CN109077067B - Biocontrol bacterium and application thereof in prevention and control of gray mold of crops - Google Patents
Biocontrol bacterium and application thereof in prevention and control of gray mold of crops Download PDFInfo
- Publication number
- CN109077067B CN109077067B CN201810933984.7A CN201810933984A CN109077067B CN 109077067 B CN109077067 B CN 109077067B CN 201810933984 A CN201810933984 A CN 201810933984A CN 109077067 B CN109077067 B CN 109077067B
- Authority
- CN
- China
- Prior art keywords
- biocontrol
- gray mold
- crops
- pepper
- biocontrol bacterium
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/10—Animals; Substances produced thereby or obtained therefrom
Landscapes
- Life Sciences & Earth Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Health & Medical Sciences (AREA)
- Plant Pathology (AREA)
- Microbiology (AREA)
- Pest Control & Pesticides (AREA)
- Biotechnology (AREA)
- Virology (AREA)
- Agronomy & Crop Science (AREA)
- Dentistry (AREA)
- Wood Science & Technology (AREA)
- Environmental Sciences (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention relates to a new biocontrol bacterium and application thereof in the prevention and treatment of gray mold of crops; the biocontrol bacterium CGMCC No.15763 can effectively prevent and treat gray mold of crops and has growth promoting effect on the crops; particularly in the aspect of pepper planting application, the control effect on pepper gray mold is obvious, the drought resistance of pepper can be improved, the growth is promoted, and the yield is increased; moreover, compared with the chemical agents in the prior art, the biocontrol bacterium is nontoxic to human and livestock, does not pollute the environment, has no residue, and is beneficial to the sustainable development of ecological environment.
Description
Technical Field
The invention relates to a biocontrol bacterium and application thereof in preventing and controlling gray mold of crops, belonging to the technical field of biological prevention and control of crops.
Background
The gray mold is a relatively serious disease in agricultural production in China, and particularly brings serious harm to the planting and generation of the peppers. The pepper is one of the favorite vegetables, and with the specialization of planting, due to the single variety and the difficulty in stubble cutting, the basic number of field botrytis cinerea is increased year by year through continuous cropping cultivation for many years, so that the occurrence of pepper botrytis cinerea is more and more serious, and the general loss rate reaches 15% -20%; in severe cases, 30 to 50 percent; the highest rate can reach 100 percent, which causes a great deal of rotting of leaves and loss of edible value; it can also cause field losses and even cause serious damage in the storage and transport of the fruit.
At present, the breeding work for resisting the gray mold has not made a breakthrough progress, and the breeding of the pepper for resisting the gray mold is basically in a stagnation state. At the present stage, main measures for preventing and treating the botrytis cinerea in the cultivation, production and after adoption of the peppers are measures such as spraying of chemical agents, but the botrytis cinerea generates drug resistance to various bactericides due to long-term use of the chemical agents. The botrytis cinerea on various vegetables in fields and protected cultivation facilities in Beijing, Shanghai and the like in China has relatively serious drug resistance, and the resistance frequency of the botrytis cinerea on systemic bactericides such as carbendazim and the like even in some regions reaches 100 percent.
Therefore, the search for new control measures is becoming a major priority for gray mold control. As a new prevention and treatment measure, compared with other methods, the biological prevention and treatment measure has the characteristics of safety, effectiveness and durability, and particularly avoids a series of problems caused by chemical prevention and treatment. The biocontrol agent has good disease control effect, is nontoxic to human and livestock, does not pollute the environment and has no residue; the killing specificity to plant diseases and insect pests is strong, natural enemies are not damaged, beneficial organisms are obtained, and ecological balance can be kept; the production raw materials and the active ingredients are natural products which are easy to degrade and can return to the nature, thereby ensuring the sustainable development.
Biological control of gray mold is becoming a focus of research, and it is desired in the art to screen out microorganisms capable of controlling gray mold.
Disclosure of Invention
In view of the above problems and/or other problems of the related art, an aspect of the present invention provides a biocontrol bacterium, wherein the biocontrol bacterium is Bacillus belief (Bacillus velezensis) deposited in the common microorganism center of the china committee for culture collection management of microorganisms with the collection number of CGMCC No. 15763.
The invention also provides a fungus medicine composition adopting the biocontrol bacterium, wherein the fungus medicine composition comprises a bactericide and the fungus powder of the biocontrol bacterium; the bactericide is any one or a mixture of a plurality of pyrrole bactericides, dicarboximide bactericides, nicotinamide bactericides or methoxyl acrylic ester bactericides.
In another aspect, the invention provides a fungal drug preparation, wherein the fungal drug preparation comprises the biocontrol bacteria and an agriculturally and pharmaceutically acceptable auxiliary, or the fungal drug preparation comprises the fungal drug composition and an agriculturally and pharmaceutically acceptable auxiliary.
The invention also provides the application of the biocontrol strain in preventing and treating crop gray mold, promoting crop growth or planting hot pepper.
The invention also provides application of the fungus medicine composition in preventing and treating gray mold of crops.
The invention also provides the application of the microbial preparation in the aspect of preventing and treating the gray mold of crops.
