A plant growth promoting rhizobacteria composition for improving salt stress tolerance of plants and application thereof

Abstract

The application discloses a kind of plant salt stress tolerance improving rhizosphere growth promoting bacteria composition and application thereof, belong to microbial application technical field.The rhizosphere growth promoting bacteria composition of the application includes Bacillus subtilis BK73 with accession number CGMCC NO.32725 and Enterobacter ludwigii FZ20 with accession number CGMCC NO.32550.Bacillus subtilis BK73 has the activity of producing ACC deaminase, can synthesize gamma-polyglutamic acid, and Enterobacter ludwigii FZ20 has the ability of nitrogen fixation, phosphorus dissolution and IAA production.Plant experiments show that the rhizosphere growth promoting bacteria composition provided by the application exerts the synergistic effect between strains, and has obvious promoting effect on the germination of corn seeds under salt stress, the growth of corn plants and the salt stress tolerance of corn plants.The application provides a new growth promoting bacteria composition for improving plant salt tolerance, and has wide application prospect.

Description

Technical Field

[0001] This invention relates to the field of microbial application technology, and in particular to a rhizosphere growth-promoting bacterial composition for improving the salt stress tolerance of plants and its application. Background Technology

[0002] Soil salinization is one of the causes of desertification and a major threat to soil function. The accumulation of salt in the soil due to salinization reduces crop yields, pasture quality, and tree survival rates, affecting farmers' and herders' livelihood choices and land use strategies, thus jeopardizing the sustainable development of agriculture, animal husbandry, and forestry. Simultaneously, salt accumulation accelerates vegetation and soil degradation, impairs the self-regulating capacity of ecosystems, endangers geological, resource, and national security, and ultimately leads to imbalances in socio-economic development. Currently, the global land area affected by salinization is approximately 1.0 × 10⁻⁶. 9 hm 2 The resulting economic losses amount to as much as 2.73 × 10⁻⁶ per year. 10 The amount is in US dollars, and the area of ​​salinized land is still increasing at an annual rate of 1.5 × 10⁻⁶. 7 -2.0×10 7 hm 2 The rate of salt accumulation in soil is increasing. Excessive accumulation of salt in any form will adversely affect plant growth and its physiological processes. Salt stress, as one of the major abiotic stresses in nature, causes osmotic stress, ion stress, and oxidative stress in plants, hindering their absorption of water and nutrients and inhibiting normal photosynthesis. This not only interferes with the normal physiological and metabolic processes of plants at various growth stages, such as seed germination, seedling growth, and reproduction, but also seriously affects the productive potential and function of plants in the ecosystem. Therefore, enhancing plant salt tolerance, promoting plant growth, and improving the utilization of saline-alkali land are not only important aspects of global land degradation and sustainable agriculture research in arid regions, but also an inevitable choice for ensuring ecological security, stabilizing economic growth, achieving a balance between arable land occupation and compensation, and solving population-resource-environment issues. Microbial measures are currently a research hotspot for saline-alkali land improvement. They utilize the combined action between plants and beneficial microorganisms in saline habitats to improve the soil. This approach enhances plant growth potential and induces salt tolerance while respecting the original genetic background of plants, and has advantages such as low cost, high efficiency, rapid results, and environmental friendliness.

[0003] Plant growth-promoting rhizobacteria (PGPR) are beneficial bacteria found in the rhizosphere soil of plants that inhibit plant diseases and promote plant growth. They are also the most widely used microorganisms in current microbial improvement measures. Numerous independent experimental studies have demonstrated, through analyzing changes in different salt tolerance indices, that PGPR can alter the physiological metabolic pathways of host plants, thereby alleviating the harm caused by salt stress. Some scholars have also classified and organized the effective PGPR strains and their salt tolerance characteristics used in existing studies, laying the foundation for the development and application of microbial fertilizers. Current research on rhizosphere growth-promoting bacteria focuses on screening strains with more prominent growth-promoting effects from known rhizosphere growth-promoting bacteria or developing new strains with growth-promoting effects. There is an urgent need to provide a rhizosphere growth-promoting bacteria composition obtained by combining different strains to achieve growth-promoting effects superior to the single-strain effects of any selected strain. Summary of the Invention

[0004] The purpose of this invention is to provide a rhizosphere growth-promoting bacterial composition and its application for improving the salt stress tolerance of plants, in order to solve the problems existing in the prior art. The rhizosphere growth-promoting bacterial composition provided by this invention can adapt to the saline-alkali environment, promote plant growth and improve the salt stress tolerance of plants under salt stress, and the growth-promoting effect is significantly stronger than that of single bacterial treatment.

