CN104059860B - One strain trichoderma pseudokiningii TZ-11 bacterial strain and application thereof - Google Patents
One strain trichoderma pseudokiningii TZ-11 bacterial strain and application thereof Download PDFInfo
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Abstract
The present invention relates to a strain trichoderma pseudokiningii (Trichoderma koningiopsis) TZ 11 bacterial strain, purposes and the preparation method of preparation thereof.The wettable powder prepared through solid fermentation by this bacterial strain, can be used for prevent and treat by GLOMERFLLA CINGULATA bacterium (Glomerella cingulata) infect the preventing and treating of a lot of anthracnose of crop such as the anthrax caused, such as Fructus Capsici, Fructus Mangifera Indicae, Fructus Musae and Folium Eriobotryae etc., especially by GLOMERFLLA CINGULATA bacterium (Glomerella cingulata) infect the diseases such as the bitter rot or anthracnose of grape caused and Strawberry anthracnose.Field test results shows, bitter rot or anthracnose of grape and Strawberry anthracnose are all had good prevention effect and strain fermentation process simple by this microbial inoculum, low production cost；Microbial inoculum spore content alive is high, and storage properties is stable.
Description
Technical field
The present invention relates to strain microbial strains and application thereof, more specifically to a strain trichoderma pseudokiningii (Trichoderma koningiopsis) TZ-11 bacterial strain, purposes and the preparation method of preparation thereof, belong to biological pesticide technical field.
Background technology
The anthrax of Fructus Vitis viniferae, Fructus Mali pumilae, pears and the Fructus Fragariae Ananssae etc. that are caused by plant pathogenic fungi GLOMERFLLA CINGULATA Pseudomonas (Glomerellaspp.), is the Major Diseases of industrialized agriculture crops.Chemical agent prevention and control are the main prophylactico-therapeutic measuress currently preventing and treating anthrax.Conventional chemical agent mainly has carbendazim (carbendazim), propiconazole (propiconazole), Bitertanol (bitertanol), imazalil (imazalil), hexaconazole (hexaconazole), Fluoxastrobin (azoxystrobin), kresoxim-methyl (kresoxim-methyl), pyrimethanil (pyrimethanil), Difenoconazole (difenoconazole), Enestroburin (enestroburin), Prochloraz (prochloraz) and pyraclostrobin (pyraclostrobin) etc..Peasant household is used mostly above-mentioned high-efficiency low-toxicity medicament on anthracnose of crop is prevented and treated, but such pharmacy effect site single (suppression mitochondrial respiratory chain electron transmission).Easily develop immunity to drugs under height prevents and treats the frequency, and between medicament, mutual resistance risk is high.The most repeatedly occurring the example that preventing and treating is failed in production, illustrate that Anthracnose Pathogen fungus all creates anti-(resistance to) property of medicine in various degree to above-mentioned main chemical agent, the research and development of surrogate-data technique are the most very urgent.
Relative to field crop disease Biological control, the Biological control research of the fruits in season disease such as Fructus Vitis viniferae, Fructus Fragariae Ananssae the most just starts in China.The most it is not used successfully to the biological pesticide special preparation of preventing and treating anthrax.nullThe research reported is mostly focused on and utilizes living microorganism to carry out prevention and control，As utilized the Trichoderma viride (Trichoderma viride) in trichoderma fungus (Trichoderma spp.)、Trichoderma harzianum (Trichoderma harzianum) and long shoot Trichoderma spp. (Trichoderma longibrachiatum)，Bacillus subtilis (Bacillus subtilis) in bacillus (Bacillus spp.)、Waxy Bacillus (Bacillus cereus)、Bacillus polymyxa (Bacillus polymyxa)、Bacillus megaterium (Bacillus megaterium) and bacillus amyloliquefaciens (Bacillus amyloliquefaciens)，Pseudomonas cepacia (Pseudomonas cepacia) in Rhodopseudomonas antibacterial (Pseudomonas spp.)、Pseudomonas antibacterial (Pseudomonas fluorescens) and pseudomonas putida (Pseudomonas putida)，Also pichia (Pichia guilliermondii) and Cryptococcus laurentii (Cryptococcus laurentii) are covered the season in yeast，Pythium oligandrum (Pythium oligandrum) etc. is prevented and treated.The research report using living microorganism preventing and treating anthracnose of crop is only limitted to toxicity mensuration, greenhouse controlling experiment and field test and measures the stage.The trichoderma pseudokiningii bacterial strain that can effectively prevent and treat anthrax that the present invention provides, does not the most mention and discloses.
