Pseudomonas rugosa resistant to tomato gray mold, leaf mold and early blight fungus and application thereof

Abstract

The present invention relates to a kind of rugosa Pseudomonas spp. which is resistant to tomato gray mold, leaf mold and early blight fungus and its application. Pseudomonas corrugata YC-2305, whose deposit number is CGMCC NO.29940; Pseudomonas rugosa of the present invention Pseudomonas corrugata Both the bacteria and fermentation liquid of YC‑2305 have strong antibacterial activity against a variety of tomato fungal diseases, and the antibacterial effect is long-lasting and stable, which has great potential for developing new, highly effective and broad-spectrum biocontrol fungicides.

Description

Technical Field

[0001] The invention relates to the technical field of microorganisms, and in particular to a Pseudomonas rugosa resistant to tomato gray mold, leaf mold and early blight fungus and an application thereof. Background Art

[0002] Tomatoes are loved by people because of their rich nutrition and good taste when eaten raw or cooked. my country is a major tomato producer, and the planting area is expanding year by year. At present, most of them are planted in greenhouses. The temperature and humidity in greenhouses are generally high, which is prone to diseases. Among them, tomato gray mold, leaf mold, and early blight are three common diseases, which harm the fruits, stems and leaves, causing fruit rot and plant wilting, causing economic losses to tomato production. At present, pesticides are mainly used for tomato diseases, which are easy to cause drug resistance, and pesticide residues harm the health of humans and animals and pollute the environment.

[0003] Endophytes are widely present in healthy plant tissues and have co-evolved with them for a long time. They are commonly found in various terrestrial and aquatic plants that have been studied so far. They are natural components of plant microecological systems and have a complex microecological relationship with plants. Endophytes can produce a variety of biological effects in plants, participate in the defense function of plants, promote rapid growth of plants, and enhance the ability of plants to resist adversity, diseases, and animal damage. At present, researchers have screened strains that can antagonize a variety of tomato diseases through a large number of experiments, and have achieved remarkable results. Summary of the invention

[0004] The invention aims to provide a Pseudomonas rugosa resistant to tomato gray mold, leaf mold and early blight fungus and application thereof.

[0005] The technical solution of the present invention is as follows.

[0006] A rugosa Pseudomonas resistant to tomato gray mold, leaf mold, and early blight, wherein the rugosa Pseudomonas is rugosa Pseudomonas YC-2305, which is deposited in the General Microbiological Center of the China Microbiological Culture Collection Administration, with a deposit address of Beijing, China, a deposit number of CGMCC NO.29940, a deposit date of March 5, 2024, and a classification name of rugosa Pseudomonas Pseudomonas corrugata .

[0007] As a preferred technical solution, the colonies of Pseudomonas rugosa YC-2305 are milky white when grown on LB agar plates, and turn yellow later. It is a Gram-negative bacterium with a smooth and bright surface, moist texture, neat edges, opaque, round spots, and a colony diameter of 1 to 2 mm.

[0008] As a preferred technical solution, the 16S rDNA sequence of the Pseudomonas rugosa YC-2305 is shown in SEQ ID NO:1.

[0009] The present invention also discloses an application of Pseudomonas rugosa resistant to tomato gray mold, leaf mold and early blight fungus.

[0010] The bacterial body and fermentation liquid of the Pseudomonas rugosa YC-2305 are used to inhibit or prevent tomato gray mold and the diseases caused by it;

[0011] The bacteria and fermentation liquid of Pseudomonas rugosa YC-2305 are used to inhibit or prevent tomato leaf mold and the diseases caused by it;

[0012] The bacterial body and fermentation liquid of the rugosa Pseudomonas YC-2305 are used for inhibiting or preventing tomato early blight fungus and the diseases caused by it.

[0013] The appearance of a yellow halo around the colony of Pseudomonas rugosa YC-2305 indicates that Pseudomonas rugosa YC-2305 can produce siderophore, which promotes plant growth and has a preventive and therapeutic effect on diseases.

[0014] As a preferred technical solution, the bacterial culture conditions of the rugosa Pseudomonas YC-2305 are: inoculate the activated strain at a 1% inoculum into LB medium with a ventilation volume of 50 mL to 75 mL / 250 mL, pH 5.5 to 8, 35°C, and 150 rpm shaking on a shaker for constant temperature shaking.

