Salt-tolerant endogenous pantoea and application thereof

Abstract

The invention discloses salt-tolerant endogenous pantoea and application thereof, and relates to the technical field of microorganisms. According to the invention, a strain of endophytic pantoea TH1 is separated from young grape fruits, and the preservation number of the endophytic pantoea TH1 is CCTCC (China Center For Type Culture Collection) NO: M 2025960. The strain can promote the growth of grapes, improve the quality of the grapes, increase the content of vanillin in the grapes, increase the content of anthocyanin, flavonoid and flavanone in the grapes and reduce the content of tannin and total phenols in the grapes. Besides, the strain has the capabilities of efficiently dissolving potassium, fixing nitrogen, dissolving phosphorus and producing IAA (indoleacetic acid) and salt resistance, can improve plant photosynthesis, improve soil environment and promote plant growth under the condition of salt stress, and can also be used for preventing and treating grape powdery mildew at the same time.

Description

Technical Field

[0001] The present invention relates to the technical field of microorganisms, in particular to a salt-tolerant endophytic Pantoea strain and application thereof. Background Art

[0002] Vanillin (4-hydroxy-3-methoxybenzoic acid, C8H8O3), also known as vanillic aldehyde or vanillin, is an aromatic aldehyde compound extracted from vanilla beans and widely used in wine, spices, and other fields. In wine production, vanillin, as a primary aromatic compound, directly influences the aroma characteristics of the wine body and has a significant sensory impact. However, the vanillin content in wine grapes is currently low. Therefore, increasing the vanillin content in grapes is of great significance for improving the taste and enhancing the aroma of wine.

[0003] Currently, the preparation methods of vanillin include: plant extraction, chemical synthesis, and microbial transformation. Among them, the plant extraction method has low yield and relatively high production costs; the chemical synthesis method will cause environmental pollution, and the generated enantiomeric impurities will affect the purity of the product; while the microbial transformation method can break through the geographical limitations of the production of raw plant materials, and the production and synthesis processes can be adapted to local conditions, saving resources to the greatest extent, making it a current research hotspot. During the microbial transformation process, the substrates for the vanillin production pathway include ferulic acid, eugenol, and isoeugenol. Among them, ferulic acid is one of the main phenolic acids in wine grapes, with low toxicity and high yield. Using it as a reaction substrate to prepare vanillin through microbial transformation has attracted widespread attention.

[0004] Plant endophytes are bacteria or fungi that colonize within plant tissues without negatively impacting plant growth. Their applications primarily include regulating crop stress tolerance, developing medicinal secondary metabolites, bioremediation of polluted environments, and enhancing plant nutrient absorption. However, using endophytes to increase vanillin content in grapes has not been reported. Summary of the Invention

[0005] In view of the above prior art, the present invention aims to provide a salt-tolerant endophytic Pantoea and its application. The present invention isolates an endophytic Pantoea from young grape fruit ( Pantoea endophytica ) TH1, which can increase the vanillin content in grape berries, promote grape growth, improve grape quality, increase the content of anthocyanins, flavonoids, and flavanones in grapes, and reduce the content of tannins and total phenols in grapes. Furthermore, this strain can promote plant growth under salt stress, enhance plant photosynthesis, reduce disease incidence, and improve plant stress resistance.

[0006] To achieve the above object, the present invention adopts the following technical solutions: In a first aspect of the present invention, there is provided an endophytic Pantoea ( Pantoea endophytica )TH1, the strain was deposited in the China Center for Type Culture Collection (CCTCC) on May 6, 2025, and the deposit address is: Wuhan University, Bayi Road, Wuchang District, Wuhan City, Hubei Province; its deposit number is: CCTCC NO: M 2025960.

[0007] The endophytic Pantoea of ​​the present invention ( Pantoea endophytica TH1 was isolated from young Cabernet Sauvignon grapes and has the following characteristics: This strain can increase the vanillin content in grapes, promoting growth and improving grape quality. It also increases the content of anthocyanins, flavonoids, and flavanones, while reducing the content of tannins and total phenols. Furthermore, this strain is highly efficient in potassium dissolution, nitrogen fixation, phosphorus solubilization, IAA production, and salt tolerance. It can enhance plant photosynthesis, improve the soil environment, and promote plant growth under salt stress conditions. It can also be used to control powdery mildew in grapes.

