CN108293480A - The method for preventing graw mold of tomato using biocontrol agent - Google Patents
The method for preventing graw mold of tomato using biocontrol agent Download PDFInfo
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- CN108293480A CN108293480A CN201810249712.5A CN201810249712A CN108293480A CN 108293480 A CN108293480 A CN 108293480A CN 201810249712 A CN201810249712 A CN 201810249712A CN 108293480 A CN108293480 A CN 108293480A
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- trichosporon cutaneum
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- 235000007688 Lycopersicon esculentum Nutrition 0.000 title claims abstract description 38
- 238000000034 method Methods 0.000 title claims abstract description 26
- 239000012681 biocontrol agent Substances 0.000 title claims abstract description 23
- 240000003768 Solanum lycopersicum Species 0.000 title description 32
- 230000002265 prevention Effects 0.000 claims abstract description 15
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- 230000004913 activation Effects 0.000 claims abstract description 5
- 241000227653 Lycopersicon Species 0.000 claims abstract 6
- 241000894006 Bacteria Species 0.000 claims description 31
- 241000223259 Trichoderma Species 0.000 claims description 25
- 238000000855 fermentation Methods 0.000 claims description 25
- 230000004151 fermentation Effects 0.000 claims description 25
- 241000589614 Pseudomonas stutzeri Species 0.000 claims description 22
- 241000223233 Cutaneotrichosporon cutaneum Species 0.000 claims description 21
- 239000007788 liquid Substances 0.000 claims description 19
- 239000002054 inoculum Substances 0.000 claims description 18
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- 239000000843 powder Substances 0.000 claims description 15
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- 239000002609 medium Substances 0.000 claims description 12
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- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 8
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 claims description 8
- 238000011081 inoculation Methods 0.000 claims description 8
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- 229910000359 iron(II) sulfate Inorganic materials 0.000 claims description 7
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- 238000009777 vacuum freeze-drying Methods 0.000 claims description 4
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- 241000123650 Botrytis cinerea Species 0.000 description 4
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- 239000001888 Peptone Substances 0.000 description 4
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- 235000013399 edible fruits Nutrition 0.000 description 2
- 229910052739 hydrogen Inorganic materials 0.000 description 2
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- 201000001880 Sexual dysfunction Diseases 0.000 description 1
- 102000013275 Somatomedins Human genes 0.000 description 1
- 241000223260 Trichoderma harzianum Species 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
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- NCYVXEGFNDZQCU-UHFFFAOYSA-N nikethamide Chemical compound CCN(CC)C(=O)C1=CC=CN=C1 NCYVXEGFNDZQCU-UHFFFAOYSA-N 0.000 description 1
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- DPJRMOMPQZCRJU-UHFFFAOYSA-M thiamine hydrochloride Chemical compound Cl.[Cl-].CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N DPJRMOMPQZCRJU-UHFFFAOYSA-M 0.000 description 1
Classifications
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G7/00—Botany in general
- A01G7/06—Treatment of growing trees or plants, e.g. for preventing decay of wood, for tingeing flowers or wood, for prolonging the life of plants
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/10—Animals; Substances produced thereby or obtained therefrom
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/30—Microbial fungi; Substances produced thereby or obtained therefrom
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Abstract
The invention belongs to biotechnology, the method for preventing graw mold of tomato using biocontrol agent is disclosed comprising following steps：Biocontrol agent is added in the water of 50 100 times of weight, is stirred evenly, is placed in 25 30 DEG C of 3 5h of activation, be then sprayed onto on tomato according to the amount of every plant of 10 20ml of sprinkling, repeat to spray after 1 week primary.The present invention carries out field control using biological prevention and control agent to graw mold of tomato, and completely without a series of problems caused by chemical prevention, and control effect is good.
Description
Technical field
The invention belongs to biotechnologies, and in particular to utilize the method for biocontrol agent prevention graw mold of tomato.
