CN107034158B - Siam bacillus B-3 and its application - Google Patents
Siam bacillus B-3 and its application Download PDFInfo
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Abstract
The invention discloses Siam bacillus B-3 and its applications.Siam bacillus B-3 disclosed by the invention is CGMCC No.13753 in the deposit number of China Committee for Culture Collection of Microorganisms's common micro-organisms center.Experiments have shown that, the microbial inoculum that culture Siam bacillus B-3 is obtained can inhibit the growth for leading to the shot hole streptomycete (Streptomyces scabies) of potato scab, sour shot hole streptomycete (Streptomyces acidiscabies) and/or Streptomycesalbidoflhaving (Streptomyces albidoflavus), show that Siam bacillus B-3 of the invention and related microbial inoculum can be used to prevent and treat potato scab.
Description
Technical field
The present invention relates in field of biotechnology, Siam bacillus B-3 and its application.
Background technique
Potato scab is typical soil-borne disease, has generation in each potato producing region in China.It is mainly shown as horse
There is subcircular to the filbert scab of unsetting suberification scab shape or patch in bell potato wedge stem surface, and it is coarse that hand touches texture.Ma Ling
Potato shot hole generally has two kinds of disease symptoms of reticulate pattern scab and breach shape scab.Under normal conditions, reticulate pattern scab and breach
Shape scab is only limitted to cortex, and in flakes, not storage tolerance, and sick potato appearance is indecency, commodity grade greatly declines scab when falling ill serious,
The quality and economic value of potato wedge are largely reduced, and potato seed can accelerate the propagation of disease with allocation and transportation in spite of illness.To China
The production of potato constitute and seriously threaten, attract extensive attention.In recent years, with the increasing of onset area and occurring degree
Greatly, caused economic loss is also considerable.
Oneself has been carried out more in-depth study to the pathogen of potato scab to external researcher, clearly should
Disease is caused by a variety of streptomycetes.And constantly there is pathogenic kind of new scab to be reported in recent years.Prevention and treatment of the researchers to the disease
Method has also carried out many beneficial explorations.Though there are many kinds of method, but not ideal being applied to field control
Chemistry or biological agent can be good at preventing and treating potato scab.Currently, potato scab is mainly with Agro-chemicals control
Based on.However chemical agent is used for a long time, one side pathogen is easy to produce drug resistance, so that control efficiency is greatly reduced；Separately
On the one hand it is be easy to cause serious environmental pollution, jeopardizes human and livestock health.Therefore, effective there is an urgent need to be carried out to potato scab
Biological control.
Existing research shows that the pathogen of potato scab is caused to be mainly shot hole streptomycete, other micro- Bai Huanglian
Mould can also cause potato scab.Shot hole streptomyces is in conditioned pathogen, i.e., in soil environment neutrality or meta-alkalescence
Under the conditions of be easy growth and cause potato disease, and in Acid soil environment then seldom occur.
Summary of the invention
The technical problem to be solved by the present invention is to how prevent and treat the shot hole of streptomycete initiation.
In order to solve the above technical problems, present invention firstly provides one plant of Siam bacillus (Bacillus
Siamensis) bacterial strain, entitled Siam bacillus (Bacillus siamensis) B-3.
Siam bacillus (Bacillus siamensis) B-3 provided by the present invention, in Chinese microorganism strain
The deposit number of preservation administration committee common micro-organisms center is CGMCC No.13753.
In order to solve the above technical problems, the present invention also provides a kind of microbial inoculums.The microbial inoculum contains Siam's gemma bar
Bacterium (Bacillus siamensis) B-3.
The active constituent of the microbial inoculum can be Siam bacillus (Bacillus siamensis) B-3.
The microbial inoculum can be the microbial inoculum for inhibiting streptomycete.The streptomycete is the streptomycete for causing shot hole.It is described
The streptomycete for causing shot hole can be shot hole streptomycete (Streptomyces scabies), sour shot hole streptomycete
(Streptomyces acidiscabies) and/or Streptomycesalbidoflhaving (Streptomyces albidoflavus).It is described
Shot hole can be potato scab.
In one embodiment of the invention, the shot hole streptomycete (Streptomyces scabies) is shot hole
Streptomycete (Streptomyces scabies) CGMCC No.4.76 and shot hole streptomycete (Streptomyces scabies)
CGMCC No.4.1765。
In one embodiment of the invention, the sour scab streptomycete (Streptomyces acidiscabies) is
Sour scab streptomycete (Streptomyces acidiscabies) CGMCC No.4.1789.
In one embodiment of the invention, the Streptomycesalbidoflhaving (Streptomyces albidoflavus) is
Streptomycesalbidoflhaving (Streptomyces albidoflavus) CGMCC No.4.3598.
