CN106967621A - Prevent and treat the complex microorganism preparations and its production method and purposes of rice seedling blight - Google Patents
Prevent and treat the complex microorganism preparations and its production method and purposes of rice seedling blight Download PDFInfo
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Abstract
Preventing and treating rice seedling blight complex microorganism preparations and its production method and purposes are acted on the present invention relates to one kind.The microbial inoculum includes Trichoderma harzianum Q2 (Trichoderma harzianum), saccharomyces cerevisiae T1 (Saccharomyces cerevisiae), bacillus amyloliquefaciens J12 (Bacillus amyloliquefaciens).By inclined-plane culture, first order seed, secondary seed and mixed culture fermentation, realizing compound bacteria most has proportioning and optimizes fermenting and producing, composite bacteria agent You effects Jun Shuo≤2.5 × 10 living9cfu/mL.The complex microorganism preparations that the present invention is provided, for preventing and treating paddy rice soil-borne disease, disease-controlling effect is obvious.
Description
Technical field
It is more particularly to a kind of to be used for rice seedling blight preventing and treating the present invention relates to a kind of complex microorganism preparations.The present invention is also
It is related to production method of the complex micro organism fungicide and application thereof.
Technical background
Rice seedling blight is that a kind of destructive disease that arid rice nursery is often sent out practises residence by soil such as sickle-like bacteria, rhizoctonias
Bacterium and bad weather, environmental condition (low temperature, saline and alkaline etc.) synthesis cause.Control is improper to be easily caused large area dead seedling, ruins
The phenomenon of seedling, not only waste of manpower, financial resources, it is often more important that miss the farming season, influence seedling quality and progress, particularly machinery are made
Industry, reduces rice yield, therefore very harmful.
At present, the preventing and treating of China's diseases and pests of agronomic crop relies primarily on chemical pesticide.The use of chemical pesticide is really to disease pest
Harmful preventing and treating has served very important, but also result in a series of environment and social concern.Chemical pesticide is agriculture face
Source pollution main source, and impact of the agricultural non-point source pollution to ecological environment be in all directions, be related to taking human as
The whole food chain at center, wherein the harm caused to water body is the most direct, the most notable.It is the drawbacks of due to chemical pesticide, micro-
Biological product is with its free from environmental pollution, agriculture and forestry of keeping ecological balance, keep sustainable development, to people and animals and pest natural enemy pole
Less or completely no toxic action the characteristics of increasingly paid close attention to by people.Therefore, the present invention is by sending out rice seedling blight
The serious soil of sick evil, carries out isolating and purifying for microorganism, filtering out has the short of money of better inhibition effect to rice seedling blight
Antibacterial, and by the antagonism between bacterial strain and the proportioning and fermentation condition of optimization strain, prepare stabilization, be efficiently combined
Microorganism formulation product.
The content of the invention
(1) technical problem to be solved
It is an object of the invention to for above-mentioned the deficiencies in the prior art, disclose a kind of complex microorganism preparations, be used for
Anti- rice seedling blight of harnessing the river.The invention also discloses the production method of the microbial inoculum and purposes.
(2) technical scheme
The present invention filters out best stabilized symbiosis strain combination from 23 different genus and species, high by individually cultivating screening
Produce strain.The strain filtered out is Trichoderma harzianum Q2 (Trichoderma harzianum), saccharomyces cerevisiae T1
(Saccharomyces cerevisiae), bacillus amyloliquefaciens J12 (Bacillus amyloliquefaciens).
