Compound microbial inoculant with growth promoting effect for moderate saline-alkali soil, preparation method and application

Abstract

This invention relates to the field of biotechnology, specifically to a compound microbial agent, preparation method, and application for promoting growth in moderately saline-alkali soils. The compound microbial agent comprises *Pseudomonas filamentosa* BG4, *Rhizobium sinense* DG1, *Arthrobacter* GCG2, *Pseudomonas* SG3, and *Arthrobacter spheroidae* GCG3. In mildly saline-alkali soils, compared to the control group, the compound microbial agent significantly increases the cumulative aboveground dry weight of corn and the yield per acre. Furthermore, its growth-promoting effect is superior in moderately saline-alkali soils compared to mildly saline-alkali soils. It can be applied to promote the growth of plants in saline-alkali soils and has broad application prospects.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, specifically to a compound bacterial agent with growth-promoting effects suitable for moderately saline-alkali land, its preparation method, and its application. Background Technology

[0002] Over the past two to three decades, soil salinization has become increasingly severe in over 80% of the agricultural soils in irrigated oases in arid Northwest my country. Soil salinization not only reduces soil productivity and leads to significant crop yield reductions by affecting the normal functioning of soil and microorganisms, but also reduces the variety of crops that can be grown, thus lowering land use efficiency. If soil salinization continues to worsen in the future, a considerable amount of farmland in arid irrigated oases will face the risk of abandonment and declining productivity, directly impacting agricultural production and food security. Currently, saline-alkali land management primarily targets coastal saline-alkali soils and alluvial plain saline-alkali soils in the eastern coastal areas. However, inland saline-alkali soils are mainly distributed in Xinjiang Uygur Autonomous Region, Qinghai, Gansu, and Inner Mongolia Autonomous Region. The causes and natural environments of these saline-alkali soils differ significantly from those in the former two regions. In recent years, the management of inland saline-alkali soils has also been concentrated in Xinjiang Uygur Autonomous Region and Inner Mongolia Autonomous Region, primarily employing physical irrigation, chemical methods, and agricultural cultivation. Biological improvement technologies are still in their initial stages.

[0003] The application of microecological agents based on native salt-tolerant microorganisms is one of the biological measures for improving saline-alkali land. Compared with physical, chemical, and engineering measures in saline-alkali land management, it has advantages such as being environmentally friendly and following the natural cycle of the ecological environment. However, its application in saline-alkali land in arid areas of Gansu Province is still in its initial stage. Therefore, exploring the application of microecological agents in typical saline-alkali land in arid areas is of great significance for improving the soil productivity of irrigated oasis agriculture in arid areas. Those skilled in the art have conducted numerous studies; for example, invention patent CN104609992B discloses a compound microbial fertilizer specifically for saline-alkali land. This compound microbial fertilizer comprises the following raw materials in weight percentages: 10%-20% phosphorus- and potassium-solubilizing compound microbial agents, 40%-60% kitchen waste, and 30%-40% diatomaceous earth. The phosphorus- and potassium-solubilizing compound microbial agent is obtained by separately culturing, centrifuging, freeze-drying, and mixing equal amounts of Bacillus megaterium, Bacillus thuringiensis, Pseudomonas fluorescens, Bacillus amyloliquefaciens, Bacillus mucilaginosus, and Azotobacter chroococcus. The kitchen waste is obtained by crushing sorted, impurity-removed, dehydrated, and degreased kitchen waste, adding the compound microbial agent and mixed enzyme preparation, and then aerobic solid-state fermentation. This compound microbial fertilizer utilizes kitchen waste and waste diatomaceous earth, and can fundamentally improve saline-alkali soil. Invention patent CN111575009A discloses a compound microbial agent for improving saline-alkali land, which is prepared by mixing single-strain solid bacteria obtained from the cultivation and fermentation of Bacillus subtilis, Bifidobacterium, Streptomyces flavus, Bacillus megaterium, silicate bacteria, halophilic bacteria, alkaliophilic bacteria, and Pseudomonas in a specific ratio. This invention's microbial agent contains a high number of viable bacteria and is particularly suitable for mixing with base fertilizer. However, none of the above-mentioned microbial fertilizers are intended to solve the soil salinity problem in Gansu Province.