The preparation method of above-mentioned microbial inoculum can include: Siam bacillus (Bacillus siamensis) B-3 is cultivated,
Obtain the microbial inoculum.
The culture is cultivated using bacteria culture media.The bacteria culture media concretely beef broth culture medium.Institute
Stating beef broth culture medium can be the culture medium being prepared using peptone, beef extract, sodium chloride.In the beef broth culture medium
The concentration of peptone, beef extract and sodium chloride can be respectively 10.0g/L, 3.0g/L, 5.0g/L.
In above-mentioned microbial inoculum, the microbial inoculum can also include carrier.The carrier can be solid carrier or liquid-carrier.It is described
Solid carrier can be mineral material, vegetable material or high-molecular compound；The mineral material can for clay, talcum, kaolin,
At least one of montmorillonite, white carbon, zeolite, silica and diatomite；The vegetable material can be in corn flour, bean powder and starch
At least one；The high-molecular compound can be polyvinyl alcohol and/or polyglycols.The liquid-carrier can for organic solvent,
Vegetable oil, mineral oil or water；The organic solvent can be decane and/or dodecane.In the microbial inoculum, the active constituent can be with
It is deposited in the form of the fermentation liquid of the living cells, living cells that are cultured, the filtrate of cell culture or cell and the mixture of filtrate
In.The dosage form of the composition can be a variety of dosage forms, such as liquor, emulsion, suspending agent, pulvis, granule, wettable powder or water
Dispersible granule.
As needed, surfactant (such as polysorbas20, Tween 80), adhesive, stabilization can be also added in the microbial inoculum
Agent (such as antioxidant), pH adjusting agent.
In order to solve the above technical problems, the present invention also provides Siam bacillus (Bacillus siamensis) or institutes
State following any applications of microbial inoculum:
A1) inhibit the application in streptomycete product in preparation；
A2) inhibiting the application in streptomycete；
A3) application in disease product caused by streptomycete is prevented and treated in preparation；
A4) application in disease caused by streptomycete is being prevented and treated.
In order to solve the above technical problems, the present invention also provides contain Siam bacillus (Bacillus siamensis)
Or the product of the microbial inoculum, the function of the product are following b1) or b2):
B1) inhibit streptomycete；
B2 disease caused by streptomycete) is prevented and treated.
The active constituent of the product can be Siam bacillus (Bacillus siamensis) or the microbial inoculum.
In the present invention, Siam bacillus (Bacillus siamensis) can be Siam bacillus
(Bacillus siamensis)B-3。
The streptomycete can be following A 1) or A2) or A3):
A1) cause the streptomycete of shot hole；
A2) cause the streptomycete of potato scab；
A2) shot hole streptomycete (Streptomyces scabies), sour shot hole streptomycete (Streptomyces
) and/or Streptomycesalbidoflhaving (Streptomyces albidoflavus) acidiscabies.
Disease caused by the streptomycete can be following B1) or B2):
B1) shot hole；
B2) potato scab.
Inhibit to cause scab it is demonstrated experimentally that Siam bacillus (Bacillus siamensis) B-3 of the invention has
The function of the streptomycete of disease can specifically inhibit shot hole streptomycete, sour shot hole streptomycete and Streptomycesalbidoflhaving: Siam's bud
Spore bacillus (Bacillus siamensis) B-3 is about 18mm to the diameter of the inhibition zone of shot hole streptomycete 4.76, to scab
The diameter of the inhibition zone of sick streptomycete 4.1765 is about 22mm, and the diameter to the inhibition zone of sour scab streptomycete 4.1789 is about
17mm, the diameter to the inhibition zone of Streptomycesalbidoflhaving 4.3598 are about 11mm.In addition, Siam bacillus of the invention
(Bacillus siamensis) B-3 is that 4.0-9.0 can be grown in pH value, illustrates it in meta-alkali and meta-acid environment
Streptomycete for inhibiting to cause shot hole.Show that Siam bacillus of the invention (Bacillus siamensis) B-3 can
To cause disease caused by the streptomycete of shot hole for preventing and treating, and do not influenced by environmental pH.
Biomaterial preservation explanation
The classification naming of biomaterial: Siam bacillus (Bacillus siamensis)
The strain number of biomaterial: B-3
Depositary institution's title of biomaterial: China Committee for Culture Collection of Microorganisms's common micro-organisms center
Depositary institution's abbreviation of biomaterial: CGMCC
The depositary institution address of biomaterial: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Chinese Academy of Sciences microorganism
Research institute, postcode: 100101
The preservation date of biomaterial: on 03 14th, 2017
The collection of biomaterial is registered on the books number: CGMCC No.13753
Detailed description of the invention
Fig. 1 is fungistatic effect of Siam bacillus B-3 to shot hole streptomycete 4.76.
