A composition for preventing and treating soil-borne diseases of crops and a method for preparing the same

Abstract

The present application relates to the technical field of crop disease prevention and treatment, and particularly relates to a composition for preventing and treating soil-borne diseases of crops and a preparation method thereof, comprising: activating and fermenting strains respectively to obtain a microbial agent mixed solution; obtaining biochar by high-temperature carbonization of corn stalks and adding the biochar to the microbial agent mixed solution to obtain biochar powder with strains adsorbed and fixed; uniformly mixing the biochar powder and a polyvinyl alcohol solution to obtain a strain suspension; adding the strain suspension dropwise to a calcium chloride solution to obtain biochar-based composite strain microspheres; preparing a chitosan / poly(N-isopropyl acrylamide) solution and determining whether the preparation of the mixed solution meets a preset standard according to a light transmittance difference value of the mixed solution; and transferring the biochar-based composite strain microspheres to the chitosan / poly(N-isopropyl acrylamide) solution to coat a film to obtain temperature-sensitive composite microbial agent microspheres, so that the environmental adaptability of the composition for preventing and treating soil-borne diseases of crops is improved.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of crop disease prevention and treatment, and particularly relates to a composition for preventing and treating soil-borne diseases of crops and a preparation method thereof. BACKGROUND

[0002] Soil-borne diseases refer to diseases caused by pathogens such as fungi, bacteria, nematodes and viruses living in the soil with plant residues, and invading crops from the roots or stems under suitable conditions.

[0003] The soil-borne disease complex microbial agent is a microbial preparation for preventing and treating soil-borne diseases, which is usually made of two or more than two microbial strains that do not antagonize each other. These strains can not only cooperate with each other in the soil, but also improve soil structure, increase soil fertility and promote crop growth.

[0004] A biocontrol agent preparation for preventing and treating soil-borne diseases is disclosed in Chinese Patent Publication No. CN111357771B, which is prepared from the following raw materials in a certain proportion by weight: Sargassum thunbergii, Bangia atropurpurea, Chlorella vulgaris, Eucheuma cottonii, Ulva pertusa, Rhodiola root, Bacillus amyloliquefaciens HAB-2, Bacillus subtilis, Trichoderma asperellum, yeast and Chaetomium aureum. The present application can effectively prevent and treat soil-borne diseases, improve soil quality and soil microbial composition, greatly improve the growth conditions of field crops and promote yield increase. It can prevent and treat soil-borne diseases such as cucumber fusarium wilt, strawberry anthracnose and rice blast, which has important significance for the sustainable development of agricultural production. It can be seen that in the above technical solution, the biocontrol agent preparation has no protective film on the outer layer, is easily disturbed by soil temperature, has poor environmental adaptability of the biocontrol agent preparation, poor effect of preventing and treating soil-borne diseases of crops, and low yield of crops. SUMMARY

[0005] Therefore, the present application provides a composition for preventing and treating soil-borne diseases of crops and a preparation method thereof to overcome the problems in the prior art that the outer layer of the complex microbial agent has no protective film and is easily disturbed by soil temperature, the environmental adaptability of the biocontrol agent preparation is poor, the effect of preventing and treating soil-borne diseases of crops is poor, and the yield of crops is low.

[0006] To achieve the above-mentioned purpose, on the one hand, the present application provides a preparation method of a composition for preventing and treating soil-borne diseases of crops, comprising:

[0007] The Bacillus subtilis and Bacillus amyloliquefaciens are inoculated into two activated culture media respectively for culture, and first seed culture solution and second seed culture solution are obtained respectively;

[0008] The first seed culture solution and the second seed culture solution are fermented and cultured respectively, and first fermentation broth and second fermentation broth are obtained respectively;

[0009] Filtering and impurity-removing the first and second fermentation bacterial solutions to obtain first and second bacterial stock solutions, and mixing the first and second bacterial stock solutions to obtain a bacterial agent mixture solution;

[0010] After high-temperature carbonization of corn stalks, the corn stalks are ground into biochar by a ball mill; the biochar is added to the bacterial agent mixture solution, and the precipitate obtained after separation and treatment of the bacterial agent mixture solution is vacuum dried to obtain biochar powder in which the Bacillus subtilis and the Bacillus amyloliquefaciens are adsorbed and fixed;

[0011] The biochar powder is mixed with a polyvinyl alcohol solution to obtain a bacterial stock suspension, and the bacterial stock suspension is added dropwise to a calcium chloride solution to obtain biochar-based composite bacterial stock microspheres.