[0004] To address the aforementioned technical problems, this invention aims to improve typical inland saline-alkali soil in Minqin County. It utilizes microorganisms in the rhizosphere of native plants in saline-alkali land to prepare salt-tolerant microecological agents, which are more suitable for the local environment and have a significant effect on improving saline-alkali land, thus having broad application prospects. Summary of the Invention

[0005] The primary objective of this invention is to provide a compound microbial agent suitable for moderately saline-alkali soils with a growth-promoting effect. The compound microbial agent comprises *Pseudomonas silesiensis* BG4, *Sinorhizobium meliloti* DG1, *Arthrobacter sp.* GCG2, and *Pseudomonas sp.* SG3. *Pseudomonas silesiensis* BG4 was deposited in shared access at the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 31645; *Sinorhizobium meliloti* DG1 was deposited in shared access at the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 31930; and *Arthrobacter sp.* GCG2 was deposited in shared access at the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC. 30147; The aforementioned Pseudomonas SG3 was deposited in the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 31646.

[0006] Preferably, the compound microbial agent also includes Arthrobacter globiformis GCG3, which was deposited on February 21, 2023, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 26629.

[0007] A second objective of this invention is to provide the application of the aforementioned compound microbial agent in promoting plant growth.

[0008] Preferably, promoting plant growth includes increasing plant height, increasing the number of spikes, leaves, internodes, and above-ground dry weight.

[0009] A third objective of this invention is to provide the application of the aforementioned compound microbial agent in increasing plant yield.

[0010] A fourth objective of this invention is to provide the application of the aforementioned compound microbial agent in improving the salt tolerance of plants.

[0011] The fifth objective of this invention is to provide the application of the aforementioned compound microbial agent in improving plant alkali tolerance.

[0012] Preferably, five bacterial solutions are obtained by separately culturing Pseudomonas filamentosa BG4, Rhizobium sinense of alfalfa DG1, Arthrobacter gamma, Pseudomonas SG3, and Arthrobacter gamma, which are then mixed in equal proportions to obtain the final product.

[0013] The sixth objective of this invention is to provide a compound microbial fertilizer specifically for saline-alkali land, wherein the compound microbial fertilizer comprises the aforementioned compound microbial agent and auxiliary materials.

[0014] The beneficial effects of this invention are as follows: This invention provides a compound microbial agent suitable for moderately saline-alkali land with a growth-promoting effect. The compound microbial agent includes *Pseudomonas silesiensis* BG4, *Sinorhizobium meliloti* DG1, *Arthrobacter sp.* GCG2, and *Pseudomonas sp.* SG3. *Pseudomonas silesiensis* BG4 was deposited in a shared manner at the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 31645; *Sinorhizobium meliloti* DG1 was deposited in a shared manner at the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 31930; and *Arthrobacter sp.* GCG2 was deposited in a shared manner at the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC. 30147; the *Pseudomonas* SG3 was deposited on March 1, 2024, in a shared manner at the Gansu Provincial Microbial Culture Collection Center, with accession number GSICC 31646; the compound microbial agent also includes *Arthrobacter globiformis* GCG3, which was deposited on February 21, 2023, at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 26629. On slightly saline-alkali land, the compound microbial agent, in the experimental groups PB, PC, and PD, increased the cumulative aboveground dry weight of maize by 7.12%, 6.68%, and 7.95% compared to the control CK, respectively, with no significant differences among the three treatments. The maize yield per mu (667 square meters) of PA and PC was significantly higher than that of CK by 25.72% and 19.58%, respectively. On moderately saline-alkali land, the experimental group PC increased the aboveground dry weight of maize by 21.1% compared to the control CK; the maize yield per mu increased by 30.1%. Meanwhile, the experimental groups PA and PB had no significant impact on maize yield compared to the negative control CK, while the experimental group PD increased maize yield by 15.7%, which was far less than that of the PC treatment. This means that the compound microbial agent described in this invention has a certain growth-promoting effect on saline-alkali land, with a better effect on moderately saline-alkali land than on mildly saline-alkali land. It can be applied to promote the growth of plants in saline-alkali land and has broad application prospects. Detailed Implementation

[0015] The following are specific embodiments of the present invention, which further describe the technical solution of the present invention, but the present invention is not limited to these embodiments.