Background technology
Graw mold of tomato is one of the important disease in tomato production, is produced to tomato in greenhouse and constitutes great threat.Children
Seedling stage and Adult plant can fall ill, and flowers and fruits base of leaf can be aggrieved, especially aggrieved most heavy with Chinese olive, generally result in the tomato underproduction 30% with
On, the yield and quality of tomato is influenced very big.There are generation in country various regions at present, have become the limitation of tomato facility cultivation
Sexual dysfunction.Currently, the anti-source due to not yet finding gray mold, breeding for disease resistance are difficult to carry out, the prevention of graw mold of tomato still relies upon
Chemical agent is prevented.However, long-term, continuous use, which makes pathogen gradually produce drug resistance, affects control effect, together
Shi Zaocheng environmental pollutions also result in the pollution by pesticides of fruit.It thus finds and screening has strong inhibition to botrytis cinerea
The microbial strains of effect are the Main ways of graw mold of tomato prevention.Chemical pesticide control plant is substituted using biological pesticide
Though disease can avoid the generation of drug resistance pathogenic strains, have that prevention mechanism is unclear, preventive effect is slower, effect is unstable equal to be lacked
Point.
Biological pesticide is mainly biocontrol agent, as a kind of emerging control measure, compared with other methods, has peace
Entirely, feature effectively, lasting is especially that of avoiding a series of problems that chemical prevention is brought.Biocontrol agent control method is to adopt
Grey mold is killed with microorganism, has begun agriculturally promoting the use of at present, relatively common be in before the onset or
Initial stage is sprayed using 300,000,000 CFU/ grams of trichoderma harzianums 300, is watered spraying, is sprayed every 5-7 days once, can be to a certain extent
Prevent the generation of gray mold.Document " chemical agent of prevention and control graw mold of tomato and biocontrol bacterial strain screening study, Niu Zhenfu, northeast
Agricultural sciences 2016 " is tested graw mold of tomato control effect in field using chemical agent and biocontrol microorganisms；Its
In, chemical agent is to the preventive effect of graw mold of tomato between 60-70%；Preventive effect model of the biocontrol bacterial strain zymotic fluid to graw mold of tomato
It encloses for 45.73-61.09%, highest preventive effect is trichoderma.
In recent years, biological control gray mold method is increasingly becoming the hot spot of research, and a large amount of antagonistic microbe is screened out
Gray mold prevented to be used.Often there is the defects of control effect is low, drug effect is slow in single bacterial strain, at present
Through there is multiple biological pesticide companies to begin one's study compound biological prevention and control agent.Compound biological prevention and control agent generally refers to include two or more micro-
The preparation of biology, selects the bacterial strain in compound biological prevention and control agent more crucial and difficult point；It is anti-between bacterial strain if selection is careless
And the reaction of mutual antagonism can be played.It is particularly urgent to develop a kind of method preventing graw mold of tomato using compound biocontrol fungicide
It cuts.
Invention content
In order to overcome in the prior art chemical prevention and control method easy to produce antagonism and pollution, single biocontrol agent control effect
The defect of difference, the present invention provides the methods for preventing graw mold of tomato using biocontrol agent.
The present invention is achieved by the following technical solution：
The method for preventing graw mold of tomato using biocontrol agent comprising following steps：
Biocontrol agent is added in the water of 50-100 times of weight, is stirred evenly, be placed in 25-30 DEG C activation 3-5h, then according to
The amount of every plant of sprinkling 10-20ml is sprayed onto on tomato, repeats to spray after 1 week primary.
Preferably,
The tomato is in gray mold early stage.
Specifically,
The biocontrol agent is prepared according to following technique：Koning trichoderma-trichosporon cutaneum mixed fermentation liquid and Amur is false single
Born of the same parents' bacteria culture fluid is according to 3:2 volume ratio is uniformly mixed, then vacuum freeze drying be made bacterium powder to get.
Preferably,
The Pseudomonas stutzeri is ATCC 17588；The koning trichoderma is ATCC 66766；The trichosporon cutaneum is ATCC
201110。
Specifically,
The preparation method of the koning trichoderma-trichosporon cutaneum mixed fermentation liquid includes the following steps：
1）Trichosporon cutaneum streak inoculation is cultivated on YPDA culture mediums, obtains single bacterium colony；Picking single bacterium colony is inoculated into YPD liquid
It is cultivated on culture medium, 30 DEG C of cultures for 24 hours, obtain trichosporon cutaneum seed liquor, for use；
2）Koning trichoderma streak inoculation is cultivated in PDA culture medium, obtains single bacterium colony；Picking single bacterium colony is inoculated into PDA liquid
It is cultivated on culture medium, 30 DEG C, 200rpm shaking table culture 36h obtain koning trichoderma seed liquor, for use；
3）First koning trichoderma seed liquor is linked into according to 10% inoculum concentration in fermentation medium, 30 DEG C cultivate 6 hours, then
Trichosporon cutaneum seed liquor is linked into fermentation medium according to the inoculum concentration of 5-7%, continues to cultivate 18 hours with 30 DEG C, obtains health
Family name's trichoderma-trichosporon cutaneum mixed fermentation liquid.
