CN110184198A - A kind of preparation method of the microbial bacterial agent of controlling plant diseases - Google Patents
A kind of preparation method of the microbial bacterial agent of controlling plant diseases Download PDFInfo
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- CN110184198A CN110184198A CN201910407764.5A CN201910407764A CN110184198A CN 110184198 A CN110184198 A CN 110184198A CN 201910407764 A CN201910407764 A CN 201910407764A CN 110184198 A CN110184198 A CN 110184198A
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Abstract
The invention discloses a kind of preparation methods of the microbial bacterial agent of controlling plant diseases, it is related to microbial control plant disease technical field, steps are as follows: 1) preparing to include following raw material: the secondary bacterium solution of Paecilomyces lilacinus, bacillus licheniformis-Sphingobacterium mixed fermentation liquid, Gram positive actinomycetes-Bacillus cercus mixed fermentation liquid, nitrogen-fixing bacteria, bacillus megaterium, photosynthetic bacteria, amino acid, humic acid, monosaccharide, chitin, modification infusorial earth, 2) bacillus licheniformis-Sphingobacterium mixed fermentation liquid；3) Gram positive actinomycetes-Bacillus cercus mixed fermentation liquid is prepared；4) mixed bacteria liquid A is prepared；5) surplus stock being added to prepare in mixed bacteria liquid A and obtains the microbial bacterial agent of controlling plant diseases, microbial bacterial agent of the invention contains multiple beneficial bacteria, and it is antibacterial with bacterium, with bacterium gram worm, it can effectively eliminate root-knot nematode and worm's ovum in soil；The growth and breeding of soil-borne disease source bacterium can effectively be inhibited.
Description
Technical field
The present invention relates to microbial control plant disease technical field, the microbial bacteria of specifically a kind of controlling plant diseases
The preparation method of agent.
Background technique
Microorganism includes: one big including bacterium, virus, fungi and some small-sized protists, microalga etc.
Class biocenose, its individual is small, close with human relation.It covers beneficial to harmful numerous kinds, relates generally to eat
The numerous areas such as product, medicine, industrial or agricultural, environmental protection, sport.In the textbook of China, it is big that microorganism is divided into following 8
Class: bacterium, virus, fungi, actinomyces, rickettsia, mycoplasma, Chlamydia, conveyor screw.Some microorganisms are that naked eyes can
With what is seen, as the mushroom, ganoderma lucidum, mushroom etc. for belonging to fungi.It is a kind of several by nucleic acid and protein etc. there are also microorganism
Kind is at " acellular organism " being grouped as, but its existence is necessarily dependent upon living cells.
Present plant prevention disease is mainly prevented and treated by chemical pesticide, but long-term largely using chemical agriculture
Medicine, bring pathogen resistance, agricultural product medicament residual and a series of problems, such as environmental pollution, seriously constrain agricultural can
Sustainable development, while threaten the health of the mankind.
Summary of the invention
The purpose of the present invention is to provide a kind of preparation methods of the microbial bacterial agent of controlling plant diseases, above-mentioned to solve
Problem.
To achieve the above object, the invention provides the following technical scheme:
A kind of preparation method of the microbial bacterial agent of controlling plant diseases, steps are as follows:
1) prepare include below according to the raw material of parts by weight: Paecilomyces lilacinus is bacterium solution 10-20 parts secondary, lichens gemma bar
3-5 parts of bacterium, 3-5 parts of Sphingobacterium, 2-4 parts of Gram positive actinomycetes, 2-4 parts of Bacillus cercus, 6-8 parts of nitrogen-fixing bacteria,
2-6 parts of bacillus megaterium, 2-4 parts of photosynthetic bacteria, 8-12 parts of amino acid, 1-3 parts of humic acid, 1-3 parts of monosaccharide, chitin 3-7
Part, 4-6 parts of modification infusorial earth；
2) bacillus licheniformis-Sphingobacterium mixed fermentation liquid is made in bacillus licheniformis and Sphingobacterium；
3) Gram positive actinomycetes-Bacillus cercus is made in Gram positive actinomycetes and Bacillus cercus to mix
Close fermentation liquid；
4) bacillus licheniformis obtained in the secondary bacterium solution of Paecilomyces lilacinus, step 2)-Sphingobacterium is mixed into hair
The mixing of Gram positive actinomycetes obtained in zymotic fluid and step 3)-Bacillus cercus mixed fermentation liquid, while stirring plus
Enter nitrogen-fixing bacteria, bacillus megaterium, photosynthetic bacteria, is uniformly mixed, obtains mixed bacteria liquid A；
5) amino acid, humic acid, monosaccharide, chitin, modification infusorial earth are added in mixed bacteria liquid A, after mixing evenly,
High temperature spray-drying is carried out, temperature is 160-180 DEG C, and feed flow rate 2-4L/min finally obtains the micro- of controlling plant diseases
Bacteria agent.
