CN109169712A - A kind of compound biocontrol fungicide and its preparation method and application - Google Patents
A kind of compound biocontrol fungicide and its preparation method and application Download PDFInfo
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- CN109169712A CN109169712A CN201811010838.3A CN201811010838A CN109169712A CN 109169712 A CN109169712 A CN 109169712A CN 201811010838 A CN201811010838 A CN 201811010838A CN 109169712 A CN109169712 A CN 109169712A
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- fermentation
- paenibacillus polymyxa
- disease
- biocontrol fungicide
- bacillus brevis
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- 230000000443 biocontrol Effects 0.000 title claims abstract description 44
- 239000000417 fungicide Substances 0.000 title claims abstract description 39
- 230000000855 fungicidal effect Effects 0.000 title claims abstract description 38
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- 230000004151 fermentation Effects 0.000 claims abstract description 67
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims abstract description 43
- 201000010099 disease Diseases 0.000 claims abstract description 42
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- 241000194105 Paenibacillus polymyxa Species 0.000 claims abstract description 39
- 241000193764 Brevibacillus brevis Species 0.000 claims abstract description 34
- 235000002566 Capsicum Nutrition 0.000 claims abstract description 31
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- 239000006002 Pepper Substances 0.000 claims abstract description 4
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Classifications
-
- 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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- Life Sciences & Earth Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Health & Medical Sciences (AREA)
- Plant Pathology (AREA)
- Microbiology (AREA)
- Pest Control & Pesticides (AREA)
- Biotechnology (AREA)
- Virology (AREA)
- Agronomy & Crop Science (AREA)
- Dentistry (AREA)
- Wood Science & Technology (AREA)
- Environmental Sciences (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
The invention discloses the preparation methods and the application in terms of prevention and treatment capsicum epidemic disease, raising pepper disease resistance performance, increase yield of hot pepper of compound biocontrol fungicide made of the fermentation liquid of a kind of fermentation liquid as beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1 compounding and the compound biocontrol fungicide.Wherein, the compound biocontrol fungicide combines the biological and ecological methods to prevent plant disease, pests, and erosion characteristic of beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1, antimicrobial spectrum is widened, in conjunction with a variety of antimicrobial mechanisms, the microecological balance of soil is sufficiently coordinated, enhance bacterial strain to the adaptability of environment, facilitates the disease-resistant advantage of collaboration between long-acting performance bacterial strain, there is better biocontrol effect than Paenibacillus polymyxa DL1 single bacterium；In addition, the compound biocontrol fungicide can adjust the micro-ecological environment of plant rhizosphere soil, and by induction plant generation system disease resistance, plant disease-resistant ability can be improved, be finally reached growth-promoting, disease-resistant integrated control effect, realize the upgrading increasing both production and income of capsicum.
Description
Technical field
The present invention relates to a kind of compound biocontrol fungicides and its preparation method and application, and in particular to a kind of by beautiful short gemma
Compound biocontrol fungicide and compound life made of the fermentation liquid of bacillus NF2 and the fermentation liquid compounding of Paenibacillus polymyxa DL1
The preparation method of anti-microbial inoculum and the application in terms of prevention and treatment capsicum epidemic disease, raising pepper disease resistance performance, increase yield of hot pepper, belong to
Bacteria agent technical field.
Background technique
Capsicum epidemic disease is commonly called as " dead seedling disease ", is caused by Phytophthora capsici (Phytophthora capsici Leonian)
A kind of soil-borne disease, germ is overwintering in the soil or in invalid body in the form of egg spore, by means of wind, rain, pours water and other farming
Activity is propagated, and can produce new sporangium after the onset, is formed zoospore and is infected again.Capsicum is from seedling stage to Adult plant
It is infected, mostly shows symptom in flowering stage, stem, leaf and fruit can fall ill.Under the conditions of broad irrigation and high temperature and humidity
It is easy pandemic, often results in the capsicum underproduction, or even have no harvest, economic loss is huge, it has also become capsicum ruins in world wide
Venereal disease of going out evil.