Preferably,
The formula of the fermentation medium is：Corn stalk powder 16g/L, dregs of beans 10g/L, ammonium chloride 5g/L, dipotassium hydrogen phosphate 2g/
L, potassium dihydrogen phosphate 1g/L, sodium chloride 0.5g/L, magnesium sulfate 0.1g/L, ferrous sulfate 0.01g/L.
Specifically,
The preparation method of the Pseudomonas stutzeri culture solution includes the following steps：
By Pseudomonas stutzeri in the flat lining out cultures of LB, single bacterium colony is obtained；Picking single bacterium colony is respectively connected to LB Liquid Cultures
It is cultivated on base, 28 DEG C, seed liquor is made for 24 hours in the shaking table culture of 200r/min；Then it is linked into expansion according to 10% inoculum concentration
In culture medium, 28 DEG C of cultures for 24 hours, obtain Pseudomonas stutzeri culture solution.
Preferably,
It is described expand culture medium formula be：Cornstarch 12g/L, yeast extract 10g/L, sodium chloride 5g/L, urea 2g/L,
Potassium dihydrogen phosphate 0.1g/L, dipotassium hydrogen phosphate 0.1g/L, ferrous sulfate 0.01g/L, manganese sulfate 0.01g/L.
Compared with prior art, the advantageous effect that the present invention obtains includes but is not limited to mainly the following aspects：
The present invention prevents graw mold of tomato using Biocontrol microorganism preparation, completely without a system caused by chemical prevention
Row problem, is conducive to the No-harmful apple orchard of crop, and peasant can not have to or reduce other prevention gray molds and improve resistance
With the measure of growth-promoting, this can not only reduce the heavy burdens for peasant planting, but also be conducive to the outlet of crop, improve product quality, increase
Add added value of product.
Without antagonism between trichosporon cutaneum and koning trichoderma, can mutual symbiosis, Trichoderma can eccrine fiber element enzyme,
It is the available carbon source of trichosporon cutaneum by straw powder cellulose decomposition, so that trichosporon cutaneum utilizes carbon source for growth, so adopting
With the mode for first accessing koning trichoderma, saccharomycete is accessed after 6 hours；Saccharomycete can also promote the growth of koning trichoderma in turn,
And the yield of enzyme of trichoderma is also greatly improved, and then improves the ability of antagonism botrytis cinerea；
Pseudomonas stutzeri, which can secrete antibacterial material, can secrete multiple active enzymes and plant stimulin, promote plant root with
And plant strain growth, enhance resistance so that plant generates antagonism to gray mold, additionally it is possible to pass through the competition of nutrition and space
To inhibit gray mold；Inventor in experiments it is found that, Pseudomonas stutzeri and Trichosporon Behrend-koning trichoderma have preferable association
Same-action can greatly improve the ability for killing gray mold, and possible cause is there is the relationship for being similar to symbiosis between them,
The metabolin of Pseudomonas stutzeri provides some necessary somatomedins for the breeding of Trichosporon Behrend, quick to promote
A large amount of antagonistic substances are bred and generated, then cause botrytis cinerea to be unable to normal growth by the effect of this substance；Specifically
The mechanism of action, inventor further in subsequent experiment can be furtherd investigate.In addition, this experiment is just for botrytis cinerea
Antagonism is studied, as the control effect to other pathogenic fungus diseases of soil biography also up for further studying.
Specific implementation mode
In order to make those skilled in the art better understand the technical solutions in the application, having below in conjunction with the application
The technical solution of the application is clearly and completely described in body embodiment, it is clear that described embodiment is only this Shen
Please a part of the embodiment, instead of all the embodiments.Based on the embodiment in the application, those of ordinary skill in the art are not having
There is the every other embodiment obtained under the premise of making creative work, should all belong to the scope of protection of the invention.
