Growth-promoting biocontrol bacterium for lettuce rhizosphere and application of growth-promoting biocontrol bacterium

Abstract

The invention discloses a growth-promoting biocontrol bacterium for lettuce rhizosphere and application of the growth-promoting biocontrol bacterium, the name of the growth-promoting biocontrol bacterium is pseudomonas urethra GPBC-2, and the preservation number of the growth-promoting biocontrol bacterium in the China Center for Type Culture Collection is CCTCC NO: M 20232551. The growth-promoting biocontrol bacterium can be used for promoting plant growth and can also be used for preventing and treating meloidogyne incognita. The growth-promoting biocontrol bacterium not only has an efficient growth-promoting effect on plants, but also has high nematode killing activity on meloidogyne incognita, can reduce the infection of the meloidogyne incognita on plant roots, and does not cause pollution to the environment.

Description

Technical Field

[0001] The present invention belongs to the technical field of agricultural microorganisms, and relates to growth-promoting and biocontrol bacteria in the lettuce rhizosphere and their applications, specifically to Pseudomonas urethralis and its applications in promoting crop growth and controlling Meloidogyne incognita Background Art

[0002] Plant growth-promoting rhizobacteria (PGPR) are a class of beneficial rhizosphere bacteria that promote plant growth by dissolving phosphorus, fixing nitrogen, secreting plant growth hormones, and antagonizing pathogenic microorganisms. On the one hand, as biological fertilizers, PGPR not only promote plant growth, enhance the resistance of plants to abiotic stresses, but also enhance soil fertility, reduce the use of chemical fertilizers, and promote the sustainable development of agriculture; on the other hand, PGPR also play an important role in inhibiting infectious diseases. There are a wide variety of infectious plant diseases, including Phytophthora blight, anthracnose, and root-knot nematode disease, etc., among which plant root-knot nematodes cause extensive damage.

[0003] Plants mainly absorb nutrients directly from the soil through their roots. When the roots of plants are infected by pathogens, it often leads to plant diseases and even death. Currently, there are various control measures for plant diseases, among which the use of chemical pesticides is more extensive. However, the long-term use of chemical pesticides will cause pathogens to develop drug resistance, cause environmental pollution, and endanger the health of humans and livestock, etc. In modern agriculture, beneficial microorganisms have gradually become safe alternatives to chemical fungicides, such as Bacillus spp. ( Bacillus ), Pseudomonas spp. ( Pseudomonas ). Although a rich variety of microbial resources have been isolated currently, only a part of the microorganisms have been made into products and applied in actual production and life. Therefore, it is still of great significance to explore efficient and novel biological resources.

[0004] It has been reported that the endophyte Pseudomonas moraviensis GF-55 can promote the growth of maize, the Pseudomonas sp. strain S10B9 has biocontrol effects on Fusarium root rot of alfalfa, and Pseudomonas aeruginosa Bc1-20 can well control melon fusarium wilt and has a growth-promoting effect. However, the current average level of growth promotion and biocontrol of Pseudomonas bacteria still needs to be improved. At the same time, there are no relevant reports on exploring the promotion of crop growth by Pseudomonas urethralis and controlling Meloidogyne incognita of crops. Summary of the Invention

[0005] The technical problem to be solved by the present invention is to overcome the deficiencies of the prior art and provide a growth-promoting and biocontrol bacteria in the lettuce rhizosphere and its applications, which not only have a highly efficient growth-promoting effect on crops, but also have a high nematicidal activity against Meloidogyne incognita, can reduce the infection of crop roots by Meloidogyne incognita, and will not cause environmental pollution.

[0006] To solve the above technical problems, the present invention adopts the following technical solutions.

[0007] A growth-promoting biocontrol bacterium in the rhizosphere of lettuce, named Pseudomonas ureae ( Pseudomonas urethralis ) GPBC-2, its preservation number in the China Center for Type Culture Collection is CCTCC NO: M 20232551, the address of the preservation unit is the China Center for Type Culture Collection (Wuhan University), No. 299, Bayi Road, Wuchang District, Wuhan City, Hubei Province, China, and the preservation date is December 13, 2023.

