CN113755389A - Bacillus belgii and application thereof - Google Patents
Bacillus belgii and application thereof Download PDFInfo
- Publication number
- CN113755389A CN113755389A CN202111166440.0A CN202111166440A CN113755389A CN 113755389 A CN113755389 A CN 113755389A CN 202111166440 A CN202111166440 A CN 202111166440A CN 113755389 A CN113755389 A CN 113755389A
- Authority
- CN
- China
- Prior art keywords
- dpt
- bacillus
- bacillus velezensis
- peanut
- suspension
- 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.)
- Granted
Links
Images
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G13/00—Protecting plants
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Biotechnology (AREA)
- Organic Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Zoology (AREA)
- Genetics & Genomics (AREA)
- Medicinal Chemistry (AREA)
- Microbiology (AREA)
- Biomedical Technology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- Virology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Toxicology (AREA)
- Environmental Sciences (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The application relates to the technical field of biocontrol strains, and particularly discloses bacillus belgii and application thereof. The preservation number of the Bacillus beleisi is CGMCC NO. 20317; a culture comprising the Bacillus belgii or a processed product thereof; a microbial agent comprising a carrier and Bacillus belgii and/or a culture or processed product thereof; a microbial fertilizer comprising bacillus belgii or a microbial agent; and the use of Bacillus belgii, its culture or processed product, microbial agent or microbial fertilizer in the preparation of a composition for ameliorating and/or preventing crop diseases. The Bacillus belgii provided by the application can be used as a biocontrol strain, and can effectively improve and prevent crop diseases.
Description
Technical Field
The application relates to the technical field of biocontrol strains, in particular to bacillus belgii and application thereof.
Background
Crops play an important role in social production as one of the main industries in China. In recent years, however, the effect of crop diseases on crop yield has been on the trend of increasing year by year.
At present, crop diseases are mainly prevented and controlled by crop rotation, deep ploughing, resistant variety cultivation, chemical agents and the like. However, the prevention and control methods all address the symptoms and do not address the root causes, and long-term use of chemical agents can cause pollution of field soil by the chemical agents, so that the quality and the yield of agricultural products are reduced, and even the safety problem of the agricultural products can occur.
Biological prevention and control are important directions for green development of agriculture, and the prevention and control of crop diseases by using biocontrol strains is a difficult problem which is urgently needed to be solved in the green development of agriculture all the time.
Disclosure of Invention
In order to improve and control crop diseases by using biocontrol strains, the application provides bacillus belgii and application thereof.
In a first aspect, the present application provides a Bacillus belief (Bacillus velezensis), which adopts the following technical scheme:
bacillus velezensis (Bacillus velezensis) with the preservation number of CGMCC NO. 20317.
The Bacillus belgii has dry wrinkles, opaqueness, milk white color, irregular or nearly circular edges and micro-protrusions on the colony surface on an LB solid culture medium.
The application provides Bacillus velezensis which can effectively inhibit the growth of hyphae of pathogenic bacteria of various crop diseases so as to reduce the influence of the pathogenic bacteria of various crop diseases on crops. The Bacillus belgii provided by the application can reduce the disease strain rate and death rate of crops, thereby effectively reducing the morbidity of various crops and having better control effect on various crop diseases.
In a second aspect, the present application provides a culture or processed product thereof comprising the above bacillus belgii.
In a third aspect, the present application provides a bacterial suspension prepared using the above Bacillus belgii.
Preferably, the concentration of the Bacillus belgii suspension is 1.0X 109-4.0×109CFU/mL。
In a fourth aspect, the present application provides a sterile fermentation broth prepared using the above-described bacillus belgii.
In a fifth aspect, the present application provides a microbial inoculum, which adopts the following technical scheme:
a microbial agent comprising the above Bacillus belgii and/or the above culture or processed product.
A microbial agent comprising the above bacillus belgii, the above bacterial suspension and/or the above sterile fermentation broth.
In a sixth aspect, the present application provides a microbial fertilizer, which adopts the following technical scheme:
a biological fertilizer comprising the above bacillus belgii or the above microbial agent.
In a seventh aspect, the present application provides the use of the above-mentioned bacillus belgii, a culture or processed product thereof, a microbial agent, or a microbial fertilizer for the preparation of a composition for ameliorating and/or preventing diseases of agricultural crops.
Preferably, the crop diseases for which the composition is used for improving and/or preventing can be one or more of southern blight of peanut, corn small leaf spot, cotton wilt, tomato leaf mold, tomato gray mold, tobacco brown spot, vegetable and flower colletotrichum gloeosporioides and grape gray mold.
In a specific embodiment, Bacillus velezensisDPT-03 bacterial suspension (1.0X 10)9CFU/mL) has a control effect on the peanut southern blight reaching 62.50 percent when inoculated for 60 days.
In a specific embodiment, a suspension of Bacillus velezensis DPT-03 (1.0X 10)9CFU/mL) has the control effect of 50.00 percent on the corn southern leaf blight at 60d of inoculation.
In a specific embodiment, a suspension of Bacillus velezensis DPT-03 (4.0X 10)9CFU/mL) has the control effect of 82.85 percent on cotton wilt after being inoculated for 25 days.
In a specific embodiment, a suspension of Bacillus velezensis DPT-03 (4.0X 10)9CFU/mL) has the control effect on tomato leaf mold reaching 82.35 percent when inoculated for 30 days.
In a specific embodiment, a suspension of Bacillus velezensis DPT-03 (4.0X 10)9CFU/mL) has the control effect of 88.46 percent on the gray mold of the tomato after being inoculated for 30 days.
Strain preservation information: the Bacillus belgii provided by the application is named as Bacillus belgii DPT-03(Bacillus velezensis DPT-03), is preserved in China general microbiological culture Collection center (CGMCC), has a preservation number of CGMCC NO.20317, and has a preservation date of 2020, 7 and 8 days. The 16S rDNA sequence is shown in SEQ ID NO 1.
In summary, the present application has the following beneficial effects:
1. the Bacillus velezensis DPT-03 provided by the application can reduce the disease rate and death rate of crops, so that the morbidity of various crops is effectively reduced, and a better control effect on various crop diseases is achieved. Especially has better control effect on one or more of southern blight of flowers, corn small spot, cotton wilt, tomato leaf mold, tomato gray mold, tobacco brown spot, vegetable and flower colletotrichum gloeosporioides and grape gray mold.
2. The culture or processed product, bacterial suspension, sterile fermentation liquor, microbial agent and microbial fertilizer prepared by the Bacillus belgii provided by the application can effectively reduce the morbidity of various crops, and have better prevention and treatment effects on various crop diseases.
Drawings
FIG. 1 is the streaking result of Bacillus velezensis DPT-03 on LB solid medium plate.
FIG. 2 is a Neighbour-join tree of Bacillus velezensis DPT-03 based on the 16S rDNA sequence.
FIG. 3 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of hyphae of pathogenic bacteria of peanut southern blight.
FIG. 4 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of hyphae of pathogenic bacteria of corn northern leaf blight.
FIG. 5 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of cotton wilt-causing pathogenic bacteria hyphae.
FIG. 6 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of pathogenic bacteria hyphae of tomato leaf mold.
FIG. 7 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of tomato gray mold pathogen hyphae.
FIG. 8 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of tobacco alternaria alternate pathogenic bacteria hypha.
FIG. 9 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of hyphae of pathogenic colletotrichum gloeosporioides.
FIG. 10 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of tobacco alternaria alternate pathogenic bacteria hyphae.
Detailed Description
The application provides a Bacillus belgii, which is named as Bacillus velezensis DPT-03(Bacillus velezensis DPT-03) and is preserved in China general microbiological culture Collection center (CGMCC), the preservation number is CGMCC NO.20317, and the preservation date is 7/8/2020. The 16S rDNA sequence is shown in SEQ ID NO 1.
