Pseudomonas, microbial inoculum prepared from pseudomonas and application of pseudomonas

Abstract

The invention discloses Pseudomonas sp., a microbial agent prepared from the Pseudomonas sp. And application of the Pseudomonas sp. And the microbial agent prepared from the Pseudomonas sp. The pseudomonas strain N2-3-1-14 disclosed by the invention is preserved in the China General Microbiological Culture Collection Center (CGMCC); the address of the preservation unit is No.3, Yard 1, Beichen West Road, Chaoyang District, Beijing; the preservation time is April 8, 2025; the preservation number is CGMCC 34128. The pseudomonas strain N2-3-1-14 and the microbial inoculum prepared from the pseudomonas strain N2-3-1-14 can effectively inhibit growth of pathogenic bacteria of pine blight and pine needle red spot, and the bacteriostasis rate is 65% or above. The strain can also significantly promote the growth of pinus sylvestris.

Description

Technical Field

[0001] The present application relates to the field of microorganisms, and in particular, to a Pseudomonas, a microbial agent prepared therefrom, and applications thereof. Background Art

[0002] Pinus sylvestris var. mongholica belongs to the genus Pinus of the family Pinaceae, and is a geographical variety of Pinus sylvestris. It has a tall tree shape, green needles, strong stress resistance, and fast growth rate. It is a common greening tree species and an important tree species for afforestation in the sand source control project and the Three-North Shelter Forest Project.

[0003] Pine shoot blight is one of the common and most widely distributed important tree branch diseases of coniferous trees worldwide, and can damage about 60 or more coniferous tree species in about 8 genera, including the genus Pinus, Abies, and Larix. This disease is also one of the important diseases that damage Pinus sylvestris var. mongholica, and is also the main manifestation of the large-scale decline of Pinus sylvestris var. mongholica plantations.

[0004] The pathogen of pine shoot blight has genotype differentiation, and the pathogen causing pine shoot blight in China is Sphaeropsis sapinea type C. Sphaeropsis sapinea can infect multiple parts of host plants, such as young shoots, needles, buds, and cones. When the terminal bud is damaged, the new shoot cannot grow normally, resulting in the wilting and bending of the young shoot, and the color of the newly grown needles gradually turns yellow and withered, developing into a blighted shoot. When the pathogen invades the branch, it can cause ulcerous spots, making the surrounding xylem show a purple-brown color. Wedge-shaped blue discoloration can be seen on the cross-section of the ulcerous branch, seriously affecting the healthy growth of the tree and even causing death. At present, the prevention and control measures for this disease mainly rely on chemical control. However, the long-term use of chemical pesticides increases the prevention and control cost, and long-term use of pesticides will also cause the pathogen to develop drug resistance, seriously damaging the stability of the ecological system. In recent years, biological control has been considered by more and more researchers as one of the best strategies for preventing and controlling plant diseases.

[0005] The biocontrol research of pine shoot blight mainly focuses on Bacillus and Trichoderma. Song Ruiqing et al. obtained the highly efficient antagonist Trichoderma viride (Trichoder mavirens) against Diplodia pinea through the introduction and screening of highly efficient wood-decaying fungi. Tang Xu used JK-AH7 (Bacillus amyloliquefaciens) to treat Pinus elliottii by spraying, and the control effect was 47.8%. The combined treatment of root irrigation with JZ-GX1 (Rahnella aquatilis) and spraying with JK-AH7 achieved a control effect of 61.1%. Yun et al. isolated a strain of Bacillus pumilus from rhizosphere soil, and after prophylactic spraying on Pinus massoniana, the control effect on pine shoot blight reached 90%. However, there is no relevant report on the prevention and control of pine shoot blight by Pseudomonas at present. Summary of the Invention

[0006] The present invention provides a Pseudomonas sp., which is named Pseudomonas strain N2-3-1-14 (hereinafter referred to as N2-3-1-14), and is deposited in the General Microbiological Center of the China Committee for Culture Collection of Microorganisms; the address of the depositary institution: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing; the deposit time is April 8, 2025; the deposit number is: CGMCC 34128.

[0007] The present invention also provides a Pseudomonas agent, which is prepared from Pseudomonas strain N2-3-1-14. Specifically, the number of viable N2-3-1-14 bacteria in the agent ≥ 10 8 CFU / mL.

