CN105505834B - Siam bacillus and its application of one plant of prevention and treatment black rot of pear and soft rot - Google Patents
Siam bacillus and its application of one plant of prevention and treatment black rot of pear and soft rot Download PDFInfo
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Abstract
The invention discloses Siam bacillus and its applications that one plant is prevented and treated black rot of pear and soft rot.The present invention separates from the Laiyang pear pericarp of surface sterilizing and obtains one plant of Siam bacillus (Bacillus siamensis), the bacterial strain has resistance of wide spectrum to various plants pathogen, rots have good control efficiency to the pear fruit as caused by ring spot and soft rot.The bacterium source is in pear fruit, to pear fruit safety；It is low that cost of material is mass produced, easy to operate, control efficiency is stablized, and can effectively solve the residual of agricultural brought by pear fruit disease pesticide control and problem of environmental pollution.There is huge application potential in terms of the biological control of pears postharvest disease, provide excellent basic bacterial strain to develop and developing microbial pesticide.
Description
Technical field
The invention belongs to microorganism fields, and in particular to one plant prevention and treatment black rot of pear and soft rot Siam bacillus and
It is applied.
Background technique
Pears are that China's cultivation history is long, area is big, one of the fruit tree that yield is high.Pears are the big fruit in China the 3rd, mesh according to statistics
Preceding national total area more than 800 ten thousand tons of annual output, accounts for the 62% of whole world pear cultivation area and yield up to more than 100 ten thousand hectares respectively
With 52%.Pear fruit is in growth and development and storage period, and the moment will receive the threat of disease, wherein ring spot
(Botryosphaeria dothidea) is one of fruit fungal disease of most serious, is the main original for causing pear fruit decayed fruit
Cause.Since Target spot pathogen is fallen ill during fruit maturation and storing in young fruit period latent infection, controls and taken precautions against and is very big
It is difficult.Pears soft rot is mainly by rhizopus stolonifer (Rhizopus stolonifer) and pyriform Mucor (Mucor piriformis)
Cause, main harm pear fruit, it is general fragmentary to occur, but when fruit wound is more, storage period temperature is high occur it is heavier.High temperature
When, disease fruit all soft corruption can be made in 5-6 days.
Fresh fruit postharvest decay is a global problem.China is every year because black rot of pear harm causes the operatic circle to subtract
About 25% is produced, decayed fruit rate is up to 80% or more when serious, causes huge economic losses.It is anti-frequently with chemical agent in production at present
Pears postharvest disease is controlled, so not only serious pollution of ecological environment, but also increases the residual quantity of toxic chemical substance in agricultural product, it is right
Human health brings potential hazard, while also easily leading to the generation of pathogen drug resistance.Biological control utilizes the antagonism between species
It acts on to reduce the generation of disease, to safety of human and livestock, environment compatibility is good, pest and disease damage is not likely to produce drug resistance, thus has and change
The advantage that fungicide does not have is learned, the best mode instead of chemical bactericide is just gradually being become.
Obtain the basis that efficient Antagonistic Fungi is biological control.Bacillus subtillis (Bacillus subtilis) is earliest
It is reported for the bacterium (Gutter and Littauer, 1953) for preventing and treating citrus disease.With going deep into for research, Ren Menyi
It screens and reports that a variety of pairs of postharvest disease of fruits and vegetables have the Antagonistic Fungi of control efficiency, mainly saccharomycete, bacillus, false unit cell
Bacterium and Trichoderma.Several plants of microorganism Antagonistic Fungis have been authorized to patent and business application, such as Biosave (Pseudomonas
Syringae, USA), " Shemer " (Metschnikowia Fructicola, Israel), Serenade (Bacillus
Subtilis, USA) and Candifruit (Candida sake, Spain).Due to can produce with the wide of high antagonistic activity
Antibacterial material, extremely strong anti-adversity ability are composed, while there is nontoxic, noresidue, no pathogenicity, to safety of human and livestock, Environmental compatibility
It is good, requirement of the consumer to agricultural product quality and safety is met, bud pole bacterium (Bacillus) plays in biological control
Particularly important effect.
Summary of the invention
It is an object of the present invention to provide one plant of Siam bacillus (Bacillus siamensis) LYLB4.