The instruments, reagents, materials and the like referred to in the following examples and comparative examples are conventional instruments, reagents, materials and the like in the related art unless otherwise specified. The test methods and the detection methods mentioned in the following examples are, unless otherwise specified, conventional test methods and conventional detection methods in the related art.
The following matters related in the present application are specifically:
the LB solid medium formulation is as follows: the volume of the extract is 1L, and the extract comprises 10g of tryptone, 5g of yeast extract, 10g of NaCl and 15g of agar.
The formula of the LB liquid culture medium is as follows: the total amount of the extract is 1L, and the extract comprises 10g of tryptone, 5g of yeast extract and 10g of NaCl.
PDA solid medium: comprises potato 200g, glucose 20g, agar 15g, and natural pH calculated by 1L.
PDA liquid culture medium: comprises potato 200g, glucose 20g, and natural pH calculated by 1L.
Wherein, the pathogenic bacteria of the southern blight of the peanuts are provided by peanut research institute in Shandong province; the pathogenic bacteria of corn small spot disease, cotton wilt, tomato leaf mold, tomato gray mold, tobacco brown spot, colletotrichum gloeosporioides and grape gray mold are provided by plant protection research institute of Chinese academy of agricultural sciences.
The present application is described in further detail below with reference to examples 1-17, comparative examples 1-9, FIGS. 1-10, and test runs one-seven.
Examples
Example 1
The embodiment provides a screening method of Bacillus velezensis DPT-03.
One, sampling site
The sampling site in this example was diseased soil in the peanut test field of army biotechnology limited in the beijing century.
Secondly, separating and purifying strains
(1) Collecting 20g of disease soil blocks at a sampling site, and taking the disease soil blocks back to a laboratory as a soil sample;
(2) weighing 10g of the soil sample obtained in the step (1), adding the weighed soil sample into a triangular flask filled with 90mL of sterile water, and then placing the triangular flask in a constant-temperature shaking table at the temperature of 28 ℃ at 150r/min for culturing for 30min to obtain a sample suspension;
(3) taking 1mL of the sample suspension obtained in the step (2), and after diluting the sample suspension according to a 10-fold proportion gradient, obtaining 10-1、10-2、10-3、10-4、10-5And 10-6Multiple dilutions of the sample;
(4) respectively taking 10 obtained in the step (3)-4、10-5And 10-6 Coating 100 mu L of the multiplied sample diluent on an LB solid culture medium flat plate, and placing the coated LB solid culture medium flat plate in a constant temperature incubator at 28 ℃ for culturing for 48 h; after the cultivation, colonies grow on an LB solid medium flat plate;
(5) picking the single colony growing on the LB solid medium flat plate in the step (4), streaking and transferring the single colony to a new LB solid medium flat plate, and putting the streaked LB solid medium flat plate in a constant temperature incubator at 28 ℃ for culturing for 48 hours; only one colony form is observed in the cultured LB solid medium flat plate, which indicates that the separation and purification are finished; and (4) storing the separated and purified LB solid medium plate at 4 ℃ for later use.
By the separation and purification of the strains in the soil sample, 8 strains are obtained in total.
Thirdly, screening of strains
(1) Respectively activating the 8 strains obtained by separation and purification, inoculating the activated strains into corresponding LB liquid culture medium, and culturing the inoculated LB liquid culture medium in a constant-temperature shaking table at the temperature of 28 ℃ at 150r/min for 24 hours; strain OD in cultured LB liquid medium600Reaching more than 1.2 to obtain tested bacterial liquid of 8 strains;
(2) screening resistant strains by using a confrontation growth method:
A. putting the peanut southern blight pathogenic bacteria on a PDA solid culture medium flat plate for activation to obtain an activated pathogenic bacteria flat plate;
B. sampling from the activated pathogenic bacteria flat plate by using a puncher to obtain pathogenic bacteria blocks, and placing the pathogenic bacteria blocks in the center of a blank PDA solid culture medium flat plate; then 2 sterilized filter paper sheets with the diameter of 6mm are parallelly placed at the position 20mm away from the center of the PDA solid culture medium flat plate, 10 mu L of activated tested bacterial liquid is added to one of the filter paper sheets to be used as a detection sample, and 10 mu L of sterile PDA liquid culture medium is added to the other filter paper sheet to be used as a reference sample; placing the PDA solid culture medium flat plate in a constant temperature incubator at 28 ℃ for culturing for 72 h; if a bacteriostatic zone appears on the cultured PDA solid medium flat plate, the strain is a bacterial strain resistant to southern blight pathogenic bacteria.
And (3) respectively screening the tested bacterial liquids obtained in the step (1) by utilizing the steps, and preliminarily judging the antibacterial capacity of 8 strains according to whether an antibacterial band is generated on a PDA solid culture medium flat plate after culture and the generated antibacterial band width.
Through screening, a strain with the bacteriostatic zone width exceeding 15mm is finally obtained and named as DPT-03, and the strain DPT-03 shows an obvious inhibiting effect on pathogenic bacteria of the southern blight. After the strain DPT-03 is cultured by using an LB liquid culture medium, the strain is preserved by glycerol at the temperature of-80 ℃.
Example 2
The embodiment provides an identification method of Bacillus velezensis DPT-03. The identified object was the strain DPT-03 selected in example 1.
Morphological characteristics of strain DPT-03
Inoculating the strain DPT-03 to an LB solid medium plate, placing the plate in a constant temperature incubator at 37 ℃ for culturing for 48h, wherein the surface of a bacterial colony is dry and wrinkled, opaque, milk white, irregular or nearly circular in edge and has microprotrusions, and a streak picture of the strain DPT-03 is shown in figure 1. The morphological characteristics of the thalli observed by a microscope are rod-shaped, single cell, gram staining positive and spore.
II, identification of strain DPT-03
The strain DPT-03 obtained by screening in example 1 is inoculated in an LB solid medium, after overnight culture, a fresh single colony is taken from a plate and placed in a 1.5mL centrifuge tube, 10uL of S2 lysate is added, the mixture is shaken and mixed evenly, the mixture is kept stand at room temperature for 20min, then diluted by 20 times, shaken and mixed evenly, 12000r/min is carried out, centrifugation is carried out for 2min, and the supernatant is taken as a template for PCR amplification.
The amplification conditions were as follows:
the forward primer was 27F: AGAGTTTGATCCTGGCTCAG;
the reverse primer is 1492R: TACGGCTACCTTGTTACGACTT, respectively;
the amplification enzyme was code # AS 11;
the amplification program is 94 ℃ for 5 min; 30s at 94 ℃; 30s at 55 ℃; 90s at 72 ℃; the cycle number is 35 times, the temperature is 72 ℃ for 7min, and the temperature is 4 ℃.
The amplification reaction system is as follows: 2x easylaq SuperMix 15 uL; 27F (10. mu.M) 1.5 uL; 1492R (10. mu.M) 1.5 uL; template 5 uL; ddH2O 7 uL; total 30 uL.
And (4) carrying out agarose gel electrophoresis on the PCR amplification product, and recovering and purifying the gel block.
The agarose gel electrophoresis configuration system is as follows: 2.16mL of distilled water; 2.64mL of 30% Acr-Bis (29: 1); 1M Tris pH 8.83.04 mL; 0.08mL of 10% SDS; 0.08mL of 10% sodium persulfate; TEMED 0.0032 mL; total 8 mL.
And carrying out sanger sequencing on the purified product to obtain a forward and reverse sequencing result, splicing the obtained gene sequence by using DNAMAN software, and comparing 16S rDNA in an www.ezbiocloud.net database to identify the type of the microorganism.