[0008] The present invention also provides the application of Pseudomonas strain N2-3-1-14 or the agent prepared from Pseudomonas strain N2-3-1-14 in preventing and controlling plant diseases and promoting plant growth. The diseases include preventing and controlling pine shoot blight caused by Sphaeropsis sapinea and red needle blight caused by Dothistroma septosporum.

[0009] Preferably, the method for preventing and controlling pine shoot blight or red needle blight is to spray Pseudomonas strain N2-3-1-14 or its agent; the application period is before the plant is infected with pine shoot blight or red needle blight.

[0010] The method for promoting plant growth is to perform root irrigation on the plant with Pseudomonas strain N2-3-1-14 or its agent.

[0011] Preferably, the plant is a pine tree; more preferably, the pine tree is Pinus sylvestris var. mongolica.

[0012] The beneficial effects of the present invention include:

[0013] The present invention provides a Pseudomonas sp. strain N2-3-1-14, which is obtained by confrontation culture experiments from more than 400 strains of bacteria isolated from the root tissue samples of Larix gmelinii. The Pseudomonas sp. strain N2-3-1-14 of the present invention and the microbial agent prepared therefrom can effectively inhibit the growth of the pathogens of pine shoot blight and pine needle red spot, and the antibacterial rate is more than 65%. Through pot control experiments, it is found that applying the microbial agent of the present invention before the pine wood is infected with pine shoot blight can achieve obvious control effects, and the average control efficacy can reach 75%. This strain can also significantly promote the growth of Pinus sylvestris var. mongolica, and is suitable for popularization and application as a biocontrol and growth-promoting microbial agent in production practice. Brief Description of the Drawings

[0014] Figure 1 It shows the growth inhibition of Sphaeropsis sapinea by the Pseudomonas sp. strain N2-3-1-14 in a petri dish in Example 1 of the present invention. Among them, A is the control treatment, and B is the treatment inoculated with the strain N2-3-1-14;

[0015] Figure 2 It is the control effect diagram of the pot control experiment in Example 3 of the present invention. Among them, A is the treatment only applying the biocontrol microbial agent N2-3-1-14, B is the treatment only applying sterile water, C is the treatment applying the biocontrol microbial agent N2-3-1-14 first and then inoculating the pathogen Sphaeropsis sapinea, and D is the treatment applying sterile water first and then inoculating the pathogen;

[0016] Figure 3 It is the measurement results of the seedling height, ground diameter, dry weight of the above-ground part and dry weight of the underground part of the pine seedlings in two groups of treatments in Example 4 of the present invention;

[0017] Figure 4 It is the contents of nitrogen, phosphorus and potassium in the needles of the pine seedlings in two groups of treatments in Example 4 of the present invention;

[0018] Figure 5 It shows the growth inhibition of Dothistroma septosporum by the Pseudomonas sp. strain N2-3-1-14 in a petri dish in Example 5 of the present invention. Among them, A is the control treatment, and B is the treatment inoculated with the strain N2-3-1-14.

[0019] Biological Deposit Information

[0020] Pseudomonas sp. strain N2-3-1-14 was deposited in the General Microbiological Center of the China Committee for Culture Collection of Microorganisms; the address of the depositary institution is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing; the deposit date was April 8, 2025; the deposit number is: CGMCC 34128. Detailed implementation manners

[0021] The present invention will be further described and described below in conjunction with embodiments. However, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. Based on the present invention and the embodiments, all other inventions and embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the scope of protection of the present invention.

[0022] Unless otherwise specified, the experimental methods used in the following embodiments are all conventional methods.

[0023] Unless otherwise specified, the materials, reagents, etc. used in the following embodiments can all be obtained from commercial channels.

[0024] Example 1: Isolation, purification and identification of root endophytic bacteria

[0025] (1) Isolation of bacteria

[0026] For the roots of Larix gmelinii collected from the Greater Khingan Mountains, the soil on the roots was brushed off with a sterile brush, stored in a sterile self-sealing bag and brought back in an ice box. After the root tissue was put into sterile water and shaken and washed twice, it was then ultrasonically washed for 10 min (160 W, 30 s / 30 s). After washing, it was soaked in 75% ethanol for 3 min, then soaked in 2.5% sodium hypochlorite for 3 min, and finally washed clean with sterile water. Take 2 g of root tissue and grind it into a homogenate with quartz sand and 2 mL of sterile water. After gradient dilution, it was spread on Tryptose Soya Agar (TSA) medium and incubated upside down at 28 °C for 2 days for counting and purification culture. Then, single colonies were picked and streaked on a new TSA plate to purify the target bacteria, and stored in a -70 °C refrigerator for later use. More than 400 strains of bacteria from root tissue were isolated and identified.