The biocontrol bacterium CGMCC No.15763 provided by the invention can effectively prevent and treat gray mold of crops and has a growth promoting effect on the crops; particularly in the aspect of pepper planting application, the control effect on pepper gray mold is obvious, the drought resistance of pepper can be improved, the growth is promoted, and the yield is increased; moreover, compared with the chemical agents in the prior art, the biocontrol bacterium is nontoxic to human and livestock, does not pollute the environment, has no residue, and is beneficial to the sustainable development of ecological environment.
Drawings
FIG. 1 is a photograph showing the results of a plate antagonism test against Botrytis cinerea.
Detailed Description
The present invention will be further described with reference to specific embodiments, but the present invention is not limited to these specific embodiments.
Materials, reagents and the like used in the following embodiments are commercially available unless otherwise specified. The specific techniques or conditions are not indicated, and the procedures or conditions are described in the literature in the art (for example, refer to J. SammBruk et al, molecular cloning, A laboratory Manual, third edition, science Press, translated by Huang Petang et al) or in accordance with the product instructions.
First, the operation process of the present inventors for obtaining the biocontrol bacterium (Bacillus velezensis) CGMCC No.15763 of the present invention by separating and screening from the natural environment will be described below.
(1) Isolation of potential biocontrol bacteria
Isolation of potential biocontrol bacteria present in vitro of plants:
shearing roots, stems and leaves of hot peppers (collected from Shanghai African garden of fruits and vegetables) by 3g respectively, weighing 3g of rhizosphere soil as a sample, and adding the sample into 27mL of sterilized water (containing sterilized glass beads) respectively;
placing the obtained sample in 180rmp shaking table, oscillating for 0.5h, standing for 5min, respectively sucking supernatant for gradient dilution, and taking dilution 104、105And 106Double dilution; taking 0.1mL of LB-coated plate from three gradient dilutions, culturing for 24-48 h at 28 ℃, selecting a plate with the colony number between 50-300, counting and selecting all colonies, purifying, inoculating into a test tube containing 5mL of LB liquid culture medium, shaking at 200rpm at 28 ℃ for culturing for 24-48 h, taking the bacterial suspension, mixing with an equal volume of 80% glycerol solution, and storing at-70 ℃ for later use (Berg et al, 2000).
Isolation of potential biocontrol bacteria present in plants:
3g of each root, stem and leaf of a pepper (collected from Shanghai African garden) is cut to serve as a sample. Firstly, the surface of a sample is disinfected and sterilized, and the specific disinfection and sterilization process is as follows: soaking the surface of the sample with 1mL of 1% sodium hypochlorite for 5min and 70% alcohol for 2min respectively, and cleaning the surface of the sample with sterile water for 3 times (taking 0.1mL of water from the last cleaning, coating the water on an R2A flat plate, culturing the water with bacteria shaking at 28 ℃ and 200rpm for 48h, or taking 0.1mL of water from the last cleaning, adding R2A culture solution, culturing the water with vibration at 28 ℃, and considering that the surface of the sample is disinfected when the water grows aseptically after 48 h).
Respectively grinding the samples with sterilized surfaces by using sterilized grinding bowls, adding 3mL of sterile water, filtering by using sterile cotton cloth, diluting by 10 times by using the sterile water, coating 0.1mL of LB plate with each point, repeating the steps for 3 times, culturing at 28 ℃ for 48 hours, counting, selecting all bacterial colonies on the plate, separating and purifying, inoculating the bacterial colonies into a test tube containing 5mL of LB liquid culture medium, culturing at 28 ℃ for 24-48 hours by shaking at 200rpm, uniformly mixing the bacterial suspension with an isovolumetric 80% glycerol solution, and storing at-70 ℃ for later use (Krechel et al, 2002).
(2) Enzyme production and metabolite activity determination primary screening biocontrol strain
The substances such as protein, chitin, glucan, cellulose and the like are components of the cell wall of the plant pathogenic fungi, can generate strains with the activities of the degrading enzymes of the substances, can degrade the cell wall of the plant pathogenic fungi preliminarily presumed, and have antagonistic activity on the plant pathogenic fungi, so the strains can be used as the standard for primarily screening the biocontrol fungi. Selecting a strain in a vigorous growth period, placing the strain on a protease enzyme activity determination culture medium, a culture medium (Chi-adsorbents) taking colloidal chitin as a unique carbon source, a beta-1, 3-glucanase enzyme activity determination culture medium and a cellulase enzyme activity determination culture medium, culturing for 3 days at 28-30 ℃ after inoculation, observing the existence of a transparent ring, and respectively recording the inner diameter and the outer diameter of the transparent ring. Strains with one or more of the above enzyme producing and secondary metabolite producing activities are retained for further screening.
(3) Plate antagonism test with Botrytis cinerea to further screen anti-Botrytis cinerea strains
Inoculating Botrytis cinerea (the part near the edge of the colony and with a sterilized hole puncher to form a dish with a diameter of 0.6 cm) cultured in an incubator at 25 deg.C for 5 days to the center of a PDA plate, inoculating the bacteria solution of biocontrol bacteria 5YN8 (viable bacteria concentration of 5 × 10) at two opposite positions with equal distance from the inoculation position with sterile toothpick9～1×1010CFU/ml), the other two opposite corners were not inoculated (as controls), and the plates were inverted and placed in an incubator at 25 ℃ for 4 days; the above treatments were performed in 3 parallel experiments simultaneously. Observing the existence and the size of the inhibition zone so as to judge the existence and the strength of the antagonism of the potential biocontrol strain to the botrytis cinerea.
