant growth by fixing nitrogen and / or secreting IAA.

[0008] The second aspect of the present invention is to provide an application of Bacillus velezensis or its fermented liquid in promoting seed germination, and / or promoting seeds to enter the whitening stage in advance, and / or promoting embryo growth, and / or promoting radicle growth, and / or increasing the fresh weight of the above-ground part of the plant, and / or increasing the fresh weight of the underground part of the plant, and / or promoting the growth of the root crown of the plant, and / or promoting the growth of the root system of the plant, and / or increasing the number of lateral roots of the plant; the Bacillus velezensis is named Bacillus velezensis QN3NO-3, registered and preserved in the China Center for Type Culture Collection, and the preservation number is CCTCC NO: M 2021304.

[0009] The third aspect of the present invention is to provide an application of Bacillus velezensis or its fermentation liquid in improving plant drought resistance; the Bacillus velezensis is named Bacillus velezensis QN3NO-3, registered and preserved in the China Center for Type Culture Collection, with a preservation number of CCTCC NO: M 2021304.

[0010] The fourth aspect of the present invention is to provide a bacterial fertilizer containing Bacillus velezensis, wherein the Bacillus velezensis is named Bacillus velezensis QN3NO-3 (hereinafter also referred to as Bacillus velezensis QN3NO-3), which is registered and preserved in the China Center for Type Culture Collection with a preservation number of CCTCC NO: M 2021304.

[0011] The "microbial fertilizer" in the present invention refers to fertilizers added with microorganisms, which can improve the nutritional conditions of crops through the life activities of microorganisms, and can also add other nutritional elements, such as nitrogen fertilizer, potassium fertilizer, phosphorus fertilizer, etc.

[0012] Preferably, the bacterial fertilizer also includes other fertilizer nutrient elements.

[0013] The bacterial fertilizer of the present invention can also be added with auxiliary materials acceptable to fertilizers according to actual use needs, such as adding carrier matrix (rice husk charcoal, etc.) to make solid fertilizer, and adding solvents to make liquid fertilizer.

[0014] Preferably, the fertilizer further comprises auxiliary materials acceptable to fertilizers.

[0015] The Velez's bacillus QN3NO-3 of the present invention has a nitrogen fixation effect and can also secrete IAA, and can promote plant growth by nitrogen fixation and / or secretion of IAA. Therefore, the bacterial fertilizer containing the Velez's bacillus of the present invention can also promote plant growth. Therefore, the fifth aspect of the present invention is to provide the use of the bacterial fertilizer as described in the fourth aspect of the present invention in promoting plant growth.

[0016] The Velez bacillus QN3NO-3 of the present invention can promote seed germination, promote seeds to enter the whitening period in advance, promote the growth of plumules and radicles, increase the fresh weight of the aboveground and underground parts of plants, promote the growth of the root crown root system of plants, and increase the number of lateral roots of plants. Therefore, the bacterial fertilizer containing the Velez bacillus of the present invention can also promote seed germination, promote seeds to enter the whitening period in advance, promote the growth of plumules and radicles, increase the fresh weight of the aboveground and underground parts of plants, promote the growth of the root crown root system of plants, and increase the number of lateral roots of plants. Therefore, the sixth aspect of the present invention is to provide the bacterial fertilizer as described in the fourth aspect of the present invention in promoting seed germination, and / or promoting seeds to enter the whitening period in advance, and / or promoting plumule growth, and / or promoting radicle growth, and / or increasing the fresh weight of the aboveground parts of plants, and / or increasing the fresh weight of the underground parts of plants, and / or promoting the growth of the root crown of plants, and / or promoting the growth of the root system of plants, and / or increasing the number of lateral roots of plants.

[0017] Lateral roots play an important role in plant drought resistance. The Velez's bacillus QN3NO-3 of the present invention can increase the number of plant lateral roots and improve the drought resistance of plants. Therefore, the bacterial fertilizer containing the Velez's bacillus of the present invention can also improve the drought resistance of plants. Therefore, the seventh aspect of the present invention is to provide the use of the bacterial fertilizer described in the fourth aspect of the present invention in improving plant drought resistance.

[0018] The Velez's bacillus QN3NO-3 of the invention has a broad-spectrum antibacterial property, and has a good inhibitory effect on tomato wilt pathogen, banana wilt pathogen No. 4 physiological subspecies, banana wilt pathogen No. 1 physiological subspecies, cucumber wilt pathogen, pepper anthracnose pathogen, strawberry anthracnose pathogen, litchi anthracnose pathogen, banana anthracnose pathogen, vegetable anthracnose pathogen, gloeosporium anthracnose pathogen, corn Curvularia leaf spot pathogen, corn Alternaria leaf blight pathogen, wheat fusarium scab pathogen, wheat fusarium scab pathogen, mango polydiscus leaf blight pathogen, rice blast pathogen, mango anthracnose pathogen, etc. Therefore, the bacterial fertilizer containing the Velez's bacillus of the invention also has an inhibitory effect on the above-mentioned pathogens.

[0019] Therefore, the eighth aspect of the present invention is to provide the use of the bacterial fertilizer described in the fourth aspect of the present invention in antagonizing tomato wilt pathogen, and / or banana wilt pathogen No. 4 physiological subspecies, and / or banana wilt pathogen No. 1 physiological subspecies, and / or cucumber wilt pathogen, and / or pepper anthracnose pathogen, and / or strawberry anthracnose pathogen, and / or litchi anthracnose pathogen, and / or banana anthracnose pathogen, and / or vegetable anthracnose pathogen, and / or gloeosporium anthracnose pathogen, and / or corn Curvularia leaf spot pathogen, and / or corn Alternaria leaf blight pathogen, and / or wheat fusarium scab pathogen, and / or wheat fusarium scab pathogen, and / or mango polytrichosporium leaf blight pathogen, and / or rice blast pathogen, and / or mango anthracnose pathogen.