[0015] As a preferred technical solution, the pH is 6-8; the antibacterial effect of the cultivated Pseudomonas rugosa is more significant.

[0016] Advantages of the present invention:

[0017] The antibacterial activity of the bacteria and fermentation liquid of the rugosa Pseudomonas YC-2305 of the present invention is determined by plate confrontation method and colony growth rate method. The rugosa Pseudomonas YC-2305 has a good antagonistic effect on tested tomato gray mold, leaf mold and early blight.

[0018] The bacterial body and fermentation liquid of the rugosa Pseudomonas YC-2305 of the present invention both have strong antibacterial activity against a variety of tomato fungal diseases, and the antibacterial effect is long-lasting and stable, and has great potential for developing new, efficient and broad-spectrum biocontrol fungicides therefrom. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 This is a colony morphology diagram of Pseudomonas rugosa YC-2305 in Example 1 of the present invention;

[0020] Figure 2 This is a graph showing the Gram test results of Pseudomonas rugosa YC-2305 in Example 1 of the present invention;

[0021] Figure 3The phylogenetic tree diagram of the Pseudomonas rugosa YC-2305 strain constructed from the 16S rDNA gene sequence in Example 1 of the present invention;

[0022] Figure 4 This is a graph showing the results of Pseudomonas rugosa YC-2305 resisting tomato leaf mold in Example 3 of the present invention;

[0023] Figure 5 This is a graph showing the results of the resistance of Pseudomonas rugosa YC-2305 to Botrytis cinerea in Example 3 of the present invention;

[0024] Figure 6 This is a graph showing the results of the resistance of Pseudomonas rugosa YC-2305 to tomato early blight fungus in Example 3 of the present invention;

[0025] Figure 7 This is a graph showing the test results of the siderophore production by Pseudomonas rugosa YC-2305 in Example 8 of the present invention. DETAILED DESCRIPTION

[0026] The present invention is further described below in conjunction with specific embodiments.

[0027] Embodiment 1:

[0028] Isolation and identification of Pseudomonas rugosa YC-2305

[0029] The Pseudomonas rugosa YC-2305 resistant to various tomato fungal diseases of the present invention was obtained by culturing and identifying the leaves of Chinese cabbage from the vegetable base of Yantai Academy of Agricultural Sciences in Shandong Province in a laboratory under the following conditions.

[0030] Isolation and purification of YC-2305: The homogenate coating method was used. The leaves of Chinese cabbage collected in the seedling stage were cut into 0.8 cm × 0.8 cm small pieces, a total of 20 pieces, soaked in 75% ethanol for 30 s in a clean bench, and then soaked in 2.5% NaClO (effective Cl-10%) solution for 3 min, and then rinsed with sterile water 4 times. 1 mL of sterile water from the last leaf washing was taken and smeared on the plate as a control to test whether the surface disinfection was thorough. The disinfected Chinese cabbage leaves were placed in a sterilized mortar, a small amount of quartz sand was added and crushed into juice, which was diluted to 1 mL, 2 mL, and 5 mL respectively, and spread on NA solid culture medium plates, repeated 3 times, and cultured at 37°C for 1 d to 2 d, and different single colonies were picked and stored for later use.

[0031] Pseudomonas rugosa YC-2305 was identified based on colony morphology, individual morphological characteristics, physiological and biochemical characteristics, and the phylogenetic tree of 16S rDNA gene sequences.

[0032] The individual morphology and colony characteristics of the bacteria are as follows: The colony of bacteria YC-2305 is milky white, turning yellow later, with a smooth and bright surface, moist texture, neat edges, opaque, round spots, and a colony diameter of 1-2 mm. The bacteria are straight or slightly curved rods, not spiral, without stipes or sheaths, and do not produce spores. Gram-negative bacteria require oxygen and carry out strict respiratory metabolism, with oxygen as the final electron acceptor. Figure 1 The colony morphology is Figure 2 Gram test results.