Summary of the invention
The first object of the present invention is that provide one to can be used for preventing and treating is infected the bacterial strain of the anthrax disease caused by GLOMERFLLA CINGULATA bacterium (Glomerella cingulata).
The second object of the present invention is to provide bacterial strain and is infected, in preventing and treating, the application caused in terms of anthrax disease by GLOMERFLLA CINGULATA bacterium (Glomerella cingulata), especially infects the application in the bitter rot or anthracnose of grape and Strawberry anthracnose caused GLOMERFLLA CINGULATA bacterium.
The third object of the present invention is a kind of wettable powder utilizing above-mentioned bacterial strains to prepare.
The fourth object of the present invention is to provide the preparation method of a kind of above-mentioned wettable powder.
One, for solve the first purpose, it is provided that technical scheme as follows:
One strain trichoderma pseudokiningii bacterial strain, it is characterized in that, the Classification And Nomenclature of this bacterial strain is trichoderma pseudokiningii TZ-11 (Trichoderma koningiopsisTZ-11), for this bacterial strain on July 8th, 2010 at the center preservation of China Committee for Culture Collection of Microorganisms's common micro-organisms, deposit number it is: CGMCCNo.3969.
Morphology and the microscopic features of bacterial strain of the present invention are as follows:
Fast in the upper growth of wort agar culture medium (MEA), cultivate 4d colony diameter 60-63mm, initial stage white under 25 DEG C of dark conditions, the later stage is celadon because producing spore, and quality is cotton-shaped；The back side is light brown, without water colo(u)r.Conidium structure is formed in a large number, and Chan Bao district is uniformly distributed.Mycelia wall is smooth or coarse, multi-branched, and tool separates, wide 2.0-3.5 μm；Conidiophore wall is smooth, and main shaft is obvious, less to branch situation estranged, wide 2.0-3.5 μm；Produce most 2-4 bunch of spore bottle stalk to be born on conidiophore or its branch, base portion column or 6.2-15.0 × 2.0-3.5 μm of slightly expanding；Conidium is oval, bottle green, and wall is smooth or the most coarse, 3.5-5.5 × 1.8-3.0 μm.
DNA sequence (this section of rRNA includes 18SrRNA fragment, ITS1,5.8SrRNA, ITS2 district complete sequence and 28SrRNA sequence fragment) corresponding to trichoderma pseudokiningii (Trichoderma koningiopsis) the TZ-11 bacterial strain rRNA gene order of the present invention is as shown in SEQ ID NO:1.
Two, bacterial strain is infected the application in terms of the anthrax disease caused, the especially application in the microbial bitter rot or anthracnose of grape of GLOMERFLLA CINGULATA and Strawberry anthracnose are prevented and treated in preventing and treating by GLOMERFLLA CINGULATA bacterium (Glomerellacingulata).
Three, a kind of wettable powder utilizing above-mentioned bacterial strains to prepare.