[0020] The ninth aspect of the present invention is to provide the use of the bacterial fertilizer described in the fourth aspect of the present invention in preventing and controlling diseases caused by tomato wilt pathogen, and / or banana wilt pathogen race 4, and / or banana wilt pathogen race 1, and / or cucumber wilt pathogen, and / or pepper anthracnose pathogen, and / or strawberry anthracnose pathogen, and / or litchi anthracnose pathogen, and / or banana anthracnose pathogen, and / or vegetable anthracnose pathogen, and / or gloeosporium anthracnose pathogen, and / or corn Curvularia leaf spot pathogen, and / or corn Alternaria leaf blight pathogen, and / or wheat fusarium sphaeroides pathogen, and / or wheat fusarium sphaeroides pathogen, and / or mango polytrichosporium leaf blight pathogen, and / or rice blast pathogen, and / or mango anthracnose pathogen.

[0021] The eleventh aspect of the present invention is to provide a method for promoting plant growth, which comprises applying the bacterial fertilizer as described in the fifth aspect of the present invention to the plants.

[0022] The Bacillus Velezii of the invention has a broad-spectrum antibacterial activity, and has a good inhibitory effect on tomato wilt pathogen, banana wilt pathogen No. 4 physiological subspecies, banana wilt pathogen No. 1 physiological subspecies, cucumber wilt pathogen, pepper anthracnose pathogen, strawberry anthracnose pathogen, litchi anthracnose pathogen, banana anthracnose pathogen, Chinese cabbage anthracnose pathogen, gloeosporium anthracnose pathogen, corn Curvularia leaf spot pathogen, corn Alternaria leaf blight pathogen, wheat fusarium scab pathogen, wheat fusarium scab pathogen, mango polydiscus leaf blight pathogen, rice blast pathogen, mango anthracnose pathogen, etc., and the strain has the ability of efficient nitrogen fixation and IAA production, and can promote the growth of plants, for example, can effectively promote seed germination, promote seeds to enter the dew white stage in advance, promote the growth of plumules and radicles, increase the fresh weight of plants, promote the growth of plant root crowns and root systems, and the number of plant lateral roots, and can also improve the drought resistance of plants. The strain also has the activity of producing cellulase (glucanase, exoglucanase, β-glucosidase, filter paper cellulase, etc.), chitinase and β-1,3-glucanase, and can efficiently degrade cellulose and destroy fungal cell walls.

[0023] The invention prepares the Bacillus Velezii into bacterial fertilizer, which can give full play to the antibacterial and nitrogen-fixing abilities and other biological characteristics of the Bacillus Velezii, reduce the dependence on pesticides and chemical fertilizers, improve the yield and quality of crops, and reduce the negative impact on the environment, and has wide application potential in the fields of agriculture and environment. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 These are the scanning electron microscopy and Gram staining results of QN3NO-3 bacteria.

[0025] Figure 2 Some physiological and biochemical experiments on strain QN3NO-3.

[0026] Figure 3These are the results of the IAA production capacity test of strain QN3NO-3.

[0027] Figure 4 is the standard curve of IAA content.

[0028] Figure 5 This is the result of the cellulose decomposition ability test of strain QN3NO-3 (Image J screenshot).

[0029] Figure 6 To determine the cellulase activity in the fermentation broth of strain QN3NO-3.

[0030] Figure 7 The germination of corn seeds after being treated with strain QN3NO-3. Left: 1-3d germination; right: 3rd day germination.

[0031] Figure 8 The antibacterial activity of volatiles of strains was determined by the double-dish method.

[0032] Fig. 9 This study was conducted to determine the antagonism of the fermentation supernatant of strain QN3NO-3 and ammonium sulfate crude protein against tomato wilt pathogen.

[0033] Fig.10 The chitinase and β-1,3-glucanase activities of the fermentation supernatant of strain QN3NO-3 were determined.

[0034] Fig.11 These are the results of a study on the broad-spectrum antibacterial activity of strain QN3NO-3. DETAILED DESCRIPTION

[0035] The present invention will be further described below with reference to the accompanying drawings in conjunction with specific embodiments to better understand the present invention. Where specific techniques or conditions are not indicated in the embodiments, the techniques or conditions described in the literature in this area or the product specifications are used. Where the manufacturer of the reagents or instruments used is not indicated, they are all conventional products that can be obtained commercially.

[0036] 1 Materials

[0037] 1.1 Sample source and processing

[0038] The fruits of live Noni plants in the Noni plantation in Qiaotou Town, Chengmai County, Hainan Province (N 19°58′9″, E 109°55′19″) were sampled at multiple points, bagged, numbered, and stored at 4°C.

[0039] 1.2 Culture medium

[0040] LB medium: 5 g yeast powder, 10 g tryptone, 10 g sodium chloride, 20 g agar, 1 L distilled water, pH 7.2-7.5;

[0041] Ashby medium: glucose 10g, potassium dihydrogen phosphate 0.2g, sodium chloride 0.2g, magnesium sulfate 0.2g, calcium sulfate 0.1g, calcium carbonate 5g, agar 20g, distilled water 1L, pH 7.0-7.2.

[0042] Cellulose Congo red medium: microcrystalline cellulose 1.88g, Congo red 1g, gelatin 2.0g, agar 20g, distilled water 1L, pH 7.0.