[0005] To achieve the above objectives, the present invention provides the following solution:

[0006] This invention provides a rhizosphere growth-promoting bacterial composition for improving the salt stress tolerance of plants, comprising Bacillus subtilis BK73 and Enterobacter ludwigii FZ20;

[0007] The Bacillus subtilis BK73 is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO.32725, deposit date November 20, 2024, and deposit address No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing.

[0008] The Enterobacter lutea FZ20 was deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO.32550, deposit date November 11, 2024, and address No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing.

[0009] The present invention also provides the application of the rhizosphere growth-promoting bacteria composition described above in the preparation of bacterial agents that improve the salt stress tolerance of plants.

[0010] The present invention also provides a growth-promoting bacterial agent for improving the salt stress tolerance of plants, comprising the aforementioned rhizosphere growth-promoting bacterial composition.

[0011] Optionally, the growth-promoting agent has an absorbance value of not less than 1 at a wavelength of 600 nm.

[0012] Optionally, the ratio of viable Bacillus subtilis BK73 to viable Enterobacter leucis FZ20 is 1:1.

[0013] The present invention also provides the use of the rhizosphere growth-promoting bacterial composition or the growth-promoting bacterial agent described herein in any of the following:

[0014] (1) Application in the production of ACC deaminase;

[0015] (2) Application in the production of γ-polyglutamic acid;

[0016] (3) Application in promoting plant growth;

[0017] (4) Application in improving the salt stress tolerance of plants.

[0018] Preferably, the plant is corn.

[0019] The present invention also provides a method for promoting plant growth and / or improving plant salt stress tolerance, comprising the step of applying the rhizosphere growth-promoting bacterial composition or the growth-promoting bacterial agent to the plant growth environment.

[0020] Preferably, the plant is corn.

[0021] Preferably, the plant grows in a saline environment.

[0022] The present invention discloses the following technical effects:

[0023] The rhizosphere growth-promoting bacterial composition provided by this invention comprises Bacillus subtilis BK73 (CGMCC NO.32725) and Enterobacter leucovorin FZ20 (CGMCC NO.32550). Bacillus subtilis BK73 and Enterobacter leucovorin FZ20 exhibit no antagonistic activity and show good adaptability to saline-alkali soils. Bacillus subtilis BK73 possesses ACC deaminase activity and can synthesize γ-polyglutamic acid, while Enterobacter leucovorin FZ20 has nitrogen-fixing, phosphorus-solubilizing, and IAA-producing capabilities. Plant experiments show that the rhizosphere growth-promoting bacterial composition provided by this invention exerts a synergistic effect between the bacterial species, significantly promoting seed germination and plant growth under salt stress, as well as enhancing the plant's tolerance to salt stress. This invention provides a novel growth-promoting bacterial composition for improving plant salt tolerance and has broad application prospects. Attached Figure Description

[0024] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0025] Figure 1 The figure shows the antagonistic reaction test results of the rhizosphere growth-promoting bacteria composition Bacillus subtilis BK73 and Enterobacter leucis FZ20;

[0026] Figure 2 Figure 1 shows the results of the determination of the ACC deaminase production capacity of Bacillus subtilis BK73 in the rhizosphere growth-promoting bacteria composition.

[0027] Figure 3 The image shows the infrared spectrum detection results of γ-polyglutamic acid standard.

[0028] Figure 4 The infrared spectrum of Bacillus subtilis BK73 fermentation broth in the rhizosphere growth-promoting bacteria composition is shown in the figure.

[0029] Figure 5 Graph showing the high performance liquid chromatography (HPLC) results of γ-polyglutamic acid standard;

[0030] Figure 6 The graph shows the high-performance liquid chromatography (HPLC) results of Bacillus subtilis BK73 fermentation broth in the rhizosphere growth-promoting bacteria composition.

[0031] Figure 7 Figure 1 shows the results of nitrogen fixation capacity detection of Enterobacter leucis FZ20 in the rhizosphere growth-promoting bacteria composition.

[0032] Figure 8 Figure 1 shows the results of phosphorus solubilization of Enterobacter leucis FZ20 in the rhizosphere growth-promoting bacteria composition.

