Plant growth-promoting rhizobacteria for enhancing salt tolerance of rice and application of plant growth-promoting rhizobacteria

Abstract

The invention provides a growth-promoting rhizobacteria for enhancing salt tolerance of rice and application of the growth-promoting rhizobacteria, and belongs to the technical field of agricultural microorganisms. The invention provides bacillus amyloliquefaciens H1-225, and the preservation number of the bacillus amyloliquefaciens H1-225 is GDMCCNO.64977. The bacillus amyloliquefaciens The bacillus amyloliquefaciens H1-225 can improve the salt stress resistance of plants and relieve the damage of salt stress to the plants. The result of the embodiment shows that the bacillus amyloliquefaciens H1-225 can promote rice to germinate under salt stress, obviously improve the plant height, root length, overground part fresh weight, root fresh weight, overground part dry weight and root dry weight under the salt stress condition, and reduce the malondialdehyde content, so that the growth of the rice under the salt stress condition is promoted, and the rice quality is improved. The damage of salt stress to the rice is relieved.

Description

Technical Field

[0001] The present invention belongs to the technical field of agricultural microorganisms, and in particular relates to a rhizosphere growth-promoting bacterium capable of enhancing the salt tolerance of rice and an application thereof. Background Art

[0002] Soil salinization is one of the most destructive environmental stresses, leading to a significant decrease in cultivated land area and crop productivity and quality. Soil salinization is mainly affected by low precipitation, weathering of primary rocks, high surface evaporation, inorganic fertilizers, saline irrigation, and poor agronomic practices. Salinity stress is caused by water-soluble cations such as potassium (K), calcium (Ca), sodium (Na), and magnesium (Mg), as well as anions such as chloride, bicarbonate, sulfate, and carbonate. Based on the EC value, soils can be divided into non-saline (0-2dS m -1 ), slightly saline-alkali land (2-4dS m -1 ), moderate saline-alkali land (4-8dS m -1 ), strong saline-alkali land (8-16dS m -1 ), and very strong salt water (16dS m -1 ).

[0003] Removing salt from heavily saline-alkali soils is complex, requiring extended time, significant labor, and capital investment. Saline soils are often treated through chemical or physical measures. Chemical measures use calcium carbonates such as lime and gypsum as soil conditioners. Physical measures include leaching, soil addition, and mulching. Biological measures involve cultivating salt-tolerant varieties of crops such as barley, rice, wheat, mung beans, cotton, and rapeseed under salt-stress conditions. According to Morton et al., few salt-tolerance genes have been explored in breeding and biotechnology research to improve crop yields under both normal and saline-alkali soils. Over time, as research deepens, the central role of salt-tolerant rhizobacteria in improving soil health and enhancing crop productivity under soil stress conditions has become increasingly apparent. Salt-tolerant rhizobacteria possess significant advantages as bioaugmentants due to their ability to produce transport proteins, osmoregulators, and compatible solutes. These properties not only contribute to increased crop yields but also significantly enhance plant resistance to pathogens, thereby ensuring that soil fertility is effectively maintained even under extreme salt stress conditions.

[0004] Currently, there is limited information on the effectiveness of Bacillus amyloliquefaciens in improving the salt tolerance of crops. Summary of the Invention

[0005] The purpose of the present invention is to provide a Bacillus amyloliquefaciens strain that can significantly improve the salt stress tolerance of plants and alleviate the damage of salt stress to plants.

[0006] The purpose of the present invention is achieved through the following technical solutions:

[0007] The present invention provides a Bacillus amyloliquefaciens H1-225, with a deposit number of GDMCC No. 64977.

[0008] The present invention provides a microbial agent, comprising the Bacillus amyloliquefaciens H1-225 described in the above technical solution.

[0009] Preferably, the OD of Bacillus amyloliquefaciens H1-225 in the microbial agent is 600 The value of ≥1.0.

[0010] The present invention provides a method for preparing the microbial agent described in the above technical solution, comprising:

[0011] The Bacillus amyloliquefaciens H1-225 is cultured in a culture medium to obtain a microbial agent.

[0012] Preferably, the culture temperature is 30-37° C.; the culture time is 24-34 h; and the culture is accompanied by shaking, with the shaking speed being 150-230 rpm.

[0013] The present invention provides the use of the Bacillus amyloliquefaciens H1-225 described in the above technical solution in promoting plant growth and / or improving plant salt tolerance.

[0014] Preferably, the promoting plant growth includes promoting the improvement of any one or more of the following indicators (1) to (6):

[0015] (1) Plant height;

[0016] (2) root length;

[0017] (3) fresh weight of aboveground parts;

[0018] (4) fresh root weight;

[0019] (5) Aboveground dry weight;

[0020] (6) Root dry weight.