Embodiment 1
The method for preventing graw mold of tomato using biocontrol agent comprising following steps：
Biocontrol agent is added in the water of 100 times of weight, is stirred evenly, is placed in 30 DEG C of activation 3h, then sprayed according to every plant
The amount of 20ml is sprayed onto on tomato, repeats to spray after 1 week primary.
The tomato is in gray mold early stage；
The biocontrol agent is prepared according to following technique：
Trichosporon cutaneum streak inoculation is cultivated on YPDA culture mediums, obtains single bacterium colony；Picking single bacterium colony is inoculated into the training of YPD liquid
It supports and is cultivated on base, 30 DEG C of cultures for 24 hours, obtain seed liquor, for use；
The formula of YPD culture mediums：Glucose 20g, peptone 20g, yeast extraction leaching powder 10g, supplement distilled water to 1000mL；
The formula of YPDA culture mediums：Glucose 20g, peptone 20g, yeast extraction leaching powder 10g, agar 20g, supplement distilled water
To 1000mL；
Koning trichoderma streak inoculation is cultivated in PDA culture medium, obtains single bacterium colony；Picking single bacterium colony is inoculated into the training of PDA liquid
It supports and is cultivated on base, 30 DEG C, 200rpm shaking table culture 36h obtain seed liquor, for use；
First by koning trichoderma seed liquor according to 10% inoculum concentration（Inoculum density is 2 × 107CFU/ml）It is linked into fermentation medium
In, 30 DEG C are cultivated 6 hours, then by trichosporon cutaneum seed liquor according to the inoculum concentration of 5-7%（Inoculum density is 0.5 × 107CFU/
ml）It is linked into fermentation medium, continues to cultivate 18 hours with 30 DEG C, obtains koning trichoderma-trichosporon cutaneum mixed fermentation liquid；It is described
The group of fermentation medium is divided into：Corn stalk powder 16g/L, dregs of beans 10g/L, ammonium chloride 5g/L, dipotassium hydrogen phosphate 2g/L, di(2-ethylhexyl)phosphate
Hydrogen potassium 1g/L, sodium chloride 0.5g/L, magnesium sulfate 0.1g/L, ferrous sulfate 0.01g/L；
By Pseudomonas stutzeri in the flat lining out cultures of LB, single bacterium colony is obtained；Picking single bacterium colony is respectively connected to LB Liquid Cultures
It is cultivated on base, 28 DEG C, seed liquor is made for 24 hours in the shaking table culture of 200r/min；Then according to 10% inoculum concentration（Inoculum density is
5×107CFU/ml）It is linked into and expands in culture medium, 28 DEG C of cultures for 24 hours, obtain Pseudomonas stutzeri culture solution；The expansion training
Support base formula be：Cornstarch 12g/L, yeast extract 10g/L, sodium chloride 5g/L, urea 2g/L, potassium dihydrogen phosphate
0.1g/L, dipotassium hydrogen phosphate 0.1g/L, ferrous sulfate 0.01g/L, manganese sulfate 0.01g/L.
By koning trichoderma-trichosporon cutaneum mixed fermentation liquid and Pseudomonas stutzeri culture solution according to 3:2 volume ratio mixing
Uniformly, then vacuum freeze drying be made bacterium powder to get.
The Pseudomonas stutzeri is ATCC 17588；The koning trichoderma is ATCC 66766；The trichosporon cutaneum is
ATCC 201110。
Embodiment 2
The method for preventing graw mold of tomato using biocontrol agent comprising following steps：
Biocontrol agent is added in the water of 50 times of weight, is stirred evenly, is placed in 28 DEG C of activation 5h, then sprayed according to every plant
The amount of 10ml is sprayed onto on tomato, repeats to spray after 1 week primary.