[0008] The second aspect of the present invention provides a bacterial agent, wherein the bacterial agent is composed of the above-mentioned endophytic Pantoea ( Pantoea endophytica ) TH1 is the active ingredient.

[0009] As a preference, the endophytic Pantoea ( Pantoea endophytica ) TH1 exists in the form of cultured live bacteria, live bacteria fermentation broth or bacterial suspension.

[0010] Furthermore, endophytic Pantoea ( Pantoea endophytica ) TH1 bacterial suspension was prepared by the following method: Endophytic Pantoea ( Pantoea endophytica ) TH1 was inoculated into LB liquid culture medium, cultured at 37 °C and 180 rpm for 24 h, and then centrifuged to collect the bacteria. The bacteria were washed with distilled water to obtain a bacterial suspension.

[0011] Furthermore, endophytic Pantoea ( Pantoea endophytica ) The inoculum size of TH1 was 1% of the volume of LB liquid culture medium; the centrifugation speed was 10,000 rpm and the centrifugation time was 10 min.

[0012] The third aspect of the present invention provides the above-mentioned endophytic Pantoea ( Pantoea endophytica ) Use of TH1 or bacterial agents in any of the following 1)-4): 1) Increase the content of vanillin in grape berries; 2) Promote the growth of grapes and improve the quality of grapes; 3) Increase the content of anthocyanins, flavonoids and flavanones in grape skin; 4) Reduce the content of tannins and total phenols in grape skin.

[0013] A fourth aspect of the present invention provides the above-mentioned endophytic Pantoea ( Pantoea endophytica ) Use of TH1 or bacterial agents in preventing and controlling grape powdery mildew and / or preparing products for preventing and controlling grape powdery mildew.

[0014] A fifth aspect of the present invention provides the above-mentioned endophytic Pantoea ( Pantoea endophytica ) Use of TH1 or a bacterial agent in any of the following 1)-4): 1) Promote plant growth under salt stress; 2) Improve the photosynthetic capacity of plants under salt stress; 3) Improve the antioxidant capacity of plants under salt stress; 4) Improve soil environment, enzyme activity and soil nutrient content under salt stress.

[0015] Preferably, the plant is grape or corn.

[0016] Preferably, the soil environment includes soil enzyme activity and soil nutrient content.

[0017] Preferably, the soil enzymes include sucrase, FDA hydrolase and alkaline phosphatase.

[0018] Preferably, the soil nutrients include soil organic carbon, soil organic matter, soil available potassium, soil available phosphorus, soil alkaline nitrogen, soil total potassium, soil total sodium, soil total nitrogen and soil total phosphorus.

[0019] Beneficial effects of the present invention: 1. The present invention isolates and screens an endophytic strain of Pantoea ( Pantoea endophytica )TH1, this strain can significantly increase the vanillin content in grapes, promote grape growth, improve grape quality, increase the content of anthocyanins, flavonoids and flavanones in grapes, and reduce the content of tannins and total phenols in grapes.

[0020] 2. As a plant endophyte, the strain of the present invention has excellent salt tolerance, nitrogen fixation, phosphorus solubilization, potassium solubilization and IAA production capabilities.

