Microbacterium alantarum and application of microbacterium alantarum in improvement of salt tolerance of crops

Abstract

The invention belongs to the technical field of microbial engineering, and particularly relates to microbacterium algal and application of the microbacterium algal to improvement of salt tolerance of crops, the microbacterium algal is preserved in the China Center for Type Culture Collection, and the preservation number is CCTCC NO: M 20242839. The strain has double growth promoting characteristics of high yield of siderophores and secretion of plant growth hormone, and can significantly relieve the inhibition effect of salt stress on crop seed germination, seedling emergence and growth. The microbacterium algal and the microbial agent provided by the invention have wide application prospects in the aspects of promoting seed germination, seedling emergence and growth of saline-alkali soil crops.

Description

Technical Field

[0001] The invention belongs to the technical field of microbial engineering, and particularly relates to a Microbacterium algeriensis strain and application thereof in improving the salt tolerance of crops. Background Art

[0002] Cultivating salt-tolerant crops in saline-alkali soils is an important way to develop and utilize saline-alkali land. Soybeans and rapeseed are dryland crops that are particularly advantageous for field cultivation in saline-alkali soils and are also recommended for cultivation in saline-alkali soils. However, soil salinization remains a major challenge facing agricultural production. Salt stress can damage cell membrane function, accumulate toxic substances, inhibit photosynthesis, and reduce nutrient acquisition, ultimately leading to cell death or even the death of the entire plant. Generally, when soil salt content exceeds 0.1%, the growth of common crops begins to be significantly affected; when soil salt content exceeds 0.3%, the yield of most crops decreases significantly. Seed germination and seedling stages are critical periods in plant growth and are also the most sensitive to salt stress. High concentrations of base ions can severely inhibit seed germination, seedling emergence, plant growth, and development, leading to problems such as difficulty in seedling emergence after sowing, post-emergence seedling death, weak growth, and missing seedlings and broken ridges.

[0003] Rhizosphere microorganisms, known as the "second genome" of plants, play a vital and irreplaceable role in helping plants resist stress, such as salt stress. Plant growth-promoting rhizosphere bacteria can directly promote plant growth under salt stress through processes such as biological nitrogen fixation, the production of auxin (IAA), and the activation of phosphorus and potassium nutrients in the soil. They can also alleviate damage to plants caused by salt stress by producing siderophores, secreting aminocyclopropane deaminase, and inducing systemic resistance, thereby improving plant adaptability to salt-stressed environments. Therefore, microorganisms or microbial agents show great potential in promoting salt-tolerant plant growth.

[0004] In view of the problems faced by soybean and rapeseed in the production process of saline-alkali land, such as difficulty in emergence, weak seedling growth, seedling death or stunted seedlings, and missing seedlings and broken ridges, it is necessary to screen out functional microorganisms with multiple life-promoting properties that can significantly promote crop seed germination, emergence, and seedling growth under salt stress conditions. Summary of the Invention

[0005] In order to solve the above problems, the present invention provides a strain of Microbacterium algeriensis and its application in improving the salt tolerance of crops. The Microbacterium algeriensis strain has the dual growth-promoting properties of high production of siderophores and secretion of plant growth hormones, which can significantly alleviate the inhibitory effects of salt stress on crop germination, emergence and growth.

[0006] To achieve the above purpose, the specific technical solutions of the present invention are as follows: The first aspect of the present invention provides a strain of Microbacterium algeriensis ( Microbacterium algeriense), the Microbacterium algeriensis was deposited in China Center for Type Culture Collection with the deposit number CCTCC NO: M 20242839.

[0007] The Microbacterium algeriensis strain is a high-yield iron carrier bacterium, and its 72-hour iron carrier secretion ability is rated as a high-yield level (++++); the strain can produce auxin (IAA, indoleacetic acid), and under the induction of L-tryptophan (500 mg / L), the average amount of indoleacetic acid secreted in 48 hours is 45.3 μg / mL.

[0008] A second aspect of the present invention provides a microbial agent, wherein the active ingredient of the microbial agent contains the above-mentioned Microbacterium algeriensis.

