Targeted biological bacterium coupling method for comprehensive treatment of salt damage, heavy metal and pesticide residues of coastal vegetables and application of targeted biological bacterium coupling method

Abstract

The invention discloses a targeted biological bacterium coupling method for comprehensive treatment of coastal vegetable salt damage, heavy metal and pesticide residues and application, and relates to the field of agricultural biotechnology and soil remediation. According to the method, bacillus subtilis, bacillus mucilaginosus, brevibacillus laterosporus, pseudomonas fluorescens, arbuscular mycorrhizal fungi and host original flora extracted from healthy vegetable rhizosphere soil are subjected to coupled symbiotic culture, and target flora with multiple functions is formed. The functional flora is compounded with a nutrient carrier containing potassium humate and fulvic acid, a seaweed extract and a specific salt-resistant additive to prepare the multifunctional water-soluble fertilizer or fungicide. After the compound fertilizer is applied to a coastal vegetable production base, the synergistic effects of degrading soil pesticide residues, passivating heavy metals, enhancing the salt tolerance of vegetables and improving the soil fertility and microbial diversity can be synchronously realized, the compound obstacle problem of coastal vegetable production is fundamentally solved, and the yield, quality and safety of vegetables are guaranteed.

Description

Technical Field

[0001] This invention relates to the fields of agricultural biotechnology and soil remediation technology, specifically to a method for constructing a targeted microbial community and its application in the comprehensive management of soil salinity, heavy metal pollution and pesticide residues in coastal vegetable production bases. Background Technology

[0002] my country's coastal areas have a large number of vegetable production bases, but these bases have long been engaged in intensive production and generally face a complex pollution problem caused by factors such as seawater intrusion and overuse of chemical fertilizers and pesticides: soil salinization leads to physiological drought in vegetables, resulting in nutrient absorption imbalance and stunted growth; heavy metals such as cadmium, lead, and arsenic in the soil are easily accumulated by vegetables and can harm human health through the food chain; continuous cropping obstacles and frequent soil-borne diseases lead to pesticide dependence, causing soil and agricultural product pollution.

[0003] Current remediation technologies are mostly piecemeal, such as using single microbial agents to degrade pesticide residues, using passivating agents to treat heavy metals, or using salt-resistant agents to alleviate salt damage. However, single salt-resistant agents can only alleviate salt damage and cannot solve the problems of heavy metals and pesticide residues; heavy metal passivating agents (such as lime and bentonite) have short-lived effects and are easily leached away by irrigation; microbial agents are mostly single strains with limited functions, only targeting a certain type of pollution, and the strains have poor compatibility with the native soil flora, and their activity is easily inhibited by salt stress; the combination of nutrient carriers and biological bacteria is unreasonable, making it difficult to meet the growth needs of the microbial community and the nutritional supply to vegetables, resulting in poor integrated remediation effects and failing to balance vegetable yield, quality, and safety. Therefore, developing a green, efficient, and sustainable technology that can simultaneously target and solve salt damage, heavy metals, and pesticide residues has become an urgent need for the sustainable development of coastal agriculture. Summary of the Invention

[0004] This invention aims to provide a targeted biological microbial coupling method and its application for the synergistic treatment of salt damage, heavy metals and pesticide residues in coastal vegetable bases.