Fig. 2 is fungistatic effect of Siam bacillus B-3 to shot hole streptomycete 4.1765.
Fig. 3 is fungistatic effect of Siam bacillus B-3 to sour scab streptomycete 4.1789.
Fig. 4 is fungistatic effect of Siam bacillus B-3 to Streptomycesalbidoflhaving 4.3598.
Fig. 5 is Siam bacillus B-3 to shot hole streptomycete 4.76,4.1765 and sour shot hole streptomycete 4.1789
With the fungistatic effect of Streptomycesalbidoflhaving 4.3598.
Fig. 6 be when co-culturing Siam bacillus B-3 to shot hole streptomycete 4.76,4.1765 and sour shot hole chain
The fungistatic effect of mould 4.1789 and Streptomycesalbidoflhaving 4.3598.
Specific embodiment
The present invention is further described in detail With reference to embodiment, and the embodiment provided is only for explaining
The bright present invention, the range being not intended to be limiting of the invention.Experimental method in following embodiments is unless otherwise specified
Conventional method.Material as used in the following examples, reagent, instrument etc., are commercially available unless otherwise specified.
Quantitative test in following embodiment, is respectively provided with three repeated experiments, and results are averaged.
Beef broth fluid nutrient medium: being dissolved in 1L distilled water for peptone 10.0g, beef extract 3.0g and sodium chloride 5.0g, adjusts
PH value is saved to 7.0；Then 121 DEG C of sterilizing 20min.
Beef broth solid medium: peptone 10.0g, beef extract 3.0g, sodium chloride 5.0g and agar 15.0g are dissolved in 1L
Distilled water adjusts pH value to 7.0；Then 121 DEG C of sterilizing 20min.
Beef broth solid plate: pouring into culture dish for the beef broth solid medium that 20mL temperature is 55 DEG C or so, cooling
It is afterwards beef broth solid plate.
Oat solid plate: taking oat 20g, and distilled water 1L is added, boils 30min；Filtrate is collected after filtered through gauze, to institute
Addition agar powder 20g in filtrate is stated, and is settled to 1L with distilled water, adjusts pH value to 7.2-7.4；Then 121 DEG C of sterilizings
20min.The oat medium that 20mL temperature is 55 DEG C or so is poured into culture dish, is oat solid plate after cooling.
ISP-2 culture medium (g/L): yeast extract 4g, malt extract 10g, glucose 4g, PH7.3, then 115 DEG C
Sterilize 15min.
ISP-2 solid plate: the ISP-2 solid medium that 20mL temperature is 55 DEG C or so is poured into culture dish, after cooling
As ISP-2 solid plate.
Potato fluid nutrient medium the preparation method comprises the following steps: take peeled potatoes 200g, be cut into small pieces, 1.0L distillation be added
Water boils 30min；Filtrate is collected after filtered through gauze, glucose 20.0g is added in Xiang Suoshu filtrate, and be settled to distilled water
1.0L, pH value is naturally, then 115 DEG C of sterilizing 15min.
Potato fluid nutrient medium 1 the preparation method comprises the following steps: take peeled potatoes 200g, be cut into small pieces, 1.0L distillation be added
Water boils 30min；Filtrate is collected after filtered through gauze, glucose 20.0g is added in Xiang Suoshu filtrate, and be settled to distilled water
1.0L adjusts pH value to 4.0；Then 115 DEG C of sterilizing 15min.
Potato fluid nutrient medium 2: " the adjusting pH value to 4.0 " in the preparation method of potato fluid nutrient medium 1 is replaced
It is changed to " adjusting pH value to 5.0 ", other steps are all the same.
Potato fluid nutrient medium 3: " the adjusting pH value to 4.0 " in the preparation method of potato fluid nutrient medium 1 is replaced
It is changed to " adjusting pH value to 6.0 ", other steps are all the same.
Potato fluid nutrient medium 4: " the adjusting pH value to 4.0 " in the preparation method of potato fluid nutrient medium 1 is replaced
It is changed to " adjusting pH value to 7.0 ", other steps are all the same.
Potato fluid nutrient medium 5: " the adjusting pH value to 4.0 " in the preparation method of potato fluid nutrient medium 1 is replaced
It is changed to " adjusting pH value to 8.0 ", other steps are all the same.
Potato fluid nutrient medium 6: " the adjusting pH value to 4.0 " in the preparation method of potato fluid nutrient medium 1 is replaced
It is changed to " adjusting pH value to 9.0 ", other steps are all the same.
