CN105028483B - Composite bactericide and its application containing bacillus subtilis BAB 1 and Boscalid - Google Patents
Composite bactericide and its application containing bacillus subtilis BAB 1 and Boscalid Download PDFInfo
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Abstract
A kind of application the present invention relates to composite bactericide containing bacillus subtilis BAB 1 and Boscalid and its in preventing and treating graw mold of tomato and leaf mold.The composite bactericide raw material contains active component, filler and/auxiliary agent；The active component is made up of bacillus subtilis BAB 1 and Boscalid.Composite bactericide of the present invention is by after bacillus subtilis BAB 1 and Boscalid compounding; form the effect having complementary advantages; efficient, the quick bacteriostasis of Boscalid and the therapeutic action to gray mold and leaf mold can be played, and can plays bacillus subtilis BAB 1 in induction tomato resistance and protective effect to gray mold and leaf mold；Stable, balance, bio-diversity a ecosystem can be formed, is reached and is improved drug effect, reduces chemical pesticide usage amount, lasting control graw mold of tomato and the popular purpose of leaf mold.
Description
Technical field
The present invention relates to a kind of composite bactericide and its application, more particularly to it is a kind of containing bacillus subtilis BAB-1 and
The composite bactericide of Boscalid and its application in preventing and treating graw mold of tomato and leaf mold.
Background technology
Gray mold is a kind of fungal disease as caused by Botrytis cinerea (Botrytis cinerea), and Botrytis cinerea can be with
The various plants of tomato, strawberry, cucumber, bottle gourd, chrysanthemum, apple, cucurbita pepo and grape etc. 200 are infected, cause the generation of gray mold.
Gray mold is a kind of global important disease, endangers and is particularly acute on facility fruits and vegetables.
Leaf mold is another fungal disease in tomato production, main harm blade, the stem that can also cause harm when serious,
Flower, fruit etc..Scab is more gradually to be crept up since lower blade, and full leaf withered curling can be caused when serious, causes plant
Early ageing, influence the yield and quality of tomato.
Due to there is no the material and kind of botrytis resistant in germ plasm resource, mainly it is aided with production based on chemical prevention
Biological and ecological control.At present, chemical prevention is to prevent and treat the important means of gray mold, but Botrytis cinerea has breeding soon, heredity
The characteristics of variation is greatly and grade of fit is high, is used continuously same medicament and is also easy to produce the resistance to the action of a drug.In production, the conventional change of preventing and treating gray mold
Learn the carbendazim of bactericide such as benzimidazole, the procymidone of dicarboximide class, the pyrimethanil etc. of anilino-pyrimidine it is equal because
Botrytis cinerea it is drug-fast generation and prevention effect substantially reduces.Dosage is arbitrarily increased if preventive effect is improved, the result is that not
But without the generation for effectively controlling gray mold, the residual risk of fruits and vegetables middle peasant is improved on the contrary, increases pollution of the agricultural chemicals to environment.
With the raising of the food-safe attention rate of people, biological control is because to person poultry safety, environment compatibility is good, germ
The features such as being not likely to produce the resistance to the action of a drug, have attracted increasing attention.Particularly bacillus subtilis, due to its can produce it is more
Kind antibacterial material, preventive effect is obvious, and existing many excellent bacillus subtilis strains are applied to production practices both at home and abroad at present.It is raw
Thing agricultural chemicals is for reducing the pollution of farmland chemical pesticide, ensureing that agricultural product quality and safety is significant.But biological control
Come with some shortcomings, such as field control effect is unstable, field efficacy effect is slow, has had a strong impact on that it is promoted the use of.
In order to make up the deficiency of biological control and chemical prevention, carry out Biocontrol microorganism and less toxic chemical agent Collaborative Control
The technical research of graw mold of tomato, can play that chemical agent is efficient, quick bacteriostasis, and and can plays Biocontrol microorganism and existed
Induce disease resistance of plant, the popular purpose of lasting control graw mold of tomato.
Boscalid (Boscalid) is the Novel tobacco acid amide fungicides developed by BASF Aktiengesellschaft, is mainly used in
Prevent and treat gray mold, powdery mildew, various rots, brown rot and root rot etc..Boscalid belongs to amber in mitochondrial respiratory chain
Sour ubiquinone reductase inhibitor, the sprouting to spore have a very strong rejection ability, and with other bactericide no interactions resistances.Separately
The toxicity of outer Boscalid is also very low, to male female rat, the oral LD of chmice acute50It is all higher than 5000mg/kg.It is anxious to male female rat
The percutaneous LD of property50>2000mg/kg.To male female chmice acute suction LD50>6.7mg/kg.It is stingless to lagophthalmos eyeball and Skin-test
Swash.Cavy is tested without sensitization.On honeybee, bird, earthworm, silkworm etc. without influence.It is after flushing liquor drying to honeybee and birds
Feeding can be put.Meanwhile on various natural enemies also without influence.
In view of this, it is special to propose the present invention.