[0009] Furthermore, the number of viable bacteria of Microbacterium algeriensis in the microbial agent is ≥10 9 CFU / mL.

[0010] Furthermore, the microbial agent is a germination promoter or a growth promoter.

[0011] The third aspect of the present invention provides a method for preparing the above-mentioned microbial agent, comprising the following steps: inoculating the Algerian Microbacterium into LB liquid culture medium, culturing at 28°C to 30°C for 16h to 18h, and collecting the bacterial liquid; centrifuging the bacterial liquid to collect the bacterial cells, adding sterile water to the bacterial cells and resuspending the bacterial cells to obtain a bacterial suspension, and adjusting the number of viable Microbacterium Algerianum in the bacterial suspension to ≥10 9 CFU / mL, the microbial agent is obtained.

[0012] A fourth aspect of the present invention provides a use of the aforementioned Microbacterium algeriensis or microbial agent in improving plant salt tolerance.

[0013] Furthermore, the improvement of plant salt tolerance is the promotion of plant seed germination, plant seed emergence, plant seedling growth or increase in plant biomass under salt stress.

[0014] Furthermore, the plant seeds are soaked in the microbial agent for 2 to 4 minutes, and the microbial agent is applied to the soil so that the number of viable bacteria of Microbacterium algeriensis per gram of soil is ≥10 9 CFU, thereby promoting plant seed germination.

[0015] Furthermore, the microbial agent is used to irrigate the roots of plants so that the number of viable bacteria of Microbacterium algeriensis per gram of soil is ≥10 9 CFU, thereby promoting the growth of plant seedlings or increasing plant biomass.

[0016] Furthermore, the plant is a dryland crop.

[0017] Furthermore, the dryland crop is soybean or rapeseed.

[0018] Compared with the prior art, the present invention has the following beneficial effects: The present invention discloses for the first time a Microbacterium algeriensis strain with growth-promoting function ( Microbacterium algeriense ) M266-2. This strain has the dual growth-promoting properties of high iron carrier production and secretion of the plant growth hormone IAA, which can significantly alleviate the inhibitory effects of salt stress on crop germination, emergence, and growth. The microbial agent prepared using this Algerian Microbacterium M266-2 can significantly increase soybean emergence rate and functional leaf area under salt stress conditions, promote seedling growth, alleviate the inhibitory effects of salt stress on soybean emergence and seedling growth, and thus enhance soybean tolerance to salt stress. It can also enhance rapeseed seed germination vigor, specifically by promoting the growth of radicles and shoots under salt stress.

[0019] As a new microbial resource, Microbacterium algeriensis ( Microbacterium algeriense ) M266-2 and the microbial agents prepared therefrom have broad application prospects in promoting seed germination, emergence and growth of dryland crops such as soybeans and rapeseed in saline-alkali lands. This provides resources and technical support for the development of microbial agents or fertilizers suitable for agricultural applications in saline-alkali lands, and provides materials and technical support for the use of saline-alkali agriculture to produce good ecological and economic benefits.

[0020] Currently, for Microbacterium algeriensis ( Microbacterium algeriense Research on the Microbacterium algeriensis (Bacillus algeriensis) has remained at the taxonomic identification stage, with no reports on its application in saline-alkali land agriculture. Therefore, the present invention not only provides a strain of Microbacterium algeriensis with clear growth-promoting and salt-tolerance properties, but also opens up new application directions for this strain in saline-alkali land agriculture.

[0021] Microbacterium algeriensis M266-2 in the present invention is also known as Microbacterium algeriense , the classification is named: Microbacterium algeriense M266-2 was deposited in the China Center for Type Culture Collection (CCTCC) on December 18, 2024. The deposit address is: No. 299, Bayi Road, Wuchang District, Wuhan City, Hubei Province, Wuhan University, Postal Code: 430072, and the deposit number is CCTCC NO: M 20242839. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0023] Figure 1 This is the colony morphology of Microbacterium algeriensis M266-2 on LB solid medium.

[0024] Figure 2 This is a phylogenetic tree constructed based on the 16S rRNA gene sequence of Microbacterium algeriensis M266-2.