[0008] Pseudomonas ureae GPBC-2 strain has the following obvious characteristics: The strain is a Gram-negative bacterium, identified as Pseudomonas by molecular biology, the cells are rod-shaped under the microscope, do not form spores, cultured on LB agar plate (pH 7.0) for 24 h, the colonies are light yellow, extremely smooth and wrinkled on the surface, easy to pick, grow well under aerobic conditions, cultured on PKO modified inorganic phosphorus (tricalcium phosphate) agar plate (pH 7.4 ± 0.2) for 48 h, a transparent circle is formed around the colonies, and the optimum growth temperature is 28 °C. The 16S rDNA sequence of the strain is shown in SEQ ID NO.1.

[0009] As a general technical concept, the present invention also provides an application of the above growth-promoting biocontrol bacterium in the rhizosphere of lettuce in promoting plant growth.

[0010] For the above application, preferably, the application includes the following steps: Prepare a growth-promoting biocontrol bacterium fermentation broth with an OD 600 not less than 1, perform root irrigation on plants, not less than 5 mL per plant, and treat once every 5d - 7d, with the number of treatments not less than 3 times.

[0011] For the above application, preferably, the preparation method of the growth-promoting biocontrol bacterium fermentation broth includes the following steps: (1) Inoculate the growth-promoting biocontrol bacterium into LB liquid medium for activation; (2) Centrifuge the activated bacterial suspension, discard the supernatant, and resuspend with sterile water to obtain the growth-promoting biocontrol bacterium fermentation broth.

[0012] For the above application, preferably, in step (1), the inoculation amount of the growth-promoting biocontrol bacterium is 1% - 3%, and the activation conditions are: temperature 26 °C - 32 °C, rotation speed 160 rpm - 220 rpm, constant temperature shaking culture for 24 h - 48 h; in the LB liquid medium, the peptone concentration is 10 g / L, the NaCl concentration is 5 g / L - 10 g / L, the yeast extract concentration is 5 g / L, pH is 7.0, and the solvent is water.

[0013] As a general technical concept, the present invention also provides an application of the above-mentioned growth-promoting biocontrol bacteria in the rhizosphere of lettuce in controlling Meloidogyne incognita.

[0014] For the above application, preferably, the application includes the following steps: Prepare a fermentation broth of the growth-promoting biocontrol bacteria with an OD 600 not less than 1, and perform root irrigation treatment on diseased plants, with each plant receiving not less than 5 mL, once every other day, and the number of treatments being not less than 3 times.

[0015] For the above application, preferably, the preparation method of the fermentation broth of the growth-promoting biocontrol bacteria includes the following steps: (1) Inoculate the growth-promoting biocontrol bacteria into an LB liquid medium for activation; (2) Centrifuge the activated bacterial suspension, discard the supernatant, and resuspend it with sterile water to obtain the fermentation broth of the growth-promoting biocontrol bacteria.

[0016] For the above application, preferably, in step (1), the inoculation amount of the growth-promoting biocontrol bacteria is 1% - 3%, and the activation conditions are: temperature 26°C - 32°C, rotation speed 160 rpm - 220 rpm, constant temperature shaking culture for 24 h - 48 h; in the LB liquid medium, the peptone concentration is 10 g / L, the NaCl concentration is 5 g / L - 10 g / L, the yeast extract concentration is 5 g / L, the pH is 7.0, and the solvent is water.

