Microbial substrate for strawberry planting and preparation method and application thereof

Abstract

The application discloses a microbial substrate for strawberry planting and a preparation method and application thereof, and the substrate is made of peat, perlite, bacteria-loaded biochar and fungi, and the volume ratio of the peat, the perlite and the bacteria-loaded biochar is (45-75):25:(10-30); the bacteria-loaded biochar is prepared by loading biochar with a biocontrol composite bacteria; and the biocontrol composite bacteria is composed of Rhodopseudomonas palustris, Bacillus subtilis and Bacillus licheniformis. The application changes the traditional single bacterial group into a mixed bacterial group, and uses biochar as a microbial loading material, so that the mixed bacterial group can better improve the prevention and treatment effect, and the biochar can provide a carbon source for the microorganisms and promote the better reproduction and growth of the microorganisms.

Description

TECHNICAL FIELD

[0001] The present application particularly relates to a microbial substrate for strawberry planting and a preparation method and application thereof. BACKGROUND

[0002] The information disclosed in this Background section is only for the purpose of increasing an understanding of the general context of the present application and does not necessarily constitute an acknowledgement or a warranty as to any prior art.

[0003] Typical diseases and pests of strawberry, such as powdery mildew, root rot, anthracnose, and verticillium wilt, can harm different parts of strawberry, such as roots, leaves, flowers, and fruits, causing a large number of rotten fruits, and even the whole strawberry plant wilting and even death. In the traditional process of strawberry planting, the combination of planting resistant varieties and chemical control is mainly used for the prevention and control of strawberry diseases and pests. Chemical fungicides become the main choice for most strawberry farmers to prevent and control strawberry diseases and pests due to their high efficiency and convenience. The fungicides widely used in production at present include azoxystrobin, prochloraz, pyraclostrobin, and difenoconazole. However, the control effect of chemical fungicides is affected by many factors, and the control effect is unstable. Long-term use of chemical fungicides can also cause pathogenic fungi to develop drug resistance, and the use of chemical fungicides is non-selective, which can easily affect non-target strains in the soil microbial community, destroy soil microbial diversity and ecological safety, and cause fruit pesticide residues, which can also harm human health. With the enhancement of people's green environmental protection consciousness and the emphasis on food safety, chemical control measures that destroy the natural environment and harm human health are gradually eliminated. Therefore, green and environmentally friendly biological control is increasingly valued because it can avoid the above problems.

[0004] Biological control mainly introduces biocontrol strains into the soil, uses biocontrol strains and their metabolites to regulate the microbial colony environment in the soil, so as to inhibit the growth and reproduction of pathogenic microorganisms, promote the growth of crops, and prevent and control diseases. The mechanism of biological control includes antibiotic action, competition, fungal hyperparasitism, growth promotion, and induced resistance system. Biological control is environmentally friendly and has the advantages of green and sustainability. At present, some research progress has been made in the biological control of strawberry diseases caused by fungal pathogens. It has been found that some fungi, bacteria, and actinomycetes can be used to prevent and control strawberry diseases and pests, and also have certain promoting effects on the growth and development of strawberry plants. At present, the fungi commonly used for biological control of strawberry diseases and pests include Penicillium griseofulvum and Streptomyces, and the bacteria include Bacillus and Pseudomonas. However, compared with traditional chemical control, biological control is relatively slow. Microorganisms need suitable environment and nutrients to maintain growth and activity, so the biological control effect of single microbial inoculant application in the field is not ideal, and it is urgent to develop efficient broad-spectrum microbial inoculants or products for the prevention and control of strawberry diseases and pests.

[0005] The current problems of biological control of strawberry diseases and pests are: (1) single bacteria can only prevent one disease, and has poor effect on other diseases; (2) the prevention effect of single bacteria is slow and the time cycle is long. SUMMARY

[0006] In view of the deficiencies of the prior art, the present application provides a microbial substrate for strawberry planting and a preparation method and application thereof.

[0007] The technical scheme adopted by the present application is as follows:

[0008] In a first aspect of the present application, a microbial substrate for strawberry planting is provided, which is made of the following raw materials: peat, perlite, bacteria-loaded biochar and fungi, the volume ratio of the peat, perlite and bacteria-loaded biochar being (45-75):25:(10-30);

[0009] The bacteria-loaded biochar is prepared by loading biochar with biocontrol composite bacteria; the biocontrol composite bacteria are composed of Rhodopseudomonas palustris Rhodopseudomonas palustris , Bacillus subtilis Bacillus subtilis and Lysinibacillus sphaericus Lysinibacillus ;

[0010] The fungi are Talaromyces Talaromyces muroii and / or Penicillium Penicillium caperatum.

[0011] In one or some embodiments of the present application, the biochar is prepared by the following method: rice husk and / or charcoal and coal gangue powder are mixed by wet method at a set mass ratio, stirred uniformly, dried to obtain wooden biochar and / or rice husk biochar; the wooden biochar and / or rice husk biochar raw material is heated, heated to 600-800℃ and then naturally cooled, and nitrogen gas is introduced as a protective gas to isolate oxygen during the cooling process, and the biochar is obtained after cooling. Further, the raw material is preferably charcoal.