[0025] Figure 3 This is the growth of Microbacterium algeriensis M266-2 on the CAS detection plate.

[0026] Figure 4 This is a graph showing the effect of Microbacterium algeriensis M266-2 on promoting soybean emergence under 1.5w / v‰ NaCl stress; Figure 4 Figure A is a photograph of a replicate sample in each treatment group on the 5th day of treatment; Figure 4 Figure B shows the photographs of all samples in each treatment group on the 9th day of treatment. The 4 samples in the left column are repeated samples of the original soil control group (CK), the 4 samples in the middle column are repeated samples of the untreated group (NaCl) under 1.5w / v‰ NaCl stress, and the 4 samples in the right column are repeated samples of the experimental group (NaCl+M266-2) treated with a microbial agent containing the M266-2 strain under 1.5w / v‰ NaCl stress.

[0027] Figure 5 Statistical chart of soybean emergence rate under the treatments of original soil (CK), original soil with 1.5w / v‰ NaCl (NaCl), and original soil with 1.5w / v‰ NaCl and application of microbial agent containing M266-2 strain (NaCl+M266-2). Lowercase letters a and b indicate significant differences among the groups.

[0028] Figure 6 This figure shows the effect of Microbacterium algeriensis M266-2 on promoting soybean seedling growth under 1.5w / v% NaCl stress; the 4 samples in the left column are repeated samples of the original soil control group (CK), the 4 samples in the middle column are repeated samples of the untreated group (NaCl) under 1.5w / v‰ NaCl stress, and the 4 samples in the right column are repeated samples of the experimental group (NaCl+M266-2) treated with a microbial agent containing the M266-2 strain under 1.5w / v‰ NaCl stress.

[0029] Figure 7 Statistical graph of soybean plant height under the treatments of original soil (CK), original soil with 1.5w / v‰ NaCl (NaCl), and original soil with 1.5w / v‰ NaCl and application of microbial agent containing strain M266-2 (NaCl+M266-2); Figure 7 Figure A shows the statistical results on the 11th day after treatment. Figure 7 Figure B shows the statistical results on the 25th day after treatment; lowercase letters a, b, and c indicate significant differences between the groups.

[0030] Figure 8 Statistical graphs of soybean aboveground fresh weight, root fresh weight, and top third leaf area in the original soil (CK), original soil supplemented with 1.5w / v‰ NaCl (NaCl), and original soil supplemented with 1.5w / v‰ NaCl and application of a microbial agent containing the strain M266-2 (NaCl+M266-2); Figure 8 Figure A is a statistical chart of fresh weight of soybean aboveground parts. Figure 8 Figure B is a statistical chart of soybean root fresh weight. Figure 8 Figure C is a statistical chart of soybean inverted trefoil area; lowercase letters a, b, and c indicate significant differences between groups.

[0031] Figure 9 This is a plate image of Microbacterium algeriensis M266-2 on LB solid medium; Figure 9 Figure A shows the results of streak culture of the monoclonal strain M266-2; Figure 9 Figure B shows that M266-2 cells were cultured for 72 hours and then coated with 10 -4 Bacterial growth on LB solid medium of rhizosphere soil suspension.

[0032] Figure 10 This figure shows the effect of M266-2 on promoting rapeseed seed germination under salt stress. The seeds on the three plates in the same row are replicate samples. DETAILED DESCRIPTION

[0033] The specific embodiments of the present invention are described in detail below, but it should be understood that the scope of protection of the present invention is not limited by the specific embodiments. Based on the embodiments in the present invention, all other embodiments obtained by those of ordinary skill in the art without making creative work are within the scope of protection of the present invention. The experimental methods described in the embodiments of the present invention are conventional methods unless otherwise specified, and the materials, reagents, etc. used in the following embodiments can be obtained from commercial sources unless otherwise specified.

[0034] LB medium: 10 g of tryptone, 5 g of yeast extract, 10 g of sodium chloride (NaCl), 1000 mL of distilled water, pH 7.0, to prepare LB liquid medium. Add 15 g of agar powder to the LB liquid medium to prepare LB solid medium, and sterilize at 121°C for 20 min.