[0033] 16S rDNA homology analysis: The genomic DNA of the strain was used as a template, and the bacterial universal primers 27-F / 1492-R were used to perform PCR amplification of the 16S rDNA gene. The primer sequences were: 27-F: 5'-AGAGTTTGATCCTGGCTCAG-3', 1492-R: 5'-GGTTACCTTGTTACGACTT-3'. The PCR product was a single band, which was purified after recovery and DNA sequencing. The 16S rDNA sequence of the strain was 1444bp in length, and the nucleotide sequence of the specific sequence is shown in SEQ ID NO: 1. The 16S rDNA sequence of the strain was submitted to the GenBank database, and the sequence number GenBank NO. SUB14112590 was obtained. Figure 3 Phylogenetic tree of Pseudomonas rugosa YC-2305 strain constructed based on 16S rDNA gene sequence.

[0034] Embodiment 2:

[0035] Optimal conditions for the cultivation and fermentation of Pseudomonas rugosa YC-2305

[0036] (1) Slant culture conditions: beef extract peptone solid medium: 3 g beef extract powder, 10 g peptone, 5 g sodium chloride, 15 g agar powder, 1000 mL water, pH 7.3±0.2. Autoclave at 121°C, 0.1 MPa for 20 min; culture temperature 37°C, culture time 2-4 days.

[0037] (2) Fermentation conditions: LB medium: 10 g tryptone, 5 g yeast extract, 10 g sodium chloride, and 1000 mL water.

[0038] Use phosphate buffer to adjust the pH of 50 mL of LB liquid culture medium to 5.5, 6.0, 6.5, 7.0, and 7.5, respectively. Sterilize with steam at 121°C and 0.1MPa for 20 minutes. After inoculation, place the culture medium in a shaker at 30°C and 150 rpm for constant temperature shaking. Calculate the number of viable bacteria according to the LB culture medium plating method to find the optimal pH value for the growth of different strains.

[0039] The activated strains were inoculated into 50 mL LB liquid medium and placed in a constant temperature incubator at 25°C, 28°C, 30°C, 35°C, and 40°C for 48 h. 1 mL of each treatment was diluted 10 times by stepwise dilution. 5 ~ 10 7 50 uL LB was taken out and spread on the plate. Each treatment was repeated 3 times. The number of live colonies on the plate was calculated after constant temperature culture at 30 ℃ for 24 h to 36 h.

[0040] Inoculate the activated strains at 1% in 250 mL Erlenmeyer flasks containing 25 mL, 50 mL, 75 mL, 100 mL, and 150 mL LB liquid culture medium. Cultivate at 30 °C, 150 rpm in a shaker. Calculate the number of viable bacteria using the LB plating method to find the optimal ventilation volume for strain growth.

[0041]

[0042] The results are shown in Table 1.

[0043] Table 1

[0044]

[0045] The experiment showed that the optimal culture conditions for Pseudomonas rugosa YC-2305 were to inoculate the activated strain at a 1% inoculum into LB medium with an aeration volume of 50 mL ~75 mL / 250 mL, and culture at a pH of 5.5 ~ 6.0, 35 ℃, and 150 rpm in a shaker.

[0046] Embodiment 3:

[0047] Analysis of the antibacterial activity of Pseudomonas rugosa YC-2305 against tomato pathogens

[0048] Antibacterial validation test on tomato leaf mold: using the plate confrontation method, a 6 mm diameter cake of tomato leaf mold was placed in the middle of a PDA solid culture medium plate, and YC-2305 was inoculated 2.5 cm around it. One strain of bacteria was inoculated on each plate, and 3 points were inoculated symmetrically. 70 μL of YC-2305 solution (concentration of 10 5 cfu / mL), repeated 3 times. Cultured in a 25 ℃ incubator, with no bacteria as control. When the control grew to the edge of the culture dish, the size of the inhibition zone was measured, see Table 2 and Figure 4 .

[0049] Table 2 Size of the inhibition zone of Pseudomonas rugosa YC-2305 against tomato leaf mold pathogen

[0050]

[0051] Antibacterial verification test on tomato early blight: Using the colony growth rate method, take the bacterial solution of Pseudomonas rugosa YC-2305 of the present invention, and add 200 uL of the concentration of 10 5 The bacterial liquid with a concentration of 100 cfu / mL was evenly spread on the sterilized PDA solid culture medium. When the bacteria were slightly dry, a 6 mm diameter indicator bacteria (tomato early blight pathogen, tomato gray mold) cake was placed in the center of the plate. The PDA solid culture medium plate inoculated with only the indicator bacteria (tomato early blight pathogen, tomato gray mold) cake was used as the control. When the indicator bacteria (tomato early blight pathogen, tomato gray mold pathogen) covered the plate, the diameter of the treated colony was measured to determine the antibacterial effect of the antagonistic bacteria YC-2305 on the indicator bacteria (tomato early blight pathogen, tomato gray mold pathogen). The results are shown in Table 3. Figure 5 , Figure 6 .