The deposit number of Siam bacillus (Bacillus siamensis) LYLB4 provided by the invention is CGMCC
No.11859。
The classification naming of above-mentioned bacterial strains LYLB4 be Siam bacillus (Bacillus siamensis), the bacterial strain in
On December 10th, 2015 is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center (abbreviation CGMCC, ground
Location: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Institute of Microorganism, Academia Sinica, postcode 100101), deposit number is
CGMCC No.11859。
It is a further object to provide Siam bacillus or its bacteria suspensions or its culture solution or its tunning
Or the new application of the microbial inoculum containing it.
The present invention provides Siam bacillus or its bacteria suspension or its culture solution or its tunning or contain its bacterium
Application of the agent in prevention and treatment ring spot and/or soft rot.
The present invention also provides Siam bacillus or its bacteria suspension or its culture solution or its tunning or containing its
Application of the microbial inoculum in the product of preparation prevention and treatment ring spot and/or soft rot.
In above-mentioned application, the ring spot and/or soft rot are black rot of pear and/or pears soft rot.
The present invention also provides Siam bacillus or its bacteria suspension or its culture solution or its tunning or containing its
Microbial inoculum antibacterial and/or it is antibacterial in application.
The present invention also provides Siam bacillus or its bacteria suspension or its culture solution or its tunning or containing its
Application of the microbial inoculum in preparation antibacterial and/or antibacterial product.
In above-mentioned application, the antibacterial and/or it is antibacterial be by inhibit mycelia growth embody.
In above-mentioned application, the bacterium is Target spot pathogen (Botryosphaeria dothidea) and/or Bacteria erwinia
(Rhizopus stolonifer) and/or dendrobium nobile alternaria (Alternaria tenuissima) and/or cotton verticillium wilt
Bacterium (Verticillium dahliae) and/or Alternaria brassicae (Alternaria brassicae) and/or ash arrhizus bacteria
(Botrytis cinerea) and/or capsicum rhizoctonia solani (Rizoctonia solani) and/or banana blight bacteria
(Fusarium oxysporum) and/or Pyricularia oryzae (Piricularia Oryzae) and/or Fusarium graminearum (Fusarium
graminearum)。
It is a still further object of the present invention to provide a kind of products.
The active constituent of product provided by the invention is above-mentioned Siam bacillus (Bacillus siamensis)
LYLB4CGMCC No.11859 or its bacteria suspension or its culture solution or its tunning or the microbial inoculum containing it；
The product have the function of it is following 1) or 2) described in:
1) ring spot and/or soft rot are prevented and treated；
2) antibacterial and/or antibacterial.
In the said goods, the antibacterial and/or it is antibacterial be by inhibit mycelia growth embody；
The bacterium is Target spot pathogen (Botryosphaeria dothidea) and/or Bacteria erwinia (Rhizopus
) and/or dendrobium nobile alternaria (Alternaria tenuissima) and/or verticillium dahliae stolonifer
(Verticillium dahliae) and/or Alternaria brassicae (Alternaria brassicae) and/or ash arrhizus bacteria
(Botrytis cinerea) and/or capsicum rhizoctonia solani (Rizoctonia solani) and/or banana blight bacteria
(Fusarium oxysporum) and/or Pyricularia oryzae (Piricularia Oryzae) and/or Fusarium graminearum (Fusarium
graminearum)。
Final object of the present invention is to provide a kind of prevention and treatment or assisting in preventing and treating pear fruit ring spot and/or soft rot
Method.
The method of prevention and treatment provided by the invention or assisting in preventing and treating pear fruit ring spot and/or soft rot includes the following steps:
Pear fruit is handled with the bacteria suspension in above-mentioned Siam bacillus or the said goods.
In the above method, Siam bacillus concentration is 5 × 10 in the bacteria suspension7CFU/ml。
The present invention separates from the Laiyang pear pericarp of surface sterilizing and obtains one plant of Siam bacillus (Bacillus
Siamensis), which has resistance of wide spectrum to various plants pathogen, to the pear fruit as caused by ring spot and soft rot
Rot that there is good control efficiency.The bacterium source is in pear fruit, to pear fruit safety；It is low that cost of material is mass produced,
Easy to operate, control efficiency is stablized, and can effectively solve the residual of agricultural brought by pear fruit disease pesticide control and environmental pollution
Problem.There is huge application potential in terms of the biological control of pears postharvest disease, provided to develop and developing microbial pesticide
Excellent basic bacterial strain.
