CN107937278A - One plant prevention rice sheath blight disease than Lay mould - Google Patents
One plant prevention rice sheath blight disease than Lay mould Download PDFInfo
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- CN107937278A CN107937278A CN201711113602.8A CN201711113602A CN107937278A CN 107937278 A CN107937278 A CN 107937278A CN 201711113602 A CN201711113602 A CN 201711113602A CN 107937278 A CN107937278 A CN 107937278A
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- sheath blight
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Abstract
The invention discloses one plant of suppression rice sheath blight disease than Lay mould, which provides a kind of nontoxic to people and animals, to eco-friendly, there is microorganism fungus kind and its Pesticidal products of good preventive effect to rice sheath blight disease.Invention increases the species of prevention rice sheath blight disease microbial bactericide.
Description
Technical field
The present invention relates to the microbial control technical field of rice sheath blight disease, more particularly to one plant of prevention rice sheath blight disease
Than Lay mould.
Background technology
Rice sheath blight disease (Rhizoctonia solani) is domestic and international most serious and is also the nosomycosis for being most difficult to administer
One of evil, its is widely distributed, it is serious to cause harm, and all causes huge economic loss to agricultural production every year.Due to banded sclerotial blight mycocriny
Raw ability is strong, host range is wide, not yet finds the highly resistance kind sick to this so far both at home and abroad.Rice is prevented in China's production at present
For banded sclerotial blight based on jinggangmeisu and some chemical pesticides, antibiotic and being repeatedly used for a long for chemical agent have made pathogen
The obvious resistance to the action of a drug is generated, causes preventive effect to decline or even lose, dosage is continuously increased, and is largely caused for a long time using pesticide
The problem of natural ecological environment pollution and rice persticide residue increase, is also more and more prominent.Since rice sheath blight disease is a kind of soil
Pass disease, pathogen is not strong to the specificity in parasitism of host, between crop varieties specific resistance difference it is very little or none (Liu Wei etc.,
2009).Biological control not only economical and effective, and without side-effects is carried out using beneficial microbe.Therefore, using there is antagonism
Microorganism carry out biological control be one have explore be worth new way.In recent years, the main rice region in the world, which has generally been carried out, adopts
The biological control research that rice sheath blight disease endangers is controlled with antagonistic microbe.
Bigger progress has been obtained at present to the biological control research of rice sheath blight disease, has reported and has screened to rice line
Blight bacterium has the yellow false list of the bacillus subtilis Bs-916, B5423-R, bacillus Drt211, NBl2, cause of strong effect
The Z88 of born of the same parents bacterium B34 and Penicillium, trichoderma, Fusariumsp, production pesticide corrosion 120 actinomyces, burkholderia SD7 and ocean are thin
The bacterial strains such as bacterium have certain antagonism to Rhizoctonia solani Kuhn.Using at most surely belonging to bacillus subtilis, Jiangsu academy of agricultural sciences
(Chen Zhiyi etc., 2003) successfully screens bacillus subtilis Bs-916, has good preventive effect, its commodity to a variety of rice diseases
" since line Qu Ning " is promoted since 1991, to the preventive effect stabilization of banded sclerotial blight more than 70%.The biology that Agricultural University Of Nanjing develops
Pesticide " wheat Fengning ", is made of bacillus subtilis B3, and Rhizoctonia solani has significant inhibitory action.Yunnan Prov Agriculture University
The principle active component of " hundred is anti-" developed jointly with China Agricultural University is bacillus subtilis B908, has obtained the Ministry of Agriculture
Register, to the prevention effect of rice sheath blight disease up to more than 70%.
In recent years, the main rice region in the world has generally been carried out using antagonistic microbe to control the biology that rice sheath blight disease endangers
Study on prevention, mainly bacillus subtilis biocontrol agent, the biocontrol bacterial strain of most of rice sheath blight disease for developing into product are equal
Still lack the novel microbial pesticide for preventing rice sheath blight disease in the laboratory research stage, in production.