[0027] (2) Screening of biocontrol functional strains

[0028] Screen biocontrol functional strains by the plate confrontation culture method. Use a puncher to take a fresh culture of Sphaeropsis sapinea (strain 88568, isolated from Pinus sylvestris var. mongolica needles and preserved in the China Forestry Microbial Culture Collection Center) cake with a diameter of about 5 mm and inoculate it in the middle of the PDA medium. After activating the bacteria obtained in step (1), draw a line on each side at a vertical distance of 2 cm from the cake. Inoculate 1 strain of bacteria on each plate, and use a PDA plate inoculated with only the Sphaeropsis sapinea cake as a control. Do three plates for each treatment (3 replicate experiments). Place the culture dishes in an incubator at 25 °C and culture for 120 h, then observe and measure the inhibition rate. Inhibition rate (%) = (colony diameter of the control group - colony diameter of the treatment group) / colony diameter of the control group × 100%.

[0029] More than 10 strains antagonistic to Diplodia pinea were successfully screened through this experiment, including the functional strain N2-3-1-14 of the present invention. An obvious inhibition zone was formed in the confrontation test of this strain. Compared with the control group, the average inhibition rate was 65.8% (the results are as Figure 1 shown in Table 1). After multiple confrontation experiments, it was confirmed that the strain N2-3-1-14 has a strong antagonistic effect on Diplodia pinea.

[0030] Table 1 Inhibition rate of strain N2-3-1-14 against Sphaeropsis sapinea

[0031] Plate number Inhibitory rate against Mycosphaerella pini (%) Plate 1 65.2 Plate 2 64.8 Plate 3 67.4 Average inhibitory rate 65.8

[0032] (3) Molecular biological identification of strain N2-3-1-14

[0033] In the workbench, pick a single colony from the streaked bacterial plate and transfer it to TSB liquid medium. Culture it at 30 °C on a shaker at 220 rpm for 24 h. Take 2 mL of the overnight culture suspension, centrifuge it at 10000 rpm for 3 min, and discard the supernatant; add 200 μL of TE Buffer (10 mmol / L Tris, 1 mmol / L EDTA, pH 8.0), vortex to fully disperse the cells, add 200 μL of extraction Buffer (2% (m / v) CTAB, 0.7 mol / L NaCl, pH 7.5), 200 μL of 3 mol / L NaCl, and mix well; freeze-thaw repeatedly 2 times in liquid nitrogen and 100 °C environment; add an equal volume of chloroform:isoamyl alcohol (24:1), mix well, centrifuge at 6000 rpm at 4 °C for 10 min, and extract 2 times; take the supernatant, add 1 / 10 volume of NaAc (3 mol / L pH 5.2), add 2 volumes of -20 °C pre-cooled absolute ethanol, centrifuge at 12000 rpm at 4 °C for 15 min; discard the supernatant, air-dry the precipitate, add 100 μL of ddH2O to dissolve the precipitate, so as to obtain the DNA solution of strain N2-3-1-14, and store it at -20 °C for standby.

[0034] The 16S rDNA amplification primers are the universal primers 27F / 1492R for bacterial 16S identification. The PCR amplification products were directly sent to Beijing Bomed Technology Development Co., Ltd., and sequenced after gel cutting and purification by the company.

[0035] PCR reaction system (25ul):

[0036] 2xPCR Master mix: 12.5ul; Primer1 (27F): 1ul; Primer2 (1492R): 1ul; DNA: 1ul; ddH20: 9.5ul.

[0037] The PCR reaction program is:

[0038]

[0039] The homology analysis of the isolated bacteria was carried out using an online identification tool (https: / / www.ezbiocloud.net / ). The 16S rRNA gene sequence data was input into the database for sequence alignment. The results showed that the homology of Pseudomonas N2-3-1-14 with Pseudomonas canadensis 2-92 T , Pseudomonas fluorescens DSM 50090 T and Pseudomonas haemolytica DSM 108987 T was 99.93%, 99.72% and 99.72% respectively.

[0040] The DNA of strain N2-3-1-14 was sent to Beijing Novogene Bioinformatics Technology Co., Ltd. Genome sequencing was carried out using the PacBio RSII and Illumina HiSeq platforms. After quality detection using NanoDrop2000, a genomic DNA library was prepared. HGAP (version 4.0) was used to assemble de novo PacBio long reads, and Pilon (version 1.23) was used for error correction with Illumina short reads. The Quast (version 5.0.2) software was used for genome assembly statistics.