And determining phylogenetic related genera of the strain DPT-03 by utilizing 16S rDNA sequence alignment, and constructing a phylogenetic tree by adopting an adjacency method based on the 16S rDNA sequence.
FIG. 2 is a Neighbour-join tree of Bacillus velezensis DPT-03 based on the 16S rDNA sequence.
As can be seen from FIG. 2, the strain DPT-03 and Bacillus velezensis have the closest genetic relationship, and the sequence similarity is 99.8%. Based on the 16S rDNA phylogenetic analysis of the strain DPT-03, the strain DPT-03 obtained by screening is determined to be Bacillus subtilis DPT-03(Bacillus velezensis DPT-03). The sequence of the 16S rDNA is shown in SEQ ID NO 1.
Example 3
This example provides a method for preparing a suspension of Bacillus velezensis DPT-03.
Taking a Bacillus velezensis DPT-03 single colony on an LB solid culture medium plate preserved at 4 ℃, inoculating the single colony in an LB liquid culture medium, and placing the single colony in a constant temperature shaking table at 37 ℃ and 180r/min for culturing for 24 hours to obtain a Bacillus velezensis DPT-03 bacterial suspension for later use.
Example 4
This example provides a method for preparing a Bacillus velezensis DPT-03 sterile fermentation broth.
Taking a Bacillus velezensis DPT-03 single colony on an LB solid culture medium plate stored at 4 ℃, inoculating the single colony in an LB liquid culture medium, and culturing in a constant temperature shaking table at 37 ℃ and 180r/min for 24 hours to obtain a Bacillus velezensis DPT-03 bacterial suspension;
taking the bacterial suspension of Bacillus velezensis DPT-03, centrifuging for 10min under the condition of 10000r/min, collecting supernatant, and filtering by using a sterile filter membrane of 0.22 mu m to obtain the Bacillus velezensis DPT-03 sterile fermentation liquor.
Detection test
Test for detection
The Bacillus velezensis DPT-03 bacterial suspension prepared in the example 3 is used as a detection object to detect the influence of the bacterial suspension on the growth of hyphae of pathogenic bacteria of southern blight of flowers, pathogenic bacteria of small leaf spot of corn, pathogenic bacteria of blight of cotton, pathogenic bacteria of leaf mold of tomato, pathogenic bacteria of gray mold of tomato, pathogenic bacteria of brown spot of tobacco, pathogenic bacteria of colletotrichum of vegetables and flowers, and pathogenic bacteria of gray mold of grape.
Detection method
(1) Activation of pathogenic bacteria: punching holes on a PDA solid culture medium plate for pathogen of southern blight of peanuts stored at 4 ℃, a PDA solid culture medium plate for pathogen of small spot disease of corn, a PDA solid culture medium plate for pathogen of blight of cotton, a PDA solid culture medium plate for pathogen of leaf mold of tomato, a PDA solid culture medium plate for pathogen of gray mold of tomato, a PDA solid culture medium plate for pathogen of red spot disease of tobacco, a PDA solid culture medium plate for pathogen of colletotrichum gloeosporioides of vegetables and flowers and a PDA solid culture medium plate for pathogen of gray mold of grapes, respectively inoculating circular fungus cakes with the diameter of 7.5mm onto the PDA liquid culture medium plate, and culturing in a constant temperature incubator at 28 ℃ for 3-4 days for later use;
(2) perforating a PDA liquid culture medium flat plate of the peanut southern blight pathogenic bacteria, the corn small spot pathogenic bacteria, the cotton wilt pathogenic bacteria, the tomato leaf mold pathogenic bacteria, the tomato gray mold pathogenic bacteria, the tobacco scab pathogenic bacteria, the vegetable and flower colletotrichum gloeosporioides pathogenic bacteria and the tobacco scab pathogenic bacteria cultured in the step (1), and respectively inoculating circular fungus cakes with the diameter of 7.5mm to the center of a new PDA solid culture medium flat plate;
then, 6mm sterilized filter paper sheets are respectively placed at the relative positions 20mm away from the center of the PDA solid culture medium flat plate, and 10 mu L of Bacillus velezensis DPT-03 bacterial suspension is added on the filter paper sheets to be used as a detection sample.
And (3) placing the PDA solid medium plate into a constant-temperature incubator at 28 ℃ for culture, and observing the bacteriostasis condition of the Bacillus velezensis DPT-03 bacterial suspension to each pathogenic bacterium at 5 days respectively.
Second, the detection result
The results of the tests are shown in FIGS. 3-10.
FIG. 3 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of hyphae of pathogenic bacteria of peanut southern blight.
FIG. 4 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of hyphae of pathogenic bacteria of corn northern leaf blight.
FIG. 5 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of cotton wilt-causing pathogenic bacteria hyphae.
FIG. 6 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of pathogenic bacteria hyphae of tomato leaf mold.
FIG. 7 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of tomato gray mold pathogen hyphae.
FIG. 8 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of tobacco alternaria alternate pathogenic bacteria hypha.
FIG. 9 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of hyphae of pathogenic colletotrichum gloeosporioides.
FIG. 10 shows the effect of Bacillus velezensis DPT-03 suspension on the growth of tobacco alternaria alternate pathogenic bacteria hyphae.
From the above fig. 3-fig. 10, it can be seen that Bacillus velezensis DPT-03 provided by the present application has a good inhibition effect on one or more of southern blight of peanut, corn small spot, cotton wilt, tomato leaf mold, tomato gray mold, tobacco brown spot, vegetable and flower colletotrichum anthracnose and grape gray mold. Therefore, the Bacillus velezensis DPT-03 provided by the application can reduce the disease rate and death rate of crops, so that the morbidity of various crops is effectively reduced, and a better control effect on various crop diseases is achieved.
Example 5
This example provides a potted peanut plant comprising a suspension of Bacillus velezensis DPT-03 bacteria. The suspension of Bacillus velezensis DPT-03 used was the suspension of Bacillus velezensis DPT-03 prepared in example 3.
The preparation method of the potted plant comprises the following steps:
(1) preparing a peanut potted plant: uniformly mixing 2kg of soil and 1g of compound fertilizer 15-15-15, placing the mixture in a flowerpot, burying 3 full and healthy peanut kernels into the soil, thinning after 15 days, and reserving 1 peanut kernel in each pot;
(2) inoculation of pathogenic bacteria: inoculating the peanut southern blight pathogenic bacteria into the soil of the peanut potted plant after 20 days, wherein the inoculation amount is 20mL, and the concentration is 1.0 multiplied by 108CFU/mL；
(3) Inoculation of the bacterial suspension: inoculating pathogenic bacteria of southern blight of peanut for 2 days, and adding water to the solution at a concentration of 1.0 × 109And inoculating the CFU/mL bacterial suspension of Bacillus velezensis DPT-03 into soil of potted peanut plants, wherein the inoculation amount is 20mL for each potted plant, and preparing the potted plants comprising the bacterial suspension of Bacillus velezensis DPT-03.
Example 6
This example provides a potted peanut plant comprising Bacillus velezensis DPT-03 sterile fermentation broth. The Bacillus velezensis DPT-03 sterile fermentation broth used was the Bacillus velezensis DPT-03 sterile fermentation broth prepared in example 4.
The preparation method of the above potting is different from the preparation method in example 5 in that:
the step (3) is as follows: inoculating sterile fermentation liquor: inoculating pathogenic bacteria of southern blight of peanut for 2 days, and adding water to the solution at a concentration of 1.0 × 109Inoculating the sterile fermentation liquor prepared from the bacterial suspension of the Bacillus velezensis DPT-03 into the soil of potted peanut plants, wherein the inoculation amount is 20mL per potted plant, and obtaining the sterile fermentation liquor containing the Bacillus velezensis DPT-03Potting.
