CN103141518B - Preparation method and application of multi-effect compound anti-biological inoculants - Google Patents
Preparation method and application of multi-effect compound anti-biological inoculants Download PDFInfo
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Abstract
The invention provides a preparation method and an application of multi-effect anti-biological inoculants for controlling crop soil-borne diseases. According to the preparation method, trichoderma viride strains and endogeny bacillus subtilis having antagonistic action to pathogenic bacteria of the crop soil-borne diseases are used as parent inoculants, and light calcium carbonate is used as a carrier to prepare the anti-biological inoculants. The anti-biological inoculants are mainly used for controlling various crop soil-borne diseases caused by pathogenic fungi including fusarium, pythium, rhizoctonia and the like, and have multiple functions of improving self resistance and increasing the output of plants. By adopting the preparation method, the anti-biological inoculants are short in production cycle, stable and lasting in control effect and pollution-free to the environment without residues to crops.
Description
Technical field
The present invention relates to a kind of biocontrol fungicide, particularly a kind of preparation method of the multiple-effect compound biocontrol fungicide for preventing and treating soil-borne disease of crop and application thereof, two described bacterial strains are respectively Trichoderma
(Trichoderma viride) and Bacillus subtillis (
bacillus subtilis).
Background technology
Along with more and more toxic chemical agricultural chemicals is limited to use, countries in the world are all in the R&D intensity strengthening biopesticide.The Antagonism fungi that Trichoderma has important biological and ecological methods to prevent plant disease, pests, and erosion to be worth as a class, just has been found that there is antagonism to plant pathogenic fungi as far back as the thirties in 20th century, and its mechanism of action comprises generation antibiotic, hyperparasitism, bacteriolysis, Competition etc.The competition that Trichoderma is strong and survival ability play an important role in the process of suppression pathogen especially soil-borne disease, are a kind of biocontrol microorganisms having very much value of exploiting and utilizing.Having developed registration in the world has a lot of Trichoderma product at present, but the Trichoderma determined due to the biocontrol mechanism of Trichoderma can only grow surely at plant root table, method of application can only be used for the premorbid prevention of crop and once morbidity, pathogen enters in plant corpus and only can't resolve by Trichoderma, especially all the more so to the biological control of soil biography class disease, this touching of rule also constraining trichoderma biopesticide is to a great extent produced and application.
What the related documents of bacillus subtilis was recorded is, and a class is prevalent in soil, microorganism in plant surface and body, there is the advantage of the poor environments such as opposing high temperature, low temperature, drying and nutritional deficiency, some bacterial strain can conduct in plant body surely to be grown and plays growth-promoting functions, main antifungal mechanism suppresses the growth of pathogen, and can surely grow in plant cell gap or cell, set up Harmonious existence relation with host plant.Surely grow the endogenetic bacteria in host, have sufficient source of nutrition, be not subject to extraneous adverse circumstance Effects of Factors, compared with other rhizosphere, phyllosphere biocontrol microorganisms, stable living environment is more conducive to the effect that it plays biological and ecological methods to prevent plant disease, pests, and erosion.
The multiple biocontrol bacterial strain compound use with different biocontrol mechanism can be significantly improved control efficiency be confirmed.As saccharomycete and bacterium compound use effectively can prevent and treat eliminating vegetable botrytis, actinomycetes mixing bacillus control strawberry Verticillium wilt etc.The Trichoderma of rhizosphere colonization and the vascular bundle endogenetic bacteria grown decided at the higher level but not officially announced are re-dubbed multifunctional bio agricultural chemicals, and there is not been reported.