[0012] At room temperature, chitosan powder is weighed and added to a 2% acetic acid solution, stirred and allowed to stand to prepare a chitosan solution with a concentration of 4%, and the viscosity of the chitosan solution is obtained.

[0013] At room temperature, poly(N-isopropyl acrylamide) is dissolved in deionized water, an initiator and a crosslinking agent are added to perform a free radical polymerization reaction, and the poly(N-isopropyl acrylamide) solution is stirred to obtain the viscosity of the poly(N-isopropyl acrylamide) solution.

[0014] At room temperature, the chitosan solution is poured into the poly(N-isopropyl acrylamide) solution at a preset pouring speed, stirred for a preset time, and the transmittance of the mixed solution is continuously obtained to determine the time when the transmittance stops changing.

[0015] When the preparation of the mixed solution does not meet the preset standard according to the transmittance difference of the mixed solution, the preset pouring speed is reduced, or the stirring speed during the preparation of the mixed solution is reduced or the mixed solution is filtered according to the uniformity characteristic value of the solution to obtain a chitosan / poly(N-isopropyl acrylamide) solution.

[0016] The biochar-based composite bacterial stock microspheres are transferred to the chitosan / poly(N-isopropyl acrylamide) solution to form a film, filtered, separated, washed, and then vacuum dried to obtain temperature-sensitive composite bacterial agent microspheres.

[0017] Further, the activation culture conditions of the Bacillus subtilis and the Bacillus amyloliquefaciens are that the pH of the culture medium is set to 7.4, the temperature is 37℃, the culture time is 18h, and the shaking speed is 180r / min.

[0018] Further, the weight ratio of the activation culture medium is that the potato is 20%, the glucose is 4%, the beef extract is 0.5%, the peptone is 2%, the sodium chloride is 2%, the agar is 1.5%, and the rest is water.

[0019] Further, the fermentation culture process of the bacillus subtilis and bacillus amyloliquefaciens comprises the following steps: inoculating the first seed culture solution and the second seed culture solution into two fermentation tanks respectively under sterile conditions, the inoculation amount is 4%, the components of the two fermentation mediums are same, the mass content of each component in 1L distilled water is as follows: soybean meal 2%, urea 0.5%, ammonium sulfate 0.2%, starch 4%, molasses 5%, potassium dihydrogen phosphate 0.8%, sodium phosphate dibasic 0.2%, magnesium sulfate 0.01%, the stirring speed is 240r / min, the culture temperature is 30-35℃, the ventilation amount is 1:0.8, and the fermentation time is 24-30h.

[0020] Further, the process of obtaining the viscosity of the chitosan solution and obtaining the viscosity of the poly(N-isopropylacrylamide) solution comprises the following steps:

[0021] Placing the chitosan solution and the poly(N-isopropylacrylamide) solution into constant temperature water baths respectively, adjusting the water bath temperature to room temperature, and keeping constant temperature;

[0022] Slowly immersing the rotor of the first rotary viscometer and the rotor of the second rotary viscometer into the chitosan solution and the poly(N-isopropylacrylamide) solution respectively;

[0023] Starting the rotary viscometer, and recording the torque value of the rotor at a preset rotating speed;

[0024] The viscosity is the ratio between the product of the rotary viscometer constant and the torque and the angular velocity;

[0025] Repeating the measurement several times, calculating the average value of the viscosity of the chitosan solution, and calculating the average value of the viscosity of the poly(N-isopropylacrylamide) solution;

[0026] Taking the average value of the viscosity of the chitosan solution as the viscosity of the chitosan solution, and taking the average value of the viscosity of the poly(N-isopropylacrylamide) solution as the viscosity of the poly(N-isopropylacrylamide) solution.

[0027] Further, the process of judging that the preparation of the mixed solution does not meet the preset standard according to the light transmittance difference of the mixed solution comprises the following steps:

[0028] Starting timing when the chitosan solution is poured into the poly(N-isopropylacrylamide) solution at a preset pouring speed at room temperature;

[0029] Obtaining the light transmittance of the mixed solution at the starting time through a transmission spectrometer;

[0030] Stopping timing when the light transmittance of the mixed solution stops changing, and obtaining the time length from the starting timing to the time when the light transmittance of the mixed solution stops changing;

[0031] calculating a light transmittance difference of the mixed solution;

[0032] comparing the light transmittance difference of the mixed solution with a first preset light transmittance difference and a second preset light transmittance difference respectively;

[0033] if the light transmittance difference of the mixed solution is less than the first preset light transmittance difference, it is determined that the preparation of the mixed solution does not meet the preset standard, and the preset pouring speed is reduced according to the ratio between the first preset light transmittance difference and the light transmittance difference of the mixed solution;

[0034] if the light transmittance difference of the mixed solution is greater than or equal to the first preset light transmittance difference and less than the second preset light transmittance difference, it is determined that the preparation of the mixed solution does not meet the preset standard, and the preparation of the mixed solution is determined again according to the solution uniformity characteristic value whether it meets the preset standard.

