CN108102991B - One plant of Bei Laisi bacillus and its application in prevention and treatment watermelon blight - Google Patents
One plant of Bei Laisi bacillus and its application in prevention and treatment watermelon blight Download PDFInfo
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/07—Bacillus
-
- 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
-
- 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
Abstract
The invention discloses one plant of Bei Laisi bacillus and its applications in prevention and treatment watermelon blight.Bacterial strain of the invention is Bei Laisi bacillus (Bacillus velezensis) strain X T1-3, it is to separate to obtain from the Dongping hu lake mud of Shandong Province Tai'an, China Committee for Culture Collection of Microorganisms's common micro-organisms center was preserved on September 26th, 2017, deposit number is CGMCC No.14746.Test result shows: Bei Laisi Bacillus strain XT1-3 biological control agent prevents and treats watermelon blight disease significant effect, and control efficiency reaches 77.03%, and watermelon growing is healthy and strong, and no phytotoxicity generates.
Description
Technical field
The invention belongs to technical field of plant disease biological control.Specifically, the present invention relates to one plant of Bei Laisi gemma
Bacillus and its application in prevention and treatment watermelon blight, and the biological control agent using Bei Laisi bacillus preparation.
Background technique
Watermelon occupies an important position always in world's horticulture, and Chinese growth of watermelon area accounts for about world's gross area
55%, total output accounts for about 70% or more.In recent years, since growth of watermelon area constantly expands, cause crop rotation difficult, cause to the west of
Cucurbit wilt etc. is that the disease of representative also aggravates year by year.Watermelon blight is by watermelon specialized form Fusarium oxysporum (Fusarium
Oxysporum f.sp.niveum) caused by destructiveness soil-borne disease, be distributed in each watermelon producing region in the world.Watermelon blight is
A kind of vascular bundle diseases can occur in entire growth period, using the bloom phase of bearing fruit and fruit expanding period as onset peak.Due to disease
Opportunistic pathogen can be into breeding in people's plant body and incubation period is long, and the disease is difficult to prevent and treat for a long time, especially in watermelon continuous crop
In the case of, disease occurs even more serious.
Prevention and treatment for watermelon blight relies primarily on chemical pesticide in production at present and is prevented and treated.High-toxic pesticide makes
With making the pathogen produce serious drug resistance, general chemical prevention not can solve problem, give soil environment band instead
Carry out adverse effect.Therefore it is necessary to develop new and effective environmentally friendly biological agent.
Bacillus is a kind of aerobic or amphimicrobian general name for producing gemma Gram-positive bacillus, and physiological characteristic is rich
Rich multiplicity, is widespread in nature, is the important microbe population of soil and plant surface rhizosphere, to the nontoxic nothing of people and animals
Evil, it is free from environmental pollution, a variety of antibiotic and enzyme can be generated, there is broad spectrum antibiotic activity.Bacillus prevents and treats as biocontrol microorganisms to be planted
The mechanism of object disease mainly has: antibiosis, nutrient competition, competition for space, hyperparasitism and induction plant generate resistance
Etc. mechanism, main Biocontrol Mechanism be with generate antagonistic substance inhibit disinfect pathogen growth or kill pathogen.These objects
The matter overwhelming majority is peptide antibiotics, has efficient, low-residual and advantages of environment protection.
Summary of the invention
The object of the present invention is to provide a kind of for efficiently preventing and treating the new microorganism of watermelon blight --- Bei Laisi bud
Spore bacillus strain XT1-3.Bei Laisi Bacillus strain of the invention is with the speed of growth is fast, sporulation quantity is big, resistance is strong, In
Plant (watermelon) surface can rapid, high volume the features such as colonizing, therefore have a good application prospect.
