Polygonum cuspidatum compound microbial fertilizer and application thereof

Abstract

The invention relates to the field of biology, in particular to a polygonum cuspidatum compound microbial fertilizer and application thereof, and a compound microbial agent is a mixed flora of bacillus subtilis, bacillus licheniformis and bacillus amyloliquefaciens. The polygonum cuspidatum compound microbial fertilizer disclosed by the invention has the following beneficial effects: (1) the organic fertilizer disclosed by the invention is fermented by virtue of synergistic fermentation of three microorganisms, so that the fermentation is more sufficient, and the content of resveratrol is relatively high; (2) the organic fertilizer contains three kinds of probiotics, so that diseases and insect pests can be obviously inhibited, and actual planting of strawberries also proves the effect; and (3) the organic fertilizer provided by the invention can reduce diseases and insect pests in strawberry planting and reduce the pesticide use amount of strawberries, and is expected to eliminate the prejudice of excessive use of pesticides for strawberry planting in consumers.

Description

Technical Field

[0001] The present invention relates to the field of biology, and more particularly, to a polygonum cuspidatum composite microbial fertilizer and its application. Background Art

[0002] The rhizomes of polygonum cuspidatum are rich in active ingredients such as resveratrol, polydatin, and emodin, and have pharmacological effects such as anti-tumor and anti-inflammatory. However, the long-term reliance on chemical fertilizers in polygonum cuspidatum cultivation has led to soil compaction, nutrient loss, and frequent occurrence of pests and diseases (such as root rot and aphids). Most existing biological fertilizers use single-strain fermentation, with single functions and low extraction rates of active ingredients. For example, although the prior art CN112174744A uses Chinese medicine residues to prepare fertilizers, it does not involve the technology of co-fermentation of compound bacteria; the prior art CN114195593A proposes an organic-inorganic compound fertilizer, but does not clarify the synergistic mechanism of active ingredient enhancement. Therefore, it is of great application value to develop a composite microbial fertilizer that can efficiently utilize polygonum cuspidatum resources and has the functions of inhibiting bacteria and increasing production. Summary of the Invention

[0003] To solve the above problems, the present invention first provides a polygonum cuspidatum composite microbial fertilizer, which comprises polygonum cuspidatum rhizomes, medicinal residues, and a compound bacterial agent, and the compound bacterial agent is a mixed bacterial population of bacillus subtilis, bacillus licheniformis, and bacillus amyloliquefaciens.

[0004] Preferably, the weight ratio of the polygonum cuspidatum rhizomes to the medicinal residues is 1:1 - 3.

[0005] Preferably, its preparation method includes mixing the raw materials and then stacking and fermenting for 20 - 30 days at a fermentation temperature of 25 - 45°C.

[0006] Preferably, the addition amount of the compound bacterial agent in the polygonum cuspidatum composite microbial fertilizer is 0.5 - 2% of the total weight of the raw materials.

[0007] The present invention also provides the application of the above-mentioned polygonum cuspidatum composite microbial fertilizer, and the application is for crop cultivation or pest and disease control.

[0008] Preferably, the crops include fruits.

[0009] Preferably, the fruits include strawberries.

[0010] Preferably, the disease is strawberry root rot.

[0011] Preferably, the pest is strawberry aphid.

[0012] Compared with the prior art, the present invention has at least the following beneficial effects:

[0013] (1) The organic fertilizer of the present invention uses three microorganisms for co-fermentation, with more sufficient fermentation and higher contents of resveratrol and polydatin;

[0014] (2) The organic fertilizer of the present invention contains 3 probiotics, which can significantly inhibit diseases and pests, and the actual cultivation of strawberries also proves this point;

[0015] (3) The organic fertilizer of the present invention can reduce diseases and pests in strawberry cultivation, reduce the amount of pesticides used for strawberries, and is expected to eliminate consumers' prejudice against excessive use of pesticides in strawberry cultivation. Detailed implementation manners

[0016] To make the technical problems, technical solutions and advantages to be solved by the present invention clearer, the following will be described in detail with reference to specific embodiments.