Based on the above technical solution, the present invention also provides following optional technical solutions:
In a kind of optinal plan: the secondary bacterium solution of the step 1) Paecilomyces lilacinus the preparation method is as follows: by pale purple
Paecilomycerol strain is inoculated in the first culture medium, and culture obtains fluorescence bacterium solution of unicellular bacillus, then by pale purple quasi- blueness
Bacterium solution of mould is inoculated in the second culture medium, and culture obtains the secondary bacterium solution of Paecilomyces lilacinus.
In a kind of optinal plan: first culture medium includes the following raw material according to parts by weight: 2-20 parts of peptone,
0.5-1.5 parts of dipotassium hydrogen phosphate, 0.5-1.8 parts of magnesium sulfate, 10-25 parts of agar, 1000 parts of sterile water, first culture medium
PH value be 7.2 ± 0.2；Second culture medium includes the following raw material according to parts by weight: 5-20 parts of peptone, yeast extract
0.5-5 parts, 5-10 parts of sodium chloride, 10-20 parts of agar, 1000 parts of distilled water；The PH value of second culture medium is 7.0.
In a kind of optinal plan: the step 2) bacillus licheniformis-Sphingobacterium mixed fermentation liquid preparation side
Method are as follows: bacillus licheniformis and Sphingobacterium seed liquor are seeded in PDB fluid nutrient medium, and in 24-28 DEG C of temperature strip
Aerobic fermentation culture 24-96h is carried out under part, obtains required bacillus licheniformis-Sphingobacterium mixed fermentation liquid.
In a kind of optinal plan: Gram positive actinomycetes described in step 3)-Bacillus cercus mixed fermentation liquid
The preparation method comprises the following steps: by Gram positive actinomycetes, Bacillus cercus using oese picking colony be seeded to respectively LB train
It supports and is cultivated in base, cultivation temperature is 32 DEG C, and hunting speed is 120rpm in reciprocating oscillator, after culture for 24 hours, is turned
Fermentation medium is connect, fermentation 24-36h is carried out, inoculum concentration is the 1-3% of fermentation total volume.
In a kind of optinal plan: monosaccharide described in step 1) is sucrose or glucose.
In a kind of optinal plan: modification infusorial earth described in step 1) the preparation method comprises the following steps: diatomite is added to PH
In the nitric acid solution of=0.01-0.5, the slurries that weight percent concentration is 20-50% are configured to, boils, cool down, filter, wash
It washs, dry, calcine, obtain modification infusorial earth, wherein drying temperature is 60-120 DEG C, dry 10-30h, calcination temperature 400-
650 DEG C, calcination time 10-20h.
Compared to the prior art, beneficial effects of the present invention are as follows:
1, the present invention is by being added multiple beneficial bacteria, antibacterial with bacterium, with bacterium gram by preparing fermented by mixed bacterium liquid
Worm further promotes the mutual synergisticing performance between each strain, can effectively eliminate root-knot nematode and worm's ovum in soil；It can be effective
Inhibit the growth and breeding of soil-borne disease source bacterium；
2, using humic acid as protective agent, using diatomite as filler, can effectively improve microbial cells germination rate and
Survival rate, to provide certain guarantee for microorganism long-acting bacteriostatic；
3, the supply that plant nutrient element is increased by addition amino acid, monosaccharide, improves nutrient availability in soil,
Promote root growth and Nutrient Absorption, protect root system and increase Genes For Plant Tolerance adverse circumstance ability, there is the ability for decomposing chitin, it can
The cell wall of pathogen is destroyed, worm's ovum shell is decomposed and protects plant.
Specific embodiment
Following embodiment can be described in detail the present invention, and each embodiment cited by the present invention is only to illustrate the present invention,
It is not used to limit the scope of the present invention.Any modification apparent easy to know or change made for the present invention are without departure from this hair
Bright spirit and scope.
Embodiment 1
A kind of preparation method of the microbial bacterial agent of controlling plant diseases, steps are as follows:
1) prepare to include below according to the raw material of parts by weight: 10 parts of the secondary bacterium solution of Paecilomyces lilacinus, bacillus licheniformis 3
Part, 3 parts of Sphingobacterium, 2 parts of Gram positive actinomycetes, 2 parts of Bacillus cercus, 6 parts of nitrogen-fixing bacteria, bacillus megaterium 2
Part, 2 parts of photosynthetic bacteria, 8 parts of amino acid, 1 part of humic acid, 1 part of monosaccharide, 3 parts of chitin, 4 parts of modification infusorial earth.