The content of the invention
It is an object of the invention to：Largely chemical pesticide control graw mold of tomato and leaf mold are used for production at present is upper
Trigger the resistance of pathogen, seriously pollute environment, cause the problems such as exceeded residues of pesticides, influence Tomato Quality, there is provided be a kind of right
Graw mold of tomato and leaf mold have the compound sterilizing containing bacillus subtilis BAB-1 and Boscalid of good prevention effect
Agent and its application.It is mutual to form advantage by after bacillus subtilis BAB-1 and Boscalid compounding for composite bactericide of the present invention
The effect of benefit, efficient, the quick bacteriostasis of Boscalid and the therapeutic action to gray mold and leaf mold can be played, again
Bacillus subtilis BAB-1 can be played in the effect such as induction tomato resistance and protection to gray mold and leaf mold.By using
The composite bactericide, a large amount of beneficial microbes are introduced in tomato planting environment, deteriorate the living environment of pathogen, with reference to cultivation
The agricultural measures such as management, stable, balance, bio-diversity a ecosystem is formed, reaches and improves drug effect, reductionization
Learn the purpose of Pesticide use amount, lasting control graw mold of tomato and leaf mold prevalence.
Bacillus subtilis BAB-1 of the present invention is screened by Inst. of Plant Protection, Hebei-Prov. Academy of Agricultural and Forestry Scie and obtained.
It is commonly micro- to be preserved in China Committee for Culture Collection of Microorganisms on July 6th, 2007 by the bacillus subtilis BAB-1
Bio-Centers, deposit number are CGMCC No.2099；Classification system is：Bacillus subtilis BAB-1 are (hereinafter referred to as
BAB-1).The bacterial strain have the characteristics that efficiently, antimicrobial spectrum it is wide, using it is simple, cost is low, non-environmental-pollution, available for tomato ash
The preventing and treating of mildew and leaf mold.Bacillus subtilis BAB-1 has natural resistance to chemical agent Boscalid, and the bacterial strain can
To be grown on LB culture medium of the Boscalid content as 1000 μ g/mL.Thus can be by bacillus subtilis BAB-1 and pyridine acyl
Bacterium amine is re-dubbed bactericide, realizes the two long-term co-existence.
Realize that technical scheme is as follows：A kind of compounding containing bacillus subtilis BAB-1 and Boscalid is killed
Microbial inoculum, its active component are made up of bacillus subtilis (Bacillus subtilis) BAB-1 and Boscalid.
Preferably, the bacillus subtilis BAB-1 is its thalline and/or its metabolite.
Preferably, the bacillus subtilis BAB-1 is its zymotic fluid or its lyophilized bacterium powder；The further preferred fermentation
The total viable count of liquid is 7 × 109~1 × 1010CFU/mL；The total viable count of the further preferred lyophilized bacterium powder is 1.8 × 1010
~2.4 × 1010CFU/g.Preferably, in the active component Boscalid and bacillus subtilis BAB-1 weight ratio for 1~
99: 99~1；Further preferred weight ratio is 1~9: 9~1；Preferable weight ratio is 1~4: 4~1.
Specifically, a kind of composite bactericide containing bacillus subtilis BAB-1 and Boscalid, in parts by weight, its
Raw material contains following components：Above-mentioned 10~80 parts of active component, 1~8 part of dispersant, 1~8 part of wetting agent, 10~80 parts of filler；
It is preferred that its raw material contains following components：40~70 parts of active component, 3~7 parts of dispersant, 3~7 parts of wetting agent, filler 20~50
Part；More preferably its raw material contains following components：50~60 parts of active component, 4~6 parts of dispersant, 4~6 parts of wetting agent, filler 30
~40 parts.
Preferably, the Boscalid is the Boscalid active compound of mass concentration 98%.
Preferably, the dispersant be lignin sulfonic acid sodium salt, sanlose, desugar sodium lignin sulfonate,
Lignosulfite, APES, ethylene glycol monobutyl ether, polyoxyethanyl alkyl ether, Nonyl pheno
One or more mixtures in ether, neopelex.
Preferably, described wetting agent is nekal (Nekal BX), wetting agent wgwin D30, fatty alcohol
APEO, dodecyl sodium sulfate, pelopon A, alkylphenol-polyethenoxy base formaldehyde condensate sulfonate
In one or more mixtures.
Preferably, the filler is one kind in white carbon, kaolin, fine particle calcium carbonate, diatomite, precipitated calcium carbonate, bentonite
Or a variety of mixture.
Specifically, a kind of composite bactericide containing bacillus subtilis BAB-1 and Boscalid, in parts by weight, contains
There is following components：The Boscalid active compound of mass concentration 98%：20.41 parts；Bacillus subtilis BAB-1 freezes bacterium powder (viable bacteria
Sum is 2.0 × 1010CFU/g)：40 parts；Dispersant sanlose：3 parts；Wetting agent：Dodecyl sodium sulfate：
4.5 part；Filler precipitated calcium carbonate：32.09 parts.Being matched somebody with somebody with this, the withered grass bud of 20% Boscalid 8,000,000,000 can be made as follows
Spore bacillus BAB-1 wettable powders.
Parts by weight of the present invention can be the unit of weight well known in the art such as μ g, mg, g, kg or its times
Number, such as 1/10,1/100,10 times, 100 times.
Composite bactericide of the present invention containing bacillus subtilis BAB-1 and Boscalid can be by art methods
It is prepared into liquid preparation or wettable powder.