Find through the experimental study for many years of this laboratory, bacillus subtilis (
bacillus subtilis) the mycelial normal growth of many plant pathogenic fungis or direct killing pathogenic bacteria can be suppressed, soil fungi being comprised to the microbial various crop fusarium wilt of the cause of diseases such as sickle-like bacteria, pythium spp, rhizoctonia, damping off, damping off etc. has control action.Carry out analysis by methods such as the recovery of resistance to rifampin Bacillus subtillis mutant strain mark and DAS-ELISA to the colonization dynamics of Bacillus subtillis in melon plant body to find, Bacillus subtillis can grow and have system running ability crop plant tissue is decided at the higher level but not officially announced, namely multiply in vivo along with the growth of plant, its metabolite to many germ spore germinations and mycelial growth inhibited, show to there is antibiotic agents in metabolite, the heat endurance that its metabolite tool is stronger and ph stability and the patience stronger to ultraviolet.Utilize Bio-control Trichoderma and endogenous bacillus subtilis to prevent and treat the different mode of action of soil-borne disease of crop and biocontrol mechanism both are worked in coordination with to use, by becoming the important means improving soil-borne disease of crop control efficiency, the deficiency of Trichoderma biological control effect instability can be solved.
Summary of the invention
The object of this invention is to provide a kind of multiple-effect biocontrol fungicide preparation method for preventing and treating crops soil-borne disease and application thereof, with other invent unlike, this preparation overcomes the deficiency of prior art, according to the occurrence characteristic of soil-borne disease, by isolating Antagonistic Trichoderma and Bacillus subtillis-Endophytic antagonistic bacteria from crop root table, be re-dubbed microorganism formulation.
The present invention finds the competitive violently survival ability of most of Antagonistic Trichoderma and the hyperparasitism to pathogen in soil by large quantity research for many years, and most of antagonism Bacillus subtillis can grow and have system running ability crop plant tissue is decided at the higher level but not officially announced, namely multiply in vivo along with the growth of plant, its metabolite to many germ spore germinations and mycelial growth inhibited, both utilizations are used the crop different ecological site mode of action simultaneously has synergistic function, control efficiency can be made to reach more than 75%
The object of the invention is to realize by following content:
The biocontrol microorganisms adopted is taken from the Trichoderma of crop root table and Nei Sheng Bacillus subtillis is female microbial inoculum, female microbial inoculum is made by being transferred in PDA test tube slant after its respectively purifying, in triangular flask, shaken cultivation is second class inoculum, make it produce resistance trichoderma harzianum chlamydospore and Bacillus subtillis spore by liquid fermentation again, after being mixed with carrier by the tunning being loaded with a large amount of spore, make multiple-effect compound biological prevention and control agent.
Described multiple-effect compound biocontrol fungicide is prepared from as follows:
A: the preparation of the female bacterial classification in inclined-plane:
Be taken as 10 grams, thing root table soil, add sterile water and be diluted to 1000 milliliters of dilutions, by dilution and PDA medium by 1: 9 part by weight mix after pour in culture dish and make flat board, cultivating 48 hours at 26 DEG C, obtaining Trichoderma through being separated; The female bacterial classification in Nei Zuo inclined-plane, test tube slant is transferred to after purifying;
PDA medium: potato 200 grams, glucose 20 grams, 20 grams, agar, rose-bengal 0.05 gram, streptomycin sulphate 0.03 gram, 1000 milliliters, water;
Get crop root 10 grams, with 0.1% mercuric chloride solution surface sterilization after 2-4 minute, again with aseptic water washing 3-5 rear grinding, add sterile water and be diluted to 1000 milliliters of dilutions, by dilution and PDA medium by 1: 9 part by weight mix after pour in culture dish and make flat board, cultivate 48 hours at 26 DEG C, through being separated raw Bacillus subtillis in acquisition; The female bacterial classification in Nei Zuo inclined-plane, test tube slant is transferred to after purifying;