[0021] Preferably, the improving the salt tolerance of plants includes alleviating the damage of salt stress to plants and / or improving the ability of plants to tolerate salt stress.

[0022] The present invention provides a method for improving the salt tolerance of plants, comprising:

[0023] After the plants grow to the two-leaf and one-heart stage, the microbial agent comprising the Bacillus amyloliquefaciens H1-225 described in the above technical solution is applied.

[0024] Preferably, the administration is performed twice, and the time interval between the two administrations is two weeks.

[0025] Beneficial effects of the present invention

[0026] The present invention provides a Bacillus amyloliquefaciens H1-225, deposit number GDMCC NO.64977. The Bacillus amyloliquefaciens H1-225 can improve the salt stress tolerance of plants and alleviate the damage of salt stress to plants. The present invention shows through the results of examples that the Bacillus amyloliquefaciens H1-225 can promote the germination of rice under salt stress, significantly improve the plant height, root length, aboveground fresh weight, root fresh weight, aboveground dry weight, root dry weight of rice under salt stress conditions, and reduce the malondialdehyde content, thereby promoting the growth of rice under salt stress conditions and alleviating the damage of salt stress to rice. In summary, the Bacillus amyloliquefaciens H1-225 provided by the present invention is beneficial to increasing the yield of crops on saline-alkali land. Furthermore, the Bacillus amyloliquefaciens H1-225 can also improve the soil physical and chemical properties of saline-alkali land to a certain extent.

[0027] Biological Deposit Description

[0028] Bacillus amyloliquefaciens H1-225, classified as Bacillus amyloliquefaciens, was deposited in Guangdong Provincial Microbiological Culture Collection Center on May 8, 2024, with the address being 5th Floor, Experimental Building, No. 100 Xianlie Middle Road, Yuexiu District, Guangzhou City, Guangdong Province, with the deposit number being GDMCCNO.64977. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] 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. 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.

[0030] Figure 1 This is a statistical result diagram of the germination rate of rice under different salt concentrations in Example 2;

[0031] Figure 2 The graph shows the phenotypic observation results of the whole rice plant of the functional bacteria H1-225 and the blank control group under a salt concentration of 5‰;

[0032] Figure 3 The graph shows the phenotypic observation results of rice potted plants under a salt concentration of 5‰ for the functional bacteria H1-225 and the blank control group;

[0033] Figure 4The graph shows the phenotypic observation results of the whole rice plant of the functional bacteria H1-225 and the blank control group under a salt concentration of 8‰;

[0034] Figure 5 The graph shows the phenotypic observation results of rice potted plants under a salt concentration of 8‰ for the functional bacteria H1-225 and the blank control group;

[0035] Figure 6 This is the result diagram of the effects of 8 functional bacteria on rice plant height;

[0036] Figure 7 This is the result diagram of the effect of 8 functional bacteria on rice root length;

[0037] Figure 8 This is the result diagram of the effects of 8 functional bacteria on the fresh weight of rice aboveground parts;

[0038] Figure 9 This is the result diagram of the effects of 8 functional bacteria on the fresh weight of rice roots;

[0039] Figure 10 This is the result diagram of the effects of 8 functional bacteria on the dry weight of rice aboveground parts;

[0040] Figure 11 This is the result diagram of the effects of 8 functional bacteria on the dry weight of rice roots;

[0041] Figure 12 This is the result diagram of the effects of 8 functional bacteria on the malondialdehyde content in rice;

[0042] Figure 13 This is the phylogenetic analysis result of Bacillus amyloliquefaciens H1-225. DETAILED DESCRIPTION

[0043] The present invention provides a Bacillus amyloliquefaciens H1-225, with a deposit number of GDMCC NO.64977. In the present invention, the Bacillus amyloliquefaciens H1-225 is derived from the rhizosphere of the halophyte Spartina alterniflora. The nucleotide sequence of 16S rDNA of the Bacillus amyloliquefaciens H1-225 is shown in SEQ ID NO.1. The Bacillus amyloliquefaciens H1-225 has been deposited in the Guangdong Provincial Microbial Culture Collection Center on May 8, 2024, with a deposit number of GDMCC NO.64977.

[0044] The Bacillus amyloliquefaciens H1-225 provided by the present invention can improve the salt stress tolerance of crops in saline-alkali land and alleviate the damage caused by salt stress to plants. The present invention shows through the results of examples that the Bacillus amyloliquefaciens H1-225 can promote the germination of rice under salt concentration stress of 5‰ and 8‰, significantly improve the plant height, root length, aboveground fresh weight, root fresh weight, aboveground dry weight, root dry weight of rice under salt stress conditions, and reduce the physiological and biochemical indicators such as malondialdehyde content, thereby promoting the growth of rice under salt stress conditions and alleviating the damage caused by salt stress to rice.