[0016] The Gansu Branch of the China Industrial Microbial Culture Collection Center is the only specialized institution in Northwest China dedicated to the preservation and management of industrial microbial strains. Its main functions include the isolation, collection, preservation, supply, and external sharing and exchange of microbial strain resources. Currently, it preserves over 2,000 strains of various microorganisms and is one of the construction units of the National Microbial Resource Platform's Industrial Microbial Strain Platform. The center provides a variety of superior strains for environmental remediation, industry, agriculture, medicine, and light industry. The center has a strain resource bank (physical specimens) and a database (website: http: / / jzk.gsmsc.cn), allowing users to conveniently and quickly browse, consult, and order the industrial microbial strains they need via the internet.

[0017] The microbial strain used in the following examples includes Pseudomonas silesiensis BG4, which was deposited in the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 31645. Those skilled in the art can find the above strain on the official website of the Gansu Provincial Microbial Culture Collection Center and can apply to purchase, exchange or obtain the above strain in other ways. In the following examples, it is abbreviated as Pseudomonas silesiensis BG4.

[0018] Sinorhizobium meliloti DG1, deposited in Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 31930, is available to those skilled in the art through the official website of Gansu Provincial Microbial Culture Collection Center after March 1, 2024. Such strains can be obtained through purchase, exchange, or other means. In the following examples, it is abbreviated as Sinorhizobium meliloti DG1.

[0019] Arthrobacter sp. DG3 was deposited in the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 30146. Since March 1, 2024, those skilled in the art can find the above strain on the official website of the Gansu Provincial Microbial Culture Collection Center and apply to obtain the above strain in a shared manner. In the following examples, it is abbreviated as Arthrobacter sp. DG3.

[0020] Arthrobacter sp. NBG2 was deposited in shared form at Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 30148. Since March 1, 2024, those skilled in the art can find the above strain on the official website of Gansu Provincial Microbial Culture Collection Center and can apply to purchase, exchange or obtain the above strain in other ways. In the following examples, it is abbreviated as Arthrobacter sp. NBG2.

[0021] Arthrobacter sp. SG2 was deposited in shared form at Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 30149. Since March 1, 2024, those skilled in the art can find the above strain on the official website of Gansu Provincial Microbial Culture Collection Center and can apply to purchase, exchange or obtain the above strain in other ways. In the following examples, it is abbreviated as Arthrobacter sp. SG2.

[0022] Pseudomonas sp. SG3 was deposited in the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 31646. Since March 1, 2024, those skilled in the art can find the above strain on the official website of the Gansu Provincial Microbial Culture Collection Center and can apply to purchase, exchange or obtain the above strain in other ways. In the following examples, it is abbreviated as Pseudomonas sp. SG3.

[0023] Arthrobacter globiformis GCG2 was deposited in shared form at Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 30147. Since March 1, 2024, those skilled in the art can find the above strain on the official website of Gansu Provincial Microbial Culture Collection Center and can apply to purchase, exchange or obtain the above strain in other ways. In the following examples, it is abbreviated as Arthrobacter globiformis GCG2.

[0024] Arthrobacter globiformis GCG3 was deposited on February 21, 2023, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 26629. The deposit address is No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing. In the following examples, it is abbreviated as Arthrobacter globiformis GCG3.

[0025] All of the above strains were isolated from plant root soil in Minqin saline-alkali land by members of the research group in October 2021.

[0026] In the following examples, the LB medium consisted of: 10 g / L tryptone, 5 g / L yeast extract, 10 g / L NaCl, and a pH of 7.0.

[0027] Example 1: Isolation and Molecular Identification of Strains

[0028] 1. Isolation of strains

[0029] Soil samples were collected from the artificially cultivated sites of various Chinese medicinal herbs, including *Nitraria tangutorum*, *Ilex chinensis*, *Scutellaria baicalensis*, *Glycyrrhiza uralensis*, and *Rheum palmatum*, at the Northwest Institute of Arid Zone Salinization Research Station of the Chinese Academy of Sciences in Minqin County, Zhangye. The topsoil was removed, and soil samples were taken from the rhizosphere soil at a depth of 15-20 cm. 100 mL of sterile water was added, and the samples were allowed to stand for 30 minutes. Then, the samples were incubated at 28℃ and rotated at 180 rpm. -1 Shake for 30 minutes to form a suspension. Add 5 mL of the suspension to 100 mL of La Shby liquid medium and incubate at 28°C and 180 rpm. -1 After 72 hours of enrichment culture, 10⁻¹⁰ samples were prepared using a conventional serial dilution method. -4 10 -5 10 -6 10 -7 After diluting the bacterial suspensions in gradients, 0.1 mL of each suspension was spread onto Ashby plates, with each gradient repeated three times. The plates were incubated at 28°C for 3–5 days. This isolation process was repeated three times. Once colonies had grown, the strains were purified further on Ashby medium using the streak plating method. The purified nitrogen-fixing bacteria were numbered sequentially as BG4, DG1, DG3, SG2, SG3, NBG2, GCG2, and GCG3.

[0030] 2. Molecular identification

[0031] The strain GCG3, isolated and purified using the above steps, was cultured using conventional methods. Total DNA from the strain was extracted and used as a template for gene amplification. Universal primers for bacterial 16S rDNA were used, and the primer sequences are as follows:

[0032] 27F:5-AGAGTTTGATCCTGGCTCAG-3

[0033] 1492R: 5-CTACGGCTACCTTGTTACGA-3

[0034] Perform PCR on a PCR amplification instrument. After the reaction, take 2 μl of the PCR product and perform 1% agarose gel electrophoresis to confirm the PCR amplification fragments. The PCR product is then recovered using the AxyPrep DNA Gel Recovery Kit; follow the kit's instructions for specific procedures.

[0035] The purified PCR products of each bacterial species were sequenced using an ABI 3730-XL sequencer. Homology comparisons with known sequences in NCBIGenBank were performed to determine the bacterial species and classify them into genus or species.

[0036] PCR gene amplification yielded approximately 1.4 kb 16S rDNA gene fragments for each strain, the gene sequences of which are shown in SEQ ID No. 1. After sequence determination, online homology comparison was performed with publicly available 16S rDNA sequences in the NCBI database.

[0037] Table 1. Similarity of strains

[0038]

[0039] The results are shown in Table 1. The strains were identified as Pseudomonas asperilesiensis BG4, Sinorhizobium meliloti DG1, Arthrobacter sp. DG3, Pseudomonas sp. SG2, Pseudomonas sp. SG3, Arthrobacter sp. NBG2, and Arthrobacter sp. GCG2, and were deposited in the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, in a shared manner.

[0040] The strain GCG3 belongs to the genus Arthrobacter and is named Arthrobacter globiformis GCG3. It was deposited on February 21, 2023, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 26629. The deposit address is No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing.

[0041] Example 2: Nitrogenase Activity Test of the Strains

[0042] Nitrogenase activity was determined in eight salt-tolerant growth-promoting bacteria isolated and purified in Example 1 using the acetylene reduction method. The specific method is as follows: One loopful of the test strain was inoculated into a serum bottle containing 5 mL of semi-solid Ashby medium and incubated at 28°C for 48 h. In a sterile environment, the serum bottle cap was replaced with a rubber stopper, and 1 mL of gas was extracted using a sterile syringe. Then, 1 mL of acetylene was injected, the bottle was sealed, and incubated at 28°C for another 36 h. 0.2 mL of the mixed gas was extracted from the serum bottle using a sterile syringe and injected into a gas chromatograph to determine the ethylene production. Nitrogenase activity was calculated using the following formula. All experiments were performed in triplicate. N = h x CV / (24.9 hst)

[0043] Where N is the concentration of ethylene produced, nmol(C2H4)·h⁻¹ -1 ·mL -1 Nitrogenase activity; hx is the peak area of ​​ethylene in the sample; C is the concentration of standard ethylene (nmol·ml)-1 V is the volume of the culture container (mL), 24.9 is a constant; hs is the peak area of ​​standard ethylene; t is the sample incubation time (h).

[0044] The results are shown in Table 2. All eight strains showed some nitrogenase activity, with *Arthrobacter globosa* GCG3 exhibiting the highest nitrogenase activity at 122.5 nmol (C2H4)·h⁻¹. -1 ·mL -1 .

[0045] Example 3: Determination of the strain's ability to secrete IAA

[0046] Qualitative determination: The ability of eight salt-tolerant growth-promoting bacteria isolated and purified in Example 1 to secrete IAA was determined. A single colony of the strain isolated in Example 1 was inoculated into 50 mL of King's B liquid medium and cultured with shaking at 28°C and 160 rpm for 12 days. After centrifugation at 10,000 rpm for 10 min at 4°C, the supernatant was collected, and 50 μL of the bacterial solution was placed on a white ceramic plate. An equal volume of Spot colorimetric solution was added. In the control, 50 μL of IAA (10 mg / L) was used instead of the bacterial solution. The plate was incubated at room temperature for 15 min, and the color change was observed. A pinkish color indicates IAA secretion, and a darker color indicates a stronger IAA secretion ability.

[0047] The results showed that the fermentation broth of the eight strains was light pink on the ceramic plate, and the color was arranged from darkest to lightest as follows: Arthrobacter DG3 > Arthrobacter GCG3 > Arthrobacter NBG2 > Arthrobacter SG2 > Rhizobium sinense of alfalfa DG1 > Arthrobacter GCG2 = Pseudomonas SG3 = Pseudomonas BG4, indicating that Arthrobacter DG3, Arthrobacter GCG3 and Arthrobacter NBG2 had a strong ability to secrete IAA.

[0048] Example 4: Determination of the Phosphate-Solubilizing Ability of the Strains

[0049] Qualitative determination: The phosphorus solubility of the eight salt-tolerant growth-promoting bacteria isolated and purified in Example 1 was determined. Single colonies of the strains isolated in Example 1 were inoculated on inorganic phosphorus medium and Mongkina organic phosphorus medium. Four regions were inoculated in each petri dish and incubated in a 28°C incubator for 7 days. The size of the phosphorus solubility zone of each strain was observed and measured, and screening was carried out based on the size of the phosphorus solubility zone.

[0050] The results are shown in Table 2. All eight strains have certain nitrogen-fixing ability and IAA production characteristics. Among them, Pseudomonas BG4, Arthrobacter SG2, Pseudomonas SG3 and Arthrobacter NBG2 have certain phosphorus-solubilizing ability.

[0051] Table 2. Nitrogen fixation capacity, IAA production, and phosphorus solubility of different strains

[0052]

[0053] Example 5: Salt and Alkali Tolerance of the Strains

[0054] The eight salt-tolerant growth-promoting bacterial strains obtained in Example 1 were inoculated onto LB plates with different salt concentrations (5%, 8%, and 10%), with the normal LB salt concentration as a control. The plates were incubated at 28°C for 24 hours to observe whether the growth of the strains was inhibited. Strains with normal growth were considered to be tolerant of the corresponding salt concentration, while those with higher concentrations were not. Similarly, the alkalinity tolerance of the strains was tested by inoculating them onto plates with different pH values ​​(8 and 9).

[0055] The results are shown in Table 3. Pseudomonas BG4, Arthrobacter DG3, Rhizobium sinense of alfalfa DG1, Arthrobacter SG2, Pseudomonas SG3, Arthrobacter NBG2, Arthrobacter GCG2, and Arthrobacter globosa GCG3 were able to tolerate a salinity of 8%, with Arthrobacter globosa GCG3 able to grow at a salinity of 10%. Pseudomonas BG4, Rhizobium sinense of alfalfa DG1, Arthrobacter SG2, Pseudomonas SG3, and Arthrobacter globosa GCG3 were able to grow in an environment with a pH of 9.

[0056] Table 3 Salt and pH tolerance of different strains

[0057]

[0058] Example 6: Effects of compound microbial agents on maize yield on saline-alkali land with different degrees of salinity

[0059] Based on their antagonistic effects, they are divided into the following four groups;

[0060] Experimental bacterial agent (PA): Pseudomonas BG4 and Arthrobacter GCG2 were cultured in LB medium at 28℃ and 200 r / min for 24 h, adjusted to the same OD value, mixed in equal proportions, and diluted with sterile water to 1×10⁻⁶. 7 cfu / ml; adjust to a total bacterial concentration of 1×10⁻⁶. 7 cfu / ml;

[0061] Experimental inoculum (PB): Pseudomonas BG4, Rhizobium sinense of alfalfa DG1, Arthrobacter DG3, Arthrobacter SG2, Pseudomonas SG3, Arthrobacter GCG2, and Arthrobacter NBG2 were cultured separately on LB medium at 28℃ and 200 r / min for 24 h, adjusted to the same OD value, and then mixed in equal proportions and diluted with sterile water to a concentration of 1×10⁻⁶. 7 cfu / ml; adjust to a total bacterial concentration of 1×10⁻⁶. 7 cfu / ml;

[0062] Experimental bacterial inoculum (PC): Pseudomonas BG4, Rhizobium sinense DG1, Pseudomonas SG3, Arthrobacter GCG2, and Arthrobacter globosa GCG3 were cultured separately in LB medium at 28℃ and 200 rpm for 24 h, adjusted to the same OD value, and then mixed in equal proportions. The mixture was then adjusted with sterile water to a concentration of 1×10⁷ CFU / ml, resulting in a total bacterial concentration of 1×10⁷ CFU / ml. 7 cfu / ml;

[0063] Experimental bacterial inoculum (PD): Pseudomonas BG4, Rhizobium sinense DG1, Arthrobacter GCG2, and Pseudomonas SG3 were cultured separately on LB medium at 28℃ and 200 r / min for 24 h, adjusted to the same OD value, and then mixed in equal proportions. The mixture was then adjusted with sterile water to a concentration of 1×10⁷ CFU / ml; the total bacterial concentration was adjusted to 1×10⁷ CFU / ml. 7 cfu / ml;

[0064] Negative control group (CK): LB medium without bacteria.

[0065] Based on the antagonistic effects of the strains, five treatments were set up: PA, PB, PC, PD, and CK. Different strains were cultured in LB medium with shaking at 28℃ and 200 r / min for 24 h, adjusted to the same OD value, and then mixed in equal proportions with sterile water to a concentration of 1×10⁻⁶. 7 CFU / ml, with equal volume applied to all treatments. Each treatment was replicated three times, with a plot size of 5 m². 2 At the sowing period, the seedlings were coated with the inoculum stock solution and then sown. After emergence, different treatments were applied by root irrigation and spraying, and sprayed once at the jointing stage, for a total of two times. Statistical data for different treatments were collected after the corn matured. Two experimental plots were selected for the experiment, one with slightly saline-alkali soil (salt content 1.4 g / kg) and the other with moderately saline-alkali soil (salt content 4.075 g / kg).

[0066] Table 4. Experimental Results of Mildly Saline-Alkali Land

[0067]

[0068] Note: Each lowercase letter in each line represents a significant difference (P<0.05) in multiple comparisons.

[0069] Table 5. Experimental Results in Moderately Saline-Alkali Land

[0070]

[0071] Note: Each lowercase letter in each line represents a significant difference (P<0.05) in multiple comparisons.

[0072] Table 4 shows that in the slightly saline-alkali land experimental groups, the cumulative aboveground dry weight of maize (PB, PC, and PD treatments) increased by 7.12%, 6.68%, and 7.95% respectively compared to the control (CK), with no significant differences among the three treatments. The maize yield per mu (667 square meters) was significantly higher in PA and PC treatments than in CK (25.72% and 19.58%, respectively). Table 5 shows that in the moderately saline-alkali land experimental group, PC treatment increased the aboveground dry weight of maize by 21.1% compared to the control (CK), and the maize yield per mu increased by 30.1%. Meanwhile, the experimental groups PA and PB had no significant effect on maize yield compared to the negative control (CK), while the PD treatment increased maize yield by 15.7%, far less than the PC treatment. Therefore, the compound microbial agent described in this invention has a certain growth-promoting effect on saline-alkali land, with a better effect in moderately saline-alkali land than in slightly saline-alkali land.