Than Lay mould, this kind of colony growth limitation, falx stem is short, top is usually expanded, and conidium presents spherical to ellipse
The feature such as circular and small is more apparent.
The present invention is in order to screen the efficient biocontrol microorganisms of Rhizoctonia solani Kuhn, using 3 opposite culture methods, mycelial growth speed
Rate method, sclerotial germination suppress method and in vitro efficiency test, to the rice terrace of the area such as Liaoning Province, Jilin Province and Heilongjiang Province collection
Soil sample is separated and screened, and carries out Identification of Species to biocontrol bacterial strain.It is intended to obtain the higher biocontrol bacterial strain of prevention effect, is
Rice sheath blight disease biological control provides foundation.
The content of the invention
To solve the above-mentioned problems, the present invention have found one plant of new biocontrol bacterial strain in prevention rice sheath blight disease.
Technical scheme is as follows：
One plant of fungi inhibited to Rhizoctonia solani Kuhn is than Lay mould JLF6-2, in September 26 in 2017
Day is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, Classification And Nomenclature：Than Lay mould
Penicillium bilaiae, preserving number are：CGMCC N0.14626.Preservation address：BeiChen West Road, Chaoyang District, BeiJing City 1
No. 3 Institute of Microorganism, Academia Sinica of institute.
It is disclosed by the invention than Lay mould, possessed morphological feature：25 DEG C of cultures 7d, colony diameter 23mm on CYA,
Central elevation, edge are narrow and complete；Quality is cotton-shaped；The green white of conidium, ellipse, 2.5 × 2.3 μm, wall is slightly coarse；Mycelia
Body white；Reverse side yellow-white；Soluble pigment lacks；7d, colony diameter 26mm, central elevation, side are cultivated on MEA for 25 DEG C
Edge is low and complete；Quality is cotton-shaped；White mycelium；Diffusate lacks；Reverse side yellow；Soluble pigment lacks；Conidiophore
Without expanding, wall is smooth on top；Penicillus single-wheel is given birth to；Conidium light green, it is subsphaeroidal, 2.6 × 2.2 μm, smooth surface；
G25N upper 25 DEG C of cultures 7d, colony diameter 18mm, growth are limited；White mycelium；Conidium light green color, it is spherical, 2.5 ×
2.2 μm, smooth surface.
Experiment proves that fungi of the invention has very strong inhibitory action than Lay mould to Rhizoctonia solani Kuhn.
The present invention suppression Rhizoctonia solani Kuhn than Lay mould, have the following advantages that：
1. provide it is nontoxic to people and animals, to eco-friendly, have to rice sheath blight disease good preventive effect geomicrobes strain and
Its Pesticidal products.
2. provide one plant of Rhizoctonia solani Kuhn than Lay mould.
3. the species of increase prevention rice sheath blight disease microbial bactericide.
Brief description of the drawings
Fig. 1：Antagonistic effect (3 points of face-off methods) than Lay mould to Rhizoctonia solani Kuhn；
Fig. 2：Antagonistic effect (mycelial growth rate method) than Lay mould to Rhizoctonia solani Kuhn；
Fig. 3：Antagonistic effect (sclerotial germination suppression method) than Lay mould to Rhizoctonia solani Kuhn；
Fig. 4：In vitro prevention effect than Lay mould zymotic fluid to rice sheath blight disease.
Embodiment
The present invention is explained in detail with specific embodiment below.
Screening, separation and the identification of bacterial strain
The separation of bacterial strain：Isolated and purified using soil dilution partition method.
The screening of bacterial strain：Method is suppressed using 3 tablet opposite culture methods, mycelial growth rate method and sclerotial germination, it is comprehensive
Screening has Rhizoctonia solani Kuhn the biocontrol microorganisms of Biocontrol Effect.