[0021] 3. The endophytic Pantoea of ​​the present invention ( Pantoea endophytica ) TH1 can not only promote plant growth under salt stress conditions, improve the photosynthetic capacity and antioxidant capacity of plants under salt stress conditions, and increase soil enzyme activity and soil nutrient content under salt stress conditions, but also improve plant resistance to powdery mildew under salt stress conditions, thereby achieving effective prevention and control of grape powdery mildew. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 : Gram staining of the TH1 strain of the present invention; Figure 2: Phylogenetic tree based on 16S rRNA sequences; Figure 3 : Colony images of endophytic Pantoea TH1 cultured in combined nitrogen fixation medium for 1 day; Figure 4 : Colony images of endophytic Pantoea TH1 cultured in inorganic phosphate bacterial medium for 4 days; Figure 5 :The colony images of endophytic Pantoea TH1 cultured in organophosphate bacterial medium for 4 days; Figure 6 : Colony images of endophytic Pantoea TH1 cultured on desilicate medium for 4 days; Figure 7 :The color development of endophytic Pantoea TH1 under Salksowski colorimetric method; Figure 8 : Quantitative determination results of IAA production by endophytic Pantoea TH1; Figure 9 : In Experimental Example 1, the growth characteristics of grape seedling potted experiments were shown; Figure 10 : In Experimental Example 1, the results of the photosynthetic capacity of grape seedling potted experiments; Figure 11 : In Experimental Example 1, the results of the active oxygen scavenging system measurement in the grape seedling potted experiment; Figure 12 : In Experimental Example 1, the results of soil enzyme determination in the grape seedling pot experiment; Figure 13 : Effect of endophytic Pantoea TH1 on the growth performance of grape berries in Experimental Example 2; Figure 14 : Effect of endophytic Pantoea TH1 on vanillin content in Experimental Example 2; Figure 15 : Effect of endophytic Pantoea TH1 on the content of secondary metabolites in grape skin in Experimental Example 2; Figure 16 : Experimental example 3, corn seedling potted experimental results; Figure 17 : Antagonistic effect diagram against grape powdery mildew in Test Example 4. DETAILED DESCRIPTION

[0023] It should be noted that the following detailed descriptions are illustrative and intended to provide further explanation of the present application. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which the present application belongs.

[0024] In order to enable those skilled in the art to more clearly understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0025] The experimental materials used in the examples of the present invention are all conventional experimental materials in the art and can be purchased through commercial channels.

[0026] In the present invention, M9 glucose culture medium: 1 g of ammonium chloride, 0.13 g of magnesium sulfate, 3 g of potassium dihydrogen phosphate, 6 g of disodium hydrogen phosphate, and 0.338 g of glucose are dissolved in 1 L of distilled water and sterilized at 115° C. for 20 minutes.

[0027] The preparation method of FA selective liquid culture medium was referred to Wu Fenghui (2020). The combined nitrogen-fixing bacteria culture medium, silicate bacteria culture medium, organophosphorus bacteria culture medium, and inorganic phosphorus bacteria culture medium were purchased from Qingdao Haibo Biotechnology Co., Ltd., with the product numbers being HB8541, HB8548-1, HB8549-1, and HB8549-2, respectively.

[0028] Example 1: Isolation and purification of strains Young Cabernet Sauvignon grapes from the Cabernet Sauvignon grape experimental field at the Horticultural Experimental Station of Shandong Agricultural University (116.16'N, 36.17'E) were cleaned of surface debris by running water. One gram of grape sample was weighed, disinfected with 75% alcohol for 30 seconds, rinsed 3-5 times with sterile distilled water, rinsed in 0.1% mercuric chloride solution for 1 minute, and then rinsed 5 times with sterile distilled water. The sterilized and cleaned young Cabernet Sauvignon grapes were placed in a high-temperature sterilized mortar with an appropriate amount of sterilized quartz sand and 9 mL of sterile water. The plant sample was ground and allowed to stand for 30 minutes. Then, 100 μL of the supernatant was spread on M9 glucose medium and incubated at 28°C.

[0029] Single colonies with different morphological characteristics were selected for purification, and the salt tolerance of the purified strains was tested using LB culture medium containing different NaCl concentrations. The strain with the best salt tolerance (which could still grow on LB culture medium with a NaCl concentration of 5%) was selected, named TH1, and stored in a glycerol tube.

[0030] Example 2: Identification of strains 1. Morphological Identification of Strains The stable strain TH1 was stained with Gram and it was purple ( Figure 1 ), confirming it is a Gram-negative bacterium. Observed under an optical microscope, it exhibits a rod-shaped morphology. Strain TH1 grows at temperatures between 23°C and 37°C, with a pH range of 4 to 9, and has an optimal growth temperature of 28°C and an optimal pH of 7.