According to above-mentioned the selection result, the present invention devises a kind of complex microorganism preparations, including Trichoderma harzianum Q2
(Trichoderma harzianum)0.5×109Cfu/mL, saccharomyces cerevisiae T1 (Saccharomyces cerevisiae) 0.5
×109Cfu/mL, bacillus amyloliquefaciens J12 (Bacillus amyloliquefaciens) 1.5 × 109cfu/mL；
Preferably,
Trichoderma harzianum Q2 (Trichoderma harzianum) 0.8 × 109Cfu/mL, saccharomyces cerevisiae T1
(Saccharomyces cerevisiae)0.8×109Cfu/mL, bacillus amyloliquefaciens J12 (Bacillus
amyloliquefaciens)1.2×109cfu/mL；
The production method of complex microorganism preparations of the present invention, comprises the following steps：
1) it is inoculated in respectively under Trichoderma harzianum Q2, saccharomyces cerevisiae T1, bacillus amyloliquefaciens J12 original strain aseptic conditions
On solid medium, Trichoderma harzianum Q2 and saccharomyces cerevisiae T1 are cultivated 4 days under the conditions of 30 DEG C；；Bacillus amyloliquefaciens J12 is in 37
Cultivated 2 days under the conditions of DEG C；
2) first order seeds culture：By step 1) culture strain aseptic condition under be inoculated in fluid nutrient medium, Ha Ci respectively
Trichoderma Q2, saccharomyces cerevisiae T1 cultivated under the conditions of 30 DEG C 4 days, bacillus amyloliquefaciens J12 cultivated 2 days under the conditions of 37 DEG C, make
First order seed, terminate each saccharomyces cerevisiae of culture, bacillus amyloliquefaciens suspension optical density OD600 values respectively reach 4.0,
3.0；
3) secondary seeds culture：The inoculum concentration for being 15% by the volume ratio of fluid nutrient medium, first order seed is inoculated with respectively
Into 100L fermentation tank, under the conditions of the cumulative volume of nutrient solution is 70L, 30 DEG C -35 DEG C in fermentation tank, mixing speed is 200r/
Min, throughput is 1：2.0, Trichoderma harzianum Q2, saccharomyces cerevisiae T1 are cultivated 3 days, and bacillus amyloliquefaciens J12 is cultivated 1 day and is made two
Level seed；
4) mixing fermentation cultures：The inoculum concentration for being 15% by the volume ratio of fluid nutrient medium, 1 is inoculated into by secondary seed
In the fermentation tank of ton, the culture medium cumulative volume in fermentation tank is 700L, carries out fermented and cultured, obtains microbial inoculum.
Wherein, step 1), 2), 3) in used culture medium Trichoderma harzianum Q2, saccharomyces cerevisiae T1 be PDA culture medium, Xie Dian
Afnyloliquefaciens J12 is LB culture mediums.
Wherein, step 4) used in the formula of culture medium be by mass percentage：Molasses 10%, fish peptone 2%, soya-bean cake
Powder 15%, surplus is water；
Fermented and cultured uses fed-batch process, and wherein feed supplement carbon source is：Glucose；Nitrogen source is：Fish peptone.
Fermentation culture stage：In starting 0~24 hour, interval ventilation is maintained at aerobic conditions fermentation, throughput 1：2.0,
Speed of agitator 200r/min, stirs 3.5 hours interval times, stirs 4 minutes, 30 DEG C -35 DEG C of temperature.
Complex microorganism preparations of the present invention can be used to prevent and treat rice seedling blight as biological prevention and control agent.
(3) beneficial effect
The present invention is used to prevent and treat rice seedling blight soil-borne disease, and action effect is obvious.
Embodiment
With reference to embodiment, the present invention is further illustrated, but is not limited to claimed model of the invention
Enclose.
The composition of the complex microorganism preparations of embodiment 1
Total bacteria count is 2.8 × 109cfu/mL
Trichoderma harzianum Q2 0.8 × 109cfu/mL
Saccharomyces cerevisiae T1 0.8 × 109cfu/mL
Bacillus amyloliquefaciens J 121.2 × 109cfu/mL
The composition of the complex microorganism preparations of embodiment 2
Total bacteria count is 2.5 × 109cfu/mL
Trichoderma harzianum Q2 1.0 × 109cfu/mL
Saccharomyces cerevisiae T1 1.0 × 109cfu/mL
Bacillus amyloliquefaciens J 120.5 × 109cfu/mL
The production method of the complex microorganism preparations of embodiment 3
1) inclined-plane cultures：Under Trichoderma harzianum Q2, saccharomyces cerevisiae T1, bacillus amyloliquefaciens J12 original strain aseptic conditions
It is inoculated in respectively on solid medium, Trichoderma harzianum Q2 and saccharomyces cerevisiae T1 are cultivated 3-4 days under the conditions of 30 DEG C；；Solve starch bud
Spore bacillus J12, is cultivated 1-2 days under the conditions of 37 DEG C.