Detailed description of the invention
Fig. 1 is the form photo of bacterial strain LYLB4.
Fig. 2 is the phylogenetic tree of the 16S rDNA building based on bacterial strain LYLB4.
Fig. 3 is the photo of bacterial strain LYLB4 and Target spot pathogen and Bacteria erwinia opposite culture.Wherein Fig. 3 A is bacterial strain LYLB4
With 7 days photos of Target spot pathogen opposite culture；Fig. 3 B is bacterial strain LYLB4 and 10 days photos of Bacteria erwinia opposite culture.
Fig. 4 is that bacterial strain LYLB4 acts on lower Target spot pathogen hypha form and Bacteria erwinia hypha form.Fig. 4 A is at LYLB4
The Target spot pathogen hypha form of reason group；Fig. 4 B is the Target spot pathogen hypha form of control group；Fig. 4 C is the soft corruption of LYLB4 processing group
Germ hypha form；Fig. 4 D is the Bacteria erwinia hypha form of control group.
Fig. 5 is preventive and therapeutic effect of the bacterial strain LYLB4 to pear fruit ring spot.
Fig. 6 is preventive and therapeutic effect of the bacterial strain LYLB4 to pear fruit soft rot.
Specific embodiment
Experimental method used in following embodiments is conventional method unless otherwise specified.
The materials, reagents and the like used in the following examples is commercially available unless otherwise specified.
Pear varieties in following embodiments are " Shuijing Pear "
Potato dextrose agar (PDA) in following embodiments: 200 grams of potato, 20 grams of glucose, agar
15 grams, it is settled to 1 liter with distilled water, natural pH.
Potato dextrose medium (PDB) in following embodiments: 200 grams of potato, 20 grams of glucose use distilled water
It is settled to 1 liter, natural pH.
Beef extract culture medium in following embodiments: beef extract 5.0g, peptone 10.0g, NaCl 5g, it is fixed with distilled water
Hold to 1 liter, pH 7.2~7.4.
The preparation of LYLB4 bacterium solution in following embodiments: LYLB4 is inoculated in beef extract culture medium, cultivates 48hr (30
DEG C, 1500r/min) after, by bacterium solution in 4 DEG C, 10000r/min be centrifuged 10min, remove supernatant, then be adjusted to need with sterile water
Concentration.
The separation and identification of embodiment 1, Siam bacillus plant
One, the separation and purifying of bacterial strain
1, by pick up from the red Xiao's pears in Pekinese, the Laiyang pear in Shandong, the Chinese pear in Hebei, Anhui Dangshan pear fruit tap water
After rinsing well, with distilled water flushing, lower pericarp is cut after drying, and pericarp is subjected to surface sterilization.Sterilization method is as follows: 70% wine
Then precision processing 40s, aseptic water washing handle 2min, aseptic water washing 3-4 times with 2.5% liquor natrii hypochloritis.
2, it aseptically, takes 100 μ L last times to rinse generated sterile water to be coated on beef extract culture medium,
30 DEG C of culture 24-72h observe whether there is or not bacterium colony generation, verify whether this sterilization method kills the micro- of material to be tested surface according to this
Biology.
3, pericarp under aseptic conditions, is cut into 0.2cm2Segment, be placed in the plate for filling beef extract culture medium, 28
It is cultivated under the conditions of DEG C.After bacterium colony occurs in tissue block, picking colony is transferred scribing line purifying culture in new beef extract culture medium, repeatedly
Bacteria purification bacterial strain is obtained several times, is inoculated in beef extract medium slant test tube, is stored in 4 DEG C of refrigerators.
It obtains 32 plants of bacteriums altogether in the pericarp of different pear fruits, carries out antagonistic bacterium by indicator bacteria of black rot of pear bacterium
Primary dcreening operation and secondary screening, wherein one plant of 1 plant of bacterium in Laiyang pear pericarp shows strong antagonistic activity, by the Strain Designation
For LYLB4.