[0033] Figure 9 Figure 1 shows the results of IAA production capacity assay of Enterobacter leucis FZ20 in the rhizosphere growth-promoting bacteria composition.

[0034] Figure 10 The figure shows the phenotypic results of the effects of Bacillus subtilis BK73, Enterobacter leuciscus FZ20 and a combination of rhizosphere growth-promoting bacteria on maize seed germination under 300 mM salt stress; where Control represents the control group.

[0035] Figure 11 The figure shows the statistical results of the effect of Bacillus subtilis BK73, Enterobacter leuciscus FZ20 and rhizosphere growth-promoting bacteria combination on the germination rate of maize seeds under 300 mM salt stress; where Control represents the control group. ###*** indicates that compared with the control group, P < 0.001; *** indicates that compared with NaCl treatment, P < 0.001.

[0036] Figure 12 Phenotypic observation results of the effects of Bacillus subtilis BK73, Enterobacter leucis FZ20 and rhizosphere growth-promoting bacteria combination on maize growth under 300 mM salt stress; where Control is the control group;

[0037] Figure 13 The graph shows the statistical results of the effects of Bacillus subtilis BK73, Enterobacter leuciscus FZ20 and rhizosphere growth-promoting bacteria combination on maize biomass (root length, stem length, stem diameter, root weight and stem weight) under 300 mM salt stress; where Control represents the control group. # This indicates that, compared with the control group, P<0.05. ## * indicates that compared with the control group, P<0.01; * indicates that compared with NaCl treatment, P<0.05; ** indicates that compared with NaCl treatment, P<0.01; *** indicates that compared with NaCl treatment, P<0.001.

[0038] Figure 14 The figure shows the statistical results of the effects of a combination of Bacillus subtilis BK73, Enterobacter leucis FZ20, and rhizosphere growth-promoting bacteria on the physiological and biochemical indicators (malondialdehyde content, proline content, POD enzyme activity, SOD enzyme activity, and soluble sugar content) of maize under 300 mM salt stress; where Control represents the control group. # This indicates that, compared with the control group, P<0.05. ## This indicates that, compared with the control group, P < 0.01. ### * indicates P < 0.001 compared to the control group; * indicates P < 0.05 compared to NaCl treatment; ** indicates P < 0.01 compared to NaCl treatment; *** indicates P < 0.001 compared to NaCl treatment.

[0039] Figure 15 Scanning electron microscope images of the colonization of maize plants by a combination of Bacillus subtilis BK73, Enterobacter leucis FZ20, and rhizosphere growth-promoting bacteria at the root zone; where Control represents the control group. Detailed Implementation

[0040] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0041] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Any stated value or intermediate value within a stated range, as well as each smaller range between any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0042] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0043] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be readily apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0044] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0045] Bacillus subtilis is a species of bacterium in the genus Bacillus. It is thermophilic, aerobic, and spore-producing, exhibiting diverse physiological characteristics. It is easily isolated and cultured, widely distributed in nature, and is a non-pathogenic bacterium that is non-toxic and harmless to humans and animals, and does not pollute the environment. It produces abundant antibacterial substances, such as lipopeptides, peptides, phospholipids, polyenes, amino acids, and nucleic acids, which have good inhibitory effects on fungi, bacteria, viruses, and mycoplasma. Simultaneously, it produces growth hormones such as auxin, gibberellin, cytokinin, abscisic acid, and ethylene, promoting plant growth and thus improving plant development. Bacillus subtilis is one of the dominant microbial populations in soil and plant microecologies, possessing strong stress resistance and antibacterial, disease-preventing, and yield-increasing effects.

[0046] *Enterobacter ludwigii* is a Gram-negative bacterium belonging to the Enterobacteriaceae family. This strain is commonly found in the microecology of water, soil, and plant roots, and possesses certain growth-promoting properties, having been studied for applications in agriculture. It helps plants grow under adverse conditions through various mechanisms, including the production of plant hormones (such as indoleacetic acid), nitrogen fixation, and the decomposition of organic matter. *Enterobacter ludwigii* can also enhance plant growth in high-salt environments by regulating osmotic balance within plants, reducing the harmful effects of salt stress. Furthermore, it exhibits strong environmental adaptability, making it a promising plant growth-promoting strain.

[0047] In the inventor's previous research, a strain of Bacillus subtilis was isolated from the rhizosphere soil of rice and named Bacillus subtilis BK73. It was deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO. 32725, deposit date November 20, 2024, and deposit address No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing.