Shot hole streptomycete (Streptomyces scabies) CGMCC No.4.76, shot hole streptomycete
(Streptomyces scabies) CGMCC No.4.1765, sour scab streptomycete (Streptomyces acidiscabies)
CGMCC No.4.1789 and the equal preservation of Streptomycesalbidoflhaving (Streptomyces albidoflavus) CGMCC No.4.3598
In China General Microbiological culture presevation administrative center (abbreviation CGMCC, address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3
Institute of Microorganism, Academia Sinica's postcode: 100101), strain catalog number is followed successively by 4.76,4.1765,4.1789 and
4.3598, the public can obtain from China General Microbiological culture presevation administrative center.Shot hole streptomycete (Streptomyces
Scabies) CGMCC No.4.76 hereinafter abbreviation shot hole streptomycete 4.76.Shot hole streptomycete (Streptomyces
Scabies) CGMCC No.4.1765 hereinafter abbreviation shot hole streptomycete 4.1765.Sour shot hole streptomycete
(Streptomyces acidiscabies) CGMCC No.4.1789 hereinafter referred to as sour shot hole streptomycete 4.1789.It is micro-
White yellow streptomycete (Streptomyces albidoflavus) CGMCC No.4.3598 hereinafter abbreviation Streptomycesalbidoflhaving
4.3598。
Embodiment 1, the separation of Siam bacillus (Bacillus siamensis) B-3 CGMCC No.13753, identification
And preservation
One, it separates
1, the processing of pedotheque
(1) 10g pedotheque is added in 90mL sterile water (to pick up from Ningxia Hui Autonomous Region's Ping Jibao kind and plant potato
Soil) and appropriate bead, 10min is vibrated, stands 30s, soil stoste is made, and (dilution is denoted as 10 at this time-1).Draw 1mL soil
The supernatant of earth stoste, which is added in the test tube for filling 9mL sterile water, mixes well that (dilution is 10 at this time-2), then from this test tube
Middle absorption 1mL be added in another test tube for filling 9mL sterile water be uniformly mixed, and so on dilution first (dilution is made
It is 10-3) and dilution second (dilution 10-4)。
2, shot hole streptomycete 4.76 is taken, oat solid plate, 28 DEG C of stationary culture 10d are inoculated in；Then solid to oat
Appropriate amounts of sterilized water is added on body plate, the spore of scraping shot hole streptomycete 4.76 is shoveled with aseptic inoculation, obtains cause of disease bacteria suspension 1
(concentration of shot hole streptomycete 4.76 is about 1.0 × 108cfu/mL)。
3, after completing step 1 and step 2,50 μ L dilution second and 50 μ L cause of disease bacteria suspensions 1 are mixed, then even spread
On beef broth solid plate, 30 DEG C of stationary culture 2d.The bacterium colony that inhibition zone can be formed is enterprising in beef broth solid plate
Row purifying agaric.
Screen one plant of bacterium is named as bacterium B-3.
Two, it identifies
1, Morphological Identification
The bacterium B-3 that step 1 obtains is seeded on beef broth solid medium, 30 DEG C of cultures, observes single colonie after 2d
Form.
The result shows that the bacterium colony of bacterium B-3 is round, surface flat, edge micro-protuberance is white, colony diameter 1-
3mm。
2,16S rDNA sequence homology analysis
The 16S rDNA of bacterium B-3 is as shown in the sequence 1 in sequence table.
By comparing, the homology highest of bacterium B-3 and Siam bacillus (Bacillus siamensis) reach
100%.
Therefore, bacterium B-3 is accredited as Siam bacillus (Bacillus siamensis).
Three, preservation
The bacterium B-3 of step 1 separation was preserved in Chinese microorganism strain preservation conservator on 03 14th, 2017
Meeting common micro-organisms center (abbreviation CGMCC, address are as follows: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3), deposit number is
CGMCC No.13753.The full name of bacterium B-3 is Siam bacillus (Bacillus siamensis) B-3CGMCC
No.13753, referred to as Siam bacillus B-3.
Embodiment 2, the expansion of Siam bacillus B-3 are numerous
Siam bacillus B-3 of embodiment 1 expands that numerous specific step is as follows:
1, Siam bacillus B-3 is activated three times on beef broth solid medium, then picking single colonie is inoculated in
In 250mL conical flask equipped with 50mL beef broth fluid nutrient medium, 30 DEG C, cultivate 2d on the constant-temperature table of 200r/min, obtain
Culture solution.
2, after completing step 1, the culture solution of step 1 is taken, 12000r/min is centrifuged 1min, abandons supernatant, collects precipitating.