[0041] The strain was identified using a web-based genomic comparison tool. The DNA sequences of the isolated strain and three strains with high homology were compared and analyzed by calculating the average nucleotide identity (ANI) using the ANI calculator on the JSpecies web server and the DNA-DNA hybridization value of NA-DNA hybridization (dDDH, version 2.1) (http: / / ggdc.dsmz.de / distcalc2.php). The ANI values were between 85.5 - 88.34%, lower than the species threshold of 95 - 96%, and the dDDH values were between 31.9 - 37.1%, also lower than the species threshold of 70%. Thus, strain N2-3-1-14 was identified as a new species of the genus Pseudomonas.

[0042] The fatty acid composition of the strain was determined using an Agilent Technologies 6890n gas chromatograph. Fatty acid extraction and component analysis were carried out according to the method recommended in the instruction manual of the Sherlock TSBA60 Library version 6.0 (MIDI) microbial identification system. The results showed that strain N2-3-1-14 contained summed feature 5, C 19:0 cycloω8c and C 17:1 ω5c, which were the main characteristics distinguishing it from other strains with high homology. At the same time, the API 20E and API50CH test strips (bioMérieux) were used to determine the biochemical properties and acid production of the strain. Compared with strains with high homology, strain N2-3-1-14 could not utilize urease and could not produce acid using sucrose and xylitol. These results further confirmed that strain N2-3-1-14 was a new species of the genus Pseudomonas and was identified as Pseudomonas sp.

[0043] The strain N2-3-1-14 of the present invention was identified as Pseudomonas sp. This strain was deposited in the General Microbiological Center of the China Committee for Culture Collection of Microorganisms; the address of the depositary institution: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing; the deposit time was April 8, 2025; the deposit number was: CGMCC 34128.

[0044] Example 2: Preparation of the biocontrol agent of Pseudomonas strain N2-3-1-14

[0045] (1) Activation of Pseudomonas N2-3-1-14: In a laminar flow hood, a small amount of the preserved bacterial liquid was picked up with an inoculation loop and streaked on a plate of TSB solid medium, and then cultured in an incubator at 28°C for 24 h; to restore the vitality of the strain, a single colony was picked and subcultured continuously for 3 generations.

[0046] (2) Liquid culture: In a laminar flow hood, use an inoculation loop to pick a single colony from a plate containing TSB solid medium and transfer it to 10 mL of TSB liquid medium for liquid culture to obtain a liquid bacterial solution. The culture conditions are: at a pH value of 7.2 - 7.5 and a culture temperature of 28°C, shake culture at 220 rpm for 16 h.

[0047] (3) Propagation of the strain: In a laminar flow hood, pour the liquid bacterial solution cultured in step (2) into 250 ml of TSB liquid medium for liquid propagation culture. The culture conditions are: at a pH value of 7.2 - 7.5 and a culture temperature of 28°C, shake culture at 220 rpm for 24 h.

[0048] (4) Preparation of the microbial agent: Centrifuge the liquid bacterial solution obtained in step (3) at 10000 rpm for 5 min to precipitate the cultured strain, discard the supernatant, dilute the precipitate with sterile water, and measure the absorbance of the liquid bacterial solution (λ = 600 nm). When the absorbance value of the bacterial solution is 1, dilute it 10 times to obtain the microbial agent. The number of active bacteria of N2 - 3 - 1 - 14 in this microbial agent is about 10 8 CFU / mL.

[0049] Example 3. Pot experiment on biological control agent N2 - 3 - 1 - 14

[0050] (1) Preparation of the pathogenic bacterium

[0051] Transfer the strain 88568 of Sphaeropsis sapinea, the pathogenic bacterium of pine shoot blight, to a TSA petri dish and culture it at 25°C for 24 h. To restore the pathogenicity of the pathogenic bacterium, pick a single colony and transfer it continuously for 3 generations. Transfer the 4th generation to a PDA plate and culture it for 72 h for inoculation experiments.