[0017] Compared with the prior art, the advantages of the present invention are as follows: (1) The growth-promoting biocontrol bacteria of the present invention is Pseudomonas ureilytica GPBC-2, which is derived from the rhizosphere soil of plants, has a simple acquisition method, and is easy to culture. This growth-promoting biocontrol bacteria strain not only has a high growth-promoting effect on plants (or crops), for example, increasing plant height, stem girth, maximum leaf length and width, and at the same time making the root system more developed, but also has a high nematicidal activity against Meloidogyne incognita, can reduce the infection of Meloidogyne incognita on plant roots, and effectively alleviate the damage of Meloidogyne incognita to plants. The production cost of this strain is low, the production process is simple, and it will not cause pollution to the environment. In the future, it can be applied to the development of economic and green growth-promoting and biocontrol dual-effect bacterial fertilizers to achieve safe and pollution-free production of crops.

[0018] (2) The root irrigation treatment of tobacco with the fermentation broth of GPBC-2 bacteria of the present invention can significantly increase its biomass and significantly improve its agronomic traits. The plant height and root length are increased by 64.56% and 24.48% respectively compared with the control group, and the fresh weights of the above-ground and underground parts and the root-shoot ratio are increased by 110.23%, 272.73% and 77.60% respectively compared with the control group.

[0019] (3) The fermentation broth of GPBC-2 bacteria used in the present invention can achieve a mortality rate of nearly 80% after treating Meloidogyne incognita for 12 h, and a mortality rate of over 90% after treating for 24 h, indicating that the GPBC-2 strain has high nematicidal activity against Meloidogyne incognita. At the same time, this strain can effectively alleviate the damage of Meloidogyne incognita to peppers.

[0020] A growth-promoting biocontrol bacterium in the rhizosphere of lettuce, and the name of the growth-promoting biocontrol bacterium in the rhizosphere of lettuce is Pseudomonas urethralis ( Pseudomonas urethralis ) GPBC-2, and its preservation number at the China Center for Type Culture Collection is CCTCC NO: M 20232551. The address of the preservation unit is the China Center for Type Culture Collection (Wuhan University), No. 299, Bayi Road, Wuchang District, Wuhan City, Hubei Province, China, and the preservation date is December 13, 2023. Description of the Drawings

[0021] Figure 1 It is the microscopic morphology diagram of Pseudomonas urethralis GPBC-2 in Example 1 of the present invention.

[0022] Figure 2 It is the colony morphology diagram of Pseudomonas urethralis GPBC-2 on the LB agar plate in Example 1 of the present invention.

[0023] Figure 3 It is the transparent circle formed by Pseudomonas urethralis GPBC-2 on the PKO agar plate in Example 1 of the present invention.

[0024] Figure 4 It is the phylogenetic tree of Pseudomonas urethralis GPBC-2 in Example 1 of the present invention.

[0025] Figure 5 It is the growth-promoting effect diagram of Pseudomonas urethralis GPBC-2 on tobacco in Example 2 of the present invention. Among them, A is the largest leaf after being treated with sterile water (left) and GPBC-2 (right), and B is the whole plant phenotype after being treated with sterile water (left) and GPBC-2 (right).

[0026] Figure 6 It is the nematicidal activity diagram of Pseudomonas urethralis GPBC-2 against Meloidogyne incognita (J2s) at 12 h and 24 h respectively in Example 3 of the present invention. Among them, A is the morphological diagram of nematodes after being treated with sterile water for 12 h, B is the morphological diagram of nematodes after being treated with 200-fold diluted avermectin for 12 h, C is the morphological diagram of nematodes after being treated with GPBC-2 for 12 h, D is the morphological diagram of nematodes after being treated with sterile water for 24 h, E is the morphological diagram of nematodes after being treated with 200-fold diluted avermectin for 24 h, and F is the morphological diagram of nematodes after being treated with GPBC-2 for 24 h.

[0027] Figure 7This is the control effect diagram of Pseudomonas ureae GPBC-2 on Meloidogyne incognita in Example 4 of the present invention. Among them, A is the phenotype of the pepper root system after treatment with sterile water (left) and GPBC-2 (right), and the swollen part is the root knot; B is the column chart of the number of root knots of peppers treated with sterile water and GPBC-2.