[0035] CAS solid medium: Dissolve 60.5 mg of chrome azurol S (CAS) in 50 mL of deionized water and mix with 10 mL of 1 mM FeCl₃ (prepared with 10 mmol / L HCl) to obtain component A. Dissolve 72.9 mg of HDTMA in 40 mL of deionized water to obtain component B. Slowly add component A to component B, sterilize at 115°C for 30 min, then mix with 900 mL of sterilized LB medium and pour onto CAS plates for later use.

[0036] CAS assay solution: Dissolve 60.5 mg of chrome azurol S (CAS) in 50 mL of deionized water and mix with 10 mL of 1 mM FeCl₃ (prepared with 10 mmol / L HCl) to obtain component A. Dissolve 72.9 mg of HDTMA in 40 mL of deionized water to obtain component B. Slowly add component A to component B, sterilize at 115°C for 30 min, and set aside.

[0037] Salkowski colorimetric solution: Mix 1 part 0.5 mol / L FeCl3 with 49 parts 35 v / v% HClO4 and store in the dark.

[0038] Soil salinization is a major challenge facing agricultural production. Salt stress can impair cell membrane function, accumulate toxic substances, inhibit photosynthesis, and reduce nutrient acquisition, ultimately leading to cell and even entire plant death. Generally, when soil salt content exceeds 0.1%, the growth of common crops begins to be significantly affected; when soil salt content exceeds 0.3%, the yield of most crops decreases significantly. Seed germination and seedling stages are critical periods in plant growth and are also the most sensitive to salt stress. High concentrations of base ions can severely inhibit seed germination, emergence, plant growth, and development, leading to problems such as difficulty in seedling emergence, post-emergence seedling death, weak growth, and missing or broken ridges. Rhizosphere microorganisms, known as the second genome of plants, play a vital and irreplaceable role in helping plants resist stress such as salt stress. Screening for functional strains with multiple growth-promoting properties that can enhance crop salt tolerance is crucial for alleviating the inhibitory effects of salt stress on crop germination, emergence, and growth.

[0039] The present invention provides a strain of Microbacterium algeriensis that significantly promotes plant growth under salt stress. Experiments have shown that this strain has the dual growth-promoting properties of high siderophore production and secretion of plant growth hormones, significantly alleviating the inhibitory effects of salt stress on crop germination, emergence, and growth. Specifically, it can promote soybean emergence after sowing, soybean seedling growth, rapeseed germination, and alleviate the inhibitory effects of salt stress on rapeseed radicle and shoot growth.

[0040] Example 1: Isolation, Identification and Preservation of Microbacterium algeriensis M266-2 Rhizosphere soil from Suaeda salsa was collected in June 2022 from a severely saline-alkali site in Binhai, Jiangsu Province. Three replicate samples were collected. A gradient dilution method was used to obtain soil suspensions at concentrations of 10⁻¹, 10⁻¹, and 10⁻¹. 100µL of each dilution was spread onto LB plates containing 2w / v‰ NaCl and incubated in a 28°C incubator for 3 days. Colonies of distinct morphology and color were selected for purification, numbering, and culture.

[0041] The purified strain was tested for its ability to produce the plant growth hormone indoleacetic acid (IAA) and siderophore, and eventually a bacterium that could secrete IAA and produce siderophore was screened out.

[0042] like Figure 1 As shown, after culturing this strain on LB solid medium at 28°C for 72 hours, colonies on the LB plate are milky white to pale yellow, with the color deepening with time. The colonies are smooth, moist, and shiny, with rounded shapes, neat edges, no wrinkles, easy to lift, and a uniform texture (non-slime). This strain is Gram-positive, aerobic, and weakly anaerobic. It is incapable of spore formation.

[0043] The 16S rRNA gene of this strain was amplified by PCR using the bacterial universal primers 27F and 1492R by the colony PCR method. The PCR product was sequenced by a sequencing company to obtain the 16S rRNA gene sequence shown in SEQ ID NO.1.