Currently, production is upper mainly using disease-resistant variety breeding, tillage control measure improvement, physical disinfection and application chemical agent
The methods of carry out the prevention and treatment of disease.But all there are more or less some problems in these methods, such as:
(1) disease-resistant variety breeding: the resistance germ plasm resource of capsicum epidemic disease is relatively deficient, and due to pathogen grade of fit
Change the degeneration that will lead to disease-resistant variety or even disease resistance is lost, so being unfavorable for playing long-acting；
(1) tillage control measure is improved: capsicum epidemic disease is prevented and treated by improvement agricultural tillage measure, it is not only time-consuming and laborious, and
The generation of epidemic disease cannot fundamentally be controlled；
(3) physical disinfection: if misoperation, easily causing soil-borne disease more serious, and needs in time to soil
Appropriate probiotic supplemented, can be only achieved better preventive effect, however probiotic supplemented this important step is but tended to by peasant
Ignore；
(4) apply chemical agent: capsicum epidemic disease is soil-borne disease, if pesticide is manured into soil using chemical pesticide control,
It will lead to soil and a series of complex processes such as physical absorption, chemisorption, migration and microbial degradation occur, seriously affected change
The control efficiency of medicine of learning to farm not only will affect the growth of capsicum if excessively increasing pesticide dosage, and it is residual to increase pesticide in pepper fruit
Allowance inhibits beneficial microbe activity in soil, and pollutes the environment, and increases production cost.
In recent years, using microorganism biocontrol microorganisms as the biological prevention of core, due to no pollution to the environment, without drug resistance
Property, capsicum root system microbiologic population, the induction features such as plant system resistant can be adjusted, be known as green Prevention Technique, by
Extensive concern.
In biological prevention, bacillus (Bacillus) is ideal biocontrol microorganisms, inhibits phytopathogen
Range it is very wide, including a variety of different types of diseases such as root, leaf, flower, fruit all have significant Biocontrol Potential.
Bacillus (Bacillus) is widely distributed, easily isolated from soil and plant, and it is inverse to generate heat-resistant
Property gemma, conducive to biocontrol agent production formulation and in the environment survival colonize and breed.
Currently, the bacillus for controlling plant diseases mainly has bacillus subtilis (Bacillus
Subtilis), bacillus laterosporus (Bacillus.laterosporus), bacillus cereus (Bacillus.cereus),
Bacillus licheniformis (Bacillus.licheniformis), bacillus thuringiensis (Bacillus.thuringiensis),
Paenibacillus polymyxa (Paenibacillus polymyxa), bacillus pumilus (Bacillus.pumilus) etc..
Bacillus not only can be used alone, and can also compound with chemical pesticide or botanical fungicide, realize advantage
It is complementary, reduce dosage, enhancing preventive effect, can be additionally used in mixed way with other Antagonistic Fungis, realize a variety of antibiotic bacterias have complementary functions,
And anti-multiple diseases, and can get the collaboration control efficiency of persistent.As it can be seen that bacillus is as biocontrol agent, with other
Microbial ratio shows bigger development potentiality and broader practice prospect.
However, no matter bacillus is single use, or compounded with chemical pesticide or botanical fungicide, also or with
Other Antagonistic Fungis are used in mixed way, and are all only shown the effect of controlling disease, are not played Plant growth promotion.
Summary of the invention
To solve the deficiencies in the prior art, the present invention common and extremely difficult prevention and treatment for capsicum greenhouse and protecting field is peppery
Green pepper epidemic disease disease, provide one kind while controlling disease, Plant growth promotion can also be played, realize capsicum upgrading,
The preparation method of the high efficiency composition biocontrol agent and the compound biocontrol fungicide that increase production, increase income.
In order to achieve the above objectives, the present invention adopts the following technical scheme that:
A kind of compound biocontrol fungicide, which is characterized in that by the fermentation liquid and mostly viscous class gemma bar of beautiful bacillus brevis NF2
The fermentation liquid of bacterium DL1 compounds.
Compound biocontrol fungicide above-mentioned, which is characterized in that the fermentation medium that two plants of bacterium use is respectively as follows:
The fermentation medium of beautiful bacillus brevis NF2: 0.2g/L MgSO4, 3g/L beef extract, 10g/L peptone,
10g/L glucose, pH 7.0-7.2,115 DEG C of high pressure sterilization 30min；
The fermentation medium of Paenibacillus polymyxa DL1: 5g/L NaCl, 10g/L peptone, 5g/L urea, 15g/L form sediment
Powder, pH 7.0-7.2,121 DEG C of high pressure sterilization 20min.