[0043] 1.3 Main reagents:

[0044] DNS reagent: weigh 10g of 3,5-dinitrosalicylic acid and dissolve it in 500ml of distilled water, add 20g of sodium hydroxide and 200g of potassium sodium tartrate, heat the solution to dissolve, add 2g of phenol and 0.5g of anhydrous sodium sulfite, cool to room temperature after the solute is dissolved, and make up to 1000ml. The reagent is stored in a brown volumetric flask and filtered after one week.

[0045] CMC substrate solution: weigh 0.625g of carboxymethyl cellulose sodium salt and dissolve it in 100ml of sodium acetate buffer solution, heat and stir until dissolved;

[0046] Microcrystalline cellulose substrate solution: weigh 0.5 g of microcrystalline cellulose and dissolve it in 100 ml of sodium acetate buffer solution;

[0047] Filter paper substrate solution: weigh 0.5 g of filter paper and soak it in 100 ml of sodium acetate buffer solution;

[0048] Standard glucose solution: Dissolve 27.0g glucose in 25ml distilled water to prepare 6mol / L glucose solution. Prepare the glucose solution standard curve according to Table 1;

[0049] 1.10 mol / L sodium hydroxide: weigh 400 g of analytical grade sodium hydroxide and dissolve it in 500 ml of distilled water, then adjust the volume to 1 L;

[0050] 2% boric acid-indicator solution: weigh 20g of boric acid solution into 1000mL of distilled water, then add 5mL of methyl red-bromocresol green mixed indicator, and adjust to slightly purple-red with dilute acid or dilute alkali. The reagent should be prepared and used immediately;

[0051] Highly efficient Kjeldahl catalyst tablets.

[0052] 1.4 Test crops:

[0053] The test material is corn, the variety is Meiyu No. 16, a sweet and glutinous variety provided by Hainan Lvchuan Seedlings Co., Ltd.

[0054] 1.5 Data processing

[0055] Image J (Version 1.38) software was used to calculate the gray area. Microsoft Excel 2007 software and one-way ANOVA were used to analyze the experimental data. Duncan's new multiple range test (DMRT) was used for significant difference analysis.

[0056] 2 Experimental methods and results

[0057] 2.1 Isolation and screening of endophytic bacteria

[0058] Sample disinfection: Wash and air-dry the noni fruit for later use. Sterilize with 75% ethanol for 5 minutes, then treat with sodium hypochlorite in the dark for 20 minutes, soak in 10% sodium bicarbonate for 10 minutes, rinse with sterile water 5 times, dry on sterile filter paper and mark.

[0059] Sample blotting experiment: The sample was placed in LB medium for 5 minutes using the tissue blotting method and cultured at 28°C for 3 days as a sterility test.

[0060] Sample grinding: After slicing the samples, grind them in a sterile mortar until they become a paste. Take 1 mL of the juice in a centrifuge tube, add 4 mL of LB medium, and culture at room temperature at 180 r / min for 1 h with vortexing.

[0061] Separation and screening: gradient dilution (10 -1 , 10 -2 , 10 -3 ) to make a suspension, after sufficient shaking, take 200 μL of the dilution and evenly inoculate it on Ashurst medium by plate spreading method, repeat 3 times, and culture it inverted in a dark box at 28°C for 5 days. Select different colonies with better morphology, better growth and faster growth rate, and purify them multiple times on new solid Ashurst medium by inoculation loop plate streak method until pure culture.

[0062] In this study, the noni fruit samples were disinfected and made into suspensions. 71 endophytic bacteria were initially isolated and purified by the dilution plate method. Five dominant bacteria were selected for nitrogen fixation and growth promotion function rescreening (refer to "2.2 Identification of nitrogen fixation ability and activity determination" for nitrogen fixation function rescreening, and refer to "2.6 Effect of strains on the biological activity of corn growth" for growth promotion function rescreening). The best strain was selected as the research object and named QN3NO-3. The colony was irregularly round, yellow-gray, opaque, smooth and wrinkled on the LB medium, with wavy edges and a central bulge. The bacteria were oval or short rod-shaped, and Gram-stained purple-red, belonging to positive bacteria ( Figure 1 ). As the culture time increases, the colonies tend to become dry and flat, with irregular wrinkles on the edges.

[0063] 2.2 Physiological and biochemical identification of strains

[0064] The antagonistic strains screened out were subjected to physiological and biochemical identification according to Bergey's Bacterial Identification Manual and Common Bacterial System Identification Manual.

[0065] (1) Single carbon source utilization experiment: In the physiological and biochemical identification of bacteria, the utilization of carbon sources by strains is an important indicator. Different bacteria have different utilization of carbon sources. The single carbon source utilization experiment is to add a single carbon source at a concentration of 1% to the basic medium of Pugo, and then inoculate the strain to be identified, culture it at 28°C for one to two weeks, and inoculate the strain on the basic medium without any carbon source as a blank control, and observe the growth of the strain on each culture medium. If it can grow, it is positive, indicating that the strain has the ability to utilize the carbon source. If it cannot grow, it is negative, indicating that it has no ability to utilize the carbon source. Carbon sources include: α-lactose, D-fructose, D-galactose, D-mannitol, glucose, mannitol, anhydrous lactose, soluble starch, trehalose, sorbitol, inositol, melezitose, rhamnose, xylose, maltose, sucrose, melibiose, etc.