[0048] Meanwhile, a strain of Enterobacter luteolyticus was isolated and named Enterobacter luteolyticus FZ20. It was deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO.32550, deposit date November 11, 2024, and deposit address No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing.

[0049] The culture medium components and preparation method involved in this invention are as follows:

[0050] LB liquid medium: 10g tryptone, 5g yeast extract, 10g NaCl, bring to a final volume of 1L with ddH2O, and autoclave at 121℃ for 30min;

[0051] ADF medium was prepared according to the literature "Methods for isolating and characterizing ACC deaminase-containing plant growth-promoting rhizobacteria[J]. Physiologia Plantarum, 2003, 118(1):10-15."

[0052] Assabbey's nitrogen-free medium: KH2PO4 0.2 g, MgSO4·7H2O 0.2 g, NaCl 0.2 g, CaCO3 5.0 g, mannitol 10 g, CaSO4·2H2O 0.1 g, bring the volume to 1 L with ddH2O, adjust the pH to 7.0-7.5, and autoclave at 121℃ for 30 min.

[0053] PKO medium: 10g glucose, 0.5g (NH4)2SO4, 0.5g yeast extract, 0.2g NaCl, 0.3g KCl, 0.1g MgSO4·7H2O, 0.003g FeSO4, 0.03g MnSO4, 5.0g Ca3(PO4)2, 20.0g agar, bring the volume to 1L with distilled water, adjust the pH to 6.8-7.0, and autoclave at 115℃ for 30min.

[0054] Example 1

[0055] I. Preparation of Rhizosphere Growth-Promoting Bacterial Composition

[0056] 1. Interstrain antagonistic reaction test

[0057] The two tested bacterial strains were streaked with a cross, and their growth was observed after 1 day. Good growth indicated no antagonistic effect, while poor growth indicated antagonistic effect.

[0058] Test results as follows Figure 1 Both Bacillus subtilis BK73 and Enterobacter leucis FZ20 showed good growth, indicating that the two strains have no antagonistic effect, can coexist, and can be cultured together.

[0059] 2. Preparation of Rhizosphere Growth-Promoting Bacterial Composition

[0060] Bacillus subtilis BK73 and Enterobacter leucis FZ20 were streaked into LB medium and cultured at 30°C for 12 hours.

[0061] Single colonies were picked and inoculated into LB liquid medium and incubated in a shaker incubator (30℃, 220 r / min) for 5 h. After the strains had grown sufficiently, the OD value of the strain suspension was measured using a UV spectrophotometer. 600 The absorbance was measured by mixing Bacillus subtilis BK73 and Enterobacter leucis FZ20 in a 1:1 ratio.

[0062] II. Detection of functional activity of rhizosphere growth-promoting bacteria

[0063] 1. Determination of ACC deaminase activity in Bacillus subtilis

[0064] The strain was inoculated onto ADF medium, and the plates were incubated upside down at 30°C to observe whether the strain grew. If it could grow on ADF medium containing only ACC (1-aminocyclopropane-1-carboxylic acid) as the sole nitrogen source, it indicates that it can produce ACC deaminase. This ACC deaminase inhibits the production of α-butanone, a precursor to ethylene, thereby reducing ethylene production and minimizing plant inhibition.

[0065] Test results as follows Figure 2 As shown, Bacillus subtilis BK73 can grow on ADF medium, indicating that it can produce ACC deaminase.

[0066] 2. Qualitative and quantitative detection of γ-PGA production by Bacillus subtilis

[0067] γ-PGA is a high-molecular-weight glutamic acid polymer obtained by polymerizing α-amine and γ-carboxyl groups via γ-amide bonds. Polyamides have a series of characteristic absorption peaks, making them easily identifiable by Fourier transform infrared (FTIR) spectroscopy. By comparing the FTIR spectra of Bacillus subtilis BK73 fermentation broth with those of γ-PGA standards, it can be determined whether there is overlap between the two spectra. If there is overlap, it indicates that Bacillus subtilis BK73 produces γ-PGA.