The tomato is in gray mold early stage；
The biocontrol agent is prepared according to following technique：
Trichosporon cutaneum streak inoculation is cultivated on YPDA culture mediums, obtains single bacterium colony；Picking single bacterium colony is inoculated into the training of YPD liquid
It supports and is cultivated on base, 30 DEG C of cultures for 24 hours, obtain seed liquor, for use；
The formula of YPD culture mediums：Glucose 20g, peptone 20g, yeast extraction leaching powder 10g, supplement distilled water to 1000mL；
The formula of YPDA culture mediums：Glucose 20g, peptone 20g, yeast extraction leaching powder 10g, agar 20g, supplement distilled water
To 1000mL；
Koning trichoderma streak inoculation is cultivated in PDA culture medium, obtains single bacterium colony；Picking single bacterium colony is inoculated into the training of PDA liquid
It supports and is cultivated on base, 30 DEG C, 200rpm shaking table culture 36h obtain seed liquor, for use；
First by koning trichoderma seed liquor according to 10% inoculum concentration（Inoculum density is 2 × 107CFU/ml）It is linked into fermentation medium
In, 30 DEG C are cultivated 6 hours, then by trichosporon cutaneum seed liquor according to the inoculum concentration of 5-7%（Inoculum density is 0.5 × 107CFU/
ml）It is linked into fermentation medium, continues to cultivate 18 hours with 30 DEG C, obtains koning trichoderma-trichosporon cutaneum mixed fermentation liquid；It is described
The group of fermentation medium is divided into：Corn stalk powder 16g/L, dregs of beans 10g/L, ammonium chloride 5g/L, dipotassium hydrogen phosphate 2g/L, di(2-ethylhexyl)phosphate
Hydrogen potassium 1g/L, sodium chloride 0.5g/L, magnesium sulfate 0.1g/L, ferrous sulfate 0.01g/L；
By Pseudomonas stutzeri in the flat lining out cultures of LB, single bacterium colony is obtained；Picking single bacterium colony is respectively connected to LB Liquid Cultures
It is cultivated on base, 28 DEG C, seed liquor is made for 24 hours in the shaking table culture of 200r/min；Then according to 10% inoculum concentration（Inoculum density is
5×107CFU/ml）It is linked into and expands in culture medium, 28 DEG C of cultures for 24 hours, obtain Pseudomonas stutzeri culture solution；The expansion training
Support base formula be：Cornstarch 12g/L, yeast extract 10g/L, sodium chloride 5g/L, urea 2g/L, potassium dihydrogen phosphate
0.1g/L, dipotassium hydrogen phosphate 0.1g/L, ferrous sulfate 0.01g/L, manganese sulfate 0.01g/L.
By koning trichoderma-trichosporon cutaneum mixed fermentation liquid and Pseudomonas stutzeri culture solution according to 3:2 volume ratio mixing
Uniformly, then vacuum freeze drying be made bacterium powder to get.
The Pseudomonas stutzeri is ATCC 17588；The koning trichoderma is ATCC 66766；The trichosporon cutaneum is
ATCC 201110。
Embodiment 3
Tomato gray mould bacterium field control：
It is that Chongqing is No. 4 anti-that tomato, which selects kind, and in greenhouse succeeding planting, control seeding row spacing is 40 × 50cm.Tomato seedling
Implantation time is January 7, carries out sprinkling biocontrol agent in 3 days 2 months gray mold early stages, sprays biocontrol microorganisms again within 10 days 2 months
Agent is sprayed according to the amount of every plant of 20ml.
Group：Experimental group：Embodiment 1；Control group 1：Using koning trichoderma-trichosporon cutaneum mixed fermentation liquid, remaining is the same as real
Apply example 1；Control group 2：Using Pseudomonas stutzeri culture solution, remaining is the same as embodiment 1；Control group 3：By koning trichoderma and silk spore ferment
Mother stock drives capable fermentation into, and koning trichoderma zymotic fluid, trichosporon cutaneum zymotic fluid and Pseudomonas stutzeri culture solution are mixed with
Bacterium powder, remaining is the same as embodiment 1.
50 plants of sprinkling every time, setting repeat to test three times；After second of sprinkling 7 days, detection graw mold of tomato disease index,
Control effect（With reference to according to GB/T 17980.28-2000 standards）, field control effect is shown in Table 1：
Table 1
| Group | Disease index | Control effect % |
| Blank control group | 23.9 | -- |
| Control group 1 | 8.6 | 64.02 |
| Control group 2 | 11.3 | 52.72 |
| Control group 3 | 6.9 | 71.13 |
| Experimental group | 4.5 | 81.17 |
Conclusion：Experimental group and control group 1-3 can prevent graw mold of tomato it can be seen from upper table 1, wherein real
The control effect for testing group is best, can reach 80% or more, and control group 2 is minimum, and only 52.72%, it is seen then that the selection of bacterial strain is to anti-
Effect influences very big；Control group 3 is although equally use three kinds of bacterial strains, the difference with experimental group biocontrol agent to essentially consist in fermentation method
Difference, but the two preventive effect produces notable difference, experimental group improves 10 percentage points compared with control group 3, it may be possible to because, first
Access koning trichoderma can generate cellulase, be the available carbon source of trichosporon cutaneum by straw powder cellulose decomposition, to make
Trichosporon cutaneum is obtained to be grown using carbon source, and trichosporon cutaneum promotes the growth of koning trichoderma, the two synergy in turn
Obviously.