2) bacillus licheniformis-Sphingobacterium mixed fermentation liquid is made in bacillus licheniformis and Sphingobacterium；
3) Gram positive actinomycetes-Bacillus cercus is made in Gram positive actinomycetes and Bacillus cercus to mix
Close fermentation liquid；
4) bacillus licheniformis obtained in the secondary bacterium solution of Paecilomyces lilacinus, step 2)-Sphingobacterium is mixed into hair
The mixing of Gram positive actinomycetes obtained in zymotic fluid and step 3)-Bacillus cercus mixed fermentation liquid, while stirring plus
Enter nitrogen-fixing bacteria, bacillus megaterium, photosynthetic bacteria, is uniformly mixed, obtains mixed bacteria liquid A；
5) amino acid, humic acid, monosaccharide, chitin, modification infusorial earth are added in mixed bacteria liquid A, after mixing evenly,
High temperature spray-drying is carried out, temperature is 160 DEG C, and feed flow rate 2L/min finally obtains the microbial bacteria of controlling plant diseases
Agent.
Wherein, the secondary bacterium solution of Paecilomyces lilacinus described in step 1) the preparation method is as follows: by Paecilomyces lilacinus bacterium
Kind is inoculated in the first culture medium, and culture obtains fluorescence bacterium solution of unicellular bacillus, then by bacterium of Paecilomyces lilacinus
Liquid is inoculated in the second culture medium, and culture obtains the secondary bacterium solution of Paecilomyces lilacinus.
First culture medium includes the following raw material according to parts by weight: 2 parts of peptone, 0.5 part of dipotassium hydrogen phosphate, sulfuric acid
0.5 part of magnesium, 10 parts of agar, 1000 parts of sterile water, the pH value of first culture medium are 7.0；Second culture medium include with
Under according to parts by weight raw material: 5 parts of peptone, 0.5 part of yeast extract, 5 parts of sodium chloride, 10 parts of agar, 1000 parts of distilled water；Institute
The pH value for stating the second culture medium is 7.0.
Bacillus licheniformis described in step 2)-Sphingobacterium mixed fermentation liquid is the preparation method comprises the following steps: by lichens gemma
Bacillus and Sphingobacterium seed liquor are seeded in PDB fluid nutrient medium, and aerobic fermentation training is carried out under the conditions of 24 DEG C of temperature
It supports for 24 hours, obtains required bacillus licheniformis-Sphingobacterium mixed fermentation liquid.
Gram positive actinomycetes described in step 3)-Bacillus cercus mixed fermentation liquid is the preparation method comprises the following steps: will leather
Lan Shi positive actinomyces, Bacillus cercus are seeded in LB culture medium respectively using oese picking colony and are cultivated, training
Supporting temperature is 32 DEG C, and hunting speed is 120rpm in reciprocating oscillator, and after culture for 24 hours, fermentation medium of transferring is carried out
For 24 hours, inoculum concentration is the 1% of fermentation total volume for fermentation.
Monosaccharide described in step 1) is sucrose.
Modification infusorial earth described in step 1) the preparation method comprises the following steps: diatomite to be added to the nitric acid solution of PH=0.01
In, the slurries that weight percent concentration is 20% are configured to, boils, cool down, filter, wash, dry, calcine, be modified
Diatomite, wherein drying temperature is 60 DEG C, and dry 10h, calcination temperature is 400 DEG C, calcination time 10h.
Embodiment 2
A kind of preparation method of the microbial bacterial agent of controlling plant diseases, steps are as follows:
1) prepare to include below according to the raw material of parts by weight: 12 parts of the secondary bacterium solution of Paecilomyces lilacinus, bacillus licheniformis-
7 parts of Sphingobacterium mixed fermentation liquid, Gram positive actinomycetes -5 parts of Bacillus cercus mixed fermentation liquid, nitrogen-fixing bacteria 6.5
Part, 2.5 parts of photosynthetic bacteria, 9 parts of amino acid, 1.5 parts of humic acid, 1.5 parts of monosaccharide, 4 parts of chitin, changes 3 parts of bacillus megaterium
4.5 parts of diatomite of property.