1 tablet opposite culture method
Using 3 tablet opposite culture methods, the Rhizoctonia solani Kuhn bacterium colony for having activated, having cultivated 48h is chosen, with beating for 5mm
Hole device is made bacteria cake and puts PDA plate center, and symmetrical 3 dislocations triangular in shape are for examination at away from rice sheath blight disease bacteria cake 30mm
Biocontrol microorganisms.27 DEG C of constant temperature incubation 72h.Observe the formation of inhibition zone and measure its size.
2 mycelial growth rate methods
Using mycelial growth rate method, the strains tested bacteria cake of purifying is moved in fluid nutrient medium, 28 DEG C of 130r
min-1Shaken cultivation 24h, as seed culture fluid.Seed culture fluid 5mL is taken to be transferred to equipped with 100mL Liquid Cultures with liquid-transfering gun
In the triangular flask (250mL) of base, 28 DEG C of 130rmin-1Shaken cultivation 120h, 8000rmin-110min is centrifuged, takes supernatant
Liquid is filtered by filter (φ=0.22 μm), obtains sterile ferment filtrate.Take the sterile ferment filtrates of 1mL and 9mL PDA culture mediums
Mix, pour into sterilizing culture dish and band bacterium solution tablet is made.The dislocation Rhizoctonia solani Kuhn bacteria cake 1 at band bacterium solution PDA plate center
It is a, 27 DEG C of constant temperature incubations.Often processing is repeated 3 times.Antibacterial circle diameter and bacteriostasis rate are measured after 48h.
Colony growth diameter (mm)=measurement diameter (mm)-bacteria cake diameter (mm)
3 sclerotial germinations suppress method
10d will be cultivated on PDA plate for examination Rhizoctonia solani Kuhn, and after Sclerotia forming, select size basically identical
Sclerotium, which is put into sterile ferment filtrate, soaks 30min, and the sclerotium after processing moves into PDA plate, per 4, ware, is soaked with sterile water
Sclerotium for control.Each processing is repeated 3 times.27 DEG C of constant temperature incubations.Micro- Microscopic observation sclerotial germination situation after 48h, statistics
Sclerotial germination rate and mycelium germination series, calculate and sprout mycelia index and inhibiting rate.
Sprout mycelia grade scale：0 grade：It is zero root to sprout mycelia number；1 grade：It is 1~20 to sprout mycelia number；2 grades：Sprout
Bacterium germination silk number is 21~50；3 grades：It is 51~80 to sprout mycelia number；4 grades：It is 81~140 to sprout mycelia number；5 grades：Sprout
Bacterium germination silk number is more than 141.
The identification of bacterial strain：
1 biocontrol fungi is identified
Carried out using morphological feature and genetic marking method, to ITS1-5.8S-ITS2rRNA genes and calmodulin base
Because fragment amplification product is sequenced.It is special to record ripe colonial morphology feature, mycelia and conidium size, form and color etc.
Sign, the morphological feature of conidiophore, reference《Chinese fungi will》Carry out taxonomic identification.Had using Tiangeng biochemical technology (Beijing)
The fungal genomic DNA extracts kit extraction bacterial strain DNA of limit company, primer sequence are as shown in table 1.
1 PCR amplification of table and the primer sequence of sequencing
Table1 The primer sequences used for amplification
PCR reaction systems：2 × Taq PCR Master Mix12.5 μ L, F primers, 1 μ L, R primer 1 μ L, 1 μ L of template DNA,
Add ddH2O to 25 μ L of cumulative volume.PCR response procedures：94 DEG C of denaturation 3min；94 DEG C of denaturation 30s, 50 DEG C of 30s and 72 DEG C of annealing are prolonged
Stretch 45s totally 34 circulations；Last 72 DEG C of extensions 5min；15 DEG C of preservation 30min.PCR product is limited by Shanghai bioengineering share
Company carries out sequencing, and the sequence measured carries out sequence analysis after NCBI carries out BLAST comparisons with MEGA7 softwares, builds
Phylogenetic tree.