Comparative example 1
This comparative example provides a peanut pot comprising a treatment agent. The preparation method is different from that of the example 5 in that:
the step (3) is as follows: inoculation of thifluzamide pesticide: and (3) inoculating thifluzamide pesticide into soil of peanut potted plants after inoculating the peanut southern blight pathogenic bacteria for 2d, wherein the inoculation amount is 20mL per potted plant, and preparing the potted plants containing the treatment agent. Wherein the thifluzamide pesticide is 240g/L thifluzamide suspending agent, and the thifluzamide pesticide is diluted by 500 times for use.
Comparative example 2
The comparative example provides a potted peanut plant. The preparation method is different from that of the example 5 in that:
the step (3) is as follows: inoculation of sterile water: inoculating the peanut southern blight pathogenic bacteria for 2d, and then inoculating sterile water into soil of the peanut pot culture, wherein the inoculation amount is 20mL per pot culture, so as to prepare the pot culture containing the sterile water.
Test 2
The peanut potted plants prepared in example 5, example 6, comparative example 1 and comparative example 2 were used as test subjects, and 15 of the potted plants were arranged in parallel to test the occurrence of diseases in each potted plant.
Detection method
And (3) respectively inoculating 20d, 40d and 60d of peanut southern blight pathogenic bacteria to the peanut pot culture, detecting the disease condition of the peanut pot culture, and counting the disease rate and the control effect of the peanut pot culture.
Second, calculating method
And respectively calculating the morbidity and the prevention and treatment effect of the potted peanut.
(1) The formula for the incidence of disease is as follows:
incidence (%) — number of diseased plants/total number of plants × 100%;
(2) the calculation formula of the control effect is as follows:
control effect (%) - (control incidence-treatment incidence)/control incidence × 100%
Third, the detection result
The results are shown in Table 1.
TABLE 1 test results of test No. two
As shown in the test results in table 1, it is understood from the test results of comparative example 2 that the onset time of the peanut plants is earlier when the peanut potting is inoculated with the peanut southern blight pathogenic bacteria alone, and the incidence rate of the peanut plants reaches 20.00% when the peanut potting is inoculated with the peanut southern blight pathogenic bacteria 20 d. According to the detection results of the embodiment 5, the embodiment 6 and the comparative example 1, it is known that the peanut plants do not have morbidity in the peanut potted plants inoculated with the peanut southern blight pathogenic bacteria 20d, the Bacillus velezensis DPT-03 bacterial suspension, the Bacillus velezensis DPT-03 sterile fermentation liquor and the treatment agent, and the control effects of the three on the peanut southern blight reach 100%. According to the results, the Bacillus velezensis DPT-03 bacterial suspension or the Bacillus velezensis DPT-03 sterile fermentation liquor provided by the application can ensure that the peanut plants do not have morbidity in a short period. Therefore, in the process of cultivating peanut plants, the Bacillus velezensis DPT-03 provided by the application can effectively prevent the peanut southern blight from occurring in the peanut plants in a short period, and can delay the morbidity time and the morbidity index of the peanut plants.
By comparing the results of the embodiment 5, the embodiment 6, the comparative example 1 and the comparative example 2 when the peanut pot culture is inoculated with the peanut southern blight pathogenic bacteria 40d, at this time, the control effect of the peanut pot culture inoculated with the Bacillus velezensis DPT-03 bacterial suspension on the peanut southern blight reaches 59.01%, and the control effect of the peanut pot culture inoculated with the Bacillus velezensis DPT-03 sterile fermentation liquor on the peanut southern blight reaches 49.98%; the peanut pot culture inoculated with the treatment medicament has the prevention and treatment effect on the southern blight of the peanuts of 59.01 percent. From the above results, when the peanut potting is inoculated with the peanut southern blight pathogenic bacteria 40d, the control effect of the peanut potting inoculated with the Bacillus velezensis DPT-03 bacterial suspension, the Bacillus velezensis DPT-03 sterile fermentation liquor and the treatment agent on the peanut southern blight is reduced to a certain extent compared with the control effect of the peanut potting inoculated with the peanut southern blight pathogenic bacteria 20d, the control effect of the peanut potting inoculated with the Bacillus velezensis DPT-03 bacterial suspension on the peanut southern blight is basically the same as the control effect of the peanut potting inoculated with the treatment agent on the peanut southern blight, and the control effect of the peanut potting inoculated with the Bacillus velezensis DPT-03 bacterial suspension on the peanut southern blight is superior to the control effect of the peanut potting inoculated with the Bacillus velezensis DPT-03 sterile fermentation liquor on the peanut southern blight. Therefore, in the process of cultivating peanut plants, the Bacillus velezensis DPT-03 bacterial suspension provided by the application can effectively improve the control effect on the southern blight of peanuts in the middle period.
By comparing the results of the embodiment 5, the embodiment 6, the comparative example 1 and the comparative example 2 when the peanut pot culture is inoculated with the peanut southern blight pathogenic bacteria for 60 days, at this time, the control effect of the peanut pot culture inoculated with the Bacillus velezensis DPT-03 bacterial suspension on the peanut southern blight reaches 62.50%, and the control effect of the peanut pot culture inoculated with the Bacillus velezensis DPT-03 sterile fermentation liquor on the peanut southern blight reaches 30.36%; the peanut pot culture inoculated with the treatment medicament has the effect of preventing and treating the southern blight of the peanuts of 57.14 percent. From the results, when the peanut potting is inoculated with the peanut southern blight pathogenic bacteria for 60 days, the control effect of the peanut potting inoculated with the Bacillus velezensis DPT-03 sterile fermentation liquor and the treatment agent on the peanut southern blight is reduced compared with the control effect of the peanut potting inoculated with the peanut southern blight pathogenic bacteria for 40 days, and particularly the control effect of the peanut potting inoculated with the Bacillus velezensis DPT-03 sterile fermentation liquor on the peanut southern blight is reduced more obviously; the control effect of the peanut pot culture inoculated with the Bacillus velezensis DPT-03 bacterial suspension on the peanut southern blight is higher than that of the peanut pot culture inoculated with the peanut southern blight pathogenic bacteria 40d, and the control effect of the peanut pot culture inoculated with the Bacillus velezensis DPT-03 bacterial suspension on the peanut southern blight is better than that of the peanut pot culture inoculated with the Bacillus velezensis DPT-03 sterile fermentation liquor on the peanut southern blight and is also better than that of the peanut pot culture inoculated with a treatment agent on the peanut southern blight. Therefore, in the process of cultivating peanut plants, thalli in the Bacillus velezensis DPT-03 bacterial suspension can be stably propagated in the potting soil, and active substances capable of inhibiting the peanut southern blight pathogenic bacteria are continuously generated, so that the peanut southern blight pathogenic bacteria can be continuously prevented and controlled, the incidence rate of the peanut southern blight can be effectively reduced, and the prevention and control effect of the peanut southern blight is improved.
Example 7
This example provides a potted corn plant comprising a suspension of Bacillus velezensis DPT-03 bacteria. The suspension of Bacillus velezensis DPT-03 used was the suspension of Bacillus velezensis DPT-03 prepared in example 3.
The preparation method of the potted plant comprises the following steps:
(1) preparing a corn pot culture: uniformly mixing 2kg of soil and 1g of compound fertilizer 15-15-15, placing the mixture in a flowerpot, burying 3 full and healthy corn kernels in the soil, thinning after 15 days, and reserving 1 corn kernel in each pot;
(2) inoculation of pathogenic bacteria: inoculating the corn small leaf spot pathogenic bacteria to the soil of the corn potted plant after 20 days, wherein the inoculation amount is 20mL, and the concentration is 1.0 multiplied by 108CFU/mL；
(3) Inoculation of the bacterial suspension: inoculating pathogenic bacteria of corn small spot for 2 days, and adding into the mixture to obtain the mixture with concentration of 1.0 × 109And inoculating the CFU/mL bacterial suspension of Bacillus velezensis DPT-03 into the soil of the potted corn plants, wherein the inoculation amount is 20mL for each potted plant, and preparing the potted plants comprising the bacterial suspension of Bacillus velezensis DPT-03.