[0073] In summary, this invention provides a compound microbial agent that is tolerant to saline-alkali soil and has a growth-promoting effect. The compound microbial agent includes *Pseudomonas silesiensis* BG4, *Sinorhizobium meliloti* DG1, *Arthrobacter sp.* GCG2, and *Pseudomonas sp.* SG3. *Pseudomonas silesiensis* BG4 was deposited in a shared access manner at the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 31645; *Sinorhizobium meliloti* DG1 was deposited in a shared access manner at the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC 31930; and *Arthrobacter sp.* GCG2 was deposited in a shared access manner at the Gansu Provincial Microbial Culture Collection Center on March 1, 2024, with the accession number GSICC. 30147; the *Pseudomonas* SG3 was deposited on March 1, 2024, in a shared manner at the Gansu Provincial Microbial Culture Collection Center, with accession number GSICC 31646; the compound microbial agent also includes *Arthrobacter globiformis* GCG3, which was deposited on February 21, 2023, at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 26629. On slightly saline-alkali land, the compound microbial agent, in the experimental groups PB, PC, and PD, increased the cumulative aboveground dry weight of maize by 7.12%, 6.68%, and 7.95% compared to the control CK, respectively, with no significant differences among the three treatments. The maize yield per mu (667 square meters) of PA and PC was significantly higher than that of CK by 25.72% and 19.58%, respectively. On moderately saline-alkali land, the experimental group PC increased the aboveground dry weight of maize by 21.1% compared to the control CK; the maize yield per mu increased by 30.1%. Meanwhile, the experimental groups PA and PB had no significant impact on maize yield compared to the negative control CK, while the experimental group PD increased maize yield by 15.7%, which was far less than that of the PC treatment. This means that the compound microbial agent described in this invention has a certain growth-promoting effect on saline-alkali land, with a better effect on moderately saline-alkali land than on mildly saline-alkali land. It can be applied to promote the growth of plants in saline-alkali land and has broad application prospects.

Claims

1. A compound microbial agent with growth-promoting effect suitable for moderately saline-alkali land, characterized in that, The compound bacterial agent is composed of Pseudomonas filamentosa ( pseudomonas silesiensis BG4, *Alfalfa rhizobium* ( Sinorhizobium meliloti DG1, Arthrobacter ( Arthrobacter sp. GCG2, Pseudomonas ( Pseudomonas sp. SG3 and Orbacterium spheroidum ( Arthrobacter globiformis The composition includes GCG3, with *Pseudomonas filamentosa* BG4 deposited on March 1, 2024, in a shared manner at the Gansu Provincial Microbial Culture Collection Center (GSICC 31645); *Rhizobium sinense* DG1 deposited on March 1, 2024, in a shared manner at the Gansu Provincial Microbial Culture Collection Center (GSICC 31930); *Arthrobacter globosa* GCG2 deposited on March 1, 2024, in a shared manner at the Gansu Provincial Microbial Culture Collection Center (GSICC 30147); *Pseudomonas sigmatis* SG3 deposited on March 1, 2024, in a shared manner at the Gansu Provincial Microbial Culture Collection Center (GSICC 31646); and *Arthrobacter globosa* GCG3 deposited on February 21, 2023, at the China General Microbiological Culture Collection Center (CGMCC No. 26629).

2. The application of the compound microbial agent as described in claim 1 in promoting corn growth.

3. The application as described in claim 2, characterized in that, The aforementioned methods for promoting corn growth include increasing plant height, increasing the number of ears, leaves, internodes, and above-ground dry weight.

4. The application of the compound microbial agent as described in claim 1 in increasing corn yield.

5. The application of the compound microbial agent as described in claim 1 in improving the salt tolerance of corn.

6. The application of the compound microbial agent as described in claim 1 in improving the alkali resistance of corn.

7. The method for preparing the compound microbial agent as described in claim 1, characterized in that, Five bacterial suspensions were obtained by culturing Pseudomonas filamentosa BG4, Rhizobium sinense of alfalfa DG1, Arthrobacter glucosinolates GCG2, Pseudomonas sG3, and Arthrobacter glucosinolates GCG3, respectively. These suspensions were then mixed in equal proportions to obtain the final product.

8. A compound microbial fertilizer specifically for saline-alkali land, characterized in that, The compound microbial fertilizer includes the compound microbial agent and auxiliary materials as described in claim 1.

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