[0017] Bacillus subtilis, Bacillus licheniformis, and Bacillus amyloliquefaciens were all purchased from the China Center for Agricultural Culture Collection (ACCC).

[0018] Example 1: Preparation of polygonum cuspidatum compound microbial fertilizer

[0019] Fermentation raw materials: polygonum cuspidatum residue and polygonum cuspidatum rhizome;

[0020] Compound microbial strain ratio: Bacillus subtilis, Bacillus licheniformis, and Bacillus amyloliquefaciens, with a weight ratio of 1:1:1;

[0021] 1 Raw material pretreatment:

[0022] 1.1 Rhizome treatment

[0023] Crushing: Crush the dried polygonum cuspidatum rhizome to 40-100 mesh.

[0024] Drying: Vacuum dry at 50-60 °C until the moisture content ≤ 8%.

[0025] 1.2 Polygonum cuspidatum residue treatment

[0026] Mix the polygonum cuspidatum residue with malt in a mass ratio of 1:10, stir and heat at 50 °C for 1.5 hours, and then perform ultrasonic treatment (260 W, 15 minutes).

[0027] Drying: Dry at 75 °C for 2 hours, dry at 100 °C for 2 hours, and cool for later use.

[0028] Take the pretreated polygonum cuspidatum rhizome raw material and the pretreated polygonum cuspidatum residue raw material and mix them in a ratio of 1:1 to prepare the fermentation substrate;

[0029] Inoculate the compound bacteria according to 1% of the weight of the fermentation substrate, and stir evenly;

[0030] Fermentation conditions: 35°C, pH 7.0, water content 50%, ferment for 21 days (turn the pile 6 times) to obtain the fermentation product.

[0031] Example 2

[0032] Fermentation raw materials: Polygonum cuspidatum residue and Polygonum cuspidatum rhizome;

[0033] Compound microbial strain ratio: Bacillus subtilis, Bacillus licheniformis, Bacillus amyloliquefaciens, with a weight ratio of 1:1:1;

[0034] 1 Raw material pretreatment:

[0035] 1.1 Rhizome treatment

[0036] Crushing: Crush the dried Polygonum cuspidatum rhizome to 40 - 100 mesh.

[0037] Drying: Vacuum dry at 50 - 60°C until the moisture content ≤ 8%.

[0038] 1.2 Polygonum cuspidatum residue treatment

[0039] Mix the Polygonum cuspidatum residue with malt in a mass ratio of 1:10, stir and heat at 50°C for 1.5 hours, and then perform ultrasonic treatment (260W, 15 minutes).

[0040] Drying: Dry at 75°C for 2 hours, then dry at 100°C for 2 hours, and cool for later use.

[0041] Take the pretreated raw materials of Polygonum cuspidatum rhizome and Polygonum cuspidatum residue and mix them in a ratio of 1:1 to prepare the fermentation substrate;

[0042] Inoculate the compound bacteria according to 0.5% of the weight of the fermentation substrate, and stir evenly;

[0043] Fermentation conditions: 30°C, pH 7.0, water content 50%, ferment for 21 days (turn the pile 6 times) to obtain the fermentation product.

[0044] Example 3

[0045] Fermentation raw materials: Polygonum cuspidatum residue and Polygonum cuspidatum rhizome;

[0046] Compound microbial strain ratio: Bacillus subtilis, Bacillus licheniformis, Bacillus amyloliquefaciens, with a weight ratio of 1:1:1;

[0047] 1 Raw material pretreatment:

[0048] 1.1 Rhizome treatment

[0049] Crushing: Crush the dried Polygonum cuspidatum rhizome to 40 - 100 mesh.

[0050] Drying: Vacuum dry at 50 - 60°C until the moisture content ≤ 8%.