[0031] 2. 16S rDNA identification of strains: Using the three-zone streak method, single colonies were picked from the plates and activated in LB medium. The bacterial suspension was then sent to Shanghai Sangon Sequencing Co., Ltd. for sequencing. After sequencing, the sequencing results were submitted to NCBI for BLAST sequence alignment. 16S rRNA sequences of strains of similar species to the sequence were obtained from GenBank. Six to eight strains with at least 98% homology were selected and a phylogenetic tree was constructed using the neighbor-joining method using MEGA64 software. Phylogenetic analysis revealed that the strain was closely related to the endophytic Pantoea. Pantoea endophytica The sequence similarity is the highest, such as Figure 2 shown.

[0032] Combined with the results of morphological, physiological and biochemical identification and 16S rDNA identification, strain TH1 was identified as endophytic Pantoea ( Pantoea endophytica ), and made a biological deposit for patent procedure, the deposit information is as follows: Reference biological material (strain): TH1; Suggested taxonomic name: Pantoea endophytica Pantoea endophytica ; Deposit number: CCTCC NO: M 2025960.

[0033] Example 3: Growth-promoting properties 1. Test method: Endogenous Pantoea Pantoea endophytica ) TH1's growth-promoting ability was tested. Among them, the growth-promoting indicators include nitrogen fixation ability, potassium solubilization ability, phosphorus solubilization ability, phosphorus solubilization ability and IAA production ability. The results are as follows Figure 3-Figure 8 The specific detection method is as follows: The determination method of nitrogen fixation ability, potassium solubilization ability, phosphorus solubilization ability and phosphorus solubilization ability is as follows: the purified endophytic Pantoea ( Pantoea endophytica ) A single TH1 colony was inoculated into LB liquid culture medium and cultured with shaking at 37°C and 180 rpm for 24-48 h for activation. Then a single colony was inoculated into the corresponding culture medium for observation.

[0034] Among them, the combined nitrogen-fixing bacteria culture medium is used to determine the nitrogen fixation ability, the silicate bacteria culture medium is used to determine the potassium solubilization ability, the organophosphorus bacteria culture medium is used to determine the phosphorus solubilization ability, and the inorganic phosphorus bacteria culture medium is used to determine the phosphorus solubilization ability.

[0035] The IAA production capacity was determined by using the Salkowski colorimetric method for qualitative analysis, using LB liquid medium containing 200 mg / L L-tryptophan without inoculation as a blank control and IAA as a positive control; and using ultraviolet spectrophotometry for quantitative analysis.

[0036] 2. Experimental results: Depend on Figure 3 It can be seen that endophytic Pantoea ( Pantoea endophytica ) TH1 grew well on the combined nitrogen fixation medium, which showed that it had good nitrogen fixation ability. Figure 4 and Figure 5 It can be seen that endophytic Pantoea ( Pantoea endophytica ) TH1 grows on inorganic phosphorus bacterial culture medium and organic phosphorus bacterial culture medium respectively, and there is an obvious phosphorus-dissolving zone around the colony, which has good phosphorus-dissolving and solubilizing effects. Figure 6 It can be seen that endophytic Pantoea ( Pantoea endophytica ) TH1 was inoculated into silicate-dissolving medium, and the colony morphology was transparent oil droplets, indicating that it had a good potassium-dissolving effect.

[0037] Depend on Figure 7 It can be seen that the color of the solution is pink, which shows that the endophytic Pantoea ( Pantoea endophytica ) TH1 has the ability to produce IAA. Figure 8 It can be seen that after 72 h, the amount of IAA secreted by strain TH1 was approximately 55.414 mg / L.

[0038] Example 4: Preparation of endophytic Pantoea TH1 bacterial suspension The purified endophytic Pantoea TH1 was inoculated into LB liquid medium at a volume fraction of 1% and cultured in a constant temperature shaking incubator at 37°C and 180 rpm for 24 h. The fermentation broth was transferred to a sterilized centrifuge tube and centrifuged at 10,000 rpm for 10 min at room temperature. The supernatant was then carefully discarded and the cells were resuspended in sterile distilled water three times. The OD values ​​were then measured. 600 , the absorbance at 600 nm was adjusted to 1, and an endophytic Pantoea TH1 bacterial suspension was prepared.

[0039] Test Example 1: Growth promotion test of Cabernet Sauvignon grape seedlings under salt stress 1. Test method: Vermiculite and vineyard soil were mixed in a mass ratio of 1:1 and sterilized to create a mixed soil. One-year-old Cabernet Sauvignon seedlings were planted in the mixed soil. When the seedlings had 5-6 functional leaves, Cabernet Sauvignon seedlings of uniform growth and size were selected for treatment.