PDA medium: potato 200 grams, glucose 20 grams, 20 grams, agar, 1000 milliliters, water;
B: the preparation of second class inoculum:
Second class inoculum culture fluid is brown sugar 20-30 gram, yeast 2-5 gram, and adjust pH is 7-9, adds water to 1000 milliliters.Be divided in 250 milliliters of triangular flasks, liquid amount is 100 milliliters, sterilizing 30 minutes at 121 DEG C, the female bacterial classification in access inclined-plane, and shaking speed is 150-200 rev/min, fermentation temperature 28-30 DEG C, and fermentation time is 2-3 days;
C: liquid fermentation:
Preparation trichoderma harzianum chlamydospore:
Culture medium prescription is: potato 150-200 gram, glucose 15-20 gram, potassium dihydrogen phosphate 1-2 gram, magnesium sulfate 1-2 gram, calcium sulphate 1-2 gram, tryptophan 3-5 gram, fructose 2-4 gram, 1000 milliliters, water: adjust pH is 5-6.5, sterilizing 30-40 minute at 121 DEG C, meets the 2-4% that bacterium amount is medium weight, fermentation temperature is 23-25 DEG C, mixing speed is 120-150rpm, and fermentation time 3-5 days obtains trichoderma harzianum chlamydospore suspension;
Preparation Bacillus subtillis spore:
Culture medium prescription is: beef extract 2-4 gram g, peptone 5-10 gram, yeast extract 2-5 gram, sucrose 10-20 gram, potassium dihydrogen phosphate 1-2 gram, magnesium sulfate 1-2 gram, 1000 milliliters, water; Tune pH value is 7-8, sterilizing 30-40 minute at 121 DEG C, meets the 2-4% that bacterium amount is medium weight, and fermentation temperature is 28-30 DEG C, and mixing speed is 120-150rpm, and fermentation time 3-5 days obtains Bacillus subtillis spore suspension;
D: the preparation of biocontrol fungicide: by gained trichoderma harzianum chlamydospore suspension by weight proportion 8-10%, interior raw Bacillus subtillis spore suspension by weight proportion 10-12% and precipitated calcium carbonate by weight proportion 20-30% and diatomite by weight proportion 50-60% make biological prevention and control agent after fully mixing.
By multiple-effect compound biocontrol fungicide with soil or water by weight proportion 1:100-300 mix, adopt bed surface mix soil spread manuer in holes or fill with root method prevent and treat soil-borne disease of crop.
The invention has the advantages that:
1. trichoderma viride chlamydospore of the present invention is as a kind of spore shape of degeneration-resistant type, and the same with the gemma of endophytic Bacillus have the ability of resisting the poor environments such as high temperature, low temperature, drying and nutritional deficiency.Two spore sterilizing kieselguhr adsorptions are made preparation, and preservation 2 years under room temperature, spore still can be sprouted.Be attributed to nature by putting again after manually cultivating, and make it to multiply further in natural microorganisms fauna, be formed with the benign cycle of beneficial microorganism in soil, be conducive to the natural equilibrium keeping root-zone soil, reach the lasting control to disease.
2. endophytic Bacillus reproduction speed is exceedingly fast, form biological barrier at crop root table and stop pathogen infection, the bacillus being simultaneously attached to root system restrained effectively the sprouting of germ spore at soil and root table secretion antibiotics, the bacillus entered in plant body suppresses by conduction and kills germ.
3. Trichoderma can grow surely at root table, Biocontrol Effect is played by competing nutrition with pathogen and capture ecological site etc., bacillus subtilis can operate in plant corpus, suppresses and kills the pathogen entered in plant body, and two class biocontrol microorganisms define the dual ecological protective screen respectively in root table and root.
4. cause the soil-borne disease pathogen of crop to be all generally invade from seedling stage, breed in host's vascular bundle, pathogen is stoped to be first barrier to the intrusion of root system in seedling stage by wooden mould biocontrol microorganisms, to enter bacillus subtilis in plant corpus for second barrier, significant to control later stage disease.
The present invention is applicable to most of trichoderma strain and bacillus, and method is simple, is suitable for large-scale industrialized production.