Above-mentioned purpose of the invention is achieved through the following technical solutions: bacterial strain provided by the present invention is Bei Laisi bud
Spore bacillus (Bacillus velezensis) strain X T1-3 is to separate to obtain from the Dongping hu lake mud of Shandong Province Tai'an,
Be preserved on September 26th, 2017 China Committee for Culture Collection of Microorganisms's common micro-organisms center (abbreviation CGMCC,
Address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Institute of Microorganism, Academia Sinica), deposit number CGMCC
No.14746.It is with following biological characteristics: on beef extract culture medium bacterium colony be creamy white, mucus, corrugated protrusion,
Microscopy, somatic cells are in the shape of a rod, can move；It is positive through Gram's staining (with Escherichia coli for control)；After spore staining
Observe that gemma is oval or column.
The cultural method or propagation method of Bei Laisi bacillus (B.velezensis) strain X T1-3 of the present invention include:
(1) Nostoc commune Vanch, which saves, uses beef extract culture medium, formula are as follows: beef extract 3g, glucose 2.5g, sodium chloride
5g, agar 17g, distilled water 1000mL, pH are adjusted to 7.2~7.4.
(2) laboratory fluids culture uses beef extract fluid nutrient medium, formula are as follows: beef extract 3g, glucose 2.5g, chlorine
Change sodium 5g, distilled water 1000mL, pH are adjusted to 7.2~7.4.
(3) solid culture based formulas: Gu the mass ratio of material and inorganic salts, the solid material and inorganic salts is 98.5: 0.5；Institute
It states solid material and is calculated as corn flour in mass ratio: bean cake powder: wheat bran=65: 25: 10；The component and weight ratio of the inorganic salts are as follows: phosphorus
Acid dihydride potassium 15%, magnesium sulfate 5%, ammonium sulfate 15%, precipitated calcium carbonate 65%.
(4) bulk fermentation culture formula: with solid culture based formulas in (3).
The present invention also provides a kind of for preventing and treating the biological control agent of watermelon blight, the biological control agent packet
Include Bei Laisi bacillus (B.velezensis) the strain X T1-3.
The present invention also provides a kind of for preventing and treating the biological control method of watermelon blight, the biological control method packet
It includes to the watermelon with wilt disease and applies above-mentioned Bei Laisi bacillus (B.velezensis) strain X T1-3 or above-mentioned biology
Prevent and treat preparation.
Present invention provides above-mentioned Bei Laisi Bacillus strain XT1-3 or above-mentioned biological control agent in prevention and treatment west
Purposes in cucurbit wilt.
The preparation method of biological control agent of the present invention, comprising the following steps:
(1) seed liquor of the Bei Laisi Bacillus strain XT1-3 is prepared；
(2) seed liquor prepared by step (1) is inoculated into solid medium, constant temperature incubation at 27~29 DEG C；
(3) sterile water is added in the culture of step (2) culture to mix, filtrate is seeded to bulk fermentation culture by filtering
Base, 27~29 DEG C of room temperature, 85% or more relative humidity fermenting cellar in carry out fermented and cultured.
In one embodiment of the invention, the preparation method comprises the following steps:
(1) spore of the Bei Laisi Bacillus strain XT1-3 is transplanted to beef extract fluid nutrient medium, 27~29 DEG C
3~5d of shaking table shaken cultivation obtains seed liquor；
(2) ratio of the seed liquor in mass ratio 10% of step (1) preparation is inoculated into solid medium, 27~29 DEG C
3~5d of lower shaken cultivation；
(3) ratio of the culture sterile water in mass ratio 1: 15 of step (2) culture is mixed, filtrate is pressed in filtering
The ratio of volume ratio 1: 6 is seeded to bulk fermentation culture medium, 27~29 DEG C of room temperature, 85% or more relative humidity fermenting cellar in send out
8~9d of ferment culture.
Wherein, the beef extract fluid nutrient medium in step (1), formula are as follows: beef extract 3g, glucose 2.5g, sodium chloride
5g, distilled water 1000mL, pH are adjusted to 7.2~7.4.