2) bacillus licheniformis-Sphingobacterium mixed fermentation liquid is made in bacillus licheniformis and Sphingobacterium；
3) Gram positive actinomycetes-Bacillus cercus is made in Gram positive actinomycetes and Bacillus cercus to mix
Close fermentation liquid；
4) bacillus licheniformis obtained in the secondary bacterium solution of Paecilomyces lilacinus, step 2)-Sphingobacterium is mixed into hair
The mixing of Gram positive actinomycetes obtained in zymotic fluid and step 3)-Bacillus cercus mixed fermentation liquid, while stirring plus
Enter nitrogen-fixing bacteria, bacillus megaterium, photosynthetic bacteria, is uniformly mixed, obtains mixed bacteria liquid A；
5) amino acid, humic acid, monosaccharide, chitin, modification infusorial earth are added in mixed bacteria liquid A, after mixing evenly,
High temperature spray-drying is carried out, temperature is 165 DEG C, and feed flow rate 2.5L/min finally obtains the microorganism of controlling plant diseases
Microbial inoculum.
Wherein, the secondary bacterium solution of Paecilomyces lilacinus described in step 1) the preparation method is as follows: by Paecilomyces lilacinus bacterium
Kind is inoculated in the first culture medium, and culture obtains fluorescence bacterium solution of unicellular bacillus, then by bacterium of Paecilomyces lilacinus
Liquid is inoculated in the second culture medium, and culture obtains the secondary bacterium solution of Paecilomyces lilacinus.
First culture medium includes the following raw material according to parts by weight: 2 parts of peptone, 0.5 part of dipotassium hydrogen phosphate, sulfuric acid
0.5 part of magnesium, 10 parts of agar, 1000 parts of sterile water, the pH value of first culture medium are 7.0；Second culture medium include with
Under according to parts by weight raw material: 5 parts of peptone, 0.5 part of yeast extract, 5 parts of sodium chloride, 10 parts of agar, 1000 parts of distilled water；Institute
The pH value for stating the second culture medium is 7.0.
Bacillus licheniformis described in step 2)-Sphingobacterium mixed fermentation liquid is the preparation method comprises the following steps: by lichens gemma
Bacillus and Sphingobacterium seed liquor are seeded in PDB fluid nutrient medium, and aerobic fermentation training is carried out under the conditions of 25 DEG C of temperature
36h is supported, required bacillus licheniformis-Sphingobacterium mixed fermentation liquid is obtained.
Gram positive actinomycetes described in step 3)-Bacillus cercus mixed fermentation liquid is the preparation method comprises the following steps: will leather
Lan Shi positive actinomyces, Bacillus cercus are seeded in LB culture medium respectively using oese picking colony and are cultivated, training
Supporting temperature is 32 DEG C, and hunting speed is 120rpm in reciprocating oscillator, and after culture for 24 hours, fermentation medium of transferring is carried out
Ferment 28h, and inoculum concentration is the 1.5% of fermentation total volume.
Monosaccharide described in step 1) is glucose.
Modification infusorial earth described in step 1) the preparation method comprises the following steps: diatomite to be added to the nitric acid solution of PH=0.01
In, the slurries that weight percent concentration is 30% are configured to, boils, cool down, filter, wash, dry, calcine, be modified
Diatomite, wherein drying temperature is 75 DEG C, and dry 15h, calcination temperature is 450 DEG C, calcination time 12h.
Embodiment 3
A kind of preparation method of the microbial bacterial agent of controlling plant diseases, steps are as follows:
1) prepare to include below according to the raw material of parts by weight: 15 parts of the secondary bacterium solution of Paecilomyces lilacinus, bacillus licheniformis-
8 parts of Sphingobacterium mixed fermentation liquid, Gram positive actinomycetes -6 parts of Bacillus cercus mixed fermentation liquid, nitrogen-fixing bacteria 7
Part, 4 parts of bacillus megaterium, 3 parts of photosynthetic bacteria, 0 part of amino acid 1,2 parts of humic acid, 2 parts of monosaccharide, 5 parts of chitin, modified diatom
5 parts of soil；
2) bacillus licheniformis-Sphingobacterium mixed fermentation liquid is made in bacillus licheniformis and Sphingobacterium；
3) Gram positive actinomycetes-Bacillus cercus is made in Gram positive actinomycetes and Bacillus cercus to mix
Close fermentation liquid；
4) bacillus licheniformis obtained in the secondary bacterium solution of Paecilomyces lilacinus, step 2)-Sphingobacterium is mixed into hair
The mixing of Gram positive actinomycetes obtained in zymotic fluid and step 3)-Bacillus cercus mixed fermentation liquid, while stirring plus
Enter nitrogen-fixing bacteria, bacillus megaterium, photosynthetic bacteria, is uniformly mixed, obtains mixed bacteria liquid A；
5) amino acid, humic acid, monosaccharide, chitin, modification infusorial earth are added in mixed bacteria liquid A, after mixing evenly,
High temperature spray-drying is carried out, temperature is 170 DEG C, and feed flow rate 3L/min finally obtains the microbial bacteria of controlling plant diseases
Agent.