[0066] (2) Single nitrogen source utilization experiment: Just like the different carbon source utilization, different bacteria also have different utilization of nitrogen sources. A single nitrogen source is added to the nitrogen source basal culture medium at a concentration of 0.5%, and a plate is made. Then the strain is inoculated. In addition, the strain is inoculated on the basal culture medium without any nitrogen source as a blank control. The strain is cultured at 28°C for one to two weeks and the growth of the strain is observed. If it can grow, it is positive, indicating that the strain has the ability to utilize the nitrogen source; if it cannot grow, it is negative, indicating that it does not have the ability to utilize the nitrogen source. The added nitrogen sources include: trypsin, asparagine, valine, histidine, oxalic acid, ammonium sulfate, ammonium acetate, ammonium chloride, ammonium molybdate tetrahydrate, ammonium nitrate, phenylalanine, glycine, cysteine, antamine, arginine, serine, tryptophan, methionine and tyrosine.

[0067] (3) Starch hydrolysis test: Nutrient agar medium is used as the basic medium, and 1% soluble starch is added to it. The antagonistic strain is inoculated onto the prepared plate using the point-inoculation method (i.e., inoculation diameter < 0.5 cm). After culturing at 28°C for one week, iodine solution is added around the colony. If there is a transparent circle around the colony, the result is positive, and the strain can produce amylase, and the diameter of the transparent circle indicates the activity of the produced amylase; if no amylase is produced, the entire plate will appear blue.

[0068] (4) Gelatin liquefaction test: The strain to be identified is inoculated into a test tube containing gelatin medium, then cultured at 28°C, and the liquefaction of gelatin is observed on the 5th, 10th, 20th, and 30th days. If liquefaction occurs, it is positive, indicating that the strain has the ability to liquefy gelatin, otherwise it is negative.

[0069] (5) Cellulose decomposition experiment: Prepare cellulose decomposition medium according to the recipe, then immerse a section of filter paper strip in the liquid medium, sterilize the medium, inoculate the strain into the medium, and culture it statically. After one month, observe whether the filter paper strip is decomposed. If it is decomposed, it is positive, indicating that cellulose decomposition enzymes are produced. Otherwise, it is negative.

[0070] (6) Nitrate reduction: Prepare nitrate reduction medium according to the formula, inoculate the strain into the medium, and culture at 28°C for 7 days and 14 days. Use the medium without the strain as a blank control. Take a blank test tube, add a small amount of the 7d and 14d culture solution to different test tubes, and add the previously prepared A solution and B solution respectively. If the solution appears pink, rose red, brown or orange, nitrate reduction is positive; if none of the above colors appear, add 1 or 2 drops of diphenylamine reagent, and the solution turns blue, it is still negative. Otherwise, it is treated as positive.

[0071] (7) Urease test: Prepare urease medium according to the formula, inoculate the strain to be tested into the urease medium, incubate it upside down at 28°C for 4 days, and observe the color change of the medium. If the strain to be tested has the ability to produce urease, the medium will turn pink, which is positive; if it does not change color, it is negative.

[0072] (8) Lipase (Tween 20, 40, 80) experiment: After preparing the lipase culture medium, sterilize the separately packaged Tween 20, 40, and 80 together, mix the Tween with the culture medium and make a plate. Inoculate the strain on the plate, culture for one to two weeks, and observe the plate. If a fuzzy halo is formed around the colony, it is positive, otherwise it is negative.

[0073] (9) Salt tolerance experiment: Prepare culture media with different NaCl concentrations (1%, 3%, 5%, 7%, 9%, 11%, 13%, 15%). The other components of the culture media are the same. The strain to be identified is inoculated on these culture media and cultured at 28°C. The culture is observed once a week for four weeks. Finally, it is recorded whether the strain grows on the culture medium to determine the upper and lower limits of the strain's tolerance to NaCl.

[0074] (10) pH tolerance test: Adjust the pH value of the liquid culture medium to 4, 5, 6, 7, 8, 9, and 10 respectively, and inoculate the strain to be tested into culture medium with different pH values ​​respectively, ensuring that other culture conditions are consistent, and culture at 28°C with shaking. Observe once a week for four weeks, and finally determine the growth of the strain on each culture medium. Determine the upper and lower limits of the pH value at which the strain can grow and the optimal pH value.

[0075] The results of the experiment showed that QN3NO-3 showed positive reactions in starch hydrolysis, gelatin liquefaction, urease test, and enzymatic test; it was only sensitive to Tween 40 in the enzyme lipid test; it had no reaction and was negative in the methyl red, malonic acid, phosphate reduction, and hydrogen sulfide tests. It can grow in pH 4-10, and the most suitable growth environment is pH 9; in the salt tolerance test, it can grow in NaCl concentrations of 1%-9%, but not in 11%-15%.

[0076] According to the results of the carbon and nitrogen source utilization experiment, QN3NO-3 can utilize carbon sources such as α-lactose, D-fructose, D-galactose, D-mannitol, glucose, mannitol, anhydrous lactose, soluble starch, trehalose, sorbitol, inositol, melezitose, rhamnose, xylose, maltose, sucrose, and melibiose; it can utilize fewer nitrogen sources, such as tryptone, asparagine, valine, histidine, and oxamic acid, but cannot utilize ammonium sulfate, ammonium acetate, ammonium chloride, ammonium molybdate tetrahydrate, ammonium nitrate, phenylalanine, glycine, cysteine, antamine, arginine, serine, tryptophan, methionine, and tyrosine.

[0077] Based on the morphological, cultural and physiological and biochemical characteristics, the bacterium was preliminarily identified as Bacillus velezensis QN3NO-3 (China Center for Type Culture Collection. Wuhan, CCTCC NO: M 2021304, March 30, 2021).