A strain is separated from root soil of a healthy pepper plant, and is confirmed to be a potential biocontrol strain through enzyme production activity detection and a flat plate antagonism test, and the strain is named as biocontrol bacteria 5YN 8.
The identification process of the biocontrol bacterium 5YN8 of the invention is as follows:
molecular biological identification
And (3) carrying out molecular biological identification on the biocontrol bacterium 5YN8 obtained by screening by adopting phylogenetic analysis according to the sequence of the 16S rRNA of the bacterium.
The biocontrol bacterium 5YN8 obtained by screening was cultured in LB medium at 28 ℃ until logarithmic phase, and centrifuged at 12000 rpm for 5 minutes to collect the cells.
The genome DNA of the collected thallus is extracted by adopting a genome DNA rapid extraction kit of Shanghai Saibaoshi Gene technology Limited.
And amplifying the 16S rRNA sequence and the gyrB gene segment of the biocontrol bacterium 5YN8 by adopting a PCR amplification kit of Tiangen Biochemical technology Co., Ltd, and specifically operating according to the kit instruction by taking the extracted genome DNA product as a template.
The forward primer sequence used for amplifying the 16S rRNA sequence was: 5'-AGAGTTTGATCACTGGCTCAG-3', the sequence of the reverse primer used for amplification is: 5'-CTACGGAGTACCTTGTTACGAC-3' are provided.
The forward primer sequence adopted for amplifying the gyrB gene fragment is as follows: 5 '-GAAGTCATCATGACCGTTCTGCAYGCNGGNGGNAARTTYGA-3', the reverse primer sequence adopted by the amplification is as follows: 5 '-AGCAGGATACGGATGTGCGAGCCRTCNACRTCNGCRTCNGTCAT-3'.
The amplified bands were recovered by cutting using DNA purification recovery kit from Tiangen Biochemical technology Ltd. The recovered sample was sent to Shanghai Bioengineering Co., Ltd for sequencing.
The sequencing result is subjected to homology comparison (similarity comparison with a 16S rRNA sequence and a gyrB gene fragment of a known bacterium) through NCBI BLAST software, and the biocontrol bacterium 5YN8 of the invention is identified as Bacillus belgii (Bacillus velezensis).
Third, preservation of strains
The biocontrol bacterium 5YN8 for preventing and treating the gray mold of crops belongs to Bacillus velezensis, is preserved in the common microorganism center of China Committee for culture Collection of microorganisms, has the preservation date of 2018, 05 and 15 days, and has the preservation number of CGMCC No. 15763.
Fourthly, the culture method and the application of the biocontrol bacteria
The biocontrol bacterium 5YN8 of the invention is streaked on an LB plate and cultured for 24h at 28 ℃, after a single bacterium grows out, the single bacterium is picked up and inoculated into a test tube containing 5ml of LB culture solution, and cultured for 12h at 28 ℃ and 200rpm as seed solution. Inoculating the seed solution to 500mL LB culture solution at a ratio of 1:100, culturing at 28 deg.C and 200rpm for 24 hr until the total viable bacteria concentration is 5 × 109～1×1010CFU/ml。
In general, the biocontrol bacterium 5YN8 of the present invention is applied by diluting the cultured bacterial liquid 100 to 200 times immediately after transplanting into the soil, irrigating roots of the transplanted crops, and simultaneously diluting 500 to 1000 times for foliar application.
Fifthly, the result of the plate antagonism test of the biocontrol bacterium 5YN8 of the invention to botrytis cinerea
The specific test method comprises the following steps: inoculating Botrytis cinerea (the part near the edge of the colony and with a sterilized hole puncher to form a dish with a diameter of 0.6 cm) cultured in an incubator at 25 deg.C for 5 days to the center of a PDA plate, inoculating the bacteria solution of biocontrol bacteria 5YN8 (viable bacteria concentration of 5 × 10) at two opposite positions with equal distance from the inoculation position with sterile toothpick9～1×1010CFU/ml), the other two opposite corners were not inoculated (as controls), and the plates were inverted and placed in an incubator at 25 ℃ for 4 days; the above treatments were performed in 3 parallel experiments simultaneously.
Referring to FIG. 1, a photograph of a plate antagonism test against Botrytis, wherein a site labeled "5 YN 8" is a plaque inoculated with biocontrol bacterium 5YN8, and a site labeled "CK" is a control.
Through measurement, the radius of bacterial plaque of the biocontrol bacteria 5YN8 on the left side and the right side is 0.72cm (the radius taking the biocontrol bacteria inoculation point as the center of a circle), the distance from the edge of the bacterial plaque of the biocontrol bacteria 5YN8 to the edge of the botrytis cinerea is 0.53cm, the radius of the bacterial plaque of the botrytis cinerea on the upper side and the lower side (the comparison position) is 3.93cm (the radius taking the center point of the circular bacterial disc as the center of a circle), an obvious transparent ring is formed at the junction of the biocontrol bacteria 5YN8 and the botrytis cinerea, and no transparent ring is formed around the filter paper sheet at the.
Therefore, from the results of the plate antagonism test described above, it is found that: 5YN8 has obvious antagonistic activity on Botrytis cinerea.