Compound biocontrol fungicide above-mentioned, which is characterized in that the compound proportion of the fermentation liquid of two plants of bacterium are as follows: 4 × 108CFU/mL
The fermentation liquid and 2.6 × 10 of beautiful bacillus brevis NF28The fermentation liquid of CFU/mL Paenibacillus polymyxa DL1 is according to 1:1 body
Product mixing.
The preparation method of compound biocontrol fungicide above-mentioned, which comprises the following steps:
(1) activated strains
The original strain for being stored in -80 DEG C is inoculated on beef-protein medium respectively through sterile working and is activated,
30 DEG C of cultivation temperature, incubation time is for 24 hours；
(2) fermentation seed liquid is prepared
The single colonie of the single colonie of beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1 is inoculated in beef respectively
It in cream peptone fluid nutrient medium, is subsequently placed in constant temperature oscillator, 30 DEG C of fermentation temperature, revolving speed 200r/min, cultivates 16h,
It is spare as fermentation seed liquid；
(3) prepare fermentation medium
The fermentation medium of beautiful bacillus brevis NF2: 0.2g/L MgSO4, 3g/L beef extract, 10g/L peptone,
10g/L glucose, pH 7.0-7.2,115 DEG C of high pressure sterilization 30min；
The fermentation medium of Paenibacillus polymyxa DL1: 5g/L NaCl, 10g/L peptone, 5g/L urea, 15g/L form sediment
Powder, pH 7.0-7.2,121 DEG C of high pressure sterilization 20min；
(4) fermentation liquid is prepared
The fermentation seed liquid of beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1 is respectively connected to respectively corresponding
Fermentation medium in, inoculum concentration 5%, 30 DEG C of fermentation temperature, revolving speed 200r/min, cultivate 48h, obtain fermentation liquid；
(5) it compounds
Fermentation liquid is diluted, with 4 × 108The fermentation liquid and 2.6 × 10 of CFU/mL beauty bacillus brevis NF28CFU/mL is more
The fermentation liquid of viscous series bacillus DL1 is according to 1:1 volume mixture to get compound biocontrol fungicide.
The invention has the beneficial effects that:
(1) compound biocontrol fungicide provided by the invention combines beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1
Biological and ecological methods to prevent plant disease, pests, and erosion characteristic, widened antimicrobial spectrum, in conjunction with a variety of antimicrobial mechanisms, sufficiently coordinated the microecological balance of soil, enhanced bacterium
Strain facilitates the disease-resistant advantage of collaboration between long-acting performance bacterial strain, has than Paenibacillus polymyxa DL1 single bacterium to the adaptability of environment
There is better biocontrol effect；
(2) compound biocontrol fungicide provided by the invention can adjust the micro-ecological environment of plant rhizosphere soil, and by luring
Plant generation system disease resistance is led, plant disease-resistant ability can be improved, is finally reached growth-promoting, disease-resistant integrated control effect, is realized
The upgrading of capsicum is increased production, increases income；
(3) compound biocontrol fungicide provided by the invention has good environment compatibility and safety.
Specific embodiment
Below in conjunction with detailed description of the invention by specific embodiments.
One, microorganism fungus kind
1, beautiful bacillus brevis NF2
Beautiful bacillus brevis NF2 separation screening is from Shaanxi Mei County Kiwi berry rhizosphere soil.Operating procedure is specific as follows:
Soil sample, which weighs 25g after drying at room temperature and is put into the sterile water that 225mL contains a small amount of bead, vibrates 30min, sample
Product suspension takes 0.1mL dilution spread beef extract-peptone plate through 80 DEG C of heating water bath 20min, and 30 DEG C of inversions are cultivated 48h, chosen
Take single colonie.
Phage gene group DNA is extracted, by 16Sr determined dna sequence, and combines bio-chemical characteristics as a result, final mirror
The bacterial strain is made as beautiful bacillus brevis (Brevibacillus formosus), we are denoted as beautiful bacillus brevis
NF2。
Then we have carried out preservation to the bacterial strain, and preservation information is as follows:
Deposit number: CGMCC No.12749, depositary institution: China General Microbiological culture presevation administrative center
(CGMCC), preservation address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, preservation date: on July 8th, 2016.