[0052] Table 3 The inhibitory effect of Pseudomonas rugosa YC-2305 on tomato early blight and tomato gray mold

[0053]

[0054] From Table 2, Table 3 and Figure 4 , Figure 5 , Figure 6 It can be seen that the size of the inhibition zone of Pseudomonas rugosa YC-2305 against tomato leaf mold is 20.1 mm, and the inhibition rates against tomato early blight and tomato gray mold are 92.16% and 89.17% respectively, indicating that Pseudomonas rugosa YC-2305 has resistance or inhibitory effects on the above pathogens.

[0055] Embodiment 4:

[0056] Analysis on the adaptability of Pseudomonas rugosa YC-2305 bacterial solution to antibacterial conditions

[0057] Thermal adaptability analysis: The bacterial liquid of Pseudomonas rugosa YC-2305 was placed at 20 ℃, 40 ℃, 60 ℃, 80 ℃, and 100 ℃ for 30 min, and 121 ℃ for 20 min. Tomato leaf mold, tomato early blight, and tomato gray mold were used as indicator bacteria. The plate confrontation method was used. A 6 mm diameter bacterial cake of tomato leaf mold was placed in the middle of a PDA solid culture medium plate, and the treated YC-2305 bacterial liquid was inoculated 2.5 cm around it. Two points were symmetrically inoculated on each plate, and 70 μL of the treatment solution (concentration of 10 5cfu / mL), repeated 3 times. The samples were cultured in a 25 ℃ constant temperature box, and the antibacterial activity of the antibacterial substances was determined with the indicator bacteria alone as the control. The results showed that there was no significant change in the inhibitory effect on tomato leaf mold, tomato early blight and tomato gray mold at 20 ℃~40 ℃. When the temperature reached above 60 ℃, the bacterial viability and inhibitory activity decreased significantly.

[0058] pH adaptability analysis: The initial pH value of the antibacterial substance crude extract of Pseudomonas rugosa YC-2305 was determined and used as the control group. The YC-2305 bacterial solution was adjusted to pH 2, 4, 6, 8, 10 and 12 with 1 mol / L HCL and NaOH, and the pH value was adjusted back to the initial value after 2 hours. Tomato leaf mold, tomato early blight and tomato gray mold were used as indicator bacteria. The plate confrontation method was used. A 6 mm diameter bacterial cake of tomato leaf mold was placed in the middle of a PDA solid culture medium plate, and the treated YC-2305 bacterial solution was inoculated 2.5 cm around it. Two points were symmetrically connected to each dish, and 70 μL of the treatment solution (concentration of 10 5 cfu / mL), repeated 3 times. The cells were cultured in a 25 ℃ constant temperature incubator, and the antibacterial activity of the antibacterial substances was determined with the indicator bacteria alone as the control. The results showed that the antibacterial effect did not change significantly in the pH range of 4 to 10, and the bacterial activity decreased significantly under strong acid and strong alkali conditions, which had a certain effect on the antibacterial effect.

[0059] Embodiment 5:

[0060] Experimental study on the control of tomato gray mold by using Pseudomonas rugosa YC-2305

[0061] 1. Preparation of pathogen inoculum: Wash the collected diseased tomato leaves, select the junction of diseased and healthy leaves and cut them into small pieces for culture on the culture medium. PDA culture medium is used for tomato gray mold. After 2 to 3 days of culture, single spores are separated and purified. Use PDA culture medium and culture in a constant temperature incubator at 28 ℃ for 14 days. After spore production, wash the spores with sterile water, filter with double gauze, obtain conidia suspension and dilute to 5×10 7 pcs / mL for later use.

[0062] 2. Preparation of YC-2305 bacterial agent: Inoculate the activated strain at a rate of 1% into LB medium with an aeration volume of 50 mL~75 mL / 250 mL, and culture at a pH of 5.5, 35°C, and 150 rpm in a shaking incubator. The resulting fermentation liquid is the YC-2305 bacterial agent.