Sixthly, the result of the greenhouse disease prevention test of the biocontrol bacterium 5YN8 of the invention on the gray mold of hot pepper
After 3-4 leaves of the pepper seedlings are finished, transplanting the pepper seedlings, and transplanting the pepper seedlings into 12 small cups (for example, transplanting the pepper seedlings into disposable plastic cups) at a time, wherein 2 plants are transplanted in each cup. After the seedlings are eased, a bacterial solution of biocontrol bacteria 5YN8 (the viable bacteria concentration is 5 multiplied by 10)7CFU/ml) is adopted for root irrigation, and 20ml of water is irrigated to each seedling; this is the experimental group (experimental group for gray mold test of pepper).
Meanwhile, in the same transplanting mode, each seedling is irrigated with 20ml of clear water to serve as a control group (a control group for pepper gray mold test).
On the same day, the test group and the control group were inoculated by spraying (Botrytis cinerea spore suspension) in such an amount that the leaf surface was completely wetted and dropped.
Moisturizing at 20-25 deg.C, and counting the disease occurrence after the control group has the disease occurrence (about 20 days).
For statistics on the incidence, the grading criteria for gray mold disease are presented below:
level 0: no disease spots;
level 1: the lesion area accounts for less than 5% of the whole leaf area;
and 3, level: the lesion area accounts for 6 to 10 percent of the whole leaf area;
and 5, stage: the lesion area accounts for 11 to 20 percent of the whole leaf area;
and 7, stage: the lesion area accounts for 21-40% of the whole leaf area;
and 9, stage: the lesion area accounts for more than 40% of the whole leaf area.
Calculation formulas for "disease severity" and "control effect" are:
disease severity { ∑ (number of disease leaves at each level × relative level value) }/(survey total × 9);
the control effect is (control group disease severity-treatment group disease severity)/control group disease severity;
see table 1 below for test results.
TABLE 1
| Disease severity (%) | Control effect (%) | |
| Experimental group | 9.04±0.028 | 90.67 |
| Control group | 96.94±0.019 | - |
Wherein the result of the severity of the disease is the mean value plus or minus standard error.
The results of the greenhouse disease prevention test for gray mold of pepper in table 1 show that the biocontrol bacterium 5YN8 of the present invention has significant control effect on gray mold of pepper (the control effect on gray mold of pepper is 90.67% after inoculation for about 20 days).
Seventhly, the greenhouse test result of the influence of the biocontrol bacterium 5YN8 on the drought resistance of the pepper
After 3-4 leaves of the pepper seedlings, transplanting the pepper seedlings, and watering 25ml of tap water every other day. After 15 days of seedling transplantation, the treatment was started, and the following two treatment groups were set. 24 seedlings were treated each.
Biocontrol bacteria 5YN8 treatment group: each seedling is watered with 20ml, after 5 days of normal watering induction, water is cut off for simulating drought treatment, and watering is carried out again after 20 days of simulating drought treatment.
Control treatment group: each seedling is watered with 20ml of clear water, after 5 days of normal watering induction, water is cut off for simulating drought treatment, and watering is carried out again after 20 days of simulating drought treatment.
The test results for the control treatment group showed: after 8 days of simulated drought treatment, the control group of peppers showed severe drought symptoms, moderate wilting symptoms appeared on all leaves (leaf curly or drooping, no recovery in the morning and evening), and stem wilting drooping;
the test results of the biocontrol bacterium 5YN8 treatment group show that: after 8 days of simulated drought treatment, the pepper only had cotyledons sagging, and a few leaf edges curled, showing slight wilting symptoms.
At the time point of simulating 8 days of drought treatment, the relative water content of the leaves, the root system reduction capacity, the MDA content of the leaves, the relative conductivity of the leaves and the chlorophyll content under drought stress were tested (each test was repeated three times).
The detection result under the drought stress condition shows that:
1) relative water content: the average relative water content of the pepper leaves of the biocontrol bacterium 5YN 8-treated group is 93.3 percent, which is 44 percent higher than the average value of the control-treated group;
2) root system reduction capacity: the root system reduction strength of the pepper of the biocontrol bacterium 5YN8 treatment group is about twice of that of the control treatment group;
3) the MDA content of the leaves is: the content of MDA in the leaves of the pepper of the biocontrol bacterium 5YN8 treatment group is 21.4 percent lower than that of the control treatment group;
4) relative conductivity of the blade: the relative conductivity of the leaves of the pepper of the biocontrol bacterium 5YN8 treatment group is 56.0 percent, which is 28.2 percent lower than that of the control treatment group;
5) chlorophyll content: the contents of chlorophyll a, chlorophyll b and total chlorophyll in leaves of Capsici fructus treated with biocontrol bacteria 5YN8 are respectively 1.16mg g-1Fw，0.85mg g-1Fw and 2.02mg g-1Fw, 16%, 20% and 20.2% higher, respectively, compared to the control treated group.
The combination of the results shows that the biocontrol bacterium 5YN8 can improve the drought resistance of the pepper by protecting the integrity of cell membranes, keeping the root activity and maintaining the high-level chlorophyll content.