[0012] The addition of the biochar screened in the present application can improve the physical and chemical properties of the substrate, optimize the morphological indexes of strawberry plants, improve the photosynthetic physiological characteristics of leaves, and explore the effects of biochar on the growth and development and photosynthetic physiology of soilless-cultivated strawberries, thereby providing a scientific basis for the reasonable application of biochar in soilless cultivation of strawberries.

[0013] In one or some embodiments of the present application, the bacteria-loaded biochar is prepared by the following method: the biocontrol composite bacteria liquid is transferred to a container containing sterilized biochar, and then the mixture of the biocontrol composite bacteria liquid and the biochar is loaded on a rotary shaker for a set time to allow the biocontrol composite bacteria to be adsorbed on the surface and pores of the biochar; after loading, the mixture is left to stand, and then the supernatant is poured off, and the obtained biochar is placed in a drying oven for drying, thereby obtaining the bacteria-loaded biochar.

[0014] In one or some embodiments of the present application, the ratio of the number of viable bacteria of Rhodopseudomonas palustris, Bacillus subtilis and Bacillus licheniformis is (2-4):(2-4):1.

[0015] Preferably, the ratio of the number of viable bacteria of Rhodopseudomonas palustris, Bacillus subtilis and Bacillus licheniformis is 2:2:1.

[0016] In one or some embodiments of the present application, the amount of fungi used is 0.7-0.8 ml of fungal spore suspension per strawberry seedling, and the concentration of the spore suspension is 1x10 5 ~2x10 5 CFU / mL.

[0017] The experimental data show that the biochar substrate added with Talaromyces and / or Penicillium performs better in plant height, stem diameter and maximum leaf area than the control group without biochar and the domestic substrate, and the performance of the biochar composite is the best, which has great practical application value.

[0018] In the second aspect of the present application, a preparation method of the microbial substrate for strawberry planting is provided, which comprises the following steps:

[0019] (1) Preparation of biochar: rice husk and / or charcoal is mixed with coal gangue powder in a set mass ratio by wet mixing, fully stirred and uniformly mixed, dried, to obtain lignin biochar and / or rice husk biochar; the lignin biochar and / or rice husk biochar raw material is heated, heated to 600-800℃, then naturally cooled, and nitrogen gas is introduced as a protective gas to isolate oxygen during the cooling process, and the biochar is obtained after cooling;

[0020] (2) Preparation of biochar material loaded with bacteria: the biocontrol composite bacteria liquid is transferred to a container containing sterilized biochar, then the mixture of the biocontrol composite bacteria liquid and the biochar is loaded on a rotary shaker for a set time, so that the biocontrol composite bacteria are adsorbed on the surface and pores of the biochar; after loading, the mixture is left to stand, then the supernatant is poured off, and the obtained biochar is placed in a drying oven for drying, to obtain the required biochar loaded with bacteria;

[0021] (3) Compounding: a set amount of peat, perlite, biochar loaded with bacteria and fungi are mixed uniformly, to obtain the microbial substrate for strawberry planting.

[0022] In one or some embodiments of the present application, in step (1), the mass ratio of the rice husk and / or charcoal to the coal gangue powder is (2-4):(1-2). The addition of coal gangue can make full use of solid waste generated in the process of coal development and processing, and make it resourceful, and the coal gangue powder has good effects on improving soil structure, water retention and fertilizer increase, etc.

[0023] Preferably, the mass ratio of the rice husk and / or charcoal to the coal gangue powder is 2:1.

[0024] In one or some embodiments of the present application, in step (1), the holding temperature is 700℃.

[0025] In one or some embodiments of the present application, in step (2), the ratio of the biocontrol composite bacteria liquid to the biochar is 0.005 mL:(0.5-1.5) g, and the number of living bacteria in each mL of the bacteria liquid is 10 9 CFU / mL, and the ratio of the number of living bacteria of the Rhodopseudomonas palustris, Bacillus subtilis and Bacillus licheniformis is (2-4):(2-4):1. The number of living bacteria in the bacteria-loaded biochar is 1.7×10 8 ~1.8×10 8 CFU / g.

[0026] In one or some embodiments of the present application, in step (2), the loading time is 12-36 h.

[0027] Preferably, the loading time is 24 h.

[0028] In one or some embodiments of the present application, in step (3), the fungus is added in the form of a bacteria liquid, and the number of living bacteria in the bacteria liquid is 1×10 5 ~2×10 5 CFU / mL.

[0029] In the third aspect of the present application, the use of the microbial substrate in promoting the growth of strawberries and preventing and treating diseases and pests is provided.

[0030] In one or some embodiments of the present application, the microbial substrate is used in promoting the growth of strawberries and preventing and treating the strawberry root rot and the strawberry anthracnose.

[0031] In the fourth aspect of the present application, a cultivation method of a strawberry soilless culture substrate is provided, which comprises the following steps: bagging the microbial substrate for strawberry planting, making the substrate into a substrate strip, stacking the substrate strip side by side on a ridge after ridging, punching small holes on the side and bottom of the substrate strip for air permeability, opening the top of the substrate strip, and laying a drip tape on the substrate strip.