Wherein, the secondary bacterium solution of Paecilomyces lilacinus described in step 1) the preparation method is as follows: by Paecilomyces lilacinus bacterium
Kind is inoculated in the first culture medium, and culture obtains fluorescence bacterium solution of unicellular bacillus, then by bacterium of Paecilomyces lilacinus
Liquid is inoculated in the second culture medium, and culture obtains the secondary bacterium solution of Paecilomyces lilacinus.
First culture medium includes the following raw material according to parts by weight: 10 parts of peptone, 1 part of dipotassium hydrogen phosphate, sulfuric acid
1.2 parts of magnesium, agar 16,1000 parts of sterile water, the pH value of first culture medium are 7.2；Second culture medium includes following
According to the raw material of parts by weight: 12 parts of peptone, 3 parts of yeast extract, 7 parts of sodium chloride, 15 parts of agar, 1000 parts of distilled water；Described
The pH value of two culture mediums is 7.0.
Bacillus licheniformis described in step 2)-Sphingobacterium mixed fermentation liquid is the preparation method comprises the following steps: by lichens gemma
Bacillus and Sphingobacterium seed liquor are seeded in PDB fluid nutrient medium, and aerobic fermentation training is carried out under the conditions of 25 DEG C of temperature
48h is supported, required bacillus licheniformis-Sphingobacterium mixed fermentation liquid is obtained.
Gram positive actinomycetes described in step 3)-Bacillus cercus mixed fermentation liquid is the preparation method comprises the following steps: will leather
Lan Shi positive actinomyces, Bacillus cercus are seeded in LB culture medium respectively using oese picking colony and are cultivated, training
Supporting temperature is 32 DEG C, and hunting speed is 120rpm in reciprocating oscillator, and after culture for 24 hours, fermentation medium of transferring is carried out
Ferment 30h, and inoculum concentration is the 2% of fermentation total volume.
Monosaccharide described in step 1) is glucose.
Modification infusorial earth described in step 1) the preparation method comprises the following steps: diatomite is added in the nitric acid solution of PH=0.3,
The slurries that weight percent concentration is 35% are configured to, boils, cool down, filter, wash, dry, calcine, obtain modified diatom
Soil, wherein drying temperature is 90 DEG C, and dry 20h, calcination temperature is 500 DEG C, calcination time 15h.
Embodiment 4
A kind of preparation method of the microbial bacterial agent of controlling plant diseases, steps are as follows:
1) prepare to include below according to the raw material of parts by weight: 18 parts of the secondary bacterium solution of Paecilomyces lilacinus, bacillus licheniformis-
9 parts of Sphingobacterium mixed fermentation liquid, Gram positive actinomycetes -7 parts of Bacillus cercus mixed fermentation liquid, nitrogen-fixing bacteria 7.5
Part, 3.5 parts of photosynthetic bacteria, 0 part of amino acid 1,2.5 parts of humic acid, 2.5 parts of monosaccharide, 6 parts of chitin, changes 5 parts of bacillus megaterium
5.5 parts of diatomite of property；
2) bacillus licheniformis-Sphingobacterium mixed fermentation liquid is made in bacillus licheniformis and Sphingobacterium；
3) Gram positive actinomycetes-Bacillus cercus is made in Gram positive actinomycetes and Bacillus cercus to mix
Close fermentation liquid；
4) bacillus licheniformis obtained in the secondary bacterium solution of Paecilomyces lilacinus, step 2)-Sphingobacterium is mixed into hair
The mixing of Gram positive actinomycetes obtained in zymotic fluid and step 3)-Bacillus cercus mixed fermentation liquid, while stirring plus
Enter nitrogen-fixing bacteria, bacillus megaterium, photosynthetic bacteria, is uniformly mixed, obtains mixed bacteria liquid A；
5) amino acid, humic acid, monosaccharide, chitin, modification infusorial earth are added in mixed bacteria liquid A, after mixing evenly,
High temperature spray-drying is carried out, temperature is 175 DEG C, and feed flow rate 3.5L/min finally obtains the microorganism of controlling plant diseases
Microbial inoculum.
Wherein, the secondary bacterium solution of Paecilomyces lilacinus described in step 1) the preparation method is as follows: by Paecilomyces lilacinus bacterium
Kind is inoculated in the first culture medium, and culture obtains fluorescence bacterium solution of unicellular bacillus, then by bacterium of Paecilomyces lilacinus
Liquid is inoculated in the second culture medium, and culture obtains the secondary bacterium solution of Paecilomyces lilacinus.