[0052] (2) Pot experiment on the control effect of biological control agent N2 - 3 - 1 - 14 prepared in Example 2

[0053] Select two - year - old healthy Mongolian pine seedlings with consistent growth conditions for pot experiments. Set the Mongolian pine seedlings into 4 treatments:

[0054] A: Apply the N2 - 3 - 1 - 14 microbial agent;

[0055] B: Only apply an equal amount of sterile water;

[0056] C: Apply the N2 - 3 - 1 - 14 microbial agent and inoculate the pathogenic bacterium 88568 after 14 days;

[0057] D: Apply an equal amount of sterile water and inoculate the pathogenic bacterium 88568 after 14 days.

[0058] Each treatment has 10 replicates. Each plant in groups A and C is sprayed with 30 ml of the product prepared in Example 2 at a concentration of 10 8CFU / mL of the N2-3-1-14 bacterial agent. Meanwhile, groups B and D were applied with an equal amount of sterile water. After 14 days of greenhouse cultivation, inoculation with Mycosphaerella pini was carried out. The bark of Pinus sylvestris var. mongolica was surface-sterilized with 75% alcohol. A small incision about 1 cm long was made on the stem of Pinus sylvestris var. mongolica at a distance of 10 cm from the soil with a sterile scalpel. Then, a fresh culture of Mycosphaerella pini fungal cake (8 mm in diameter) was placed at the wound of the plant stems in groups C and D, and fixed with a moist sterile cotton and sealing film. Groups A and B were replaced with sterile PDA culture medium of the same size.

[0059] On the 15th day after inoculation with the pathogenic bacterium, the disease incidence of the potted Pinus sylvestris var. mongolica seedlings was counted. The disease severity was divided into grades 0-4 (0 = asymptomatic; 1 = <25% of the upper crown turned brown; 2 = 25.1%-50% of the upper crown turned brown; 3 = 50.1-75% of the upper crown turned brown; 4 = >75% of the upper crown was brown). The disease index and the relative biocontrol effect of the biocontrol bacterium were calculated as follows.

[0060]

[0061] On the 15th day of the inoculation experiment, the disease incidence of Pinus sylvestris var. mongolica was observed and recorded as Figure 2 shown, and the disease index is shown in Table 2. The average disease index of the Pinus sylvestris var. mongolica samples treated only with the pathogenic bacterium (group D) was 60, and the average disease index of the samples treated with the N2-3-1-14 bacterial agent first and then inoculated with the pathogenic bacterium (group C) was 15. It shows that the strain N2-3-1-14 can effectively enhance the resistance of pine seedlings to Botryosphaeria dothidea, and the relative control effect reaches 75%.

[0062] Table 2 Disease severity and disease index of potted Pinus sylvestris var. mongolica control experiment in 2023

[0063]

[0064] Combined with the control effect of first applying the N2-3-1-14 bacterial agent and then inoculating the pathogenic bacterium 14 days later in this example, it can be expected that applying the biocontrol bacterial agent of the present invention before the infection of the Botryosphaeria dothidea pathogen can achieve good control effects.

[0065] Example 4: Pot experiment on promoting growth of the biocontrol bacterial agent N2-3-1-14

[0066] Healthy two-year-old Pinus sylvestris var. mongolica seedlings with consistent growth conditions were selected for pot experiments. The Pinus sylvestris var. mongolica was set with two treatments:

[0067] A: Applying the growth-promoting bacterial suspension (treatment group);

[0068] B: Only applying an equal amount of sterile water (control group).

[0069] Each treatment had 10 replicates. Each plant in group A was applied with 30 mL of the N2-3-1-14 bacterial agent with a concentration of 10 8 CFU / mL prepared in Example 2, and group B was applied with an equal amount of sterile water. The irrigation root method was used for both groups, and they were cultured under greenhouse conditions for 5 months, with sterile water being regularly watered.

[0070] After 5 months of inoculation, a ruler was used to measure the height of the pine seedlings from the base of the ground diameter to the base of the apical bud (accurate to 0.1 cm) as the seedling height, and a vernier caliper was used to measure the diameter at the dry soil mark of the seedling stem (accurate to 0.002 cm) as the seedling ground diameter. The seedlings were cut at the base of the ground diameter and placed in an 80 °C oven to dry to a constant weight, and the dry weights of the above-ground and underground parts of the seedlings were weighed respectively.

[0071] The determination of nitrogen in the needles of pine seedlings was based on the forestry industry standard LY / T 1228-2015, the determination of phosphorus was based on the forestry industry standard LY / T 1232-2015, and the determination of potassium was based on the national standard LY / T 1234-2015.