[0032] Compared with the related art known to the present inventors, one of the technical solutions of the present application has the following beneficial effects:

[0033] The present application changes the traditional single flora into a mixed flora, and uses biochar as a microbial loading material. The mixed flora can better improve the prevention and control effect, and the biochar can provide carbon source for the microorganisms to promote the better reproduction and growth of the microorganisms. The mixed biological flora is composed of Bacillus subtilis, Rhodopseudomonas palustris and Bacillus licheniformis. Each bacterium in the flora has its unique function and metabolic way, and the prevention and control effect on different strawberry diseases has certain difference. The construction of the mixed flora can make up for the deficiency of single bacteria in the prevention and control effect on a certain disease. Different strains of the mixed biological flora can produce synergistic effect, promote the generation of beneficial metabolites, and jointly inhibit the growth of harmful microorganisms. Different strains depend on each other and promote each other, so as to improve the prevention and control effect on strawberry diseases. In addition, the interaction of different strains also improves the adaptability to the environment, so that the survival, growth and reproduction ability is more stable, and it is easier to form a stable community in the strawberry planting soil (substrate) and improve the ecological environment. Therefore, compared with single bacteria, the mixed biological flora has more stable and excellent prevention and control effect on strawberry diseases, can prevent and control more types of strawberry diseases, and some biocontrol bacteria can also degrade or biotransform organic pollutants or heavy metals introduced into the strawberry planting soil (substrate) in the early stage due to pesticide application, so as to control the generation of harmful substances from the source, effectively improve the quality of strawberries, and have environmental, social and economic benefits in actual strawberry planting and production. The present application loads the mixed biological flora on the biochar. The biochar is a solid material rich in carbon and highly aromatic left after the pyrolysis combustion of biomass. The present application uses coal-based solid waste and agricultural waste such as straw, rice husk and charcoal as carbon-containing raw materials to prepare biochar. As a new type of soil conditioner, biochar has various application potentials and plays a significant role in improving soil properties and increasing microbial activity. It has a porous structure and high surface area, can provide shelter for microorganisms, and can also control the entry of harmful substances into strawberry fruits through adsorption of existing or potential harmful substances in the soil (substrate). The application of biochar into the strawberry planting soil (substrate) can play a role in carbon sequestration, increase the carbon content of the soil (substrate) and thus increase its fertility. In addition, biochar is rich in nutrients, which can provide carbon source, nitrogen source and other nutrients for microorganisms, thereby enhancing the survival ability and metabolic activity of functional bacteria, and thus greatly improving the yield of strawberries. BRIEF DESCRIPTION OF DRAWINGS

[0034] The accompanying drawings, which form a part of the present application, are used to provide a further understanding of the present application, and the illustrative embodiments of the present application and their description serve the purpose of explaining the present application. The present application shall not be limited by the accompanying drawings.

[0035] Figure 1 : Inhibition rate of Bacillus subtilis on Fusarium oxysporum.

[0036] Figure 2 : Inhibition rate of Bacillus subtilis on Colletotrichum gloeosporioides.

[0037] Figure 3 : Verticillium dahliae strains.

[0038] Figure 4 The inhibition rate of Rhodopseudomonas palustris against Verticillium dahliae.

[0039] Figure 5 The inhibition rate of Bacillus subtilis against Verticillium dahliae.

[0040] Figure 6 Inhibition rate of lysine-containing Bacillus against Verticillium dahliae.

[0041] Figure 7 The inhibition rates of Bacillus subtilis and Rhodopseudomonas palustris against Fusarium oxysporum.

[0042] Figure 8 The inhibition rates of Bacillus subtilis and Rhodopseudomonas palustris against Colletotrichum anthracis.

[0043] Figure 9 Results of the plate antagonism experiment.

[0044] Figure 10 The inhibitory effect of biocontrol bacteria on strawberry anthracnose.

[0045] Figure 11 The inhibitory effect of biocontrol bacteria on strawberry root rot.

[0046] Figure 12 Experiments on strawberry potted plants with different treatments.

[0047] Figure 13 Experiment on greenhouse strawberry cultivation. Detailed Implementation

[0048] It should be noted that the following detailed descriptions are exemplary and intended to provide further illustration of the invention. 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 invention pertains.

[0049] It should be noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the exemplary embodiments of the present invention. As used herein, the singular form is intended to include the plural form as well, unless the context clearly indicates otherwise. Furthermore, it should be understood that when the terms "comprising" and / or "including" are used in this specification, they indicate the presence of features, steps, operations, and / or combinations thereof.

[0050] 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.

[0051] Material sources:

[0052] Imported peat, domestic peat, and perlite manufacturers are all from Shandong Yide Import and Export Co., Ltd.

[0053] Bacillus subtilis Bacillus subtilis Rhodopseudomonas palustris Rhodopseudomonas palustris、 Bacillus licheniformis Lysinibacillus : Purchased from the laboratory of China University of Mining and Technology.

[0054] Talaromyces Talaromyces muroii : Talaromyces muroii, preserved in the China General Microbiological Culture Collection Center on August 24, 2022, located at No. 3, Beichen West Road, Chaoyang District, Beijing, with the preservation number CGMCC No. 40277 and the classification name Talaromyces muroii.

[0055] Penicillium Penicillium caperatum : Penicillium MR-16, preservation number GDMCC No: 62082, purchased from the Ecological Research Institute of Shandong Province.

[0056] Other materials or reagents can be obtained through commercial channels.

[0057] The entire experimental process is divided into indoor potting experiments and greenhouse strawberry planting experiments.

[0058] Indoor potting experiment:

[0059] 1. Strain selection

[0060] The purpose of this step is to verify that the antagonistic effect of mixed strains on pathogenic bacteria is better than that of single strains, providing data support for the next step of strain compounding and soil phase inhibition experiment.