[0045] The present invention provides a microbial agent, comprising the Bacillus amyloliquefaciens H1-225 described in the above technical solution. In the present invention, the OD of the Bacillus amyloliquefaciens H1-225 in the microbial agent is 600 The value of ≥1.0.

[0046] The present invention provides a method for preparing the microbial agent described in the above technical solution, comprising:

[0047] The Bacillus amyloliquefaciens H1-225 is cultured in a culture medium to obtain a microbial agent.

[0048] As an optional embodiment of the present invention, the culture medium includes TSB liquid culture medium; the culture temperature can be 30-37°C, or 30, 31, 32, 33, 34, 35, 36 or 37°C; the culture time can be 24-34h, or 24, 25, 26, 27, 28, 29, 30, 31, 32, 33 or 34h; the culture process is accompanied by oscillation, and the oscillation speed can be 150-230rpm, or 150, 160, 170, 180, 190, 200, 210, 220 or 230rpm.

[0049] After the culture is completed, the culture fluid of Bacillus amyloliquefaciens H1-225 is obtained. The culture fluid of Bacillus amyloliquefaciens H1-225 can be directly used as a microbial agent; the bacteria in the culture fluid of Bacillus amyloliquefaciens H1-225 can also be separated and resuspended to obtain a bacterial suspension as a microbial agent. The present invention has no special limitation on the separation method, and any conventional method in the art can be used. As an optional embodiment of the present invention, the separation method can be centrifugation. After the bacteria are separated and obtained in the present invention, the bacteria are resuspended with sterile water to obtain a bacterial suspension. As an optional embodiment of the present invention, the OD value of Bacillus amyloliquefaciens H1-225 in the bacterial suspension is 600 The value of is ≥1.0 and can also be 1.0.

[0050] The present invention provides the use of the Bacillus amyloliquefaciens H1-225 described in the above technical solution in promoting plant growth and / or improving plant salt tolerance. As an optional embodiment of the present invention, the plant can be rice. As an optional embodiment of the present invention, the promoting plant growth includes promoting plant growth under salt stress conditions. The promoting plant growth includes promoting the improvement of any one or more of the following indicators (1) to (6): (1) plant height; (2) root length; (3) aboveground fresh weight; (4) root fresh weight; (5) aboveground dry weight; (6) root dry weight. In the present invention, the improving plant salt tolerance includes alleviating the damage of salt stress to plants and / or improving the ability of plants to tolerate salt stress. The present invention shows through the results of the examples that the Bacillus amyloliquefaciens H1-225 can promote rice growth under 5‰ and 8‰ salt concentration stress conditions, and improve rice plant height, root length, aboveground fresh weight, root fresh weight, aboveground dry weight and root fresh weight. Furthermore, the Bacillus amyloliquefaciens H1-225 can also significantly reduce the malondialdehyde content of rice under salt stress conditions, thereby helping to improve the salt stress tolerance of rice.

[0051] The present invention provides a method for improving the salt tolerance of plants, comprising:

[0052] After the plant grows to the stage of two leaves and one heart, the microbial agent comprising the Bacillus amyloliquefaciens H1-225 described in the above technical solution is applied. As an optional embodiment of the present invention, the timing of the application can be within 1 to 2 weeks after the plant grows to the stage of two leaves and one heart. As an optional embodiment of the present invention, the number of applications can be 2 times; the time interval between the 2 applications is two weeks; the application amount of the Bacillus amyloliquefaciens H1-225 in each application is 10 7 CFU / g soil, that is, 10 7 CFU of Bacillus amyloliquefaciens H1-225.

[0053] In order to further illustrate the present invention, the technical solution provided by the present invention is described in detail below with reference to the accompanying drawings and embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0054] The experimental methods in the following examples, unless otherwise specified, are conventional methods. The experimental materials used in the following examples, unless otherwise specified, were purchased from conventional biochemical reagent stores. The quantitative tests in the following examples were all repeated three times, and the results were averaged.

[0055] Example 1 Construction of a bacterial strain resource library

[0056] Halophyte materials: Suaeda salsa, Sesbania dasyphylla, and Tamarix chinensis were collected from a farm near the Chinese Elk Park in Dafeng District, Yancheng City (32°59′46″N, 120°49′48″E); the halophyte Spartina alterniflora was collected from the mudflats near the Yellow Sea National Forest Park in Dafeng District, Yancheng City (32°52′25″N, 120°54′24″E).