Eighthly, the greenhouse test result of the biocontrol bacterium 5YN8 for promoting the growth of the pepper
After 3-4 leaves of the pepper seedlings are finished, transplanting the pepper seedlings, and transplanting the pepper seedlings into 12 small cups (for example, transplanting the pepper seedlings into disposable plastic cups) at a time, wherein 2 plants are transplanted in each cup.
After the seedlings are eased, a bacterial solution of biocontrol bacteria 5YN8 (the viable bacteria concentration is 5 multiplied by 10)7CFU/ml) is adopted for root irrigation, and 20ml of water is irrigated to each seedling; this is the experimental group (experimental group for pepper growth promotion test).
Meanwhile, in the same transplanting mode, each seedling is irrigated with 20ml of clear water to serve as a control group (a control group for a pepper growth promotion test).
After 45 days of treatment, pepper seedlings were observed for growth and the biomass of each plant was counted: stem thickness, leaf number, plant height, chlorophyll, results are shown in table 2 below.
TABLE 2
The values in the tables are mean. + -. standard error, as is the case with the tables below.
As can be seen from the results in table 2, the biomass (stem thickness, leaf number, plant height, chlorophyll) of the experimental group treated with the biocontrol bacterium 5YN8 was significantly increased, and the total biomass was 87.8% higher than that of the control group. Therefore, the result shows that the biocontrol bacterium 5YN8 has the growth promoting effect on pepper planting.
Ninth, the field test result of the biocontrol bacterium 5YN8 of the invention for preventing and controlling gray mold of various crops (hot pepper, tomato, cucumber and strawberry)
The field test is carried out in the area of Huaiyin region of Huai Yingzhen Daniancun of Huai-Yin of Jiangsu province, is divided into 2 processing areas, a 2m wide protection row is arranged in the middle, field management is carried out according to the conventional cultivation management technology, and the area of each processing area is 20m2The length is 10m, the width is 2m, and the periphery of each treatment area is provided with a drainage irrigation ditch, so that the drainage irrigation can be realized, and the drainage irrigation is convenient.
On the day of seedling transplantation, bacterial solution of biocontrol bacterium 5YN8 (viable bacteria concentration is 5 × 10)7CFU/ml) is adopted for root irrigation, and 20ml of water is irrigated to each seedling; this is the experimental group (experimental group for preventing and controlling gray mold in field).
Meanwhile, in the same transplanting mode, each seedling is irrigated with 20ml of clear water to serve as a control group (a control group for field gray mold control).
After the seedlings are eased, spraying and inoculating the botrytis cinerea spore suspension until the surfaces of the leaves are completely wet and drip. After the control had developed, the disease was observed.
See table 3 below for the field test results for control of gray mold of pepper.
TABLE 3
| Disease severity (%) | Control effect (%) | |
| Experimental group | 11.04±0.011 | 88.48 |
| Control group | 95.83±0.024 | - |
See table 4 below for the results of field trials for control of tomato gray mold.
TABLE 4
| Disease severity (%) | Control effect (%) | |
| Experimental group | 13.05±0.034 | 86.44 |
| Control group | 96.27±0.025 | - |
See table 5 below for the field test results for control of cucumber gray mold.
TABLE 5
| Disease severity (%) | Control effect (%) | |
| Experimental group | 12.11±0.014 | 87.11 |
| Control group | 93.94±0.030 | - |
See table 6 below for the results of field trials for control of tomato gray mold.
TABLE 6
| Disease severity (%) | Control effect (%) | |
| Experimental group | 11.37±0.041 | 87.97 |
| Control group | 94.48±0.026 | - |
As can be seen from the results of tables 3, 4, 5 and 6, the biocontrol bacterium 5YN8 of the present invention has a control effect of more than 85% on gray mold of various crops (pepper, tomato, cucumber and strawberry), and has a significant control effect, especially the best control effect on gray mold of pepper.
Ten, the field test result of the growth promoting effect of the biocontrol bacterium 5YN8 on various crops (hot pepper, tomato, cucumber and strawberry)
The field test is carried out in the area of Huaiyin region of Huai Yingzhen Daniancun of Huai-Yin of Jiangsu province, is divided into 2 processing areas, a 2m wide protection row is arranged in the middle, field management is carried out according to the conventional cultivation management technology, and the area of each processing area is 20m2The length is 10m, the width is 2m, and the periphery of each treatment area is provided with a drainage irrigation ditch, so that the drainage irrigation can be realized, and the drainage irrigation is convenient.
Performing field management according to conventional cultivation management technique, and using bacterial solution of biocontrol bacteria 5YN8 (viable bacteria concentration is 5 × 10)7CFU/ml) is adopted for root irrigation, and 20ml of water is irrigated to each seedling; this is the experimental group (experimental group of field growth promotion test).
Meanwhile, in the same manner, each seedling was irrigated with 20ml of clear water as a control group (control group for field growth promotion test).
After 5 weeks of treatment, the seedlings of the crop were observed for changes and the biomass of each plant was counted: stem thickness, leaf number, plant height, chlorophyll.
See table 7 below for field test results on growth promoting effect of pepper.
TABLE 7
See table 8 below, the results of field trials on tomato growth promotion.
TABLE 8
See table 9 below, the results of field trials on cucumber growth promoting effects.
TABLE 9
See table 10 below, field test results for strawberry growth promoting effect.