Solid medium in step (2) includes solid material and inorganic salts, and the mass ratio of the solid material and inorganic salts is 98.5:
0.5；The solid material is calculated as corn flour in mass ratio: bean cake powder: wheat bran=65: 25: 10；The component and weight ratio of the inorganic salts
Are as follows: potassium dihydrogen phosphate 15%, magnesium sulfate 5%, ammonium sulfate 15%, precipitated calcium carbonate 65%.
The solid medium of the same step of bulk fermentation culture medium (2) in step (3).
Experiment show Bei Laisi Bacillus strain of the invention have the speed of growth is fast, sporulation quantity is big, resistance is strong,
Plant (especially watermelon) surface can rapid, high volume colonize the features such as, therefore have a good application prospect.By the shellfish Lay
The biological control agent of this bacillus preparation can efficiently prevent and treat watermelon blight, be a kind of biology of great application prospect
Prevent and treat preparation.Test result shows: Bei Laisi bacillus (B.velezensis) strain X T1-3 biological control agent prevention and treatment west
Cucurbit wilt disease significant effect, control efficiency reaches 77.03%, and watermelon growing is healthy and strong, and no phytotoxicity generates.
Specific embodiment
The invention will now be further described with reference to specific embodiments, the advantages and features of the present invention will be with description and
It is apparent.But these embodiments are merely exemplary, and it is not intended to limit the scope of the present invention in any way.Those skilled in the art
Member it should be understood that without departing from the spirit and scope of the invention can details to technical solution of the present invention and form into
Row modifications or substitutions, but these modifications and replacement are within the scope of the invention.
Embodiment 1
1, the isolation and purification of Bei Laisi bacillus (B.velezensis) XT1-3
Bei Laisi bacillus (B.velezensis) XT1-3 of the invention is from lake mud using dilution-plate method peace
The separation of plate method of scoring obtains, separation method are as follows:
(1) separation of bacillus
Lake mud sample is acquired on April 21st, 2013 from Shandong Province Tai'an Dongping hu.Weigh 1g soil sample (lake mud) in
In 100mL sterile water, it is placed in 150rpm in 30 DEG C of shaking tables and shakes 10min, be subsequently placed in 60 DEG C of water-baths and be incubated for 30min, take
100μL 10-2、10-3、10-4Dilution is coated on LB culture medium flat plate, and each gradient is coated with three in parallel, is cultivated at 28 DEG C
The microbial bacteria of different shape after 2d on picking beef extract culture medium falls within and crosses on beef extract culture medium flat plate, timing
Observe bacterium colony growing state.Then plate streak is used, Bacillus strain is purified, number saves respectively.
(2) screening of the efficient antagonistic Bacillus of watermelon blight
1. primary dcreening operation: using opposite culture method, PDA (Potato Dextrose Agar) plate is prepared, with punch in bud
Spore bacillus, watermelon blight pathogen edge take diameter be 5mm bacteria cake, be implanted in respectively plate it is opposite two sides center, 28
DEG C constant temperature incubation observes bacillus to the inhibiting effect of pathogen day by day.
2. secondary screening: the Bacillus strain with efficient antagonistic activity screened being carried out secondary screening, mainly by resistance to
Warm nature, resistance to acid and alkali, drug-resistant test screen the preferable Bacillus strain of patience, carry out potting controlling experiment and field
Test, and the strain X T1-3 obtained to screening is identified.
The present inventor obtains one plant of Bei Laisi gemma bar that can efficiently prevent and treat watermelon blight by a large amount of screening operations
Bacterium (B.velezensis) XT1-3.It is demonstrated experimentally that the Bei Laisi fermentation of bacillus liquid is shown in prevention and treatment watermelon blight
Very efficient control efficiency.Thus, Bei Laisi bacillus of the invention is the Bei Laisi gemma with wide application prospect
Bacillus new strains can be used for preparing the biological control agent of prevention and treatment watermelon blight.