Comparative example 3
This comparative example provides a potted corn plant. The preparation method differs from example 7 in that:
the step (3) is as follows: inoculating carbendazim pesticide: inoculating the pathogenic bacteria of the corn small spot disease for 2 days, and then inoculating the carbendazim pesticide into soil of the corn pot culture, wherein the inoculation amount is 20mL per pot culture, so as to obtain the corn pot culture. Wherein, the carbendazim pesticide is diluted by 500 times for use.
Comparative example 4
This comparative example provides a potted corn plant. The preparation method differs from example 7 in that:
the step (3) is as follows: inoculation of sterile water: inoculating the corn small spot pathogen for 2d, and then inoculating sterile water into soil of the corn pot culture, wherein the inoculation amount is 20mL per pot culture, so as to prepare the pot culture containing the sterile water.
Test III
The corn pots prepared in example 7, comparative example 3, and comparative example 4 were used as test subjects, and 30 of each of the pots were arranged in parallel to test the incidence of diseases in each pot.
Detection method
And respectively inoculating 30d and 60d of pathogenic bacteria of the corn small spot disease to the corn pot culture, detecting the disease condition of the corn pot culture, and counting the disease incidence and the prevention and treatment effect of the corn pot culture.
Second, calculating method
The calculation formula of the morbidity and the prevention and treatment effect is the same as the corresponding formula in the second detection test.
Third, the detection result
The results are shown in Table 2.
TABLE 2 test results of test III
As shown in the test results in Table 2, the test results of comparative example 4 revealed that the incidence of corn plants in the case of potting corn plants inoculated with 30 days of the pathogenic bacteria of northern leaf blight reached 23.30%. The detection results of the example 7 and the comparative example 3 show that the morbidity of the corn plants in the corn pot culture inoculated with the corn small spot disease pathogenic bacteria 30d, the Bacillus velezensis DPT-03 bacterial suspension and the treatment agent is respectively 10.00% and 13.30%, and the control effects respectively reach 57.08% and 42.92%. According to the results, the Bacillus velezensis DPT-03 bacterial suspension provided by the application can reduce the morbidity of corn plants in a short period. Therefore, in the process of culturing the corn plants, the Bacillus velezensis DPT-03 bacterial suspension provided by the application can effectively improve the situation that the corn plants have corn small spot in a short period.
By comparing the results of the example 7, the comparative example 3 and the comparative example 4 when the corn pot culture is inoculated with the corn northern leaf blight pathogenic bacteria for 60 days, the control effect of the corn pot culture inoculated with the Bacillus velezensis DPT-03 bacterial suspension on the northern leaf blight is 50.00 percent, and the control effect of the corn pot culture inoculated with the treatment agent on the northern leaf blight is 43.37 percent. From the above results, it can be seen that when the maize potted plant is inoculated with the maize small spot disease pathogenic bacteria for 60d, the control effect of the maize potted plant inoculated with the Bacillus velezensis DPT-03 bacterial suspension on the maize small spot disease is reduced to a certain extent compared with the control effect of the maize small spot disease pathogenic bacteria for 30d, and the control effect is still better than that of the maize potted plant inoculated with the treatment agent on the maize small spot disease. Therefore, in the process of cultivating corn plants, the Bacillus velezensis DPT-03 bacterial suspension provided by the application can effectively reduce the incidence rate of the corn small spot disease and has a good prevention and treatment effect on the corn small spot disease.
Example 8
This example provides a cotton potting comprising a suspension of Bacillus velezensis DPT-03 bacteria. The suspension of Bacillus velezensis DPT-03 used was the suspension of Bacillus velezensis DPT-03 prepared in example 3.
The preparation method of the potted plant comprises the following steps:
(1) preparing a cotton pot: uniformly mixing 2kg of soil and 1g of compound fertilizer 15-15-15, placing the mixture in a flowerpot, burying 3 full and healthy cotton grains in the soil, thinning after 15 days, and keeping 1 cotton seedling in each pot;
(2) inoculation of pathogenic bacteria: inoculating the pathogenic bacteria of cotton wilt to the soil of potted cotton after 20 days, wherein the inoculation amount is 20mL, and the concentration is 1.0 multiplied by 108CFU/mL；
(3) Inoculation of the bacterial suspension: inoculating pathogenic bacteria of cotton wilt for 10 days, and inoculating to the solution with concentration of 1.0 × 109Inoculating the suspension of the Bacillus velezensis DPT-03 bacteria of CFU/mL into the soil of the cotton potted plant, wherein the inoculation amount is 20mL per potted plant, and the strain is prepared to comprise the Bacillus velezensis DPT-03 bacteriaAnd (5) potting the suspension.
Example 9
This example provides a cotton potting comprising a suspension of Bacillus velezensis DPT-03 bacteria.
The preparation method of the above potting is different from the preparation method in example 8 in that:
the concentration of the Bacillus velezensis DPT-03 bacterial suspension inoculated in the step (3) is 2.0 multiplied by 109CFU/mL。
Example 10
This example provides a cotton potting comprising a suspension of Bacillus velezensis DPT-03 bacteria.
The preparation method of the above potting is different from the preparation method in example 8 in that:
the concentration of the Bacillus velezensis DPT-03 bacterial suspension inoculated in the step (3) is 4.0 multiplied by 109CFU/mL。
Comparative example 5
The present comparative example provides a cotton potted plant. The preparation method is different from that of the example 8 in that:
the step (3) is as follows: inoculation of sterile water: inoculating the cotton wilt pathogen for 10 days, and inoculating sterile water into soil of the cotton potted plants, wherein the inoculation amount is 20mL per potted plant, so as to obtain the potted plants containing the sterile water.
Test four
The cotton potted plants prepared in example 8, example 9, example 10 and comparative example 5 were used as test subjects, and the disease of each potted plant was examined by setting 30 potted plants in parallel.
Detection method
And respectively inoculating cotton blight pathogenic bacteria to the cotton potted plants for 25 days, detecting the disease conditions of the cotton potted plants, and counting the disease rate and the prevention and treatment effect of the cotton potted plants.
Second, calculating method
The calculation formula of the morbidity and the prevention and treatment effect is the same as the corresponding formula in the second detection test.
Third, the detection result
The results are shown in Table 3.
TABLE 3 detection results of detection test four
As shown in the test results in Table 3, it is understood that the incidence of cotton potting at 25d after inoculation of cotton wilt pathogens by cotton potting reaches 31.5% according to the test results of comparative example 5. On the other hand, the detection results of example 8, example 9 and example 10 show that the incidence rates of cotton potted plants inoculated with the Bacillus velezensis DPT-03 bacterial suspension are respectively 16.2%, 9.0% and 5.4% and the control effects respectively reach 48.57%, 71.42% and 82.85% when the cotton potted plants are inoculated with the cotton wilt pathogen 25 d. According to the results, in the process of cultivating the potted cotton plants, the Bacillus velezensis DPT-03 bacterial suspension provided by the application can effectively reduce the incidence rate of cotton wilt and has a good prevention and control effect on the cotton wilt. And by comparing the detection results of the embodiment 8, the embodiment 9 and the embodiment 10, it can be known that the concentration of the Bacillus velezensis DPT-03 bacterial suspension is increased, and the prevention and treatment effect of the Bacillus velezensis DPT-03 on cotton wilt can be further improved.