Two, the identification of bacterial strain LYLB4
1, the Morphological Identification of bacterial strain LYLB4
The morphological feature of bacterial strain LYLB4: gram-positive bacteria, bacterium colony is round, milky, neat in edge, and surface wettability is glutinous
Thick, bacterium colony is gradual drying two days later for growth, and fold (Fig. 1) occurs in surface.
2, the identification of the Physiology and biochemistry of bacterial strain LYLB4
The physio-biochemical characteristics of bacterial strain LYLB4 are as shown in table 1, and bacterial strain LYLB4 belongs to aerobic bacteria, and methyl red experiment is negative,
Contact enzyme reaction, gelatin hydrolysis, Starch Hydrolysis, VP experiment, nitrate reduction experiment are the positive, can utilize grape well
Sugar and sucrose, it is weaker to the utilization of mannitol, it is grown under the conditions of 28~37 DEG C best.According to " common bacteria system identification hand
Volume ", bacterial strain LYLB4 is initially identified as bacillus (Bacillus) bacterium.
The physio-biochemical characteristics of table 1, bacterial strain LYLB4
*+-: growth is bad*+: growth
3, the Molecular Identification of bacterial strain LYLB4
The total DNA of bacterial strain LYLB4 is extracted as template, PCR amplification is carried out using universal primer 27f and 1492r, is contained
There is the segment of the conserved region bacterial strain LYLB416S rDNA.Primer sequence is as follows:
27f (upstream primer): 5 '-AGAGTTTGATCCTGGCTCAG-3 '；
1492r (downstream primer): 5 '-TACGGTTACCTTGTTACGACTT-3 '.
The sequence of the 16s rDNA for the bacterial strain LYLB4 that PCR amplification obtains is as shown in sequence 1.Based on 16S rDNA sequence
Phylogenetic tree is as shown in Figure 2.
4, the preservation of bacterial strain
According to morphological feature, Physiology and biochemistry and Molecular Identification result and analysis, the classification naming of bacterial strain LYLB4 is Siam's bud
Born of the same parents bacillus (Bacillus siamensis), the bacterial strain are preserved in Chinese microorganism strain preservation management on December 10th, 2015
(abbreviation CGMCC, address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, the Chinese Academy of Sciences are micro- for committee's common micro-organisms center
Biological study institute, postcode 100101), deposit number is CGMCC No.11859.
The application of embodiment 2, bacterial strain LYLB4 in antibacterial or antibacterial
One, antagonistic activity of the bacterial strain LYLB4 to black rot of pear bacterium and Bacteria erwinia
Activate pathogenic strains black rot of pear bacterium (Botryosphaeria dothidea) and soft respectively in PDA culture medium
Maize ear rot bacterium (Rhizopus stolonifer) is beatened to take bacteria cake with the punch of 6mm, respectively obtains black rot of pear bacteria cake and soft rot
Bacteria cake.Using opposite culture method, black rot of pear bacteria cake and soft rot bacteria cake are seeded in the PDA plate side of diameter 9cm respectively,
Opposite side streak inoculation bacterial strain LYLB4, two vaccinations are at a distance of 3-4cm, and to be individually seeded in the pathogen on PDA plate
Strain black rot of pear bacterium and Bacteria erwinia compare.3 repetitions are all provided with, are cultivated under the conditions of 25 DEG C, when full ware is covered in control, are surveyed
Amount processing colony diameter and antibacterial bandwidth, bacteriostatic activity is using inhibiting rate=[(control colony radius-processing bacterium radius)/right
According to colony radius] × 100%.
As a result as shown in figure 3, bacterial strain LYLB4 shows strong bacteriostasis to Target spot pathogen, bacteriostasis rate reaches 78%
(Fig. 3 A) above.Bacteria erwinia growth rapidly, can expire ware for one day, there is apparent antibacterial band (0.5- around bacterial strain LYLB4
1cm), after ten days still clear (Fig. 3 B).Illustrate that bacterial strain LYLB4 has two kinds of pears pathogen black rot of pear bacterium and Bacteria erwinia
Apparent inhibiting effect.