2, Paenibacillus polymyxa DL1
Paenibacillus polymyxa DL1 separation screening belongs to endogenetic bacteria from the bast of Dali County greenhouse production pepper root.Behaviour
It is specific as follows to make step:
The interconnecting piece for taking healthy pepper plant stem and root, is cleaned up with tap water, is then impregnated with 2% sodium hypochlorite
5min is sterilized, then with aseptic water washing 3 times, is later torn stem epidermis with sterile razor blade, it will be subepidermal with sterile tweezers
Organize and be affixed on PDA culture medium surface, picking single colonie after 28 DEG C of cultures.
Phage gene group DNA is extracted, by 16Sr determined dna sequence, and combines bio-chemical characteristics as a result, final mirror
Making the bacterial strain is Paenibacillus polymyxa (Paenibacillus polymyxa), we are denoted as Paenibacillus polymyxa
DL1。
Then we have carried out preservation to the bacterial strain, and preservation information is as follows:
Deposit number: CGMCC No.16311, depositary institution: China General Microbiological culture presevation administrative center
(CGMCC), preservation address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, preservation date: on August 17th, 2018.
Two, pathogen is tested
Phytophthora blight of pepper: biological husbantry research institute of Shaanxi Province provides.
Three, culture medium
1, microorganism fungus kind minimal medium
Beef-protein medium: beef extract 3g, peptone 5g, glucose 10g, agar powder 15g, distilled water
1000mL, pH 7.0-7.2,121 DEG C of high pressure sterilization 20min.
2, pathogen minimal medium
PDA culture medium: potato 200g is cut into 1cm fritter and 500mL boiling is added to boil 30min, and filtered through gauze adds after taking filtrate
Enter glucose 20g, kept the skin wet after dissolution to 1000mL, pH is naturally, 115 DEG C of high pressure sterilization 15min.
Four, compound biocontrol fungicide is prepared
1, activated strains
- 80 DEG C of original strain (beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1) will be stored in through sterile behaviour
It is inoculated on beef-protein medium and activates respectively, 30 DEG C of cultivation temperature, incubation time is for 24 hours.
2, the antagonistic effect of biocontrol microorganisms
Using tablet face-off method, antagonistic effect is carried out to beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1.
As a result: not finding there is antagonism between two plants of bacterium.
Therefore, beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1 can be used in mixed way.
3, fermentation seed liquid is prepared
The single colonie of the single colonie of beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1 is inoculated in beef respectively
It in cream peptone fluid nutrient medium, is subsequently placed in constant temperature oscillator, 30 DEG C of fermentation temperature, revolving speed 200r/min, cultivates 16h,
It is spare as fermentation seed liquid.
4, fermentative medium formula optimizes
(1) beautiful bacillus brevis NF2
30 DEG C of fermentation temperature, revolving speed 200r/min, 48h is cultivated, the nutrition in fermentation medium is optimized by orthogonal experiment
Ingredient (is shown in Table 1).
Nutritional ingredient in the beautiful bacillus brevis NF2 orthogonal experiment optimization fermentation medium of table 1
The results show that the fermentative medium formula after the optimization of beautiful bacillus brevis NF2 are as follows:
0.2g/L MgSO4, 3g/L beef extract, 10g/L peptone, 10g/L glucose, pH 7.0-7.2, viable count stablize
≥2.0×108CFU/mL meets agricultural microbial agent and executes standard GB20287-2006.
(2) Paenibacillus polymyxa DL1
30 DEG C of fermentation temperature, revolving speed 200r/min, 48h is cultivated, the nutrition in fermentation medium is optimized by orthogonal experiment
Ingredient (is shown in Table 2).
2 Paenibacillus polymyxa DL1 orthogonal experiment of table optimizes the nutritional ingredient in fermentation medium
The results show that the fermentative medium formula after the optimization of Paenibacillus polymyxa DL1 is 5 g/L NaCl, 10g/L
Peptone, 5g/L urea, 15g/L starch, pH 7.0-7.2, viable count stablize >=2.0 × 108CFU/mL meets agricultural micro- life
Object microbial inoculum executes standard GB20287-2006.