Example 11
This example provides a tomato pot culture comprising a suspension of Bacillus velezensis DPT-03 bacteria. The suspension of Bacillus velezensis DPT-03 used was the suspension of Bacillus velezensis DPT-03 prepared in example 3.
The preparation method of the potted plant comprises the following steps:
(1) preparing a tomato pot: sowing tomato seeds in a sterilization matrix, and transplanting peanut seedlings into a pot when 6 true leaves grow out of tomato seedlings; a plastic basin with the diameter of about 15cm is adopted, a nylon net with 20 meshes is padded at the bottom of the basin, and each basin is filled with about 2kg of soil;
(2) inoculation of pathogenic bacteria: inoculating tomato leaf mold pathogenic bacteria into soil of potted tomato plant, wherein the inoculation amount is 20mL, and the concentration is 1.0 multiplied by 108CFU/mL；
(3) Inoculation of the bacterial suspension: inoculating tomato leaf mold pathogenic bacteria for 24hThe concentration is 1.0 × 109And inoculating the CFU/mL bacterial suspension of Bacillus velezensis DPT-03 into soil of potted tomato plants, wherein the inoculation amount is 20mL for each potted plant, and preparing the potted plants comprising the bacterial suspension of Bacillus velezensis DPT-03.
Example 12
This example provides a tomato pot culture comprising a suspension of Bacillus velezensis DPT-03 bacteria.
The preparation method of the above potting is different from the preparation method in example 11 in that:
the concentration of the Bacillus velezensis DPT-03 bacterial suspension inoculated in the step (3) is 2.0 multiplied by 109CFU/mL\。
Example 13
This example provides a tomato pot culture comprising a suspension of Bacillus velezensis DPT-03 bacteria.
The preparation method of the above potting is different from the preparation method in example 11 in that:
the concentration of the Bacillus velezensis DPT-03 bacterial suspension inoculated in the step (3) is 4.0 multiplied by 109CFU/mL\。
Comparative example 6
This comparative example provides a tomato pot. The preparation method is different from that of the example 11 in that:
the step (3) is as follows: inoculation of sterile water: inoculating tomato leaf mold pathogenic bacteria for 24h, and inoculating sterile water into the soil of the tomato pot plant, wherein the inoculation amount is 20mL per pot plant, so as to obtain the pot plant containing the sterile water.
Test five
The tomato pots prepared in example 11, example 12, example 13, and comparative example 6 were used as test subjects, and 10 of each pot was set in parallel to test the incidence of diseases in each pot.
Detection method
And respectively inoculating tomato leaf mold pathogenic bacteria to the tomato pot culture for 30 days, detecting the disease condition of the tomato pot culture, and counting the disease incidence and the control effect of the tomato pot culture.
Second, calculating method
The calculation formula of the morbidity and the prevention and treatment effect is the same as the corresponding formula in the second detection test.
Third, the detection result
The results are shown in Table 4.
TABLE 4 test results of test No. five
As shown in the test results in Table 4, the test results of the comparative example 6 show that the tomato potted plant has an incidence rate of 76.5% when the tomato potted plant is inoculated with the tomato leaf mold pathogenic bacteria for 30 days. On the other hand, the detection results of example 11, example 12 and example 13 show that the incidence rates of tomato potted plants inoculated with the suspension of Bacillus velezensis DPT-03 and inoculated with the tomato leaf mold pathogen 30d are respectively 25.2%, 19.8% and 13.5%, and the control effects respectively reach 67.06%, 74.12% and 74.12%. According to the results, in the process of culturing potted tomatoes, the Bacillus velezensis DPT-03 bacterial suspension provided by the application can effectively reduce the morbidity of the tomato leaf mold and has a good control effect on the tomato leaf mold. And by comparing the detection results of the embodiment 11, the embodiment 12 and the embodiment 13, it can be known that the concentration of the Bacillus velezensis DPT-03 bacterial suspension is increased, and the control effect of the Bacillus velezensis DPT-03 on tomato leaf mold can be further improved.
Example 14
This example provides a tomato pot culture comprising a suspension of Bacillus velezensis DPT-03 bacteria. The suspension of Bacillus velezensis DPT-03 used was the suspension of Bacillus velezensis DPT-03 prepared in example 3.
The preparation method of the potted plant comprises the following steps:
(1) preparing a tomato pot: sowing tomato seeds in a sterilization matrix, and transplanting peanut seedlings into a pot when 6 true leaves grow out of tomato seedlings; a plastic basin with the diameter of about 15cm is adopted, a nylon net with 20 meshes is padded at the bottom of the basin, and each basin is filled with about 2kg of soil;
(2) inoculation of pathogenic bacteria: pathogenic bacteria of tomato gray moldInoculating to soil of potted tomato with an inoculum size of 20mL and a concentration of 1.0 × 108CFU/mL；
(3) Inoculation of the bacterial suspension: inoculating tomato gray mold pathogenic bacteria for 24h, and adding the solution at a concentration of 1.0 × 109And inoculating the CFU/mL bacterial suspension of Bacillus velezensis DPT-03 into soil of potted tomato plants, wherein the inoculation amount is 20mL for each potted plant, and preparing the potted plants comprising the bacterial suspension of Bacillus velezensis DPT-03.
Example 15
This example provides a tomato pot culture comprising a suspension of Bacillus velezensis DPT-03 bacteria.
The preparation method of the above potting is different from the preparation method in example 14 in that:
the concentration of the Bacillus velezensis DPT-03 bacterial suspension inoculated in the step (3) is 2.0 multiplied by 109CFU/mL\。
Example 16
This example provides a tomato pot culture comprising a suspension of Bacillus velezensis DPT-03 bacteria.
The preparation method of the above potting is different from the preparation method in example 14 in that:
the concentration of the Bacillus velezensis DPT-03 bacterial suspension inoculated in the step (3) is 4.0 multiplied by 109CFU/mL\。
Comparative example 7
This comparative example provides a tomato pot. The preparation method is different from that of the example 14 in that:
the step (3) is as follows: inoculation of sterile water: inoculating tomato leaf mold pathogenic bacteria for 24h, and inoculating sterile water into the soil of the tomato pot plant, wherein the inoculation amount is 20mL per pot plant, so as to obtain the pot plant containing the sterile water.
Test six
The tomato pots prepared in example 14, example 15, example 16, and comparative example 7 were used as test subjects, and 10 of each of the pots were set in parallel to test the incidence of diseases in each pot.
Detection method
And respectively inoculating tomato gray mold pathogenic bacteria to the tomato pot culture for 30 days, detecting the disease condition of the tomato pot culture, and counting the disease incidence and the control effect of the tomato pot culture.
Second, calculating method
The calculation formula of the morbidity and the prevention and treatment effect is the same as the corresponding formula in the second detection test.
Third, the detection result
The results are shown in Table 5.
TABLE 5 test results of test six
As shown in the test results in Table 5, the test results of comparative example 7 revealed that the tomato potting rate reached 70.2% when the tomato potting was inoculated with the tomato gray mold pathogen for 30 days. On the other hand, the results of the tests in examples 14, 15 and 16 revealed that the tomato potted plants inoculated with the suspension of Bacillus velezensis DPT-03 and inoculated with the same pathogen 30d for tomato gray mold had incidence rates of 18.9%, 13.5% and 8.1%, respectively, and the control effects of 73.08%, 80.77% and 88.46%, respectively. According to the results, in the process of culturing potted tomatoes, the Bacillus velezensis DPT-03 bacterial suspension provided by the application can effectively reduce the incidence rate of the botrytis cinerea and has a good control effect on the botrytis cinerea. And by comparing the detection results of the embodiment 14, the embodiment 15 and the embodiment 16, it can be known that the concentration of the Bacillus velezensis DPT-03 bacterial suspension is increased, and the control effect of the Bacillus velezensis DPT-03 on the gray mold of tomatoes can be further improved.