3, after completing step 2, the precipitating of step 2 is taken, is resuspended with 50mL sterile water, obtains bacteria suspension (Siam bacillus
The concentration of B-3 is about 1.0 × 1011cfu/mL)。
The fungistatic effect of embodiment 3, Siam bacillus B-3
Fungistatic effect of Siam bacillus B-3 to germ of embodiment 1 is detected, experiment in triplicate, is averaged.Often
Specific step is as follows for secondary experiment:
One, inhibitory effect of Siam bacillus B-3 to shot hole streptomycete 4.76
1, shot hole streptomycete 4.76 is taken, oat solid plate, 28 DEG C of stationary culture 10d are inoculated in.
2, after completing step 1, the oat solid plate is taken, appropriate amounts of sterilized water is added, is then shoveled and is scraped with aseptic inoculation
The spore of the oat solid planar surface, obtain cause of disease bacteria suspension 1 (concentration of shot hole streptomycete 4.76 is about 1.0 ×
108cfu/mL)。
3, after completing step 2,50 μ L cause of disease bacteria suspensions 1 is taken, ISP-2 solid plate is spread evenly across.
4, after completing step 3, the ISP-2 solid plate that pathogen suspension 1 is coated with obtained by step 3 is taken, first in its center
The aseptic filter paper that diameter is 5mm is placed, the bacterium that then point connects that step 3 obtains in 10 μ L embodiments 2 on the aseptic filter paper is hanged
Liquid.30 DEG C of stationary cultures after cultivating 2d, observe fungistatic effect.
Siam's bacillus B-3 microbial inoculum in step 4 is replaced with into sterile water, other steps are constant, as negative right
According to.
Siam bacillus B-3 is shown in Fig. 1 to the fungistatic effect of shot hole streptomycete 4.76.The result shows that Dian Jie Siam bud
After spore bacillus B-3 culture, there is the appearance of inhibition zone (diameter of inhibition zone is about 18mm)；And after point connects sterile Aquaponic, no suppression
The appearance of bacterium circle.Therefore, Siam bacillus B-3 has certain inhibitory effect to shot hole streptomycete 4.76.
Two, inhibitory effect of Siam bacillus B-3 to shot hole streptomycete 4.1765
According to the method for step 1, shot hole streptomycete 4.76 is replaced with into shot hole streptomycete 4.1765, other steps
It is constant, Siam bacillus B-3 is obtained to the inhibitory effect of shot hole streptomycete 4.1765.
Siam bacillus B-3 is shown in Fig. 2 to the fungistatic effect of shot hole streptomycete 4.1765.The result shows that Dian Jie Siam
After bacillus B-3 culture, there is the appearance of inhibition zone (diameter of inhibition zone is about 22mm)；And after point connects sterile Aquaponic, nothing
The appearance of inhibition zone.Therefore, Siam bacillus B-3 has certain inhibitory effect to shot hole streptomycete 4.1765.
Three, inhibitory effect of Siam bacillus B-3 to sour scab streptomycete 4.1789
According to the method for step 1, shot hole streptomycete 4.76 is replaced with into sour scab streptomycete 4.1789, other steps
It is constant, Siam bacillus B-3 is obtained to the inhibitory effect of sour scab streptomycete 4.1789.
Siam bacillus B-3 is shown in Fig. 3 to the fungistatic effect of sour scab streptomycete 4.1789.The result shows that Dian Jie Siam
After bacillus B-3 culture, there is the appearance of inhibition zone (diameter of inhibition zone is about 17mm)；And after point connects sterile Aquaponic, nothing
The appearance of inhibition zone.Therefore, Siam bacillus B-3 has certain inhibitory effect to sour scab streptomycete 4.1789.
Four, inhibitory effect of Siam bacillus B-3 to Streptomycesalbidoflhaving 4.3598
According to the method for step 1, shot hole streptomycete 4.76 is replaced with into Streptomycesalbidoflhaving 4.3598, other steps
It is constant, Siam bacillus B-3 is obtained to the inhibitory effect of Streptomycesalbidoflhaving 4.3598.
Siam bacillus B-3 is shown in Fig. 4 to the fungistatic effect of Streptomycesalbidoflhaving 4.3598.The result shows that Dian Jie Siam
After bacillus B-3 culture, there is the appearance of inhibition zone (diameter of inhibition zone is about 11mm)；And after point connects sterile Aquaponic, nothing
The appearance of inhibition zone.Therefore, Siam bacillus B-3 has certain inhibitory effect to Streptomycesalbidoflhaving 4.3598.