Four, the preparation method of a kind of above-mentioned wettable powder, specifically includes following steps:
(A) acquisition of solid fermentation culture
(1) after Testa Tritici and wood sawdust being mixed in the ratio of 3:7 (V/V), it is separately added into sucrose and the ammonium carbonate of 0.1% (W/W), and add water that to make water content be 60～65%, pH is adjusted to 7.5-8.0, again mixing after thicken polyethylene plastic bag in every bag of subpackage 1.5kg, tighten bag mouth, and moist heat sterilization 60min at 121 DEG C.At level trough 2 sterilizing newspapers of solid fermentation tray bottom place mat, adding solid fermentation culture medium 1.5kg after sterilizing, charging thickness is about 5cm.The trichoderma pseudokiningii TZ-11 bacterial strain that will be transferred in advance in PDA plate, be placed in incubator 28 DEG C cultivate 7d after, with the spore under aseptic washing, with after microscopic counting, spore liquid concentration is allocated as: 1.0 × 107Individual/mL.After this spore liquid of 100mL is uniformly inoculated into 1.5kg solid fermentation culture medium, at 2 sterilizing newspapers of fermentation dish top cover and tighten sealing with jag, finally it is positioned over fermentation in 28 DEG C of moisturizing incubators, yeasting relative humidity reaches more than 80%, fermentation 8-10d, tunning lives spore amount up to 1.0-2.0 × 109cfu/g；
(2) the above-mentioned tunning appropriate distilled water of addition is placed in 150rpm vibration washing 8min on shaking table, then filters with 325 mesh standard sieves and collect filtrate, 3 times repeatedly, finally filtrate is centrifuged at 3,000 rpm 15min, collects precipitate standby.
(B) microbial inoculum processing technique
(1) will suspend through solid fermentation, collected after centrifugation to precipitate 0.5% (W/W) methocel solution (solution sterilized water is prepared), and the content of spore of living in suspension will be adjusted to >=5.0 × 108cfu/mL；
(2) after sodium metnylene bis-naphthalene sulfonate, calcium dodecyl benzene sulfonate, precipitated calcium carbonate and kieselguhr are pressed the weight ratio mixing of 5:4:1:90, fineness is made to reach more than 325 mesh by comminution by gas stream, be subsequently adding the spore suspension prepared, after become the wettable powder containing trichoderma pseudokiningii TZ-11 bacterial strain work spore through 50 DEG C of airflow dryings, blender mixing and packaging post-treatment.Spore content >=1.0 × 10 alive of wherein said trichoderma pseudokiningii TZ-11 bacterial strain wettable powder8cfu/g。
Beneficial effects of the present invention
(1) the present invention is to solve the problem of anthrax preventing and treating in the arable farming such as Fructus Vitis viniferae and Fructus Fragariae Ananssae, in conjunction with agricultural production actual demand, the biocontrol bacterial strain trichoderma pseudokiningii TZ-11 bacterial strain that one strain is new is provided, the wettable powder prepared through solid fermentation by this bacterial strain, can be used for preventing and treating and infected, by GLOMERFLLA CINGULATA bacterium (Glomerella cingulata), the anthrax caused, such as Fructus Capsici, Fructus Mangifera Indicae, the preventing and treating of a lot of anthracnose of crop such as Fructus Musae and Folium Eriobotryae, especially infected the diseases such as the bitter rot or anthracnose of grape caused and Strawberry anthracnose by GLOMERFLLA CINGULATA bacterium (Glomerella cingulata).Field test results shows, this microbial inoculum all has good prevention effect to bitter rot or anthracnose of grape and Strawberry anthracnose.
(2) strain fermentation process is simple, low production cost；Microbial inoculum spore content alive is high, and storage properties is stable.
Trichoderma pseudokiningii TZ-11 (the Trichoderma koningiopsis TZ-11) bacterial strain of the present invention (is called for short CGMCC on July 8th, 2010 at China Committee for Culture Collection of Microorganisms's common micro-organisms center, address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Institute of Microorganism, Academia Sinica) preservation, deposit number is: CGMCCNo.3969.
Detailed description of the invention
Embodiment is set forth below, and the present invention is further described, but is not so limited present disclosure.
Embodiment 1 this example demonstrates that the method obtaining trichoderma pseudokiningii (Trichoderma koningiopsis) TZ-11 bacterial strain.