[0028] Figure 8 This is the growth promotion effect diagram of Pseudomonas ureae GPBC-2 on peppers in Example 4 of the present invention. Among them, A is the phenotype of the above-ground part of the pepper treated with sterile water, and B is the phenotype of the above-ground part of the pepper treated with GPBC-2. Detailed implementation mode

[0029] The present invention will be further described below in conjunction with the specification drawings and specific preferred embodiments, but the protection scope of the present invention is not limited thereby. The materials and instruments used in the following embodiments are all commercially available.

[0030] Example 1 A growth-promoting biocontrol bacterium in the rhizosphere of lettuce of the present invention, and the name of this growth-promoting biocontrol bacterium in the rhizosphere of lettuce is Pseudomonas ureae GPBC-2 ( Pseudomonas urethralis GPBC-2), its preservation number in the China Center for Type Culture Collection is CCTCC NO: M 20232551, the address of the preservation unit is located at the China Center for Type Culture Collection (Wuhan University), No. 299, Bayi Road, Wuchang District, Wuhan City, Hubei Province, China, and the preservation date is December 13, 2023.

[0031] This strain is a Gram-negative bacterium, and is identified as Pseudomonas by molecular biology. Its main biological characteristics are: as Figure 1 shown, the cells are rod-shaped under the microscope and do not form spores; as Figure 2 shown, after culturing on an LB agar plate (pH 7.0) for 24 h, the colonies are light yellow, the surface is extremely smooth and wrinkled, easy to pick, and grow well under aerobic conditions; as Figure 3 shown, after culturing on a PKO modified inorganic phosphorus (tricalcium phosphate) agar plate (pH 7.4 ± 0.2) for 48 h, a transparent circle is formed around the colonies; the optimum growth temperature is 28°C. The 16S rDNA sequence of the strain is as shown in SEQ ID NO.1.

[0032] The method for isolating and purifying the growth-promoting biocontrol bacterium in this example includes the following steps: (1) Sampling: Collect soil samples from the rhizosphere of lettuce in Lianhua Town, Yuelu District, Changsha City, Hunan Province (but not limited to this), and transport them back to the laboratory in time. The soil samples are placed in 50 mL sterile centrifuge tubes and placed in a foam ice box. During transportation, the temperature of the soil samples is maintained at about 4°C.

[0033] (2)Isolation of plant rhizosphere soil microorganisms: Add 30 mL of PBS buffer containing surfactant TWEEN-20 (final concentration 1‰) to a 50 mL centrifuge tube containing plant roots and rhizosphere soil. After tightening the tube cap, vortex for 2 min. Transfer the vortexed soil homogenate to a 15 mL sterile centrifuge tube (10 mL / tube), centrifuge at 3900 rpm at room temperature for 5 min, discard the supernatant, add 2 mL of PBS buffer without TWEEN-20 to the tube, resuspend by pipetting, and serially dilute the resuspended solution to 10 -4 , take 50 μL of the diluted solution and spread it on a PKO agar plate, and culture at 28 °C for 3 d.

[0034] (3)Strain purification: Select colonies with clear zones, and perform purification operations on an LB agar plate by streaking in zones, and culture at 28 °C for 24 h until single colonies grow.

[0035] (4)Re-purification and qualitative detection of phosphate-solubilizing characteristics: Pick the purified single colonies, streak in zones on a PKO agar plate for re-purification, culture at 28 °C for 48 h, and if obvious clear zones appear around the colonies, it indicates that the strain has phosphate-solubilizing characteristics.

[0036] (5)Sequencing and construction of phylogenetic tree: Pick single colonies on the PKO plate for colony PCR, amplify the full length of the 16S rDNA of the strain using universal primers (27F / 1492R), send the original PCR solution to Tsingke Biotechnology Co., Ltd. for sequencing; perform sequence alignment of the sequenced DNA sequence on the NCBI website, and construct a phylogenetic tree using MEGA 11. As Figure 4 shown, the strain of the present invention is Pseudomonas urethralis.