[0005] The technical solution of the present invention to achieve the above objectives is as follows: An embodiment of the present invention provides a targeted biological-microbe coupling method for the comprehensive management of salt damage, heavy metals, and pesticide residues in coastal vegetables, comprising the following steps: a. Construct a targeted biological strain resource bank and screen for functional single strains; b. Purify and compound the single strain; c. Couple and symbiotically culture the compound microbial community with the host original microbial community extracted from the rhizosphere soil of healthy vegetables; d. The coupled-cultured composite functional microbial community is mixed with a nutrient carrier containing salt-tolerant adjuvants to prepare a microbial agent or water-soluble fertilizer for vegetable cultivation. The targeted biological microbial coupling method proposed in this invention improves the compatibility and environmental adaptability of the microbial community by screening and compounding multifunctional single strains and combining them with native soil host microbial communities for coupled symbiotic culture. The combination of the compound functional microbial community with a nutrient carrier containing salt-resistant adjuvants can not only enable the microbial community to degrade pesticide residues, passivate heavy metals, and improve soil structure, but also alleviate salt damage and supplement the nutrients needed for vegetable growth through salt-resistant adjuvants, thereby achieving synergistic treatment of salt damage, heavy metals, and pesticide residues, while improving the stress resistance and growth performance of vegetables. Optionally, the functional single strain mentioned in step a includes at least three of Bacillus subtilis, Bacillus mucilaginosus, Bacillus brevis, Pseudomonas fluorescens, and arbuscular mycorrhizal fungi. By screening five strains with specific functions, Bacillus subtilis can degrade organophosphate pesticide residues and inhibit pathogens; Bacillus mucilaginosus can activate soil nutrients and passivate heavy metals; Bacillus lateralis can degrade pyrethroid pesticide residues and enhance the stress resistance of vegetables; Pseudomonas fluorescens can secrete siderophores, promote nutrient absorption and reduce the availability of heavy metals; and Arbuscular mycorrhizal fungi can form a symbiotic relationship with vegetable roots, expand the absorption area and enhance salt tolerance. The combination of at least three strains ensures complementary functions and achieves synergistic management. Optionally, the concentration requirements for each strain in the functional microbial community are: Bacillus subtilis ≥ 5 × 10⁻⁶ 8 CFU / ml, Bacillus subtilis ≥3×10 8 CFU / ml, Bacillus retrosporum ≥2×10 8 CFU / ml, Fluorescent Pseudomonas ≥1×10⁻⁶ 8 CFU / ml, arbuscular mycorrhizal fungi ≥50 spores / g, host original flora ≥1×10 8 CFU / ml. Define the minimum concentration standard for each strain to ensure that the bacterial community has sufficient activity and functional strength, and avoid the decline in treatment effect due to insufficient concentration; control the concentration of the original host bacterial community to ensure its balanced symbiosis with the compound bacterial community and enhance the colonization ability of the compound bacterial community in the soil. Optionally, the nutrient carrier described in step d includes potassium humate, seaweed extract, amino acid chelated trace elements, potassium dihydrogen phosphate, sucrose or molasses, and salt-resistant agents. The components of the nutrient carrier complement each other: humic acid and potassium fulvate improve soil structure and promote microbial reproduction; seaweed extract enhances the stress resistance of vegetables; amino acid chelates and trace elements are easily absorbed by vegetables, supplementing the nutrients needed for growth; potassium dihydrogen phosphate provides phosphorus and potassium nutrition and improves photosynthetic efficiency; sucrose or molasses provides a carbon source for microbial growth and maintains microbial activity; salt-resistant adjuvants specifically alleviate salt damage, ensuring that each component synergistically supports microbial function and vegetable growth. Optionally, the salt-resistant adjuvant comprises, by weight, 1-1.5 parts urea, 0.003-0.006 parts naphthaleneacetic acid, 0.02-0.06 parts humic acid, 0.3-0.6 parts mannitol, 0.1-0.4 parts ammonium sulfate, 0.01-0.06 parts proline, 0.01-0.04 parts arginine, 0.005-0.01 parts surfactant, and 90-100 parts water. The salt-resistant adjuvant has a precise ratio of each component: urea and ammonium sulfate provide nitrogen sources to alleviate nutrient deficiency under salt stress; naphthaleneacetic acid