First culture medium includes the following raw material according to parts by weight: 20 parts of peptone, 1.5 parts of dipotassium hydrogen phosphate, sulphur
1.8 parts of sour magnesium, 25 parts of agar, 1000 parts of sterile water, the pH value of first culture medium are 7.3；Second culture medium includes
Below according to the raw material of parts by weight: 20 parts of peptone, 0.5 part of yeast extract, 10 parts of sodium chloride, 20 parts of agar, distilled water 1000
Part；The pH value of second culture medium is 7.0.
Bacillus licheniformis described in step 2)-Sphingobacterium mixed fermentation liquid is the preparation method comprises the following steps: by lichens gemma
Bacillus and Sphingobacterium seed liquor are seeded in PDB fluid nutrient medium, and aerobic fermentation training is carried out under the conditions of 28 DEG C of temperature
80h is supported, required bacillus licheniformis-Sphingobacterium mixed fermentation liquid is obtained.
Gram positive actinomycetes described in step 2)-Bacillus cercus mixed fermentation liquid is the preparation method comprises the following steps: will leather
Lan Shi positive actinomyces, Bacillus cercus are seeded in LB culture medium respectively using oese picking colony and are cultivated, training
Supporting temperature is 32 DEG C, and hunting speed is 120rpm in reciprocating oscillator, and after culture for 24 hours, fermentation medium of transferring is carried out
Ferment 32h, and inoculum concentration is the 3% of fermentation total volume.
Monosaccharide described in step 1) is sucrose.
Modification infusorial earth described in step 1) the preparation method comprises the following steps: diatomite is added in the nitric acid solution of PH=0.5,
The slurries that weight percent concentration is 45% are configured to, boils, cool down, filter, wash, dry, calcine, obtain modified diatom
Soil, wherein drying temperature is 110 DEG C, and dry 25h, calcination temperature is 600 DEG C, calcination time 18h.
Embodiment 5
A kind of preparation method of the microbial bacterial agent of controlling plant diseases, steps are as follows:
1) prepare to include below according to the raw material of parts by weight: 20 parts of the secondary bacterium solution of Paecilomyces lilacinus, bacillus licheniformis-
10 parts of Sphingobacterium mixed fermentation liquid, Gram positive actinomycetes -8 parts of Bacillus cercus mixed fermentation liquid, nitrogen-fixing bacteria 8
Part, 6 parts of bacillus megaterium, 4 parts of photosynthetic bacteria, 2 parts of amino acid 1,3 parts of humic acid, 3 parts of monosaccharide, 7 parts of chitin, modified diatom
6 parts of soil；
2) bacillus licheniformis-Sphingobacterium mixed fermentation liquid is made in bacillus licheniformis and Sphingobacterium；
3) Gram positive actinomycetes-Bacillus cercus is made in Gram positive actinomycetes and Bacillus cercus to mix
Close fermentation liquid；
4) bacillus licheniformis obtained in the secondary bacterium solution of Paecilomyces lilacinus, step 2)-Sphingobacterium is mixed into hair
The mixing of Gram positive actinomycetes obtained in zymotic fluid and step 3)-Bacillus cercus mixed fermentation liquid, while stirring plus
Enter nitrogen-fixing bacteria, bacillus megaterium, photosynthetic bacteria, is uniformly mixed, obtains mixed bacteria liquid A；
5) amino acid, humic acid, monosaccharide, chitin, modification infusorial earth are added in mixed bacteria liquid A, after mixing evenly,
High temperature spray-drying is carried out, temperature is 180 DEG C, and feed flow rate 4L/min finally obtains the microbial bacteria of controlling plant diseases
Agent.
Wherein, the secondary bacterium solution of Paecilomyces lilacinus described in step 1) the preparation method is as follows: by Paecilomyces lilacinus bacterium
Kind is inoculated in the first culture medium, and culture obtains fluorescence bacterium solution of unicellular bacillus, then by bacterium of Paecilomyces lilacinus
Liquid is inoculated in the second culture medium, and culture obtains the secondary bacterium solution of Paecilomyces lilacinus.
First culture medium includes the following raw material according to parts by weight: 20 parts of peptone, 1.5 parts of dipotassium hydrogen phosphate, sulphur
1.8 parts of sour magnesium, 25 parts of agar, 1000 parts of sterile water, the pH value of first culture medium are 7.4；Second culture medium includes
Below according to the raw material of parts by weight: 20 parts of peptone, 0.5 part of yeast extract, 10 parts of sodium chloride, 20 parts of agar, distilled water 1000
Part；The pH value of second culture medium is 7.0.