Opposite culture measures the inhibitory action than Lay mould to rice banded sclerotial blight pathogen
Biocontrol effect of 2 biocontrol bacterial strain of table to Rhizoctonia solani Kuhn
Table2 Bio-control strains antagonistic effect ofR.solani
Note：Data are the mean+SD of 3 repetitions；Significance analysis is using minimum range method (LSD, P ＜
0.05)。
Note:The experimental data in the table were Mean±SD(repeated
3times). Different lowercase letters indicated significant differences using
LSD test at 0.05 level.
In vitro prevention effect of 3 bacterial strain fermentation liquor of table to rice sheath blight disease
Table3 Control effect offermentation broth onR.solani
Note：Significance analysis is using minimum range method (LSD, P ＜ 0.05).
Note:Different lowercase letters indicated significant differences
using LSD test at 0.05level.
The ratio Lay mould JLF6-2 of the present invention has Rhizoctonia solani Kuhn preferable prevention effect, passes through 3 opposite cultures
Method, mycelial growth rate method, sclerotial germination suppress method and in vitro efficiency test, the results showed that than Lay mould to Rhizoctonia solani Kuhn
Inhibition zone be respectively 38.30mm；Mycelial growth inhibition rate is respectively 77.30%；Sclerotial germination inhibiting rate is respectively
86.67%.Protection processing in, than Lay mould 12h preventive effect be 67.24%；In treatment processing, than Lay mould in 8h
Preventive effect be 64.41%.
The foregoing is only a preferred embodiment of the present invention, but protection scope of the present invention be not limited thereto,
Any one skilled in the art the invention discloses technical scope in, technique according to the invention scheme and its
Inventive concept is subject to equivalent substitution or change, should be covered by the protection scope of the present invention.
Claims (4)
1. the ratio Lay mould JLF6-2 of one plant of suppression rice sheath blight disease, was preserved in China Microbiological on 26th in September in 2017
Culture presevation administration committee common micro-organisms center, preserving number are：14626.
2. suppress the ratio Lay mould JLF6-2 of rice sheath blight disease as claimed in claim 1, it is characterised in that it is to rice banded sclerotial blight
Germ has good antagonistic effect.
3. suppress the ratio Lay mould JLF6-2 of rice sheath blight disease as claimed in claim 1, it is characterised in that it is in prevention rice
Application in banded sclerotial blight microbial bactericide.
4. suppress the ratio Lay mould JLF6-2 of rice sheath blight disease as claimed in claim 1, it is characterised in that it is in rice banded sclerotial blight
Application in disease prevention pesticide.
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| CN201711113602.8A CN107937278A (en) | 2017-11-13 | 2017-11-13 | One plant prevention rice sheath blight disease than Lay mould |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109439594A (en) * | 2018-12-12 | 2019-03-08 | 沈阳农业大学 | A kind of Death Valley bacillus preventing and treating false smut |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2017137353A1 (en) * | 2016-02-09 | 2017-08-17 | Basf Se | Mixtures and compositions comprising paenibacillus strains or metabolites thereof and other biopesticides |
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2017
- 2017-11-13 CN CN201711113602.8A patent/CN107937278A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2017137353A1 (en) * | 2016-02-09 | 2017-08-17 | Basf Se | Mixtures and compositions comprising paenibacillus strains or metabolites thereof and other biopesticides |
Non-Patent Citations (1)
| Title |
|---|
| 曾金凤: "青霉Z88菌株对水稻纹枯病菌的抗生作用", 《福建农业大学学报》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109439594A (en) * | 2018-12-12 | 2019-03-08 | 沈阳农业大学 | A kind of Death Valley bacillus preventing and treating false smut |
| CN109439594B (en) * | 2018-12-12 | 2022-01-18 | 沈阳农业大学 | Bacillus vallismortis for preventing and treating false smut |
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