promotes root growth and enhances water and fertilizer absorption capacity; humic acid regulates soil osmotic pressure; mannitol, proline, and arginine act as osmotic regulators to enhance the water retention capacity of vegetable cells and reduce salt ion toxicity; surfactants improve adjuvant dispersibility and absorption efficiency of vegetable leaves / roots, synergistically enhancing the salt resistance effect. Optionally, the nutrient carrier in each ton of finished water-soluble fertilizer comprises: 40-60 kg of potassium humate, 20-40 kg of seaweed extract, 5-15 kg of amino acid chelated trace elements, 15-25 kg of potassium dihydrogen phosphate, 10-20 kg of sucrose or molasses, and 30-50 kg of salt-resistant adjuvant. The dosage range of each nutrient carrier in each ton of finished water-soluble fertilizer is clearly defined to ensure that the nutrient supply matches the activity requirements of the microbial community and avoid the effect of use due to excessive or insufficient components. The dosage range is set to take into account the degree of soil pollution and the differences in vegetable varieties in different coastal areas, so as to improve the product adaptability. This invention also provides a multifunctional targeted biological bacterial water-soluble fertilizer or bacterial agent prepared by any of the methods described above. This product integrates the synergistic effects of compound functional microbial communities, nutrient components, and salt-resistant adjuvants. The form can be selected according to the application scenario: water-soluble fertilizer (liquid) or microbial agent (solid granules). The water-soluble fertilizer is convenient for drip irrigation and irrigation, while the microbial agent is convenient for soil base application. It is flexible in use and can be directly applied to coastal vegetable cultivation and soil remediation. This invention also provides a cultivation method to improve the yield, quality and safety of coastal vegetables. Based on the above-mentioned multifunctional targeted biological bacteria water-soluble fertilizer or bacterial agent, the multifunctional targeted biological bacteria water-soluble fertilizer or bacterial agent is applied regularly during the critical growth and development period of vegetables. Targeting the growth characteristics of coastal vegetables, the product is applied during critical periods to ensure timely colonization and function of the microbial community in the soil. The precise supply of nutrients and salt-resistant adjuvants effectively mitigates the negative effects of salt damage, heavy metals, and pesticide residues, thereby specifically increasing vegetable yield, improving quality (such as vitamin content and taste), reducing pollutant residues, and ensuring food safety. Optionally, the vegetable is a leafy vegetable or a fruit vegetable; the key growth and development period includes the seedling stage, the vigorous growth stage and the pre-harvest stage; the application method is to dilute it 500-800 times and then apply it to the roots by irrigation or drip irrigation, and to spray the salt-resistant adjuvant on the leaves during periods of significant salt stress. Clearly define the applicable vegetable types to expand the scope of technology application; select key periods in line with the growth patterns of vegetables, apply during the seedling stage to promote root development and establish stress resistance, supplement nutrition and enhance the treatment effect during the vigorous growth period, and consolidate quality and reduce residues before harvest; limit the dilution ratio and application method to ensure that the product effect is fully realized, avoid excessive concentration that burns crops or excessive concentration that is ineffective, and apply salt-resistant adjuvants on the leaves during the salt stress period to specifically alleviate salt damage and improve the precision of treatment. The embodiments of the present invention also provide the application of any of the above-described methods or the multifunctional targeted biological bacteria water-soluble fertilizers or agents in the remediation of coastal salinized, heavy metal polluted, and pesticide-residual soils. Expanding the application scenarios of the technical solution, in addition to its direct application in vegetable cultivation, it can also be used for the ecological restoration of polluted coastal soils. Through the degradation and passivation effects of the compound microbial community and the improvement effect of the nutrient carrier, it can reduce soil salinity, heavy metal activity and pesticide residue content, restore soil ecological function, lay the foundation for subsequent crop planting, and enhance the practical value and promotion prospects of the technology.