Watch 10
From the results of tables 7, 8, 9 and 10, it can be seen that the biocontrol bacterium 5YN8 of the present invention has significant growth promoting effect on various crops (pepper, tomato, cucumber and strawberry), each biomass (stem thickness, leaf number, plant height, chlorophyll) is significantly increased, and the total biomass is higher than that of the control group by more than 82%.
In conclusion, the biocontrol bacterium 5YN8 can effectively prevent and treat gray mold of crops and has a growth promoting effect on the crops; particularly in the aspect of pepper planting application, the control effect on pepper gray mold is remarkable, and the drought resistance of pepper can be improved, the growth can be promoted, and the yield can be improved.
After the biocontrol bacterium 5YN8 has the control effect on the gray mold of crops, a conventional bactericide (chemical agent) for controlling the gray mold can be compounded with the bacterium powder of the biocontrol bacterium 5YN8 to obtain a bacterium-medicine composition.
The conventional bactericides for preventing and treating the gray mold comprise pyrrole bactericides, dicarboximide bactericides, nicotinamide bactericides and methoxy acrylate bactericides.
Therefore, any one or more of the above conventional bactericides can be compounded with the bacterial powder of the biocontrol bacterium 5YN8 to obtain the bacterial-medicine composition, and the skilled person can mix the proportions according to the specific needs and the drug effects and can verify the composition through a limited number of experiments.
In addition, after the biocontrol bacterium 5YN8 of the invention is known to have the control effect on the gray mold of crops, a fungicide preparation for controlling the gray mold of crops can be obtained by combining the biocontrol bacterium 5YN8 of the invention with an auxiliary agent acceptable in pesticides.
Similarly, the above-mentioned fungicide composition (composition of fungicide and fungus powder of biocontrol bacterium 5YN8 of the present invention) can be combined with an agriculturally acceptable adjuvant to obtain a fungicide preparation for controlling gray mold of crops.
It should be understood that although the present description refers to embodiments, not every embodiment contains only a single technical solution, and such description is for clarity only, and those skilled in the art should make the description as a whole, and the technical solutions in the embodiments can also be combined appropriately to form other embodiments understood by those skilled in the art.
The above-listed detailed description is only a specific description of a possible embodiment of the present invention, and they are not intended to limit the scope of the present invention, and equivalent embodiments or modifications made without departing from the technical spirit of the present invention should be included in the scope of the present invention.
Claims (6)
1. A biocontrol bacterium, which is characterized in that: the biocontrol bacterium is Bacillus velezensis (Bacillus velezensis) which is preserved in the China general microbiological culture Collection center of the China Committee for culture Collection of microorganisms with the preservation number of CGMCC No. 15763.
2. A fungal pharmaceutical composition using the biocontrol bacterium of claim 1, wherein: the microbial drug composition comprises a bactericide and the biocontrol microbial powder of claim 1; the bactericide is any one or a mixture of a plurality of pyrrole bactericides, dicarboximide bactericides, nicotinamide bactericides or methoxyl acrylic ester bactericides.
3. A bacterial preparation, which is characterized in that: the fungal drug preparation comprises the biocontrol bacterium and an agriculturally pharmaceutically acceptable auxiliary agent of claim 1, or the fungal drug preparation comprises the fungal drug composition of claim 2 and an agriculturally pharmaceutically acceptable auxiliary agent.
4. The use of the biocontrol bacterium of claim 1 in the control of gray mold in crops, in the growth promotion of crops or in the planting of peppers.
5. The use of a fungicide composition according to claim 2 for the control of gray mold in crops.
6. The use of a fungal preparation according to claim 3 for the control of gray mold in crops.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810933984.7A CN109077067B (en) | 2018-08-16 | 2018-08-16 | Biocontrol bacterium and application thereof in prevention and control of gray mold of crops |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810933984.7A CN109077067B (en) | 2018-08-16 | 2018-08-16 | Biocontrol bacterium and application thereof in prevention and control of gray mold of crops |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN109077067A CN109077067A (en) | 2018-12-25 |
| CN109077067B true CN109077067B (en) | 2021-04-30 |
Family
ID=64793489
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810933984.7A Active CN109077067B (en) | 2018-08-16 | 2018-08-16 | Biocontrol bacterium and application thereof in prevention and control of gray mold of crops |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN109077067B (en) |