[0044] SEQ ID NO.1:

[0045] The nucleotide sequence of primer 27F is shown in SEQ ID NO. 2, and the nucleotide sequence of primer 1492R is shown in SEQ ID NO. 3; SEQ ID NO.2: 5'-AGAGTTTGATCMTGGCTCAG-3'; SEQ ID NO. 3: 5'-TACGGYTACCTTGTTACGACTT-3'.

[0046] The 16S rRNA gene sequence of the strain was uploaded to EzBioCloud (https: / / www.ezbiocloud.net / ) for homology comparison. The results showed that the 16S rRNA gene sequence of the strain was consistent with that of Microbacterium algeriensis ( Microbacterium algeriense ) The sequence similarity of G1(Type) is 99.57%, and further phylogenetic tree (such as Figure 2 As shown in Figure 2), the strain is also closer to the evolutionary distance of Microbacterium algeriensis, and the strain was identified as Microbacterium algeriensis ( Microbacterium algeriense ), named M266-2 in the present invention.

[0047] Example 2: Determination of the growth-promoting properties of Microbacterium algeriensis M266-2 Preparation of microbial inoculant: strain M266-2 was inoculated into LB liquid medium, cultured at 28°C and 180 rpm for 17 h, centrifuged, and resuspended in sterile water. The OD600 value of the bacterial suspension was adjusted to 1 to obtain a microbial inoculant with a viable count of 10 Microbacterium algeriensis in the inoculant. 9 CFU / mL.

[0048] To determine the IAA production capacity of the strain, 250 µL of the microbial inoculum was added to 5 mL of LB liquid medium containing 500 mg / L L-tryptophan and incubated at 28°C with shaking at 180 rpm for 48 hours. After centrifugation, 100 µL of the supernatant was added to a 96-well plate containing 100 µL of Salkowski colorimetric solution. Uninoculated culture medium was used as a blank control. The suspension was incubated in the dark for 30 minutes, and the absorbance at 530 nm was measured. The IAA content in the suspension was calculated using an IAA standard curve. The results are shown in Table 1. After 48 hours of incubation, strain M266-2 secreted as much as 45.3 ± 10.9 µg / mL of IAA.

[0049] Qualitative test of the siderophore secretion of the strain: Pipette 2 μL of microbial inoculum onto the center of the CAS plate, repeat 3 times, place the plate in a 28°C constant temperature incubator, and incubate for 4 days. Observe the size of the yellow halo outside the colony. The stronger the ability to secrete siderophores, the larger the halo. Figure 3As shown, M266-2 has a strong ability to secrete siderophores, and the value of its yellow halo diameter D / colony diameter d is 4.20.

[0050] Quantitative determination of siderophore secretion by strains: 300 μL of microbial inoculum was added to 3 mL of LB medium, cultured at 28°C and 180 rpm with shaking for 72 h, centrifuged, and the supernatant was mixed with equal volumes of CAS detection solution and allowed to stand for 1 h. The absorbance of the sample at 680 nm (As) was measured. The absorbance at 680 nm measured after mixing blank LB medium with CAS detection solution was taken as Ar.

[0051] ReferencesManjanatha MG, Loynachan TE, Atherly AG, Tn5 mutagenesis ofChinese Rhizobiumfredii For siderophore overproduction. Soil Biology and Biochemistry, 1992, 24(2): 151-155. Bacteria are classified as having high siderophore production capacity: the As / Ar ratio ranges from 1.0 to 0, with intervals of 0.2, and a + is added for every 0.2 decrease. The As / Ar ratio of strain M266-2 is 0.362, which is in the range of 0.2 to 0.4. Its siderophore secretion capacity is rated as ++++ (Table 1), reaching the high-yield level. Therefore, M266-2 is classified as a high-yield siderophore-producing bacterium.

[0052] Table 1 IAA and siderophore secretion ability of strain M266-2 Example 3: Microbacterium algeriensis M266-2 can promote soybean emergence under salt stress Soil collection: Soil from the 0 cm to 20 cm farmland plough layer was collected from the Xinyang Agricultural Experimental Station in Yancheng City, Jiangsu Province. The soluble salt content of the soil was 0.5 w / v‰.