[0061] This example will study the antagonistic effect of three functional microorganisms (Bacillus subtilis, Rhodopseudomonas palustris, and Bacillus licheniformis) on three pathogenic microorganisms (strawberry root rot - Fusarium oxysporum, strawberry anthracnose - Colletotrichum gloeosporioides, and strawberry yellow wilt - Verticillium dahliae), and calculate the inhibition rate. Further determine the compounding scheme of mixed bacteria, study the antagonistic effect of mixed bacteria and its inhibition rate. This example uses the plate confrontation method to screen biocontrol strains. After activating the biocontrol strains preserved in the laboratory on LB plates, they are inoculated into LB liquid medium for activation. The pathogenic fungus mycelium block is inoculated in the center of the PDA plate, and the activated biocontrol strains are inoculated around it, repeated three times, with normal growing pathogenic fungus plates as controls. Observe and record the size and growth rate of the colonies in a timely manner during the culture process, and measure the inhibition zone diameter and calculate the inhibition rate after 7 days (due to different growth periods, the culture time of Verticillium dahliae is 28 days).

[0062] Before improvement, the example carried out the single species of Bacillus subtilis respectively on Fusarium oxysporum, Colletotrichum gloeosporioides and L. digitata. The experimental antagonistic data are as follows:

[0063] At 6d, the inhibition rate of Bacillus subtilis on Fusarium oxysporum was 27.8%. As shown. Figure 1

[0064] At 6d, the inhibition rate of Bacillus subtilis on Colletotrichum gloeosporioides was 51.9%. As shown. Figure 2

[0065] At 6d, the inhibition rate of Rhodopseudomonas palustris on L. digitata was more than 90%, the inhibition rate of Bacillus subtilis on L. digitata was more than 85%, and the inhibition rate of Bacillus licheniformis on L. digitata was 30-40%, as shown in Figure 3 、 Figure 4 、 Figure 5 and Figure 6 .

[0066] After improvement, the example carried out the antagonistic experiment of Bacillus subtilis and Rhodopseudomonas palustris on Fusarium oxysporum, and the results are as follows:

[0067] At 6d, the inhibition rate of Bacillus subtilis and Rhodopseudomonas palustris on Fusarium oxysporum was 60%. As shown. Figure 7

[0068] Antagonistic experiment of Bacillus subtilis and Rhodopseudomonas palustris on Colletotrichum gloeosporioides:

[0069] At 6d, the inhibition rate of Bacillus subtilis and Rhodopseudomonas palustris on Colletotrichum gloeosporioides was 70%. As shown. Figure 8

[0070] The antagonistic data of plate experiment showed that when two kinds of mixed bacteria were added, the antagonistic effect of each functional bacteria on the pathogenic bacteria was enhanced compared with the original single strain.

[0071] 2. Soil phase inhibition experiment

[0072] The experimental steps mainly verify the inhibition effect of mixed strains on pathogenic bacteria after adding sterilized substrate.

[0073] The mixed substrate was weighed, the moisture content was adjusted to 10%, and it was placed in a sterilization pot and sterilized at 121℃ for 60min. After three treatments, it was ready for use. The sterilized medium was added to the enriched culture of the biocontrol strain, and the OD600 of the bacterial suspension was adjusted to 1.0. Then, the mixed bacterial solution was prepared according to different proportions. Among them, the unit of the mixed bacterial proportion was based on the adjustment of the OD600 of the bacterial suspension to about 1.0 after the single strain was cultured in liquid medium to the logarithmic growth phase, that is, the number of viable bacteria of the single strain (10​​​​9 After the number of viable cells of each strain in the mixed microbial flora was substantially consistent, the strains were mixed according to the volume ratio to prepare the mixed microbial flora, so that the number of viable cells of each strain in the mixed microbial flora remained in the corresponding proportion. After spores of the pathogenic bacteria were scraped, the spores were cultured in PDA liquid medium, and then the pathogenic bacteria spores were adjusted to a concentration of CFU = 1.0 x 10 6 1.0 x 10 7 (OD600 = 0.1).

[0074] The spore solution of the pathogenic bacteria was added to the conical flask with the sterilized substrate, and then the single-strain solution, the double-strain compound solution, and the triple-strain compound solution were added to different conical flasks, respectively. A conical flask without the biocontrol bacteria solution was used as a control, and each group was repeated three times. The conical flasks were placed in a 30°C incubator for culture, and the moisture content was observed regularly, and water was added in time. Samples were taken at 3d, 7d, and 15d, respectively, gradient dilution was performed, and plate counting was performed to count the bacteria and pathogenic bacteria, and the growth of the pathogenic bacteria and the activity of the functional microorganisms were observed, and the change of the microorganisms was analyzed. According to the plate coating results of 3d, 7d, and 15d, the following conclusions were obtained: in terms of the bacteriostatic effect, the compound of Rhodopseudomonas palustris and Bacillus subtilis, the compound of Rhodopseudomonas palustris and Bacillus licheniformis, and the triple-strain compound (the ratio of the number of viable cells of Rhodopseudomonas palustris, Bacillus subtilis, and Bacillus licheniformis was 2:2:1) had good inhibitory effects on the pathogenic bacteria of strawberry root rot, anthracnose, and Labyrinthula lactis. As shown in FIGS. Figure 9 、 Figure 10 and Figure 11

