CN108676745A - Solution xylose lysine bacillus LX024 and its application of one plant of prevention wheat base rot disease and head blight - Google Patents
Solution xylose lysine bacillus LX024 and its application of one plant of prevention wheat base rot disease and head blight Download PDFInfo
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- CN108676745A CN108676745A CN201810493252.0A CN201810493252A CN108676745A CN 108676745 A CN108676745 A CN 108676745A CN 201810493252 A CN201810493252 A CN 201810493252A CN 108676745 A CN108676745 A CN 108676745A
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- 239000012681 biocontrol agent Substances 0.000 claims abstract description 28
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Abstract
The present invention discloses a kind of solution xylose lysine bacillus LX024 and its application of prevention wheat base rot disease and head blight.The bacterial strain is solution xylose lysine bacillus(Lysinibacillus xylanilyticus), it is preserved in China General Microbiological culture presevation administrative center, preservation date September in 2017 19 days, deposit number is CGMCC No.14645.The biocontrol agent made of LX024 strain liquid fermented and cultureds has better effects to prevention wheat base rot disease and head blight.There are a degree of control effect, preventive effect to respectively reach 65.01% and 65.81% prevention wheat base rot disease and head blight using biocontrol agent LX024 in Wheat Seedling and growth period.
Description
Technical field
The invention belongs to field of agricultural microorganism, are related to one plant of biological and ecological methods to prevent plant disease, pests, and erosion solution for preventing wheat base rot disease and head blight
Xylose lysine bacillus LX024 and its application.
Background technology
Wheat scab and base rot disease are seriously affected by the microbial global disease for endangering wheat of F.graminearum schw
The yield and quality of wheat.Wherein wheat base rot disease be it is a kind of invaded by a variety of pathogens, the one kind for mixing generation compound is invaded
Metachromia soil-borne disease, also referred to as reaping hook root rot and reaping hook base rot disease etc..The disease is a kind of global important disease,
There is more than 10 country to report it.In China, the harm of Disease of Wheatbrown Foot Rot was once reported in Jiangsu, big according to Henan Agriculture
The investigation of wheat soil-borne diseases seminar of plant protection institute is learned, which generally sends out in China the Yellow River and Huai He River wheat main producing region each province
It is raw.Wheat scab is caused by Fusarium graminearum synthesis species, is happened at warm moist and one kind of semi-humid region extensively
Destructive disease.The Yangtze River in China middle and lower reaches area of wheat and the Major Epidemic area that eastern part of Northeast spring wheat area is the disease.Recently as complete
The morbidity range of ball climate change, wheat scab has spread to varieties in Huang-Huai-Hai Plain south.Illness wheat seriously affects wheat production
Amount and quality also jeopardize people, animal health.
Currently, mainly taking chemical prevention to the disease control of wheat base rot disease and head blight, breeding resistant variety, adding
The measures such as strong field management.But since breeding cycle is long and the limitation of field management, chemical prevention is most common prevention
Method.Though chemical prevention can effectively control the development of the field state of an illness, secondary dirt is easily caused to ecological environment and Wheat Production
Dye, therefore, environmentally friendly biological prevention gradually draws attention.Biological control strategy has biological control source resource
Extensively, the features such as environmentally friendly, every country and area scholar and researcher all try to explore and assess biology prevent
Control application prospect and potential value of the method on Fusarium graminearum means of prevention.Therefore biological control is red as novel wheat
Mildew and base rot disease control measure are worth with good production application.
Invention content
The purpose of the present invention is for the life not having to wheat scab and base rot disease in the prior art compared with high preventive effect
The present situation of anti-preparation, provide it is a kind of prevention wheat base rot disease and head blight solution xylose lysine bacillus LX024 and its
Using.