PDA culture medium prescription is: Rhizoma Solani tuber osi (peeling) 200g/L, glucose 20g/L, water 1L, and agar 20g/L, pH are natural.By peeling potatoes, being cut into small pieces, the about 1L that adds water in pot boils half an hour, filters with double gauze, takes its filtrate and adds glucose, and adds water and supply 1L.
Comprise the following steps:
1) separation of bacterial strain, purification and preservation.
Sample is gathered on kyoto grape in grape growing region, Jurong City of Jiangsu Province, the sample of collection is carried out on PDA plate separate tissue by superclean bench, colony characteristics timely picking different shape list bacterium colony according to fungus is on PDA plate, purification 2 times, record bacterial strain number, puts in 4 DEG C of refrigerators standby after forwarding to the bacterial strain of purification cultivate well on PDA inclined-plane.Then the fungal bacterial strain switching above-mentioned separation preserved activates on PDA plate, uses the antagonistic strain of plate opposite culture method screening GLOMERFLLA CINGULATA bacterium (Glomerella cingulata), evaluate the antagonistic activity of bacterial strain according to the size of inhibition zone.
2) bacterial strain kind preservation are identified.
After strain passage purification the strongest for antagonistic activity is cultivated, bacterial strain is carried out Molecular Identification, qualification result display bacterial strain is trichoderma pseudokiningii (Trichoderma koningiopsis), to its named trichoderma pseudokiningii (Trichoderma koningiopsis) TZ-11 bacterial strain, and the lyophilization pipe making trichoderma pseudokiningii (Trichoderma koningiopsis) TZ-11 bacterial strain carries out the cold storage of original strain.
The colonial morphology of above-mentioned strain and microscopic features are: fast in the upper growth of wort agar culture medium (MEA), cultivate 4d colony diameter 60-63mm, initial stage white under 25 DEG C of dark conditions, and the later stage is celadon because producing spore, and quality is cotton-shaped；The back side is light brown, without water colo(u)r.Conidium structure is formed in a large number, and Chan Bao district is uniformly distributed.Mycelia wall is smooth or coarse, multi-branched, and tool separates, wide 2.0-3.5 μm；Conidiophore wall is smooth, and main shaft is obvious, less to branch situation estranged, wide 2.0-3.5 μm；Produce most 2-4 bunch of spore bottle stalk to be born on conidiophore or its branch, base portion column or 6.2-15.0 × 2.0-3.5 μm of slightly expanding；Conidium is oval, bottle green, and wall is smooth or the most coarse, 3.5-5.5 × 1.8-3.0 μm.
DNA sequence (this section of rRNA includes 18SrRNA fragment, ITS1,5.8SrRNA, ITS2 district complete sequence and 28SrRNA sequence fragment) corresponding to trichoderma pseudokiningii (Trichoderma koningiopsis) the TZ-11 bacterial strain rRNA gene order of the present invention is as shown in SEQIDNO:1..
The embodiment 2 bacterial strain TZ-11 of the preventing and treating anthrax of the present invention prepares wettable powder
Preparation method comprises the following steps:
(A) acquisition of solid fermentation culture
(1) after Testa Tritici and wood sawdust being mixed in the ratio of 3:7 (V/V), it is separately added into sucrose and the ammonium carbonate of 0.1% (W/W), and add water that to make water content be 60～65%, pH is adjusted to 7.5-8.0, again mixing after thicken polyethylene plastic bag in every bag of subpackage 1.5kg, tighten bag mouth, and moist heat sterilization 60min at 121 DEG C.At level trough 2 sterilizing newspapers of solid fermentation tray bottom place mat, adding solid fermentation culture medium 1.5kg after sterilizing, charging thickness is about 5cm.The trichoderma pseudokiningii TZ-11 bacterial strain that will be transferred in advance in PDA plate, be placed in incubator 28 DEG C cultivate 7d after, with the spore under aseptic washing, with after microscopic counting, spore liquid concentration is allocated as: 1.0 × 107Individual/mL.After this spore liquid of 100mL is uniformly inoculated into 1.5kg solid fermentation culture medium, at 2 sterilizing newspapers of fermentation dish top cover and tighten sealing with jag, finally it is positioned over fermentation in 28 DEG C of moisturizing incubators, yeasting relative humidity reaches more than 80%, fermentation 8-10d, tunning lives spore amount up to 1.0-2.0 × 109cfu/g；(2) the above-mentioned tunning appropriate distilled water of addition is placed in 150rpm vibration washing 8min on shaking table, then filters with 325 mesh standard sieves and collect filtrate, 3 times repeatedly, finally filtrate is centrifuged at 3,000 rpm 15min, collects precipitate standby.