Bacillus licheniformis described in step 2)-Sphingobacterium mixed fermentation liquid is the preparation method comprises the following steps: by lichens gemma
Bacillus and Sphingobacterium seed liquor are seeded in PDB fluid nutrient medium, and aerobic fermentation training is carried out under the conditions of 28 DEG C of temperature
96h is supported, required bacillus licheniformis-Sphingobacterium mixed fermentation liquid is obtained.
Gram positive actinomycetes described in step 3)-Bacillus cercus mixed fermentation liquid is the preparation method comprises the following steps: will leather
Lan Shi positive actinomyces, Bacillus cercus are seeded in LB culture medium respectively using oese picking colony and are cultivated, training
Supporting temperature is 32 DEG C, and hunting speed is 120rpm in reciprocating oscillator, and after culture for 24 hours, fermentation medium of transferring is carried out
Ferment 36h, and inoculum concentration is the 3% of fermentation total volume.
Monosaccharide described in step 1) is glucose.
Modification infusorial earth described in step 1) the preparation method comprises the following steps: diatomite is added in the nitric acid solution of PH=0.5,
The slurries that weight percent concentration is 50% are configured to, boils, cool down, filter, wash, dry, calcine, obtain modified diatom
Soil, wherein drying temperature is 120 DEG C, and dry 30h, calcination temperature is 650 DEG C, calcination time 20h.
Comparative example 1
Compared with Example 3, bacillus licheniformis-Sphingobacterium is not made in bacillus licheniformis and Sphingobacterium
Mixed fermentation liquid, other are same as Example 3；
Comparative example 2
Compared with Example 3, Gram-positive unwrapping wire will not be made in Gram positive actinomycetes and Bacillus cercus
Bacterium-Bacillus cercus mixed fermentation liquid, other are same as Example 3；
Comparative example 3
Compared with Example 3, bacillus licheniformis-sphingol bar neither is made in bacillus licheniformis and Sphingobacterium
Bacterium mixed fermentation liquid, but it is wax-like Gram positive actinomycetes-will not to be made in Gram positive actinomycetes and Bacillus cercus
Bacillus mixed fermentation liquid, other are same as Example 3；
Field controling test method of the microbial bacterial agent of the application preparation to Cucumber root-knot nematode disease: big Tanaka cucumber moves
Processing in 10,17,24,31 days uses the identical microbial bacteria agent prescription for diluting dosage or bacterial manure formula to cucumber after planting field planting
Pouring root or fertilizer treatment are carried out, each processing is arranged 3 cells and repeats；20 plants of root system grab sample.
Investigation grade scale: 0 grade: health of root, no root knot；1 grade: root system has a small amount of root knot, accounts for the 1-25% of full root system；
2 grades: root knot number is medium on root system, accounts for the 26-50% of full root system；3 grades: root knot number is more on root system, accounts for the 51- of full root system
75%；4 grades: there are many root knot number on root system, account for the 76-100% of full root system.
Disease index and control efficiency is calculated by formula.
Disease index (%)=[Σ (each disease grade diseased plant number × disease grade series)/(investigation total strain number × highest series)] ×
100%；
Control efficiency (%)=[(control disease index-processing disease index)/control disease index] × 100%
Experimental result such as following table
| Group | Disease index/% | Preventive effect/% |
| Comparative example 1 | 48.9 | 33.1 |
| Comparative example 2 | 49.5 | 33.4 |
| Comparative example 3 | 66.1 | 11.2 |
| Embodiment 3 | 8.4 | 88.5 |
As can be seen from the above results, the application pass through preparation bacillus licheniformis-Sphingobacterium mixed fermentation liquid
With Gram positive actinomycetes-Bacillus cercus mixed fermentation liquid, the collaboration that can further improve between each strain is matched
Cooperation is used, and the diseases and insect pests resistance of plant is further increased.
The above, the only specific embodiment of the disclosure, but the protection scope of the disclosure is not limited thereto, it is any
Those familiar with the art can easily think of the change or the replacement in the technical scope that the disclosure discloses, and should all contain
It covers within the protection scope of the disclosure.Therefore, the protection scope of the disclosure should be subject to the protection scope in claims.