The purpose of the present invention can be achieved through the following technical solutions:
The solution xylose lysine bacillus LX024 of one plant of prevention wheat base rot disease and head blight, the identified Pseudomonas is in solution
Xylose lysine bacillus LX024(Lysinibacillus xylanilyticus), the biological material specimens preservation
And it survives, biomaterial Classification And Nomenclature is solution xylose lysine bacillus Lysinibacillus xylanilyticus,
The deposit date is Septembers in 2017 19 days, and depositary institution is China Committee for Culture Collection of Microorganisms's common micro-organisms center
(CGMCC), depositary institution address is Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, and culture presevation number is CGMCC
No.14645。
Above-mentioned solution xylose lysine bacillus LX024 is in prevention wheat base rot disease and head blight or prepares for preventing
Control the application in wheat base rot disease and the biocontrol agent of head blight.
It is a kind of to be used to prevent wheat base rot disease and the biocontrol agent of head blight, above-mentioned solution wood is included in the biocontrol agent
Sugared lysine bacillus LX024.As a kind of optimal technical scheme, viable bacteria total concentration in the biocontrol agent not less than 1 ×
108 CFU/mL。
The preparation method of the microbial inoculum is:By the seed bacterium solution of above-mentioned solution xylose lysine bacillus LX024 with 1:500 bodies
Product is than being seeded to fermented and cultured in LB fermentation mediums, 28 DEG C of 150 ~ 200 40 ~ 48 h of rpm shaken cultivations, then 5000 ~
6000 rpm centrifuge 10 ~ 15min, take and precipitate up to the biocontrol agent LX024, are diluted with aqua sterilisa and microbial inoculum is made,
Viable bacteria total concentration is 1 × 10 in finished product microbial inoculum8~1×109 CFU/mL。
Certainly, the solution xylose lysine bacillus LX024 systems of the prevention wheat scab and base rot disease are utilized
Standby biocontrol agent is not limited to above method.It is every can mass propgation LX024, and keep the active method of its biological and ecological methods to prevent plant disease, pests, and erosion available
In the claimed biocontrol agent of the preparation present invention.
The preparation for the solution xylose lysine bacillus LX024 seed bacterium solutions that the preserving number is CGMCC No. 14645
Method is:The solution xylose lysine bacillus LX024 inoculations that preserving number is CGMCC No. 14645 are cultivated to LB
OD values in liquid at 28 DEG C of 180 rpm shaken cultivations to 600 nm of culture terminate to cultivate when being 0.8.
A method of prevention wheat base rot disease and head blight carry out at inoculation wheat with the biocontrol agent
Reason.
Wheat is inoculated under greenhouse and crop field environment respectively after the biocontrol agent is diluted 100-200 times
Processing, when morbidity, start to count the state of an illness.Solve preventive effects of the xylose lysine bacillus LX024 to wheat base rot disease and head blight
Respectively 65.01 and 65.81%.
Beneficial effects of the present invention:
The present invention is specific to the biological prevention and control agent of wheat scab and base rot disease exploitation.Due to being biological agent, do not have completely
There is a series of problems caused by the use because of chemical pesticide, thus be conducive to the No-harmful apple orchard of wheat, peasant can not have to
Or the dosage of other chemical pesticides is reduced, this can not only reduce expenses for peasant, but also reduce the residual of chemical agent.
Greenhouse experiments and field experiment show:The solution xylose lysine bacillus LX024(CGMCC No. 14645)Energy
Significantly reduce the generation of wheat scab and base rot disease.The biocontrol agent prepared using the bacterium is diluted 100 ~ 200 times and is filled
Root processing, to the preventive effect of wheat base rot disease and head blight respectively up to 65.01 and 65.81%.
Biological sample preservation information
One plant of solution xylose lysine bacillus LX024, Classification And Nomenclature areLysinibacillus xylanilyticus, in
September is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center on 19th within 2017(CGMCC), preservation address
For Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Institute of Microorganism, Academia Sinica, culture presevation number is CGMCC No.