[0037] (6)Cryopreservation: Pick an appropriate amount of purified strain and inoculate it into LB liquid medium with an inoculation amount of 2%, culture at 28 °C and 180 rpm with constant shaking for 24 h. Take 2 mL and transfer it to a sterile centrifuge tube, add 800 μL of the bacterial suspension and mix it evenly with 50% glycerol (protective agent) at a ratio of 1:1, and then cryopreserve it in a -80 °C ultra-low temperature refrigerator.

[0038] The formulations of the culture medium and buffer in the above steps are as follows: LB liquid medium: Peptone 10 g / L, NaCl 10 g / L, Yeast extract 5 g / L, pH 7.0, and the solvent is pure water.

[0039] LB agar medium: Add agar powder on the basis of LB liquid medium, and the content is 20 g / L.

[0040] PKO improved inorganic phosphorus agar medium: Glucose 10 g / L, (NH4)2SO4 0.5 g / L, NaCl 0.3 g / L, MgSO4·7H2O 0.3 g / L, MnSO4·H2O 0.03 g / L, KCl 0.3 g / L, FeSO4·7H2O 0.03 g / L, Ca3(PO4)2 5 g / L, yeast extract 0.5 g / L, agar powder 15 g / L, pH 7.4 ± 0.2, and the solvent is pure water.

[0041] PBS buffer: NaH2PO4 6.33 g / L, Na2HPO4 8.5 g / L, pH 6.5, and the solvent is pure water.

[0042] The above-mentioned medium and buffer are sterilized at 121°C under high temperature and high pressure for 15 min.

[0043] The PCR system and procedure in the above steps are as follows: (1) 50 μL system: Template (i.e., picking a small amount of bacteria), 1 μL of 27F, 1 μL of 1492R, 23 μL of sterile water, and 25 μL of 2×FineTaqTM PCR SuperMix (+dye); (2) Procedure: Pre-denaturation at 94°C for 10 min, (denaturation at 94°C for 30 sec, annealing at 52°C for 30 sec, extension at 72°C for 1.5 min, 30 - 35 cycles), sufficient extension at 72°C for 5 min, and constant temperature at 4°C.

[0044] Example 2 An application of the growth-promoting biocontrol bacteria in the rhizosphere of lettuce of the present invention in promoting crop growth, specifically, it is the experimental method for promoting the growth of tobacco using Pseudomonas ureae GPBC-2 in Example 1. This growth-promoting biocontrol bacteria can promote the growth of crops. In this example, tobacco is used as a model crop for research. It includes the following steps: (1) Activation: Take an appropriate amount of bacterial liquid from the cryopreservation tube containing Pseudomonas strains (such as the strains cryopreserved in Example 1), inoculate it into LB liquid medium (20 mL of medium in a 50 mL conical flask), the inoculation amount is 2%, at 28°C, 180 rpm, and incubate with constant shaking for 24 h.

[0045] (2) Preparation of the bacterial strain fermentation broth: Centrifuge the cultured bacterial liquid (12000 rpm, 4°C, 2 min), discard the supernatant, and resuspend the bacterial cells with sterile water to OD 600 = 1 to obtain the bacterial strain fermentation broth.

[0046] (3)Pot experiment: The tobacco plants were irrigated with the strain fermentation broth at a rate of 10 mL / plant, and sterile water was used as the negative control. The treatment was carried out every 5 days. After 30 days, the agronomic trait indicators and related biomass of the tobacco were measured and recorded to evaluate the growth rate and health status of the tobacco under different treatment conditions. Graphs were plotted using Office 2021 - Excel, and the data were statistically analyzed using IBM SPSS Statistics 27.

[0047] The results are as Figure 5 shown. Compared with the sterile water treatment group, after treatment with the GPBC - 2 strain fermentation broth, the agronomic traits of the tobacco were significantly improved, and the related biomass increased significantly. The specific data are shown in Tables 1 and 2, indicating that the strain fermentation broth has an obvious growth - promoting effect on the tobacco, and the strain GPBC - 2 is a highly efficient plant growth - promoting rhizobacterium.