[0053] Surface disinfection of soybean seeds: Disinfect the soybean seeds with 75% alcohol for 1 minute, wash them once with sterile water, then disinfect them with 2.5v / v% sodium hypochlorite for 2 minutes, wash them three times with sterile water, and use the water after the last wash to spread on the plate. After culturing for 4 days, no bacteria will grow on the plate, indicating that the disinfection is thorough.

[0054] Experimental treatments: 150 g (dry soil) of soil was weighed into a culture cup. Three treatments were set up: treatment with 1.5 w / v‰ NaCl (NaCl), treatment with 1.5 w / v‰ NaCl and microbial inoculum (NaCl + M266-2), and a control treatment without NaCl and inoculation (CK).

[0055] In the NaCl+M266-2 group, microbial agents were applied to the soil to make the inoculation concentration of M266-2 per gram of soil 10 9 Soybean seeds that had been surface sterilized were immersed in the microbial agent prepared in Example 2 and stirred to allow the Microbacterium algeriensis M266-2 strain to fully adhere to the surface of the seeds.

[0056] Soybean sowing and cultivation: Sow 5 soybean seeds of different treatments in a culture cup at a sowing depth of 1 cm. Cultivate soybeans in a smart greenhouse, control the daytime temperature at 25°C~30°C, and the nighttime temperature at 18°C~24°C, and add water by weighing every day to keep the soil moisture content at 60% SWHC. After the soybeans grow to a single leaf, count the emergence rate. Thin out the seedlings, retaining 2 seedlings of the same growth per cup. Then, follow the 10 9 CFU / g soil inoculation rate Microbial agents were applied by root irrigation to enhance the colonization effect of the strain. The emergence of soybean seedlings was observed every day during the culture period. The emergence of soybean seedlings after 5 and 9 days of culture was as follows: Figure 4 As shown in Figure 5 shown.

[0057] from Figure 4 and Figure 5 As can be seen from the results, NaCl treatment significantly inhibited soybean germination, but microbial inoculant application completely eliminated the inhibitory effect of NaCl stress on soybean germination. Specifically, NaCl reduced the soybean germination rate from 85% in the control treatment to 45%, while the microbial inoculant application increased the soybean germination rate to 90%, comparable to the control treatment without NaCl stress.

[0058] Example 4: Strain M266-2 promotes soybean seedling growth under salt stress Soil collection, soybean seed surface disinfection, experimental treatment, soybean sowing and culture conditions are the same as in Example 3. After 25 days of culture, the growth of soybeans is as follows: Figure 6 As shown in Figure 2, under NaCl stress, the growth of soybeans after microbial inoculation was significantly better than that of the non-inoculated treatment. The height of the aboveground part of soybeans in each treatment group was calculated. Figure 7 ; Determine the aboveground biomass, root biomass, and inverted third leaf area of ​​soybeans, and obtain Figure 8 .

[0059] NaCl treatment significantly reduced soybean plant height, aboveground fresh weight, root fresh weight and top third leaf area ( P<0.05), indicating that soybean growth was significantly inhibited under NaCl stress. The application of microbial agents effectively alleviated the inhibitory effect of NaCl stress on soybean growth, and in some indicators, the microbial agent treatment was even superior to the unstressed control treatment. Specifically, after 11 days of cultivation, the microbial agent application under NaCl stress restored the plant height to 9.07 cm, which is comparable to the control level ( Figure 7 Figure A). After 25 days of cultivation, under no NaCl stress, the soybean plant height was 23.5 cm; under NaCl treatment, the soybean plant height decreased to 20.1 cm; after the application of microbial inoculant, the soybean plant height was 28.6 cm, which was 22.0% higher than the no-NaCl control group ( Figure 7 Figure B). After 25 days of cultivation, under NaCl stress, the application of microbial agents restored the aboveground fresh weight and the area of ​​the third leaf to the same level as the control treatment without NaCl stress ( Figure 8 Figure A, Figure 8 Figure C), the root fresh weight recovered to 61.1% of the non-salt stress control treatment ( Figure 8 Figure B).