2, bacterial strain is identified
(1) Microbiological Characteristics
On beef extract culture medium bacterium colony be creamy white, mucus, corrugated protrusion, microscopy, somatic cells are in the shape of a rod, energy
Movement；It is positive through Gram's staining (with Escherichia coli for control)；Observe that gemma is oval or column after spore staining.
(2) molecular biological characteristic
The 16s rDNA result of gene sequence determination of the bacterial strain is following (SEQ-1):
CCTCACCGACTTCGGGTGTTACAAACTCTCGTGGTGTGACGGGCGGTGTGTACAAGGCCCGGGAACGT
ATTCACCGCGGCATGCTGATCCGCGATTACTAGCGATTCCAGCTTCACGCAGTCGAGTTGCAGACTGCGATCCGAA
CTGAGAACAGATTTGTGGGATTGGCTTAACCTCGCGGTTTCGCTGCCCTTTGTTCTGTCCATTGTAGCACGTGTGT
AGCCCAGGTCATAAGGGGCATGATGATTTGACGTCATCCCCACCTTCCTCCGGTTTGTCACCGGCAGTCACCTTAG
AGTGCCCAACTGAATGCTGGCAACTAAGATCAAGGGTTGCGCTCGTTGCGGGACTTAACCCAACATCTCACGACAC
GAGCTGACGACAACCATGCACCACCTGTCACTCTGCCCCCGAAGGGGACGTCCTATCTCTAGGATTGTCAGAGGAT
GTCAAGACCTGGTAAGGTTCTTCGCGTTGCTTCGAATTAAACCACATGCTCCACCGCTTGTGCGGGCCCCCGTCAA
TTCCTTTGAGTTTCAGTCTTGCGACCGTACTCCCCAGGCGGAGTGCTTAATGCGTTAGCTGCAGCACTAAGGGGCG
GAAACCCCCTAACACTTAGCACTCATCGTTTACGGCGTGGACTACCAGGGTATCTAATCCTGTTCGCTCCCCACGC
TTTCGCTCCTCAGCGTCAGTTACAGACCAGAGAGTCGCCTTCGCCACTGGTGTTCCTCCACATCTCTACGCATTTC
ACCGCTACACGTGGAATTCCACTCTCCTCTTCTGCACTCAAGTTCCCCAGTTTCCAATGACCCTCCCCGGTTGAGC
CGGGGGCTTTCACATCAGACTTAAGAAACCGCCTGCGAGCCCTTTACGCCCAATAATTCCGGACAACGCTTGCCAC
CTACGTATTACCGCGGCTGCTGGCACGTAGTTAGCCGTGGCTTTCTGGTTAGGTACCGTCAAGGTGCCGCCCTATT
TGAACGGCACTTGTTCTTCCCTAACAACAGAGCTTTACGATCCGAAAACCTTCATCACTCACGCGGCGTTGCTCCG
TCAGACTTTCGTCCATTGCGGAAGATTCCCTACTGCTGCCTCCCGTAGGAGTCTGGGCCGTGTCTCAGTCCCAGTG
TGGCCGATCACCCTCTCAGGTCGGCTACGCATCGTCGCCTTGGTGAGCCGTTACCTCACCAACTAGCTAATGCGCC
GCGGGTCCATCTGTAAGTGGTAGCCGAAGCCACCTTTTATGTCTGAACCATGCGGTTCAGACAACCATCCGGTATT
AGCCCCGGTTTCCCGGAGTTATCCCAGTCTTACAGGCAGGTTACCCACGTGTTACTCACCCGTCCGCCGCTAACAT
CAGGGAGCAAGCTCCC。
(3) cellular morphology and physical and chemical experiment result
The cellular morphology and physical and chemical experiment result of table 1 Bei Laisi bacillus (B.velezensis) XT1-3
| Experimental project | As a result | Experimental project | As a result |
| Gram's staining | It is positive | Carbohydrate produces acid | + |
| Cell shape | It is rod-shaped | Glucose | + |
| 1 μm of cell dia ﹥ | - | Xylose | + |
| Form gemma | + | L-arabinose | + |
| Gemma expands | - | Mannitol | - |
| Gemma is round | - | Lactose | - |
| Parasporal crystal | - | Glucose fermentation produces gas | + |
| Catalase | + | Utilize citrate | + |
| Oxidizing ferment | + | 50 DEG C of growths | + |
| Anaerobic growth | - | PH 5.7 is grown | + |
| VP test | + | 7%NaCl growth | + |
| VP ﹤ pH 6 | + | Starch Hydrolysis | + |
| VP ﹥ pH 7 | - | Decompose casein | + |
| Nitrate reduction | + | Gelatin hydrolysate | + |
Strain X T1-3 is accredited as Bei Laisi bacillus, is preserved in Chinese microorganism strain on September 26th, 2017
Preservation administration committee common micro-organisms center, deposit number are CGMCC No.14746.