[0048] Table 1 Effects of GPBC - 2 strain fermentation broth on the agronomic traits of tobacco

[0049] Table 2 Effects of GPBC - 2 strain fermentation broth on the biomass of tobacco

[0050] Example 3 An in vitro screening method for a growth - promoting biocontrol bacterium in the rhizosphere of lettuce of the present invention to control Meloidogyne incognita, using Pseudomonas ureae GPBC - 2 of Example 1, includes the following steps: (1)Activation: Take an appropriate amount of bacterial liquid from the cryopreservation tube containing Pseudomonas ureae GPBC - 2 (such as the strain cryopreserved in Example 1), inoculate it into the LB liquid medium (20 mL of medium in a 50 mL conical flask), with an inoculation amount of 2%, culture at 28 °C and 180 rpm with constant shaking for 24 h.

[0051] (2)Preparation of the strain fermentation broth: Centrifuge the cultured bacterial liquid (12000 rpm, 4 °C, 2 min), discard the supernatant, and resuspend the bacterial cells with sterile water to OD 600 = 2 to obtain the strain fermentation broth.

[0052] (3)Obtaining second - stage juveniles (J2s) of Meloidogyne incognita: Rinse the tobacco roots, pick nematode egg masses with forceps, surface - sterilize them with 1.0% NaClO, place them in a 90 mm glass petri dish, add sterile water, and incubate at room temperature for 3 days. Then collect the hatched J2s of Meloidogyne incognita. Observe and count the concentration under a microscope, and adjust it to 50 individuals / 20 μL.

[0053] (4)In vitro screening test: Add 80 μL of the treatment solution and 20 μL of the suspension of Meloidogyne incognita (containing 50 J2s) into a 96-well cell culture plate. Wrap it with tin foil and place it in an incubator at 28 °C for static incubation. Place the 96-well plate on the stage of an inverted microscope at 12 h and 24 h respectively to observe and record the death of J2s, and calculate the lethality rate. Judgment criterion: The nematodes that are rigid and motionless are dead, and those that are curved and soft are alive. In this experiment, sterile water was used as the negative control, and a 200-fold dilution of 1.8% abamectin emulsifiable concentrate (Zhiqingsong brand) was used as the positive control. Each treatment was repeated 3 times.

[0054]

[0055] The results are as Figure 6 shown. After treating Meloidogyne incognita with the GPBC-2 fermentation broth, J2s showed obvious lethality. After co-incubating the GPBC-2 fermentation broth with J2s for 12 h and 24 h, the lethality rates were 77.15% and 90.55% respectively. The specific data are shown in Table 3, indicating that the GPBC-2 fermentation broth has good nematicidal activity.

[0056] Table 3 Corrected lethality rate of GPBC-2 against J2s of Meloidogyne incognita

[0057] Note: Different lowercase letters in the same column of the table indicate significant differences (P < 0.05) among treatments at the 0.05 level.

[0058] Example 4 Application of the growth-promoting and biocontrol bacteria in the rhizosphere of lettuce of the present invention in controlling Meloidogyne incognita, specifically, a pot experiment method for controlling Meloidogyne incognita using Pseudomonas ureae GPBC-2 of Example 1. Meloidogyne incognita can infect peppers and cause serious diseases. In this pot experiment, Qiemen peppers (susceptible variety) were used as the model crop for research. It includes the following steps: (1) Activation: Take an appropriate amount of bacterial liquid from the cryopreservation tube containing Pseudomonas ureae strain (such as the strain cryopreserved in Example 1), inoculate it into LB liquid medium (20 mL of medium in a 50 mL conical flask), with an inoculation amount of 2%, at 28 °C, 180 rpm, and incubate with constant shaking for 24 h.

[0059] (2) Preparation of the bacterial strain fermentation broth: Centrifuge the cultured bacterial liquid (12000 rpm, 4 °C, 2 min), discard the supernatant, and resuspend the bacterial cells with sterile water to OD 600 = 1.5 to obtain the bacterial strain fermentation broth.