[0075] 3. Preparation of biochar

[0076] The blocky coal gangue was crushed into pieces with a diameter of less than 2 cm by a rock crusher, and then was ball milled with water for 3h. After being taken out, it was dried at 80°C. The caked coal gangue powder was fully ground into powder by a ceramic grinding mortar. The rice husk and charcoal were mixed with the coal gangue by wet method at a mass ratio of 2:1, 5mL of water was added per gram, and they were fully stirred and uniformly dried at 80°C. The modified biochar raw material was heated to the target temperature at a heating gradient of 2.5°C / min by a tube furnace, and was kept at the target temperature for 30min. Then it was naturally cooled, and nitrogen gas with a flow rate of 0.15L / min was introduced as a protective gas to isolate oxygen during the cooling process. In this way, the biochar at 700°C was prepared, and the biochar with a mesh size of 35-100 was obtained after cooling and sieving.

[0077] 4. Preparation of biochar material loaded with bacteria

[0078] ​The mixed bacterial population of biocontrol strains (Rhodopseudomonas palustris: Bacillus subtilis: Bacillus licheniformis = 2:2:1) was immobilized on biochar by adsorption method: the enriched culture of biocontrol strains was mixed in a certain proportion, and the ratio of bacterial liquid to biochar was 0.005:1 (ml / g), i.e. 0.5%. The number of viable bacteria in each milliliter of bacterial liquid was 10 9 CFU / mL, and the mixed bacterial liquid was prepared. The mixed bacterial liquid was transferred to a conical flask containing sterilized biochar, and then the mixture of mixed bacterial liquid and biochar was loaded on a rotary shaker for 24 h to allow the bacteria to adsorb on the surface and pores of the biochar. After 24 h of loading, the mixture was allowed to stand, and then the supernatant was poured off. The obtained biochar was placed in an electric heating air drying oven at 30°C for drying, and the required bacterial-loaded biochar material was obtained. The pH and electrical conductivity of the three-bacterial compound biochar were:

[0079]

[0080] 5. Matrix mixing

[0081] Imported peat, perlite, wood charcoal and rice hull charcoal (both wood charcoal and rice hull charcoal are bacterial-loaded wood charcoal, from the preparation of bacterial-loaded biochar material in Experiment 4) were mixed to prepare wood charcoal-based matrix and rice hull charcoal-based matrix, respectively. Among them, MT represents wood charcoal, DKT represents rice hull charcoal, and CK is the control. The volume ratio of the matrix is as follows:

[0082]

[0083] 6. Laboratory pot experiment

[0084] According to the proportions listed in the above table, different volume percentages of biochar (all loaded with bacteria), peat and perlite were taken to prepare different cultivation matrices. Peat and perlite (3:1) mixed matrix was used as the control group for strawberry cultivation experiment. The prepared cultivation matrix was taken 2 L and placed in a one-gallon plastic pot. Each treatment had 6 replicates, and one pot was a replicate. After the strawberry seedlings were planted and watered, they were placed on a culture shelf equipped with LED light source. The strawberry seedlings were selected from Xiangye virus-free seedlings. After the different treatment matrices were air-dried, the bulk density, porosity, pH and EC were measured. After 30 days, the plant height, leaf number, leaf length, leaf width and petiole length of the strawberry seedlings in different treatments were investigated, and the net photosynthetic rate (Pn), transpiration rate (Tr), stomatal conductance (Gs), intercellular CO2 concentration (Ci) and water use efficiency (WUE) of the leaves were measured using a CIRAS-3 portable photosynthesis system (UK, PPSystem), as shown in Figure 12 .

[0085] Experimental results:

[0086] (1) After 1 month of pot culture, the bulk density, porosity, pH and electrical conductivity (EC) of different pots were measured, and the data results are as follows:

[0087]

[0088] (2) The plant height, leaf number, leaf length, leaf width and petiole length of strawberry seedlings under different treatments are shown in the following table:

[0089]

[0090] Plant height: After adding biochar, the plant height of rice husk charcoal and wood charcoal was higher than that of the control group, but with the increase of biochar amount, the plant height first showed an upward trend (10%-20%), and then began to decline (after 20%-30%), indicating that the addition of biochar promoted plant growth, but as the amount increased, the promotion effect would not be obvious. And with the same proportion of biochar addition, the performance of wood charcoal was better than that of rice husk charcoal.

[0091] Leaf number: After adding biochar, the leaf number of rice husk charcoal and wood charcoal was higher than that of the control group, but with the increase of biochar amount, the leaf number first showed an upward trend (10%-20%), and then began to decline (after 20%-30%), indicating that the addition of biochar promoted plant growth, but as the amount increased, the promotion effect would not be obvious. And with the same proportion of biochar addition, the performance of wood charcoal was better than that of rice husk charcoal.

[0092] Leaf length: After adding biochar, the leaf length of rice husk charcoal and wood charcoal was higher than that of the control group, but with the increase of biochar amount, the leaf length first showed an upward trend (10%-20%), and then began to decline (after 20%-30%), indicating that the addition of biochar promoted plant growth, but as the amount increased, the promotion effect would not be obvious. And with the same proportion of biochar addition, the performance of wood charcoal was better than that of rice husk charcoal.

[0093] Leaf width and petiole length also showed the above trend.