Five, Siam bacillus B-3 is to shot hole streptomycete 4.76,4.1765, sour shot hole streptomycete 4.1789 and micro-
The mixing inhibitory effect of white yellow streptomycete 4.3598
1, shot hole streptomycete 4.76 is taken, oat solid plate, 28 DEG C of stationary culture 10d are inoculated in；Then to the swallow
Appropriate amounts of sterilized water is added on wheat solid plate, the spore of scraping shot hole streptomycete 4.76 is shoveled with aseptic inoculation, obtains pathogen
(concentration of shot hole streptomycete 4.76 is about 1.0 × 10 to suspension 18cfu/mL)。
2, shot hole streptomycete 4.1765 is taken, oat solid plate, 28 DEG C of stationary culture 10d are inoculated in；Then to described
Appropriate amounts of sterilized water is added on oat solid plate, the spore of scraping shot hole streptomycete 4.1765 is shoveled with aseptic inoculation, obtains disease
(concentration of shot hole streptomycete 4.1765 is about 1.0 × 10 to opportunistic pathogen suspension 28cfu/mL)。
3, sour scab streptomycete 4.1789 is taken, oat solid plate, 28 DEG C of stationary culture 10d are inoculated in；Then to described
Appropriate amounts of sterilized water is added on oat solid plate, the spore for scraping sour scab streptomycete 4.3598 is shoveled with aseptic inoculation, obtains disease
(concentration of sour scab streptomycete 4.1789 is about 1.0 × 10 to opportunistic pathogen suspension 38cfu/mL)。
4, Streptomycesalbidoflhaving 4.3598 is taken, oat solid plate, 28 DEG C of stationary culture 10d are inoculated in；Then to described
Appropriate amounts of sterilized water is added on oat solid plate, the spore of scraping Streptomycesalbidoflhaving 4.3598 is shoveled with aseptic inoculation, obtains disease
(concentration of Streptomycesalbidoflhaving 4.3598 is about 1.0 × 10 to opportunistic pathogen suspension 48cfu/mL)。
5, after completing step 1, step 2, step 3 and step 4, by cause of disease bacteria suspension 1, cause of disease bacteria suspension 2, cause of disease bacteria suspension
3 and cause of disease bacteria suspension 4 in equal volume mix, obtain plastc ring first.
6, after completing step 5,150 μ L plastc ring first is taken, ISP-2 solid plate is spread evenly across.
7, after completing step 6, the ISP-2 solid plate that plastc ring first is coated with obtained by step 6 is taken, first in its center
The aseptic filter paper that diameter is 5mm is placed, the bacterium that then point connects that step 3 obtains in 10 μ L embodiments 2 on the aseptic filter paper is hanged
Liquid.30 DEG C of stationary cultures after cultivating 2d, observe fungistatic effect.
" bacteria suspension that step 3 obtains in embodiment 2 " in step 7 is replaced with into sterile water, other steps are constant, make
For negative control.
Siam bacillus B-3 is to shot hole streptomycete 4.76,4.1765 and sour shot hole streptomycete 4.1789 and micro-
The fungistatic effect of white yellow streptomycete 4.3598 is shown in Fig. 5.The result shows that having inhibition zone after Dian Jie Siam bacillus B-3 culture
Occur (diameter of inhibition zone is about 11mm)；And after point connects sterile Aquaponic, the appearance of no inhibition zone.Therefore, Siam's gemma bar
Bacterium B-3 is to shot hole streptomycete 4.76,4.1765 and sour shot hole streptomycete 4.1789 and 4.3598 groups of Streptomycesalbidoflhaving
At Mixed Microbes have certain inhibitory effect.
Five, fungistatic effect is co-cultured
1, in step 41.
2, in step 42.
3, in step 43.
4, in step 44.
5, after completing step 1, step 2, step 3 and step 4, by 50 μ L cause of disease bacteria suspensions 1,50 μ L cause of disease bacteria suspensions 2,50
The bacteria suspension mixing that step 3 obtains in μ L cause of disease bacteria suspension 3,50 μ L cause of disease bacteria suspensions 4 and 10 μ L embodiments 2, obtains Mixed Microbes
Suspension second.
6, after completing step 5, plastc ring second is taken, ISP-2 solid plate is spread evenly across.30 DEG C of stationary cultures, training
After supporting 2d, fungistatic effect is observed.
It co-cultures fungistatic effect and sees Fig. 6.The result shows that during co-cultivation, only Siam bacillus B-3 growth,
And what shot hole streptomycete 4.76,4.1765, sour shot hole streptomycete 4.1789 and Streptomycesalbidoflhaving 4.3598 were not grown
Sign.Therefore, Siam bacillus B-3 is to shot hole streptomycete 4.76,4.1765, sour shot hole streptomycete 4.1789 and micro- white
Yellow streptomycete 4.3598 has inhibitory effect.