[0057] Using the macro-culture omics method, the rhizosphere samples of halophytes were diluted with buffer (PBS-S) to a limiting dilution gradient of 10 -6 , 10 -7 The rhizosphere sample suspension was cultured in TSB liquid medium in a 96-well plate. After 14 days of culture, a 96-well plate with a microbial growth probability of 30% to 50% was selected. Under a sterile environment, 80 μL of bacterial solution was aspirated from the wells with growing microorganisms and added to a PCR tube containing 80 μL of 40% glycerol aqueous solution. The mixture was gently pipetted and mixed, and the culture was labeled and stored in a -80°C freezer to obtain a library of halophytic rhizosphere bacterial strains.

[0058] The experimental results showed that 3991 effective fungi were isolated from the rhizosphere of four halophytes: Sesbania truncatula, Suaeda salsa, Spartina alterniflora and Tamarix chinensis.

[0059] Example 2 Screening of functional strains

[0060] 1. Determination of critical salt concentration for rice seed germination

[0061] 1. Select plump rice seeds of uniform size for surface disinfection.

[0062] After soaking in an ethanol aqueous solution with a volume fraction of 75% for 1 minute, the ethanol on the surface of the seeds was washed off with sterile water, and then soaked in a sodium hypochlorite aqueous solution with a mass fraction of 2% for 30 minutes, and rinsed with sterile water 6 times.

[0063] 2. Soak the rice seeds surface-sterilized in step 1 in sterile sodium chloride solutions of different salt concentrations (0 mM, 40 mM, 80 mM, 120 mM, 160 mM, 200 mM, 240 mM) respectively. Each treatment was repeated three times, with 10 seeds in each repeat. Soak in the dark at 25°C for 4 h.

[0064] 3. Place the seeds soaked in step 2 in a culture dish containing sterile moistened filter paper of the corresponding salt concentration and place them in a 25°C light incubator for germination with a day / night ratio of 16h / 8h for 7 days.

[0065] 4. Complete step 3, calculate the germination rates of rice seeds with different treatments (germination rate (%) = (number of normally germinated seeds on the 7th day of germination / number of test seeds) × 100), and find the critical salt concentration for rice seed germination.

[0066] Experimental results: According to the statistical results of rice germination rate under different salt concentrations ( Figure 1 ), and 200 mM was determined to be the critical salt stress concentration for rice germination.

[0067] 2. Screening of Fungal Pores

[0068] Test strains: 3991 effective pores isolated in Example 1.

[0069] 1. Place the surface-sterilized rice seeds in a 240 mM sodium chloride solution at 25°C in the dark for 4 hours. The control group was soaked in sterile water.

[0070] 2. Place the soaked seeds in step 1 in culture dishes containing sterile water and 240 mM sodium chloride solution and lined with filter paper, with 3 seeds in each culture dish, and perform three sets of biological replicates.

[0071] 3. Pipette 10 μL of bacterial solution from each PCR tube of the halophyte bacterial strain resource library constructed in Example 1 into a 2 mL centrifuge tube containing 1 mL of TSB liquid medium, gently shake and mix, and culture in a shaker at 30°C and 170 rpm until the OD 600 =1.0.

[0072] 4. Add 100 μL of the corresponding bacterial solution from step 3 directly above each treated rice seed in step 2. That is, add the bacterial solution obtained from step 3 above one effective bacterial well directly above each treated rice seed, number them and distinguish them. Place them in a 25°C light incubator for germination with a day / night ratio of 16h / 8h for 4 days.

[0073] 5. Observe the germination of different treatments in step 4 and preliminarily screen out the fungus holes that can promote rice germination under salt stress conditions.

[0074] Bacillus amyloliquefaciens SQR9 (accession number is CGMCC NO.5808, published in ZL201710797445.0) was used as a positive control, salt-only treatment without bacteria was used as a negative control, and neither salt nor bacteria was used as CK.

[0075] Experimental results: Based on the statistical results of the germination rate of rice after inoculation with different bacterial wells at a salt concentration of 240mM, 33 bacterial wells that can promote rice germination under salt stress were screened out.

[0076] 3. Purification of strains

[0077] Based on the initial screening results of the germination test under salt stress, the fungus pores that can promote rice germination under 240mM salt stress were found for isolation and purification of single bacteria.

[0078] 1. Pipette the bacterial solution from the wells obtained in the initial screening and spread it on a TSB plate. Incubate it upside down at 30℃ for 24 hours.

[0079] 2. Select the single colonies that grow rapidly and have different morphologies in step 1, and continue to streak the remaining colonies on TSB plates until single colonies appear.

[0080] 3. Each single colony in step 2 was then streaked three times on TSB plates.

[0081] 4. Pick a single colony from step 3 and place it in a test tube containing 3 mL of TSB liquid medium. Shake the culture at 30°C and 170 rpm until the liquid becomes turbid.

[0082] 5. Pipette 80 μL of the bacterial solution from step 4 into a 40% glycerol aqueous solution tube and store in a -80°C refrigerator.