[0094] Therefore, it can be preliminarily indicated that the wood charcoal substrate loaded with mixed bacteria has a promoting effect on the growth of strawberries.

[0095] (3) The chlorophyll content and nitrogen content of strawberry leaves under different treatments are shown in the following table:

[0096]

[0097] With the increase of wood charcoal and rice husk charcoal addition amount, the chlorophyll content and nitrogen content first showed an upward trend, and then began to decline (after 20%). 20% is the best addition amount, and the effect of wood charcoal is better than that of rice husk charcoal.

[0098] (4) The net photosynthetic rate (Pn), transpiration rate (Tr), stomatal conductance (Gs), intercellular CO2 concentration (Ci) and water use efficiency (WUE) of strawberry leaves under different treatments are shown in the following table:

[0099]

[0100] The net photosynthetic rate, transpiration rate and stomatal conductance of the leaves increased first and then decreased with the increase of the amount of rice husk carbon, and increased with the increase of the amount of wood carbon; the intercellular CO2 concentration decreased with the increase of the amount of rice husk carbon, and increased first and then decreased with the increase of the amount of wood carbon; the water use efficiency increased first and then decreased with the increase of the amount of the two kinds of biochar. Therefore, the leaf photosynthetic parameters under the treatment of biochar were generally higher than those of the control, and the effect of adding 10%-20% wood carbon was better.

[0101] In summary, through the determination of the physical and chemical properties of the substrate after adding different proportions of biochar loaded with bacteria, and the monitoring of the morphological indicators and photosynthetic physiological parameters of strawberry after potting, it is preliminarily determined that adding 10%-20% biochar loaded with bacteria to the substrate is beneficial to the growth of strawberry, and the effect of adding 10%-20% wood carbon is better than that of rice husk carbon.

[0102] 7. Greenhouse strawberry planting experiment

[0103] Based on the laboratory potting experiment, in this embodiment, wood carbon and biochar loaded with mixed bacterial colonies will be selected, and the addition proportion will be 10% and 20% by volume respectively. Six treatments are set, namely domestic control substrate (GC), imported control substrate (CK), 10% wood carbon (MT10), 20% wood carbon (MT20), 10% rice husk carbon (DT10) and 20% rice husk carbon (DT20), 6 rows are planted in each treatment as 6 replicates, a total of 36 rows. Among them, in each treatment, 3 rows (3 replicates) only carry out general water and fertilizer management, and do not inoculate microorganisms in the whole process; the remaining 3 rows (3 replicates) add spore suspension with a concentration of 10 5 CFU / ml of Talaromyces muroii (basket fungus) and Penicillium caperatum (Penicillium fungus) after the present budding period, and observe whether the co-action of the three functional bacteria and two growth-promoting bacteria originally loaded on the biochar has a growth-promoting effect on strawberry planting and production.

[0104] The pH, electrical conductivity (EC) and total nitrogen (TN) contents of the substrates were measured before and during the flowering and fruiting stages of strawberry cultivation. Morphological indicators (petiole length, leaf length, and leaf width) were monitored during the budding and flowering and fruiting stages. The net photosynthetic rate (Pn) of the leaves was determined using a CIRAS-3 portable photosynthesis system (UK, PPSystem). The SPAD value of 10 strawberry plants in each treatment was determined using a portable chlorophyll meter to represent the chlorophyll content. The dry weight of the strawberry leaves was then measured in the laboratory, and the specific leaf mass (g / m 2 ) was calculated. The remaining leaves were dried at 80 ℃, crushed, and digested. The nitrogen content was determined using the Kjeldahl method, and the photosynthetic nitrogen use efficiency was then calculated.

[0105] The substrate and biochar were mixed in the following proportions: the ratio of biochar to substrate was consistent with the pot experiment.

[0106]

[0107] Cultivation method:

[0108] The imported substrate was mixed with wood charcoal and rice husk charcoal in the above-mentioned volume proportions, and then packed by hand. The substrate strip used was 1 m long, 30 cm wide, and 30 cm high, with a volume of 35 L. According to the experimental design, 15 ridges were formed in a 0.8 mu greenhouse, with a ridge distance of 1.2 m, a ridge height of 20 cm, and a ridge length of 10 m. There were 20 packs of substrate strips per ridge, arranged in two rows with 10 packs in each row. There were 4 strawberry plants per pack, with a plant distance of 25 cm, and a total of 80 strawberry plants per ridge. The strawberry variety used was the same as that used in the pot experiment. After the ridges were formed, the substrate strips were placed side by side on the ridges, and 10-odd small holes were drilled on the sides and bottoms of the substrate strips for ventilation. The top of the substrate strip was fully open, and the drip tape was laid flat on the substrate strip. Figure 13

[0109] (1) Effect of different treatments on substrate nitrogen content

[0110] The TN contents of the substrates used before and after strawberry cultivation are shown in the table below. The addition of biochar increased the nitrogen content of the substrate to some extent, indicating that biochar can provide a certain amount of nitrogen element. Compared with the control group (CK), the total nitrogen content increased after the addition of 10% and 20% biochar, indicating that biochar can provide a nitrogen source for the substrate.