14645。
Specific implementation mode
Embodiment 1
Screening technique:Solve xylose lysine bacillus LX024(CGMCC No. 14645)It is small by Jiangsu Province Huai'an harbour town
It detaches and obtains in wheat rhizosphere soil.Wheat root is cut into the small pieces of 1cm, impregnates 5 min, the leaching of 70% alcohol with 1% sodium hypochlorite
Steep 1-2 min, sterile water wash 3 times(It takes the 100 μ l of solution that last time is cleaned to be applied on R2A solid mediums to cultivate, if
Asepsis growth then thinks that surface sterilization is clean after 48 h, sterile).It takes the sample that 3 g are disinfected to be placed in bacterium mortar, is added 27
0.85% sterile ml NaCl macerates tissue, grinding, with 0.85% sterile NaCl gradient dilutions.Take 10-1,10-2,
Each 100 μ l of dilution of 10-3 three gradients are applied on R2A, 28 DEG C of 48 h of culture.The maximum single bacterium colony of picking is inoculated into newly
Fresh R2A solid mediums obtain purifying bacterial strain through transferring repeatedly.By bacterial strain after purification with 40% glycerine be stored in -70 DEG C it is ultralow
In temperature refrigerator.
Identification method:By morphological feature, the bacterial strain is identified in bio-chemical characteristics and 16S rDNA sequence analyses.
Morphological feature:Gram-positive bacteria, it is rod-shaped, gemma is generated, bacterium colony is circle, and faint yellow, surface is smooth.
Gene magnification and sequence analysis:Xylose lysine bacillus LX024 is solved to cultivate extremely for 28 DEG C in LB culture mediums
Logarithmic phase centrifuges 5 min with 12 000 r/min and collects thalline, using precious bioengineering(Dalian)The genome of Co., Ltd
DNA rapid extraction kits, extract the genomic DNA of bacterium, using the DNA product of extraction as template, are expanded with bacterial 16 S rRNA
Increase universal primer U8-27 (5 '-AGAGTTTGATC (AC) TGGCTCAG-3 '), L1494-1514 (5 '-CTACGG (AG)
TACCTTGTTACGAC-3 ') 16S rDNA genetic fragments are amplified from genomic DNA.PCR product serves the raw work biology in sea
Engineering Co., Ltd is sequenced.By BLAST softwares tetraploid rice, knot are carried out to measuring 16S rRNA gene orders
Therefore the bacterial strain is accredited as solution xylose lysine bacillus by fruit abridged table(Lysinibacillus xylanilyticus).
Comparison result is sequenced in 1 LX024 of table
Bacterial strain | Most like bacterial strain and accession number | Similarity (%) |
LX024 | Lysinibacillus xylanilyticus KX703024.1 | 99 |
The preparation of 2 LX024 biocontrol agents of embodiment
It will solution xylose lysine bacillus LX024(CGMCC No. 14645)28 DEG C 180 in inoculation to LB culture solutions
It after 16 h of rpm shaken cultivations, is sampled in superclean bench every 2 h and surveys its OD value at 600 nm, when OD values are 0.8
When terminate to cultivate, this bacterium solution is as seed bacterium solution.Seed bacterium solution with 1:500 volume ratios, which are seeded in LB fermentation cultures, ferments
Culture, 28 DEG C, 180 rpm, 48 h of culture, then 6000 rpm centrifugations, 10 min, take and precipitate up to the biocontrol agent
LX024.It is diluted biocontrol agent LX024 with aqua sterilisa, viable bacteria total concentration is 1 × 10 in gained biocontrol agent8 ~ 1×
10 9 CFU/mL。
The preventive effect of 3 greenhouse of embodiment and plot experiment biocontrol microorganisms LX024 to wheat scab and base rot disease
Experiment 1:Greenhouse test detects preventive effects of the LX024 to wheat base rot disease(2016.09-2016.12)
Place:Huaiyingong College gives birth to academy of sciences greenhouse
Greenhouse pot culture is tested:Wheat seed is first used to aseptic water washing twice, repeats to use after being sterilized within two minutes with 75% alcohol washes
Aseptic water washing three times, the wheat seed after disinfection cultivate germination in culture dish.In sowing to basin alms bowl, two leaves are grown to wholeheartedly
Phase is inoculated with biocontrol bacteria.It is inoculated with pathogen after a week, observation statistics is carried out after 7d one month.Greenhouse(25 DEG C of temperature)It cultivates small
Wheat seeding is per three plants of basin, and in two one heart stages of leaf, biocontrol microorganisms LX024 bacterium solutions and control group root irrigation are each to handle in triplicate,
It is inoculated with pathogen after 7d, takes moisturizing 48h, greenhouse-grown after inoculation at 25 DEG C.Daily to wheat basal part of stem base rot disease into
Row observation survey.