Families Citing this family (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109880778B (en) * | 2019-04-24 | 2022-09-09 | 贵州大学 | Composite microbial inoculum with growth promoting and yield increasing effects on capsicum and application thereof |
| CN110295129B (en) * | 2019-07-23 | 2021-03-19 | 山东省农药科学研究院 | Biocontrol bacterium for preventing and treating gray mold and powdery mildew of cucumber and application thereof |
| CN110804570B (en) * | 2019-11-20 | 2022-04-08 | 中国农业大学 | Bacillus beijerinckii for simultaneously degrading zearalenone and aflatoxin and application thereof |
| CN111394281B (en) * | 2020-04-16 | 2022-08-23 | 山东省科学院生态研究所 | Bacillus belgii BV03 microbial agent and preparation method and application thereof |
| CN116445376B (en) * | 2023-06-15 | 2023-11-14 | 中国农业科学院农业资源与农业区划研究所 | Bacillus bailii and application thereof |
Citations (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20100052797A (en) * | 2008-11-11 | 2010-05-20 | 한국화학연구원 | Bacillus velezensis pb3 strain and method for the biological control of plant diseases using same |
| KR20100055625A (en) * | 2008-11-18 | 2010-05-27 | 한국화학연구원 | Bacillus velezensis s3-5 strain and method for the biological control of plant diseases using same |
| KR20120010017A (en) * | 2010-07-23 | 2012-02-02 | 대한민국(농촌진흥청장) | A novel Bacillus velezensis B-42 for multi function |
| CN102391040A (en) * | 2011-07-19 | 2012-03-28 | 杨凌绿力生物工程有限公司 | New organic microbial fertilizer and preparation process thereof |
| CN103173386A (en) * | 2013-03-12 | 2013-06-26 | 山东农业大学 | Bio-control strain G1 for preventing and controlling pepper phytophthora blights and applications thereof |
| CN106035381A (en) * | 2016-06-27 | 2016-10-26 | 中国烟草总公司郑州烟草研究院 | Bactericide composition containing benzamide bactericides and biocontrol bacterium, preparation and applications of bactericide composition and preparation |
| CN106172503A (en) * | 2016-06-27 | 2016-12-07 | 中国烟草总公司郑州烟草研究院 | A kind of bacterium drug composition, preparation and application thereof containing dithiocarbamate antibacterial and biocontrol microorganisms |
| CN106479934A (en) * | 2016-11-23 | 2017-03-08 | 河北省科学院生物研究所 | A kind of ash arrhizus bacteria antagonistic strain and its screening technique and application |
| CN106967632A (en) * | 2017-02-23 | 2017-07-21 | 南京农业大学 | It is a kind of efficiently to prevent and treat various crop gray mold and the microorganism mixture of capsicum resistance be lifted |
| CN107242249A (en) * | 2017-04-19 | 2017-10-13 | 黑龙江中医药大学 | It is a kind of to prevent and treat ginseng and the gemma bacteria composition of American Ginseng black spot and gray mold and application thereof |
| KR101803678B1 (en) * | 2017-03-21 | 2017-12-01 | 충북대학교 산학협력단 | Primer sets for detecting Dyella thiooxydans ATSB10 and uses thereof |
| CN107446847A (en) * | 2017-08-14 | 2017-12-08 | 云南农业大学 | One plant of Bei Laisi bacillus GT11 and its application |
| CN107964514A (en) * | 2017-09-08 | 2018-04-27 | 华夏春秋科技发展（北京）有限责任公司 | A kind of Bei Laisi bacillus and its application on plant |
| CN108004185A (en) * | 2018-01-09 | 2018-05-08 | 中国农业科学院植物保护研究所 | One plant of tool diseases prevention, growth-promoting, drought resisting function plant endogenesis Bei Laisi bacillus and its application |
| CN108330089A (en) * | 2018-03-13 | 2018-07-27 | 华中农业大学 | A kind of active Bei Laisi bacillus HNt7 of broad-spectrum antibacterial and preparation method and application |
-
2018
- 2018-08-16 CN CN201810933984.7A patent/CN109077067B/en active Active
Patent Citations (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20100052797A (en) * | 2008-11-11 | 2010-05-20 | 한국화학연구원 | Bacillus velezensis pb3 strain and method for the biological control of plant diseases using same |
| KR20100055625A (en) * | 2008-11-18 | 2010-05-27 | 한국화학연구원 | Bacillus velezensis s3-5 strain and method for the biological control of plant diseases using same |
| KR20120010017A (en) * | 2010-07-23 | 2012-02-02 | 대한민국(농촌진흥청장) | A novel Bacillus velezensis B-42 for multi function |
| CN102391040A (en) * | 2011-07-19 | 2012-03-28 | 杨凌绿力生物工程有限公司 | New organic microbial fertilizer and preparation process thereof |
| CN103173386A (en) * | 2013-03-12 | 2013-06-26 | 山东农业大学 | Bio-control strain G1 for preventing and controlling pepper phytophthora blights and applications thereof |
| CN106172503A (en) * | 2016-06-27 | 2016-12-07 | 中国烟草总公司郑州烟草研究院 | A kind of bacterium drug composition, preparation and application thereof containing dithiocarbamate antibacterial and biocontrol microorganisms |
| CN106035381A (en) * | 2016-06-27 | 2016-10-26 | 中国烟草总公司郑州烟草研究院 | Bactericide composition containing benzamide bactericides and biocontrol bacterium, preparation and applications of bactericide composition and preparation |
| CN106479934A (en) * | 2016-11-23 | 2017-03-08 | 河北省科学院生物研究所 | A kind of ash arrhizus bacteria antagonistic strain and its screening technique and application |
| CN106967632A (en) * | 2017-02-23 | 2017-07-21 | 南京农业大学 | It is a kind of efficiently to prevent and treat various crop gray mold and the microorganism mixture of capsicum resistance be lifted |