5, fermentation liquid is prepared
The fermentation medium of beautiful bacillus brevis NF2, pH 7.0- are prepared according to the fermentative medium formula after optimization
7.2,115 DEG C of high pressure sterilization 30min.
The fermentation medium of Paenibacillus polymyxa DL1, pH 7.0- are prepared according to the fermentative medium formula after optimization
7.2,121 DEG C of high pressure sterilization 20min.
The fermentation seed liquid of beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1 is respectively connected to corresponding hair
In ferment culture medium, inoculum concentration 5%, cultivates 48h, obtains fermentation liquid by 30 DEG C of fermentation temperature, revolving speed 200r/min.
Through detecting, the concentration of beautiful bacillus brevis NF2 is 4 × 10 in fermentation liquid9CFU/mL, Paenibacillus polymyxa
The concentration of DL1 is 2.6 × 108CFU/mL。
6, biocontrol agent compound proportion optimizes
The fermentation liquid of beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1 is distinguished into gradient dilution to 108、107、
106, then according to 1:1 volume mixture, analyzed by orthogonal experiment and determine two plants of optimal conditions of mixture ratios of bacterium (being shown in Table 3).
The orthogonal experiment of 3 biocontrol agent compound proportion of table is analyzed
The results show that as beautiful bacillus brevis NF2 (4 × 108CFU/mL) and Paenibacillus polymyxa DL1 (2.6 ×
108When CFU/mL) according to 1:1 volume mixture, single plant yield of hot pepper highest.
7, biocontrol agent is compounded
By the fermentation liquid (4 × 10 of beautiful bacillus brevis NF29CFU/mL 4 × 10) are diluted to8 CFU/mL。
By the fermentation liquid (2.6 × 10 of Paenibacillus polymyxa DL18CFU/mL 2.6 × 10) are diluted to8 CFU/mL。
By the fermentation liquid after dilution according to 1:1 volume mixture to get compound biocontrol fungicide.
Five, the biocontrol effect of compound biocontrol fungicide
Field trial is completed in Shaanxi Province Weinan City Dali County modern agriculture Demonstration Base greenhouse, and the time 2018 3
Month in July, 2018, the every processing cell 10m of Hot Pepper Seedling2, 4 groups of repetitions.When transplanting Hot Pepper Seedling respectively using mostly viscous class gemma
Bacillus DL1 single bacterium (2.0 × 108CFU/mL) and compound biocontrol fungicide does root irrigation, starts within 10 days after transplanting to be inoculated with pathogen
Phytophthora capsici disease spores (105CFU/mL, 25mL) it is infected, every 2 months, class gemma bar was glued in corresponding application more later
50mL/ plants of capsicums of bacterium DL1 single bacterium and compound biocontrol fungicide, cooperate normal field management.To apply clear water as negative control,
To apply 800 times of dilutions of Mancozeb as positive control.The incidence for observing capsicum epidemic disease, 30 from there is capsicum epidemic disease
Incidence is investigated after it, and calculates disease incidence, disease index and control efficiency (being shown in Table 4).
The control efficiency of 4 single bacterium of table and compound biocontrol fungicide to capsicum epidemic disease
The results show that compound biocontrol fungicide has good control efficiency to capsicum epidemic disease, relative control effect be can achieve
70.4%, and shown compared to Paenibacillus polymyxa DL1 single bacterium and cooperate with disease-resistant advantage.
Six, the disease-resistant growth-promoting functions of compound biocontrol fungicide
After applying Paenibacillus polymyxa DL1 single bacterium and compound biocontrol fungicide, capsicum top the 3rd Newborn Leaves reciprocal are taken
Piece detects the content of the malonaldehyde (MDA) of pepper plant and the activity (being shown in Table 5) of defensive ferment using spectrophotometer method,
Wherein, defensive ferment includes: phenylalanine lyase (PAL), superoxide dismutase (SOD), polyphenol oxidase (PPO), peroxide
Change hydrogen enzyme (CAT), peroxidase (POD).
The growth-promoting effect of 5 single bacterium of table and compound biocontrol fungicide to capsicum
The results show that compound biocontrol fungicide can dramatically increase the activity of capsicum PAL, PPO, CAT and POD, therefore can
By Scavenging active oxygen, protection membrane stability, positive response is generated to stress and enhances the ability that pepper plant resists pathogen,
Improve pepper plant itself disease resistance.