Two, the antimicrobial spectrum of bacterial strain LYLB4
With dendrobium nobile alternaria (Alternaria tenuissima), verticillium dahliae (Verticillium
Dahliae), Alternaria brassicae (Alternaria brassicae), ash arrhizus bacteria (Botrytis cinerea), capsicum are vertical
Blight bacterium (Rizoctonia solani), banana blight bacteria (Fusarium oxysporum), Pyricularia oryzae
(Piricularia Oryzae) and Fusarium graminearum (Fusarium graminearum) are used as test plant pathogen, research
The antimicrobial spectrum of bacterial strain LYLB4.Specific step is as follows:
Activate dendrobium nobile alternaria (Alternaria tenuissima), verticillium dahliae respectively in PDA culture medium
(Verticillium dahliae), Alternaria brassicae (Alternaria brassicae), ash arrhizus bacteria (Botrytis
Cinerea), capsicum rhizoctonia solani (Rizoctonia solani), banana blight bacteria (Fusarium oxysporum), rice
Seasonal febrile diseases bacterium (Piricularia Oryzae) and Fusarium graminearum (Fusarium graminearum), and with the punch of 6mm
It is beatened to take bacteria cake, respectively obtains above-mentioned each germ bacteria cake.Using opposite culture method, respectively by above-mentioned each germ pure culture biscuits involvng inoculation in diameter
The PDA plate side of 9cm, opposite side streak inoculation bacterial strain LYLB4, two vaccinations are individually connect at a distance of 3-4cm with pathogen
Kind compares on PDA plate.3 repetitions are all provided with, are cultivated under the conditions of 25 DEG C, when full ware is covered in control, measure each cause of disease
Colony diameter and antibacterial bandwidth, bacteriostatic activity is using inhibiting rate=[(control colony radius-processing bacterium radius)/control bacterium colony
Radius] × 100%.
The results are shown in Table 2, and bacterial strain LYLB4 also shows strong inhibitory effect to 8 kinds of test plant disease fungus, suppression
Bacterium rate is 50% or more.Illustrate that bacterial strain LYLB4 has the bacteriostasis of wide spectrum, can be used for the biological control of multiple diseases.
Table 2, LYLB4 are to the suppression result of different phytopathogens
Three, the influence that bacterial strain LYLB4 grows black rot of pear bacterium and Bacteria erwinia mycelia
Pathogenic strains activation black rot of pear bacterium and Bacteria erwinia are distinguished in PDA culture medium, are beaten with the punch of 6mm and are taken bacterium
Cake respectively obtains black rot of pear bacteria cake and soft rot bacteria cake.After black rot of pear bacteria cake and soft rot bacteria cake are ground, it is inoculated with respectively
In 1/10PDB culture medium, 25 DEG C, after 150rpm/min is cultivated 2 days, it is 1 × 10 that concentration, which is added,7The LYLB4 bacterium of CFU/ml is outstanding
Liquid, and compareed with the black rot of pear bacterium being individually seeded in 1/10PDB culture medium and Bacteria erwinia.It is all provided with 3 weights
It is multiple, microscopy mycelia upgrowth situation after being cultivated under the conditions of 25 DEG C.
As a result as shown in figure 4, Severe distortion occurs for Target spot pathogen hypha form under bacterial strain LYLB4 effect, mycelia is expanded
At beading, mycelia deformity, thickness are uneven, and some grows the top deformity of mycelia, rupture after forming expansion bubble, intracellular matter
Leak out (Fig. 4 A)；And compare mycelia even thickness and surface it is smooth, grow vigorous (Fig. 4 B).Illustrate that bacterial strain LYLB4 is serious
Hypha form is destroyed, has the function of inhibiting the growth of Target spot pathogen mycelia.
Under bacterial strain LYLB4 effect, Bacteria erwinia can hardly be grown, mycelia fracture, self-dissolving (Fig. 4 C), and compare germ
Mycelia grows vigorous (Fig. 4 D).Illustrate that bacterial strain LYLB4 can completely inhibit the growth of Bacteria erwinia mycelia.
The application of embodiment 3, bacterial strain LYLB4 in prevention and treatment pear fruit ring spot
One, preventive and therapeutic effect of the bacterial strain LYLB4 to pear fruit ring spot
1, pears pre-process
It chooses that appearance is neat, no disease and pests harm, atraumatic fruit, is first rinsed well with tap water, then with 75% alcohol
It is spare with aseptic filter paper to dry surface after 2min for spray disinfectant.