2) first order seeds culture：By step 1) culture strain aseptic condition under be inoculated in fluid nutrient medium, Ha Ci respectively
Trichoderma Q2, saccharomyces cerevisiae T1 cultivated under the conditions of 30 DEG C 3-4 days, bacillus amyloliquefaciens J12 cultivate 1-2 under the conditions of 37 DEG C
My god, first order seed is made, terminates each saccharomyces cerevisiae of culture, bacillus amyloliquefaciens suspension optical density OD600 values and respectively reaches
4.0、3.0；
3) secondary seeds culture：The inoculum concentration for being 10% by the volume ratio of fluid nutrient medium, first order seed is inoculated with respectively
Into 70L fermentation tank, under the conditions of the cumulative volume of nutrient solution is 100L, 25 DEG C -35 DEG C in fermentation tank, mixing speed is 120r/-
200min, throughput is 1：1.5-2.0, Trichoderma harzianum Q2, saccharomyces cerevisiae T1 are cultivated 3-4 days, bacillus amyloliquefaciens J12 cultures
1-2 days obtained secondary seeds；
4) mixing fermentation cultures：The inoculum concentration for being 10-15% by the volume ratio of fluid nutrient medium, secondary seed is inoculated with
Into 1 ton of fermentation tank, the culture medium cumulative volume in fermentation tank is 700L, carries out fermented and cultured, obtains microbial inoculum,
Wherein, step 1), 2), 3) in used culture medium Trichoderma harzianum Q2, saccharomyces cerevisiae T1 be PDA culture medium, Xie Dian
Afnyloliquefaciens J12 is LB culture mediums.
Wherein, step 4) used in the formula of culture medium be by mass percentage：Molasses 8-10%, fish peptone 2-5%,
Beancake powder 10-15%, surplus is water；
Fermented and cultured uses fed-batch process, and wherein feed supplement carbon source is：Glucose, sucrose；Nitrogen source is：Fish protein
Peptone, beancake powder.
Fermentation culture stage：In starting 0~24 hour, interval ventilation is maintained at aerobic conditions fermentation, throughput 1：1.8,
Mixing speed 150r-200/min, stirs interval time 3-5 hour, stirs 3-5 minutes, 25 DEG C -35 DEG C of temperature.
The production method of the complex microorganism preparations of embodiment 4
1) it is inoculated in respectively under Trichoderma harzianum Q2, saccharomyces cerevisiae T1, bacillus amyloliquefaciens J12 original strain aseptic conditions
On solid medium, Trichoderma harzianum Q2 and saccharomyces cerevisiae T1 are cultivated 4 days under the conditions of 30 DEG C；Bacillus amyloliquefaciens J12 is in 37
Cultivated 2 days under the conditions of DEG C；
2) first order seeds culture：By step 1) culture strain aseptic condition under be inoculated in fluid nutrient medium, Ha Ci respectively
Trichoderma Q2, saccharomyces cerevisiae T1 cultivated under the conditions of 30 DEG C 4 days, bacillus amyloliquefaciens J12 cultivated 2 days under the conditions of 37 DEG C, make
First order seed, terminate each saccharomyces cerevisiae of culture, bacillus amyloliquefaciens suspension optical density OD600 values respectively reach 4.0,
3.0；
3) secondary seeds culture：The inoculum concentration for being 15% by the volume ratio of fluid nutrient medium, first order seed is inoculated with respectively
Into 100L fermentation tank, under the conditions of the cumulative volume of nutrient solution is 70L, 30 DEG C -35 DEG C in fermentation tank, mixing speed is 200r/
Min, throughput is 1：2.0, Trichoderma harzianum Q2, saccharomyces cerevisiae T1 are cultivated 3 days, and bacillus amyloliquefaciens J12 is cultivated 1 day and is made two
Level seed；
4) mixing fermentation cultures：The inoculum concentration for being 15% by the volume ratio of fluid nutrient medium, 1 is inoculated into by secondary seed
In the fermentation tank of ton, the culture medium cumulative volume in fermentation tank is 700L, carries out fermented and cultured, obtains microbial inoculum.
Wherein, step 1), 2), 3) in used culture medium Trichoderma harzianum Q2, saccharomyces cerevisiae T1 be PDA culture medium, Xie Dian
Afnyloliquefaciens J12 is LB culture mediums
Wherein, step 4) used in the formula of culture medium be by mass percentage：Molasses 10%, fish peptone 2%, soya-bean cake
Powder 15%, surplus is water；
Fermented and cultured uses fed-batch process, and wherein feed supplement carbon source is：Glucose；Nitrogen source is：Fish peptone.
Fermentation culture stage：In starting 0~24 hour, interval ventilation is maintained at aerobic conditions fermentation, throughput 1：2.0,
Speed of agitator 200r/min, stirs 3.5 hours interval times, stirs 4 minutes, 30 DEG C -35 DEG C of temperature.