[0111]

[0112] ​(2) The leaf petiole length, leaf length and leaf width of the strawberry at the present budding stage and the flowering and fruiting stage under different treatments are shown in the following table. The results show that the leaf petiole length, leaf length and leaf width in the treatment group with the addition of biochar are higher than those in the control group (CK). Meanwhile, three parallel treatments of each group are added with Talaromyces muroii (Talaromyces) and Penicillium sp. (Penicillium) after planting, and the leaf performance is described later.

[0113]

[0114] (3) The leaf net photosynthetic rate of the strawberry under different treatments is shown in the following table. The data results show that the leaf net photosynthetic rate of the treatment group with the addition of biochar is higher than that of the control group (CK) at the present budding stage and the flowering and fruiting stage, and the treatment group with the addition of woody charcoal is higher than that with the addition of rice husk charcoal, and the leaf net photosynthetic rate with the addition of 20% woody charcoal is the highest at the present budding stage and the flowering and fruiting stage.

[0115]

[0116] (4) The leaf light response characteristics of the strawberry at the present budding stage and the flowering and fruiting stage under different treatments are shown in the following table. The maximum net photosynthetic rate, light saturation point, light compensation point and dark respiration of the biochar substrate loaded with mixed bacteria are higher than those of the control group (CK), and the light saturation point of the biochar addition is much higher than that of the control group. At the present budding stage, the light saturation point and the light compensation point of 10% woody charcoal are the largest, indicating that the plant with the addition of 10% biochar has the strongest and most sufficient photosynthesis, and the maximum net photosynthetic rate of 20% woody charcoal is the largest. At the flowering and fruiting stage, the maximum net photosynthetic rate and the light saturation point of 10% woody charcoal are the largest. In particular, it can be seen that the addition of 10% woody charcoal can obviously enhance the photosynthetic rate of the strawberry crop.

[0117]

[0118] 8. Fungus inoculation

[0119] Through literature query and experimental research, the present inventors found that Talaromyces muroii (Talaromyces) and Penicillium caperatum (Penicillium) have no experimental and data support for the synergistic effect with Bacillus subtilis, Rhodopseudomonas palustris and Bacillus licheniformis and the performance on the growth of strawberries after being loaded with biochar. Therefore, three parallel groups of each treatment are set as not inoculated with fungi, inoculated with Talaromyces muroii (Talaromyces) and inoculated with Penicillium caperatum (Penicillium) after the present budding stage, and there are 18 treatments (the woody charcoal and the rice husk charcoal of the following treatments are loaded with bacteria), which are as follows:

[0120] Treatment 1: Imported peat + perlite + 30 ml water (40 substrate packs);

[0121] Treatment 2: Imported peat + perlite + 30 ml basket fungus (10 substrate packs);

[0122] Treatment 3: Imported peat + perlite + 30 ml Penicillium fungus (10 substrate packs);

[0123] Treatment 4: Imported peat + perlite + 10% wooden charcoal + 30 ml water (40 substrate packs);

[0124] Treatment 5: Imported peat + perlite + 10% wooden charcoal + 30 ml basket fungus (10 substrate packs);

[0125] Treatment 6: Imported peat + perlite + 10% wooden charcoal + 30 ml Penicillium fungus (10 substrate packs);

[0126] Treatment 7: Imported peat + perlite + 20% wooden charcoal + 30 ml water (40 substrate packs);

[0127] Treatment 8: Imported peat + perlite + 20% wooden charcoal + 30 ml basket fungus (10 substrate packs);

[0128] Treatment 9: Imported peat + perlite + 20% wooden charcoal + 30 ml Penicillium fungus (10 substrate packs);

[0129] Treatment 10: Imported peat + perlite + 10% rice husk charcoal + 30 ml water (40 substrate packs);

[0130] Treatment 11: Imported peat + perlite + 10% rice husk charcoal + 30 ml basket fungus (10 substrate packs);

[0131] Treatment 12: Imported peat + perlite + 10% rice husk charcoal + 30 ml Penicillium fungus (10 substrate packs);

[0132] Treatment 13: Imported peat + perlite + 20% rice husk charcoal + 30 ml water (40 substrate packs);

[0133] Treatment 14: Imported peat + perlite + 20% rice husk charcoal + 30 ml basket fungus (10 substrate packs);

[0134] Treatment 15: Imported peat + perlite + 20% rice husk charcoal + 30 ml Penicillium fungus (10 substrate packs);

[0135] Treatment 16: Domestic peat + perlite + 30 ml water (40 substrate packets);

[0136] Treatment 17: Domestic peat + perlite + 30ml basket bacteria (10 substrate packets);

[0137] Treatment 18: Domestic peat + perlite + 30ml Penicillium (10 substrate packets).

[0138] Except for the inoculation of Basilaria and Penicillium, the other materials and bacterial composition were the same as those in the aforementioned greenhouse experiment.

[0139] Experimental data show that adding Talaromyces muroii (basket-shaped bacteria) and Penicillium caperatum The biochar substrate of (Penicillium) showed better performance in terms of plant height, stem diameter, and maximum leaf area than the control group (GK) and the domestic substrate (GC) without biochar.