[0068] FTIR detection results as follows Figure 3 and Figure 4 As shown, comparison Figure 3 and Figure 4 The two infrared spectra show that the characteristic absorption peaks of the two samples are basically consistent. At 3426.15 cm⁻¹, the extract... -l The absorption peak at this point is the NH symmetric stretching oscillation band. Due to the large number of hydrogen bonds on the γ-PGA molecule, VNH shifts to lower frequencies, resulting in a stronger and broader peak; wavenumber (σ / cm) -1 The 2000-1500 mM range is mainly characterized by stretching vibrations of double bonds such as C=C, C=O, and C=N. The extract measured 1636.00 cm⁻¹. -1 The absorption peak at 1403.47 cm⁻¹ corresponds to the stretching vibration band of C=O; the extract is at 1403.47 cm⁻¹. -1 The absorption peak at 1223.68 cm⁻¹ is due to the symmetric vibration of the carboxyl group (COOH). -1 This indicates the presence of -CH2 or -CH3 (bending vibration) in the molecular structure, at 2935.60 cm⁻¹. -1 The band at 900-500 cm⁻¹ represents the stretching vibration of saturated CH bonds; the broad absorption peak in the low-frequency region (900-500 cm⁻¹) is due to the planar rocking vibration and in-plane bending vibration of (CH₂)n (n>4). Spectral analysis combined with comparison to standard spectra confirms that this polymer is γ-PGA.

[0069] Next, high performance liquid chromatography (HPLC) was used to quantitatively analyze γ-PGA in the fermentation broth of Bacillus subtilis BK73.

[0070] HPLC identification results as follows Figure 5 and Figure 6As shown, by comparing the HPLC spectra of Bacillus subtilis BK73 fermentation solution and a commercial γ-polyglutamic acid standard, it was found that the UV spectra and the HPLC retention times of the target metabolite were consistent with the γ-polyglutamic acid standard. The results indicate that the metabolite secreted by Bacillus subtilis BK73 is γ-polyglutamic acid.

[0071] 3. Assay of the functional activity of Enterobacter leukemia FZ20

[0072] Enterobacter leucis FZ20 was activated and cultured in a shaker at 220 rpm and 30°C for 12 h. The nitrogen fixation, phosphorus solubilization, and IAA production capabilities of Enterobacter leucis FZ20 were then determined.

[0073] Nitrogen fixation: The selected strains were inoculated onto Assumption a nitrogen-free medium. Large, raised, slimy colonies were nitrogen-fixing bacteria.

[0074] Phosphate dissolution: The selected strains were inoculated onto PKO medium plates and cultured at 30°C for 3-5 days. The presence of a clear zone was then observed.

[0075] IAA production: Inoculate the selected strains with 100-400 μg / mL solution. -1 L-tryptophan was cultured in LB medium at (25±2)℃ for 48 h. After culture, the LB bacterial culture was centrifuged (8000 r / min, 10 min), and the supernatant was collected for analysis. 2 mL of the supernatant and 4 mL of Salkowski's reagent (50 mL 35% H2SO4 and 1 mL 0.5 mol·L⁻¹) were taken. -1 Mix the two solutions (FeCl3 solution and FeCl3 solution) thoroughly and let stand in the dark for 30 minutes. Observe; if it turns pink, it confirms the formation of IAA.

[0076] Nitrogen fixation test results as follows Figure 7 As shown, the phosphorus dissolution test results are as follows: Figure 8 As shown, the IAA test results are as follows: Figure 9 As shown above, the results indicate that Enterobacter leucis FZ20 possesses nitrogen fixation, phosphorus solubilization, and IAA production capabilities.

[0077] III. Detection of the plant growth-promoting function of rhizosphere growth-promoting bacteria composition

[0078] 1. Corn seed germination experiment

[0079] Using OD respectively 600The seeds were soaked in a solution of Bacillus subtilis BK73, Enterobacter rumenella FZ20, or a mixture of Bacillus subtilis BK73 and Enterobacter rumenella FZ20 (live count 1:1) overnight at 22°C. The next day, the liquid on the surface of the seeds was absorbed, and the seeds were placed in covered glass petri dishes with double layers of filter paper. 100 corn seeds were evenly placed in each dish, and 20 mL of 300 mmol / L NaCl solution was added to each dish. Each group was repeated 3 times.

[0080] Then, the petri dishes were placed in an incubator for a light duration of 16h / 8h (day / night) and a temperature of 22℃ to conduct a seed germination experiment. The germination status of the corn seeds was observed regularly. During the experiment, the NaCl treatment solution was changed every 24 hours to ensure the consistency of salt concentration between treatments.

[0081] Test results as follows Figure 10 and Figure 11 As shown, the results indicate that the growth-promoting bacterial composition has a better effect on maize seed germination than a single bacterial strain.