In addition, we also count the yield of hot pepper of application compound biocontrol fungicide, the results show that applying compound life
After anti-microbial inoculum, the yield of every plant of capsicum reaches 0.394kg, and folding per mu yield about 1600kg about increases production per acre compared with clear water control group
39.5%, it is seen then that compound biocontrol fungicide of the invention has good effect of increasing production.
In conclusion compound biocontrol fungicide of the invention, not only has significant control efficiency to capsicum epidemic disease, is conducive to
The amount of application of chemical pesticide is reduced, pesticide residue is reduced, preserves the ecological environment, but also can induce and improve pepper plant itself
Disease-resistant performance increases capsicum per mu yield, is conducive to upgrading increasing both production and income, increases economic efficiency.
It should be noted that the above embodiments do not limit the invention in any form, it is all to use equivalent replacement or equivalent change
The mode changed technical solution obtained, falls within the scope of protection of the present invention.
Claims (5)
1. a kind of compound biocontrol fungicide, which is characterized in that by the fermentation liquid and Paenibacillus polymyxa of beautiful bacillus brevis NF2
The fermentation liquid of DL1 compounds.
2. compound biocontrol fungicide according to claim 1, which is characterized in that the fermentation medium difference that two plants of bacterium use
Are as follows:
The fermentation medium of beautiful bacillus brevis NF2: 0.2g/L MgSO4, 3g/L beef extract, 10g/L peptone, the Portugal 10g/L
Grape sugar, pH 7.0-7.2,115 DEG C of high pressure sterilization 30min；
The fermentation medium of Paenibacillus polymyxa DL1: 5g/L NaCl, 10g/L peptone, 5g/L urea, 15g/L starch, pH
7.0-7.2 121 DEG C of high pressure sterilization 20min.
3. compound biocontrol fungicide according to claim 2, which is characterized in that the compound proportion of the fermentation liquid of two plants of bacterium are as follows: 4
×108The fermentation liquid and 2.6 × 10 of CFU/mL beauty bacillus brevis NF28The fermentation liquid of CFU/mL Paenibacillus polymyxa DL1
According to 1:1 volume mixture.
4. the preparation method of compound biocontrol fungicide described in claims 1 to 3 any one, which is characterized in that including following step
It is rapid:
(1) activated strains
The original strain for being stored in -80 DEG C is inoculated on beef-protein medium respectively through sterile working and is activated, is cultivated
30 DEG C of temperature, incubation time is for 24 hours；
(2) fermentation seed liquid is prepared
The single colonie of the single colonie of beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1 is inoculated in beef extract egg respectively
It in white peptone fluid nutrient medium, is subsequently placed in constant temperature oscillator, 30 DEG C of fermentation temperature, revolving speed 200r/min, cultivates 16h, as
Fermentation seed liquid is spare；
(3) prepare fermentation medium
The fermentation medium of beautiful bacillus brevis NF2: 0.2g/LMgSO4, 3g/L beef extract, 10g/L peptone, 10g/L grape
Sugar, pH 7.0-7.2,115 DEG C of high pressure sterilization 30min；
The fermentation medium of Paenibacillus polymyxa DL1: 5g/L NaCl, 10g/L peptone, 5g/L urea, 15g/L starch, pH
7.0-7.2 121 DEG C of high pressure sterilization 20min；
(4) fermentation liquid is prepared
The fermentation seed liquid of beautiful bacillus brevis NF2 and Paenibacillus polymyxa DL1 is respectively connected to corresponding hair
In ferment culture medium, inoculum concentration 5%, cultivates 48h, obtains fermentation liquid by 30 DEG C of fermentation temperature, revolving speed 200r/min；
(5) it compounds
Fermentation liquid is diluted, with 4 × 108The fermentation liquid and 2.6 × 10 of CFU/mL beauty bacillus brevis NF28The mostly viscous class of CFU/mL
The fermentation liquid of bacillus DL1 is according to 1:1 volume mixture to get compound biocontrol fungicide.
5. compound biocontrol fungicide described in claims 1 to 3 any one prevention and treatment capsicum epidemic disease, improve pepper disease resistance performance and
Increase the application in terms of yield of hot pepper.
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