The present invention also provides the preparation method of the above-mentioned composite bactericide containing bacillus subtilis BAB-1 and Boscalid
Including：After various components are mixed according to said ratio, carry out primary air stream crushing (grain fineness reaches 80 mesh)；Mixing
After stirring, then secondary air flow crushing (grain fineness reaches 325 mesh) is carried out, produced.
Bacillus subtilis BAB-1 of the present invention freezes bacterium powder can be by bacillus subtilis BAB-1 zymotic fluids by existing
Technology conventional method is made.The bacillus subtilis BAB-1 zymotic fluids can be by prior art such as Chinese patent
The method that CN200710121275.0 is recorded is made；Or it is made by the following method.
A kind of preparation method of bacillus subtilis BAB-1 zymotic fluids, comprises the following steps：BAB-1 strains are existed first
Activated in LB flat boards, picking single bacterium colony is scoring in Ke Shi bottles, is washed out lawn with sterilized water after 3 days and is seeded to seeding tank (such as 1
Ton tank) seed liquor is used as, seed liquor is inoculated into fermentation tank (such as 5 ton tanks) after 31 DEG C of culture 12h is fermented, tank under 48h, i.e.,
Obtain bacillus subtilis BAB-1 zymotic fluids.
After tank under the bacillus subtilis BAB-1 zymotic fluids, centrifuged and freezed, that is, obtain bacillus subtilis
BAB-1 freezes bacterium powder.
Present invention additionally comprises the composite bactericide containing bacillus subtilis BAB-1 and Boscalid in preventing and treating plant
Application in terms of gray mold and leaf mold；The plant includes but is not limited to water fruits and vegetables and flowers, such as tomato, strawberry, Huang
The various plants of melon, cucurbita pepo, grape, lettuce, romaine lettuce, Chinese cabbage, eggplant, capsicum, fresh kidney beans, bottle gourd, chrysanthemum, apple etc. 200；It is preferred that
For tomato.Composite bactericide of the present invention containing bacillus subtilis BAB-1 and Boscalid is to graw mold of tomato and leaf
The preventing and treating of mildew has remarkable result, and prevention effect is respectively up to 88.6% and 87.5%.
The application method of composite bactericide of the present invention containing bacillus subtilis BAB-1 and Boscalid includes：
By after the compound sterilizing dilution agent containing bacillus subtilis BAB-1 and Boscalid to the plant (such as tomato)
Cauline leaf is sprayed, to prevent and treat gray mold and leaf mold；It is preferred that prevented and treated in the growth period of the plant (such as tomato).
Usually, its is effective when in use for the composite bactericide containing bacillus subtilis BAB-1 and Boscalid
Component content is at least 300 μ g/mL；The μ g/mL of preferably 500 μ g/mL~600.
Usually, make after the composite bactericide containing bacillus subtilis BAB-1 and Boscalid is diluted with water
With.
The advantage of the invention is that：By using to person poultry safety, have ash arrhizus bacteria and Fulvia fulva Ciferri compared with high inhibition
The bacillus subtilis BAB-1 of effect, itself and less toxic chemical agent Boscalid are compounded by a certain percentage, it is mutual to reach advantage
The dosage of chemical bactericide is mended, reduced, while prevention effect is improved, ecological environment is improved, reduces chemical agent
Residual quantity in vegetables.
Embodiment
Following examples are used to illustrate the present invention, but are not limited to the scope of the present invention.
Such as indicated without special, the compound sterilizing containing bacillus subtilis BAB-1 and Boscalid described in following examples
Raw material Boscalid in agent is the Boscalid active compound of mass concentration 98%, is provided by Jiangyin Suli Chemicals Co., Ltd.；
Raw material Bacillus subtilis BAB-1 is that bacillus subtilis (Bacillus subtilis) BAB-1 freezes bacterium powder (total viable count
For 2.0 × 1010CFU/g), provided by Inst. of Plant Protection, Hebei-Prov. Academy of Agricultural and Forestry Scie.
Embodiment 1
The present embodiment provides a kind of composite bactericide containing bacillus subtilis BAB-1 and Boscalid, containing following
Component：Active component 60kg；Dispersant：APES 5Kg, ethylene glycol monobutyl ether 1Kg；Wetting agent：Dodecane
Base benzene sulfonic acid sodium salt 4Kg, AEO 3Kg；Filler：White carbon 5Kg, kaolin 25Kg；Wherein active component by
The Boscalid active compound of mass concentration 98% and bacillus subtilis BAB-1 freeze bacterium powder, and (total viable count is 2.0 × 1010CFU/g)
Composition, the two weight ratio are 4: 1.
Embodiment 2
The present embodiment provides a kind of composite bactericide containing bacillus subtilis BAB-1 and Boscalid, with embodiment 1
It is 2: 1 that active component Boscalid, which is differed only in, with both bacillus subtilis BAB-1 weight ratio.
Embodiment 3
The present embodiment provides a kind of composite bactericide containing bacillus subtilis BAB-1 and Boscalid, with embodiment 1
It is 1: 1 that active component Boscalid, which is differed only in, with both bacillus subtilis BAB-1 weight ratio.
Embodiment 4
The present embodiment provides a kind of composite bactericide containing bacillus subtilis BAB-1 and Boscalid, with embodiment 1
It is 1: 2 that active component Boscalid, which is differed only in, with both bacillus subtilis BAB-1 weight ratio.
