CN109868250B - Bacillus belgii for inhibiting viruses and promoting plant growth and application thereof - Google Patents
Bacillus belgii for inhibiting viruses and promoting plant growth and application thereof Download PDFInfo
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Abstract
The Bacillus belief propagation strain is characterized in that the strain is named as Bacillus belief propagation (SNB) 55 and the application of the strain in preventing and treating plant virus diseases and plant fungal diseases and promoting plant growth. The invention has the beneficial effects that: the bacillus belief in inhibiting viruses and promoting plant growth and the application thereof have the advantages of no toxicity, no pathogenicity, safety to people and livestock, environmental friendliness, strong stress resistance and stable bacteriostatic activity; can inhibit bacillus cereus, tobacco mosaic virus and tomato yellow leaf curl virus, cucumber fusarium wilt, tobacco target spot pathogen, phytophthora capsici, rice sheath blight, tomato botrytis cinerea and other plant pathogenic bacteria, can secrete indoleacetic acid, promotes seed germination and plant growth, is a multifunctional biocontrol bacterium, and has wide application prospect.
Description
Technical Field
The invention belongs to the technical field of microbial prevention and treatment, relates to a microbial prevention and treatment technology for Tomato virus diseases, and particularly relates to a Bacillus subtilis (Bacillus velezensis) for inhibiting Tobacco Mosaic Virus (TMV) and Tomato Yellow Leaf Curl Virus (TYLCV) and application thereof.
Background
Plant viral diseases, known as "plant cancers," are the second largest plant disease after fungal plant disease, causing significant economic losses to agricultural production. Plant viruses are various, and among them, Tobacco Mosaic Virus (TMV) is one of the major toxic sources causing solanaceous plants such as Tomato and tobacco. In recent years, Tomato Yellow Leaf Curl Viruses (TYLCV) are rapidly spread in China and tend to increase year by year, and pose a serious threat to Tomato production in China. For a long time, the prevention and treatment of plant virus diseases mainly comprises chemical prevention and treatment and cultivation of antiviral varieties, in the aspect of chemical agent prevention and treatment, the number of currently satisfactory practical varieties of plant virus resisting agents is small, particularly, the number of specific therapeutic agents is small, the reported agents are not ideal in field practical application prevention effect, the chemical prevention and treatment also has many problems, and a large amount of chemical pesticides are applied to cause the problems of pesticide residue, food safety, ecological environment pollution and the like. Therefore, the cultivation of antiviral varieties is considered to be an economical and effective way for preventing and treating tobacco mosaic virus. However, limited resistant materials with commercial value greatly limit the progress of breeding work, and no high-quality and high-resistance variety can be popularized in a large area at present. With the aggravation of the harm of plant virus diseases, the long-term use of a large amount of chemical pesticides, the problems of environmental pollution, pesticide residues and the like, the development of a new, efficient and safe microbial preparation for controlling the plant virus diseases becomes the need of agricultural safety and sustainable development.
Antiviral active substances generated by microbial metabolism have the advantages of strong activity, safety and high efficiency, and antiviral substances such as polysaccharide, nucleic acid, protein, micromolecule and the like obtained by screening and separating from microbial resources become hot spots of current research. Some products are formed and play an important role in agricultural production, wherein lentinan is successfully registered as an antiviral agent in China and is industrially produced. Especially, the development of antiviral agricultural antibiotics has made breakthrough progress, and ningnanmycin and pyriminomycin are the antiviral agricultural antibiotics which are the most successful in research. The ningnanmycin is separated from streptomyces noursensis Sichang variety, is a novel cytosine nucleoside antibiotic, has the advantages of broad-spectrum and high-efficiency inhibition, low toxicity, low residue and the like, is widely applied to agricultural production, and has the prevention effect on TMV infection of 58.1 percent at the concentration of 500 mug/mL. It has been reported that ningnanmycin induces increased anti-TMV activity in plants. The pyriminomycin isolated from streptomyces hygroscopicus shows potent anti-plant viral activity and has been successfully registered and industrialized in china as an anti-plant viral agent. Most other anti-plant virus active substances and biocontrol strains are in the laboratory research stage, so that novel microbial pesticides for preventing and treating plant virus diseases are still lacking in production.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides the Bacillus belief which is nontoxic, non-pathogenic, safe to human and livestock, environment-friendly, strong in stress resistance, stable in bacteriostatic activity, capable of inhibiting plant viruses and promoting plant growth, and is applied to the aspects of preventing plant virus diseases and plant fungal diseases, promoting plant growth and the like, so that the aims of efficiently inhibiting the viruses and promoting the plant growth without damaging the natural environment are fulfilled, and sustainable and harmonious development is further realized.