Embodiment
Below in conjunction with specific embodiment, technical scheme of the present invention is further described.
embodiment 1
A: the preparation of the female bacterial classification in inclined-plane:
Be taken as 10 grams, thing root table soil, add sterile water and be diluted to 1000 milliliters of dilutions, by dilution and PDA medium by 1: 9 part by weight mix after pour in culture dish and make flat board, cultivating 48 hours at 26 DEG C, obtaining Trichoderma through being separated; The female bacterial classification in Nei Zuo inclined-plane, test tube slant is transferred to after purifying;
PDA medium: potato 200 grams, glucose 20 grams, 20 grams, agar, rose-bengal 0.05 gram, streptomycin sulphate 0.03 gram, 1000 milliliters, water;
Get crop root 10 grams, with 0.1% mercuric chloride solution surface sterilization after 2 minutes, grind with after aseptic water washing 3 times again, add sterile water and be diluted to 1000 milliliters of dilutions, by dilution and PDA medium: pour in culture dish after the part by weight by 1: 9 mixes and make flat board, cultivate 48 hours at 26 DEG C, through being separated raw Bacillus subtillis in acquisition; The female bacterial classification in Nei Zuo inclined-plane, test tube slant is transferred to after purifying;
PDA medium: potato 200 grams, glucose 20 grams, 20 grams, agar, 1000 milliliters, water;
B: the preparation of second class inoculum:
Second class inoculum culture fluid is 20 grams, brown sugar, 5 grams, yeast, and adjust pH is 7, adds water to 1000 milliliters; Be divided in 250 milliliters of triangular flasks, liquid amount is 100 milliliters, sterilizing 30 minutes at 121 DEG C, the female bacterial classification in access inclined-plane, and shaking speed is 150 revs/min, fermentation temperature 28 DEG C, and fermentation time is 3 days;
C: liquid fermentation:
Preparation trichoderma harzianum chlamydospore: culture medium prescription is potato 150 grams, glucose 15 grams, potassium dihydrogen phosphate 2 grams, 2 grams, magnesium sulfate, 2 grams, calcium sulphate, tryptophan 5 grams, 4 grams, fructose, 1000 milliliters, water; Adjust pH is sterilizing 30 minutes at 5,121 DEG C, connect that bacterium amount is medium weight 4%, and fermentation temperature is 23 DEG C, and mixing speed is 150rpm, and fermentation time 5 days, obtains trichoderma harzianum chlamydospore suspension;
Preparation Bacillus subtillis spore: culture medium prescription is beef extract 2 grams, peptone 10 grams, yeast extract 2 grams, sucrose 10 grams, potassium dihydrogen phosphate 1 gram, 1 gram, magnesium sulfate, 1000 milliliters, water; PH value is adjusted to be sterilizing 30 minutes at 7,121 DEG C, connect that bacterium amount is medium weight 2%, fermentation temperature is 28 DEG C, and mixing speed is 120rpm, and fermentation time 5 days, obtains Bacillus subtillis spore suspension;
D: the preparation of biocontrol fungicide: by gained trichoderma harzianum chlamydospore suspension by weight proportion 8%, interior raw Bacillus subtillis spore suspension by weight proportion 12% with precipitated calcium carbonate by weight proportion 20%, diatomite by weight proportion 60% fully mix after make biological prevention and control agent.
Show for melon crop soil-borne disease controlling experiment result: when cucumber is sowed, with this preparation and soil by weight proportion 1:100 mix, dress feeding block seedlings raising, bear fruit early stage, again with this preparation and water by weight proportion 1:300 mix, fill with root once, control efficiency reaches 82.12%.When watermelon is sowed, with this preparation and soil by weight proportion 1:100 mix, dress feeding block seedlings raising, bears fruit early stage, then with this preparation and water by weight proportion 1:300 mix, filling root once, control efficiency 76.44%.
embodiment 2
A: the preparation of the female bacterial classification in inclined-plane:
Be taken as 10 grams, thing root table soil, add sterile water and be diluted to 1000 milliliters of dilutions, by dilution and PDA medium by 1: 9 part by weight mix after pour in culture dish and make flat board, cultivating 48 hours at 26 DEG C, obtaining Trichoderma through being separated; The female bacterial classification in Nei Zuo inclined-plane, test tube slant is transferred to after purifying.