Claims (7)
1. a kind of preparation method of the microbial bacterial agent of controlling plant diseases, which is characterized in that steps are as follows:
1) prepare include below according to the raw material of parts by weight: Paecilomyces lilacinus is bacterium solution 10-20 parts secondary, bacillus licheniformis-
6-10 parts of Sphingobacterium mixed fermentation liquid, Gram positive actinomycetes -4-8 parts of Bacillus cercus mixed fermentation liquid, fixed nitrogen
6-8 parts of bacterium, 2-6 parts of bacillus megaterium, 2-4 parts of photosynthetic bacteria, 8-12 parts of amino acid, 1-3 parts of humic acid, 1-3 parts of monosaccharide, chitin
3-7 parts of matter, 4-6 parts of modification infusorial earth；
2) bacillus licheniformis-Sphingobacterium mixed fermentation liquid is made in bacillus licheniformis and Sphingobacterium；
3) Gram positive actinomycetes-Bacillus cercus is made in Gram positive actinomycetes and Bacillus cercus and mixes hair
Zymotic fluid；
By bacillus licheniformis obtained in the secondary bacterium solution of Paecilomyces lilacinus, step 2-Sphingobacterium mixed fermentation liquid with
And Gram positive actinomycetes obtained in step 4)-Bacillus cercus mixed fermentation liquid mixing, fixed nitrogen is added while stirring
Bacterium, bacillus megaterium, photosynthetic bacteria are uniformly mixed, obtain mixed bacteria liquid A；
5) amino acid, humic acid, monosaccharide, chitin, modification infusorial earth are added in mixed bacteria liquid A, after mixing evenly, are carried out high
Temperature spray drying, temperature are 160-180 DEG C, and feed flow rate 2-4L/min finally obtains the microbial bacteria of controlling plant diseases
Agent.
2. the preparation method of the microbial bacterial agent of controlling plant diseases according to claim 1, which is characterized in that step 1)
Described in the secondary bacterium solution of Paecilomyces lilacinus the preparation method is as follows: Paecilomyces lilacinus strain is inoculated in the first culture medium
In, culture obtains fluorescence bacterium solution of unicellular bacillus, and bacterium solution of Paecilomyces lilacinus is then inoculated in the second culture medium
In, culture obtains the secondary bacterium solution of Paecilomyces lilacinus.
3. the preparation method of the microbial bacterial agent of controlling plant diseases according to claim 2, which is characterized in that described
One culture medium includes the following raw material according to parts by weight: 2-20 parts of peptone, 0.5-1.5 parts of dipotassium hydrogen phosphate, magnesium sulfate 0.5-
1.8 parts, 10-25 parts of agar, 1000 parts of sterile water, the pH value of first culture medium are 7.2 ± 0.2；Second culture medium
Including below according to the raw material of parts by weight: 5-20 parts of peptone, 0.5-5 parts of yeast extract, 5-10 parts of sodium chloride, 10-20 parts of agar,
1000 parts of distilled water；The pH value of second culture medium is 7.0.
4. the preparation method of the microbial bacterial agent of controlling plant diseases according to claim 1, which is characterized in that step 2
Described in bacillus licheniformis-Sphingobacterium mixed fermentation liquid the preparation method comprises the following steps: by bacillus licheniformis and sphingol bar
Bacterium seed liquor is seeded in PDB fluid nutrient medium, and aerobic fermentation culture 24-96h is carried out under the conditions of 24-28 DEG C of temperature, is obtained
To required bacillus licheniformis-Sphingobacterium mixed fermentation liquid.
5. the preparation method of the microbial bacterial agent of controlling plant diseases according to claim 4, which is characterized in that step 3)
Described in Gram positive actinomycetes-Bacillus cercus mixed fermentation liquid the preparation method comprises the following steps: by Gram positive actinomycetes,
Bacillus cercus is seeded in LB culture medium respectively using oese picking colony and is cultivated, and cultivation temperature is 32 DEG C,
Hunting speed is 120rpm in reciprocating oscillator, and after culture for 24 hours, fermentation medium of transferring carries out fermentation 24-36h, inoculum concentration
For the 1-3% for the total volume that ferments.
6. the preparation method of the microbial bacterial agent of controlling plant diseases according to claim 1, which is characterized in that step 1)
Described in monosaccharide be sucrose or glucose.
7. the preparation method of the microbial bacterial agent of controlling plant diseases according to claim 6, which is characterized in that step 1)
Described in modification infusorial earth the preparation method comprises the following steps: diatomite is added in the nitric acid solution of PH=0.01-0.5, be configured to weight
Percent concentration is the slurries of 20-50%, boils, cools down, filters, washs, dries, calcines, obtain modification infusorial earth, wherein doing
Dry temperature is 60-120 DEG C, and dry 10-30h, calcination temperature is 400-650 DEG C, calcination time 10-20h.
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| CN113248308A (en) * | 2021-05-20 | 2021-08-13 | 施可丰化工股份有限公司 | Microbial agent for effectively preventing and treating root knot nematode disease and preparation method thereof |
| CN114394863A (en) * | 2022-01-18 | 2022-04-26 | 山东省农业科学院 | Microbial water-soluble fertilizer for preventing and treating peanut diseases and insect pests and preparation method thereof |
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