(B) microbial inoculum processing technique
(1) will suspend through solid fermentation, collected after centrifugation to precipitate 0.5% (W/W) methocel solution (solution sterilized water is prepared), and the content of spore of living in suspension will be adjusted to >=5.0 × 108cfu/mL；(2) after sodium metnylene bis-naphthalene sulfonate, calcium dodecyl benzene sulfonate, precipitated calcium carbonate and kieselguhr are pressed the weight ratio mixing of 5:4:1:90, fineness is made to reach more than 325 mesh by comminution by gas stream, be subsequently adding the spore suspension prepared, after become the wettable powder containing trichoderma pseudokiningii TZ-11 bacterial strain work spore through 50 DEG C of airflow dryings, blender mixing and packaging post-treatment.Spore content >=1.0 × 10 alive of wherein said trichoderma pseudokiningii TZ-11 bacterial strain wettable powder8cfu/g。
Implement 3 test example
(1) the TZ-11 bacterial strain wettable powder indoor virulence to GLOMERFLLA CINGULATA bacterium (Glomerella cingulata)
TZ-11 bacterial strain wettable powder is as follows to the Toxicity Determination method of GLOMERFLLA CINGULATA bacterium:
Respectively with sterilized water by 1.0 × 108Cfu/gTZ-11 bacterial strain wettable powder and 1.0 × 106Cfu/g pythium oligandrum wettable powder (Beijing is than Ao Rui bio tech ltd) is diluted to finite concentration successively, again 1mL medicinal liquid and 9mLPDA culture medium are mixed in culture dish, making the PDA culture medium containing graded series drug concentrations, each medicament graded series concentration in PDA culture medium is both designed as 1.0 × 105、5.0×104、1.0×104、5.0×103、1.0×103、5.0×102With 1.0 × 102Cfu/mL, uses sterilized water to compare, and each process is repeated 4 times.The GLOMERFLLA CINGULATA bacterium of reservation is transplanted in PDA plate, 96h is activated at 25 DEG C, then the bacterium cake of a diameter of 5mm is produced at nearly colony edge card punch, and be transplanted in the PDA plate of above-mentioned pastille and blank, cultivate 96h for 25 DEG C, when the 4/5 of bacterium colony length in compareing to about plate diameter, decussation method is used to measure colony diameter.Calculate colony diameter meansigma methods, and calculate mycelial growth average inhibition according to the following formula: mycelial growth average inhibition={ (comparison colony diameter average-process colony diameter average)/(comparison colony diameter average-inoculation bacterium cake diameter) } × 100%.Use DPS data handling system, calculate regression equation, EC that GLOMERFLLA CINGULATA bacterium mycelial growth is suppressed by each medicament50And 95% confidence limit.
The TZ-11 bacterial strain wettable powder Toxicity Determination result to GLOMERFLLA CINGULATA bacterium:
Toxicity Determination result (table 1) shows, the EC that GLOMERFLLA CINGULATA bacterium mycelial growth is suppressed by TZ-11 bacterial strain wettable powder and pythium oligandrum wettable powder50It is respectively as follows: 8.2979 × 102With 1.4434 × 103μ g/mL, differs about 1.7 times, and the activity that GLOMERFLLA CINGULATA bacterium mycelial growth is suppressed by TZ-11 bacterial strain wettable powder is higher than pythium oligandrum wettable powder.