Embodiment 2
1, fermentation of bacillus process
Beef extract Liquid Culture based formulas are as follows: beef extract 3g, glucose 2.5g, sodium chloride 5g, distilled water 1000mL, pH
It is adjusted to 7.2~7.4.
Solid culture based formulas: Gu the mass ratio of material, inorganic salts and water, the solid material and inorganic salts is 98.5: 0.5；Institute
It states solid material and is calculated as corn flour in mass ratio: bean cake powder: wheat bran=65: 25: 10；The component and weight ratio of the inorganic salts are as follows: phosphorus
Acid dihydride potassium 15%, magnesium sulfate 5%, ammonium sulfate 15%, precipitated calcium carbonate 65%.Preparation method: suitable water (10-15 is added
Times quality) dissolution inorganic salts, obtain inorganic salt solution；Inorganic salt solution is mixed with solid material, and the training of water final adjustment solid is added
The water content for supporting base is 40-60%, and solid medium is held in the hand, has water in webs but not drippage is advisable.
The same solid medium of bulk fermentation culture medium.
The a large amount of solid fermentation process of Bei Laisi bacillus (B.velezensis) strain X T1-3:
1. strain seed liquor culture
By a small amount of spore of Bei Laisi Bacillus strain (B.velezensis) XT1-3 picking from test tube slant, move to
In beef extract fluid nutrient medium, 28 DEG C of shaking table 3~5d of shaken cultivation, this is seed liquor.
2. the culture of solids manufacture strain
Seed liquor is inoculated into solid medium (500mL triangular flask) in 10% ratio, 28 DEG C of constant temperature incubations 3~
5d, intermediate multiple oscillation.
3. a large amount of solid fermentations
2. the culture sterile water of middle solid culture is pressed into 1: 15 dilution proportion, and is filtered with sterile gauze, is removed thick
Slag, as production bacterium solution, inoculative proportion are inoculated in bulk fermentation culture medium by 1: 6.The raw material of inoculation is placed in (28 DEG C of fermenting cellar
With 85% or more relative humidity) in 8~9d of fermented and cultured, solid fermentation culture active bacteria can reach 1,500,000,000/gram.
Prevention and treatment for clear Bei Laisi bacillus (B.velezensis) strain X T1-3 fermentation liquid to watermelon blight
Effect provides foundation for its popularization and application.We have carried out crop field examination in Heze city of Shandong province Dongming County in June, 2016
It tests, is now reported as follows test result:
1 material to be tested and method
1.1 reagent agent
Bei Laisi bacillus (B.velezensis) strain X T1-3 by the production of 2 fermentation process of the embodiment of the present invention is solid
Body fermentation culture medium (1,500,000,000/gram)；25% carbendazol wettable powder (production of Shandong rural area Biotechnology Co., Ltd, city
It sells).
1.2 materials to be tested and controlling object
Material to be tested is watermelon；Controlling object is watermelon blight.