The pH value growth scope of embodiment 4, Siam bacillus B-3
1, Siam bacillus B-3 of embodiment 1 is activated three times on beef broth solid medium, then picking single bacterium
It falls and is inoculated in 5mL potato fluid nutrient medium, 30 DEG C, cultivate 1d on the constant-temperature table of 200r/min, obtain initial bacterium solution.It will
Initial bacterium solution inoculation (inoculum concentration is 2% (percent by volume)) is extremely equipped with the 250mL conical flask of 50mL potato fluid nutrient medium 1
In, 30 DEG C, cultivate 2d on the constant-temperature table of 200r/min, obtain culture solution.
2, after completing step 1, the culture solution is taken, its light absorption value at 600nm wavelength is detected.
The result shows that light absorption value of the culture solution at 600nm wavelength is 3.75.As it can be seen that Siam bacillus B-3 exists
Growing state in potato fluid nutrient medium 1 is good.
According to above-mentioned steps, potato fluid nutrient medium 1 is replaced with into potato fluid nutrient medium 2, potato liquid respectively
Body culture medium 3, potato fluid nutrient medium 4, potato fluid nutrient medium 5 and potato fluid nutrient medium 6, obtained culture solution
Light absorption value at 600nm wavelength is followed successively by 4.21,4.17,4.16,4.10 and 3.96.As it can be seen that Siam bacillus B-3 is in horse
Bell potato fluid nutrient medium 1, potato fluid nutrient medium 2, potato fluid nutrient medium 3, potato fluid nutrient medium 4, potato
Growing state in fluid nutrient medium 5 and potato fluid nutrient medium 6 is good and almost the same.
The above results show that the pH value growth scope of Siam bacillus B-3 is 4.0-9.0.
<110>Institute of Microorganism, Academia Sinica
<120>Siam bacillus B-3 and its application
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 1340
<212> DNA
<213>Siam bacillus (Bacillus siamensis)
<400> 1
acttcgggtg ttacaaactc tcgtggtgtg acgggcggtg tgtacaaggc ccgggaacgt 60
attcaccgcg gcatgctgat ccgcgattac tagcgattcc agcttcacgc agtcgagttg 120
cagactgcga tccgaactga gaacagattt gtgggattgg cttaacctcg cggtttcgct 180
gccctttgtt ctgtccattg tagcacgtgt gtagcccagg tcataagggg catgatgatt 240
tgacgtcatc cccaccttcc tccggtttgt caccggcagt caccttagag tgcccaactg 300
aatgctggca actaagatca agggttgcgc tcgttgcggg acttaaccca acatctcacg 360
acacgagctg acgacaacca tgcaccacct gtcactctgc ccccgaaggg gacgtcctat 420
ctctaggatt gtcagaggat gtcaagacct ggtaaggttc ttcgcgttgc ttcgaattaa 480
accacatgct ccaccgcttg tgcgggcccc cgtcaattcc tttgagtttc agtcttgcga 540
ccgtactccc caggcggagt gcttaatgcg ttagctgcag cactaagggg cggaaacccc 600
ctaacactta gcactcatcg tttacggcgt ggactaccag ggtatctaat cctgttcgct 660
ccccacgctt tcgctcctca gcgtcagtta cagaccagag agtcgccttc gccactggtg 720
ttcctccaca tctctacgca tttcaccgct acacgtggaa ttccactctc ctcttctgca 780
ctcaagttcc ccagtttcca atgaccctcc ccggttgagc cgggggcttt cacatcagac 840
ttaagaaacc gcctgcgagc cctttacgcc caataattcc ggacaacgct tgccacctac 900
gtattaccgc ggctgctggc acgtagttag ccgtggcttt ctggttaggt accgtcaagg 960
tgccgcccta tttgaacggc acttgttctt ccctaacaac agagctttac gatccgaaaa 1020
ccttcatcac tcacgcggcg ttgctccgtc agactttcgt ccattgcgga agattcccta 1080
ctgctgcctc ccgtaggagt ctgggccgtg tctcagtccc agtgtggccg atcaccctct 1140
caggtcggct acgcatcgtc gccttggtga gccgttacct caccaactag ctaatgcgcc 1200
gcgggtccat ctgtaagtgg tagccgaagc caccttttat gtctgaacca tgcggttcaa 1260
acaaccatcc ggtattagcc ccggtttccc ggagttatcc cagtcttaca ggcaggttac 1320
ccacgtgtta ctcacccgtc 1340
Claims (4)
1. Siam bacillus (Bacillus siamensis) B-3, general in China Committee for Culture Collection of Microorganisms
The deposit number at logical microorganism center is CGMCC No.13753.