[0006] The beneficial effects of this invention are: Collaborative governance: A single solution addresses the three major challenges of salt, heavy metals, and pesticide residues simultaneously, overcoming the limitations of traditional technologies with their single function.

[0007] Highly targeted: The microbial community is designed with specific targets, with Bacillus lateralis specifically passivating heavy metals, and salt-resistant adjuvants precisely alleviating salt stress, with clearly defined functions.

[0008] Eco-friendly: Through microbial remediation and ecological regulation, the use of chemical fertilizers and pesticides is significantly reduced, and the soil micro-ecology is restored.

[0009] 4. Improve quality and increase yield: While ensuring safety, improve vegetable yield and nutritional quality by promoting plant growth and secondary metabolism. Detailed Implementation

[0010] The present invention will be further described below with reference to the embodiments. Example

[0011] Preparation of multifunctional targeted bio-based water-soluble fertilizer: 1. Activation and propagation of strains: Bacillus subtilis, Bacillus mucilaginosus, Bacillus brevichorus, and Pseudomonas fluorescens were activated and fermented at high density, and arbuscular mycorrhizal fungi were propagated separately.

[0012] 2. Extraction of original host microbiota: Samples were collected from the rhizosphere soil of healthy spinach or bok choy, and the original host microbiota were obtained through enrichment culture and centrifugation purification.

[0013] 3. Coupled symbiotic culture: The four bacterial strains obtained in step 1 were mixed according to the ratio of viable cells, and then mixed with the original host flora at a volume ratio of 10:1. The mixture was cultured at 28℃ and pH 6.5 for 72 hours with aeration. Finally, arbuscular mycorrhizal fungal spores were added and stirred thoroughly to form a concentrated solution of the composite functional flora. The concentration of each strain met the following requirements: Bacillus subtilis ≥ 5 × 10⁻⁶. 8 CFU / ml, Bacillus subtilis ≥3×10 8 CFU / ml, Bacillus retrosporum ≥2×10 8 CFU / ml, Fluorescent Pseudomonas ≥1×10⁻⁶ 8 CFU / ml, arbuscular mycorrhizal fungi ≥50 spores / g, host original flora ≥1×10 8 CFU / ml.

[0014] 4. Formulation preparation: The concentrated solution of the coupled cultured compound functional microbial community is evenly mixed with the nutrient carrier to prepare water-soluble fertilizer.

[0015] The salt-resistant additives, by weight, include: 1 part urea, 0.003 parts naphthaleneacetic acid, 0.02 parts humic acid, 0.3 parts mannitol, 0.1 parts ammonium sulfate, 0.01 parts proline, 0.01 parts arginine, 0.005 parts surfactant, and 90 parts water.

[0016] Nutrient carrier mixing: Weigh out 40 kg of potassium humate, 20 kg of seaweed extract, 5 kg of amino acid chelated trace elements, 15 kg of potassium dihydrogen phosphate, 10 kg of sucrose or molasses, and 30 kg of salt-resistant agent per ton of finished product, and mix at 100 r / min for 30 min.

[0017] Preparation of water-soluble fertilizer: Add compound functional microbial concentrate at a dosage of 50L per ton of finished product, mix at 120r / min for 60min to obtain water-soluble fertilizer. Example

[0018] Field application trials and efficacy verification of the water-soluble fertilizer prepared in Example 1.

[0019] An experiment was conducted at a leafy vegetable base along the coast of Jiangsu Province, with experimental and control groups set up. The experimental group was a planting group that received water-soluble fertilizer, while the control group was a conventional planting group that did not receive the multifunctional targeted biological water-soluble fertilizer and salt-resistant adjuvant of this invention. Other cultivation and management measures were the same as those for the experimental group. Water-soluble fertilizer application method: During the seedling stage and vigorous growth stage, the water-soluble fertilizer was diluted 500 times and applied to the roots twice; during the high-risk period of salt damage, the salt-resistant adjuvant was sprayed on the leaves once with a foliar spray diluted 200 times.

[0020] Experimental results: 1. Salt damage relief: The vegetable plants in the experimental group grew normally, with bright green leaves, and the salt damage symptoms were significantly milder than those in the control group.