[0051] 1.2 Polygonum cuspidatum residue treatment

[0052] Mix the Polygonum cuspidatum residue and malt in a mass ratio of 1:10, stir and heat at 50 °C for 1.5 hours, and then perform ultrasonic treatment (260 W, 15 minutes).

[0053] Drying: Dry at 75 °C for 2 hours, dry at 100 °C for 2 hours, and reserve after cooling.

[0054] Take the pre-treated raw material of Polygonum cuspidatum rhizome and the pre-treated raw material of Polygonum cuspidatum residue and mix them in a ratio of 1:1 to prepare the fermentation substrate;

[0055] Inoculate the composite bacteria according to 1.5% of the weight of the fermentation substrate, and stir evenly;

[0056] Fermentation conditions: 40 °C, pH 7.0, water content 50%, ferment for 21 days (turn the pile 6 times) to obtain the fermentation product.

[0057] Comparative Example 1

[0058] Compared with Example 1, the difference is that Bacillus subtilis + Bacillus licheniformis (1:1) is used to replace the composite bacteria for fermentation, and parameters such as the total inoculation amount of the bacteria are the same as those in Example 1.

[0059] Comparative Example 2

[0060] Compared with Example 1, the difference is that Bacillus amyloliquefaciens + Bacillus licheniformis (1:1) is used to replace the composite bacteria for fermentation, and parameters such as the total inoculation amount of the bacteria are the same as those in Example 1.

[0061] Comparative Example 3

[0062] Compared with Example 1, the difference is that Bacillus subtilis + Bacillus amyloliquefaciens (1:1) is used to replace the composite bacteria for fermentation, and parameters such as the total inoculation amount of the bacteria are the same as those in Example 1.

[0063] Comparative Example 4

[0064] Compared with Example 1, the difference is that single Bacillus subtilis is used to replace the composite bacteria for fermentation, and parameters such as the total inoculation amount of the bacteria are the same as those in Example 1.

[0065] Comparative Example 5

[0066] Compared with Example 1, the difference is that single Bacillus licheniformis is used to replace the composite bacteria for fermentation, and parameters such as the total inoculation amount of the bacteria are the same as those in Example 1.

[0067] Comparative Example 6

[0068] Compared with Example 1, the difference is that single Bacillus amyloliquefaciens is used to replace the composite bacteria for fermentation, and parameters such as the total inoculation amount of the bacteria are the same as those in Example 1.

[0069] Control Group 1

[0070] (1) Raw material pretreatment:

[0071] 1.1 Rhizome treatment

[0072] Crushing: Crush the dried rhizomes of Polygonum cuspidatum to 40 - 100 mesh.

[0073] Drying: Vacuum dry at 50 - 60 °C until the moisture content ≤ 8%.

[0074] 1.2 Treatment of Polygonum cuspidatum residue

[0075] Mix the Polygonum cuspidatum residue with malt in a mass ratio of 1:10, stir and heat at 50 °C for 1.5 hours, and then perform ultrasonic treatment (260 W, 15 minutes).

[0076] Take the pretreated raw materials of Polygonum cuspidatum rhizomes and the pretreated raw materials of Polygonum cuspidatum residue and mix them in a ratio of 1:1 to prepare the substrate for enzymatic hydrolysis;

[0077] (2) Resveratrol extraction

[0078] (2.1) Enzymatic hydrolysis and conversion: Add water to the substrate for enzymatic hydrolysis at a solid - liquid ratio of 1:4, adjust the pH to 4.5 - 5.0, add 3% cellulase (50,000 U / g) + 0.2% pectinase (50,000 U / g) based on the weight of the substrate for enzymatic hydrolysis, and perform enzymatic hydrolysis at 45 - 55 °C for 2 - 3 hours. After enzymatic hydrolysis, raise the temperature to 85 - 100 °C to inactivate the enzyme, and filter to obtain the enzymatic hydrolysate.