[0082] 2. Corn pot experiment

[0083] Select plump and uniformly sized corn seeds, soak them in 75% alcohol for 2 minutes to disinfect them, then treat them with 10% sodium hypochlorite for 10 minutes, and rinse them thoroughly with sterile water 5-6 times.

[0084] After disinfection, the seeds were soaked in sterile water for 24 hours. The disinfected seeds were then placed on water-soaked filter paper plates for germination. After 1-2 days of observation, seeds that had germinated and grown uniformly were selected and transferred to individual flowerpots. Each flowerpot contained only one seed. After transfer, an appropriate amount of sterile water was added to each flowerpot. The experimental group used 5 mL of bacterial fermentation broth (OD200). 600 =1.0) treatment. Incubate the flowerpots in an incubator at 22°C for 21 days, ensuring that sterile water is added regularly to maintain sufficient humidity.

[0085] Initially, the seedlings were irrigated daily with a 300 mM NaCl solution for 7 days. Then, every 2 days, 5 mL of bacterial fermentation broth was evenly poured around the roots. The corn seedlings were cultured at 22°C for 21 days under a 16 h light / 8 h dark light cycle, followed by various biomass and physiological and biochemical index tests.

[0086] Phenotypic observation results of maize seedlings are as follows: Figure 12 As shown in the figure, phenotypic observation revealed that the rhizosphere growth-promoting bacterial combination (Bacillus subtilis BK73 + Enterobacter leucis FZ20) was more effective than single-strain bacteria in promoting the growth of maize seedlings. The test results are as follows: Figure 13 and Figure 14As shown, biomass (root length, root weight, stem diameter, stem length, stem weight) and physiological and biochemical indicators (malondialdehyde content, proline content, POD enzyme activity, SOD enzyme activity, and soluble sugar content) were measured. The results showed that Bacillus subtilis and Enterobacter leucovorin, when applied individually to maize, significantly promoted maize seedling growth and stress tolerance. When Bacillus subtilis and Enterobacter leucovorin were combined and applied to maize seedlings, the combined bacteria significantly increased maize biomass, and was significantly superior to single-bacterial treatment in most physiological and biochemical indicators.

[0087] 3. Observation of the bacterial strain's root colonization

[0088] The results are as follows Figure 15 As shown, electron microscopy revealed that the number of rhizosphere microorganisms in the rhizosphere growth-promoting bacterial composition (Bacillus subtilis BK73 + Enterobacter leucis FZ20) was significantly greater than that in the single-bacterial group. This indicates that the rhizosphere growth-promoting bacterial composition provided by this invention plays a role in the physiological functions of plant rhizosphere and enhances the salt stress tolerance of maize seedlings.

[0089] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A rhizosphere growth-promoting bacterial composition for improving salt stress tolerance in maize, characterized in that, It is composed of Bacillus subtilis BK73 and Enterobacter ludwigii FZ20; The Bacillus subtilis BK73 is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO.32725, deposit date November 20, 2024, and deposit address No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing. The Enterobacter lutea FZ20 was deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO.32550, deposit date November 11, 2024, and address No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing.

2. The use of the rhizosphere growth-promoting bacterial composition of claim 1 in the preparation of a bacterial agent to improve the salt stress tolerance of maize.

3. A growth-promoting bacterial agent for improving the salt stress tolerance of maize, characterized in that, The rhizosphere growth-promoting bacteria composition according to claim 1 is the active ingredient.

4. The growth-promoting bacterial agent as described in claim 3, characterized in that, The growth-promoting agent has an absorbance value of not less than 1 at a wavelength of 600 nm.

5. The growth-promoting bacterial agent as described in claim 3, characterized in that, The ratio of viable Bacillus subtilis BK73 to viable Enterobacter leucis FZ20 is 1:

1.

6. The use of a rhizosphere growth-promoting bacterial composition as described in claim 1 or a growth-promoting bacterial agent as described in any one of claims 3-5 in any of the following: (1) Application in promoting plant growth in saline environments; (2) Application in improving the salt stress tolerance of plants; The plant in question is corn.

7. A method for promoting plant growth and / or improving plant salt stress tolerance, characterized in that, The step includes applying the rhizosphere growth-promoting bacterial composition of claim 1 or the growth-promoting bacterial agent of any one of claims 3-5 to the plant growth environment; The plant in question is corn; The plant grows in a saline environment.

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