The name of a strain of Bacillus belgii for inhibiting viruses and promoting plant growth is Bacillus belgii (Bacillus velezensis) SNB55, the preservation unit is China general microbiological culture Collection center, the preservation number is CGMCC NO.17315, and the preservation date is 2019, 3 and 8 days. And (4) storage address: the institute of microbiology, national academy of sciences No. 3, Xilu No.1, Beijing, Chaoyang, Beijing.
As a preferable scheme, the strain culture of the Bacillus beilaisi SNB55 for inhibiting virus and promoting plant growth is characterized in that bacterial colonies on a beef extract peptone agar plate are white to milky, nearly circular, rough, irregular in edges, opaque, wrinkled in surface, sticky in texture and textured; after the liquid is cultured, a white film can be formed on the surface of the liquid by standing, and the liquid is uniform in turbidity and does not cake.
More preferably, the strain of Bacillus beilis SNB55 for inhibiting virus and promoting plant growth is characterized in that the strain has rod shape, size of (0.5-0.6) Mumx (1.5-3.2) Mum, short chain or moniliform arrangement, flagella and motion ability; gram positive staining; the spore is oval, the mesogenic cyst and the cyst are expanded, and no parasporal crystal exists.
More preferably, the optimum growth temperature of the bacillus beijerinckii SNB55 for inhibiting viruses and promoting plant growth is 26-28 ℃.
More preferably, the fermentation product of the bacillus beijerinckii SNB55 capable of inhibiting viruses and promoting plant growth can inhibit plant viruses and plant pathogenic bacteria; the plant virus and plant pathogenic bacteria include tobacco mosaic virus, tomato yellow leaf curl virus, Bacillus cereus, cucumber fusarium wilt, tobacco target spot pathogen, phytophthora capsici, rice sheath blight and tomato gray mold.
More preferably, the bacillus beijerinckii SNB55 capable of inhibiting viruses and promoting plant growth can secrete indoleacetic acid to promote seed germination and plant growth.
More preferably, the bacillus beijerinckii SNB55 for inhibiting viruses and promoting plant growth and the fermentation product thereof are nontoxic, non-pathogenic, safe to human and livestock, environment-friendly, strong in stress resistance and stable in bacteriostatic activity.
The application of the Bacillus beijerinckii SNB55 for inhibiting viruses and promoting plant growth in preventing and treating plant virus diseases and plant fungal diseases.
As a preferable scheme, the Bacillus beijerinckii SNB55 for inhibiting virus and promoting plant growth is applied to promoting plant growth.
More preferably, the bacillus beijerinckii SNB55 for inhibiting virus and promoting plant growth is applied to control tobacco mosaic virus disease, tomato yellow leaf curl virus disease, cucumber fusarium wilt, tobacco target spot, pepper phytophthora blight, rice sheath blight and tomato gray mold.
The invention has the beneficial effects that: the bacillus beiLeisi capable of inhibiting virus and promoting plant growth and the application thereof provided by the invention have the following advantages:
the Bacillus beijerinckii SNB55 for inhibiting viruses and promoting plant growth and the fermentation product thereof are nontoxic and non-pathogenic, are safe to people and livestock, environment-friendly, strong in stress resistance and stable in bacteriostatic activity, are multifunctional biocontrol bacteria, and have wide application prospects.
Secondly, the Bacillus belgii SNB55 for inhibiting viruses and promoting plant growth and the fermentation product thereof can inhibit plant viruses and plant pathogenic bacteria in a broad spectrum, wherein the plant viruses and the plant pathogenic bacteria comprise tobacco mosaic virus, tomato yellow leaf curl virus, Bacillus cereus, cucumber fusarium wilt, tobacco target spot pathogen, phytophthora capsici, rice sheath blight and tomato botrytis cinerea. Especially has stronger inhibiting effect on tobacco mosaic virus and tomato yellow leaf curl virus, the fermentation liquor is prepared into a biological water repellent, and the tomato virus disease caused by the tobacco mosaic virus and the tomato yellow leaf curl virus can be well prevented and treated by spraying, root irrigation and seed coating in the seedling stage.
And thirdly, the Bacillus belgii SNB55 for inhibiting viruses and promoting plant growth can secrete indoleacetic acid (IAA) and promote tomato seed germination, rooting and seedling growth.
The Bacillus beijerinckii SNB55 for inhibiting virus and promoting plant growth increases the variety of microbial antiviral agents for preventing and treating plant virus diseases.