[0079] (2.2) Solvent extraction: Extract twice by refluxing with 60% ethanol (volume ratio) at a solid - liquid ratio of 1:8 for 2 hours each time. The extract is concentrated by vacuum rotary evaporation and dried to obtain the crude extract.

[0080] Example 4: Determination of the content of active ingredients

[0081] Method: Use the HPLC method to determine the resveratrol content of Examples 1 - 3, Comparative Examples 1 - 6, and Control Group 1, and then calculate the relative yield of resveratrol. The calculation of the relative yield of resveratrol is based on Control Group 1, and the results are shown in Table 1;

[0082] Table 1. Relative yield of resveratrol

[0083]

[0084]

[0085] Note: Different letters a, b, c, d in the table indicate significant differences from other groups (P < 0.05);

[0086] The results in Table 1 show that the yield of resveratrol in the three-strain complex fermentation group was significantly higher than that in Control Group 1 (p < 0.01), indicating that fermentation is conducive to the extraction of resveratrol.

[0087] Example 5: Antibacterial Test

[0088] Method: The final products of Examples 1 - 3, Comparative Examples 1 - 6, and Control Group 1 were formulated into a concentration of 10 mg / mL, and the inhibition rates of the fermentation end products of Examples 1 - 3, Comparative Examples 1 - 6, and Control Group 1 against three common strawberry pathogens (Botrytis cinerea, Fusarium sp., and Rhizoctonia solani Kuhn) were determined using the inhibition zone method. The results are shown in Table 2;

[0089] Table 2. Inhibition Rates of Fermentation Products against Three Common Strawberry Pathogens (%)

[0090] Group Botrytis cinerea Fusarium Rhizoctonia solani Example 1 <![CDATA[99.14+0.04 a > <![CDATA[99.91+0.02 a > <![CDATA[98.74+0.31 a > Example 2 <![CDATA[98.11+0.14 a > <![CDATA[93.21+0.04 a > <![CDATA[88.26+0.22 a > Example 3 <![CDATA[93.02+0.07 a > <![CDATA[94.11+0.06 a > <![CDATA[87.45+0.56 a > Comparative Example 1 <![CDATA[82.01+0.24 b > <![CDATA[85.61+0.09 b > <![CDATA[78.36+0.21 b > Comparative Example 2 <![CDATA[81.32+0.30 b > <![CDATA[82.14+0.16 b > <![CDATA[80.02+0.34 b > Comparative Example 3 <![CDATA[85.17+0.14 b > <![CDATA[83.04+0.21 b > <![CDATA[81.68+0.50 b <!-- 4 -->]]> Comparative Example 4 <![CDATA[78.64+0.19 b > <![CDATA[72.52+0.37 c > <![CDATA[74.47+0.09 c > Comparative Example 5 <![CDATA[83.33+0.08 b > <![CDATA[69.99+0.12 c > <![CDATA[76.03+0.11 c > Comparative Example 6 <![CDATA[79.12+0.16 b > <![CDATA[71.90+0.08 c > <![CDATA[74.12+0.24 c > Control Group 1 <![CDATA[66.12+0.31 c > <![CDATA[55.21+0.14 d > <![CDATA[65.42+0.41 d >

[0091] Note: Different letters a, b, c, d in the table indicate significant differences from other groups (P < 0.05); the results in Table 2 show that the inhibition rates of the fermentation products of Examples 1 - 3 against strawberry pathogens were significantly higher than those in Control Group 1 (p < 0.01).

[0092] Example 6: Planting Experiment

[0093] · Experimental Design: Strawberry seedlings were grouped and treated with the fermentation fertilizers of Examples 1 - 3, Comparative Examples 1 - 6, and Control Group 1, as well as a commercially available ordinary fertilizer (Control Group 2);

[0094] The incidence of root rot in strawberry planting (the pathogens include 20 species such as Rhizoctonia solani, Fusarium sp., and Pythium sp.), the incidence of fruit pests, and the relative yield (yield increase) were statistically analyzed as shown in Table 3; the relative yield was calculated based on the yield of Control Group 2.