[0035] Further, the solution uniformity characteristic value is determined according to the viscosity of the chitosan solution, the viscosity of the poly(N-isopropyl acrylamide) solution and the time length for the light transmittance of the mixed solution to reach stability.

[0036] Further, the process of determining again whether the preparation of the mixed solution meets the preset standard according to the solution uniformity characteristic value comprises:

[0037] comparing the solution uniformity characteristic value with a preset solution uniformity characteristic value;

[0038] if the solution uniformity characteristic value is less than the preset solution uniformity characteristic value, it is determined that the preparation of the mixed solution meets the preset standard, and the mixed solution meeting the preset standard is filtered to obtain a chitosan / poly(N-isopropyl acrylamide) solution;

[0039] if the solution uniformity characteristic value is greater than or equal to the preset solution uniformity characteristic value, it is determined that the preparation of the mixed solution does not meet the preset standard, and the stirring speed in the preparation process of the mixed solution is reduced according to the difference between the solution uniformity characteristic value and the preset solution uniformity characteristic value.

[0040] Further, there are several speed reduction modes for the reduction of the stirring speed in the preparation process of the mixed solution, and each speed reduction mode has a different reduction amplitude of the stirring speed in the preparation process of the mixed solution.

[0041] On the other hand, the present application also provides a composition for preventing and treating soil-borne diseases of crops, which is composed of the following raw materials: biochar, Bacillus subtilis, Bacillus amyloliquefaciens, polyvinyl alcohol, chitosan and poly(N-isopropyl acrylamide).

[0042] Compared with the prior art, the present application has the beneficial effects that: the present application activates and ferments Bacillus subtilis and Bacillus amyloliquefaciens respectively, mixes the bacterial stock solution to obtain a microbial agent mixed solution, adsorbs and solidifies the composite bacterial strain by using biochar to improve the survival rate of the bacterial strain, forms biochar-based composite bacterial strain microspheres through a polyvinyl alcohol solution, and finally transfers the biochar-based composite bacterial strain microspheres to a chitosan / poly(N-isopropyl acrylamide) solution to provide a protective layer for the biochar-based composite bacterial strain microspheres, protect the activity of the bacterial strain, and improve the environmental adaptability of the composite microbial agent.

[0043] Further, the present application has the prevention and treatment effect on various soil-borne diseases by formulating a composite microbial agent including Bacillus subtilis and Bacillus amyloliquefaciens, and improves the utilization rate of the composition.

[0044] Further, the present application grinds the corn stalks after carbonization through a ball mill to increase the specific surface area of the biochar, thereby improving the adsorption capacity of the biochar.

[0045] Further, the present application uses biochar as a composite bacterial strain carrier, not only adsorbs and fixes the bacterial strain through the porous structure of the biochar, but also provides nutrients and living environment for the bacterial strain, thereby improving the stability of the bacterial strain.

[0046] Further, the present application determines whether the preparation of the mixed solution meets the preset standard according to the light transmittance difference of the mixed solution, thereby ensuring the qualification of the mixed solution.

[0047] Further, the present application sets the solution uniformity characteristic value to provide an evaluation standard for the preparation of the mixed solution, thereby improving the scientificity of the test of the preparation process of the mixed solution.

[0048] Further, the present application determines whether the preparation of the mixed solution meets the preset standard according to the solution uniformity characteristic value twice, thereby improving the reliability of the preparation process of the mixed solution.