| KR101803678B1 (en) * | 2017-03-21 | 2017-12-01 | 충북대학교 산학협력단 | Primer sets for detecting Dyella thiooxydans ATSB10 and uses thereof |
| CN107242249A (en) * | 2017-04-19 | 2017-10-13 | 黑龙江中医药大学 | It is a kind of to prevent and treat ginseng and the gemma bacteria composition of American Ginseng black spot and gray mold and application thereof |
| CN107446847A (en) * | 2017-08-14 | 2017-12-08 | 云南农业大学 | One plant of Bei Laisi bacillus GT11 and its application |
| CN107964514A (en) * | 2017-09-08 | 2018-04-27 | 华夏春秋科技发展（北京）有限责任公司 | A kind of Bei Laisi bacillus and its application on plant |
| CN108004185A (en) * | 2018-01-09 | 2018-05-08 | 中国农业科学院植物保护研究所 | One plant of tool diseases prevention, growth-promoting, drought resisting function plant endogenesis Bei Laisi bacillus and its application |
| CN108330089A (en) * | 2018-03-13 | 2018-07-27 | 华中农业大学 | A kind of active Bei Laisi bacillus HNt7 of broad-spectrum antibacterial and preparation method and application |
Non-Patent Citations (7)
| Title |
|---|
| Bacillus velezensis, a potential and effcient biocontrol agent in control of pepper gray mold caused by Botrytis cinerea;Chun-Hao Jiang et al;《Biological Control》;20180801;第126卷;第147-157页 * |
| Complete genome sequence of Bacillus velezensis S3-1, a potential biologicalpesticide with plant pathogen inhibiting and plant promoting capabilities;Qing Jina et al;《Journal of Biotechnology》;20170712;第259卷;第199-203页 * |
| Effects of Bacillus velezensis strain BAC03 in promoting plant growth;Qingxiao Meng et al;《Biological Control》;20160324;第98卷;第18-26页 * |
| 番茄灰霉病菌颉颃菌株的筛选及功能基因的分析;索雅丽等;《中国植保导刊》;20100820;第30卷(第8期);第7-10页 * |
| 贝莱斯芽孢杆菌（Bacillus velezensis）研究进展;蔡高磊等;《北方园艺》;20180630(第12期);第162-167页 * |
| 贝莱斯芽孢杆菌L-1 对梨灰霉和青霉病菌的抑制作用评价及全基因组分析;孙平平等;《微生物学报》;20180528;第58卷(第9期);第1637-1646页 * |
| 贝莱斯芽孢杆菌的分类、拮抗功能及其应用研究进展;张德锋等;《微生物学通报》;20200409;https://doi.org/10.13344/j.microbiol.china.190947 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN109077067A (en) | 2018-12-25 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN109077067B (en) | Biocontrol bacterium and application thereof in prevention and control of gray mold of crops | |
| CN109749943B (en) | Trichoderma asperellum and application thereof | |
| CN105907680B (en) | A kind of bacillus subtilis J-5 and its bacteria agent and application | |
| CN104630071A (en) | Polysporus trichoderma and application thereof | |
| CN112322560B (en) | Bacillus belgii and application thereof in prevention and control of pear diseases | |
| CN104694397A (en) | Chaetomium globosum and application thereof | |
| CN108148794A (en) | A kind of the bacillus subtilis DYr3.3 and preparation method and application of broad-spectrum antibacterial activity | |
| CN111778173B (en) | Bacillus subtilis Pro1A2, microbial inoculum and preparation method thereof, and application of bacillus subtilis Pro1A2 in cultivation of melons | |
| CN115058358A (en) | Salt-tolerant bacillus and application thereof | |
| Zakharchenko et al. | Effect of rhizosphere bacteria Pseudomonas aureofaciens on the resistance of micropropagated plants to phytopathogens | |
| CN113151044B (en) | Pseudomonas for preventing and treating diseases of radix pseudostellariae and separation, screening and identification method and application | |
| CN109055265B (en) | Biocontrol bacterium and application thereof in prevention and control of gummy stem blight of crops | |
| CN109609403B (en) | Biocontrol bacterium and application thereof in prevention and control of downy mildew of crops | |
| CN115960777B (en) | Bacillus pseudomycoides and application thereof in prevention and treatment of vegetable epidemic disease | |
| CN115873760A (en) | Bacillus amyloliquefaciens and biocontrol microbial inoculum and application thereof | |
| CN104962501A (en) | Preparation and application of bacterial strain resistant to gray mould of vegetables and fruits and antagonist | |
| CN111808778B (en) | Bacillus wegener for preventing and treating verticillium wilt and culture method thereof, microbial inoculum and preparation method and application thereof | |
| CN111548951B (en) | Bacillus subtilis Pro6A5, microbial inoculum and preparation method thereof, and application of bacillus subtilis Pro6A5 in cultivation of melons | |
| CN108384736A (en) | A kind of bacterium and its screening technique having inhibiting effect to citrus rubber germ | |
| CN100557013C (en) | The bacterial strain PX 35 of control curing glasshouse vegetable epidemic disease | |
| CN105886447A (en) | Bacillus strain DR011 and plant drought-enduring inducting agent prepared from bacillus strain DR011 | |
| CN103865825B (en) | Prevent and treat the biocontrol microorganisms of sesame blight, separation method, microbial inoculum and application thereof | |
| CN103865824B (en) | A kind of prevent and treat sesame blight biocontrol microorganisms, separation method, microbial inoculum and application thereof | |
| CN102329758B (en) | Biological prevention bacterial strain BS1 (bacillus cereus) for preventing and controlling greenhouse cucumber downy mildew and application thereof | |
| CN111334453B (en) | Biocontrol bacterium and application thereof in prevention and control of sheath blight of crops |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |