A stress-resistant and growth-promoting microbial agent and its preparation method and application

Abstract

The present invention discloses a stress-resistant and growth-promoting microbial agent and its preparation method and application, which belong to the field of microbial technology. The stress-resistant and growth-promoting microbial agent of the present invention comprises Bacillus siamese, Burkholderia bivariate, and Sphingomonas yunnanensis. After the bacterial liquid of the three strains is mixed in equal proportions, they cooperate with each other to achieve significant stress-resistant and growth-promoting effects, especially in improving stress resistance indicators during the growth process of wheat plants, which is manifested in increasing the activity of the antioxidant enzyme SOD in the plants, increasing the content of the osmotic regulating substance proline, and reducing the content of MDA. The microbial agent of the present invention, when applied in conjunction with conventional fertilization, significantly promotes the growth of wheat seedlings under salt stress and increases wheat yield.

Description

Technical Field

[0001] The present invention belongs to the field of microbial technology, and in particular relates to a stress-resistant and growth-promoting microbial agent, a preparation method and an application thereof. Background Art

[0002] wheat( Triticum aestivum L. ) is an annual or biennial herbaceous plant of the genus Triticeae in the Poaceae family. The caryopsis of wheat is edible and it is one of the most important food crops at present.

[0003] Soil salinization seriously impacts plant growth and crop yields. Many agricultural irrigated lands are plagued by salt stress, and the problem is becoming increasingly serious due to irrational land use and environmental pollution. Salinized soils have poor physical and chemical properties and are prone to compaction. This not only inhibits enzyme activity but also affects microbial activity. This ultimately leads to suppressed plant growth and reduced soil fertility. As global climate change intensifies, agricultural production faces increasingly severe abiotic stresses such as drought and salinization. Traditional chemical fertilizers and plant growth regulators have limited effectiveness under adverse conditions and may cause environmental pollution. Microbial agents, as environmentally friendly biofertilizers, can promote plant growth and enhance plant stress resistance through various mechanisms.

[0004] However, the microbial agents currently on the market generally have problems such as insufficient stress resistance, single function, and unstable field effects. There are no microbial agents specifically for wheat to resist salt stress. Therefore, there is an urgent need to develop a microbial agent that can improve wheat's stress resistance and promote growth. Summary of the Invention

[0005] In response to the problems existing in the prior art, the present invention provides a stress-resistant and growth-promoting microbial agent for wheat growing in saline-alkali soil. Through the synergistic effect of a specific functional bacterial strain combination and other synergistic ingredients, the growth performance of wheat under adverse conditions of saline-alkali soil is significantly improved.

[0006] In order to achieve the above technical objectives, the technical solution adopted by the present invention is:

[0007] A stress-resistant and growth-promoting microbial agent, comprising the following raw materials in parts by weight:

[0008] 15-25 parts of functional microbial agents, 3-5 parts of trehalose-proline composite nanoparticles, 35-45 parts of composite carriers, and 2-3 parts of trace element-containing substances.

[0009] Furthermore, the functional microbial agent includes Bacillus siamese, Burkholderia bifida, and Sphingomonas yunnanensis; the Bacillus siamese was purchased from the China Center for Type Culture Collection, with a deposit number of CCTCC AB2022048 and an original deposit date of January 7, 2022; the Burkholderia bifida was purchased from the China General Microbiological Culture Collection Center, with a deposit number of CGMCC No. 1.10511 and an original deposit date of March 29, 2010; the Sphingomonas yunnanensis was purchased from the China General Microbiological Culture Collection Center, with a deposit number of CGMCC No. 1.15275 and an original deposit date of June 20, 2015.

[0010] Furthermore, the preparation method of the functional microbial agent is:

[0011] (1) Thaw Bacillus siamese and Burkholderia bifida and activate them in LB solid medium. Pick out single colonies and inoculate them into LB liquid medium. Cultivate at 27℃ and 180r / min until OD 600 =0.6 to obtain seed solution, and 1% of the seed solution was inoculated into 250 mL of LB liquid culture medium and cultured at 27 ° C and 180 r / min. The number of viable bacteria in the culture solution was detected, and the number of viable bacteria reached 1×10 8 cfu / mL and then the culture was terminated to obtain Bacillus siamensis bacterial solution and Burkholderia bifida bacterial solution respectively;

[0012] (2) Place Sphingomonas Yunnanensis in nutrient broth agar medium for activation, pick a single colony and inoculate it into nutrient broth liquid medium, and culture at 30℃ and 200r / min until OD 600 =0.6 to obtain seed solution, and 1% of the seed solution was inoculated into 250 mL of LB liquid medium and cultured at 30°C and 200 r / min. The number of viable bacteria was detected and the number reached 1×10 8 cfu / mL and the culture was terminated to obtain the Sphingomonas yunnanensis bacterial liquid;

[0013] (3) The Bacillus siamese bacterial solution and the Burkholderia bifida bacterial solution in step (1) and the Sphingomonas yunnanensis bacterial solution in step (2) are mixed in a volume ratio of 1:1:1 to obtain a functional microbial agent.

[0014] Furthermore, the preparation method of the trehalose-proline composite nanoparticles is:

[0015] Trehalose and L-proline were dissolved in deionized water at a mass ratio of 2:1 to prepare a 10% solution; 0.1% chitosan was then added as a stabilizer; high-pressure homogenization was performed five times at 100 MPa to obtain composite nanoparticles with a particle size of 50-100 nm; and powdered trehalose-proline composite nanoparticles were obtained by spray drying.

[0016] Furthermore, the composite carrier includes humic acid, bentonite and biochar, and the mass ratio of the three is 3:2:1.

[0017] Furthermore, the trace element-containing substance includes chelated zinc, chelated iron, and chelated manganese, and the mass ratio of the three is 4:3:2.

[0018] A method for preparing a stress-resistant and growth-promoting microbial agent comprises the following steps:

[0019] The three strains were expanded and cultured separately to obtain bacterial liquids of the three strains, which were mixed in equal volumes to obtain a functional microbial agent. The functional microbial agent was first mixed with a composite carrier, and then trehalose-proline composite nanoparticles and trace element substances were added and mixed thoroughly. The moisture content was adjusted to 25-30%, and the mixture was fermented at 30°C for 48 hours, dried at low temperature to a moisture content of less than 10%, and crushed to pass 80 mesh to obtain a stress-resistant and growth-promoting microbial agent.

[0020] The present invention also provides the use of the stress-resistant and growth-promoting microbial agent in promoting wheat growth and effectively improving the salt-alkali resistance of wheat.

[0021] The raw materials used in the present invention are all commercially available.

[0022] Beneficial effects

[0023] (1) The microbial agent of the present invention is a synergistic mixture of three functional strains of Bacillus siamese, Burkholderia bifida, and Sphingomonas yunnanensis, which are mixed in equal proportions to effectively improve the activity of antioxidant enzyme SOD in plants, increase osmotic regulating substances such as proline, and reduce membrane lipid peroxidation - MDA content, significantly promote the growth of wheat seedlings under salt stress, and increase wheat yield.

[0024] (2) This invention is the first to combine trehalose and proline through nanotechnology to form stable particles. These particles act as a microbial protectant and plant stress resistance inducer, slowly releasing the particles under saline-alkali conditions. Furthermore, the nanoparticles are more easily absorbed and utilized by microorganisms and plant roots. Furthermore, the trace elements iron, manganese, and zinc are added to the microbial agent in chelated form, which can be effectively absorbed and utilized by plants. The synergistic effect with the composite microbial community comprehensively improves the stress resistance and growth performance of wheat plants.

[0025] (3) The microbial agent of the present invention, when applied in combination with conventional fertilization, significantly promoted the growth and stress resistance of wheat seedlings under salt stress, and increased wheat yield. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 This is a diagram showing the growth status of the strains selected in the present invention on LB medium; Note: 1 represents Bacillus siamensis, 2 represents Burkholderia bifida, and 3 represents Sphingomonas yunnanensis. DETAILED DESCRIPTION

[0027] The technical solution of the present invention will be further described below with reference to specific embodiments, but is not limited thereto.

[0028] Example 1

[0029] A stress-resistant and growth-promoting microbial agent, comprising the following raw materials in parts by weight:

[0030] 15 parts of functional microbial agents, 3 parts of trehalose-proline composite nanoparticles, 35 parts of composite carriers, and 2 parts of trace element-containing substances.

[0031] The functional microbial agent includes Bacillus siamensis, Burkholderia bifida, and Sphingomonas yunnanensis; the preservation number of the Bacillus siamensis is CCTCC AB 2022048; the preservation number of the Burkholderia bifida is CGMCC No. 1.10511; and the preservation number of the Sphingomonas yunnanensis is CGMCC No. 1.15275.

[0032] The preparation method of the functional microbial agent is as follows:

[0033] (1) Thaw Bacillus siamese and Burkholderia bifida and activate them in LB solid medium. Pick out single colonies and inoculate them into LB liquid medium. Cultivate at 27℃ and 180r / min until OD 600 =0.6 to obtain seed solution, and 1% of the seed solution was inoculated into 250 mL of LB liquid culture medium and cultured at 27 ° C and 180 r / min. The number of viable bacteria in the culture solution was detected, and the number of viable bacteria reached 1×10 8 cfu / mL and then the culture was terminated to obtain Bacillus siamensis bacterial solution and Burkholderia bifida bacterial solution respectively;

[0034] (2) Place Sphingomonas Yunnanensis in nutrient broth agar medium for activation, pick a single colony and inoculate it into nutrient broth liquid medium, and culture at 30℃ and 200r / min until OD 600 =0.6 to obtain seed solution, and 1% of the seed solution was inoculated into 250 mL of LB liquid medium and cultured at 30°C and 200 r / min. The number of viable bacteria was detected and the number reached 1×108 cfu / mL and the culture was terminated to obtain the Sphingomonas yunnanensis bacterial liquid;

[0035] (3) The Bacillus siamese bacterial solution and the Burkholderia bifida bacterial solution in step (1) and the Sphingomonas yunnanensis bacterial solution in step (2) are mixed in a volume ratio of 1:1:1 to obtain a functional microbial agent.

[0036] The preparation method of the trehalose-proline composite nanoparticles is as follows:

[0037] Trehalose and L-proline were dissolved in deionized water at a mass ratio of 2:1 to prepare a 10% solution; 0.1% chitosan was then added as a stabilizer; high-pressure homogenization was performed five times at 100 MPa to obtain composite nanoparticles with a particle size of 50-100 nm; and powdered trehalose-proline composite nanoparticles were obtained by spray drying.

[0038] The composite carrier includes humic acid, bentonite and biochar, and the mass ratio of the three is 3:2:1.

[0039] The trace element-containing substance includes chelated zinc, chelated iron and chelated manganese, and the mass ratio of the three is 4:3:2.

[0040] A method for preparing a stress-resistant and growth-promoting microbial agent comprises the following steps:

[0041] The three strains were expanded and cultured separately to obtain bacterial liquids of the three strains, which were mixed in equal volumes to obtain a functional microbial agent. The functional microbial agent was first mixed with a composite carrier, and then trehalose-proline composite nanoparticles and trace element substances were added and mixed thoroughly. The moisture content was adjusted to 25-30%, and the mixture was fermented at 30°C for 48 hours, dried at low temperature to a moisture content of less than 10%, and crushed to pass 80 mesh to obtain a stress-resistant and growth-promoting microbial agent.

[0042] Example 2

[0043] A stress-resistant and growth-promoting microbial agent, comprising the following raw materials in parts by weight:

[0044] 20 parts of functional microbial agents, 4 parts of trehalose-proline composite nanoparticles, 40 parts of composite carriers, and 2 parts of trace element-containing substances.

[0045] The functional microbial agent includes Bacillus siamensis, Burkholderia bifida, and Sphingomonas yunnanensis; the preservation number of the Bacillus siamensis is CCTCC AB 2022048; the preservation number of the Burkholderia bifida is CGMCC No. 1.10511; and the preservation number of the Sphingomonas yunnanensis is CGMCC No. 1.15275.

[0046] The preparation method of the functional microbial agent is as follows:

[0047] (1) Thaw Bacillus siamese and Burkholderia bifida and activate them in LB solid medium. Pick out single colonies and inoculate them into LB liquid medium. Cultivate at 27℃ and 180r / min until OD 600 =0.6 to obtain seed solution, and 1% of the seed solution was inoculated into 250 mL of LB liquid culture medium and cultured at 27 ° C and 180 r / min. The number of viable bacteria in the culture solution was detected, and the number of viable bacteria reached 1×10 8 cfu / mL and then the culture was terminated to obtain Bacillus siamensis bacterial solution and Burkholderia bifida bacterial solution respectively;

[0048] (2) Place Sphingomonas Yunnanensis in nutrient broth agar medium for activation, pick a single colony and inoculate it into nutrient broth liquid medium, and culture at 30℃ and 200r / min until OD 600 =0.6 to obtain seed solution, and 1% of the seed solution was inoculated into 250 mL of LB liquid medium and cultured at 30°C and 200 r / min. The number of viable bacteria was detected and the number reached 1×10 8 cfu / mL and the culture was terminated to obtain the Sphingomonas yunnanensis bacterial liquid;

[0049] (3) The Bacillus siamese bacterial solution and the Burkholderia bifida bacterial solution in step (1) and the Sphingomonas yunnanensis bacterial solution in step (2) are mixed in a volume ratio of 1:1:1 to obtain a functional microbial agent.

[0050] The preparation method of the trehalose-proline composite nanoparticles is as follows:

[0051] Trehalose and L-proline were dissolved in deionized water at a mass ratio of 2:1 to prepare a 10% solution; 0.1% chitosan was then added as a stabilizer; high-pressure homogenization was performed five times at 100 MPa to obtain composite nanoparticles with a particle size of 50-100 nm; and powdered trehalose-proline composite nanoparticles were obtained by spray drying.

[0052] The composite carrier includes humic acid, bentonite and biochar, and the mass ratio of the three is 3:2:1.

[0053] The trace element-containing substance includes chelated zinc, chelated iron and chelated manganese, and the mass ratio of the three is 4:3:2.

[0054] A method for preparing a stress-resistant and growth-promoting microbial agent comprises the following steps:

[0055] The three strains were expanded and cultured separately to obtain bacterial liquids of the three strains, which were mixed in equal volumes to obtain a functional microbial agent. The functional microbial agent was first mixed with a composite carrier, and then trehalose-proline composite nanoparticles and trace element substances were added and mixed thoroughly. The moisture content was adjusted to 25-30%, and the mixture was fermented at 30°C for 48 hours, dried at low temperature to a moisture content of less than 10%, and crushed to pass 80 mesh to obtain a stress-resistant and growth-promoting microbial agent.

[0056] Example 3

[0057] A stress-resistant and growth-promoting microbial agent, comprising the following raw materials in parts by weight:

[0058] 25 parts of functional microbial agents, 5 parts of trehalose-proline composite nanoparticles, 45 parts of composite carriers, and 3 parts of trace element-containing substances.

[0059] The functional microbial agent includes Bacillus siamensis, Burkholderia bifida, and Sphingomonas yunnanensis; the preservation number of the Bacillus siamensis is CCTCC AB 2022048; the preservation number of the Burkholderia bifida is CGMCC No. 1.10511; and the preservation number of the Sphingomonas yunnanensis is CGMCC No. 1.15275.