The wettable powder of the table 1TZ-11 bacterial strain Toxicity Determination result to GLOMERFLLA CINGULATA bacterium (Glomerella cingulata)
(2) the TZ-11 bacterial strain wettable powder field control effectiveness test to bitter rot or anthracnose of grape
Test processing method: test site is positioned at vineyard, Jiangsu Hilly Ground Zhenjiang Agriculture Science Research Institute, and kind is huge peak, horizontal rack formula is cultivated, the age of tree about 20 years.Soil is Hepar Equi soil, and middle fertility, the content of organic matter 1.75%, during test, orchard management routine.Test sets: TZ-11 bacterial strain wettable powder 1000 and 2000 times of liquid, 8000 times of liquid of pythium oligandrum wettable powder (as recommended dose, need 30 minutes in advance preparation stock solution before spraying), and with clear water spraying for comparison, spray amount is 900kg/hm2, each plot area 100m2, random district group arranges, and repeats for 4 times.Test in the 1st spray medicine on June 3, prevent and treat once every 10d, altogether preventing and treating 5 times later.After 5 medicines, 14d investigates the incidence of each community, select 4 strain Fructus Vitis viniferaes during investigation the most at random, in upper, middle and lower, orientation, 5, left and right take 1 fruit ear, totally 20 fruit ears respectively, a situation arises to record disease in fruit ear axle, fruit grain and fruit stem, calculates disease and refers to and prevention effect.
Field controling test result (table 2) shows, TZ-11 bacterial strain wettable powder 1000 and 2000 times of liquid are respectively 88.08% and 73.37%, significant difference to the preventive effect of bitter rot or anthracnose of grape；8000 times of liquid of pythium oligandrum wettable powder are 80.27% to the preventive effect of bitter rot or anthracnose of grape, and preventive effect is less than 1000 times of liquid of TZ-11 bacterial strain wettable powder, significant difference.
The table 2TZ-11 bacterial strain wettable powder field control effectiveness test result to bitter rot or anthracnose of grape
Note: different after data lowercase alphabet differential is different reaches significant level (p < 0.05)
(3) the TZ-11 bacterial strain wettable powder field control effectiveness test to Strawberry anthracnose
Test processing method: test site is positioned at the Fructus Fragariae Ananssae Demonstration Garden of white rabbit town, Jurong City of Jiangsu Province cloud rabbitweed certain kind of berries cooperative society, and kind is red cheek.Test sets: TZ-11 bacterial strain wettable powder 1000 and 2000 times of liquid, 8000 times of liquid of pythium oligandrum wettable powder (as recommended dose, need 30 minutes in advance preparation stock solution before spraying), and with clear water spraying for comparison, spray amount is 900kg/hm2, each plot area 50m2, random district group arranges, and repeats for 4 times.Test in the 1st spray medicine on May 3rd, 2013, prevent and treat once every 10d, altogether preventing and treating 6 times later.After 6 medicines, 14d investigates the incidence of each community, and in sampling survey about 100 strain of 5, each community during investigation, a situation arises to record every strain Strawberry anthracnose, calculates disease and refers to and prevention effect.
Field controling test result (table 3) shows, TZ-11 bacterial strain wettable powder 1000 and 2000 times of liquid are respectively 85.63% and 69.92%, significant difference to the preventive effect of Strawberry anthracnose；8000 times of liquid of pythium oligandrum wettable powder are 80.38% to the preventive effect of Strawberry anthracnose, and preventive effect is less than 1000 times of liquid of TZ-11 bacterial strain wettable powder, significant difference.
The table 3TZ-11 bacterial strain wettable powder field control effectiveness test result to Strawberry anthracnose
Note: different after data lowercase alphabet differential is different reaches significant level (p < 0.05).
Sequence table
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Claims (4)
1. a strain trichoderma pseudokiningii bacterial strain, it is characterised in that the Classification And Nomenclature of this bacterial strain be trichoderma pseudokiningii TZ-11 (Trichoderma
koningiopsisTZ-11), this bacterial strain on July 8th, 2010 at the center preservation of China Committee for Culture Collection of Microorganisms's common micro-organisms, deposit number is: CGMCC No.3969.