[0060] (3)Obtaining second-stage juveniles (J2s) of Meloidogyne incognita: Rinse the roots of tobacco plants grown in this laboratory. Use tweezers to pick up nematode egg masses, surface disinfect them with 1.0% NaClO, place them in a 90 mm glass petri dish, add sterile water, and incubate at room temperature for 3 days. Then collect the hatched J2s of Meloidogyne incognita. Observe and count the concentration under a microscope, and adjust it to 250 individuals / mL.

[0061] (4)When cultivating Qiemen peppers until the four true leaf stage, irrigate the roots with the GPBC-2 fermentation broth. The OD of the fermentation broth 600 = 2, 10 mL per plant, and treat once every 1 day for a total of 3 times. Use sterile water as the negative control and a 200-fold dilution of abamectin as the positive control. Two days after the third treatment, inoculate a suspension of J2s of Meloidogyne incognita (250 individuals / mL) 2 cm deep near the roots, 4 mL per plant. Each treatment is repeated 3 times. After 30 days, observe the growth phenotype of the plants and record the fresh weight of the roots to evaluate the growth-promoting effect of the GPBC-2 fermentation broth on Qiemen peppers; through fuchsin staining, observe and record the number of root knots, calculate the reduction rate of root knots, and evaluate the control effect of the GPBC-2 fermentation broth on Meloidogyne incognita.

[0062]

[0063] The results are as Figure 7 and Figure 8 shown: The GPBC-2 fermentation broth can effectively prevent the occurrence of Meloidogyne incognita diseases, with a reduction rate of root knots of 35.84%. After inoculating nematodes, the GPBC-2 fermentation broth alleviated the damage of nematodes to peppers, thereby promoting the growth of Qiemen peppers. The leaves in the above-ground part were larger and plumper than those treated with sterile water and a 200-fold dilution of abamectin. Treating with the GPBC-2 fermentation broth can significantly promote the root development of crops, the roots are more developed, and the fresh weight of the roots is also significantly higher than that treated with sterile water and a 200-fold dilution of abamectin. For specific data, see Table 4.

[0064] Table 4 Control effect of GPBC-2 on Meloidogyne incognita diseases

[0065] Note: Different lowercase letters in the same column of the table indicate significant differences between treatments at the 0.05 level (P < 0.05).

[0066] In summary, the present invention provides Pseudomonas ureae GPBC-2 and its application in promoting crop growth and controlling Meloidogyne incognita of crops. This strain was deposited at the China Center for Type Culture Collection on December 13, 2023, with the deposit number CCTCC NO: M 20232551, and the deposit address is the China Center for Type Culture Collection (Wuhan University), No. 299, Bayi Road, Wuchang District, Wuhan City, Hubei Province, China.

[0067] The present invention uses the method of irrigating the roots with the fermentation broth of strain GPBC-2 and finds that the fermentation broth of this strain can significantly improve the agronomic traits of tobacco. For example, it significantly increases the plant height, root length, maximum leaf length and width, and stem diameter, which are increased by 64.56%, 24.48%, 7.97%, 3.17% and 18.45% respectively compared with the control group. At the same time, it can significantly increase the fresh weight of the above-ground and underground parts of tobacco and significantly increase the root-shoot ratio, which are increased by 110.23%, 272.73% and 77.60% respectively compared with the control group, indicating that strain GPBC-2 is a highly efficient plant growth-promoting rhizobacterium.

[0068] After the present invention co-incubates strain GPBC-2 with Meloidogyne incognita (J2s) for 12 h and 24 h, obvious lethal effects are observed. Especially at 24 h, the lethality rate is as high as 90.55%, which indicates that strain GPBC-2 has high nematicidal activity against Meloidogyne incognita. At the same time, this strain can reduce the infection of nematodes on the plant roots and effectively alleviate the damage of Meloidogyne incognita to crops (Qiemen pepper was used as the test crop in the example).