1.3 experimental field situations
Experimental field it is located at Heze city of Shandong province, Datun Town, Dongming County watermelon producing region.In this area, due to continuous cropping continuous cropping, watermelon
Wilt disease causes very huge economic loss than more serious, to peasant.Test soil types is moisture soil, and the content of organic matter is
0.63%, pH value 7.4, all experimental plot cultivation conditions and management measure are consistent, and watermelon blight is in morbidity when application
Initial stage.
1.4 experimental designs and arrangement
This test totally 3 processing, respectively 20 times of fermentation liquid of Bei Laisi bacillus (B.velezensis) strain X T1-3
Liquid, 25% carbendazol wettable powder, 500 times of liquid are not administered clear water and compare, 4 repetitions of each processing, each processing 140
Strain.To diseased plant and its neighbouring plant pouring root and full field foliar spray is carried out in early stage, 10 days are primary, and continuous 2 times.It is every every time
Diseased plant irrigation liquid 0.25kg；Full field is spraying, every 666.7m2It is 30~50L with liquid volume.On June 1st, 2016 and June 10
Each pouring root, sprinkling are primary, and spray instrument is 16 type sprayer of workers and peasants-.
1.5 pilot surveies and calculation method
1.5.1 meteorological condition
1st application (June 1) same day is cloudy, and 3 grades of wind-force, the highest temperature is 30 DEG C, and the lowest temperature is 19 DEG C, relatively
Humidity is 67%.2nd application (June 10), the same day is cloudy, and 3-4 grades of wind-force, the highest temperature is 33 DEG C, the lowest temperature 22
DEG C, relative humidity 70%.
1.5.2 drug effect and security survey
Efficacy survey: 14d is investigated after last time is administered.Each processing investigates 120 plants, investigates plant vines base
Portion, Investigate incidence record disease index and calculate disease index.
Security survey: the safety of 7d and 14d observation watermelon after being administered in first time occurs if any phytotoxicity, retouches in detail
It states symptom of chemical damage and determines phytotoxicity degree by phytotoxicity grading standard.
Stage division (as unit of single plant):
0 grade: watermelon plant health, disease-free spot.
1 grade: occurring slight scab in watermelon plants stems, but grow normal.
3 grades: scab is obvious in watermelon plants stems, and growth is slight suppressed.
5 grades: scab is obvious in watermelon plants stems, and plant top is wilted, slow growth or rigid.
7 grades: 50% or more browning of watermelon plant is withered.
9 grades: 50% or more browning of watermelon plant is withered or withered.
1.5.3 drug effect calculation method
Drug effect is calculated by formula (1), (2):
In formula: CK0--- disease index before blank district is administered；
CK1--- disease index after blank district application；
PT0————Disease index before chemicals treatment is administered；
PT1————Disease index after chemicals treatment application.
1.5.4 to the influence of watermelon growing
Observing medicament, whether there is or not phytotoxicity to watermelon, record the type and extent of phytotoxicity.In addition, also to record to watermelon other
Beneficial effect (such as stimulating growth etc.).
Phytotoxicity is recorded with following manner:
If (a) phytotoxicity can be measured or calculate, absolute value representation, such as plant height are used.
(b) in other cases, phytotoxicity degree and frequency can be estimated by following two methods:
1. according to the phytotoxicity degree of phytotoxicity stage division record watermelon, with-,+, ++, +++, ++++it indicates.
Phytotoxicity stage division:
-: without phytotoxicity；
+: slight phytotoxicity does not influence plant growth；
++: moderate phytotoxicity, it is resilient, it not will cause crop failure；
+++: moderate phytotoxicity influences crop normal growth, causes a degree of loss to crop yield and quality；
The serious phytotoxicity of ++++:, plant growth are obstructed, and yield and quality loss is serious.
2. evaluating its phytotoxicity percentage by chemicals treatment district compared with blank district.Meanwhile wanting accurate description crop
Symptom of chemical damage (dwarfing, chlorisis, deformity etc.).