[0049] Further, the present application combines the biochar-based composite bacterial strain microspheres with the chitosan / poly(N-isopropyl acrylamide) solution to prepare a composite microbial agent microsphere with temperature sensitivity, which can release the bacterial strain only under specific temperature conditions to realize precise prevention and treatment, thereby improving the prevention and treatment effect of the composition. BRIEF DESCRIPTION OF DRAWINGS

[0050] Figure 1 The flowchart of the preparation method of the composition for preventing and treating soil-borne diseases of crops in the embodiment of the present application;

[0051] Figure 2 The flowchart of determining whether the preparation of the mixed solution meets the preset standard in the embodiment of the present application;

[0052] Figure 3 A flow chart for determining whether the preparation of the mixed solution meets the preset standard in the embodiment of the present application;

[0053] Figure 4 A flow chart for determining the reducing mode of the stirring speed in the preparation of the mixed solution in the embodiment of the present application. DETAILED DESCRIPTION

[0054] In order to make the objects and advantages of the present application more clear, the present application will be further described in conjunction with the embodiments. It should be understood that the specific embodiments described herein are only used to explain the present application, and are not used to limit the present application.

[0055] The preferred embodiments of the present application will be described below with reference to the accompanying drawings. It should be understood by those skilled in the art that the embodiments are only used to explain the technical principles of the present application, and are not used to limit the protection scope of the present application.

[0056] Please refer to Figure 1 , Figure 2 , Figure 3 and Figure 4 , which are respectively a flow chart of the preparation method of the composition for preventing and treating soil-borne diseases of crops in the embodiment of the present application; a flow chart for determining whether the preparation of the mixed solution meets the preset standard in the embodiment of the present application; a flow chart for secondly determining whether the preparation of the mixed solution meets the preset standard in the embodiment of the present application; and a flow chart for determining the reducing mode of the stirring speed in the preparation of the mixed solution in the embodiment of the present application.

[0057] The embodiment of the present application provides a preparation method of a composition for preventing and treating soil-borne diseases of crops, comprising:

[0058] Step S1, Bacillus subtilis and Bacillus amyloliquefaciens are inoculated into two activated culture media respectively for shaking culture, and first seed culture solution and second seed culture solution are obtained respectively;

[0059] Step S2, the first seed culture solution and the second seed culture solution are inoculated into two fermentation culture media respectively under sterile conditions for fermentation culture, and after the viable cell count of each strain fermentation liquor meets the standard, first fermentation liquor and second fermentation liquor are obtained respectively;

[0060] Step S3, the first fermentation liquor and the second fermentation liquor are filtered and impurities are removed, and first strain stock solution and second strain stock solution are obtained respectively; the first strain stock solution and the second strain stock solution are mixed in a 1:1 ratio to obtain a microbial agent mixed solution;

[0061] Step S4, after the corn straw is carbonized at high temperature and then ground by a ball mill, biochar is formed; the biochar is added to the microbial agent mixed solution, after shaking bed shaking, standing, centrifugal separation, vacuum drying is performed on the precipitate obtained by centrifugation, and biochar powder in which the Bacillus subtilis and the Bacillus amyloliquefaciens are adsorbed and fixed is obtained;

[0062] Step S5, after the biochar powder in which the Bacillus subtilis and the Bacillus amyloliquefaciens are adsorbed and fixed is uniformly mixed with a polyvinyl alcohol solution, a microbial agent suspension is obtained; the microbial agent suspension is added dropwise into a calcium chloride solution, cross-linked and solidified for 24 hours, filtered and separated, and biochar-based composite microbial agent microspheres are obtained.

[0063] Step S6, at room temperature, 4g of chitosan powder is weighed and poured into 100ml of 2% acetic acid solution, stirred uniformly, and after standing, a chitosan solution with a concentration of 4% is prepared, and the viscosity of the chitosan solution is obtained.

[0064] Step S7, at room temperature, poly(N-isopropyl acrylamide) is dissolved in deionized water, an initiator and a crosslinking agent are added, and a free radical polymerization reaction is carried out at 65°C under nitrogen protection, and the poly(N-isopropyl acrylamide) solution is obtained, and the viscosity of the poly(N-isopropyl acrylamide) solution is obtained.

[0065] Step S8, at room temperature, the chitosan solution is poured into the poly(N-isopropyl acrylamide) solution at a preset pouring speed of 10ml / min, stirred for a preset time of 45min, and the transmittance of the mixed solution is continuously obtained to determine the time when the transmittance stops changing.

[0066] Step S9, when the preparation of the mixed solution does not meet the preset standard according to the transmittance difference of the mixed solution, the preset pouring speed is reduced, or the stirring speed in the preparation process of the mixed solution is reduced or the mixed solution is filtered according to the uniformity characteristic value of the solution, and a chitosan / poly(N-isopropyl acrylamide) solution is obtained.

[0067] Step S10, the biochar-based composite microbial agent microspheres are transferred to the chitosan / poly(N-isopropyl acrylamide) solution for film coating, filtered, separated, washed, and then vacuum dried to obtain temperature-sensitive composite microbial agent microspheres.