[0069] It can be seen from this that Pseudomonas ureae GPBC-2 is a plant rhizosphere bacterium with both highly efficient growth-promoting and biocontrol effects, providing a theoretical basis and agricultural microbial germplasm resources for the development of multifunctional biological fertilizers for promoting crop growth and controlling Meloidogyne incognita, and having profound practical significance and broad application prospects.

[0070] The above are only the preferred embodiments of the present invention and do not impose any form of limitation on the present invention. Although the present invention has been disclosed above with preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art can make many possible changes and modifications to the technical solution of the present invention by using the above-disclosed methods and technical contents, or modify it into equivalent embodiments with equivalent changes. Therefore, any simple modification, equivalent replacement, equivalent change and modification made to the above embodiments based on the technical essence of the present invention without departing from the technical solution of the present invention still fall within the scope of protection of the technical solution of the present invention.

Claims

1. A growth-promoting and biocontrol bacterium in the rhizosphere of lettuce, characterized in that, The name of the growth-promoting biocontrol bacterium is Pseudomonas urethralis ( Pseudomonas urethralis ) GPBC-2, and its preservation number at the China Center for Type Culture Collection is CCTCC NO: M 20232551.

2. Use of the growth-promoting and biocontrol bacterium in the rhizosphere of lettuce as claimed in claim 1 in promoting plant growth.

3. The application according to claim 2, characterized in that, The said use comprises the following steps: Preparation of OD 600 Apply the growth-promoting biocontrol bacterial fermentation broth with a concentration not less than 1 to the roots of plants by irrigation, with each plant receiving not less than 5 mL, and treat once every 5 to 7 days for no less than 3 times.

4. The application according to claim 3, characterized in that The preparation method of the fermentation broth of the growth-promoting and biocontrol bacterium comprises the following steps: (1) Inoculate the growth-promoting and biocontrol bacterium into LB liquid medium for activation; (2) Centrifuge the activated bacterial suspension, discard the supernatant, and resuspend it with sterile water to obtain the fermentation broth of the growth-promoting and biocontrol bacterium.

5. The application according to claim 4, characterized in that, In step (1), the inoculation amount of the growth-promoting and biocontrol bacterium is 1% - 3%, and the activation conditions are: temperature 26°C - 32°C, rotation speed 160 rpm - 220 rpm, constant temperature shaking culture for 24 h - 48 h; in the LB liquid medium, the peptone concentration is 10 g / L, the NaCl concentration is 5 g / L - 10 g / L, the yeast extract concentration is 5 g / L, the pH is 7.0, and the solvent is water.

6. Use of the growth-promoting and biocontrol bacterium in the rhizosphere of lettuce as claimed in claim 1 in controlling Meloidogyne incognita.

7. The application according to claim 6, wherein The said use comprises the following steps: Preparation of OD 600 Use a biocontrol bacteria fermentation broth with a growth promotion effect of not less than 1 to perform root irrigation on diseased plants, with not less than 5 mL per plant, once every other day, and the number of times is not less than 3 times.

8. The application according to claim 7, characterized in that, The preparation method of the fermentation broth of the growth-promoting and biocontrol bacterium comprises the following steps: (1) Inoculate the growth-promoting and biocontrol bacterium into LB liquid medium for activation; (2) Centrifuge the activated bacterial suspension, discard the supernatant, and resuspend it with sterile water to obtain the fermentation broth of the growth-promoting and biocontrol bacterium.

9. The application according to claim 8, characterized in that In step (1), the inoculation amount of the growth-promoting and biocontrol bacterium is 1% - 3%, and the activation conditions are: temperature 26°C - 32°C, rotation speed 160 rpm - 220 rpm, constant temperature shaking culture for 24 h - 48 h; in the LB liquid medium, the peptone concentration is 10 g / L, the NaCl concentration is 5 g / L - 10 g / L, the yeast extract concentration is 5 g / L, the pH is 7.0, and the solvent is water.

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