PDA medium: potato 200 grams, glucose 20 grams, 20 grams, agar, rose-bengal 0.05 gram, streptomycin sulphate 0.03 gram, 1000 milliliters, water;
Get crop root 10 grams, with 0.1% mercuric chloride solution surface sterilization after 3 minutes, grind with after aseptic water washing 4 times again, add sterile water and be diluted to 1000 milliliters of dilutions, by dilution and PDA medium by 1: 9 part by weight mix after pour in culture dish and make flat board, cultivate 48 hours at 26 DEG C, through being separated raw Bacillus subtillis in acquisition; The female bacterial classification in Nei Zuo inclined-plane, test tube slant is transferred to after purifying.
PDA medium: potato 200 grams, glucose 20 grams, 20 grams, agar, 1000 milliliters, water;
B: the preparation of second class inoculum:
Second class inoculum culture fluid is 25 grams, brown sugar, 3 grams, yeast, and adjust pH is 8, adds water to 1000 milliliters; Be divided in 250 milliliters of triangular flasks, liquid amount is 100 milliliters, sterilizing 30 minutes at 121 DEG C, the female bacterial classification in access inclined-plane, and shaking speed is 180 revs/min, fermentation temperature 29 DEG C, and fermentation time is 3 days;
C: liquid fermentation:
Preparation trichoderma harzianum chlamydospore:
Culture medium prescription is: potato 180 grams, glucose 18 grams, potassium dihydrogen phosphate 1.5 grams, 1.5 grams, magnesium sulfate, 1.5 grams, calcium sulphate, tryptophan 4 grams, 3 grams, fructose, 1000 milliliters, water; Adjust pH is sterilizing 30 minutes at 6,121 DEG C, connect that bacterium amount is medium weight 3%, and fermentation temperature is 24 DEG C, and mixing speed is 130rpm, and fermentation time 4 days, obtains trichoderma harzianum chlamydospore suspension;
Preparation Bacillus subtillis spore: culture medium prescription is: beef extract 3 grams, peptone 8 grams, yeast extract 4 grams, sucrose 15 grams, potassium dihydrogen phosphate 1.5 grams, 1.5 grams, magnesium sulfate, 1000 milliliters, water; PH value is adjusted to be sterilizing 35 minutes at 7.5,121 DEG C, connect that bacterium amount is medium weight 3.5%, fermentation temperature is 29 DEG C, and mixing speed is 140rpm, fermentation time 4 days, obtains interior raw Bacillus subtillis spore suspension;
D: the preparation of biocontrol fungicide: by gained trichoderma harzianum chlamydospore suspension by weight proportion 9%, interior raw Bacillus subtillis spore suspension by weight proportion 11% with precipitated calcium carbonate by weight proportion 25% diatomite by weight proportion 55% fully mix after make biological prevention and control agent.
When muskmelon is sowed, with this preparation and soil by weight proportion 1:200 be mixed in seedling bed, bear fruit early stage, then with this preparation and water by weight proportion 1:300 mix, filling root once, prevents and treats soil-borne disease effect 77.06%.When capsicum is sowed, with this preparation and soil by weight proportion 1:300 be mixed in seedling bed, bear fruit early stage, then with this preparation and water by weight proportion 1:300 mix, filling root once, anti-soil-borne disease effect 75.36%.