[0068] In this embodiment, the molar ratio of chitosan to N-isopropyl acrylamide is 1:0.25.

[0069] In this embodiment, the initiator can be potassium persulfate or ammonium persulfate; the crosslinking agent can be N,N'-methylene bisacrylamide.

[0070] In this embodiment, the range of the number of viable bacteria of each strain of fermentation broth that meets the standard is 4x108 cfu / g-5x10 8 cfu / g.

[0071] In this embodiment, the preset time is selected in the range of [30 min, 42 min].

[0072] Specifically, the activation culture conditions of the Bacillus subtilis and the Bacillus amyloliquefaciens are that the pH of the culture medium is set to 7.4, the temperature is 37℃, the culture is 18h, and the shaking speed is 180r / min.

[0073] Specifically, the total number of viable bacteria in the microbial agent mixed solution is not less than 3x10 8 cfu / g.

[0074] Specifically, the weight ratio of the activation culture medium is potato 20%, glucose 4%, beef extract 0.5%, peptone 2%, sodium chloride 2%, agar 1.5%, and the rest is water.

[0075] Specifically, the fermentation culture process of the Bacillus subtilis and the Bacillus amyloliquefaciens includes: under sterile conditions, the first seed culture solution and the second seed culture solution are inoculated into two fermentation tanks respectively, the inoculation amount is 4%, the components of the two fermentation media are the same, and the mass content of each component in 1L distilled water is: soybean meal 2%, urea 0.5%, ammonium sulfate 0.2%, starch 4%, molasses 5%, potassium dihydrogen phosphate 0.8%, sodium hydrogen phosphate 0.2%, magnesium sulfate 0.01%, the stirring speed is 240r / min, the culture temperature is 30℃-35℃, the aeration amount is 1:0.8, and the fermentation time is 24h-30h.

[0076] Specifically, the process of obtaining the viscosity of the chitosan solution and obtaining the viscosity of the poly(N-isopropyl acrylamide) solution includes:

[0077] Step S701, the chitosan solution and the poly(N-isopropyl acrylamide) solution are respectively placed in a constant temperature water bath, the water bath temperature is adjusted to room temperature, and the constant temperature is maintained;

[0078] Step S702, slowly immerse the rotor of the first rotary viscometer and the rotor of the second rotary viscometer into the chitosan solution and the poly(N-isopropyl acrylamide) solution respectively, and avoid generating bubbles;

[0079] Step S703, start the rotary viscometer, and record the torque value of the rotor at a preset speed of 30r / min;

[0080] Step S704, the viscosity is the ratio between the product of the rotary viscometer constant and the torque and the angular velocity;

[0081] Step S705, repeating the measurement several times, calculating the average value of the viscosity of the chitosan solution, calculating the average value of the viscosity of the poly(N-isopropylacrylamide) solution;

[0082] Step S706, recording the average value of the viscosity of the chitosan solution as the viscosity of the chitosan solution; recording the average value of the viscosity of the poly(N-isopropylacrylamide) solution as the viscosity of the poly(N-isopropylacrylamide) solution.

[0083] Specifically, the process of determining that the preparation of the mixed solution does not meet the preset standard according to the transmittance difference of the mixed solution comprises:

[0084] Timing starts when the chitosan solution is poured into the poly(N-isopropylacrylamide) solution at a preset pouring speed at room temperature;

[0085] The transmittance of the mixed solution is obtained by a transmission spectrometer;

[0086] When the transmittance of the mixed solution stops changing, timing stops, and the time length from the start of timing to the transmittance of the mixed solution reaching stability is obtained;

[0087] The absolute value of the difference between the transmittance of the mixed solution when the transmittance stops changing and the transmittance of the mixed solution at the beginning is recorded as the transmittance difference of the mixed solution;

[0088] The transmittance difference of the mixed solution is calculated;

[0089] The transmittance difference of the mixed solution is compared with a first preset transmittance difference 3.29% and a second preset transmittance difference 5.26%, respectively;

[0090] If the transmittance difference of the mixed solution is less than the first preset transmittance difference, it is determined that the preparation of the mixed solution does not meet the preset standard, and the preset pouring speed is reduced according to the ratio between the first preset transmittance difference and the transmittance difference of the mixed solution;

[0091] If the transmittance difference of the mixed solution is greater than or equal to the first preset transmittance difference and less than the second preset transmittance difference, it is determined that the preparation of the mixed solution does not meet the preset standard, and the preparation of the mixed solution is determined whether it meets the preset standard according to the solution uniformity characteristic value.