[0040] The experiment has 4 treatment groups, namely: Single inoculation treatment (TH1): 500 mL of clean water was poured into each pot of seedlings once every 5 days. After 3 days of watering, 50 mL of the endophytic Pantoea TH1 suspension prepared in Example 4 was added to each pot once a week, for a total of two waterings.

[0041] Single salt stress treatment (NaCl): 500 mL of 250 mM NaCl solution was poured into each pot of seedlings once every 5 days; 3 days after pouring the NaCl solution, 50 mL of clean water was added to each pot once a week, for a total of two pours of clean water.

[0042] Salt stress + bacteria treatment (NaCl + TH1): 500 mL of 250 mM NaCl solution was poured into each pot of seedlings once every 5 days. Three days after pouring with NaCl solution, 50 mL of the endophytic Pantoea TH1 bacterial suspension prepared in Example 4 was added to each pot once a week, for a total of two pourings.

[0043] Control treatment (CK): 500 mL of clean water was poured into each pot of seedlings once every 5 days; after 3 days of watering, 50 mL of clean water was added to each pot once a week, for a total of two waterings.

[0044] Each pot of seedlings was watered with 50 mL of water once a week for a total of two times. Each treatment group had 14 biological replicates and was treated for 28 days.

[0045] Sampling was taken to measure various indicators, including: growth indicators (plant height, stem diameter, root length, aboveground fresh weight, root fresh weight, aboveground dry weight, root dry weight), photosynthetic indicators (net photosynthetic rate, stomatal conductance, intercellular CO2 concentration, transpiration rate, water use efficiency, chlorophyll a content, chlorophyll b content, total chlorophyll content, carotenoid content), MDA content, hydrogen peroxide content, superoxide anion content, antioxidant enzyme (ascorbate peroxidase activity, peroxidase activity, superoxide dismutase activity) activity, and antioxidant substance (reduced glutathione) content.

[0046] In addition, the sucrase activity, FDA hydrolase activity, alkaline phosphatase, soil organic carbon content, soil organic matter content, soil available potassium content, soil available phosphorus content, soil alkaline nitrogen content, soil total potassium content, soil total sodium content, soil total nitrogen content and soil total phosphorus content of the treated soil were measured.

[0047] 2. Experimental Results Depend on Figure 9 It can be seen that under salt stress conditions, the endophytic Pantoea ( Pantoea endophytica ) The number of new leaves of Cabernet Sauvignon seedlings treated with TH1 bacterial suspension increased significantly, and the leaves were dark green. The plant height, stem diameter and root length were significantly increased, and the fresh weight and dry weight were also significantly increased. Figure 10 It can be seen that endophytic Pantoea ( Pantoea endophytica ) TH1 can not only significantly increase the content of chlorophyll a, chlorophyll b, total chlorophyll and carotenoids, but also increase the net photosynthetic rate and reduce the intercellular CO2 concentration. Therefore, it can be seen that strain TH1 can not only significantly increase the content of photosynthetic pigments in Cabernet Sauvignon seedlings, but also improve the photosynthesis of plants. Figure 11It can be seen that endophytic Pantoea ( Pantoea endophytica ) TH1 application can significantly enhance the active oxygen scavenging system. Figure 12 It can be seen that the inoculation of the strain significantly improved the soil environment under salt stress conditions.

[0048] Test Example 2: Field test on Cabernet Sauvignon fruit growth and secondary metabolites 1. Test method: This experiment was conducted at the Horticulture Experimental Station of Shandong Agricultural University, using perennial Cabernet Sauvignon grapes as the research subjects. On July 25, 2023, before veraison, 14 uniform bunches of fruit were selected for each treatment.

[0049] This experiment set up four treatment groups, as follows: Control group (CK): 20 mL of sterile water was evenly sprayed on the surface of the grape fruit for each bunch of grapes, once every other week, for a total of three treatments.