[0078] Table 1 Analysis of physiological and biochemical characteristics of QN3NO-3 strain

[0079]

[0080]

[0081] 2.3 Identification of nitrogen fixation ability and activity determination

[0082] Use a sterilized toothpick to pick the strain to be tested into Ashby nitrogen-free solid medium and subculture for 6 generations. If it can grow well, it means it has the ability to fix nitrogen, and if it cannot grow, it means it does not have the ability to fix nitrogen.

[0083] The nitrogen fixation activity of the strain was determined by the Kjeldahl method. The dominant nitrogen-fixing bacteria were picked with an inoculation loop and fermented in LB culture medium for 2 days. The fermentation liquid was centrifuged at 10,000 rpm for 10 minutes at 4°C to remove the supernatant. The bacteria were placed in a mortar and quickly ground with liquid nitrogen. 1 g was weighed and placed at the bottom of a dry digestion tube. A small amount of water was added to moisten it, and 5 mL of concentrated sulfuric acid and 2 high-efficiency Kjeldahl nitrogen determination catalyst tablets were added. The digestion tube was digested, and when the solution in the digestion tube turned grayish white with a hint of green, it was continued to be digested for a period of time. The digestion tube was removed. The nitrogen content in each digested culture solution was measured using a fully automatic Kjeldahl nitrogen analyzer.

[0084] The nitrogen content in the bacterial suspension was determined using a Kjeltec 8200 Kjeldahl nitrogen analyzer. Under the action of a catalyst, the sample was digested with concentrated sulfuric acid to convert all organic nitrogen into inorganic ammonium salts, and then the ammonium salts were converted into ammonia under alkaline conditions, distilled out with water vapor and absorbed by excess boric acid solution. It was then titrated with standard hydrochloric acid, and the nitrogen content in the sample was calculated to be 152.75 mg / L, and the protein content was 954.69 mg (protein content = nitrogen content / 16%), indicating that the strain has a high-efficiency nitrogen fixation ability.

[0085] 2.4 IAA production capacity identification and activity determination

[0086] Prepare liquid LB containing 100 mg / L α-aminoindolylpropionic acid, inoculate the strain to be tested, set up 3 replicates, and culture at 180 r / min at room temperature for 2 days. Pipette 50 μL of the cultured bacterial suspension onto a white droplet plate, and set up 2 groups of controls: 50 μL of uninoculated LB as a negative control, 50 μL of 50 μg / mL IAA solution as a positive control, and add 50 μL of Salkowski colorimetric solution to all three at the same time. Let stand at room temperature away from light for 15 min. If there is no color change, it is negative and does not produce IAA; if it turns pink, it is an IAA-producing bacterium and the IAA content can be determined.

[0087] Prepare IAA stock solution (100 μg / mL), and dilute IAA solutions with concentrations of 0, 10, 20, 30, 40, 50, and 60 μg / mL in 25 mL constant volume bottles, and measure OD using a UV spectrophotometer. 530nm , draw the IAA standard curve. Pipette the bacterial suspension into a centrifuge tube and place it in a centrifuge with an initial speed of 12000r / min for 10min, centrifuge and discard the precipitate, take 4mL of the supernatant and mix it with an equal volume of Salkowski colorimetric solution, and use the blank control without adding the supernatant. The determination method is the same as above, and the IAA yield of the strain is calculated according to the IAA standard curve.

[0088] According to the Salkowski colorimetric method ( Figure 3 ), the mixture of strain QN3NO-3 and the IAA positive control group was pink, while the uninoculated negative control group had no color change, indicating that strain QN3NO-3 can secrete IAA. The results of the ultraviolet spectrophotometer were used to draw the IAA ( Figure 4 ), the amount of IAA secreted by QN3NO-3 in LB medium containing L-tryptophan was calculated to be 40.61±0.43μg / mL.

[0089] 2.5 Identification of cellulose decomposition ability and determination of cellulase activity

[0090] Use an inoculation loop to streak the bacterial sample on the cellulose Congo red medium. After culturing at 28°C for 2-3 days, observe whether a transparent circle is produced around the colony in the medium after the colony grows. Set three replicates. The production of a transparent circle indicates that the strain can produce cellulase. The cellulase activity can be preliminarily judged based on the size of the transparent circle. After taking pictures, use Image J software to measure the gray area (pix) of the irregular transparent circle on the plate and the colony area (ar), calculate the ratio of the two, and calculate the AR / ar ratio. The size of the ratio can preliminarily reflect the activity of the strain in producing cellulase. Excel 2007 and variance analysis software were used for experimental data conversion and analysis.

[0091] Calculate the "ratio" formula:

[0092] Ratio (AR / ar) = colony area (pix) / transparent zone area (pix).

[0093] According to the results of plate streaking observation ( Figure 5 ), a large area of ​​transparent circles was produced on the cellulose Congo red medium, indicating that QN3NO-3 can produce cellulase. Using Image J (Version 1.38) software, the average irregular transparent circle area (AR) on the plate was 12690.33 pix, and the average colony area (ar) was 823.33 pix, and the ratio AR / ar was 15.41. It can be seen that the strain has strong cellulase production activity and can be used for identification and subsequent experiments.

[0094] The bacterial suspension cultured for 2 days (10 8 CFU / mL) was inoculated into a 250mL flask containing 100mL CMC liquid culture medium and cultured at 28°C with shaking at 150r / min for 2 days. The fermentation broth was centrifuged at 10000rmp for 5 minutes at 4°C, and the supernatant was used as an enzyme extract. The activities of exoglucanase, endoglucanase and β-glucosidase were determined by measuring the reducing sugar release of microcrystalline cellulose (MCC), carboxymethyl cellulose (CMC) and salicylic acid (SAL). Total cellulase activity was expressed as filter paper cellulase activity (FPA) and was determined by the filter paper analysis method described by Ghose (1987) using Whatman No. 1 filter paper as the substrate. Cellulase activity was determined according to the dinitrosalicylic acid (DNS) method at 540nm, and monosaccharide analysis was performed. Different concentrations of D-glucose solutions were used as calibration curves. All samples were measured at 540nm using a UV-visible spectrophotometer. The unit (U) of enzyme activity was defined as the amount of enzyme that released 1 μmol (2 μmol for β-glucosidase) of reducing sugar (in terms of glucose) per ml of fermentation supernatant per minute.

