Method for preparing functional biological flora by coupling target biological flora and host original flora and application of functional biological flora in ginseng

Abstract

The invention discloses a method for preparing a functional biological flora in a targeted biological bacterium and host original flora coupling mode and application of the functional biological flora in ginseng, and relates to the field of agricultural microbial technology and traditional Chinese medicinal material cultivation, and the method comprises the following steps: a, constructing a targeted biological strain resource library, and screening a functional single strain; b, purifying and compounding the single strain to form a compound flora; the core of the method is in vitro pre-coupling, exogenous targeting bacteria and host original flora are subjected to symbiotic adaptation in advance in a controlled environment to form a stable micro-ecological complex, the micro-ecological complex can be rapidly colonized and establish advantages after being applied to soil, the industrial pain point that a traditional microbial agent is living but not long and long but not effective is solved, and the selected strains are scientific in combination and easy to control. The bacillus subtilis and the brevibacillus laterosporus are used for resisting diseases and promoting growth, the bacillus mucilaginosus and the pseudomonas fluorescens are used for activating nutrients and removing heavy metal toxicity, and the arbuscular mycorrhizal fungi are used for expanding the root system absorption area.

Description

Technical Field

[0001] This invention relates to the fields of agricultural microbial technology and cultivation of Chinese medicinal herbs, specifically to a method for preparing functional microbial communities by coupling targeted microorganisms with the original host microbiota and its application in ginseng. Background Technology

[0002] As a rare Chinese medicinal herb, the core of ginseng's medicinal value lies in the content of secondary metabolites such as ginsenosides. Modern intensive cultivation faces severe challenges such as continuous cropping obstacles, soil microecological degradation, and dependence on chemical fertilizers and pesticides, which directly lead to the inhibition of ginsenoside synthesis and a significant decrease in medicinal properties. At the same time, heavy metals such as cadmium, lead, and arsenic in the soil accumulate through the root system, posing a safety risk to the medicinal materials and seriously restricting the sustainable development of the ginseng industry.

[0003] In existing technologies, the application of microbial fertilizers is a common means of improving soil health. However, traditional microbial agents are mostly mechanical mixtures of single-function bacteria or limited species. After these exogenous microorganisms are introduced into the complex field soil environment, they face problems such as difficulty in colonization, low survival rate, and unstable function. The fundamental reason is that they fail to form ecological synergy with the original complex "host original microbial community" in the crop rhizosphere. This "niche isolation" makes it difficult for the beneficial functions to be exerted in a sustained and efficient manner, and it is difficult to systematically solve the quality and safety bottlenecks in ginseng cultivation.

[0004] Therefore, developing a method and products for constructing functional microbial communities that can integrate with the host environment in advance and achieve niche synergy is of great significance for improving the quality and safety of ginseng. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides a method for formulating functional microbial communities through a targeted coupling of microorganisms with the original host microbial community, and its application in ginseng. This method solves the problem that traditional microbial agents are mostly mechanical mixtures of single-function bacteria or limited species. When these exogenous microorganisms are introduced into complex field soil environments, they face problems such as difficulty in colonization, low survival rate, and unstable function. The root cause is that they fail to form ecological synergy with the original and complex "original host microbial community" in the crop rhizosphere. This "niche isolation" leads to the technical problem that beneficial functions cannot be sustained and effectively exerted.

[0006] To achieve the above objectives, the present invention provides the following technical solution: a method for preparing functional microbial communities by coupling targeted microorganisms with the original host microbiota, comprising the following steps: a. Construct a targeted microbial strain resource bank and screen for functional single strains; b. Purify and compound the single strain to form a compound bacterial community; c. Couple and symbiotically culture the compound microbial community with the original host microbial community extracted from the rhizosphere soil of healthy ginseng; d. The coupled cultured microbial community is mixed with a nutrient carrier to prepare a microbial agent or water-soluble fertilizer for ginseng cultivation.

[0007] Preferably, 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.

[0008] Preferably, the functional single strain includes Bacillus subtilis, Bacillus mucilaginosus, Bacillus brevis, Pseudomonas fluorescens, and arbuscular mycorrhizal fungi.

[0009] Preferably, the conditions for the coupled symbiotic culture in step c include: a culture temperature of 25-30℃, a pH value of 6.5-7.5, aeration culture, and a culture time of 48-72 hours.

[0010] Preferably, the concentration of each microorganism in the functional microbial community meets the following requirements: Bacillus subtilis ≥5×10 8 CFU / ml; Bacillus mucilaginosus ≥3×10 8 CFU / ml; Lateral spores ≥2×10 8 CFU / ml; Fluorescent Pseudomonas ≥1×10 8 CFU / ml; Arbuscular mycorrhizal fungi ≥50 spores / gram; Host original microbiota ≥1×10 8 CFU / ml.

[0011] Preferably, the nutrient carrier in step d includes potassium humate, seaweed extract, amino acid chelated trace elements, potassium dihydrogen phosphate, and sucrose or molasses.

[0012] Preferably, in each ton of finished water-soluble fertilizer, the nutrient carrier 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.