Embodiment 5
The present embodiment provides a kind of composite bactericide containing bacillus subtilis BAB-1 and Boscalid, with embodiment 1
It is 1: 4 that active component Boscalid, which is differed only in, with both bacillus subtilis BAB-1 weight ratio.
Embodiment 6
The present embodiment provides a kind of composite bactericide containing bacillus subtilis BAB-1 and Boscalid, with embodiment 1
It is 1: 99 that active component Boscalid, which is differed only in, with both bacillus subtilis BAB-1 weight ratio.
Embodiment 7
The present embodiment provides a kind of composite bactericide containing bacillus subtilis BAB-1 and Boscalid, with embodiment 1
It is 99: 1 that active component Boscalid, which is differed only in, with both bacillus subtilis BAB-1 weight ratio.
Embodiment 8
The present embodiment provides a kind of composite bactericide containing bacillus subtilis BAB-1 and Boscalid, with 100kg
Meter, contains following components：The Boscalid active compound of mass concentration 98%：20.41kg；Bacillus subtilis BAB-1 freezes bacterium powder
(total viable count is 2.0 × 1010CFU/g)：40kg；Dispersant sanlose：3kg；Wetting agent dodecyl sodium sulfonate
Sodium：4.5kg；Filler precipitated calcium carbonate：32.09kg.
Embodiment 9
The preparation method for the composite bactericide being formulated as described in Example 1 is present embodiments provided, is specifically included：According to
Than various components are mixed, primary air stream crushing is carried out, grain fineness reaches 80 mesh；After mixing, then through air-flow powder
Broken machine carries out two airflow levels crushing, and grain fineness reaches 325 mesh, is made.
The preparation method of composite bactericide described in embodiment 2-8 is same as Example 9.
Experimental example 1
Virulence experiments of the bacillus subtilis BAB-1 with Boscalid to botrytis cinerea
1 experiment material
It is botrytis cinerea (Botrytis cinerea) for examination cause of disease bacteria strain, by Hebei Prov. Academy of Agricultural ＆. Forest Sciences plant
Protective strategy is provided, using PDA culture medium (potato 200g, glucose 20g, agar 10g, distilled water 1000mL) in 25 DEG C ±
Preculture 3d, standby under the conditions of 1 DEG C.The Boscalid active compound of mass concentration 98%, is carried by Jiangyin Suli Chemicals Co., Ltd.
For.Bacillus subtilis (Bacillus subtilis) BAB-1 zymotic fluids (total viable count is 8,000,000,000 CFU/mL), by Hebei province
Agricultural and forest science institute Plant Protection Institute provides.Embodiment 1-5 prepare containing bacillus subtilis BAB-1 and Boscalid
Composite bactericide, its active component Boscalid and both bacillus subtilis BAB-1 weight ratio respectively 4:1、2:1、1:1、
1:2、1:4。
2 experimental methods
The Boscalid active compound of mass concentration 98% is dissolved in proper amount of acetone by 2.1, and the mother liquor that concentration is 10000 μ g/mL is made,
Then be serially diluted with sterilized water, mixed with PDA culture medium by 1: 9 volume ratio, be respectively prepared concentration for 10,5,1,
0.5th, 0.1,0.05 μ g/mL drug containing flat board.
2.2 are diluted bacillus subtilis BAB-1 zymotic fluids with sterilized water, and 1: 9 volume ratio is pressed with PDA culture medium
Example mixing, the drug containing flat board that bacillus subtilis bacteria concentration is 10,5,1,0.5,0.1 and 0.05 μ g/mL is respectively prepared.
Composite bactericide containing bacillus subtilis BAB-1 and Boscalid prepared by 2.3 Example 1-5 is with sterile
Water is diluted, and is mixed respectively with PDA culture medium, and the drug containing flat board that concentration is 10,5,1,0.5,0.1,0.05 μ g/mL is made.
Concentration refers to the concentration of active component in the composite bactericide (i.e. bacillus subtilis BAB-1 and Boscalid) herein.
2.4 in the above-mentioned bacterium that a diameter of 5mm is taken through being beaten on the preculture botrytis cinerea PDA plate of 3 days with card punch
Cake, bacteria cake bacterium is inoculated into the drug containing flat board center of the preparation of above-mentioned steps 2.1~2.3 respectively down；And with without medicament
PDA plate is blank control, is operated in the same way；After incubated 3d under the conditions of 25 DEG C ± 1 DEG C, crossing method measurement
Colony diameter.
3 results calculate
The inhibiting rate of each processing is calculated using EXCEL, according to the concentration and inhibiting rate of each processing, utilizes DPSv6.55 data
Processing software obtains regression coefficient (b), intercept (a), coefficient correlation, EC50(processing when effectively suppressing pathogen growth 50% is dense
Degree, unit is μ g/mL).Then virulence regression equation is write out：Y=bx+a, wherein y represent the probit value of processing inhibiting rate；X tables
Show the logarithm value of the concentration of processing.