[0095] The results of cellulase activity assay were as follows Figure 6As shown, after 2 days of cultivation, the exoglucanase activity was 61.81 IU / mL, the endoglucanase activity was 47.11 IU / mL, the β-glucosidase activity was 53.31 IU / mL, and the filter paper cellulase activity was 29.47 IU / mL, indicating that the QN3NO-3 strain has good enzyme activities such as endoglucanase, exoglucanase, and β-glucosidase, indicating that the strain can efficiently degrade cellulose.

[0096] 2.6 Effects of the biological activity of strains on corn growth

[0097] The corn seeds were placed in a refrigerator at 4°C for vernalization for 3 days, taken out and placed in 10% (v / v) sodium hypochlorite for surface disinfection for 15 min, and then washed repeatedly with sterile water for 5 times. -1 ) for 60 minutes, then arrange and place in a culture dish with sterilized filter paper and culture at 28℃ in the dark for 3 days, wait for germination, and count the germination rate. Then transfer to a transparent incubator for 7 days, collect the whole seedlings, and slowly wash the filter paper attachments in clean water, record the length and fresh weight of the roots and aboveground parts, and calculate the seedling growth indicators (aboveground fresh weight g, underground fresh weight g, plumule length cm, radicle length cm, lateral root number pcs, root-crown ratio). The control group was the culture solution without inoculation of the strain, and each treatment was repeated 3 times, with 10 corn seeds each time.

[0098] Germination rate = (number of germinated seeds / total number of seeds) × 100%; root-to-shoot ratio = root fresh weight / aerial fresh weight.

[0099] Corn seeds were treated with bacterial suspension (10 -1 ) treatment, with the culture medium as the control. After 3 days of culture, the seed germination rates were 93.99% and 77.78% respectively. On the first day of culture, 93.99% of the seeds in the bacterial suspension treatment group had whitened, while only 66.67% in the control group. On the second day, the QN3NO-3 bacterial suspension promoted the growth of the radicle of corn seeds, and the embryo also expanded rapidly. 80% of the seeds in the culture medium treatment group whitened. There was a significant difference between the two treatment groups, indicating that the strain can make corn seeds enter the whitening period ahead of time, has good stability, and has a germination effect on corn ( Figure 7 ).

[0100] The quality of corn seedlings greatly affects the later growth and yield. Robust seedlings generally have stronger biological activity. After the corn seeds were inoculated with Bacillus Velez QN3NO-3 and grown for 7 days, the bioactivity indicators of corn seedlings were measured. The results showed that compared with the uninoculated control group, the corn seedlings inoculated with QN3NO-3 had a significant promoting effect on the length and fresh weight of the plumule and radicle, and also had a certain incremental effect on the number of lateral roots. The QN3NO-3 strain increased the aboveground and underground fresh weight of corn seedlings by 35.29% and 80% respectively compared with the uninoculated seedlings, and the root-crown ratio also increased from 0.60 to 0.80, significantly promoting the growth of the underground root system of corn seedlings, and then promoting the growth of the aboveground part. The plumule of the strain treatment group was 6.04cm, which was 2.22cm longer than that of the culture medium treatment; the average length of the radicle was 11.45cm, which was 1.56 times that of the culture medium group, significantly enhancing the functional activity of the root system. Lateral roots play an important role in the drought tolerance of corn. The number of lateral roots in the culture medium group at the seedling stage was 8.07, and after the strain treatment, it was 8.3. The above results show that QN3NO-3 can promote the growth and development of corn seedlings and play an important role in root growth.

[0101] 2.7 Determination of broad-spectrum antibacterial activity of strains by plate confrontation culture method

[0102] Test pathogens: tomato wilt pathogen F.oxysporum f.sp.lycopersic, banana wilt pathogen F.oxysporum f.sp.cubense Race 4 (ATCC 76255), banana wilt pathogen Fusarium oxysporum Schlecht.f.sp.cubense Race 1 (ATCC MYA-3244), cucumber wilt pathogen Fusarium oxysporum sp.Cucumebrium (ACCC 30220), pepper anthracnose pathogen Colletotrichum capsici (ATCC 96157), strawberry anthracnose pathogen Colletotrichum fragariae (ATCC 58718), litchi anthracnose pathogen Colletotrichum gloeosporioides (ATCC 58691), banana anthracnose pathogen Colletotrichum musae (ATCC 96167), and vegetable anthracnose pathogen Colletotrichum higginsianum (KACC 40807), Colletotrichum gloeosporioides (ATCC 58222), Curvularia ulina (ATCC 60937), Alternaria tenuissima (ATCC 26276), Fusarium graminearum Schwabe (ATCC MYA-4620), Cryphonectra aparaitica, Pestalotiopsis mangiferae (MTCC 3412), Magnaporthe grisea (ATCC 208987) and Colletotrichum gloeosporioides (ATCC MYA-4131).