[0050] Group C1: concentration was 1×10 6 Endophytic Pantoea TH1 bacterial suspension at a concentration of CFU / mL; Group C2: concentration was 1×10 8 Endophytic Pantoea TH1 bacterial suspension at a concentration of CFU / mL; Group C3: concentration of 1×10 9 Endophytic Pantoea TH1 bacterial suspension with a concentration of CFU / mL.

[0051] The preparation method of the endophytic Pantoea TH1 suspension was the same as that in Example 4. 20 mL of the inoculum per bunch of grapes was evenly sprayed onto the surface of the grape fruit, spraying once every other week for a total of three treatments.

[0052] On August 31, 2023, when the grapes reach maturity, the grapes will be collected and various grape indicators will be measured, including: single grape weight and 100-grain weight, grape diameter and vertical diameter, grape skin anthocyanin content, grape skin tannin content, grape total phenol, flavonoid and flavanol content, and vanillin content. The method for detecting anthocyanin content in grape peels was based on the experimental methods of Lee J et al. (2017) and Ribereau-Gayon et al. (1965).

[0053] Grape peel tannin content was determined as follows: 1 g of grape peel was weighed into a 50 mL centrifuge tube, 20 mL of water was added, and the mixture was boiled in a water bath for 30 minutes. The extract was then filtered through a double layer of filter paper into a 50 mL beaker. 1 mL of the extract was transferred to a 10 mL centrifuge tube, 0.5 mL of Fc-Folin phenol reagent and 6 mL of water were added, and the mixture was mixed thoroughly. Immediately, 1 mL of 20% Na₂CO₃ solution was added, shaken, and the volume was made up to 10 mL with deionized water. After standing at room temperature for 30 minutes, the absorbance was measured at a UV wavelength of 680 nm (Geng Nana et al., 2013).

[0054] The assay steps for total phenolics, flavonoids, and flavanols in grapes were as follows: 1 g of grape peel was weighed into a 10 mL centrifuge tube, 4 mL of 1% hydrochloric acid-methanol solution was added, and extraction was performed under ultrasonic sealing for 30 minutes. The volume was then adjusted to 8 mL with water. The solution was incubated at 4°C overnight in the dark for 24 hours. The resulting solution was then centrifuged at 10,000 rpm at 4°C for 10 minutes. The original solution was stored in a refrigerator at 4°C in the dark. The determination of total phenolics and flavonoids was based on Petra et al., 2019; the determination of flavanols was based on Wang Yanping et al., 2017.

[0055] Vanillin content: Vanillin content was determined by high performance liquid chromatography. The specific steps are: (1) Preparation of vanillin standard solution: Weigh the vanillin standard separately, dissolve it in methanol (HPLC grade) to prepare a standard solution with a concentration of 0.01 mg / mL.

[0056] (2) Sample pretreatment: Grape pulp and peel samples (0.5 g each) were ground in liquid nitrogen, transferred to a pre-chilled centrifuge tube, and 5 mL of methanol was added. The tubes were shaken in the dark for 24 hours. After removal, the tubes were centrifuged at 10,000 rpm for 10 minutes, and the supernatant was filtered. 400 μL of the supernatant was aspirated and diluted to 2 mL with distilled water. An equal volume of dichloromethane was added, and the tubes were shaken for 9 hours. The supernatant was centrifuged and the extract was aspirated. The extract was evaporated in a metal bath at 50°C, and 300 μL of methanol was added for re-dissolution. 200 μL of the extract was then loaded onto the sample tube. Before loading, the sample was ultrasonically extracted for 30 minutes before loading onto the instrument.

[0057] (3) The vanillin content was detected using a SHIMADZU Nexera UHPLC LC-30A high performance liquid chromatograph. The detection wavelength of vanillin was 280 nm, the chromatographic column was a Hypersil BDS C18 column (250 mm*4.6 mm, 5 μm), the column temperature was 25°C, the flow rate was 1 mL / min, the injection volume was 20 μL, and the mobile phase was methanol:1% glacial acetic acid = 35:65 (V / V).

[0058] 2. Experimental conclusions: Depend on Figure 13 It can be seen that the bacterial suspension of TH1 strain can significantly increase the single fruit weight, 100-grain weight, and the horizontal and vertical diameters of the grape fruit.