2. the bacterial strain described in claim 1 preventing and treating by GLOMERFLLA CINGULATA bacterium (Glomerella cingulata) infect the application in terms of the anthrax disease caused.
3. the wettable powder prepared with the bacterial strain described in claim 1.
4. the method preparing wettable powder according to claim 3, it is characterised in that comprise the following steps:
(A) acquisition of solid fermentation culture
(1) after Testa Tritici and wood sawdust being mixed in the ratio of 3:7 (V/V), it is separately added into sucrose and the ammonium carbonate of 0.1% (W/W), and add water that to make water content be 60 ~ 65%, pH is adjusted to 7.5-8.0, again mixing after thicken polyethylene plastic bag in every bag of subpackage 1.5kg, tighten bag mouth, and moist heat sterilization 60 min at 121 DEG C；At level trough 2 sterilizing newspapers of solid fermentation tray bottom place mat, adding the solid fermentation after sterilizing and cultivate based 1.5 kg, charging thickness is about 5 cm；
The trichoderma pseudokiningii TZ-11 bacterial strain that will be transferred in advance in PDA plate, be placed in incubator 28 DEG C cultivate 7d after, with the spore under aseptic washing, with after microscopic counting, spore liquid concentration is allocated as: 1.0 × 107Individual/mL；After this spore liquid of 100mL is uniformly inoculated into 1.5 kg solid fermentation culture medium, at 2 sterilizing newspapers of fermentation dish top cover and tighten sealing with jag, finally it is positioned over fermentation in 28 DEG C of moisturizing incubators, yeasting relative humidity reaches more than 80%, fermentation 8-10d, tunning lives spore amount up to 1.0-2.0 × 109cfu/g；
(2) the above-mentioned tunning appropriate distilled water of addition is placed in 150rpm vibration washing 8min on shaking table, then filters with 325 mesh standard sieves and collect filtrate, 3 times repeatedly, finally filtrate is centrifuged at 3,000 rpm 15min, collects precipitate standby；
(B) microbial inoculum processing technique
(1) will suspend through solid fermentation, collected after centrifugation to precipitate 0.5% (W/W) methocel solution, wherein solution sterilized water is prepared, and the content of spore of living in suspension is adjusted to >=5.0 × 108cfu/mL；
(2) after sodium metnylene bis-naphthalene sulfonate, calcium dodecyl benzene sulfonate, precipitated calcium carbonate and kieselguhr are pressed the weight ratio mixing of 5:4:1:90, fineness is made to reach more than 325 mesh by comminution by gas stream, be subsequently adding the spore suspension prepared, after become the wettable powder containing trichoderma pseudokiningii TZ-11 bacterial strain work spore through 50 DEG C of airflow dryings, blender mixing and packaging post-treatment.
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| CN102586130B (en) * | 2011-01-14 | 2015-01-07 | 辽宁省农业科学院 | Bio-control bacterial strain and bio-control bactericide capable of preventing grape anthracnose and application thereof |
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| CN102876605B (en) * | 2012-09-23 | 2014-02-19 | 江苏丘陵地区镇江农业科学研究所 | Bacillus subtilis DJ-6 and application thereof in prevention and treatment of grape disease |
| CN102876603B (en) * | 2012-09-23 | 2013-11-13 | 江苏丘陵地区镇江农业科学研究所 | Bacillus amyloliquefaciens Mg116 and application thereof |
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| CN107629970A (en) * | 2017-11-13 | 2018-01-26 | 中国科学院天津工业生物技术研究所 | A kind of microorganism and application for being used to prevent and treat ginseng crythrodermia |
| CN107629970B (en) * | 2017-11-13 | 2019-12-20 | 中国科学院天津工业生物技术研究所 | Microorganism for preventing and treating ginseng erythroderma and application |
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