[0083] Based on the preliminary screening results of crop germination experiments under salt stress, a single bacteria resource library was constructed.

[0084] Experimental results: 8 strains were purified that can promote rice germination under salt stress.

[0085] Example 3 Potted plant verification of functional strains

[0086] Strains to be tested: the 8 strains screened in Example 2 and Bacillus amyloliquefaciens SQR9.

[0087] Soil material for the test: saline-alkali soil in Cangdong Farm, Yancheng City, Jiangsu Province, divided into two salinity gradients (5‰ and 8‰), the soil type is paddy soil, the pH of 5‰ saline-alkali soil is 7.84, and the pH of 8‰ saline-alkali soil is 7.80.

[0088] 1. Preparation of bacterial suspension

[0089] 1. The 8 functional strains obtained by screening and the control strain SQR9 (i.e., Bacillus amyloliquefaciens SQR9) were inoculated into TSB liquid culture medium and cultured at 30°C and 170 rpm for 24 h to obtain the culture liquid of each strain, referred to as the culture liquid for short.

[0090] 2. After culturing for 24 hours, centrifuge the bacterial suspension and resuspend it with sterile water to make the OD 600 The value of is 1.0.

[0091] 2. Germination and seedling cultivation of rice seeds

[0092] 1. First, the surface of the rice seeds needs to be sterilized. This step is performed in a clean bench. Select plump and basically uniform-sized Nanjing 46 rice seeds and sterilize them with an alcohol solution with a volume fraction of 75% ethanol for 1 minute. Then, soak the rice seeds in a sodium hypochlorite solution with a mass fraction of 2% sodium hypochlorite for 30 minutes. Rinse the rice seeds with sterile ultrapure water six times to wash away the sodium hypochlorite solution attached to the seed surface.

[0093] 2. Place the soaked rice seeds in a petri dish containing sterile water and covered with sterile filter paper, and germinate them in the dark at 25°C for 4 days.

[0094] 3. Select the rice seedlings with the same growth in step 2 and transplant them into the rice seedling soil for seedling cultivation. When the rice grows to the stage of two leaves and one heart, select the rice seedlings with the same growth and transplant them into the designated soil for subsequent experiments.

[0095] 3. Potted Experiment Design

[0096] This experiment was conducted in the greenhouse of the Baima Research Base of Nanjing Agricultural University in Nanjing, Jiangsu Province from November 2023 to January 2024, with an experimental period of 28 days.

[0097] Two salt gradients were set up in this experiment: 5‰ and 8‰. Each pot was loaded with 500g of soil. The experiment set up a control group and a treatment group. The control group pots were not inoculated with bacteria (blank control group) and inoculated with SQR9 (i.e. Bacillus amyloliquefaciens SQR9, as the positive control group), and the treatment groups were inoculated with 8 functional strains.

[0098] 1. Select rice seedlings with consistent growth from step 2 and transplant them into saline soil with two salinity gradients (5‰ and 8‰). Plant five rice seedlings per pot, and set up three control pots for each treatment. Cultivate at room temperature and water appropriately to maintain soil moisture.

[0099] 2. One week after rice transplanting, 8 functional bacteria suspensions and the bacterial suspension of strain SQR9 were used to irrigate the seedling roots, maintaining the accession concentration of each bacteria at 10 7 CFU / g soil, that is, 10 7 The specific steps are as follows: one rice treatment group is inoculated with one bacterium, the blank control group is not inoculated with bacteria, and sterile water equal to the amount of bacterial suspension is added, and the positive control group is inoculated with Bacillus amyloliquefaciens SQR9.

[0100] 3. After two weeks, add the bacterial suspension in the same manner. Irrigate each pot of the control group with the same volume of sterile water. Continue growing for two weeks at 30°C with 16 / 8h light conditions.

[0101] 4. After completing step 3, measure the various physiological and biochemical indicators of the rice plants.

[0102] The malondialdehyde (MDA) content was detected using a commercially available kit (purchased from Nanjing Jiancheng Biotechnology Co., Ltd.).

[0103] The test results of various physiological and biochemical indicators of rice plants are shown in Tables 1-2 and Figures 2 to 12 In the figure, CK is the blank control group, SQR9 is the positive control group, and TJ3-65, TJ3-85, TJ3-92, TJ3-104, TJ3-120, J2-15, J2-16, and H1-225 are the strain numbers of the eight functional bacteria.