[0021] 2. Heavy metal content: The Cd and Pb contents in the experimental group of bok choy were reduced by 38.5% and 32.1% respectively compared with the control group.

[0022] 3. Pesticide residues: The amount of chlorpyrifos residues detected in the soil of the experimental group was 45.0% lower than that of the control group.

[0023] 4. Yield and quality: The vegetable yield in the experimental group increased by 12.7%, and the vitamin C content increased by 18.3%.

[0024] As can be seen from the above results, this invention has successfully developed a green product and technical solution that can systematically solve the complex obstacles in coastal vegetable production bases by coupling specific functional bacterial groups with the host's original bacterial groups and combining them with a nutrient carrier that integrates salt resistance. This provides effective technical support for ensuring the safe production and supply of vegetables in coastal areas. Example

[0025] This embodiment discloses the preparation process of multifunctional targeted biological bacterial water-soluble fertilizers with different ratios: 1. Activation and propagation of strains: Bacillus subtilis, Bacillus mucilaginosus, Bacillus brevichorus, and Pseudomonas fluorescens were activated and fermented at high density, and arbuscular mycorrhizal fungi were propagated separately.

[0026] 2. Extraction of original host microbiota: Samples were collected from the rhizosphere soil of healthy spinach or bok choy, and the original host microbiota were obtained through enrichment culture and centrifugation purification.

[0027] 3. Coupled symbiotic culture: The four bacterial strains obtained in step 1 were mixed according to the ratio of viable cells, and then mixed with the original host flora at a volume ratio of 10:1. The mixture was cultured at 30℃ and pH 7 for 72 hours with aeration. Finally, arbuscular mycorrhizal fungal spores were added and stirred thoroughly to form a concentrated solution of the composite functional flora. The concentration of each strain met the following requirements: Bacillus subtilis ≥ 5 × 10⁻⁶. 8 CFU / ml, Bacillus subtilis ≥3×10 8 CFU / ml, Bacillus retrosporum ≥2×10 8 CFU / ml, Fluorescent Pseudomonas ≥1×10⁻⁶ 8 CFU / ml, arbuscular mycorrhizal fungi ≥50 spores / g, host original flora ≥1×10 8 CFU / ml.

[0028] 4. Formulation preparation: The concentrated solution of the coupled cultured compound functional microbial community is evenly mixed with the nutrient carrier to prepare water-soluble fertilizer.

[0029] The salt-resistant additives, by weight, include: 1.5 parts urea, 0.006 parts naphthaleneacetic acid, 0.06 parts humic acid, 0.6 parts mannitol, 0.4 parts ammonium sulfate, 0.06 parts proline, 0.04 parts arginine, 0.01 parts surfactant, and 100 parts water.

[0030] Nutrient carrier mixing: per ton of finished product, mix 60 kg of potassium humate, 40 kg of seaweed extract, 15 kg of amino acid chelated trace elements, 25 kg of potassium dihydrogen phosphate, 20 kg of sucrose or molasses, and 50 kg of salt-resistant agent at 100 r / min for 60 min.

[0031] Preparation of water-soluble fertilizer: Add compound functional microbial concentrate at a dosage of 50L per ton of finished product, mix at 120r / min for 60min to obtain water-soluble fertilizer.

[0032] The above embodiments are only for illustrating the technical concept and features of the present invention, and are intended to enable those skilled in the art to understand the content of the present invention and implement it accordingly. They should not be construed as limiting the scope of protection of the present invention. All equivalent changes or modifications made in accordance with the spirit and essence of the present invention should be covered within the scope of protection of the present invention.