[0060] The preparation method of the functional microbial agent is as follows:

[0061] (1) Thaw Bacillus siamese and Burkholderia bifida and activate them in LB solid medium. Pick out single colonies and inoculate them into LB liquid medium. Cultivate at 27℃ and 180r / min until OD 600 =0.6 to obtain seed solution, and 1% of the seed solution was inoculated into 250 mL of LB liquid culture medium and cultured at 27 ° C and 180 r / min. The number of viable bacteria in the culture solution was detected, and the number of viable bacteria reached 1×10 8 cfu / mL and then the culture was terminated to obtain Bacillus siamensis bacterial solution and Burkholderia bifida bacterial solution respectively;

[0062] (2) Place Sphingomonas Yunnanensis in nutrient broth agar medium for activation, pick a single colony and inoculate it into nutrient broth liquid medium, and culture at 30℃ and 200r / min until OD 600 =0.6 to obtain seed solution, and 1% of the seed solution was inoculated into 250 mL of LB liquid medium and cultured at 30°C and 200 r / min. The number of viable bacteria was detected and the number reached 1×10 8 cfu / mL and the culture was terminated to obtain the Sphingomonas yunnanensis bacterial liquid;

[0063] (3) The Bacillus siamese bacterial solution and the Burkholderia bifida bacterial solution in step (1) and the Sphingomonas yunnanensis bacterial solution in step (2) are mixed in a volume ratio of 1:1:1 to obtain a functional microbial agent.

[0064] The preparation method of the trehalose-proline composite nanoparticles is as follows:

[0065] Trehalose and L-proline were dissolved in deionized water at a mass ratio of 2:1 to prepare a 10% solution; 0.1% chitosan was then added as a stabilizer; high-pressure homogenization was performed five times at 100 MPa to obtain composite nanoparticles with a particle size of 50-100 nm; and powdered trehalose-proline composite nanoparticles were obtained by spray drying.

[0066] The composite carrier includes humic acid, bentonite and biochar, and the mass ratio of the three is 3:2:1.

[0067] The trace element-containing substance includes chelated zinc, chelated iron and chelated manganese, and the mass ratio of the three is 4:3:2.

[0068] A method for preparing a stress-resistant and growth-promoting microbial agent comprises the following steps:

[0069] The three strains were expanded and cultured separately to obtain bacterial liquids of the three strains, which were mixed in equal volumes to obtain a functional microbial agent. The functional microbial agent was first mixed with a composite carrier, and then trehalose-proline composite nanoparticles and trace element substances were added and mixed thoroughly. The moisture content was adjusted to 25-30%, and the mixture was fermented at 30°C for 48 hours, dried at low temperature to a moisture content of less than 10%, and crushed to pass 80 mesh to obtain a stress-resistant and growth-promoting microbial agent.

[0070] Comparative Example 1

[0071] Compared with Example 3, this comparative example is the same as Example 3, except that the volume ratio of the siam Bacillus solution, the biphasic Burkholderia solution, and the yunnan Sphingomonas solution is 1:2:1, and the other raw materials and steps are the same as Example 3.

[0072] Comparative Example 2

[0073] Compared with Example 3, this comparative example is the same as Example 3, except that the volume ratio of the siam Bacillus solution, the biphasic Burkholderia solution, and the yunnan Sphingomonas solution is 2:1:1. The other raw materials and steps are the same as Example 3.

[0074] Comparative Example 3

[0075] Compared with Example 3, this comparative example is the same as Example 3, except that the volume ratio of the siam Bacillus solution, the Burkholderia bivariata solution, and the Sphingomonas yunnanensis solution is 1:1:2. The other raw materials and steps are the same as Example 3.

[0076] Comparative Example 4

[0077] Compared with Example 3, this comparative example uses only the siam Bacillus subtilis bacterial solution and the biphasic Burkholderia bacterial solution in a volume ratio of 1:1. The other raw materials and steps are the same as those in Example 3.