embodiment 3
A: the preparation of the female bacterial classification in inclined-plane:
Be taken as 10 grams, thing root table soil, add sterile water and be diluted to 1000 milliliters of dilutions, by dilution and PDA medium by 1: 9 part by weight mix after pour in culture dish and make flat board, cultivating 48 hours at 26 DEG C, obtaining Trichoderma through being separated; The female bacterial classification in Nei Zuo inclined-plane, test tube slant is transferred to after purifying;
PDA medium: potato 200 grams, glucose 20 grams, 20 grams, agar, rose-bengal 0.05 gram, streptomycin sulphate 0.03 gram, 1000 milliliters, water;
Get crop root 10 grams, with 0.1% mercuric chloride solution surface sterilization after 4 minutes, grind with after aseptic water washing 5 times again, add sterile water and be diluted to 1000 milliliters of dilutions, by dilution and PDA medium by 1: 9 part by weight mix after pour in culture dish and make flat board, cultivate 48 hours at 26 DEG C, through being separated raw Bacillus subtillis in acquisition; The female bacterial classification in Nei Zuo inclined-plane, test tube slant is transferred to after purifying;
PDA medium: potato 200 grams, glucose 20 grams, 20 grams, agar, 1000 milliliters, water;
B: the preparation of second class inoculum:
Second class inoculum culture fluid is 30 grams, brown sugar, 2 grams, yeast, and adjust pH is 9, adds water to 1000 milliliters.Be divided in 250 milliliters of triangular flasks, liquid amount is 100 milliliters, sterilizing 30 minutes at 121 DEG C, the female bacterial classification in access inclined-plane, and shaking speed is 200 revs/min, fermentation temperature 30 DEG C, and fermentation time is 2 days;
C: liquid fermentation:
Preparation trichoderma harzianum chlamydospore: culture medium prescription is: potato 200 grams, glucose 20 grams, potassium dihydrogen phosphate 1 gram, 1 gram, magnesium sulfate, 1 gram, calcium sulphate, tryptophan 3 grams, 2 grams, fructose, 1000 milliliters, water; Adjust pH is sterilizing 40 minutes at 6.5,121 DEG C, connect that bacterium amount is medium weight 2%, and fermentation temperature is 25 DEG C, and mixing speed is 120rpm, and fermentation time 3 days, obtains trichoderma harzianum chlamydospore suspension;
Preparation Bacillus subtillis spore: culture medium prescription is: beef extract 4 grams, peptone 5 grams, yeast extract 5 grams, sucrose 20 grams, potassium dihydrogen phosphate 2 grams, 2 grams, magnesium sulfate, 1000 milliliters, water; PH value is adjusted to be sterilizing 40 minutes at 8,121 DEG C, connect that bacterium amount is medium weight 4%, fermentation temperature is 230 DEG C, and mixing speed is 150rpm, fermentation time 3 days, obtains interior raw Bacillus subtillis spore suspension;
D: the preparation of biocontrol fungicide: by gained trichoderma harzianum chlamydospore suspension by weight proportion 10%, interior raw Bacillus subtillis spore suspension by weight proportion 10% with precipitated calcium carbonate by weight proportion 30% and diatomite by weight proportion 50% fully mix after make biological prevention and control agent.
When corn seeding, with this preparation and soil by weight proportion 1:100 mix, spread manuer in holes, control of maize stem rot effect reaches 78.36%.
When ginseng transplantation of seedlings in 2 years, be first evenly mixed into ginseng bed plow layer with 200 grams, the every curtain of this preparation (10 square metres), Zhan Yehou with this preparation and water by weight proportion 1:300 mix, fill with root once, control ginseng root disease effect is 77.97%.