[0104] Figure 2 The phenotypic observation results of the whole rice plants of the functional bacteria H1-225 and the blank control group under a salt concentration of 5‰ are shown. The three plants on the left are the blank control group; the three plants on the right are the functional bacteria H1-225 treatment group. Figure 3 The phenotypic observation results of rice potted plants treated with functional bacteria H1-225 and blank control group under 5‰ salt concentration are shown. The two pots on the left are blank control group; the two pots on the right are treated with functional bacteria H1-225. Figure 4 The phenotypic observation results of the whole rice plants of the functional bacteria H1-225 and the blank control group under a salt concentration of 8‰ are shown. The three plants on the left are the blank control group; the three plants on the right are the functional bacteria H1-225 treatment group. Figure 5 These are the phenotypic observation results of rice potted plants treated with functional bacteria H1-225 and blank control group at a salt concentration of 8‰. The two pots on the left are blank control group, and the two pots on the right are functional bacteria H1-225 treatment group.

[0105] Figure 6 Figure 1 is a graph showing the effects of eight functional bacteria on rice plant height; A shows the effects of eight functional bacteria on rice plant height at a salt concentration of 5‰; and B shows the effects of eight functional bacteria on rice plant height at a salt concentration of 8‰.

[0106] Figure 7 Figure 1 is a graph showing the effects of eight functional bacteria on rice root length; A shows the effects of eight functional bacteria on rice root length at a salt concentration of 8‰; and B shows the effects of eight functional bacteria on rice root length at a salt concentration of 5‰.

[0107] Figure 8 Figure 1 is a graph showing the effects of eight functional bacteria on the fresh weight of the aboveground part of rice. Figure A shows the effects of eight functional bacteria on the fresh weight of the aboveground part of rice at a salt concentration of 5‰. Figure B shows the effects of eight functional bacteria on the fresh weight of the aboveground part of rice at a salt concentration of 8‰.

[0108] Figure 9 Figure 1 is a graph showing the effects of eight functional bacteria on the fresh weight of rice roots; A shows the effects of eight functional bacteria on the fresh weight of rice roots at a salt concentration of 5‰; and B shows the effects of eight functional bacteria on the fresh weight of rice roots at a salt concentration of 8‰.

[0109] Figure 10 Figure 1 is a graph showing the effects of eight functional bacteria on the dry weight of the aboveground part of rice. Figure A shows the effects of eight functional bacteria on the dry weight of the aboveground part of rice at a salt concentration of 5‰. Figure B shows the effects of eight functional bacteria on the dry weight of the aboveground part of rice at a salt concentration of 8‰.

[0110] Figure 11 Figure 1 is a graph showing the effects of eight functional bacteria on the dry weight of rice roots; A shows the effects of eight functional bacteria on the dry weight of rice roots at a salt concentration of 5‰; and B shows the effects of eight functional bacteria on the dry weight of rice roots at a salt concentration of 8‰.

[0111] Figure 12 The figure shows the effects of 8 functional bacteria on the malondialdehyde content of rice. A shows the effects of 8 functional bacteria on the malondialdehyde content of rice at a salt concentration of 5‰. B shows the effects of 8 functional bacteria on the malondialdehyde content of rice at a salt concentration of 8‰.

[0112] Table 1. Test results of various physiological and biochemical indicators of rice plants in each treatment group under 5‰ salt concentration (mean)

[0113]

[0114]

[0115] Table 2 Results of various physiological and biochemical indices of rice plants in each treatment group under 8‰ salt concentration (mean)

[0116]

[0117] From Tables 1 to 2 and Figures 2 to 12 The experimental results are as follows:

[0118] 1. As Figures 2 to 5 As shown in the results, under the influence of salt stress, the growth and development of rice is severely inhibited. At a salt concentration of 5‰, rice growth basically stagnated, but inoculation with the functional bacteria H1-225 helped the rice to recover. At a salt concentration of 8‰, rice growth was severely inhibited, with leaves turning yellow and curling, but inoculation with the functional bacteria H1-225 helped the rice to survive.

[0119] 2. If Figure 6 As shown in the figure, with the increase of salt concentration, the plant height of rice under high salt stress (8‰) was significantly lower than that under low salt stress (5‰). Under both salt concentrations, the plant height of rice seedlings treated with the fungus was significantly higher than that of rice seedlings under the blank control treatment (CK) and SQR9 treatment.

[0120] 1. At a salt concentration of 5‰, the height of rice plants inoculated with functional bacteria H1-225 increased by 17.9% compared with the blank control.

[0121] 2. At a salt concentration of 8‰, the height of rice plants inoculated with functional bacteria H1-225 increased by 27.9% compared with the blank control.

[0122] 3. Figure 7 As shown in the figure, with the increase of salt concentration, the root length of rice seedlings under high salt stress (8‰) was significantly shorter than that under low salt stress (5‰). At both salt concentrations, the root length of rice seedlings treated with the fungus was significantly longer than that of the blank control treatment (CK) and the SQR9 treatment.

[0123] 1. At a salt concentration of 5‰, the root length of rice inoculated with functional bacteria H1-225 increased by 55.2% compared with the blank control.