Inhibiting rate (%)=(control bacterium colony net growth-processing bacterium colony net growth diameter)/control bacterium colony net growth
× 100%
The coefficient of synergism of different compound drugs is calculated with reference to Wadley (1967) formula：
SR=EC50(th)/EC50(ob)
It is shared in the mixture to represent two kinds of Boscalid (A), bacillus subtilis (B) bactericide respectively by a, b in above formula
Percentage；EC50And EC (A)50(B) A and B actual observation EC is represented respectively50Value, EC50(th) represent that two kinds of bactericide of A, B are pressed
a:The mixed theoretical EC of b50Value, EC50(ob) a is pressed for two kinds of bactericide of A, B:The mixed actual observation EC of b50Value.SR>
1.5, represent synergy；SR<0.5, represent antagonism；SR represents summation action between 0.5 and 1.5.
4 experimental results
It is shown in Table 1.As a result show, withered grass gemma is contained described in Boscalid, bacillus subtilis BAB-1 and embodiment 1-5
The composite bactericide of bacillus BAB-1 and Boscalid has good inhibitory action to botrytis cinerea respectively；And implement
Composite bactericide described in example 1-5 is all higher than 1.5 to the coefficient of synergism of botrytis cinerea, shows described in embodiment 1-5 containing withered
Boscalid has synergy work with bacillus subtilis BAB-1 in careless bacillus BAB-1 and Boscalid composite bactericide
With.
The composite bactericide containing bacillus subtilis BAB-1 and Boscalid of the different ratio of table 1 is to graw mold of tomato
The co-toxicity of bacterium
| Medicament | Virulence regression equation | EC50 | It is related | Synergy |
| (μg/mL) | Coefficient | Coefficient | ||
| Boscalid (A) | Y=0.4591x+4.9027 | 1.6291 | 0.9975 | - |
| Bacillus subtilis BAB-1 (B) | Y=1.3081x+6.9959 | 0.0298 | 0.839 | - |
| (the A of embodiment 1:B=4:1) | Y=0.6651x+5.6944 | 0.0903 | 0.9552 | 1.54 |
| (the A of embodiment 2:B=2:1) | Y=0.6430x+5.8558 | 0.0467 | 0.9384 | 1.85 |
| (the A of embodiment 3:B=1:1) | Y=1.3254x+6.9616 | 0.0331 | 0.8406 | 1.77 |
| (the A of embodiment 4:B=1:2) | Y=1.5315x+7.5243 | 0.0225 | 0.845 | 1.97 |
| (the A of embodiment 5:B=1:4) | Y=1.3223x+7.1449 | 0.0239 | 0.86 | 1.55 |
" A in table 1:B=4:Compounding containing bacillus subtilis BAB-1 and Boscalid described in 1 " expression embodiment 1 is killed
Microbial inoculum wherein Boscalid and bacillus subtilis BAB-1 part by weight are 4:1.Other are similar therewith.
Experimental example 2
Synergistic effect of the composite bactericide containing bacillus subtilis BAB-1 and Boscalid to Fulvia fulva is surveyed
It is fixed
1 experiment material
Experimental pharmacy：The compound sterilizing containing bacillus subtilis BAB-1 and Boscalid prepared according to embodiment 8
Agent.
Comparison medicament 1：(BASF European Co., Ltd provides the Boscalid water dispersible granules of mass concentration 50%, lot number
140820000)。
Comparison medicament 2：Bacillus subtilis BAB-1 zymotic fluids (Inst. of Plant Protection, Hebei-Prov. Academy of Agricultural and Forestry Scie provides,
Total viable count is 8,000,000,000 CFU/mL).
It is Fulvia fulva (Fulvia fulva (Cooke) Cif.) for examination cause of disease bacteria strain, by agricultural section of Hebei province
Institute's Plant Protection Institute provides, and uses PDA culture medium (potato 200g, glucose 20g, agar 10g, distilled water 1000mL)
The preculture 20d under the conditions of 24 DEG C ± 1 DEG C, produce spore after it is standby.
2 experimental methods
Comparison medicament 1 is dissolved in proper amount of acetone by 2.1, and the mother liquor that concentration is 10000 μ g/mL is made, is then entered with sterilized water
Row is serially diluted, and is mixed with PDA culture medium by 1: 9 volume ratio, it is 10,5,1,0.5,0.1,0.05 μ that concentration, which is respectively prepared,
G/mL drug containing flat board.
2.2 are diluted bacillus subtilis BAB-1 zymotic fluids with sterilized water, and 1: 9 volume ratio is pressed with PDA culture medium
Example mixing, the drug containing flat board that bacillus subtilis bacteria concentration is 10,5,1,0.5,0.1,0.05 μ g/mL is respectively prepared.
The 2.3 composite bactericide nothings containing bacillus subtilis BAB-1 and Boscalid that will be prepared according to embodiment 8
Bacterium water dilutes, and is mixed respectively with PDA culture medium by 1: 9 volume ratio, it is 10,5,1,0.5,0.1,0.05 μ g/ that concentration, which is made,
ML drug containing flat board.Concentration refers to active component in the composite bactericide (i.e. bacillus subtilis BAB-1 and pyridine acyl bacterium herein
Amine) concentration.
2.4 above-mentioned through on the preculture Fulvia fulva bacterium PDA plate of 20 days, add the tween water of 10mL 0.5%,
Leaf mold spore is scraped with rod is applied, using microscope, spore suspension is adjusted to 1 × 105Individual/mL, using pipettor to above-mentioned
0.1mL spore suspensions are injected in drug containing flat board prepared by step 2.1~2.3, are then smoothened with painting rod.And with without medicament
PDA plate is blank control, is operated in the same way；After incubated 20h under the conditions of 24 DEG C ± 1 DEG C, in low-powered microscope
The sum and sprouting number of lower investigation spore, about 100 spores of often processing investigation.