[0063] 3. Experiment on the control of tomato gray mold with YC-2305 bacterial agent:

[0064] The tomato variety was Yanfen 201. The seeds were sown in plug trays, with 10 seedlings per replicate for a total of 3 replicates. The treatment was performed when the tomato had 4 to 6 leaves.

[0065] Pathogen inoculation treatment: Spray the conidia suspension of Botrytis cinerea on the surface and back of tomato true leaves, about 1 mL of conidia suspension per plant, with a conidia concentration of 5×10 7 Before inoculation, keep the air moist for 12 hours to make the relative humidity of the indoor air reach 90%~100%. After inoculation, the humidity should be kept above 85% for 8 hours, and then normal management can be carried out.

[0066] YC-2305 preventive treatment: spray YC-2305 on the surface and back of tomato true leaves, about 1 mL of the agent per plant, with a spore concentration of 10 5 After the inoculum dried, the conidia suspension of Botrytis cinerea was inoculated, with about 1 mL of conidia per strain and a spore concentration of 5×10 7 / mL, the humidity was maintained above 85% for 8 h after inoculation, and then normal management was carried out.

[0067] YC-2305 treatment: Spray the conidia suspension of Botrytis cinerea to each plant with about 1 mL of the suspension at a spore concentration of 5×10 7 After inoculation, the humidity was kept above 85% for 8 h, and then YC-2305 was sprayed on the surface and back of the true leaves of tomatoes, about 1 mL of the agent per plant, and the spore concentration of the bacterial solution was 10 5 cfu / mL and then managed normally.

[0068] Control treatment: Use the spray method to spray LB culture medium on the surface and back of tomato true leaves, about 1 mL per plant. After spraying, the humidity is maintained above 85% for 8 hours, and then normal management is performed.

[0069] The disease index was investigated 7 days after treatment.

[0070] Level 0: no lesions; Level 1: the lesion area accounts for less than 5% of the total leaf area; Level 3: the lesion area accounts for 6% to 15% of the total leaf area; Level 5: the lesion area accounts for 16% to 25% of the total leaf area; Level 7: the lesion area accounts for 26% to 50% of the total leaf area; Level 9: the lesion area accounts for more than 50% of the total leaf area.

[0071] Disease index = [∑(number of diseased leaves at each level × relative level value) / (total number of leaves investigated × 9)] × 100

[0072] Control effect (%) = (CK treatment disease index - YC-2305 treatment disease index) / CK treatment disease index × 100%.

[0073] Experimental results: The disease investigation results showed that on the 7th day after inoculation with tomato gray mold, each treatment had disease to varying degrees. The prevention and treatment treatments with the Pseudomonas rugosa YC-2305 agent showed strong disease prevention effects. The disease index of the prevention treatment was 8.9, and the disease index of the treatment treatment was 12.22, both lower than that of the control treatment. The prevention efficiencies were 82.56% and 76.09%, respectively, indicating that prevention was better than treatment (Table 4) and had good application value.

[0074] Table 4 Control effect of Pseudomonas rugosa YC-2305 on tomato gray mold

[0075]

[0076] Embodiment 6:

[0077] Experiment on the control of tomato leaf mold by using Pseudomonas rugosa YC-2305

[0078] The tomato variety was Yanfen 201. The seeds were sown in plug trays, with 10 seedlings per replicate for a total of 3 replicates. The treatment was performed when the tomato had 3 to 4 leaves.

[0079] Pathogen inoculation treatment: Spray the tomato leaf mold conidia suspension on the surface and back of the tomato true leaves using the spray method. Each plant should have about 1 mL of conidia suspension, and the conidia concentration of the conidia suspension should be 5×10 7 Moisturize for 24 hours before inoculation to make the relative humidity of indoor air reach 90%~100%, and keep the humidity above 85% for 8 hours after inoculation, and then manage normally.

[0080] YC-2305 preventive treatment: spray YC-2305 on the surface and back of tomato true leaves, about 1 mL of the agent per plant, with a spore concentration of 10 5 After the inoculum dried, the conidia suspension of tomato leaf mold was inoculated, with about 1 mL of bacterial solution per plant, and the spore concentration of the bacterial solution was 5×10 7 / mL, the humidity was maintained above 85% for 8h after inoculation, and then normal management was carried out.