[0078] Comparative Example 5

[0079] Compared with Example 3, this comparative example uses only the siam Bacillus bacterial solution and the yunnan Sphingomonas bacterial solution in a volume ratio of 1:1. The other raw materials and steps are the same as those in Example 3.

[0080] Comparative Example 6

[0081] Compared with Example 3, this comparative example uses only the Burkholderia bispecifica solution and the Sphingomonas yunnanensis solution at a volume ratio of 1:1. The remaining raw materials and steps are the same as those of Example 3.

[0082] Comparative Example 7

[0083] Compared with Example 3, this comparative example is the same as Example 3 except that only the siam Bacillus subtilis liquid is used. The other raw materials and steps are the same as those of Example 3.

[0084] Comparative Example 8

[0085] Compared with Example 3, this comparative example uses only the biphasic Burkholderia bacterial liquid, and the other raw materials and steps are the same as those of Example 3.

[0086] Comparative Example 9

[0087] Compared with Example 3, this comparative example uses only the Sphingomonas Yunnanensis bacterial liquid, and the other raw materials and steps are the same as those of Example 3.

[0088] Performance Testing

[0089] Compatibility determination between strains

[0090] The compatibility between strains was determined by the filter paper method: the three strains were inoculated into LB liquid culture medium, placed in a shaker for 24 hours, and the culture solution was adjusted to OD 600 = 1.0, spread 100 µL onto LB medium. After drying, place a 5 mm sterile filter paper on the plate and add 3 µL of the bacterial suspension of the other strain. Incubate the plate at 30°C for three times. After 2 days, observe for the formation of inhibition zones to determine the compatibility of the three strains. Note: 1 represents Bacillus siamese, 2 represents Burkholderia bifida, and 3 represents Sphingomonas yunnanensis.

[0091] Depend on Figure 1As shown, no antagonistic zone was produced between the three strains, indicating that Bacillus siamensis, Burkholderia bifida, and Sphingomonas yunnanensis were not antagonistic to each other and were compatible with each other.

[0092] Planting trials

[0093] The experiment was conducted in a moderately saline-alkali soil area in the Yellow River Delta National Agricultural Science and Technology Park. The wheat variety used was Yannong 1212.

[0094] Experimental Design:

[0095] The experiment set up 14 treatment groups, including a blank control group (CK0), a conventional fertilization group (CK1), and microbial agents T1-T12 prepared using Examples 1-3 and Comparative Examples 1-9 of the present invention. Each treatment group was repeated 3 times, and the area of each plot was 10m 2 Each treatment group was randomly arranged, with protection rows set up around it. The specific fertilization conditions of each treatment group were as follows:

[0096] CK0: no fertilizer treatment;

[0097] CK1: conventional fertilization treatment (urea 340 kg / hm 2 , diammonium phosphate 500 kg / hm 2 , potassium sulfate 150 kg / hm 2 );

[0098] T1: 50% conventional fertilization + microbial agent of Example 1;

[0099] T2: 50% conventional fertilizer + microbial agent from Example 2;

[0100] T3: 50% conventional fertilizer + microbial agent of Example 3;

[0101] T4: 50% conventional fertilizer + microbial agent of comparative example 1;

[0102] T5: 50% conventional fertilizer + microbial agent of comparative example 2;

[0103] T6: 50% conventional fertilizer + microbial agent of comparative example 3;

[0104] T7: 50% conventional fertilizer + microbial agent of Comparative Example 4;

[0105] T8: 50% conventional fertilizer + microbial agent of comparative example 5;

[0106] T9: 50% conventional fertilizer + microbial agent of comparative example 6;

[0107] T10: 50% conventional fertilizer + microbial agent of comparative example 7;

[0108] T11: 50% conventional fertilizer + microbial agent of Comparative Example 8;

[0109] T12: 50% conventional fertilizer + microbial agent of comparative example 9;

[0110] Wheat was planted with equal row spacing, the sowing amount per mu was 15 kg, the application amount of the microbial agent in the embodiment and the comparative example was 3 kg / mu, and the conventional fertilizer was applied in a one-time base application manner. The conventional fertilization treatment mentioned also adopted a one-time base application manner, and the management measures of each treatment group were the same as the conventional field management measures.

[0111] Measurement indicators and methods:

[0112] (1) Determination of wheat growth indicators: One week before wheat harvest, wheat plants in each treatment group were sampled using the five-point method. Five plants were measured in each treatment group. The height above the root of the wheat was measured with a ruler, and the average value was finally taken.

[0113] (2) Determination of wheat stress resistance index: When the wheat was 30 days old, wheat leaves were collected from each treatment group using the five-point sampling method and then mixed. The proline content of the wheat plants was determined by the sulfosalicylic acid method, the malondialdehyde (MDA) content was determined by the thiobarbituric acid (TBA) reaction method, and the superoxide dismutase (SOD) content was determined by the nitro blue tetrazolium (NBT) photoreduction method.

[0114] (3) Determination of wheat yield and component indicators: When wheat was harvested, three 1m×1m sample plots were randomly selected for each treatment group, and the number of effective ears, number of grains per ear, and 1000-grain weight were counted. Each treatment group was repeated three times, and the yield was converted.

[0115] The specific test results are shown in the following table:

[0116] Table 1 Results of wheat plant growth and stress resistance indexes under different treatments

[0117]

[0118] As shown in Table 1, the height of wheat plants increased significantly after application of the microbial agent of the present invention compared with CK0 and CK1, indicating that the microbial agent of the present invention can significantly promote the growth of wheat plants on saline-alkali land. At 30 days, the proline content of wheat plants treated with Example 3 of the present invention reached 133.4 μg g -1, significantly increased by 113.8% and 77.4% compared to CK0 and CK1. Under salt stress conditions, plants increase their ability to resist stress by increasing the amount of substances that maintain osmotic pressure, such as proline, thereby maintaining water balance. This suggests that the microbial agent of the present invention can alter wheat's defense mechanism to cope with free radicals, increasing the proline content of wheat plants and improving their stress resistance.

[0119] As shown in Table 1, the MDA data show that, compared with the blank control group and the conventional fertilization group, the MDA content in the wheat plants after application of the microbial agent of the present invention was much lower than that in the wheat plants in CK0 and CK1. The MDA content reflects the level of lipid peroxidation in the plant membrane and the degree of cell membrane damage; the higher the content, the greater the degree of membrane damage. These results demonstrate that the application of the microbial agent of the present invention can reduce the MDA content of wheat plants, improve their stress resistance, and reduce the degree of membrane damage.

[0120] Under soil salt stress, plants produce harmful substances such as reactive oxygen species (ROS) due to dehydration, leading to an imbalance in the body's ROS levels. Superoxide dismutase (SOD) is a key component of the antioxidant enzyme system, protecting cell membranes from damage caused by ROS and inhibiting membrane lipid peroxidation, thereby alleviating stress damage to plant cells. The SOD data in Table 1 show that SOD activity in wheat plants significantly increased after application of the microbial agent according to the present invention, compared to the blank control group and the conventional fertilization group, indicating that the microbial agent according to the present invention is beneficial in improving wheat plants' stress resistance.

[0121] Table 2 Wheat yield and its components under different treatments

[0122]

[0123] As can be seen from the data in Table 2, there are significant differences in the number of ears, number of grains per ear, 1000-grain weight and yield of wheat at the harvest stage under different treatments. Compared with the blank control group and the conventional fertilization group, after applying the microbial agent of the embodiment of the present invention, the number of ears, number of grains per ear, 1000-grain weight and theoretical yield of wheat at the harvest stage are significantly increased, indicating that the application of the microbial agent of the present invention improves the stress resistance of wheat plants and can significantly promote the absorption and utilization of nutrients and the formation of yield of wheat plants in saline-alkali land.

[0124] It should be noted that the above embodiments are only some of the preferred embodiments of the present invention, and not all of them. Obviously, based on the above embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative work should fall within the scope of protection of the present invention.

Claims

1. A stress-resistant and growth-promoting microbial agent, characterized in that: The microbial agent comprises the following raw materials in parts by weight: 15-25 parts of functional microbial agent, 3-5 parts of trehalose-proline composite nanoparticles, 35-45 parts of composite carrier, and 2-3 parts of trace element-containing substance; The functional microbial agent includes Bacillus siamensis, Burkholderia bifida, and Sphingomonas yunnanensis; the deposit number of Bacillus siamensis is CCTCC AB 2022048; the deposit number of Burkholderia bifida is CGMCC No. 1.10511; and the deposit number of Sphingomonas yunnanensis is CGMCC No. 1.15275. The preparation method of the functional microbial agent is as follows: (1) Thaw Bacillus siamese and Burkholderia bifida and activate them in LB solid medium. Pick out single colonies and inoculate them into LB liquid medium. Cultivate at 27℃ and 180r / min until OD 600 =0.6 to obtain seed solution, and 1% of the seed solution was inoculated into 250 mL of LB liquid culture medium and cultured at 27 ° C and 180 r / min. The number of viable bacteria in the culture solution was detected, and the number of viable bacteria reached 1×10 8 cfu / mL and then the culture was terminated to obtain Bacillus siamensis bacterial solution and Burkholderia bifida bacterial solution respectively; (2) Place Sphingomonas Yunnanensis in nutrient broth agar medium for activation, pick a single colony and inoculate it into nutrient broth liquid medium, and culture at 30℃ and 200r / min until OD 600 =0.6 to obtain seed solution, and 1% of the seed solution was inoculated into 250 mL of LB liquid medium and cultured at 30°C and 200 r / min. The number of viable bacteria was detected and the number reached 1×10 8 cfu / mL and the culture was terminated to obtain the Sphingomonas yunnanensis bacterial liquid; (3) The Bacillus siamese bacterial solution and the Burkholderia bifida bacterial solution in step (1) and the Sphingomonas yunnanensis bacterial solution in step (2) are mixed in a volume ratio of 1:1:1 to obtain a functional microbial agent.

2. The stress-resistant and growth-promoting microbial agent according to claim 1, characterized in that The preparation method of the trehalose-proline composite nanoparticles is as follows: Trehalose and L-proline were dissolved in deionized water at a mass ratio of 2:1 to prepare a 10% solution; 0.1% chitosan was then added as a stabilizer; high-pressure homogenization was performed five times at 100 MPa to obtain composite nanoparticles with a particle size of 50-100 nm; and powdered trehalose-proline composite nanoparticles were obtained by spray drying.

3. The stress-resistant and growth-promoting microbial agent according to claim 1, characterized in that: The composite carrier includes humic acid, bentonite and biochar, and the mass ratio of the three is 3:2:

1.

4. The stress-resistant and growth-promoting microbial agent according to claim 1, characterized in that The trace element-containing substance includes chelated zinc, chelated iron and chelated manganese, and the mass ratio of the three is 4:3:

2.

5. A method for preparing the stress-resistant and growth-promoting microbial agent according to any one of claims 1 to 4, characterized in that: The method comprises the following steps: The three strains were expanded and cultured separately to obtain bacterial liquids of the three strains, which were mixed in equal volumes to obtain a functional microbial agent. The functional microbial agent was first mixed with a composite carrier, and then trehalose-proline composite nanoparticles and trace element substances were added and mixed thoroughly. The moisture content was adjusted to 25-30%, and the mixture was fermented at 30°C for 48 hours, dried at low temperature to a moisture content of less than 10%, and crushed to pass 80 mesh to obtain a stress-resistant and growth-promoting microbial agent.

6. A use of the stress-resistant and growth-promoting microbial agent according to claim 1, characterized in that: The microbial agent is used for promoting the growth of wheat and effectively improving the stress resistance of wheat growing in saline-alkali land.

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