[0072] The measurement results of the seedling height, ground diameter, dry weights of the above-ground and underground parts of the pine seedlings in the two groups of treatments were as Figure 3 shown ( Figure 3 where * represents a significant difference at the P<0.05 level). It can be seen from Figure 3 that the seedling height of the treatment group increased by 15.42% compared with the control group, the ground diameter increased by 7.02% compared with the control group, the above-ground dry weight increased by 16.36% compared with the control group, and the underground dry weight increased by 23.10% compared with the control group. Except for the ground diameter, the other indexes reached a significant difference level compared with the control group.

[0073] After 5 months of inoculation, the contents of nitrogen, phosphorus, and potassium in the needles of the pine seedlings in the two groups of treatments were measured simultaneously, and the results were as Figure 4 shown ( Figure 4 where * represents a significant difference at the P<0.05 level). It can be seen from Figure 4 that the nitrogen content in the leaves of the pine seedlings in the treatment group increased by 26% compared with the control group, the phosphorus content increased by 53% compared with the control group, and the potassium content increased by 14% compared with the control group, and the differences with the control group all reached a significant level.

[0074] Thus, it can be seen that under greenhouse conditions, the N2-3-1-14 bacterial agent can not only promote the growth of the biomass of pine seedlings, but also promote their absorption of nutrient elements, and has a significant growth-promoting effect.

[0075] Example 5, Inhibitory effect of strain N2-3-1-14 on the pine needle red spot pathogen Dothistroma septosporum

[0076] Take a fresh 3 mm × 6 mm culture block of Septoria acicola (isolated from Pinus koraiensis needles and preserved in the China Forestry Microbial Culture Collection Center), and inoculate it in the middle of the PDA medium. After activating the Pseudomonas strain N2-3-1-14 obtained in Example 1, spread it on the surface of the medium more than 1 cm away from the culture block. Inoculate 1 strain of bacteria on each plate, and use the PDA plate inoculated with only the Septoria acicola culture block as a control. Conduct three plates for each treatment (3 replicate experiments). Place the petri dishes in an incubator at 25 °C and observe and measure the inhibition rate after culturing for 120 h. The calculation method of the antibacterial rate refers to the formula in step (2) of Example 1.

[0077] The results are as Figure 5 shown. It can be seen from the figure that an obvious antibacterial zone is formed in the confrontation test of the functional strain N2-3-1-14 of the present invention. Compared with the control group, the average antibacterial rate is 83.6% (the results are shown in Table 3). After multiple confrontation experiments, it is confirmed that the strain N2-3-1-14 has a strong antagonistic effect on the pathogen Septoria acicola of pine needle red spot disease.

[0078] Table 3 Antibacterial rate of strain N2-3-1-14 against Septoria acicola

[0079] Plate number Inhibitory rate against Discocrea dothidea (%) Plate 1 85.7 Plate 2 82.3 Plate 3 82.9 Average inhibitory rate 83.6

Claims

1. A Pseudomonas sp., characterized in that The Pseudomonas sp. is named Pseudomonas sp. N2-3-1-14 and is deposited in the General Microbiology Center of China Culture Collection Administration; the deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing; the deposit date is April 8, 2025; and the deposit number is CGMCC 34128.

2. A Pseudomonas bacterial agent, characterized in that The bacterial agent is prepared from the Pseudomonas strain N2-3-1-14 according to claim 1.

3. The Pseudomonas agent according to claim 2, characterized in that The number of active bacteria of N2-3-1-14 in the bacterial agent is ≥10 8 CFU / mL.

4. Use of the Pseudomonas according to claim 1 or the microbial agent according to any one of claims 2 to 3 in preventing and controlling plant diseases and promoting plant growth, wherein the diseases include preventing and controlling pine tip blight caused by Sphaeropsis sapinea and pine needle red spot caused by Dothistroma septosporum.

5. The use according to claim 4, characterized in that The method for preventing and controlling pine tip blight or pine needle red spot is spraying the Pseudomonas strain N2-3-1-14 or its bacterial agent.

6. The use according to claim 5, characterized in that The application period is before the plants are infected with pine dieback or pine needle red spot.

7. The use according to claim 4, characterized in that The method for promoting plant growth comprises the following steps: using the Pseudomonas strain N2-3-1-14 or its bacterial agent to perform root irrigation treatment on plants.

8. The use according to any one of claims 4 to 7, characterized in that: The plant is a pine tree.

9. The use according to claim 8, characterized in that The pine tree is Pinus sylvestris var. mongolica.

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