[0081] YC-2305 treatment: Spray the tomato leaf mold conidia suspension with about 1 mL of the suspension per plant, with a spore concentration of 5×10 7After inoculation, the humidity was kept above 85% for 8 h, and then YC-2305 was sprayed on the surface and back of the true leaves of tomatoes, about 1 mL of the agent per plant, and the spore concentration of the bacterial solution was 10 5 cfu / mL and then managed normally.

[0082] The disease index was investigated 14 days after treatment.

[0083] Level 0: no symptoms; Level 1: chlorotic to yellow spots appear on the inoculated leaves; Level 3: a thin and sparse layer of mildew appears on the spots on the inoculated leaves; Level 5: an obvious mildew layer appears on the spots on the inoculated leaves; Level 7: a dense mildew layer appears on the spots on the inoculated leaves, and the upper leaves are also infected; Level 9: in addition to the dense mildew layer on the spots on the inoculated leaves, the mildew layer on the upper leaves is also very obvious or the diseased leaves begin to turn yellow.

[0084] Disease index = [∑(number of diseased leaves at each level × relative level value) / (total number of leaves surveyed × highest level value)] × 100

[0085] Control effect (%) = (CK treatment disease index - YC-2305 treatment disease index) / CK treatment disease index × 100%.

[0086] Experimental results: The results of the disease investigation showed that on the 14th day after inoculation with tomato leaf mold, each treatment had disease to varying degrees. The prevention and treatment treatments with the Pseudomonas rugosa YC-2305 agent showed strong disease prevention effects. The disease index of the prevention treatment was 7.5, and the disease index of the treatment treatment was 11.11, both lower than that of the control treatment. The prevention efficiencies were 86.5% and 80%, respectively, indicating that prevention was better than treatment (Table 5) and had good application value.

[0087] Table 5 Control effect of Pseudomonas rugosa YC-2305 on tomato leaf mold

[0088]

[0089] Embodiment 7:

[0090] Experiment on the control of tomato early blight by using Pseudomonas rugosa YC-2305

[0091] The tomato variety was Yanfen 201. The seeds were sown in plug trays, with 10 seedlings per replicate for a total of 3 replicates. The treatment was carried out when the tomato reached the 4-leaf stage.

[0092] Pathogen inoculation treatment: Spray the conidia suspension of tomato early blight fungus on the surface and back of tomato true leaves, about 1 mL of conidia suspension per plant, and the conidia concentration of the conidia suspension was 5×10 6Before inoculation, keep the relative humidity of the indoor air at 90% to 100% for 24 hours, and keep the humidity above 85% for 24 hours after inoculation, and then manage normally.

[0093] YC-2305 preventive treatment: spray YC-2305 on the surface and back of tomato true leaves, about 1 mL of the agent per plant, with a spore concentration of 10 5 After the inoculum dried, the conidia suspension of tomato early blight fungus was inoculated, with about 1 mL of conidia per plant and a conidia concentration of 5×10 6 / mL, the humidity was maintained above 85% for 24h after inoculation, and then managed normally.

[0094] YC-2305 treatment: Spray the conidia suspension of tomato early blight fungus with about 1 mL of conidia suspension per plant. The conidia concentration of the conidia suspension is 5×10 6 After inoculation, the humidity was kept above 85% for 24 h, and then YC-2305 was sprayed on the surface and back of the true leaves of tomatoes, about 1 mL of the agent per plant, and the spore concentration of the bacterial solution was 10 5 cfu / mL and then managed normally.

[0095] The disease index was investigated 7 days after treatment.

[0096] Level 0: no lesions on leaves; Level 1: lesion area accounts for less than 1 / 4 of the pinnate leaves; Level 2: lesion area accounts for 1 / 4 to 1 / 2 of the pinnate leaves; Level 3: lesion area accounts for 1 / 2 to 3 / 4 of the pinnate leaves, and nearly half of the leaflets are dead; Level 4: lesion area accounts for more than 3 / 4 of the pinnate leaves, and more than half or all of the leaflets are dead.

[0097] Disease index = [∑(number of diseased leaves at each level × relative level value) / (total number of leaves surveyed × highest level value)] × 100

[0098] Control effect (%) = (CK treatment disease index - YC-2305 treatment disease index) / CK treatment disease index × 100%.