[0060] Therefore, the microbial agent containing the M266-2 strain can significantly alleviate the inhibitory effect of NaCl stress on soybean seedling growth, as shown by: after 25 days of cultivation, the aboveground fresh weight and the area of ​​the third leaf are restored to the same level as the salt-free stress treatment, and even the plant height exceeds that of the salt-free stress control treatment. It also has a significant effect on alleviating the inhibitory effect of NaCl stress on root growth.

[0061] Example 5: Stable colonization of Microbacterium algeriensis M266-2 in the soybean rhizosphere under salt stress Soil collection, soybean seed surface disinfection, experimental treatment, soybean sowing and culture conditions were the same as in Example 3. After 25 days of soybean culture, the soybean rhizosphere soil was collected. For the rhizosphere soil of the group treated with NaCl and microbial agent (NaCl + M266-2), the original rhizosphere soil 10 was prepared by gradient dilution method. -4 , 10 -5 , 10 -6 For each dilution of the suspension, take 100 μL of the suspension on an LB plate and spread the suspension evenly with a spreader. After incubation at 28°C for 3 days, observe the growth of the M266-2 strain.

[0062] According to the colony morphology characteristics of M266-2 ( Figure 9 ), observe the bacterial growth on the LB solid medium coated with soil suspension after 3 days of culture ( Figure 9(Figure B) A toothpick was used to pick individual colonies with morphological characteristics identical to those of strain M266-2 from LB solid medium coated with the soil suspension. These colonies were named NaCl+M266-2_1RS, NaCl+M266-2_2RS, NaCl+M266-2_3RS, and NaCl+M266-2_5RS, respectively. These colonies were then placed in a PCR reaction system and the 16S rRNA gene was amplified using universal bacterial primers 27F and 1492R. After verifying the integrity and length of the PCR products using polysorbate gel electrophoresis, the PCR products were sent to a sequencing company for sequencing. The resulting sequences were uploaded to the EzBioCloud database (https: / / www.ezbiocloud.net / ) for homology comparison.

[0063] The results showed that the 16S rRNA gene sequence of the colonies on the test plate was consistent with the model strain Microbacterium algeriense The sequence similarity of G1 was 99.07% to 99.36% (Table 2), and its taxonomic status was consistent with that of strain M266-2, confirming that the single colony on the test plate also belonged to Microbacterium algeriense (Microbacterium algeriensis).

[0064] Based on the colony morphology characteristics of M266-2 and the 16S rRNA gene identification results of the colonies, single colonies with the same colony morphology as M266-2 on the LB solid medium coated with the soil suspension were counted.

[0065] The results showed that strain M266-2 could still colonize in the soybean rhizosphere soil when soybeans were harvested, with the number of colonies per gram of soil reaching 0.68×10 7 CFU~1.70×10 7 CFU.

[0066] Table 2 Colonization detection of soybean M266-2 in rhizosphere soil after 25 days of cultivation under salt stress Example 6: Microbacterium algeriensis M266-2 promotes rapeseed seed germination under salt stress Preparation of Petri dishes: Place two layers of sterile filter paper in 9 cm diameter Petri dishes, prepare 15 in total.

[0067] The preparation method of the microbial agent is the same as that in Example 2.

[0068] Disinfection of rapeseed seeds: Select seeds with intact seed coat and uniform size, soak the seeds in 75v / v% ethanol for 30 seconds, and wash once with sterile water; then soak the seeds in 15w / v% H2O2 for 30 seconds, and wash three times with sterile water.

[0069] Seed sowing: The salt concentration was set to 150 mM NaCl (treatment labeled 150NaCl). The microbial inoculant was applied to the 150 mM NaCl solution at three doses: 1 v / v%, 10 v / v%, and 50 v / v% of the 150 mM NaCl solution. Sterile water was used as a control (treatment labeled H2O). Three replicates were set for each treatment. 10 mL of each treatment solution was added to a Petri dish lined with two layers of filter paper. Sterilized seeds were sown in each dish, with 15 seeds per dish. The dishes were incubated at 25°C, and radicle and shoot lengths were measured after 5 days.

[0070] Table 3 Radicle length and shoot length of rapeseed under different treatments Note: Lowercase letters a, b, and c in the table indicate significant differences between the groups.

