Microbial preparation for straw decomposition and soil nutrient improvement and application thereof

Abstract

The invention discloses a microbial preparation for straw decomposition and soil nutrient improvement and application thereof, and relates to the technical field of agricultural waste recycling and soil improvement. The streptomyces sp. PLXSD-1 capable of dissolving phosphorus, fixing nitrogen and promoting straw decomposition is separated from straw returning soil, the streptomyces and a tobacco leaf extract are compounded to prepare the microbial preparation, and the microbial preparation can promote straw degradation and improve the soil nutrient condition. The tobacco leaf extract is used for improving the activity of streptomycete, promoting rapid decomposition of straw and improving the nitrogen and phosphorus supply capacity of soil. Specifically, compared with single streptomycete, after the streptomycete is compounded with a tobacco leaf extract, the phosphate solubilizing capacity is improved by 19.43%, and the straw degradation rate is remarkably increased by 25.82%. In addition, the streptomyces PLXSD-1 and the tobacco leaf extract are combined to achieve a synergistic effect on improvement of the straw degradation rate.

Description

Technical Field

[0001] This invention relates to the field of agricultural waste resource utilization and soil improvement technology, and in particular to a microbial preparation for straw decomposition and soil nutrient enhancement and its application. Background Technology

[0002] The wheat-corn double-cropping rotation system widely implemented in North China generates a large amount of crop straw annually. Returning straw to the field is the most direct way to utilize straw resources. It not only reduces the environmental pressure caused by burning or off-site disposal but also increases soil organic carbon content and improves soil structure through organic matter input, making it an important measure for achieving sustainable agricultural development. However, after straw is returned to the field, especially in the autumn and winter seasons when temperatures gradually decrease, the decomposition rate is slow. This not only affects the sowing and emergence of subsequent crops but may also increase the risk of pests and diseases due to pathogens or insect eggs carried by uncomposted straw. Therefore, how to accelerate the straw decomposition process has become a key issue that urgently needs to be addressed in the promotion of straw return technology.

[0003] Soil nutrient supply capacity directly determines crop yield and quality. Phosphorus, as one of the key elements limiting crop growth, exists in the soil primarily as insoluble inorganic phosphorus (such as calcium phosphate, iron phosphate, and aluminum phosphate) or highly stable organic phosphorus (such as phytates). Only a tiny fraction of the total soil phosphorus is readily available for direct absorption and utilization by plants, insufficient to meet the needs of normal crop growth and development. Although phosphate fertilizers are applied to increase crop yield, most phosphorus is fixed or precipitated in the soil, resulting in a utilization rate of less than 20%. This not only wastes resources but also exacerbates soil environmental risks. Furthermore, nitrogen, another essential macronutrient for plant growth and development, also faces low utilization efficiency. Conventional nitrogen fertilizers typically have a utilization rate below 40%. Excessive application not only leads to nitrogen loss but also causes ecological and environmental problems such as greenhouse gas emissions, groundwater nitrate pollution, and soil acidification. Therefore, combining efficient straw decomposition with soil nutrient enhancement to achieve soil nutrient cycling and organic matter transformation is a systemic requirement for achieving sustainable agricultural development, reducing fertilizer input, and improving soil health.

[0004] Soil microorganisms are the core drivers of soil nutrient cycling and organic matter transformation. For example, phosphorus-solubilizing microorganisms can activate insoluble inorganic and organic phosphorus by secreting metabolites such as organic acids, phosphatases, and phytases, thereby increasing the available phosphorus content in the soil; nitrogen-fixing microorganisms can fix atmospheric nitrogen into ammonium nitrogen, providing a continuous biologically available nitrogen source for crops; and cellulose-degrading microorganisms play a crucial role in the rapid decomposition of straw, the accumulation of soil organic matter, and the improvement of soil physical structure. However, the resources of functional microorganisms that can simultaneously degrade cellulose, dissolve organic and inorganic phosphorus, and fix nitrogen are relatively scarce, making it difficult for existing microbial fertilizers to meet the multiple needs of soil phosphorus activation, nitrogen supply, and straw decomposition. In the prior art, CN104894025A discloses a Streptomyces strain that can be used for straw decomposition, increasing the decomposition rate, and promoting plant seed germination.

[0005] In recent years, the application of plant-derived natural bioactive substances in agriculture has gradually attracted attention. For example, tobacco leaf extract, due to its content of alkaloids, polyphenols, and other secondary metabolites, possesses antioxidant, biostimulatory, and certain antibacterial activities. However, existing research mainly focuses on the regulatory effects of tobacco leaf extract on plant growth or pests and diseases. Currently, there is no evidence of combining Streptomyces and tobacco leaf extract to promote rapid straw decomposition or enhance soil nitrogen and phosphorus supply. Summary of the Invention

[0006] In view of the above-mentioned prior art, the purpose of this invention is to provide a microbial preparation for straw decomposition and soil nutrient enhancement, and its application. This invention isolates a Streptomyces strain from straw-returned soil that possesses phosphorus-solubilizing, nitrogen-fixing, and straw-decomposition-promoting properties. Streptomyces sp. PLXSD-1. A microbial preparation was prepared by combining Streptomyces with tobacco extract. The tobacco extract enhances the activity of Streptomyces, accelerating straw decomposition and improving soil nitrogen and phosphorus supply. Specifically, compared to Streptomyces alone, the combination with tobacco extract increased phosphorus solubility by 19.43%, with organic phosphorus increasing by 17.43% and inorganic phosphorus by 21.43%, and the straw degradation rate significantly increased by 25.82%.

[0007] To achieve the above objectives, the present invention adopts the following technical solution: In a first aspect, the present invention provides a strain of Streptomyces. Streptomyces sp. PLXSD-1, the Streptomyces Streptomyces sp. PLXSD-1 was deposited on June 17, 2025, at the China General Microbiological Culture Collection Center (CGMCC) and classified as Streptomyces. Streptomyces sp. The accession number is CGMCC NO: 34931.

[0008] In a second aspect, the present invention provides the aforementioned Streptomyces.Streptomyces sp. The PLXSD-1 is used in any of the following (1)-(3): (1) Promotes straw decomposition; (2) Used for phosphorus solubilization and nitrogen fixation to improve soil nutrient availability; (3) Prepare microbial agents for straw decomposition and soil nutrient enhancement.

[0009] A third aspect of the present invention provides a microbial preparation for straw decomposition and soil nutrient enhancement, comprising the aforementioned Streptomyces. Streptomyces sp. PLXSD-1 and tobacco leaf extract.

[0010] As a preferred choice, in microbial preparations, Streptomyces Streptomyces sp. The viable count of PLXSD-1 was 5.0 × 10⁻⁶. 8 cfu / mL -7.0×10 8 cfu / mL.

[0011] Preferably, the above-mentioned microbial preparation for straw decomposition and soil nutrient enhancement is prepared by the following method: Streptomyces Streptomyces sp. PLXSD-1 was inoculated into sodium carboxymethyl cellulose liquid enrichment medium and fermented to obtain Streptomyces fermentation broth; tobacco extract was added to the Streptomyces fermentation broth and mixed to obtain a microbial preparation.

[0012] Furthermore, the components of the sodium carboxymethyl cellulose liquid enrichment medium are: CMC-Na 8-12g, (NH4)2SO4 1.5-2.5g, KH2PO4 0.8-1.2g, MgSO4·7H2O 0.4-0.6g, NaCl 0.4-0.6g, yeast extract 0.8-1.2g, and distilled water 1000 mL.

[0013] Furthermore, during the fermentation process, the temperature is 27-32℃, the rotation speed is 150-250rpm, and the time is 2-4 days.

[0014] Furthermore, the amount of tobacco extract added is 0.005%-0.01% of the mass of the Streptomyces fermentation broth.

[0015] In a fourth aspect, the present invention provides the application of the above-mentioned microbial preparation for straw decomposition and soil nutrient enhancement in the following (1) or (2): (1) Promotes rapid decomposition of straw; (2) Enhance soil nitrogen and phosphorus supply capacity.

[0016] The beneficial effects of this invention are: This invention isolated a Streptomyces strain from soil containing straw that has been returned to the field. This strain possesses phosphorus-solubilizing, nitrogen-fixing, and straw-promoting decomposition properties. Streptomyces sp. PLXSD-1. A microbial preparation was prepared by combining Streptomyces with tobacco extract. The tobacco extract enhances the activity of Streptomyces, accelerating straw decomposition and improving soil nitrogen and phosphorus supply capacity. Specifically, compared with Streptomyces alone, the combination with tobacco extract increased phosphorus solubility by 19.43%, including a 17.43% increase in organic phosphorus and a 21.43% increase in inorganic phosphorus, and significantly increased straw degradation rate by 25.82%.

[0017] In addition, the present invention uses Streptomyces Streptomyces sp. The combination of PLXSD-1 and tobacco extract has a synergistic effect in improving the degradation rate of straw. Attached Figure Description

[0018] Figure 1 Example 1: Filter paper enzyme activity diagram of cellulose-degrading bacteria; Figure 2 Example 1: Straw degradation rate diagram of cellulose-degrading bacteria; Figure 3 Phylogenetic tree of 16S rDNA of Streptomyces PLXSD-1; Figure 4 : In Experiment 1, the growth of Streptomyces PLXSD-1 on nitrogen-free nitrogen-fixing medium. Detailed Implementation

[0019] It should be noted that the following detailed descriptions are illustrative and intended to provide further explanation of this application. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains.

[0020] To enable those skilled in the art to better understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0021] The experimental materials used in the embodiments of this invention are all conventional experimental materials in the art and can be purchased through commercial channels.

[0022] In this invention, the tobacco leaf extract was purchased from Ningbo (Xi'an) Biomedical Technology Co., Ltd., and the pure water extraction ratio was 100:1.

[0023] The components of the sodium carboxymethyl cellulose (CMC-Na) liquid enrichment medium are: 10 g CMC-Na, 2.0 g (NH4)2SO4, 1 g KH2PO4, 0.5 g MgSO4·7H2O, 0.5 g NaCl, 1 g yeast extract powder, and 1000 mL distilled water.

[0024] The components of sodium carboxymethyl cellulose (CMC-Na) solid culture medium are: 10 g CMC-Na, 2.0 g (NH4)2SO4, 1 g KH2PO4, 0.5 g MgSO4·7H2O, 0.5 g NaCl, 1 g yeast extract powder, 20 g agar, and 1000 mL distilled water.

[0025] The inorganic phosphorus-solubilizing medium consists of: 10 g glucose, 0.5 g MgSO4·7H2O, 0.5 g (NH4)2SO4, 0.3 g KCl, 0.3 g NaCl, 22 g Ca3(PO4)2, 0.1 g MnSO4, 20 g agar, and 1000 mL distilled water.

[0026] The composition of the phytate organic phosphorus soluble medium is as follows: 10 g glucose, 0.5 g (NH4)2SO4, 2 g calcium phytate, 0.5 g MgSO4·7H2O, 0.3 g KCl, 0.1 g MnSO4, 20 g agar, and 1000 mL distilled water.

[0027] The nitrogen-free nitrogen-fixing medium consists of: 10 g sucrose, 1 g KH2PO4, 0.5 g MgSO4·7H2O, 0.3 g NaCl, 1 g CaCO3, 20 g agar, and 1000 mL distilled water.

[0028] Example 1: Isolation and Identification of Streptomyces In fields where straw return to the field has been carried out for 5 consecutive years, topsoil samples of 0-20cm were taken during the wheat overwintering period (after corn straw return to the field). After removing stones and fallen leaves from the soil, corn straw cellulose degrading strains were screened.

[0029] 1. Strains are isolated and purified: 5g of sieved soil was poured into a 250mL Erlenmeyer flask. 45mL of sterile sodium carboxymethyl cellulose (CMC-Na) liquid enrichment medium was added to the flask. After sealing, the flask was placed in a constant temperature shaker at 30℃ and 200rpm for 48 hours. The flask was then removed and allowed to stand for 10 minutes. After filtering through gauze, the supernatant was collected and diluted with sterile water to a final concentration of 10. -3 10 -4 and 10 -5 After three dilutions, 100 µL of the diluted bacterial solution was taken and spread onto sodium carboxymethyl cellulose (CMC-Na) solid medium. Three plates were spread for each dilution gradient and incubated upside down at 30 °C for 3 days until colonies grew. Then, the strain was purified by incubating upside down at 30 °C for 3 days until a single strain grew.

[0030] 2. Strain screening (1) Preliminary screening of strains: The Congo red staining method-hydrolysis zone assay was used for initial screening of straw cellulose-degrading strains. The specific steps were as follows: The purified strain was inoculated onto sodium carboxymethyl cellulose solid medium and cultured at 30°C for 3 days. After staining with 1 g / L Congo red for 30 min, the stain was discarded. Then, the strain was eluted with 1 mol / L NaCl solution for 30 min. The diameter of the transparent zone (H) and the diameter of the colony (C) were measured with a ruler, and the H / C ratio was calculated. The results are shown in Table 1.

[0031] Table 1. Degradation capacity of cellulose-degrading bacteria on CMC-Na solid medium As can be seen from Table 1, strains 1-1, 1-2, 1-3, 1-4, and 1-5 have higher H / C values, indicating stronger straw cellulose degradation capabilities.

[0032] (2) Secondary screening of strains: The strains obtained from the initial screening were rescreened using the DNS colorimetric method-filter paper disintegration test. The specific steps were as follows: Using filter paper as a substrate, the reaction was carried out at 50℃ for 1 h, and the glucose production was determined by the DNS method. The cellulose-degrading bacteria obtained from the initial screening were added to sodium carboxymethyl cellulose (CMC-Na) liquid enrichment medium and fermented at 30℃ and 180 rpm for 3 days to obtain the bacterial fermentation broth. The viable cell count was adjusted to 1×10⁻⁶ with sterile water. 7 The concentration of cfu / mL was determined by centrifugation of the fermentation broth, and the supernatant was collected to obtain the crude enzyme solution. 0.5 mL of the crude enzyme solution, 1.0 mL of pH 4.8 acetate buffer, and 50 mg of filter paper strips were mixed and reacted at 50°C for 1 h. The reaction was terminated by boiling in a water bath. The amount of glucose produced was determined by the DNS method, and the results are as follows: Figure 1 As shown.

[0033] Depend on Figure 1 It can be seen that strains 1-1, 1-2, 1-3, and 1-4 have higher filter paper enzyme activity and stronger straw cellulose degradation ability.

[0034] (3) Application verification using straw degradation rate, the specific steps are as follows: The straw was crushed to 1-2 cm and sterilized for later use. 5 mL of fermentation broth and 45 mL of inorganic salt liquid culture medium were mixed, and 5 g of straw was added. The mixture was then incubated at 30℃ and 180 rpm with shaking. The inorganic salt liquid culture medium consisted of: (NH4)2SO4 2.0 g, KH2PO4 1 g, MgSO4·7H2O 0.5 g, NaCl 0.5 g, yeast extract powder 1 g, and distilled water 1000 mL. An uninoculated straw culture medium was used as a control group. Each group was divided into three replicates. The degradation capacity was evaluated using the straw degradation rate (%). The results are shown below. Figure 2 As shown.

[0035] The straw degradation rate (%) is calculated as follows: [(initial straw mass - degraded straw mass) / initial straw mass] × 100.

[0036] Depend on Figure 2 It can be seen that strain 1-1 has the strongest ability to degrade straw cellulose, and it has been identified as the target strain.

[0037] 3. Strain identification Biological identification was performed on the screened strain 1-1, and its phylogenetic tree was constructed, such as... Figure 3 As shown. Based on biological identification and phylogenetic analysis, this bacterium was identified as Streptomyces.

[0038] 4. Strain preservation The isolated Streptomyces strains were preserved, and the preservation information is as follows: Strain name: Streptomyces; Latin name: Streptomyces sp. ; Strain number: PLXSD-1; Preservation institution: China General Microbiological Culture Collection Center (CGMCC); Deposit date: June 17, 2025; Registered with the China Museum Collection Center: CGMCC NO: 34931.

[0039] Experimental Example 1: Determination of nitrogen fixation and phosphorus solubilization capabilities of Streptomyces PLXSD-1 1. Using the spot inoculation method, the purified Streptomyces PLXSD-1 single strain was inoculated onto inorganic phosphate-solubilizing medium and phytate organic phosphate-solubilizing medium, respectively, and cultured at 30℃ for 3-7 days. The presence or absence of a clear transparent zone (phosphate-solubilizing zone) around the colony was used as the criterion. After culture, the colony diameter and the diameter of the transparent zone were measured, and the results are shown in Table 2.

[0040] Table 2. Phosphorus solubilization capacity (organic and inorganic phosphorus) of Streptomyces PLXSD-1 As shown in Table 2, Streptomyces PLXSD-1 has the ability to dissolve both organic and inorganic phosphorus.

[0041] 2. Using the spot inoculation method, the purified Streptomyces PLXSD-1 single strain was inoculated onto nitrogen-free nitrogen-fixing medium and cultured at 30°C for 3–7 days. Figure 4 It can be seen that Streptomyces PLXSD-1 can form colonies / mycelia on nitrogen-free nitrogen-fixing medium, which proves that the strain has nitrogen-fixing ability.

[0042] Experimental Example 2: This experiment consisted of four treatment groups: Treatment group 1: CK group, water control group; Treatment group 2: Streptomyces PLXSD-1 group; Treatment group 3: Tobacco extract group; Treatment Group 4: Microbial Preparation Group; The microbial preparation is prepared by the following method: Streptomyces PLXSD-1 is inoculated into sodium carboxymethyl cellulose (CMC-Na) liquid enrichment medium and fermented in liquid form at 30℃ and 200 rpm for 3 days to obtain Streptomyces fermentation broth; Tobacco leaf extract is added to the Streptomyces fermentation broth at a dosage of 0.005% (mass ratio), and mixed well to obtain the microbial preparation; wherein, the microbial preparation contains Streptomyces Streptomyces sp. The viable count of PLXSD-1 was 6.0 × 10⁻⁶. 8 cfu / mL.

[0043] The filter paper enzyme activity, phosphorus solubilization capacity, and straw degradation rate of the four treatment groups were measured, and the results are shown in Table 3. Filter paper enzyme activity: The filter paper enzyme activity of each treatment group was determined by the DNS colorimetric method-filter paper disintegration test, with the specific steps being the same as in Example 1. Straw degradation rate: The substances of each treatment group were mixed with inorganic salt liquid culture medium, straw was added, and the mixture was shaken and cultured for 7 days. The straw degradation rate was then calculated, with the specific steps being the same as in Example 1. Phosphorus solubilization capacity: The substances of each treatment group were added to inorganic phosphorus-solubilizing medium and phytate organic phosphorus-solubilizing medium, respectively, and the diameter (H) of the clear zone of the colonies in each treatment group was measured, with the specific steps being the same as in Example 1.

[0044] Table 3. Functional indicators of Streptomyces PLXSD-1 combined with tobacco extract. As shown in Table 3, compared with single strains, the combination of strains and tobacco extract increased filter paper enzyme activity by 23.61%, enhanced phosphorus solubilization capacity by 19.43% (organic phosphorus by 17.43% and inorganic phosphorus by 21.43%), and significantly increased straw degradation rate by 25.82%. This indicates that tobacco extract can enhance the activity of Streptomyces.Streptomyces sp. PLXSD-1 has the ability to solubilize phosphorus and degrade cellulose, and accelerates the decomposition of straw.

[0045] Regarding straw degradation rate, treatment with only tobacco extract for 7 days resulted in a 0.1% increase in straw degradation rate compared to the control group (CK); treatment with only Streptomyces PLXSD-1 for 7 days resulted in a 15.2% increase in straw degradation rate compared to the CK; while treatment with the microbial preparation obtained by combining tobacco extract and Streptomyces PLXSD-1 for 7 days, as described in this application, resulted in a 21.5% increase in straw degradation rate compared to the CK. This demonstrates that tobacco extract and Streptomyces PLXSD-1 have a synergistic effect in improving straw degradation rate.

[0046] The above description is merely a preferred embodiment of this application and is not intended to limit this application. Various modifications and variations can be made to this application by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this application should be included within the protection scope of this application.

Claims

1. A strain of Streptomyces Streptomyces sp. PLXSD-1, with accession number CGMCC NO: 34931.

2. The Streptomyces as described in claim 1 Streptomyces sp. The PLXSD-1 is used in any of the following (1)-(3): (1) Promotes straw decomposition; (2) Used for phosphorus solubilization and nitrogen fixation to improve soil nutrient availability; (3) Prepare microbial agents for straw decomposition and soil nutrient enhancement.

3. A microbial preparation for straw decomposition and soil nutrient enhancement, characterized in that, Including the Streptomyces as described in claim 1 Streptomyces sp. PLXSD-1 and tobacco leaf extract.

4. The microbial preparation for straw decomposition and soil nutrient enhancement as described in claim 3, characterized in that, In microbial preparations, Streptomyces Streptomyces sp. The viable count of PLXSD-1 was 5.0 × 10⁻⁶. 8 cfu / mL -7.0×10 8 cfu / mL.

5. The microbial preparation for straw decomposition and soil nutrient enhancement as described in claim 3, characterized in that, It is prepared by the following method: Streptomyces Streptomyces sp. PLXSD-1 was inoculated into sodium carboxymethyl cellulose liquid enrichment medium and fermented to obtain Streptomyces fermentation broth; tobacco extract was added to the Streptomyces fermentation broth and mixed to obtain a microbial preparation.

6. The microbial preparation for straw decomposition and soil nutrient enhancement as described in claim 5, characterized in that, The components of the sodium carboxymethyl cellulose liquid enrichment medium are: CMC-Na 8-12 g, (NH4)2SO4 1.5-2.5 g, KH2PO4 0.8-1.2 g, MgSO4·7H2O 0.4-0.6 g, NaCl 0.4-0.6 g, yeast extract 0.8-1.2 g, and distilled water 1000 mL.

7. The microbial preparation for straw decomposition and soil nutrient enhancement as described in claim 5, characterized in that, During the fermentation process, the temperature is 27-32℃, the rotation speed is 150-250rpm, and the time is 2-4 days.

8. The microbial preparation for straw decomposition and soil nutrient enhancement as described in claim 5, characterized in that, The amount of tobacco extract added is 0.005%-0.01% of the mass of the Streptomyces fermentation broth.

9. The application of the microbial preparation for straw decomposition and soil nutrient enhancement according to any one of claims 3-8 in (1)-(2): (1) Promotes rapid decomposition of straw; (2) Enhance soil nitrogen and phosphorus supply capacity.

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