3 results calculate
Spore germination rate and inhibiting rate is calculated as follows, empirically the method for example 1 calculate virulence regression equation, coefficient correlation,
EC50(effectively suppressing concentration for the treatment of during pathogen growth 50%, unit is μ g/mL) and coefficient of synergism.
Germination rate (%)=sprouting number/always investigate number × 100%
Inhibiting rate (%)=(control germination rate-processing germination rate)/control germination rate × 100%
4 experimental results
It is shown in Table 2.As a result show, contain bacillus subtilis described in Boscalid, bacillus subtilis BAB-1 and embodiment 8
The composite bactericide of bacterium BAB-1 and Boscalid has good inhibitory action to Fulvia fulva bacterium respectively；And implement
Composite bactericide described in example 8 is 1.90 to the coefficient of synergism of Fulvia fulva, shows to contain bacillus subtilis described in embodiment 8
Boscalid has synergistic effect with bacillus subtilis BAB-1 in the composite bactericide of bacterium BAB-1 and Boscalid.
Joint poison of composite bactericide of the table 2 containing bacillus subtilis BAB-1 and Boscalid to Fulvia fulva
Power
" A in table 2:B=1:Compounding containing bacillus subtilis BAB-1 and Boscalid described in 2 " expression embodiments 8 is killed
Microbial inoculum wherein Boscalid and bacillus subtilis BAB-1 part by weight are 1:2.
Experimental example 3
Field efficacy of the composite bactericide containing bacillus subtilis BAB-1 and Boscalid to graw mold of tomato
Test place Hebei province Baoding Xushui County Gao Lin villages and small towns White Tower paving village.Stubble is cucumber before sample plot, tomato planting
More than 15 years, gray mold occurred over the years.It is holy for east for examination tomato variety, straw mat insulation is covered outside experiment greenhouse, is rolled daytime.When
When temperature of shed is too low, heated up using stove fire.On January 18th, 2014 is colonized, 3000 plants of tomato seedlings of every mu of field planting.
1 experiment material
Experimental pharmacy：The compounding containing bacillus subtilis BAB-1 and Boscalid prepared according to the method for embodiment 8 is killed
Microbial inoculum.Comparison medicament 1：(BASF European Co., Ltd provides the Boscalid water dispersible granules of mass concentration 50%, lot number
140820000.Comparison medicament 2：Bacillus subtilis BAB-1 dry powder (Inst. of Plant Protection, Hebei-Prov. Academy of Agricultural and Forestry Scie provides,
Total viable count is 20,000,000,000 CFU/g).Comparison medicament 3：Concentration is 400g/L pyrimethanils (Bayer Bitterfeld GmbH crop science company
There is provided, lot number 20131127).Comparison medicament 4：The difenoconazole water dispersible granule of mass concentration 10% (is first just made up to (Suzhou)
Thing protection Co., Ltd provides, lot number SJ14BWG004).Blank control：Water.
2 experimental methods
Experiment sets 7 processing altogether：After experimental pharmacy is diluted with water, respectively by amount of formulation 750g/hm2、1125g/hm2With
1500g/hm2Spraying medication；After comparison medicament 1 (50% Boscalid water dispersible granules) is diluted with water, by amount of formulation 720g/
hm2Spraying medication；After comparison medicament 2 (20,000,000,000 CFU/mL bacillus subtilis BAB-1 dry powder) is diluted with water, by 3600g/
hm2Spraying medication；After comparison medicament 3 (concentration 400g/L pyrimethanils) is diluted with water, by amount of formulation 1500mL/hm2Spray
Mist medication；After comparison medicament 4 (mass concentration is 10% difenoconazole water dispersible granule) is diluted with water, by amount of formulation
600g/hm2Spraying medication；Using clear water as blank control.Experimental plot is arranged using random district's groups, and often processing sets 4 repetitions, small
Area area 13m2.First time medicine was applied March 1, March 10 and March 17 apply second and third time medicine respectively, during experiment
Share medicine 3 times.Sprayed using " bodyguard " board knapsack hand sprayer, spray uniformly thoughtful, water consumption 900L/hm2。
Gray mold sporadicly occurs before dispenser, and radix is considered as zero, and 7d investigates incidence after last medication.Calculate disease index
And prevention effect.Take 5 points of investigation at random per cell, investigate 2 plants, every plant of whole fruits of investigation, records disease fruit number at every.Disease fruit
Grade scale is：
0 grade：Disease-free spot；
1 grade：Remain petal morbidity or column cap morbidity；
3 grades：Sepal rots or column cap morbidity spreads to areola portion；
5 grades：There is infiltration spot in areola portion but without mould layer；
7 grades：There is mould layer in areola portion but does not expand to other positions；
9 grades：The mould layer in areola portion expands to other positions.
Diseased fruit rate (%)=(the total fruit number of disease fruit number/investigation) × 100
Disease index=∑ [(disease fruit number × relative disease series)/(investigating total fruit number × highest series)] × 100
Prevention effect (%)=[(control disease index-processing disease index)/control disease index] × 100
3 experimental results are shown in Table 3.