[0092] Specifically, the determination process of the solution uniformity characteristic value comprises:

[0093] a ratio of the viscosity of the chitosan solution to a preset viscosity of the chitosan solution is multiplied by a first evaluation coefficient to obtain a first evaluation value, wherein the first evaluation coefficient is 0.34; a ratio of the viscosity of the poly(N-isopropylacrylamide) solution to a preset viscosity of the poly(N-isopropylacrylamide) solution is multiplied by a second evaluation coefficient to obtain a second evaluation value, wherein the first evaluation coefficient is 0.37; and a ratio of a time length for the transmittance of the mixed solution to reach stability to a preset time length of 15 minutes is multiplied by a third evaluation coefficient to obtain a third evaluation value, wherein the third evaluation coefficient is 0.44;

[0094] a sum of the first evaluation value, the second evaluation value and the third evaluation value is recorded as a solution uniformity characteristic value.

[0095] In the embodiment, the preset viscosity of the chitosan solution is selected in a range of [0.6η1, 0.8η1], wherein η1 is an average value of the viscosity of the chitosan solution.

[0096] The preset viscosity of the poly(N-isopropylacrylamide) solution is selected in a range of [0.7η2, 0.9η2], wherein η2 is an average value of the viscosity of the poly(N-isopropylacrylamide) solution.

[0097] Specifically, a process of determining whether the preparation of the mixed solution meets the preset standard according to the solution uniformity characteristic value twice comprises:

[0098] comparing the solution uniformity characteristic value with a preset solution uniformity characteristic value of 0.87;

[0099] If the solution uniformity characteristic value is less than the preset solution uniformity characteristic value, it is determined that the preparation of the mixed solution meets the preset standard, and the mixed solution meeting the preset standard is filtered to obtain a chitosan / poly(N-isopropylacrylamide) solution.

[0100] If the solution uniformity characteristic value is greater than or equal to the preset solution uniformity characteristic value, it is determined that the preparation of the mixed solution does not meet the preset standard, and the stirring speed in the preparation process of the mixed solution is reduced according to a difference between the solution uniformity characteristic value and the preset solution uniformity characteristic value.

[0101] Specifically, the reduction of the stirring speed in the preparation process of the mixed solution is provided with a plurality of speed reduction modes, wherein,

[0102] If the solution characteristic difference is less than a first preset solution characteristic difference of 0.23, the stirring speed in the preparation process of the mixed solution is reduced by using a first speed adjustment coefficient of 0.98;

[0103] If the solution characteristic difference value is greater than or equal to the first preset solution difference value and less than the second preset solution difference value 0.46, a second speed adjustment coefficient 0.96 is used to reduce the stirring speed in the preparation process of the mixed solution.

[0104] If the solution characteristic difference value is greater than or equal to the second preset solution difference value, a third speed adjustment coefficient 0.94 is used to reduce the stirring speed in the preparation process of the mixed solution; the solution characteristic difference value is the difference between the solution uniformity characteristic value and the preset solution uniformity characteristic value.

[0105] In another aspect, an embodiment of the present application is a composition for preventing and treating soil-borne diseases of crops, which is composed of the following raw materials: biochar, Bacillus subtilis, Bacillus amyloliquefaciens, polyvinyl alcohol, chitosan and poly(N-isopropyl acrylamide).

[0106] The composition for preventing and treating soil-borne diseases of crops prepared by the embodiment of the present application can be used in combination with other fertilizers for application in soil, or the seeds or seedlings can be soaked in the composition before planting, so as to achieve the purpose of preventing and treating soil-borne diseases.

[0107] Example 1

[0108] Taking tomatoes as an example, the tomato field is divided into three areas, which are respectively recorded as a test group, a comparison group and a blank group, wherein the test group is the composition of the present application mixed with traditional fertilizer at a ratio of 1:2; the comparison group is the microbial agent in the prior art mixed with traditional fertilizer at a ratio of 1:2; the blank group is traditional fertilizer; when the tomatoes are picked, the damage rate due to soil-borne diseases is counted, and the statistical results are shown in Table 1 as follows:

[0109] Table 1 Relationship table between soil-borne disease incidence and yield of composition mixed with traditional fertilizer at a ratio of 1:2

[0110] Test group Comparison group Blank group Incidence of bacterial wilt (%) 8.3 16.2 21.7 Tomato yield (kg / acre) 3985 3421 2946