[0013] Preferably, the amino acid chelated trace elements include amino acid chelates of at least three elements selected from iron, manganese, zinc, copper, boron, and molybdenum.

[0014] A method for coupling targeted microorganisms with the host's original microbial community to formulate functional microbial communities for use in ginseng, wherein the application is used to increase the content of ginsenosides in ginseng and / or reduce the residual amounts of heavy metals cadmium, lead, and arsenic.

[0015] Preferably, the application increases the ginsenoside content in ginseng by more than 10% and / or reduces the residue of heavy metals cadmium, lead, and arsenic by more than 30%.

[0016] Beneficial effects This invention provides a method for formulating functional microbial communities through coupling targeted microorganisms with the host's original microbial community, and its application in ginseng. The core of this invention lies in "in vitro pre-coupling," which allows exogenous targeted bacteria to pre-adapt to the host's original microbial community under controlled conditions, forming a stable "micro-ecological consortium." After being applied to the soil, it can rapidly colonize and establish dominance, solving the industry pain point of traditional microbial agents being "active but not growing, growing but ineffective." The selected strain combination is scientific: Bacillus subtilis and Bacillus laterosporus promote disease resistance and growth, Bacillus mucilage and Pseudomonas fluorescens activate nutrients and detoxify heavy metals, and arbuscular mycorrhizal fungi expand the root absorption area. After coupling with the host microbial community, they constitute a... A powerful and self-stable rhizosphere microecological engine, by improving the rhizosphere microecology and activating the plant's secondary metabolic pathways, can significantly increase the content of ginsenosides by more than 10%, fundamentally guaranteeing and enhancing the medicinal value of ginseng. The functional microbial community, through various mechanisms such as microbial adsorption, extracellular precipitation, ion competition, and valence state transformation, significantly reduces the bioavailability and accumulation of heavy metals cadmium, lead, and arsenic by more than 30%, providing key technical support for the production of safe and high-quality Chinese medicinal materials. The specific nutrient carrier is not only the "food" for microorganisms but also the "tonic" for plants, realizing the comprehensive effect of "promoting fertilizer with microorganisms, nourishing microorganisms with fertilizer, and synergistic effect of microorganisms and fertilizers", significantly improving fertilizer utilization. Detailed Implementation

[0017] The present invention will be further described in detail below through specific embodiments. These embodiments are for illustrative purposes only and are not intended to limit the scope of the invention.

[0018] Example 1: Formulation of functional microbial communities and preparation of water-soluble fertilizer Strains were prepared by selecting Bacillus subtilis, Bacillus mucilaginosus, Bacillus breviculatus, and Pseudomonas fluorescens, as well as commercially available arbuscular mycorrhizal fungi inoculants, and conducting high-density liquid (fungi are solid) fermentation respectively. Host microbiota extraction: Rhizosphere soil was collected from the Changbai Mountain healthy ginseng planting base, and a mixed microbial suspension was obtained by gradient centrifugation and membrane filtration. Coupled culture: The compound bacterial solution obtained in step 1 (volume ratio 2:1:1:1:0.5) was mixed with the host bacterial suspension in step 2 at a volume ratio of 3:1 and placed in a 30L fermenter. The mixture was then cultured for 60 hours at 28℃, pH 7.0, aeration rate of 1.0 vvm, and stirring at 200 rpm. Preparation of finished product: The high-concentration bacterial solution after coupling culture was mixed with the nutrient carrier shown in Table 1 below to prepare 1 ton of finished water-soluble fertilizer, and the final microbial indicators were tested. Table 1: Composition and Function of Nutrient Carrier per Ton of Water-Soluble Fertilizer Components Dosage (kg) Main functions Potassium humate 50 Stimulates microbial activity, improves soil structure, and promotes root development. seaweed extract 30 Provides natural plant hormones and trace elements to enhance stress resistance. Amino acid chelated trace elements (Fe, Zn, Mn, B) 10 Directly supplement plants with easily absorbed micronutrients to prevent nutrient deficiencies. Potassium dihydrogen phosphate 20 Provides essential elements such as phosphorus and potassium, promoting energy metabolism and substance transport. Molasses 15 Provide microorganisms with readily available carbon and energy sources. Table 2: Detection Results of Microbiological Indicators in Finished Water-Soluble Fertilizers Microbial species Target concentration Measured concentration Bacillus subtilis <![CDATA[≥ 5×10 8 CFU / ml]]> <![CDATA[7.2×10 8 CFU / ml]]> Bacillus mucilaginosus <![CDATA[≥ 3×10 8 CFU / ml]]> <![CDATA[4.5×10 8 CFU / ml]]> Lateral spores of Bacillus brevis <![CDATA[≥ 2×10 8 CFU / ml]]> <![CDATA[3.1×10 8 CFU / ml]]> Fluorescent Pseudomonas <![CDATA[≥ 1×10 8 CFU / ml]]> <![CDATA[1.8×10 8 CFU / ml]]> Arbuscular mycorrhizal fungi ≥ 50 spores / gram 95 spores / gram Host's original microbiota (total bacterial count) <![CDATA[≥ 1×10 8 CFU / ml]]> <![CDATA[2.5×10 8 CFU / ml]]> Example 2: Application and Effect Verification in Ginseng Cultivation To verify the actual effect of the product of this invention, a field trial lasting one growth cycle was conducted at a ginseng planting base in Fusong County, Jilin Province.