Prevention effect of the embodiment 5 to rice seedling blight
Method：Using the turfy soil of sterilizing as matrix, alms bowl nursery of nursing young plants in hothouses fills soil 400g per basin and sows 20 seeds,
Each processing is repeated 4 times, and seed treatment is pressed respectively by the complex microorganism preparations of example one and 50% carbendazol wettable powder
0.3% seed dressing.It is placed in greenhouse and grows, is kept the skin wet daily depending on rice seedlings growing state.Rhizoctonia solani Kuhn inoculation is using bacterium soil
Method：The Rhizoctonia solani Kuhn germ for cultivating 10d is inoculated in 1: 4 paddy rice, on sand culture medium, after 1 week by 2% connect bacterium amount with
The turfy soil of sterilizing is well mixed, and is sub-packed in pot for growing seedlings, is then again sowed the rice paddy seed handled through different agents respectively
In seedlings nursing plate, put in 25-28 DEG C of greenhouse of temperature and cultivate.20d investigates the incidence of disease of each basin rice plant, system after each processing is emerged
Count the prevention effect of each processing.25d carries out seedling quality investigation after each processing basin rice seedling, will pull out rice seedling 10 at random per basin
Strain, measures its plant height, root length, fresh weight, dry weight.
Bacterium number × 100 of seedling number/investigation of the incidence of disease (%)=morbidity
Prevention effect (%)=(the control incidence of disease-processing incidence of disease)/control incidence of disease × 100
Processing：
Handle the microbial inoculum of 1 example one
Handle 2 carbendazim
CK clear water is compareed
The prevention effect to seedling quality of the microbial rice seedling blight of the miliary damping-off of table 1
As seen from the data in Table 1, carbendazim is fallen ill with the complex microorganism preparations configured according to example one to rice seedling blight
To be significantly stronger than using carbendazim processing with the seedling quality of preventive effect difference very little, but administration microbial inoculum, in current country for agriculture
Medicine is applied under the policy of zero growth rate and a series of harm for being brought using agricultural chemicals, it is seen that complex microorganism preparations have wide
Application prospect.
Claims (6)
1. according to a kind of complex microorganism preparations of claim 1, it is characterised in that the microbial inoculum includes Trichoderma harzianum Q2
(Trichoderma harzianum)0.5×109Cfu/mL, saccharomyces cerevisiae T1 (Saccharomyces cerevisiae) 0.5
×109Cfu/mL, bacillus amyloliquefaciens J12 (Bacillus amyloliquefaciens) 1.5 × 109cfu/mL。
2. complex microorganism preparations according to claim 1, it is characterised in that the microbial inoculum includes Trichoderma harzianum Q2
(Trichoderma harzianum)0.8×109Cfu/mL, saccharomyces cerevisiae T1 (Saccharomyces cerevisiae) 0.8
×109Cfu/mL, bacillus amyloliquefaciens J12 (Bacillus amyloliquefaciens) 1.2 × 109cfu/mL。
3. complex microorganism preparations according to claim 1, it is characterised in that the microbial inoculum includes Trichoderma harzianum Q2
(Trichoderma harzianum)1.0×109Cfu/mL, saccharomyces cerevisiae T1 (Saccharomyces cerevisiae) 1.0
×109Cfu/mL, bacillus amyloliquefaciens J12 (Bacillus amyloliquefaciens) 0.5 × 109cfu/mL。
4. the production method of a kind of complex microorganism preparations as described in any one of claims 1 to 3, it is characterized in that comprising as follows
Step：
1) inclined-plane cultures：Under Trichoderma harzianum Q2, saccharomyces cerevisiae T1, bacillus amyloliquefaciens J12 original strain aseptic conditions respectively
It is inoculated on solid medium, Trichoderma harzianum Q2 and saccharomyces cerevisiae T1 are cultivated 3-4 days under the conditions of 30 DEG C；；Solve starch gemma bar
Bacterium J12, is cultivated 1-2 days under the conditions of 37 DEG C；
2) first order seeds culture：By step 1) culture strain aseptic condition under be inoculated in fluid nutrient medium, Trichoderma harzianum respectively
Q2, saccharomyces cerevisiae T1 cultivated under the conditions of 30 DEG C 3-4 days, bacillus amyloliquefaciens J12 cultivated 1-2 days under the conditions of 37 DEG C, make
First order seed, terminate each saccharomyces cerevisiae of culture, bacillus amyloliquefaciens suspension optical density OD600 values respectively reach 4.0,
3.0；
3) secondary seeds culture：The inoculum concentration for being 10% by the volume ratio of fluid nutrient medium, first order seed is inoculated into respectively
In 70L fermentation tank, under the conditions of the cumulative volume of nutrient solution is 100L, 25 DEG C -35 DEG C in fermentation tank, mixing speed is 120r/-
200min, throughput is 1：1.5-2.0, Trichoderma harzianum Q2, saccharomyces cerevisiae T1 are cultivated 3-4 days, bacillus amyloliquefaciens J12 cultures
1-2 days obtained secondary seeds；
4) mixing fermentation cultures：The inoculum concentration for being 10-15% by the volume ratio of fluid nutrient medium, 1 is inoculated into by secondary seed
In the fermentation tank of ton, the culture medium cumulative volume in fermentation tank is 700L, carries out fermented and cultured, obtains microbial inoculum,
Wherein, step 1), 2), 3) in used culture medium Trichoderma harzianum Q2, saccharomyces cerevisiae T1 be PDA culture medium, solve starch bud
Spore bacillus J12 is LB culture mediums；
Wherein, step 4) used in the formula of culture medium be by mass percentage：Molasses 8-10%, fish peptone 2-5%, soya-bean cake
Powder 10-15%, surplus is water；
Fermented and cultured uses fed-batch process, and wherein feed supplement carbon source is：Glucose, sucrose；Nitrogen source is：Fish peptone,
Beancake powder.