2. a kind of microbial inoculum, it is characterised in that: the microbial inoculum contains Siam bacillus (Bacillus described in claim 1
Siamensis) B-3.
3. Siam bacillus (Bacillus siamensis) B-3 described in claim 1 or microbial inoculum as claimed in claim 2
Following any applications:
A1) inhibit the application in streptomycete product in preparation；
A2) inhibiting the application in streptomycete；
A3) application in disease product caused by streptomycete is prevented and treated in preparation；
A4) application in disease caused by streptomycete is being prevented and treated;
The streptomycete is shot hole streptomycete (Streptomyces scabies), sour shot hole streptomycete
(Streptomyces acidiscabies) and/or Streptomycesalbidoflhaving (Streptomyces albidoflavus)；
The shot hole streptomycete (Streptomyces scabies) is shot hole streptomycete (Streptomyces
Scabies) CGMCC No.4.76 or shot hole streptomycete (Streptomyces scabies) CGMCC No.4.1765；
The acid scab streptomycete (Streptomyces acidiscabies) is sour scab streptomycete (Streptomyces
Acidiscabies) CGMCC No.4.1789；
The Streptomycesalbidoflhaving (Streptomyces albidoflavus) is Streptomycesalbidoflhaving (Streptomyces
Albidoflavus) CGMCC No. 4.3598；Disease caused by the streptomycete is potato scab.
4. containing bacterium described in Siam bacillus described in claim 1 (Bacillus siamensis) B-3 or claim 2
The product of agent；The function of the product is following b1) or b2):
B1) inhibit streptomycete；
B2 disease caused by streptomycete) is prevented and treated；
The streptomycete is shot hole streptomycete (Streptomyces scabies), sour shot hole streptomycete
(Streptomyces acidiscabies) and/or Streptomycesalbidoflhaving (Streptomyces albidoflavus)；
The shot hole streptomycete (Streptomyces scabies) is shot hole streptomycete (Streptomyces
Scabies) CGMCC No.4.76 or shot hole streptomycete (Streptomyces scabies) CGMCC No.4.1765；
The acid scab streptomycete (Streptomyces acidiscabies) is sour scab streptomycete (Streptomyces
Acidiscabies) CGMCC No.4.1789；
The Streptomycesalbidoflhaving (Streptomyces albidoflavus) is Streptomycesalbidoflhaving (Streptomyces
Albidoflavus) CGMCC No. 4.3598；
Disease caused by the streptomycete is potato scab.
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| CN107384839B (en) * | 2017-09-05 | 2020-06-30 | 农业部沼气科学研究所 | Bacillus siamensis BERC-11 and application thereof |
| CN108060100B (en) * | 2017-12-29 | 2021-01-29 | 中国科学院南海海洋研究所 | Multifunctional Siamese bacillus and preparation and application of bioactive substance thereof |
| CN108034618B (en) * | 2018-01-24 | 2021-01-01 | 吉林省农业科学院 | Siam bacillus strain and application thereof |
| CN110117554B (en) * | 2018-02-05 | 2023-03-07 | 呼伦贝尔恒屹农牧业股份有限公司 | Microbial agent, preparation method and application thereof |
| CN108849986B (en) * | 2018-07-09 | 2020-09-15 | 江苏师范大学 | Application of bacillus simmer in preventing and treating white rot |
| CN108753630B (en) * | 2018-07-28 | 2020-10-09 | 福建三炬生物科技股份有限公司 | Paecilomyces gossypii strain Sanju-18 and application thereof |
| CN109161500B (en) * | 2018-09-06 | 2022-01-07 | 中国农业科学院饲料研究所 | Siamese bacillus ZJ-2018-1, mycotoxin degrading microbial inoculum and application |
| CN109207404B (en) * | 2018-10-23 | 2022-02-18 | 山西农业大学 | Siam bacillus YJ15 and application thereof |
| CN109362787A (en) * | 2018-12-07 | 2019-02-22 | 江苏师范大学 | Application of Siam bacillus in prevention and treatment nematode |
| CN111944714B (en) * | 2020-07-27 | 2021-04-27 | 天津农学院 | Siamese bacillus HZY-54 growing in anthurium and application thereof |
| CN111718881B (en) * | 2020-08-13 | 2022-05-27 | 石河子大学 | Siamese bacillus and application thereof |
| KR102525964B1 (en) * | 2021-06-14 | 2023-04-28 | 대한민국 | Bacterial wilt, bacterial canker, and clavibacter michiganensis subsp. michiganensis inhibition micoorganism bacillus siamensis k203 strain and use thereof |
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