[0059] Vanillin plays a key role in the growth, ripening and fruit quality formation of grapes. The present invention uses high performance liquid chromatography to determine the vanillin content in grape pulp and peel. The retention time of the vanillin standard solution is 6.94 minutes, and the content is calculated based on the peak area. Figure 14 It can be seen that the vanillin content in grape skin and pulp increased significantly after treatment with endophytic Pantoea TH1 compared with CK.

[0060] Tannins, anthocyanins, flavanones and total phenols in grape skin are key components that determine the quality of the fruit. Figure 15 It can be seen that compared with the CK group, the spraying concentration of 1×10 8 The bacterial suspension of endophytic Pantoea TH1 at a CFU / mL level can significantly increase the content of anthocyanins, flavonoids and flavanones, and reduce the content of tannins and total phenols.

[0061] Test Example 3: Experiment on promoting corn growth under salt stress 1. Test method: Vermiculite and cornfield soil were mixed in a mass ratio of 1:1 and sterilized to obtain mixed soil. Corn seedlings were planted in the mixed soil. When the seedlings grew to two leaves and one heart, corn seedlings of uniform growth and size were selected for treatment.

[0062] The experiment has 4 treatment groups, namely: Single inoculation treatment (TH1): Initially, the soil was irrigated thoroughly with clean water. After 3 days, 20 mL of the endophytic Pantoea TH1 suspension prepared in Example 4 was added to each pot of seedlings. The soil was irrigated once a week for a total of three times.

[0063] Single salt stress treatment (NaCl): At the beginning, the soil was drenched with the same amount of NaCl solution (250 mM) as that in the single inoculation treatment. After 3 days, each pot of seedlings was watered with 20 mL of clean water. Watering was done once a week for a total of three times.

[0064] Salt stress + bacteria treatment (NaCl + TH1): Initially, the soil was drenched with an equal amount of NaCl solution (250 mM) as in the single inoculation treatment. After 3 days, each pot of seedlings was irrigated with 20 mL of the endophytic Pantoea TH1 suspension prepared in Example 4. Watering was carried out once a week for a total of three times.

[0065] Control treatment (CK): At the beginning, the soil was irrigated with the same amount of clean water as that of the single inoculation treatment. After 3 days, each pot of seedlings was irrigated with 20 mL of clean water. Watering was done once a week for a total of three times.

[0066] Eight biological replicates were set up for each treatment. Samples were taken 27 days after treatment to measure various parameters, including plant height, stem diameter, aboveground fresh weight, belowground fresh weight, root length, dry weight, chlorophyll a content, chlorophyll b content, total chlorophyll content, and carotenoid content.

[0067] 2. Test results: like Figure 16 As shown, under salt stress conditions, the endophytic Pantoea ( Pantoea endophytica Corn seedlings inoculated with the TH1 bacterial suspension showed significantly higher biomass (total fresh weight, aboveground fresh weight, underground fresh weight, dry weight, plant height, and root length) than uninoculated corn seedlings. Furthermore, inoculation with the bacterial strain significantly increased the content of photosynthetic pigments in corn seedlings under salt stress.

[0068] Test Example 4: Antagonistic test on grape powdery mildew under salt stress 1. Test method: Vermiculite and vineyard soil were mixed in a mass ratio of 1:1 and sterilized to create a mixed soil. One-year-old Cabernet Sauvignon seedlings were planted in the mixed soil. When the seedlings had 5-6 functional leaves, Cabernet Sauvignon seedlings of uniform growth and size were selected for treatment.

[0069] Preparation of powdery mildew pathogen spore suspension: The pathogen of grape powdery mildew is Uncinaria officinalis. Grape leaves infected with powdery mildew were collected and powdery mildew spores were gently brushed into sterile water. After mixing, a spore suspension was obtained. The spore concentration in the suspension was adjusted to ≥ 3×10 6 / mL, set aside.

[0070] The experiment has 4 treatment groups, namely: Pathogen + bacterial agent treatment (powdery mildew + TH1): 500 mL of water was poured into each pot of seedlings, and 20 mL of powdery mildew pathogen spore suspension was sprayed on the front and back of the grape leaves. Three days later, 50 mL of the endophytic Pantoea TH1 bacterial suspension prepared in Example 4 was applied to each pot.