Fermentation culture stage：In starting 0~24 hour, interval ventilation is maintained at aerobic conditions fermentation, throughput 1：1.8, stirring
Speed 150r-200/min, stirs interval time 3-5 hour, stirs 3-5 minutes, 25 DEG C -35 DEG C of temperature.
5. a kind of production method as claimed in claim 4 for being used to prevent and treat the complex microorganism preparations of rice seedling blight, it is special
Levy and be：
1) solid is inoculated in respectively under Trichoderma harzianum Q2, saccharomyces cerevisiae T1, bacillus amyloliquefaciens J12 original strain aseptic conditions
On culture medium, Trichoderma harzianum Q2 and saccharomyces cerevisiae T1 are cultivated 4 days under the conditions of 30 DEG C；；Bacillus amyloliquefaciens J12 is in 37 DEG C of bars
Cultivated 2 days under part；
2) first order seeds culture：By step 1) culture strain aseptic condition under be inoculated in fluid nutrient medium, Trichoderma harzianum respectively
Q2, saccharomyces cerevisiae T1 cultivated under the conditions of 30 DEG C 4 days, bacillus amyloliquefaciens J12 cultivated 2 days under the conditions of 37 DEG C, be made one
Level seed, terminates each saccharomyces cerevisiae of culture, bacillus amyloliquefaciens suspension optical density OD600 values and respectively reaches 4.0,3.0；
3) secondary seeds culture：The inoculum concentration for being 15% by the volume ratio of fluid nutrient medium, first order seed is inoculated into respectively
In 100L fermentation tank, under the conditions of the cumulative volume of nutrient solution is 70L, 30 DEG C -35 DEG C in fermentation tank, mixing speed is 200r/
Min, throughput is 1：2.0, Trichoderma harzianum Q2, saccharomyces cerevisiae T1 are cultivated 3 days, and bacillus amyloliquefaciens J12 is cultivated 1 day and is made two
Level seed；
4) mixing fermentation cultures：The inoculum concentration for being 15% by the volume ratio of fluid nutrient medium, 1 ton is inoculated into by secondary seed
In fermentation tank, the culture medium cumulative volume in fermentation tank is 700L, carries out fermented and cultured, obtains microbial inoculum.
Wherein, step 1), 2), 3) in used culture medium Trichoderma harzianum Q2, saccharomyces cerevisiae T1 be PDA culture medium, solve starch bud
Spore bacillus J12 is LB culture mediums；
Wherein, step 4) used in the formula of culture medium be by mass percentage：Molasses 10%, fish peptone 2%, beancake powder
15%, surplus is water；
Fermented and cultured uses fed-batch process, and wherein feed supplement carbon source is：Glucose；Nitrogen source is：Fish peptone.
Fermentation culture stage：In starting 0~24 hour, interval ventilation is maintained at aerobic conditions fermentation, throughput 1：2.0, stirring
Rotating speed 200r/min, stirs 3.5 hours interval times, stirs 4 minutes, 30 DEG C -35 DEG C of temperature.
6. a kind of being used for as described in any one of claims 1 to 3 prevents and treats the production of the complex microorganism preparations of rice seedling blight
Method is obvious to rice seedling blight prevention effect.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110896966A (en) * | 2019-12-19 | 2020-03-24 | 河南农贝得农业科技有限公司 | Microbial agent for crop take-all and preparation method thereof |
| CN116083251A (en) * | 2023-02-23 | 2023-05-09 | 天津坤禾生物科技集团股份有限公司 | Mixed microbial agent and preparation method and application thereof |
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