2, LYLB4 bacteria suspension is handled
Two depth wound as consistent as possible is symmetrically made a call in fruit surface with aseptic card punch (diameter 6mm), in wound
It is inside inoculated with 50 μ L 5 × 10 respectively7The LYLB4 bacteria suspension (LYLB4 processing group) and equivalent sterile water (control group) of CFU/ml, often
8 fruits are handled, are repeated 3 times, are placed at room temperature for one day.
3, it is inoculated with pathogen
The ring spot fungus block of the wound inoculation diameter 6mm of LYLB4 processing group and control group respectively in step 2, connects
Kind processing after be put at 25 DEG C in PE closed box, under the conditions of RH > 90% under the conditions of cultivate, periodically observe and record as a result, and counting
It calculates average attack rate (%) and average lesion diameter (cm).
Bacterial strain LYLB4 to the preventive and therapeutic effect of pear fruit ring spot as shown in Fig. 5, table 3, LYLB4 processing group and control group phase
Than the incidence of pear fruit ring spot obviously weakens.After access Target spot pathogen 3 days, control group starts to fall ill, at LYLB4
Reason can completely inhibit the generation of pear fruit ring spot；With the increase of inoculation time, control group onset speed is very fast, access disease
After bacterium 5 days, control group 100% is fallen ill, and the generation that LYLB4 processing still can inhibit pear fruit ring spot with 100%；Access disease
After bacterium 7 days, LYLB4 processing group starts scab occur；After 9 days, control group rot area be greater than 50%, diameter in 6cm or so, and
The pear fruit lesion diameter of LYLB4 processing group is average in 1.7cm, reduces 71.7%, LYLB4 processing than control and effectively slows down
The occurrence and development of black rot of pear have good control efficiency.
Table 3, LYLB4 are to the preventive and therapeutic effect of pear fruit ring spot
Two, preventive and therapeutic effect of the various concentration LYLB4 bacteria suspension to pear fruit ring spot
1, pears pre-process
It chooses that appearance is neat, no disease and pests harm, atraumatic fruit, is first rinsed well with tap water, then with 75% alcohol
It is spare with aseptic filter paper to dry surface after 2min for spray disinfectant.
2, the LYLB4 bacteria suspension processing of various concentration
Two depth wound as consistent as possible is symmetrically made a call in fruit surface with aseptic card punch (diameter 6mm), wherein
It is inoculated with 5 × 10 respectively in one wound7CFU/ml、5×106CFU/ml、5×105CFU/ml and 5 × 104CFU/ml LYLB4 bacterium
50 μ L of liquid, and it is control that equivalent sterile water, which is added, in another wound.8 fruits of every processing, are repeated 3 times, are placed at room temperature for one
It.
3, it is inoculated with pathogen
The Target spot pathogen of the wound inoculation diameter 6mm of each LYLB4 processing group and control group respectively in step 2
Block, inoculation processing after be put at 25 DEG C in PE closed box, under the conditions of RH > 90% under the conditions of cultivate, periodically observe and record as a result,
And it calculates average attack rate (%) and is averaged lesion diameter (mm).
Various concentration LYLB4 bacteria suspension is as shown in table 4 to the preventive and therapeutic effect of pear fruit ring spot, the LYLB4 of various concentration
Bacteria suspension is handled pear fruit 9 days, with the reduction of YLB4 bacteria suspension concentration, is gradually weakened to the preventive and therapeutic effect of black rot of pear, 5 ×
107CFU/ml LYLB4 processing group effect is best, postpones and slow down the generation of black rot of pear, has good preventive and therapeutic effect.
Table 4, various concentration LYLB4 are to the preventive and therapeutic effect of pear fruit ring spot
| Processing | Scab begins existing | Lesion diameter cm | Inhibiting rate % |
| 5×107CFU/ml | 7th day | 1.76±0.66 | 70.5 |
| 5×106CFU/ml | 6th day | 3.72±0.53 | 37.8 |
| 5×105CFU/ml | 5th day | 5.05±0.61 | 15.6 |
| 5×104CFU/ml | 5th day | 5.25±0.75 | 12.2 |
| CK | 3rd day | 5.98±1.08 |
The application of embodiment 4, bacterial strain LYLB4 in prevention and treatment pear fruit soft rot
1, pears pre-process
It chooses that appearance is neat, no disease and pests harm, atraumatic fruit, is first rinsed well with tap water, then with 75% alcohol
It is spare with aseptic filter paper to dry surface after 2min for spray disinfectant.