2 results
Control efficiency of 2.1 reagent agents to watermelon blight
Reagent agent is to the control efficiency of watermelon blight as shown in table 2-3.Table 3 the result shows that, Bei Laisi bacillus
(B.velezensis) strain X T1-3 solid fermentation culture and 25% carbendazol wettable powder have to watermelon blight compared with
Good control efficiency, wherein the former control efficiency reaches 75% or more.
From the point of view of the disease index of watermelon blight field trial (table 3), 14d after last time is administered, Bei Laisi gemma
Bacillus (B.velezensis) XT1-3 control efficiency is 77.03%, hence it is evident that is higher than 25% carbendazol wettable powder 53.63%
Control efficiency.
Watermelon blight disease incidence before 2 three kinds of chemicals treatments of table
Control efficiency of the 3 three kinds of chemicals treatments of table to watermelon blight
2.2 watermelon security surveys
Through application 7d and 14d observation, for each chemicals treatment district compared with check plot, watermelon growing is normal, and no phytotoxicity generates, and says
Ming Beilaisi bacillus solid fermentation culture is for examination concentration to watermelon safety.
3 brief summaries
3.1 from the point of view of disease index and control efficiency, and Bei Laisi bacillus (B.velezensis) strain X T1-3 (is protected
Hiding CGMCC No.14746) there is preferable control efficiency to watermelon blight, the control efficiency of 14d after last time application
Up to 75% or more.
3.2 in terms of watermelon growing way, and Bei Laisi bacillus (B.velezensis) strain X T1-3 has watermelon apparent
Promote growth result, without phytotoxicity, securely and reliably.
SEQUENCE LISTING
<110>Shandong Dai Meng obtains Biotechnology Co., Ltd
<120>one plants of Bei Laisi bacillus and its application in prevention and treatment watermelon blight
<130> 0
<160> 1
<170> PatentIn version 3.3
<210> 1
<211> 1376
<212> DNA
<213>the 16s rDNA gene order of Bei Laisi bacillus (Bacillus velezensis) strain X T1-3
<400> 1
cctcaccgac ttcgggtgtt acaaactctc gtggtgtgac gggcggtgtg tacaaggccc 60
gggaacgtat tcaccgcggc atgctgatcc gcgattacta gcgattccag cttcacgcag 120
tcgagttgca gactgcgatc cgaactgaga acagatttgt gggattggct taacctcgcg 180
gtttcgctgc cctttgttct gtccattgta gcacgtgtgt agcccaggtc ataaggggca 240
tgatgatttg acgtcatccc caccttcctc cggtttgtca ccggcagtca ccttagagtg 300
cccaactgaa tgctggcaac taagatcaag ggttgcgctc gttgcgggac ttaacccaac 360
atctcacgac acgagctgac gacaaccatg caccacctgt cactctgccc ccgaagggga 420
cgtcctatct ctaggattgt cagaggatgt caagacctgg taaggttctt cgcgttgctt 480
cgaattaaac cacatgctcc accgcttgtg cgggcccccg tcaattcctt tgagtttcag 540
tcttgcgacc gtactcccca ggcggagtgc ttaatgcgtt agctgcagca ctaaggggcg 600
gaaaccccct aacacttagc actcatcgtt tacggcgtgg actaccaggg tatctaatcc 660
tgttcgctcc ccacgctttc gctcctcagc gtcagttaca gaccagagag tcgccttcgc 720
cactggtgtt cctccacatc tctacgcatt tcaccgctac acgtggaatt ccactctcct 780
cttctgcact caagttcccc agtttccaat gaccctcccc ggttgagccg ggggctttca 840
catcagactt aagaaaccgc ctgcgagccc tttacgccca ataattccgg acaacgcttg 900
ccacctacgt attaccgcgg ctgctggcac gtagttagcc gtggctttct ggttaggtac 960
cgtcaaggtg ccgccctatt tgaacggcac ttgttcttcc ctaacaacag agctttacga 1020
tccgaaaacc ttcatcactc acgcggcgtt gctccgtcag actttcgtcc attgcggaag 1080
attccctact gctgcctccc gtaggagtct gggccgtgtc tcagtcccag tgtggccgat 1140
caccctctca ggtcggctac gcatcgtcgc cttggtgagc cgttacctca ccaactagct 1200
aatgcgccgc gggtccatct gtaagtggta gccgaagcca ccttttatgt ctgaaccatg 1260
cggttcagac aaccatccgg tattagcccc ggtttcccgg agttatccca gtcttacagg 1320
caggttaccc acgtgttact cacccgtccg ccgctaacat cagggagcaa gctccc 1376
Claims (8)
- Bei Laisi bacillus 1. (Bacillus velezensis) strain X T1-3, the deposit number of the bacterial strain is CGMCC No.14746。
- 2. purposes of the Bei Laisi Bacillus strain XT1-3 described in claim 1 in prevention and treatment watermelon blight.