Claims

1. A targeted biological-microbial coupling method for the integrated management of salt damage, heavy metals, and pesticide residues in coastal vegetables, characterized in that, Includes the following steps: a. Construct a targeted biological strain resource bank and screen for functional single strains; b. Purify and compound the single strain; c. Couple and symbiotically culture the compound microbial community with the host original microbial community extracted from the rhizosphere soil of healthy vegetables; d. The coupled-cultured composite functional microbial community is mixed with a nutrient carrier containing salt-tolerant adjuvants to prepare a microbial agent or water-soluble fertilizer for vegetable cultivation.

2. The targeted biological-microbial coupling method for the comprehensive management of salt damage, heavy metals, and pesticide residues in coastal vegetables according to claim 1, characterized in that, The functional single strain mentioned in step a includes at least three of Bacillus subtilis, Bacillus mucilaginosus, Bacillus brevis, Pseudomonas fluorescens, and arbuscular mycorrhizal fungi.

3. The targeted biological-microbial coupling method for the comprehensive management of salt damage, heavy metals, and pesticide residues in coastal vegetables according to claim 2, characterized in that, The required concentration of each strain in the functional microbial community is: Bacillus subtilis ≥ 5 × 10⁻⁶. 8 CFU / ml, Bacillus subtilis ≥3×10 8 CFU / ml, Bacillus retrosporum ≥2×10 8 CFU / ml, Fluorescent Pseudomonas ≥1×10⁻⁶ 8 CFU / ml, arbuscular mycorrhizal fungi ≥50 spores / g, host original flora ≥1×10 8 CFU / ml.

4. The targeted biological-microbial coupling method for the integrated management of salt damage, heavy metals, and pesticide residues in coastal vegetables according to claim 1, characterized in that, The nutrient carriers mentioned in step d include potassium humate, seaweed extract, amino acid chelated trace elements, potassium dihydrogen phosphate, sucrose or molasses, and salt-resistant agents.

5. The targeted biological-microbial coupling method for the comprehensive management of salt damage, heavy metals, and pesticide residues in coastal vegetables according to claim 4, characterized in that, The salt-resistant adjuvant comprises, by weight, 1-1.5 parts urea, 0.003-0.006 parts naphthaleneacetic acid, 0.02-0.06 parts humic acid, 0.3-0.6 parts mannitol, 0.1-0.4 parts ammonium sulfate, 0.01-0.06 parts proline, 0.01-0.04 parts arginine, 0.005-0.01 parts surfactant, and 90-100 parts water.

6. The targeted biological-microbial coupling method for the comprehensive management of salt damage, heavy metals, and pesticide residues in coastal vegetables according to claim 5, characterized in that, The nutrient carrier in each ton of finished water-soluble fertilizer comprises: 40-60 kg of potassium humate, 20-40 kg of seaweed extract, 5-15 kg of amino acid chelated trace elements, 15-25 kg of potassium dihydrogen phosphate, 10-20 kg of sucrose or molasses, and 30-50 kg of salt-resistant adjuvant.

7. A multifunctional targeted biological bacterial water-soluble fertilizer or bacterial agent prepared by the method described in any one of claims 1-6.

8. A cultivation method for improving the yield, quality, and safety of coastal vegetables, based on the multifunctional targeted biological microbial water-soluble fertilizer or microbial agent described in claim 7, characterized in that, During the critical growth and development period of vegetables, apply the aforementioned multifunctional targeted biological microbial water-soluble fertilizer or microbial agent regularly.

9. The cultivation method according to claim 8, characterized in that, The vegetables mentioned are leafy vegetables or fruit vegetables; The critical growth and development periods include the seedling stage, the vigorous growth period, and the pre-harvest stage. The application method is to dilute it 500-800 times and then apply it to the roots by irrigation or drip irrigation, and to spray the salt-resistant adjuvant on the leaves during periods of significant salt stress.

10. The application of the method according to any one of claims 1-6 or the multifunctional targeted biological bacterial water-soluble fertilizer or bacterial agent according to claim 7 in the remediation of coastal salinized, heavy metal polluted and pesticide residue soils.