[0071] As shown in Table 3 and Figure 10 As shown in the figure, 150mM NaCl significantly reduced the length of radicle and shoot compared with the control treatment of sterile water, which indicates that NaCl stress significantly inhibited the growth of radicle and shoot. Inoculation of 10v / v% microbial agent can significantly ( P <0.05) alleviated the inhibitory effect of NaCl on radicle and shoot growth. Specifically, NaCl stress caused the radicle length to drop significantly from 342.9 mm in the control to 8.1 mm, a decrease of 97.6%. After inoculation with 10 v / v% of the microbial agent, the radicle length increased to 21.2 mm, an increase of 1.61 times compared with the NaCl treatment. NaCl stress caused the shoot length to drop significantly from 164.3 mm in the control to 21.3 mm, a decrease of 87.0%. After inoculation with 10 v / v% of the microbial agent, the shoot length increased to 44.9 mm, an increase of 1.10 times compared with the NaCl treatment. Although inoculation with 1 v / v% and 50 v / v% of the microbial agent also increased the radicle length and shoot length compared with the NaCl treatment, neither reached a significant level ( P >0.05). Therefore, inoculation with 10 v / v% microbial agent can significantly alleviate the inhibitory effect of NaCl on the growth of rapeseed radicle and shoot.

[0072] It should be noted that when numerical ranges are mentioned in the present invention, it should be understood that both endpoints of each numerical range and any value between the two endpoints may be selected. Since the steps and methods used are the same as those in the embodiments, in order to avoid redundancy, the present invention describes preferred embodiments. Although preferred embodiments of the present invention have been described, those skilled in the art may make additional changes and modifications to these embodiments once they understand the basic inventive concepts. Therefore, the appended claims are intended to be interpreted as including the preferred embodiments and all changes and modifications that fall within the scope of the present invention.

[0073] Obviously, those skilled in the art may make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if such changes and modifications fall within the scope of the claims and their equivalents, the present invention is intended to include such changes and modifications.

Claims

1. A strain of Microbacterium algeriensis ( Microbacterium algeriense ), characterized in that, The Microbacterium algeriensis is deposited in the China Center for Type Culture Collection with a deposit number of CCTCC NO: M 20242839.

2. A microbial agent, characterized in that: The active ingredient of the microbial agent contains the Microbacterium algeriensis according to claim 1.

3. The microbial agent according to claim 2, characterized in that The number of viable bacteria of Microbacterium algeriensis in the microbial agent is ≥10 9 CFU / mL.

4. The microbial agent according to claim 2, characterized in that The microbial agent is a growth-promoting agent or a bud-promoting agent.

5. A method for preparing the microbial agent according to claim 2, characterized in that: The method comprises the following steps: inoculating the Algerian Microbacterium into LB liquid culture medium, culturing at 28°C to 30°C for 16 hours to 18 hours, and collecting the bacterial liquid; centrifuging the bacterial liquid to collect the bacterial cells, adding sterile water to the bacterial cells and resuspending the bacterial cells to obtain a bacterial suspension, and adjusting the number of viable bacteria of the Algerian Microbacterium in the bacterial suspension to ≥10 9 CFU / mL, the microbial agent is obtained.

6. Use of the Microbacterium algeriensis according to claim 1 or the microbial agent according to claim 2 for improving plant salt tolerance, characterized in that: The improvement of plant salt tolerance is to promote plant seed germination, plant seed emergence, plant seedling growth or increase plant biomass under salt stress.

7. The use according to claim 6, characterized in that Soak the plant seeds with the microbial agent for 2 to 4 minutes, and apply the microbial agent to the soil so that the number of viable bacteria of Microbacterium algeriensis per gram of soil is ≥10 9 CFU, thereby promoting plant seed germination.

8. The use according to claim 6, characterized in that The plant roots are irrigated with the microbial agent so that the number of viable bacteria of the Algerian Microbacterium per gram of soil is ≥10 9 CFU, thereby promoting the growth of plant seedlings or increasing plant biomass.

9. The use according to claim 6, characterized in that The plants are dryland crops.

10. The use according to claim 9, characterized in that The dryland crops are soybeans or rapeseed.

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