[0095] Table 3. Strawberry Planting Conditions

[0096] Group Incidence of root rot (%) Incidence of pest damage (%) Relative yield Example 1 <![CDATA[10.3+1.06 a > <![CDATA[8.7+0.95 a > <![CDATA[1.22+0.02 a > Example 2 <![CDATA[11.4+0.76 a > <![CDATA[9.1+0.54 a > <![CDATA[1.15 + 0.10 a > Example 3 <![CDATA[11.7 + 1.91 a > <![CDATA[9.3+2.31 a > <![CDATA[1.13+0.04 a > Comparative Example 1 <![CDATA[16.6+2.35 b > <![CDATA[15.1 + 1.78 b > <![CDATA[1.08+0.06 b > Comparative Example 2 <![CDATA[18.7+0.66 b > <![CDATA[16.2+2.11 b > <![CDATA[1.07+0.05 b > Comparative Example 3 <![CDATA[16.1+2.16 b > <![CDATA[15.4+0.86 b > <![CDATA[1.08+0.02 b > Comparative Example 4 <![CDATA[21.3+0.98 c > <![CDATA[21.5+0.76 c > <![CDATA[1.06+0.04 b > Comparative Example 5 <![CDATA[23.1+2.11 c > <![CDATA[24.6 + 0.71 c > <![CDATA[1.05+0.07 b > Comparative Example 6 <![CDATA[21.9+3.10 c > <![CDATA[22.1+0.84 c > <![CDATA[1.06+0.02 b > Control Group 1 <![CDATA[27.3+2.14 d > <![CDATA[28.4+1.11 d > <![CDATA[1.01 + 0.03 c > Control Group 2 <![CDATA[37.6+3.12 d > <![CDATA[32.1+0.11 d > <![CDATA[1.00+0.01 c >

[0097] Note: Different letters a, b, c, and d in the table indicate significant differences from other groups (P < 0.05); the results are shown in Table 3. Among them, the polygonum cuspidatum composite microbial fertilizer in Examples 1-3 can reduce the diseases and pests in strawberry cultivation, reduce the amount of pesticides used for strawberries, and is expected to eliminate the prejudice of consumers about excessive pesticide use in strawberry cultivation. The results of Example 1 and Control Group 1 show that the polygonum cuspidatum composite microbial fertilizer of the present application also contains other beneficial components in addition to resveratrol, which can help reduce the diseases and pests in strawberry cultivation while increasing the yield.

[0098] The above are the preferred embodiments of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and modifications can be made, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. A polygonum cuspidatum compound microbial fertilizer, characterized in that, The polygonum cuspidatum composite microbial fertilizer contains polygonum cuspidatum rhizomes, medicinal residues and a composite microbial agent, and the composite microbial agent is a mixed flora of bacillus subtilis, bacillus licheniformis and bacillus amyloliquefaciens.

2. The fertilizer according to claim 1, characterized in that, The weight ratio of the polygonum cuspidatum rhizomes to the medicinal residues is 1:1-3.

3. The fertilizer according to claim 1, characterized in that, Its preparation method includes mixing the raw materials and then piling them up for fermentation for 20-30 days, and the fermentation temperature is 25-45°C.

4. The fertilizer according to claim 1, characterized in that, The addition amount of the composite microbial agent in the polygonum cuspidatum composite microbial fertilizer is 0.5-2% of the total weight of the raw materials.

5. Use of the polygonum cuspidatum composite microbial fertilizer according to any one of claims 1-4, characterized in that, The application is for crop planting or disease and pest control.

6. The application according to claim 5, characterized in that The crops include fruits.

7. The application according to claim 6, wherein The fruits include strawberries.

8. The application according to claim 7, characterized in that The disease is strawberry root rot.

9. The application according to claim 5, characterized in that, The pest is strawberry aphid.

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