[0124] 2. At a salt concentration of 8‰, the root length of rice inoculated with functional bacteria H1-225 increased by 61.3% compared with the blank control.

[0125] 4. Figure 8 As shown in the figure, with increasing salt concentration, the aboveground fresh weight of rice seedlings under high salt stress (8‰) was significantly lower than that under low salt stress (5‰). At both salt concentrations, the aboveground fresh weight of rice seedlings inoculated with fungi was significantly higher than that in the blank control (CK) and SQR9 treatments.

[0126] 1. At a salt concentration of 5‰, the fresh weight of the aboveground part of rice inoculated with functional bacteria H1-225 increased by 432.6% compared with the blank control.

[0127] 2. At a salt concentration of 8‰, the fresh weight of the aboveground part of rice inoculated with functional bacteria H1-225 increased by 358.1% compared with the blank control.

[0128] 5. If Figure 9 As shown in the figure, with the increase of salt concentration, the fresh weight of rice roots under high salt stress (8‰) was significantly lower than that under low salt stress (5‰). At both salt concentrations, the fresh weight of rice roots inoculated with fungi was significantly higher than that in blank control (CK) and SQR9 treatments.

[0129] 1. At a salt concentration of 5‰, the fresh weight of rice roots inoculated with functional bacteria H1-225 increased by 184.0% compared with the blank control.

[0130] 2. At a salt concentration of 8‰, the fresh weight of rice roots inoculated with functional bacteria H1-225 increased by 443.7% compared with the blank control.

[0131] 6. If Figure 10 As shown in the figure, with increasing salt concentration, the aboveground dry weight of rice seedlings under high salt stress (8‰) was significantly lower than that under low salt stress (5‰). At both salt concentrations, the aboveground dry weight of rice seedlings inoculated with the fungus was significantly higher than that in the blank control (CK) and SQR9 treatments.

[0132] 1. At a salt concentration of 5‰, the dry weight of the aboveground part of rice inoculated with functional bacteria H1-225 increased by 198.8% compared with the blank control.

[0133] 2. At a salt concentration of 8‰, the dry weight of the aboveground part of rice inoculated with functional bacteria H1-225 increased by 212.1% compared with the blank control.

[0134] 7. If Figure 11 As shown in the figure, with the increase of salt concentration, the dry weight of rice roots under high salt stress (8‰) was significantly lower than that under low salt stress (5‰). At both salt concentrations, the dry weight of rice roots inoculated with fungi was significantly higher than that in blank control (CK) and SQR9 treatments.

[0135] 1. At a salt concentration of 5‰, the dry weight of rice roots inoculated with functional bacteria H1-225 increased by 121.8% compared with the blank control.

[0136] 2. At a salt concentration of 8‰, the dry weight of rice roots inoculated with functional bacteria H1-225 increased by 192.0% compared with the blank control.

[0137] When plants are stimulated, they produce large amounts of superoxide free radicals, which cause membrane lipid peroxidation. Therefore, membrane lipid peroxidation is an important indicator of plant cell membrane damage. The membrane lipid peroxidation process produces malondialdehyde (MDA), the content of which can directly reflect the peroxidation level of the cytoplasmic membrane.

[0138] 8. If Figure 12 As shown in the figure, under salt stress conditions, the malondialdehyde content in plant leaves increased significantly. At two salt concentrations, the malondialdehyde content in rice leaves treated with the fungus was significantly lower than that in the blank control treatment (CK) and the SQR9 treatment.

[0139] 1. At a salt concentration of 5‰, the malondialdehyde (MDA) content of rice inoculated with functional bacteria H1-225 was reduced by 72.0% compared with the blank control.

[0140] 2. At a salt concentration of 8‰, the malondialdehyde (MDA) content of rice inoculated with functional bacteria H1-225 decreased by 46.1% compared with the blank control.

[0141] Example 4 16S phylogenetic identification of strain H1-225

[0142] 1. The 16S rDNA sequence of strain H1-225 was amplified and sequenced. The sequencing result is shown in SEQ ID NO. 1, specifically:

[0143] TGCAAGTCGAGCGGACAGATGGGAGCTTGCTCCCTGATGTTAGCGGCGGA

[0144] CGGGTGAGTAACACGTGGGTAACCTGCCTGTAAGACTGGGATAACTCCGG

[0145] GAAACCGGGGCTAATACCGGATGGTTGTTTGAACCGCATGGTTCAGACATA

[0146] AAAGTGGCTTCGGCTACCACTTACAGATGGACCCGCGGCGCATTAGCTAG

[0147] TTGGTGAGGTAACGGCTCACCAAGGCGACGATGCGTAGCCGACCTGAGAG

[0148] GGTGATCGGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAG

[0149] GCAGCAGTAGGGAATCTTCCGCAATGGACGAAAGTCTGACGGAGCAACGC

[0150] CGCGTGAGTGATGAAGGTTTTCGGATCGTAAAGCTCTGTTGTTAGGGAAG

[0151] AACAAGTGCCGTTCAAATAGGGCGGCACCTTGACGGTACCTAACCAGAAA

[0152] GCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGC

[0153] GTTGTCCGGAATTATTGGGCGTAAAGGGCTCGCAGGCGGTTTCTTAAGTCT

[0154] GATGTGAAAGCCCCCGGCTCAACCGGGGAGGGTCATTGGAAACTGGGGA

[0155] ACTTGAGTGCAGAAGAGGAGAGTGGAATTCCACGTGTAGCGGTGAAATGC

[0156] GTAGAGATGTGGAGGAACACCAGTGGCGAAGGCGACTCTCTGGTCTGTAA

[0157] CTGACGCTGAGGAGCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTG

[0158] GTAGTCCACGCCGTAAACGATGAGTGCTAAGTGTTAGGGGGTTTCCGCCCC

[0159] TTAGTGCTGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGGTCGCA

[0160] AGACTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCA

[0161] TGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCT

[0162] CTGACAATCCTAGAGATAGGACGTCCCCTTCGGGGGCAGAGTGACAGGTG

[0163] GTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCA

[0164] ACGAGCGCAACCCTTGATCTTAGTTGCCAGCATTCAGTTGGGCACTCTAAG

[0165] GTGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATC

[0166] ATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGACAGAACAAAG

[0167] GGCAGCGAAACCGCGAGGTTAAGCCAATCCCACAAATCTGTTCTCAGTTC

[0168] GGATCGCAGTCTGCAACTCGACTGCGTGAAGCTGGAATCGCTAGTAATCGC

[0169] GGATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCG

[0170] TCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTTATGGAGCCAGCCGCCGAA.

[0171] 2. The sequencing results were analyzed by BLAST comparison to construct a phylogenetic tree ( Figure 13 ) and preliminarily identified the strain species based on the name with the highest homology to the strain to be tested. Strain H1-225 belongs to Bacillus amyloliquefaciens, so it was named Bacillus amyloliquefaciens H1-225.

[0172] 3. Preservation of Bacillus amyloliquefaciens H1-225

[0173] Bacillus amyloliquefaciens H1-225 was deposited with the Guangdong Provincial Microbiological Culture Collection Center (CDMCC; Address: 5th Floor, Dayuan Laboratory Building, 100 Xianlie Middle Road, Yuexiu District, Guangzhou, Guangdong Province) on May 8, 2024, with the deposit number GDMCC No. 64977. Bacillus amyloliquefaciens H1-225 is referred to as Bacillus H1-225.

[0174] Although the above embodiment provides a detailed description of the present invention, it is only a part of the embodiments of the present invention, not all of the embodiments. People can also obtain other embodiments based on this embodiment without creativity, and these embodiments all fall within the scope of protection of the present invention.

Claims

1. A Bacillus amyloliquefaciens H1-225, characterized in that: Deposit number: GDMCC NO.64977.

2. A microbial agent, characterized in that: It comprises the Bacillus amyloliquefaciens H1-225 described in claim 1.

3. The microbial agent according to claim 2, characterized in that: The OD of Bacillus amyloliquefaciens H1-225 in the microbial agent 600 The value of is ≥1.

0.

4. A method for preparing the microbial agent according to claim 2 or 3, characterized in that: include: The Bacillus amyloliquefaciens H1-225 is cultured in a culture medium to obtain a microbial agent.

5. The preparation method according to claim 4, characterized in that: The culture temperature is 30-37° C.; the culture time is 24-34 hours; the culture process is accompanied by shaking, and the shaking speed is 150-230 rpm.

6. Use of the Bacillus amyloliquefaciens H1-225 according to claim 1 in promoting plant growth and / or improving plant salt tolerance.

7. The use according to claim 6, characterized in that: The promoting of plant growth includes promoting the improvement of any one or more of the following indicators (1) to (6): (1) Plant height; (2) root length; (3) fresh weight of aboveground parts; (4) fresh root weight; (5) Aboveground dry weight; (6) Root dry weight.

8. The use according to claim 6, characterized in that: The improving the salt tolerance of plants includes alleviating the damage of salt stress to plants and / or improving the ability of plants to tolerate salt stress.

9. A method for improving the salt tolerance of plants, characterized in that: include: After the plants grow to the two-leaf and one-heart stage, the microbial agent comprising the Bacillus amyloliquefaciens H1-225 of claim 1 is applied.

10. The method according to claim 9, characterized in that: The administration was performed twice, and the interval between the two administrations was two weeks.

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