Example 17
This example provides a case where a suspension of Bacillus velezensis DPT-03 was used in a pilot field. The suspension of Bacillus velezensis DPT-03 used was the suspension of Bacillus velezensis DPT-03 prepared in example 3.
Test field address: zhang Lou cun, Zheng Yang county, Henan.
Basic conditions of the test field: the test field is a continuous cropping field for peanuts for many years, the land parcel is flat, and the southern blight of the peanuts is serious.
The fertility of the test field: the fertility status of 0-20 cm soil in the test field is as follows: 11.60g/kg of organic matter, 154.39mg/kg of alkaline hydrolysis nitrogen and quick-acting phosphorus (P)2O5)56.62mg/kg, quick-acting potassium (K)2O)168.0mg/kg。
The method for applying the Bacillus velezensis DPT-03 bacterial suspension to the experimental field is as follows:
(1) spraying Bacillus velezensis DPT-03 bacterial suspension into soil at a concentration of 10L/mu by utilizing an agricultural sprayer at one time, wherein the concentration of the Bacillus velezensis DPT-03 bacterial suspension is 1 multiplied by 109cfu/mL, and then plowing the test field;
(2) and (5) sowing peanuts in the test field after ploughing.
Comparative example 8
This comparative example provides a case where the treatment agent was used in the test field.
This comparative example differs from example 17 in that a treatment agent was used instead of the suspension of Bacillus velezensis DPT-03 in example 17. The treatment agent is thifluzamide pesticide. Wherein the thifluzamide pesticide is 240g/L thifluzamide suspending agent, the dosage per mu is 100ml, and the thifluzamide suspending agent is diluted by 100 times for use.
Comparative example 9
This comparative example provides a case where sterile water was used in the test field.
This comparative example differs from example 17 in that sterile water is used instead of the suspension of Bacillus velezensis DPT-03 in example 17.
Seven detection test
The cases provided in example 17, comparative example 8 and comparative example 9 were used as the test subjects. And detecting the field control effect of the pathogens of the southern blight in each case.
Detection method
Randomly selecting groups in the test field, selecting 3 groups in each case, wherein each group has an area of 180m2And counting the morbidity, disease index and prevention and treatment effect of each case.
Second, calculating method
The calculation formula of the morbidity and the prevention and treatment effect is the same as the corresponding formula in the second detection test.
Third, the detection result
The results are shown in Table 6.
TABLE 6 detection results of detection test seven
| Detecting an object | Investigation of plant number (strain) | Number of affected plants | Incidence (%) | Control effect (%) |
| Example 17 | 300 | 33b | 11.00b | 63.61 |
| Comparative example 8 | 300 | 37b | 12.33b | 62.21 |
| Comparative example 9 | 300 | 80a | 26.67a | - |
As shown in the test results in Table 6, the incidence of peanut plants in the test field inoculated with sterile water was 26.67% according to the test result of comparative example 9, while the incidence of peanut plants in the test field inoculated with Bacillus velezensis DPT-03 suspension and the treatment agent was 11.00% and 12.33% respectively and the control effect was 63.61% and 62.21% respectively according to the test results of example 17 and comparative example 8. According to the results, the Bacillus velezensis DPT-03 bacterial suspension can reduce the incidence rate and disease index of the southern blight of the peanuts in the test field and improve the control effect of the southern blight of the peanuts in the test field. Meanwhile, the seasonal prevention and control effect of the Bacillus velezensis DPT-03 bacterial suspension on the southern blight of the peanuts in the test field is higher than the prevention and control effect of the treatment agent on the southern blight of the peanuts in the test field, and the Bacillus velezensis DPT-03 bacterial suspension on the southern blight of the peanuts in the test field has higher application potential and safety on the continuity and safety of biological prevention and control.
The present embodiment is only for explaining the present application, and it is not limited to the present application, and those skilled in the art can make modifications of the present embodiment without inventive contribution as needed after reading the present specification, but all of them are protected by patent law within the scope of the claims of the present application.
Sequence listing
<110> Beijing century Ames bioengineering, Inc
Beijing Century Amms Biotechnology Co.,Ltd.
<120> Bacillus belgii and application thereof
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1514
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 1
agagtttgat cctggctcag gacgaacgct ggcggcgtgc ctaatacatg caagtcgagc 60
ggacagatgg gagcttgctc cctgatgtta gcggcggacg ggtgagtaac acgtgggtaa 120
cctgcctgta agactgggat aactccggga aaccggggct aataccggat ggttgtttga 180
accgcatggt tcagacataa aaggtggctt cggctaccac ttacagatgg acccgcggcg 240
cattagctag ttggtgaggt aacggctcgc caaggcgacg atgcgtagcc gacctgagag 300
ggtgatcggc cacactggga ctgagacacg gcccagactc ctacgggagg cagcagtagg 360
gaatcttccg caatggacga aagtctgacg gagcaacgcc gcgtgagtga tgaaggtttt 420
cggatcgtaa agctctgttg ttagggaaga acaagtgccg ttcaaatagg gcggcacctt 480
gacggtacct aaccagaaag ccacggctaa ctacgtgcca gcagccgcgg taatacgtag 540
gtggcaagcg ttgtccggaa ttattgggcg taaagggctc gcaggcggtt tcttaagtct 600
gatgtgaaag cccccggctc aaccggggag ggtcattgga aactggggaa cttgagtgca 660
gaagaggaga gtggaattcc acgtgtagcg gtgaaatgcg tagagatgtg gaggaacacc 720
agtggcgaag gcgactctct ggtctgtaac tgacgctgag gagcgaaagc gtggggagcg 780
aacaggatta gataccctgg tagtccacgc cgtaaacgat gagtgctaag tgttaggggg 840
tttccgcccc ttagtgctgc agctaacgca ttaagcactc cgcctgggga gtacggtcgc 900
aagactgaaa ctcaaaggaa ttgacggggg cccgcacaag cggtggagca tgtggtttaa 960
ttcgaagcaa cgcgaagaac cttaccaggt cttgacatcc tctgacaatc ctggagatag 1020
gacgtcccct tcgggggcag agtgacaggt ggtgcatggt tgtcgtcagc tcgtgtcgtg 1080
agatgttggg ttaagtcccg caacgagcgc aacccttgat cttagttgcc agcattcagt 1140
tgggcactct aaggtgactg ccggtgacaa accggaggaa ggtggggatg acgtcaaatc 1200
atcatgcccc ttatgacctg ggctacacac gtgctacaat ggacagaaca aagggcagcg 1260
aaaccgcgag gttaagccaa tcccacaaat ctgttctcag ttcggatcgc agtctgcaac 1320
tcgactgcgt gaagctggaa tcgctagtaa tcgcggatca gcatgccgcg gtgaatacgt 1380
tcccgggcct tgtacacacc gcccgtcaca ccacgagagt ttgtaacacc cgaagtcggt 1440
gaggtaacct tttaggagcc agccgccgaa ggtgggacag atgattgggg tgaagtcgta 1500
acaaggtagc cgta 1514
Claims (10)
1. Bacillus belgii (B.), (B.), (B.beijerinckii)Bacillus velezensis) The preservation number is CGMCC NO. 20317.
2. A Bacillus belgii bacterium according to claim 1 (B: (B))Bacillus velezensis) The method is characterized in that: the Bacillus belgii has dry wrinkles, opaqueness, milk white color, irregular or nearly circular edges and micro-protrusions on the colony surface on an LB solid culture medium.