[0103] Prepare a PDA culture medium plate, use a hole puncher to take out the pathogenic bacteria cake (Φ=5mm) that has been cultured for 5 days and has good growth, and place it in the center of the PDA plate. Draw a "cross" with the center of the plate as the center, and inoculate the test strain at 2.5cm from the center on the four sides of the "cross". Inoculate 4 places on each plate, set 3 replicates, and inoculate only the pathogenic bacteria as a control. After incubating at 28℃ for 5-7 days, calculate the inhibition rate, which is (control colony diameter-treated colony diameter) / control colony diameter×100%. The results are shown in Table 2 and Fig.11As shown, strain QN3NO-3 has a broad-spectrum resistance to 17 host crop / plant pathogens such as banana, tomato, mango, rubber, and rice.

[0104] Table 2 Results of the study on the broad-spectrum antibacterial activity of strains

[0105]

[0106] 2.8 Determination of antibacterial activity of volatiles from strains by double-dish method

[0107] The method for determining the antibacterial activity of volatile antibacterial substances from microorganisms uses the "double-dish-to-dish method". The QN3NO-3 strain was streaked on the LB medium and incubated in an incubator at 28°C for 2 days. Then, the tomato wilt pathogen and banana wilt pathogen No. 4 physiological subspecies cake (Φ=5mm) were inoculated on the center of the PDA plate, and then respectively inlaid with the strain QN3NO-3 grown for 2 days. The two plates were sealed with sealing film, and the plate inlaid with the empty LB culture dish was used as a control. Continue to incubate in an incubator at 28°C for 7 days. Repeat 3 times. It was found that the QN3NO-3 strain can release volatile substances to inhibit the growth of tomato wilt pathogens and banana wilt pathogens, and the diameter of the pathogens was measured using ImageJ software.

[0108] Inhibition rate = (control colony diameter - treated colony diameter) / control colony diameter × 100%

[0109] The inhibition rates were 49.22% and 42.16% respectively.

[0110] 2.9 Study on the supernatant and crude protein of the fermentation broth

[0111] (1) Antagonistic effect of QN3NO-3 fermentation supernatant and ammonium sulfate crude protein on tomato wilt pathogen

[0112] The bacterial suspension cultured for 2 days (10 8 CFU / mL) was inoculated into a 250 mL flask containing 100 mL CMC liquid culture medium and cultured at 28°C with shaking at 150 r / min for 2 days. The fermentation broth was centrifuged at 10,000 rmp for 5 minutes at 4°C, and the supernatant was the supernatant of QN3NO-3 fermentation broth, and the antibacterial rate was determined.

[0113] Using ammonium sulfate graded precipitation method, centrifuge 1L fermentation broth in batches at high speed in a centrifuge under the centrifugal conditions of 4°C, 12000r / min, 30min. After discarding the precipitate, slowly add the corresponding amount of ammonium sulfate to the supernatant in batches to a final concentration of 20%, 30%, 50%, 70%, and 80% solution, stir until clear, and obtain supernatants of different saturation concentrations and place them in a 4°C refrigerator for precipitation for 24h. After being divided into 50mL centrifuge tubes, place them in a centrifuge at 4°C, 12000r / min for 30min, and collect the precipitate. Add a small amount of 10mmol / L PBS buffer to the above precipitates in a beaker until dissolved, place them in a dialysis bag with the same concentration of PBS buffer and let them stand overnight at 4°C. The solution after dialysis and desalination is sterilized with a filter to obtain the fermentation broth crude protein solution, and the antibacterial rate is determined.

[0114] Determination of antibacterial rate: With sterile water as the control, the antibacterial activity of the supernatant of QN3NO-3 fermentation broth and the crude protein solution of the fermentation broth was determined by a plate punching antibacterial test. A hole was punched at 25 mm from the center of the PDA plate with a hole diameter of 7 mm, and 100 μL of each treatment solution was added to each hole. Each treatment was repeated 3 times, and pathogenic fungi (tomato wilt pathogen) were added to the center of the plate. After the control plate was cultured at room temperature and was full of fungi, the diameter of the pathogens of the QN3NO-3 fermentation broth treatment solution and the crude protein solution with different concentration gradients was measured using ImageJ software. The antibacterial rate = (control colony diameter - treatment colony diameter) / control colony diameter × 100%.

[0115] The results are as follows Fig. 9 As shown in the figure, the supernatant of QN3NO-3 can antagonize tomato wilt pathogen, with an inhibition rate of 49.11%. Different concentrations of ammonium sulfate crude protein also have a significant inhibitory effect on tomato wilt pathogen, with an inhibition rate of more than 46.39%.

[0116] (2) Determination of chitinase and β-1,3-glucanase activities in the supernatant of the fermentation broth

[0117] The QN3NO-3 strain was incubated in sterile LB liquid medium at 28°C and 180 rpm for 5 days, and the bacterial suspension was collected and centrifuged at 8000 rpm for 15 minutes. The supernatant was collected every day to measure the chitinase activity and β-1,3-glucanase activity.

[0118] The results are as follows Fig.10As shown. The cell wall components of pathogenic fungi are mainly composed of chitin and glucan. Experimental data show that Bacillus Velez QN3NO-3 can produce chitinase and β-1,3-glucanase at different culture periods. After incubation for 5 days, the activities of chitinase and β-1,3-glucanase were the highest, 19.79±0.52U / mL and 15.19±0.38U / g, respectively. Combined with the antibacterial experiment, it is shown that chitinase can weaken or destroy the structural integrity of the fungal cell wall by degrading chitin, directly inhibiting the growth and development of a variety of plant pathogenic fungi. In addition, β-1,3-glucanase hydrolyzes another key component of the fungal cell wall by degrading β-1,3-glucan. These two enzymes lead to osmotic imbalance, cell lysis and fungal growth inhibition. The activities of chitinase and β-1,3-glucanase provide a basis for the antibacterial mechanism of biological control agents. Understanding the existence and activity of these enzymes can further support the development and optimization of biological control strategies for plant pathogen management.