Although above having used general explanation, specific implementation mode and experiment, the present invention is described in detail,
On the basis of the present invention, it can be made some modifications or improvements, this will be apparent to those skilled in the art.Cause
This, these modifications or improvements, belong to the scope of protection of present invention without departing from theon the basis of the spirit of the present invention.
Claims (8)
1. utilizing the method for biocontrol agent prevention graw mold of tomato comprising following steps：
Biocontrol agent is added in the water of 50-100 times of weight, is stirred evenly, be placed in 25-30 DEG C activation 3-5h, then according to
The amount of every plant of sprinkling 10-20ml is sprayed onto on tomato, repeats to spray after 1 week primary.
2. according to the method described in claim 1, it is characterized in that, the tomato is in gray mold early stage.
3. method according to claim 1 or 2, which is characterized in that the biocontrol agent is prepared according to following technique：
By koning trichoderma-trichosporon cutaneum mixed fermentation liquid and Pseudomonas stutzeri culture solution according to 3:2 volume ratio is uniformly mixed, then
Vacuum freeze drying be made bacterium powder to get.
4. according to the method described in claim 3, it is characterized in that, the Pseudomonas stutzeri is ATCC 17588；The health
Family name's trichoderma is ATCC 66766；The trichosporon cutaneum is ATCC 201110.
5. according to the method described in claim 3, it is characterized in that, the system of the koning trichoderma-trichosporon cutaneum mixed fermentation liquid
Preparation Method includes the following steps：
1）Trichosporon cutaneum streak inoculation is cultivated on YPDA culture mediums, obtains single bacterium colony；Picking single bacterium colony is inoculated into YPD liquid
It is cultivated on culture medium, 30 DEG C of cultures for 24 hours, obtain trichosporon cutaneum seed liquor, for use；
2）Koning trichoderma streak inoculation is cultivated in PDA culture medium, obtains single bacterium colony；Picking single bacterium colony is inoculated into PDA liquid
It is cultivated on culture medium, 30 DEG C, 200rpm shaking table culture 36h obtain koning trichoderma seed liquor, for use；
3）First koning trichoderma seed liquor is linked into according to 10% inoculum concentration in fermentation medium, 30 DEG C cultivate 6 hours, then
Trichosporon cutaneum seed liquor is linked into fermentation medium according to the inoculum concentration of 5-7%, continues 30 DEG C and cultivates 18 hours, obtain Kang Shi
Trichoderma-trichosporon cutaneum mixed fermentation liquid.
6. according to the method described in claim 5, it is characterized in that, the formula of the fermentation medium is：Corn stalk powder
16g/L, dregs of beans 10g/L, ammonium chloride 5g/L, dipotassium hydrogen phosphate 2g/L, potassium dihydrogen phosphate 1g/L, sodium chloride 0.5g/L, magnesium sulfate
0.1g/L, ferrous sulfate 0.01g/L.
7. according to the method described in claim 3, it is characterized in that, the preparation method of the Pseudomonas stutzeri culture solution includes
Following steps：
By Pseudomonas stutzeri in the flat lining out cultures of LB, single bacterium colony is obtained；Picking single bacterium colony is respectively connected to LB Liquid Cultures
It is cultivated on base, 28 DEG C, seed liquor is made for 24 hours in the shaking table culture of 200r/min；Then it is linked into expansion according to 10% inoculum concentration
In culture medium, 28 DEG C of cultures for 24 hours, obtain Pseudomonas stutzeri culture solution.
8. the method according to the description of claim 7 is characterized in that the formula for expanding culture medium is：Cornstarch 12g/
L, yeast extract 10g/L, sodium chloride 5g/L, urea 2g/L, potassium dihydrogen phosphate 0.1g/L, dipotassium hydrogen phosphate 0.1g/L, sulfuric acid
Ferrous 0.01g/L, manganese sulfate 0.01g/L.
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