Wheat base rot disease investigates foundation《Wheat scab Forecast Techniques specification》(GB/T 15796-2011) combines this reality
The actual conditions tested record and analyze test data.
0 grade, outer layer leaf sheath non-evident sympton;
1 grade, the first leaf sheath it is brown it is withered less than leaf sheath length 1/4;
2 grades, the first leaf sheath it is brown it is withered less than leaf sheath length 1/4-1/2;
3 grades, the first leaf sheath it is brown it is withered less than leaf sheath length 1/2-3/4;
4 grades, the first leaf sheath whole removing green is rotted;
5 grades, third leaf sheath has apparent brown withered or complete stool withered,
Disease severity (disease index)=∑(Sick series × disease grade plant the number)/(Highest disease series × total plant number)×
100%
Biocontrol effect=(Control group disease severity-processing group disease severity)/ control group disease severity × 100%
Note:Calculate different disposal disease index and preventive effect, and using DPS Data Analysis Software progress significance difference analysis and
Multiple range test.
Investigation result after biocontrol agent LX024 is handled 15 days shows that biocontrol agent LX024 is to wheat base rot disease
Biological control effect(Hereinafter referred to as preventive effect)Reach 65.01%(Table 2).
Biocontrol effects of the 2 biocontrol agent LX024 of table to wheat base rot disease
Processing | Disease severity % | Preventive effect % |
LX024 microbial inoculums | 21.57±2.33b | 65.01 |
Control | 61.63±3.05a | --- |
Experiment 2:Field plot trial detects preventive effects of the LX024 to wheat scab(2016.10-2017.05)
Place:Huaiyingong College agronomic trials field
Experimental field preceding stubble is corn, and the soil texture is sandy loam.On October 24th, 2014:Wheat cultivation, thickness of sowing 100
Grain m-1, line space 25cm applies seed manure between two every trades(Composite fertilizer)10g;On November 14th, 2014:Wheat thinning, thinning is extremely
70-80 plants of m-1;November 23:Tillering fertilizer is applied, urea 5g is in the ranks applied.Other management are according to Huai'an high-yield culturing mode
It carries out.
Experimental plot 15m × 10m=150m2, experimental plot surrounding plantation protection row.2 processing of this experiment, each handle 4 weights
It is multiple.
This experiment selects wheat breed for Huaihe River wheat 27;Gibberellic hypha Inoculating date is jan flowering wheat, carries out single flower inoculation
Inoculation, each kind is not fallen ill before being inoculated with, and 2 little Hua of middle part small ear are inoculated with per fringe, and each cell that handles is inoculated with 20 fringes, connects
It is inoculated with pathogen after kind biocontrol microorganisms LX024 and moisturizing 1d;After 21d, each cell that handles samples 10 plants.It compares and each
Processing weighs 4 times.
3 wheat scab disease severity of table is classified
Severity gradation | Head blight Disease symptoms describe |
0 | Fringe portion is without clearly visible disease symptom |
1 | Only it is inoculated with small ear or the morbidity of adjacent small ear |
2 | Morbidity small ear accounts for the 1/4 or less of spike length |
3 | Morbidity small ear accounts for the 1/4-1/2 of spike length |
4 | Morbidity small ear accounts for 1/2 of spike length or more |
Disease severity (disease index)=∑(Sick series × disease grade plant the number)/(Highest disease series × total plant number)×
100%
Biocontrol effect=(Control group disease severity-processing group disease severity)/ control group disease severity × 100%
Note:Calculate different disposal disease index and preventive effect, and using DPS Data Analysis Software progress significance difference analysis and
Multiple range test.