[0111] Example 2

[0112] Taking corn as an example under the condition of large diurnal temperature difference, the corn field is divided into three areas, which are respectively recorded as a test group, a comparison group and a blank group, wherein the test group is the composition of the present application mixed with traditional fertilizer at a ratio of 1:2; the comparison group is the microbial agent in the prior art mixed with traditional fertilizer at a ratio of 1:2; the blank group is only traditional fertilizer; when the corn is picked, the damage rate due to soil-borne diseases is counted, and the statistical results are shown in Table 2 as follows:

[0113] Table 2 Relationship table between soil-borne disease incidence and yield under the condition of large diurnal temperature difference

[0114] Test group Comparison group Blank group Incidence of bacterial wilt (%) 12.7 22.3 57.3 Corn yield (kg / acre) 843 579 367

[0115] The two tables are summarized to conclude that the tomato in the test area is greatly increased in yield per mu by adopting appropriate proportion mixing of the composition and the traditional fertilizer than the conventional fertilization, the composition has the prevention and treatment effect on the bacterial wilt and the effect is obvious, under the condition of large day and night temperature difference, the test group applying the temperature-sensitive composite microbial agent microspheres still has obvious effect on the prevention and treatment of the bacterial wilt, can still improve the crop yield, and improves the environmental applicability of the composite microbial agent.

[0116] So far, the technical solutions of the present application have been described in combination with the preferred embodiments shown in the drawings, but those skilled in the art can easily understand that the protection scope of the present application is obviously not limited to these specific embodiments. Those skilled in the art can make equivalent changes or replacements to the related technical features without departing from the principles of the present application, and the technical solutions after the changes or replacements will fall within the protection scope of the present application.

[0117] The above description is only the preferred embodiments of the present application and is not used to limit the present application; for those skilled in the art, the present application can have various changes and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principles of the present application shall be included in the protection scope of the present application.

Claims

1. A method of preparing a composition for controlling soil-borne diseases of crops, characterized by, The application relates to a preparation method of a biochar-based composite microbial seed microsphere. The Bacillus subtilis and the Bacillus amyloliquefaciens are respectively inoculated into two activated culture mediums to obtain a first seed culture solution and a second seed culture solution; The first seed culture solution and the second seed culture solution are respectively subjected to fermentation culture to obtain a first fermentation bacterial solution and a second fermentation bacterial solution; The first fermentation bacterial solution and the second fermentation bacterial solution are filtered and impurity-removed to obtain a first bacterial seed stock solution and a second bacterial seed stock solution, and the first bacterial seed stock solution and the second bacterial seed stock solution are mixed in proportion to obtain a microbial agent mixed solution; Corn stalks are high-temperature carbonized and ground into biochar by a ball mill; the biochar is added into the microbial agent mixed solution, and the precipitate obtained by separating and treating the microbial agent mixed solution is vacuum-dried to obtain biochar powder in which the Bacillus subtilis and the Bacillus amyloliquefaciens are adsorbed and fixed; The biochar powder is mixed with a polyvinyl alcohol solution to obtain a microbial seed suspension, and the microbial seed suspension is added dropwise into a calcium chloride solution to obtain the biochar-based composite microbial seed microsphere; At room temperature, chitosan powder is added into 2% acetic acid solution, and the mixture is stirred and placed to prepare a chitosan solution with a concentration of 4%, and the viscosity of the chitosan solution is obtained; At room temperature, poly-N-isopropyl acrylamide is dissolved in deionized water, an initiator and a crosslinking agent are added to perform a free radical polymerization reaction, the mixture is stirred to obtain a poly-N-isopropyl acrylamide solution, and the viscosity of the poly-N-isopropyl acrylamide solution is obtained; At room temperature, the chitosan solution is poured into the poly-N-isopropyl acrylamide solution at a preset pouring speed, the mixture is stirred for a preset time, and the transmittance of the mixed solution is continuously obtained to determine the time length when the transmittance stops changing; At room temperature, the time is started when the chitosan solution is poured into the poly-N-isopropyl acrylamide solution at a preset pouring speed; The transmittance of the mixed solution at the starting time is obtained by a transmission spectrometer; When the transmittance of the mixed solution stops changing, the time counting is stopped, and the time length from the starting time to the time when the transmittance of the mixed solution stops changing is obtained; The transmittance difference of the mixed solution is calculated; The transmittance difference of the mixed solution is compared with a first preset transmittance difference and a second preset transmittance difference respectively; If the transmittance difference of the mixed solution is less than the first preset transmittance difference, it is determined that the preparation of the mixed solution does not meet the preset standard, and the preset pouring speed is reduced according to the ratio between the first preset transmittance difference and the transmittance difference of the mixed solution; If the transmittance difference of the mixed solution is greater than or equal to the first preset transmittance difference and less than the second preset transmittance difference, it is determined that the preparation of the mixed solution does not meet the preset standard, and whether the preparation of the mixed solution meets the preset standard is determined according to the solution uniformity characteristic value twice; The process of determining whether the preparation of the mixed solution meets the preset standard according to the solution uniformity characteristic value twice comprises the following steps: The solution uniformity characteristic value is compared with a preset solution uniformity characteristic value; If the solution uniformity characteristic value is less than the preset solution uniformity characteristic value, it is determined that the preparation of the mixed solution meets the preset standard, and the mixed solution meeting the preset standard is filtered to obtain a chitosan / poly-N-isopropyl acrylamide solution; If the solution uniformity characteristic value is greater than or equal to the preset solution uniformity characteristic value, it is determined that the preparation of the mixed solution does not meet the preset standard, and the stirring speed in the preparation process of the mixed solution is reduced according to the difference between the solution uniformity characteristic value and the preset solution uniformity characteristic value; The solution uniformity characteristic value is determined according to the viscosity of the chitosan solution, the viscosity of the poly-N-isopropyl acrylamide solution, and the time length for the transmittance of the mixed solution to reach stability; The reduction of the stirring speed in the preparation process of the mixed solution is provided with several speed reduction modes, and each speed reduction mode has a different reduction amplitude of the stirring speed in the preparation process of the mixed solution; The biochar-based composite microbial inoculum microspheres are transferred into a chitosan / poly-N-isopropyl acrylamide solution for film coating, and then filtered, separated, washed, and vacuum dried to obtain temperature-sensitive composite microbial inoculum microspheres.