[0071] Salt stress + pathogen treatment (NaCl + powdery mildew): 500 mL of 250 mM NaCl solution was applied to each pot of seedlings, and 20 mL of powdery mildew pathogen spore suspension was sprayed on the front and back of the grape leaves. After 3 days, 50 ml of clean water was applied to each pot.

[0072] Salt stress + pathogen + bacterial agent treatment (NaCl + powdery mildew + TH1): 500 mL of 250 mM NaCl solution was applied to each pot of seedlings, and 20 mL of powdery mildew pathogen spore suspension was sprayed on the front and back of the grape leaves. Three days later, 50 mL of the endophytic Pantoea TH1 bacterial suspension prepared in Example 4 was applied to each pot.

[0073] Control treatment (CK+pathogen): 500 mL of clean water was poured into each pot of seedlings, and 20 mL of powdery mildew pathogen spore suspension was sprayed on the front and back of the grape leaves. After 3 days, 50 mL of clean water was applied to each pot.

[0074] Fourteen biological replicates were set up for each treatment group to observe the growth and resistance of the plants to powdery mildew under each treatment.

[0075] 2. Test results: The results are as follows Figure 17 As shown in the results, Cabernet Sauvignon seedlings inoculated with an endophytic Pantoea TH1 suspension under pathogen stress significantly increased plant height and alleviated powdery mildew stress compared to uninoculated Cabernet Sauvignon seedlings. Furthermore, under salt stress, the damage caused by the powdery mildew pathogen to Cabernet Sauvignon seedlings increased. However, under dual stress of salt and pathogens, inoculation with an endophytic Pantoea TH1 suspension still significantly alleviated powdery mildew stress and significantly increased plant height. These results indicate that endophytic Pantoea TH1 can enhance plant resistance to powdery mildew under salt stress.

[0076] The above description is merely a preferred embodiment of the present application and is not intended to limit the present application. Various modifications and variations are possible for those skilled in the art. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present application shall be included within the scope of protection of the present application.

Claims

1. An endophytic Pantoea ( Pantoea endophytica )TH1, its deposit number is CCTCC NO: M 2025960.

2. A bacterial agent, characterized in that The bacterial agent is the endophytic Pantoea ( Pantoea endophytica ) TH1 is the active ingredient.

3. The bacterial agent according to claim 2, wherein Among the bacterial agents, endophytic Pantoea ( Pantoea endophytica ) TH1 exists in the form of cultured live bacteria, live bacteria fermentation broth or bacterial suspension.

4. The use according to claim 3, characterized in that The bacterial suspension was prepared by the following method: Endophytic Pantoea ( Pantoea endophytica ) TH1 was inoculated into LB liquid culture medium, cultured at 37°C and 180 rpm for 24 h, and then centrifuged to collect the bacteria, which were resuspended in distilled water to obtain a bacterial suspension.

5. The endophytic Pantoea according to claim 1 ( Pantoea endophytica ) Use of the bacterial agent described in TH1 or any one of claims 2 to 4 in any one of the following 1) to 4): 1) Increase the content of vanillin in grape berries; 2) Promote the growth of grapes and improve the quality of grapes; 3) Increase the content of anthocyanins, flavonoids and flavanones in grape skin; 4) Reduce the content of tannins and total phenols in grape skin.

6. The endophytic Pantoea according to claim 1 ( Pantoea endophytica ) Use of the bacterial agent described in TH1 or any one of claims 2 to 4 in preventing and controlling grape powdery mildew and / or in preparing a product for preventing and controlling grape powdery mildew.

7. The endophytic Pantoea according to claim 1 ( Pantoea endophytica ) Use of the bacterial agent described in TH1 or any one of claims 2 to 4 in any one of the following 1) to 4): 1) Promote plant growth under salt stress; 2) Improve the photosynthetic capacity of plants under salt stress; 3) Improve the antioxidant capacity of plants under salt stress; 4) Improve soil environment under salt stress.

8. The use according to claim 7, characterized in that The plant is grape or corn.

9. The use according to claim 7, characterized in that Soil environment includes soil enzyme activity and soil nutrient content; soil enzymes include sucrase, FDA hydrolase and alkaline phosphatase.

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