Experimental example 4
Field efficacy of the composite bactericide containing bacillus subtilis BAB-1 and Boscalid to leaf muld of tomato
Test place Hebei province Baoding Xushui County Gao Lin villages and small towns White Tower paving village.Stubble is cucumber before sample plot, tomato planting
More than 15 years, leaf mold occurred over the years.It is Caesar for examination tomato variety.Test and straw mat insulation is covered outside greenhouse, roll daytime.When
When temperature of shed is too low, heated up using stove fire.On January 18th, 2014 is colonized, 3000 plants of tomato seedlings of every mu of field planting.
1 experiment material
With experimental example 3.
2 experimental methods
Experimental pharmacy, comparison medicament and dosage are the same as experimental example 3.
First time medicine was applied March 4, apply second and third time medicine respectively on March 14 and March 21, during experiment altogether
Medication 3 times.Sprayed using " bodyguard " board knapsack hand sprayer, spray uniformly thoughtful, water consumption 900L/hm2。
Leaf mold sporadicly occurs radix and is considered as zero before dispenser, and 7d investigates incidence after last medication.Calculate disease index
And prevention effect.Take 3 points of investigation at random per cell, every 5 plants of investigation, every plant point of upper, middle and lower part take one it is representative
Compound leaf, whole blade area percentage is accounted for every vanelets lesion area on compound leaf and is classified, stage division is：
0 grade：Disease-free spot；
1 grade：Lesion area accounts for less than the 5% of whole blade area；
3 grades：Lesion area accounts for the 5%~10% of whole blade area；
5 grades：Lesion area accounts for the 11%~20% of whole blade area；
7 grades：Lesion area accounts for the 21%~50% of whole blade area；
9 grades：Lesion area accounts for more than the 51% of whole blade area.
Disease index=∑ [(disease fruit number × relative disease series)/(investigating total fruit number × highest series)] × 100
Prevention effect (%)=[(control disease index-processing disease index)/control disease index] × 100
3 experimental results are shown in Table 3.
As shown in table 3, field experiment result shows：Comparison medicament 1 (use by the Boscalid water dispersible granules of single dose 50%, mu
48g is measured, dilutes 1250 times) it is respectively 83.6% and 82.3% to the prevention effect of graw mold of tomato and leaf mold.Comparison medicament 2
(the bacillus subtilis BAB-1 dry powder of single dose 20,000,000,000, mu dosage 240g, diluting 250 times) is prevented graw mold of tomato and leaf mold
Control effect difference 76.7% and 77.8%；Experimental drug (the answering containing bacillus subtilis BAB-1 and Boscalid of embodiment 8
With bactericide, mu dosage 100g, 600 times are diluted), respectively 88.6% He best to the preventive effect of graw mold of tomato and leaf mold
87.5%；Preventing and treating of the comparison medicament 3 (400g/L pyrimethanils, mu dosage 100mL, diluting 600 times) to graw mold of tomato
Effect is 62.3%.
Experimental drug (composite bactericide of the embodiment 8 containing bacillus subtilis BAB-1 and Boscalid) is in mu dosage
When 100 grams (600 times of dilution), the dosage of wherein chemical bactericide Boscalid is about the 83.3% of its single dose Boscalid；Institute
The dosage for stating bacillus subtilis BAB-1 in composite bactericide is about the 16.7% of its single dose bacillus subtilis BAB-1, but right
The prevention effect of gray mold is suitable with its single dose Boscalid, but is better than its single dose Boscalid to the prevention effect of leaf mold.
Experimental drug (composite bactericide of the embodiment 8 containing bacillus subtilis BAB-1 and Boscalid) is in mu dosage
When 50 grams (1200 times of dilution), the dosage of wherein chemical bactericide Boscalid is about the 41.7% of its single dose Boscalid；Institute
The dosage for stating bacillus subtilis BAB-1 in composite bactericide is about the 8.3% of its single dose bacillus subtilis BAB-1, described
Composite bactericide is respectively 78.7% and 78.0% to the preventive effect of graw mold of tomato and leaf mold, with comparison medicament 1, comparison medicament
1 two single doses are suitable to the prevention effect of gray mold and leaf mold；Conventional dose is significantly higher than to the prevention effect of gray mold
400g/L pyrimethanils (mu dosage 100mL, 600 times of dilutions)；To the prevention effect and the benzene of conventional dose 10% of leaf mold
Ether methyl cyclic-azole water dispersible granules (40 grams of mu dosage, 1500 times of dilutions) are not significantly different.
Table 3 is to graw mold of tomato and the field control effect of leaf mold
Various embodiments above is not the concrete restriction to the present invention, as long as the scope limited according to claim, at this specially
Under the enlightenment of profit, with reference to the basic general knowledge of this area, the compounding microbial inoculum is used in the disease control of the crops such as tomato, all belonged to
In protection scope of the present invention.