[0140]

[0141] In summary, this invention utilizes the advantages of different existing microorganisms through mixing and compounding, and innovatively discovers that mixed strains have a good promoting effect on strawberry plant growth, making up for the shortcomings of traditional biological control methods where single-strain control of certain diseases is insufficient. This is achieved through three functional bacteria (Bacillus subtilis, Rhodopseudomonas palustris, and Bacillus lysine) and two fungi (…). Talaromyces muroii The combined effects of *Bacillus subtilis* and *Penicillium sp.* showed that multi-strain formulations had a better promoting effect on strawberry plant growth. Experimental data indicated that adding a mixed strain of *Bacillus subtilis*, *Rhodopseudomonas palustris*, and *Bacillus lysine* to biochar, with the addition of 10% lignin charcoal, was most effective for strawberry plant growth. When added separately... Talaromyces muroii (basket-shaped bacteria) and Penicillium caperatum When Penicillium is present, the three functional bacteria mentioned above synergistically couple with the two fungi to jointly promote plant growth, and the plant growth is most significant under the condition of 20% rice husk charcoal. Therefore, this invention has found that when Rhodopseudomonas palustris: Bacillus subtilis: Bacillus lysine is in a ratio of 2:2:1, adding... Talaromyces muroii Bassula ovata, and rice husk charcoal as a load carrier, are the most effective at promoting strawberry growth.

[0142] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.

Claims

1. A microorganism substrate for strawberry cultivation, characterized by, The substrate is made of peat, perlite, bacteria-loaded biochar and fungi, and the volume ratio of the peat, the perlite and the bacteria-loaded biochar is (45-75):25:(10-30); The bacteria-loaded biochar is prepared by loading biochar with biocontrol composite bacteria; the biocontrol composite bacteria are composed of Bacillus subtilis, Rhodopseudomonas palustris and Bacillus licheniformis; The fungus is a basket fungus Talaromyces muroii and / or a penicillium Penicillium caperatum ; The live bacteria quantity ratio of the Rhodopseudomonas palustris, the Bacillus subtilis and the Bacillus licheniformis is (2-4):(2-4):

1.

2. The microbial substrate for strawberry cultivation according to claim 1, characterized by, The biochar is prepared by the following method: rice husk and / or charcoal and coal gangue powder are mixed by wet method at a set mass ratio, fully stirred and uniformly mixed, dried, to obtain lignin biochar and / or rice husk biochar; the lignin biochar and / or rice husk biochar raw material is heated, heated to 600-800℃, then naturally cooled, and nitrogen is introduced as a protective gas to isolate oxygen during the period, and the biochar is obtained after cooling.

3. The microbial substrate for strawberry cultivation according to claim 1, characterized by, The bacteria-loaded biochar is prepared by the following method: the biocontrol composite bacteria liquid is transferred to a container containing sterilized biochar, then the mixture of the biocontrol composite bacteria liquid and the biochar is loaded on a rotary shaker for a set time, so that the biocontrol composite bacteria are adsorbed on the surface and pores of the biochar; after loading, the mixture is left to stand, then the supernatant is poured off, and the obtained biochar is placed in a drying oven for drying, to obtain the bacteria-loaded biochar.

4. The method for preparing a microbial substrate for strawberry cultivation according to any one of claims 1 to 3, characterized in that, The method comprises the following steps: (1) Preparation of biochar: rice husk and / or charcoal and coal gangue powder are mixed by wet method at a set mass ratio, fully stirred and uniformly mixed, dried, to obtain lignin biochar and / or rice husk biochar; the lignin biochar and / or rice husk biochar raw material is heated, heated to 600-800℃, then naturally cooled, and nitrogen is introduced as a protective gas to isolate oxygen during the period, and the biochar is obtained after cooling; (2) Preparation of bacteria-loaded biochar material: the biocontrol composite bacteria liquid is transferred to a container containing sterilized biochar, then the mixture of the biocontrol composite bacteria liquid and the biochar is loaded on a rotary shaker for a set time, so that the biocontrol composite bacteria are adsorbed on the surface and pores of the biochar; after loading, the mixture is left to stand, then the supernatant is poured off, and the obtained biochar is placed in a drying oven for drying, to obtain the bacteria-loaded biochar; (3) Compounding: a set amount of peat, perlite, bacteria-loaded biochar and fungi are mixed uniformly, to obtain the microbial substrate for strawberry planting.

5. The method for preparing a microbial substrate for strawberry cultivation according to claim 4, characterized by, In step (1), the mass ratio of the rice husk and / or charcoal to the coal gangue powder is (2-4):(1-2).

6. The method for preparing a microbial substrate for strawberry cultivation according to claim 5, characterized by, In step (2), the ratio of the biocontrol complex bacteria liquid to the biochar is 0.005 mL:(0.5-1.5) g, and the number of living bacteria contained in each milliliter of the bacteria liquid is 10 9 CFU / mL, and the ratio of the number of living bacteria of the Rhodopseudomonas palustris, Bacillus subtilis and Lysinibacillus sphaericus is (2-4):(2-4):

1. In step (2), the loading time is 12-36h.

7. Use of the microbial substrate of any one of claims 1-3 in promoting the growth of strawberries and preventing and treating diseases and pests.

8. The use according to claim 7, characterized in that The diseases and pests are strawberry root rot and strawberry anthracnose.

9. A method of growing a soilless strawberry growing medium, characterized by, The method comprises the following steps: bagging the microbial substrate for strawberry planting according to any one of claims 1-3, making into a substrate strip, stacking the substrate strip side by side on the ridge after ridging, punching small holes on the side and bottom of the substrate strip respectively for ventilation, opening all the holes above the substrate strip, and laying a drip tape belt on the substrate strip.

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