Investigation result after the processing 35 days of biocontrol agent LX024 the 1st time shows that biocontrol agent LX024 is to wheat
The biological control effect of head blight(Hereinafter referred to as preventive effect)Reach 65.81%(Table 4).
Biocontrol effects of the 4 biocontrol agent LX024 of table to wheat scab
Processing | Disease severity % | Preventive effect % |
LX024 microbial inoculums | 20.94±1.56b | 65.81 |
Control | 61.24 ±1.43a | --- |
Claims (7)
1. the solution xylose lysine bacillus LX024 of one plant of prevention wheat base rot disease and head blight, the identified bacterial strain category
In solution xylose lysine bacillus LX024(Lysinibacillus xylanilyticus), the preservation on the 19th of September in 2017
In China Committee for Culture Collection of Microorganisms's common micro-organisms center, culture presevation number is CGMCC No.14645.
2. solution xylose lysine bacillus LX024 described in claim 1 is in prevention wheat base rot disease and head blight
Using.
3. solution xylose lysine bacillus LX024 described in claim 1 is being prepared for preventing wheat base rot disease and red
Application in the biocontrol agent of mildew.
4. a kind of for preventing wheat base rot disease and the biocontrol agent of head blight, which is characterized in that include in the biocontrol agent
Solution xylose lysine bacillus LX024 described in claim 1.
5. according to claim 4 for preventing wheat base rot disease and the biocontrol agent of head blight, which is characterized in that should
Viable bacteria total concentration in biocontrol agent is not less than 1 × 108 CFU/mL。
6. application of the biocontrol agent in prevention wheat base rot disease and head blight described in claim 4 or 5.
7. a kind of method of prevention wheat base rot disease and head blight, which is characterized in that with the biological and ecological methods to prevent plant disease, pests, and erosion described in claim 4 or 5
Microbial inoculum carries out inoculation processing to wheat.
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CN111996125A (en) * | 2020-08-18 | 2020-11-27 | 河南省农业科学院植物保护研究所 | Application of chaetomium globosum in preventing and treating wheat stem basal rot |
CN113925064A (en) * | 2021-11-01 | 2022-01-14 | 海南大学 | Application of B-lysine-resistant bacillus in inhibition of growth of pitaya soft rot germs |
CN114134087A (en) * | 2021-12-22 | 2022-03-04 | 安徽农业大学 | Streptomyces algicidal and application thereof in prevention and treatment of wheat stem basal rot |
CN114908012A (en) * | 2022-05-19 | 2022-08-16 | 中国水产科学研究院黑龙江水产研究所 | Bacterial strain beneficial to symbiosis of rice and fish and application thereof |
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Cited By (7)
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CN111996125A (en) * | 2020-08-18 | 2020-11-27 | 河南省农业科学院植物保护研究所 | Application of chaetomium globosum in preventing and treating wheat stem basal rot |
CN111996125B (en) * | 2020-08-18 | 2022-03-04 | 河南省农业科学院植物保护研究所 | Application of chaetomium globosum in preventing and treating wheat stem basal rot |
CN113925064A (en) * | 2021-11-01 | 2022-01-14 | 海南大学 | Application of B-lysine-resistant bacillus in inhibition of growth of pitaya soft rot germs |
CN114134087A (en) * | 2021-12-22 | 2022-03-04 | 安徽农业大学 | Streptomyces algicidal and application thereof in prevention and treatment of wheat stem basal rot |
CN114134087B (en) * | 2021-12-22 | 2023-07-14 | 安徽农业大学 | Streptomyces alginolyticus and application thereof in prevention and control of wheat stem basal rot |
CN114908012A (en) * | 2022-05-19 | 2022-08-16 | 中国水产科学研究院黑龙江水产研究所 | Bacterial strain beneficial to symbiosis of rice and fish and application thereof |
CN114908012B (en) * | 2022-05-19 | 2022-10-28 | 中国水产科学研究院黑龙江水产研究所 | Bacterial strain beneficial to symbiosis of rice and fish and application thereof |
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