[0119] 2.10 Pesticide allergy test

[0120] The drug sensitivity test of QN3NO-3 strain was carried out by agar dilution method: agar dilution plates containing various fungicides (3 concentrations, 3 repetitions for each concentration) were prepared for QN3NO-3 streaking inoculation, and the growth of the strain was observed. The results are shown in Table 3. According to the results, QN3NO-3 strain can grow in 7 alternative broad-spectrum fungicides, including methyl thiophanate, zhongshengmycin, mancozeb, carbendazim, dichlorvos, metalaxyl, and benomyl. When the concentration of mancozeb is too high, the growth is inhibited and the growth is poor. When the concentration of carbendazim reaches 500%, the growth is excellent. This shows that QN3NO-3 strain has certain development potential as a microbial source fertilizer in the agricultural industry where pesticide residues are serious. It can replace fungicides to inhibit bacterial and fungal pathogens, and can also reproduce normally in farmland contaminated by pesticide residues to promote plant growth.

[0121] Table 3 Results of the study on the pesticide resistance of strains

[0122]

[0123]

[0124] Note: +: slight growth, ++: normal growth, +++: substantial growth, -: no growth.

[0125] The specific embodiments of the present invention are described in detail above, but they are only examples, and the present invention is not limited to the specific embodiments described above. For those skilled in the art, any equivalent modification and substitution of the practical is also within the scope of the present invention. Therefore, the equalization transformation and modification made without departing from the spirit and scope of the present invention should be included in the scope of the present invention.

Claims

1. Use of Bacillus velezensis or its fermentation liquid in promoting plant growth; the Bacillus velezensis is named Bacillus velezensis QN3NO-3, registered and preserved in China Center for Type Culture Collection, with the deposit number of CCTCCNO: M 2021304.

2. The use according to claim 1, characterized in that The Bacillus Velezii promotes plant growth by fixing nitrogen and / or secreting IAA.

3. Use of Bacillus Velez or its fermented liquid in promoting seed germination, and / or promoting seeds to enter the whitening stage in advance, and / or promoting embryo growth, and / or promoting radicle growth, and / or increasing the fresh weight of the above-ground part of the plant, and / or increasing the fresh weight of the underground part of the plant, and / or promoting the growth of the root crown of the plant, and / or promoting the growth of the root system of the plant, and / or increasing the number of lateral roots of the plant; or use of Bacillus Velez or its fermentation liquid in improving drought resistance of plants; The Bacillus velezensis is named Bacillus velezensis QN3NO-3, registered and preserved in the China Center for Type Culture Collection, with the preservation number being CCTCC NO: M 2021304.

4. A bacterial fertilizer containing Bacillus velezensis, wherein the Bacillus velezensis is named Bacillus velezensis QN3NO-3, registered and preserved in the China Center for Type Culture Collection, and the preservation number is CCTCC NO: M 2021304.

5. The bacterial fertilizer according to claim 4, characterized in that The bacterial fertilizer also includes other fertilizer nutrient elements; the bacterial fertilizer also includes auxiliary materials acceptable to fertilizers.

6. Use of the bacterial fertilizer as claimed in claim 4 or 5 in promoting plant growth.

7. The use according to claim 6, characterized in that Bacillus velezensis promotes plant growth by fixing nitrogen and / or secreting IAA.

8. The use of the bacterial fertilizer as claimed in claim 4 or 5 in promoting seed germination, and / or promoting seeds to enter the whitening stage in advance, and / or promoting plumule growth, and / or promoting radicle growth, and / or increasing the fresh weight of plant aerial parts, and / or increasing the fresh weight of plant underground parts, and / or promoting plant root crown growth, and / or promoting plant root system growth, and / or increasing the number of plant lateral roots; or Use of the bacterial fertilizer as claimed in claim 4 or 5 in improving plant drought resistance.

9. The use of the bacterial fertilizer as claimed in claim 4 or 5 in antagonizing tomato wilt pathogen, and / or banana wilt pathogen No. 4 physiological subspecies, and / or banana wilt pathogen No. 1 physiological subspecies, and / or cucumber wilt pathogen, and / or pepper anthracnose pathogen, and / or strawberry anthracnose pathogen, and / or litchi anthracnose pathogen, and / or banana anthracnose pathogen, and / or vegetable anthracnose pathogen, and / or gloeosporium anthracnose pathogen, and / or corn Curvularia leaf spot pathogen, and / or corn Alternaria leaf blight pathogen, and / or wheat fusarium, and / or mango polytrichosporium leaf blight pathogen, and / or rice blast pathogen, and / or mango anthracnose pathogen; or The use of the bacterial fertilizer as claimed in claim 4 or 5 in preventing and controlling diseases caused by tomato wilt pathogen, and / or banana wilt pathogen race 4, and / or banana wilt pathogen race 1, and / or cucumber wilt pathogen, and / or pepper anthracnose pathogen, and / or strawberry anthracnose pathogen, and / or litchi anthracnose pathogen, and / or banana anthracnose pathogen, and / or vegetable anthracnose pathogen, and / or gloeosporium anthracnose pathogen, and / or corn Curvularia leaf spot pathogen, and / or corn Alternaria leaf blight pathogen, and / or wheat fusarium sphaeroides pathogen, and / or mango polytrichosporium leaf blight pathogen, and / or rice blast pathogen, and / or mango anthracnose pathogen.

10. A method for promoting plant growth, characterized in that: Apply the bacterial fertilizer as claimed in claim 4 or 5 to plants.

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