- 3. a kind of biological control agent, including Bei Laisi Bacillus strain XT1-3 described in claim 1.
- 4. the preparation method of biological control agent as claimed in claim 3, characterized in that the following steps are included:(1) seed liquor of Bei Laisi Bacillus strain XT1-3 is prepared；(2) seed liquor prepared by step (1) is inoculated into solid medium, constant temperature incubation at 27~29 DEG C；(3) sterile water is added in the culture of step (2) culture to mix, filtrate is seeded to fermentation medium, in room temperature by filtering 27~29 DEG C, carry out fermented and cultured in the fermenting cellar of 85% or more relative humidity.
- 5. the preparation method of biological control agent as claimed in claim 4, characterized in thatSolid medium in the step (2) includes solid material and inorganic salts, and the mass ratio of the solid material and inorganic salts is 98.5: 0.5；The solid material is calculated as corn flour in mass ratio: bean cake powder: wheat bran=65: 25: 10；The component and weight ratio of the inorganic salts Are as follows: potassium dihydrogen phosphate 15%, magnesium sulfate 5%, ammonium sulfate 15%, precipitated calcium carbonate 65%；The solid medium of the same step of fermentation medium (2) in step (3).
- 6. the preparation method of biological control agent as claimed in claim 4, characterized in that the following steps are included:(1) spore of Bei Laisi Bacillus strain XT1-3 is transplanted to beef extract fluid nutrient medium, 27~29 DEG C of shaking table oscillations 3~5d of culture obtains seed liquor；(2) ratio of the seed liquor in mass ratio 10% of step (1) preparation is inoculated into solid medium, is shaken at 27~29 DEG C Swing 3~5d of culture；(3) ratio of the culture sterile water in mass ratio 1: 15 of step (2) culture is mixed, filtering, filtrate is pressed into volume Ratio than 1: 6 is seeded to fermentation medium, 27~29 DEG C of room temperature, 85% or more relative humidity fermenting cellar in fermented and cultured 8 ~9d.
- 7. purposes of the biological control agent as claimed in claim 3 in prevention and treatment watermelon blight.
- 8. a kind of biological control method of watermelon blight, characterized in that apply claim 1 institute to the watermelon with wilt disease The Bei Laisi Bacillus strain XT1-3 or biological control agent as claimed in claim 3 stated.
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| CN111304135B (en) * | 2020-04-10 | 2022-06-07 | 宁波市农业科学研究院 | Bacillus and application thereof in plant disease control |
| CN113755358B (en) * | 2020-11-30 | 2023-07-04 | 山西农业大学 | Bacillus bailii ZJ1 strain and fermentation broth preparation and use methods |
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| CN113755393B (en) * | 2021-10-09 | 2022-12-23 | 河南省科学院生物研究所有限责任公司 | Bacillus beilesensis HP-24 and application thereof in preparation of bacterial liquid for preventing and treating bacterial fruit blotch of melons |
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