2, LYLB4 bacteria suspension is handled
Two depth wound as consistent as possible is symmetrically made a call in fruit surface with aseptic card punch (diameter 6mm), in wound
It is inside inoculated with 50 μ L 5 × 10 respectively7The LYLB4 bacteria suspension (LYLB4 processing group) and equivalent sterile water (control group) of CFU/ml, often
8 fruits are handled, are repeated 3 times, are placed at room temperature for one day.
3, it is inoculated with pathogen
The wound inoculation 1 × 10 of LYLB4 processing group and control group respectively in step 25The ramose root of crawling of spores/ml
It is put into PE closed box after mould 50 μ L inoculation of (Rhizopus stolonifer) suspension processing at 25 DEG C, item under the conditions of RH > 90%
It cultivates, is periodically observed and recorded as a result, and calculating average attack rate (%) and being averaged lesion diameter (mm) under part.
Bacterial strain LYLB4 to the preventive and therapeutic effect of pear fruit soft rot as shown in Fig. 6, table 5, rhizopus stolonifer inoculation after, control group
Rapidly, after access Bacteria erwinia 2 days, control group lesion diameter has reached 6-7cm, and LYLB4 processing group can completely inhibit for morbidity
The generation of pear fruit soft rot；After 4 days, control group pear fruit all rots, and oozes out a large amount of liquid, and be covered with black mycelia and
Spore；And LYLB4 processing group still 100% inhibit pear fruit soft rot generation；LYLB4 processing group starts to fall ill after 10 days.
LYLB4 has good preventive and therapeutic effect to pear fruit soft rot.Table 5, LYLB4 are to the preventive and therapeutic effect of pear fruit soft rot
Claims (9)
1. one plant of Siam bacillus (Bacillus siamensis) LYLB4, deposit number is CGMCC No.11859.
2. Siam bacillus described in claim 1 or its bacteria suspension or its culture solution or its tunning or the bacterium containing it
Application of the agent in prevention and treatment ring spot and/or soft rot.
3. Siam bacillus described in claim 1 or its bacteria suspension or its culture solution or its tunning or the bacterium containing it
Application of the agent in the product of preparation prevention and treatment ring spot and/or soft rot.
4. application according to claim 2 or 3, it is characterised in that: the ring spot and/or soft rot are black rot of pear
And/or pears soft rot.
5. Siam bacillus described in claim 1 or its bacteria suspension or its culture solution or its tunning or the bacterium containing it
Agent antibacterial and/or it is antibacterial in application；
The bacterium is Target spot pathogen (Botryosphaeria dothidea) and/or Bacteria erwinia (Rhizopus
) and/or dendrobium nobile alternaria (Alternaria tenuissima) and/or verticillium dahliae stolonifer
(Verticillium dahliae) and/or Alternaria brassicae (Alternaria brassicae) and/or ash arrhizus bacteria
(Botrytis cinerea) and/or capsicum rhizoctonia solani (Rizoctonia solani) and/or banana blight bacteria
(Fusarium oxysporum) and/or Pyricularia oryzae (Piricularia Oryzae) and/or Fusarium graminearum (Fusarium
graminearum)。
6. Siam bacillus described in claim 1 or its bacteria suspension or its culture solution or its tunning or the bacterium containing it
Application of the agent in preparation antibacterial and/or antibacterial product；
The bacterium is Target spot pathogen (Botryosphaeria dothidea) and/or Bacteria erwinia (Rhizopus
) and/or dendrobium nobile alternaria (Alternaria tenuissima) and/or verticillium dahliae stolonifer
(Verticillium dahliae) and/or Alternaria brassicae (Alternaria brassicae) and/or ash arrhizus bacteria
(Botrytis cinerea) and/or capsicum rhizoctonia solani (Rizoctonia solani) and/or banana blight bacteria
(Fusarium oxysporum) and/or Pyricularia oryzae (Piricularia Oryzae) and/or Fusarium graminearum (Fusarium
graminearum)。
7. a kind of product, active constituent is Siam bacillus described in claim 1 (Bacillus siamensis)
LYLB4 CGMCC No.11859 or its bacteria suspension or its culture solution or its tunning or the microbial inoculum containing it；
The product have the function of it is following 1) or 2) described in:
1) ring spot and/or soft rot are prevented and treated；
2) antibacterial and/or antibacterial；
The bacterium is Target spot pathogen (Botryosphaeria dothidea) and/or Bacteria erwinia (Rhizopus
) and/or dendrobium nobile alternaria (Alternaria tenuissima) and/or verticillium dahliae stolonifer
(Verticillium dahliae) and/or Alternaria brassicae (Alternaria brassicae) and/or ash arrhizus bacteria
(Botrytis cinerea) and/or capsicum rhizoctonia solani (Rizoctonia solani) and/or banana blight bacteria
(Fusarium oxysporum) and/or Pyricularia oryzae (Piricularia Oryzae) and/or Fusarium graminearum (Fusarium
graminearum)。
8. a kind of method of prevention and treatment or assisting in preventing and treating pear fruit ring spot and/or soft rot, includes the following steps: to use claim
Bacteria suspension in product described in Siam bacillus or claim 7 described in 1 handles pear fruit.