[0099] Experimental results: The disease investigation results showed that on the 7th day after inoculation with tomato early blight, each treatment had disease to varying degrees. The prevention and treatment treatments with Pseudomonas rugosa YC-2305 showed strong disease prevention effects. The disease index of the prevention treatment was 12.5, and the disease index of the treatment treatment was 18.75, both lower than that of the control treatment. The prevention efficiencies were 75% and 62.5%, respectively, indicating that prevention was better than treatment (Table 6) and had good application value.

[0100] Table 6 Control effect of Pseudomonas rugosa YC-2305 on tomato early blight

[0101]

[0102] Embodiment 8:

[0103] Siderophore production ability of Pseudomonas rugosa YC-2305

[0104] The culture medium (CAS medium) for siderophore detection is used for culture detection. The main components of CAS medium are: chrome azuro blue S (CAS), hexadecyl trimethylammonium bromide (HDTMA), ferric chloride hexahydrate, sodium dihydrogen phosphate dihydrate, disodium hydrogen phosphate dodecahydrate, ammonium chloride, potassium dihydrogen phosphate, sodium chloride, agar, pH 6.8±0.1, and the plate is spread. This plate is the siderophore detection plate. The activated Pseudomonas rugosa YC-2305 was inoculated on the siderophore detection plate, and after culturing at 28℃ for 72 hours, a yellow halo was observed around the colony to see if it produced a yellow halo. Since siderophore competes for the iron ions chelated by EDTA in the culture medium, the culture medium changes from blue to yellow. Therefore, if a yellow halo appears around the colony, it indicates that it produces siderophore.

[0105] The results showed that a yellow halo appeared around the colonies of Pseudomonas rugosa YC-2305, and that Pseudomonas rugosa YC-2305 could produce siderophores, which can promote plant growth and prevent diseases ( Figure 7 ).

[0106] The above shows and describes the basic principles, main features and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The above embodiments and the description in the specification are only for illustrating the principles of the present invention. Without departing from the spirit and scope of the present invention, the present invention may have various changes and improvements, which fall within the scope of protection of the present invention. The scope of protection of the present invention is defined by the attached claims and their equivalents.

Claims

1. A Pseudomonas rugosa resistant to tomato gray mold, leaf mold, and early blight fungus, characterized in that: The rugosa Pseudomonas is rugosa Pseudomonas YC-2305, which is deposited in the General Microbiological Center of the China Microbiological Culture Collection Administration, with a deposit address of CGMCC NO.29940 in Beijing, China, a deposit date of March 5, 2024, and a classification name of Pseudomonas rugosa Pseudomonas corrugata .

2. The Pseudomonas rugosa resistant to tomato gray mold, leaf mold, and early blight fungus according to claim 1, characterized in that: When the strain of Pseudomonas rugosa YC-2305 grows on LB agar plates, the colonies are milky white and turn yellow later. It is a Gram-negative bacterium with a smooth and bright surface, moist texture, neat edges, opaque, round spots, and a colony diameter of 1 to 2 mm.

3. The Pseudomonas rugosa resistant to tomato gray mold, leaf mold, and early blight fungus according to claim 1, characterized in that: The 16S rDNA sequence of the Pseudomonas rugosa YC-2305 is shown in SEQ ID NO:

1.

4. A use of Pseudomonas rugosa against tomato gray mold, leaf mold and early blight as claimed in any one of claims 1 to 3, characterized in that: The bacterial body and fermentation liquid of the Pseudomonas rugosa YC-2305 are used to inhibit or prevent tomato gray mold and the diseases caused by it; Or the bacteria and fermentation liquid of the Pseudomonas rugosa YC-2305 are used to inhibit or prevent tomato leaf mold and the diseases caused by it; Or the bacterial body and fermentation liquid of the rugosa Pseudomonas YC-2305 are used to inhibit or prevent tomato early blight fungus and the diseases caused by it.

5. The use of Pseudomonas rugosa against tomato gray mold, leaf mold and early blight as claimed in claim 4, characterized in that: The bacterial culture conditions of the rugosa Pseudomonas YC-2305 are as follows: the activated strain is inoculated at a 1% inoculum into an LB medium with an aeration volume of 50 mL to 75 mL / 250 mL, pH 5.5 to 8, 35°C, and a constant temperature shaking culture on a shaker at 150 rpm.

6. The use of Pseudomonas rugosa against tomato gray mold, leaf mold and early blight as claimed in claim 5, characterized in that: The pH is 6-8.

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