Although above the present invention is described in detail with a general description of the specific embodiments,
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 (16)
1. a kind of composite bactericide containing bacillus subtilis BAB-1 and Boscalid, its active component by Boscalid and
Bacillus subtilis (Bacillus subtilis) BAB-1 is formed；
Boscalid and bacillus subtilis BAB-1 weight ratio are 1~4: 4~1 in the active component；
The Boscalid is the Boscalid active compound of mass concentration 98%；The bacillus subtilis BAB-1 is its freeze-dried vaccine
Powder, the total viable count of the lyophilized bacterium powder is 2.0 × 1010CFU/g。
2. composite bactericide according to claim 1, it is characterised in that Boscalid and withered grass bud in the active component
Spore bacillus BAB-1 weight ratio is 4: 1.
3. composite bactericide according to claim 1, it is characterised in that Boscalid and withered grass bud in the active component
Spore bacillus BAB-1 weight ratio is 2: 1.
4. composite bactericide according to claim 1, it is characterised in that Boscalid and withered grass bud in the active component
Spore bacillus BAB-1 weight ratio is 1: 1.
5. composite bactericide according to claim 1, it is characterised in that Boscalid and withered grass bud in the active component
Spore bacillus BAB-1 weight ratio is 1: 2.
6. composite bactericide according to claim 1, it is characterised in that Boscalid and withered grass bud in the active component
Spore bacillus BAB-1 weight ratio is 1: 4.
7. according to the composite bactericide described in claim any one of 1-6, it is characterised in that in parts by weight, its raw material contains
Following components：10~80 parts of active component, 1~8 part of dispersant, 1~8 part of wetting agent, 10~80 parts of filler.
8. composite bactericide according to claim 7, it is characterised in that in parts by weight, its raw material contains following components：
40~70 parts of active component, 3~7 parts of dispersant, 3~7 parts of wetting agent, 20~50 parts of filler.
9. composite bactericide according to claim 8, it is characterised in that in parts by weight, its raw material contains following components：
50~60 parts of active component, 4~6 parts of dispersant, 4~6 parts of wetting agent, 30~40 parts of filler.
10. composite bactericide according to claim 7, it is characterised in that the dispersant is lignin sulfonic acid sodium salt, carboxylic
Methylcellulose sodium salt, desugar sodium lignin sulfonate, lignosulfite, APES, ethylene glycol monobutyl
One or more mixtures in ether, polyoxyethanyl alkyl ether, NPE, neopelex；Institute
The wetting agent stated is nekal, AEO, dodecyl sodium sulfate, neopelex
One or more mixtures in salt, alkylphenol-polyethenoxy base formaldehyde condensate sulfonate；The filler is white carbon, height
One or more mixtures in ridge soil, diatomite, precipitated calcium carbonate, bentonite.
11. composite bactericide according to claim 8 or claim 9, it is characterised in that the dispersant is sodium lignin sulfonate
Salt, sanlose, desugar sodium lignin sulfonate, lignosulfite, APES, ethylene glycol list
One or more mixing in butyl ether, polyoxyethanyl alkyl ether, NPE, neopelex
Thing；Described wetting agent is nekal, AEO, dodecyl sodium sulfate, detergent alkylate sulphur
One or more mixtures in acid sodium-salt, alkylphenol-polyethenoxy base formaldehyde condensate sulfonate；The filler is hard charcoal
One or more mixtures in black, kaolin, diatomite, precipitated calcium carbonate, bentonite.
12. composite bactericide according to claim 1, it is characterised in that in parts by weight, contain following components：Quality
The Boscalid active compound of concentration 98%：20.41 parts；Total viable count is 2.0 × 1010CFU/g bacillus subtilis BAB-1 is freezed
Bacterium powder：40 parts；Dispersant sanlose：3 parts；Wetting agent：Dodecyl sodium sulfate：4.5 part；Filler lightweight carbonic acid
Calcium：32.09 parts.
13. the preparation method of any one of the claim 1-12 composite bactericides, it is characterised in that comprise the following steps：Press
Various components are mixed according to the proportioning, carry out one-level crushing, grain fineness reaches 80 mesh；After mixing, then through gas
Flow pulverizer and carry out two level crushing, grain fineness reaches 325 mesh, produced.
14. application of any one of the claim 1-12 composite bactericides in terms of preventing and treating plant botrytis and leaf mold.
15. application according to claim 14, it is characterised in that the plant includes but is not limited to water fruits and vegetables and flower
Grass.
16. application according to claim 15, it is characterised in that the plant is tomato.
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| CN105838656B (en) * | 2016-06-15 | 2019-09-24 | 东北农业大学 | The degeneration-resistant bacillus of broad-spectrum disease resistance growth-promoting and its application for preventing and treating graw mold of tomato and leaf mold |
| CN106818881A (en) * | 2016-12-30 | 2017-06-13 | 西北农林科技大学 | A kind of complex preparation of bacillus subtilis and chemical pesticide and preparation method thereof |
| CN106942276A (en) * | 2017-03-16 | 2017-07-14 | 河北省农林科学院植物保护研究所 | Bacillus subtilis BAB 1 prevents and treats the application of cucurbits powdery mildew |
| CN107079928B (en) * | 2017-05-19 | 2020-07-10 | 安徽省农业科学院植物保护与农产品质量安全研究所 | Bactericidal composition containing bacillus amyloliquefaciens and boscalid |
| CN107494568B (en) * | 2017-08-14 | 2021-04-06 | 河北省农林科学院植物保护研究所 | Compound synergistic bactericidal composition for preventing and treating main diseases of tomatoes and pears and application thereof |
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