Bibliography:
1. the biological safety of Bio-control Trichoderma detects. Scientia Agricultura Sinica, 2006 (4)
2. the influence research of nutrition elements on biocontrol efficiency of Trichoderma against melon wilt. plant protection journal, 2004 (4)
3. trichoderma viride T23 is on cucumber fusarium axysporum control efficiency and Defense Enzyme Activities impact. Plant Pathology, 2005 (2)
4. bacillus subtilis (Bacillus subtilis) B6 is to the Biocontrol Effect of Muskmelon Fusarium wilt. Journal of Fruit Science 2008 (6)
Claims (2)
1. a preparation method for multiple-effect compound biocontrol fungicide, is characterized in that comprising following concrete steps:
(1) preparation of the female bacterial classification in inclined-plane:
Be taken as 10 grams, thing root table soil, add sterile water and be diluted to 1000 milliliters of dilutions, by dilution and PDA medium by 1: 9 part by weight mix after pour in culture dish and make flat board, cultivating 48 hours at 26 DEG C, obtaining Trichoderma through being separated; The female bacterial classification in Nei Zuo inclined-plane, test tube slant is transferred to after purifying;
PDA medium: potato 200 grams, glucose 20 grams, 20 grams, agar, rose-bengal 0.05 gram, streptomycin sulphate 0.03 gram, 1000 milliliters, water;
Get crop root 10 grams, with 0.1% mercuric chloride solution surface sterilization after 2-4 minute, again with aseptic water washing 3-5 rear grinding, add sterile water and be diluted to 1000 milliliters of dilutions, by dilution and PDA medium by 1: 9 part by weight mix after pour in culture dish and make flat board, cultivating 48 hours at 26 DEG C, obtaining endophytic Bacillus subtilis through being separated; The female bacterial classification in Nei Zuo inclined-plane, test tube slant is transferred to after purifying;
PDA medium: potato 200 grams, glucose 20 grams, 20 grams, agar, 1000 milliliters, water;
(2) preparation of second class inoculum:
Second class inoculum culture fluid is brown sugar 20-30 gram, and yeast 2-5 gram adds water to 1000 milliliters, adjusts pH value to be 7-9; Be divided in 250 milliliters of triangular flasks, liquid amount is 100 milliliters, sterilizing 30 minutes at 121 DEG C, the female bacterial classification in access inclined-plane, and shaking speed is 150-200 rev/min, fermentation temperature 28-30 DEG C, and fermentation time is 2-3 days;
(3) liquid fermentation:
Preparation trichoderma harzianum chlamydospore:
Culture medium prescription is: potato 150-200 grams, glucose 15-20 grams, potassium dihydrogen phosphate 1-2 gram, 1-2 gram, magnesium sulfate, 1-2 gram, calcium sulphate, tryptophan 3-5 grams, 2-4 grams, fructose, 1000 milliliters, water; PH value is adjusted to be sterilizing 30-40 minute at 5-6.5,121 DEG C, connect that bacterium amount is medium weight 2-4%, fermentation temperature is 23-25 DEG C, and mixing speed is 120-150rpm, and fermentation time 3-5 days, obtains trichoderma harzianum chlamydospore suspension;
Prepare Bacillus globigii spores:
Culture medium prescription is: beef extract 2-4 gram, peptone 5-10 gram, yeast extract 2-5 gram, sucrose 10-20 gram, potassium dihydrogen phosphate 1-2 gram, 1-2 gram, magnesium sulfate, 1000 milliliters, water; PH value is adjusted to be 7-8, sterilizing 30-40 minute at 121 DEG C, connect that bacterium amount is medium weight 2-4%, fermentation temperature is 28-30 DEG C, and mixing speed is 120-150rpm, and fermentation time 3-5 days, obtains Bacillus globigii spores suspension;
(4) preparation of biocontrol fungicide: by gained trichoderma harzianum chlamydospore suspension by weight proportion 8-10%, Bacillus globigii spores suspension by weight proportion 10-12% and precipitated calcium carbonate by weight proportion 20-30% and diatomite by weight proportion 50-60% fully mix after make biological prevention and control agent.
2. an application for the multiple-effect compound biocontrol fungicide as described in claim 1, is characterized in that: by multiple-effect composite bacteria agent capable by weight proportion 100-300 doubly adopt bed surface to mix soil spread manuer in holes or fill with root method control soil-borne disease of crop.
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| CN109628323B (en) * | 2018-12-28 | 2019-09-27 | 云南星耀生物制品有限公司 | Trichoderma Viride method and its water dispersible granules |
| CN111838190A (en) * | 2019-04-24 | 2020-10-30 | 山东大学苏州研究院 | Biocontrol microbial inoculum for preventing and treating stem base rot and gummosis as well as preparation method and application thereof |
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