9. according to the method described in claim 8, it is characterized by: in the bacteria suspension Siam bacillus concentration be 5 ×
107CFU/ml。
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| CN106282067B (en) * | 2016-08-24 | 2019-10-11 | 李丽丽 | Multifunctional agricultural complex micro organism fungicide and probiotics and application |
| CN107034158B (en) * | 2017-04-27 | 2019-11-19 | 中国科学院微生物研究所 | Siam bacillus B-3 and its application |
| CN108034618B (en) * | 2018-01-24 | 2021-01-01 | 吉林省农业科学院 | Siam bacillus strain 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 |
| CN110964654A (en) * | 2018-09-29 | 2020-04-07 | 福建省农业科学院农业生物资源研究所 | Bacillus antagonistic to fusarium wilt and application thereof |
| CN109207404B (en) * | 2018-10-23 | 2022-02-18 | 山西农业大学 | Siam bacillus YJ15 and application thereof |
| CN109468243B (en) * | 2018-11-13 | 2021-07-30 | 河北农业大学 | Bacillus siamensis 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 |
| CN111909877A (en) * | 2020-08-21 | 2020-11-10 | 四川农业大学 | Application of Siamese bacillus B11 in prevention and/or treatment of chestnut branch blight |
| CN111925963A (en) * | 2020-08-21 | 2020-11-13 | 四川农业大学 | Application of Siamese bacillus B11 in prevention and/or treatment of cotton bamboo blast tip disease |
| CN111961622A (en) * | 2020-08-21 | 2020-11-20 | 四川农业大学 | Siamese bacillus B11 and application thereof in prevention and/or treatment of hybrid bamboo stem-based rot disease |
| CN111849843A (en) * | 2020-08-21 | 2020-10-30 | 四川农业大学 | Application of Siamese bacillus B11 in prevention and/or treatment of bamboo shoot dry blight |
| CN113151059B (en) * | 2021-03-19 | 2022-09-23 | 北京林业大学 | Multifunctional Siamese bacillus and application thereof |
| CN113308392B (en) * | 2021-04-17 | 2023-06-27 | 中国热带农业科学院热带生物技术研究所 | Application of Nori endophytic Siamese bacillus |
| CN113549578B (en) * | 2021-08-03 | 2023-05-09 | 华南农业大学 | Bacillus siamensis BsNlG13 for inhibiting Pyricularia oryzae and promoting seed germination and application thereof |
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| CN114891670B (en) * | 2022-04-28 | 2023-09-19 | 岭南师范学院 | Siamese bacillus ZJh with broad-spectrum antibacterial property and application thereof |
| CN115109723B (en) * | 2022-06-23 | 2023-06-23 | 湖北工业大学 | Bacillus californicus ST-1 and application thereof in pear disease control or storage and preservation |
| CN116253600B (en) * | 2022-11-30 | 2024-02-02 | 云南省农业科学院农业环境资源研究所 | Organic fertilizer and method for promoting vegetable growth and/or preventing and controlling vegetable diseases |
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