3. A culture comprising the Bacillus belgii of claim 1 or a processed product thereof.
4. A bacterial suspension prepared using the Bacillus belgii of claim 1.
5. A sterile fermentation broth prepared using the Bacillus belgii of claim 1 or the bacterial suspension of claim 5.
6. A microbial inoculant characterized by: the microbial agent comprises Bacillus belgii of claim 1 and/or the culture or processed product of claim 3.
7. A microbial inoculant characterized by: the microbial agent comprises the Bacillus belgii of claim 1, the bacterial suspension of claim 4, and/or the sterile fermentation broth of claim 5.
8. A microbial fertilizer is characterized in that: the microbial fertilizer comprises the Bacillus belgii of claim 1, the microbial agent of claim 6, or the microbial agent of claim 7.
9. Use of a bacillus beijerinckii according to claim 1, a culture or a processed product according to claim 3, a microbial agent according to claim 6 or 7, or a microbial fertilizer according to claim 8 for the preparation of a composition for the amelioration and/or prevention of a disease in a crop.
10. Use according to claim 9, characterized in that: the crop diseases for improving and/or preventing the composition are one or more of southern blight of peanuts, small spot of corns, blight of cotton, leaf mold of tomatoes, gray mold of tomatoes, tobacco brown spot, colletotrichum gloeosporioides of vegetables and flowers and gray mold of grapes.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111166440.0A CN113755389B (en) | 2021-09-30 | 2021-09-30 | Bacillus bailii and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111166440.0A CN113755389B (en) | 2021-09-30 | 2021-09-30 | Bacillus bailii and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN113755389A true CN113755389A (en) | 2021-12-07 |
| CN113755389B CN113755389B (en) | 2023-05-02 |
Family
ID=78798729
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202111166440.0A Active CN113755389B (en) | 2021-09-30 | 2021-09-30 | Bacillus bailii and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113755389B (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114703098A (en) * | 2022-04-06 | 2022-07-05 | 陕西嘉益蓝德生物工程有限公司 | Fermentation preparation and application of Bacillus belgii fungicide for preventing and treating cotton wilt |
| CN114990020A (en) * | 2022-06-17 | 2022-09-02 | 湖北省农业科学院中药材研究所 | Biocontrol strain and application thereof |
| CN115975848A (en) * | 2022-07-30 | 2023-04-18 | 北京世纪阿姆斯生物工程有限公司 | Tropical bacillus and application thereof |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| 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 |
| CN108265012A (en) * | 2016-12-30 | 2018-07-10 | 北京绿色农华作物科技有限公司 | A kind of Bei Laisi Bacillus strains and its microbial inoculum and application |
| CN109266586A (en) * | 2018-10-18 | 2019-01-25 | 淮海工学院 | Bei Laisi bacillus BMF 03 and application thereof and fermentation process |
| CN110283742A (en) * | 2019-06-17 | 2019-09-27 | 北京农业生物技术研究中心 | The Bei Laisi bacillus of one plant of broad-spectrum disease resistance and its application |
| CN112195142A (en) * | 2020-11-17 | 2021-01-08 | 山东省花生研究所 | Biocontrol bacillus beleisi ZHX-7 and application thereof |
| CN112458012A (en) * | 2020-11-24 | 2021-03-09 | 新疆农业科学院微生物应用研究所（中国新疆-亚美尼亚生物工程研究开发中心） | Bacillus belgii microbial agent and application thereof |
-
2021
- 2021-09-30 CN CN202111166440.0A patent/CN113755389B/en active Active
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108265012A (en) * | 2016-12-30 | 2018-07-10 | 北京绿色农华作物科技有限公司 | A kind of Bei Laisi Bacillus strains and its microbial inoculum and 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 |
| CN109266586A (en) * | 2018-10-18 | 2019-01-25 | 淮海工学院 | Bei Laisi bacillus BMF 03 and application thereof and fermentation process |
| CN110283742A (en) * | 2019-06-17 | 2019-09-27 | 北京农业生物技术研究中心 | The Bei Laisi bacillus of one plant of broad-spectrum disease resistance and its application |
| CN112195142A (en) * | 2020-11-17 | 2021-01-08 | 山东省花生研究所 | Biocontrol bacillus beleisi ZHX-7 and application thereof |
| CN112458012A (en) * | 2020-11-24 | 2021-03-09 | 新疆农业科学院微生物应用研究所（中国新疆-亚美尼亚生物工程研究开发中心） | Bacillus belgii microbial agent and application thereof |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114703098A (en) * | 2022-04-06 | 2022-07-05 | 陕西嘉益蓝德生物工程有限公司 | Fermentation preparation and application of Bacillus belgii fungicide for preventing and treating cotton wilt |
| CN114990020A (en) * | 2022-06-17 | 2022-09-02 | 湖北省农业科学院中药材研究所 | Biocontrol strain and application thereof |
| CN114990020B (en) * | 2022-06-17 | 2023-01-31 | 湖北省农业科学院中药材研究所 | Biocontrol strain and application thereof |
| CN115975848A (en) * | 2022-07-30 | 2023-04-18 | 北京世纪阿姆斯生物工程有限公司 | Tropical bacillus and application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN113755389B (en) | 2023-05-02 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN113151062B (en) | Bacillus belgii LJBV19 and application thereof | |
| CN101760438B (en) | Broad-spectrum antifungal plant endophytic bacillus subtillis and application thereof | |
| CN105441366B (en) | Methylotrophic bacillus ZBL-1 is applied in preventing cotton verticillium wilt | |
| CN113755389B (en) | Bacillus bailii and application thereof | |
| CN113652382B (en) | Bacillus bailii for preventing and treating peanut southern blight, microbial agent, microbial fertilizer and application of microbial fertilizer | |
| CN113717901A (en) | Bacillus belgii and application thereof in prevention and treatment of various vegetable diseases | |
| CN110129240B (en) | Bacillus amyloliquefaciens and application thereof in preventing and treating celery soft rot | |
| CN108148794A (en) | A kind of the bacillus subtilis DYr3.3 and preparation method and application of broad-spectrum antibacterial activity | |
| CN110317747B (en) | Bacillus amyloliquefaciens JT68 and application thereof in prevention and treatment of tea anthracnose | |
| CN106701631B (en) | Streptomyces roseoflavus and application thereof | |
| CN113444651B (en) | Saffron endophytic fungus and application thereof in preventing and treating bulb rot | |
| CN115261283A (en) | Bacillus cereus and application thereof in prevention and control of dry farming potato diseases | |
| CN111040976A (en) | Bacillus amyloliquefaciens and application thereof | |
| CN116855407A (en) | Streptomyces ka Wu Ershi Y274 and application thereof | |
| CN108641989A (en) | One plant of Methylotrophic bacillus and its application | |
| CN105567600A (en) | Pathogen verticillium antagonistic bacterium and application thereof | |
| CN108220211B (en) | Acinetobacter oleophilic NMB17 and application thereof in plant disease control | |
| CN114806960A (en) | Bacillus cereus YX53 and application thereof in preventing and treating fusarium root rot of tobacco and promoting growth | |
| CN114806928A (en) | Pepper endophytic Bacillus beleisi PEB23 and application thereof | |
| CN111778174B (en) | Bacillus subtilis with inhibiting effect on citrus sand skin disease and screening method thereof | |
| CN109182216B (en) | Marine streptomyces SCFJ-05 with inhibition effect on succulent plant stem rot | |
| CN115960777B (en) | Bacillus pseudomycoides and application thereof in prevention and treatment of vegetable epidemic disease | |
| CN117384801A (en) | Paenibacillus polymyxa J-04-06 and application thereof | |
| CN110982764A (en) | Bacillus tequilensis S12 for preventing and treating rice blast and application thereof | |
| CN114933980B (en) | Streptomyces shallowus HJB-XTBG45 for preventing and treating rhizoma polygonati root rot and application thereof |
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 |