[0019] Experimental design: Two ginseng plots with similar soil background values ​​were selected as the experimental group and the control group, with three replicates in each group.

[0020] Solution: Experimental group: The water-soluble fertilizer prepared in Example 1 was used to perform root irrigation with a 500-fold diluted solution once each during the leaf expansion, flowering and fruiting stages of ginseng, with 500L applied per acre each time.

[0021] Control group: The same amount of water was applied at the same time, and other field management measures were exactly the same as those in the experimental group.

[0022] Sampling and testing: Representative ginseng samples were collected during the harvest period and sent to a qualified third-party testing institution to test the total amount of ginsenosides and the content of heavy metals. The results are shown in Table 3 below.

[0023] Table 3: Effects of the water-soluble fertilizer of this invention on the quality and safety of ginseng detection indicators control group experimental group range of change Total ginsenosides (mg / g) 3.42±0.15 3.92±0.13 +14.6% Cadmium (Cd) residue (mg / kg) 0.248±0.021 0.157±0.015 -36.7% Lead (Pb) residue (mg / kg) 0.88±0.07 0.56±0.05 -36.4% Arsenic (As) residue (mg / kg) 0.38±0.03 0.24±0.02 -36.8% The experimental results fully demonstrate that the application of the targeted biological bacteria water-soluble fertilizer prepared by this invention can significantly increase the content of the core medicinal components of ginseng and simultaneously reduce the residue of various harmful heavy metals, achieving the dual goals of "quality improvement" and "safety protection", with excellent results.

[0024] It should be noted that in this paper, relational terms such as first and second are used only to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any such actual relationship or order between these entities or operations.

Claims

1. A method for preparing a functional bio-bacterial community by targeting the coupling mode of biological bacteria with host original bacterial community, characterized in that, The method comprises the following steps: a. Constructing a target biological bacterial resource library, and screening functional single strains; b. Purifying and compounding the single strains to form a compounded bacterial population; c. Coupling and symbiotic culturing the compounded bacterial population with a host original bacterial population extracted from the rhizosphere soil of healthy ginseng; d. Mixing the compounded bacterial population after coupling and symbiotic culturing with a nutrient carrier to prepare a microbial inoculum or a water-soluble fertilizer for ginseng cultivation.

2. The method of claim 1, wherein the method is characterized in that, The functional single strains in step a. comprise at least three of Bacillus subtilis, Bacillus mucilaginosus, Brevibacillus laterosporus, Pseudomonas fluorescens and arbuscular mycorrhizal fungi.

3. The method of claim 2, wherein the method is characterized by, The functional single strains comprise Bacillus subtilis, Bacillus mucilaginosus, Brevibacillus laterosporus, Pseudomonas fluorescens and arbuscular mycorrhizal fungi.

4. The method of claim 3, wherein the method is characterized by, The coupling and symbiotic culturing conditions in step c. comprise a culture temperature of 25-30℃, a pH value of 6.5-7.5, air culture, and a culture time of 48-72 hours.

5. The method of claim 4, wherein the method is characterized by, In the functional biological bacterial population, the concentration of each microorganism meets the following requirements: Bacillus subtilis ≥ 5 x 10 8 CFU / ml; Bacillus mucilaginosus > 3 x 10 8 CFU / ml; Brevibacillus laterosporus ≥ 2 x 10 8 CFU / ml; Pseudomonas fluorescens > 1 x 10 8 CFU / ml; Arbuscular mycorrhizal fungi ≥ 50 spores / gram; Host original flora > 1 x 10 8 CFU / ml.

6. The method of claim 1, wherein the method is characterized by, The nutrient carrier in step d. comprises humic acid fulvic acid potassium, seaweed extract, amino acid chelated trace elements, potassium dihydrogen phosphate and sucrose or molasses.

7. The method of claim 6, wherein the method is characterized by, In each ton of finished water-soluble fertilizer, the composition of the nutrient carrier comprises: Humic acid fulvic acid potassium 40-60 kg; Seaweed extract 20-40 kg; Amino acid chelated trace elements 5-15 kg; Potassium dihydrogen phosphate 15-25 kg; Sucrose or molasses 10-20 kg.

8. The method of claim 7, wherein the method is characterized by, The amino acid chelated trace elements comprise amino acid chelates of at least three of iron, manganese, zinc, copper, boron and molybdenum.

9. The use of a functional bio-bacterial population on ginseng according to any one of claims 1-8, wherein the bio-bacterial population is coupled to the original bacterial population of the host. The application is used for increasing the content of ginsenosides in ginseng medicinal materials and / or reducing the residual amount of heavy metals cadmium, lead and arsenic.

10. The use of a functional bio-bacterial population prepared by the coupling of a target bio-bacterial population and a host original bacterial population on ginseng according to claim 9, characterized in that, The application increases the content of ginsenosides in ginseng medicinal materials by more than 10% and / or reduces the residual amount of heavy metals cadmium, lead and arsenic by more than 30%.