2. The method of preparing a composition for controlling soil-borne diseases of crops according to claim 1, characterized in that, The activation culture conditions of the Bacillus subtilis and the Bacillus amyloliquefaciens are both set as follows: the pH of the culture medium is 7.4, the temperature is 37°C, the culture time is 18h, and the shaking speed is 180r / min.

3. The method for preparing the composition for controlling soil-borne crop diseases according to claim 2, characterized in that, The weight ratio of the activation culture medium is as follows: potato 20%, glucose 4%, beef extract 0.5%, peptone 2%, sodium chloride 2%, agar 1.5%, and the rest is water.

4. The method for preparing the composition for controlling soil-borne crop diseases according to claim 3, characterized in that, The fermentation culture process of the Bacillus subtilis and the Bacillus amyloliquefaciens includes the following steps: under sterile conditions, the first seed culture liquid and the second seed culture liquid are inoculated into two fermentation tanks respectively, and the inoculation amount is 4% in each tank; the two fermentation culture media have the same components, and the mass content of each component in 1L distilled water is as follows: soybean meal 2%, urea 0.5%, ammonium sulfate 0.2%, starch 4%, molasses 5%, potassium dihydrogen phosphate 0.8%, sodium hydrogen phosphate 0.2%, and magnesium sulfate 0.01%; the stirring speed is 240r / min, the culture temperature is 30°C~35°C, the aeration amount is 1:0.8, and the fermentation time is 24h~30h.

5. The method for preparing the composition for controlling soil-borne crop diseases according to claim 4, characterized in that, The process for obtaining the viscosity of the chitosan solution and the viscosity of the poly-N-isopropyl acrylamide solution includes the following steps: The chitosan solution and the poly-N-isopropyl acrylamide solution are respectively placed in a constant-temperature water bath, the water bath temperature is adjusted to room temperature, and the temperature is kept constant; The rotors of the first and second rotary viscometers are slowly immersed in the chitosan solution and the poly-N-isopropyl acrylamide solution respectively; The rotary viscometer is started, and the torque value of the rotor at a preset rotating speed is recorded; The viscosity is the ratio between the product of the rotary viscometer constant and the torque and the angular velocity; The average value of the viscosity of the chitosan solution is calculated by repeating the measurement several times, and the average value of the viscosity of the poly-N-isopropyl acrylamide solution is calculated; The average value of the viscosity of the chitosan solution is recorded as the viscosity of the chitosan solution, and the average value of the viscosity of the poly-N-isopropyl acrylamide solution is recorded as the viscosity of the poly-N-isopropyl acrylamide solution.

Images