Drawings
FIG. 1 is a colony morphology of a strain of Bacillus beijerinckii SNB55 for inhibiting viruses and promoting plant growth according to the present invention;
FIG. 2 is a plot of the zone of inhibition of Bacillus cereus by a strain of Bacillus belgii SNB55 fermentation broth and fermentation filtrate for inhibiting viruses and promoting plant growth according to the present invention;
FIG. 3 is a control group of cigarette heart-leaves inoculated with TMV buffer solution in a test for TMV inhibition on cigarette heart-leaves measured by a pot culture test in the example of the present invention;
FIG. 4 is a pot test assay of the inhibitory effect of fermentation filtrates on TMV on cigarette leaf tobacco according to the example of the present invention, in which SNB55 fermentation filtrate was mixed with 2-fold TMV buffer and inoculated into cigarette leaf tobacco;
FIG. 5 shows the rooting of the seeds of control group 2 in the test of the effect of the fermentation filtrate on the growth of tomato seeds and seedlings according to the example of the present invention;
FIG. 6 shows the rooting of the seeds of experiment group 2 in the test of the effect of the fermentation filtrate on the growth of tomato seeds and seedlings according to the example of the present invention;
FIG. 7 shows the growth of seedlings in control group 3 in the test of the effect of fermentation filtrate on the growth of tomato seeds and seedlings in the example of the present invention;
FIG. 8 shows the growth of seedlings in experiment 3 in the test of the effect of fermentation filtrate on the growth of tomato seeds and seedlings in the example of the present invention.
Reference numerals: 1-fermentation liquor bacteriostasis zone, and 2-fermentation filtrate bacteriostasis zone.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are some, but not all, embodiments of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
The Bacillus belgii strain has the bacterial strain name of Bacillus belgii (Bacillus velezensis) SNB55, the preservation unit is China general microbiological culture Collection center, the preservation number is CGMCC NO.17315, and the preservation date is 3, 8 and 2019. And (4) storage address: the institute of microbiology, national academy of sciences No. 3, Xilu No.1, Beijing, Chaoyang, Beijing.
1. Culturing of bacterial strains
(1) Composition of culture medium
The plate and slant culture medium is beef extract peptone agar culture medium: 3g of beef extract, 5g of peptone, 5g of sodium chloride, 20g of agar, and adding distilled water to 1000mL, wherein the pH value is 7.0-7.2.
Fermentation medium: 3g of beef extract, 5g of peptone and 5g of sodium chloride, and adding distilled water to 1000mL, wherein the pH value is 7.0-7.2.
Bacillus cereus culture medium: beef extract 3g, peptone 3g, Na2HPO40.1g of agar, 18-20 g of agar, and distilled water till the volume is 1000mL, wherein the pH value is 7.8-8.0.
(2) Bacillus belgii SNB55 culture and fermentation filtrate preparation
Slant culture: transferring the strain SNB55 to a fresh slant culture medium, and culturing for 1d at 28 ℃ to obtain mature slant spores;
fermentation culture: under the aseptic condition, inoculating the mature slant spores with the inoculation amount of 5% into a 250mL triangular flask filled with 50mL fermentation medium, and carrying out shake culture on a rotary shaking table at the temperature of 28 ℃ and the rotating speed of 150r/min for 46h to obtain fermentation liquor;
preparing sterile fermentation filtrate: centrifuging the fermentation broth at 10000r/min for 20min, collecting supernatant, and purifying with
Filtering the mixture by using the bacterial filter to obtain sterile fermentation filtrate.
2. Strain detection
(1) Detection item and method
1) And (3) detecting colony morphology: the strain SNB55 was streaked out in zones on a plate medium, cultured at 28 ℃ for 5 days, and observed for single colony morphology.
2) The strain SNB55 inhibits the activity detection of Bacillus cereus: will contain 109Mixing 700 mu L of cfu/mL bacillus cereus suspension and 70mL of melted bacillus cereus culture medium, pouring the mixture into a square plate to prepare a flat plate, adding 200 mu L of the fermentation liquid into one Oxford cup, adding 200 mu L of the sterile fermentation filtrate into the other Oxford cup, culturing at 28 ℃ for 16h, and measuring the diameter of a bacteriostatic circle.
3) Pot culture assay to determine the inhibitory effect of fermentation filtrate on TMV on snuff: the inhibition effect of the fermentation filtrate on the TMV is measured by artificially inoculating the TMV on the potted healthy heart-leaf tobacco by adopting a juice friction inoculation method, the experiment is divided into 4 experiment groups and 1 control group, each group contains 30 plants and is repeated for 3 times, and the specific treatment method is shown in the following table 1. After inoculation for 5d, the number of dead spots is investigated, and the inhibition rate of the lesion spots is calculated.
TABLE 1 potted plant test determination of inhibition of fermentation filtrate on tobacco leaf on TMV experiment grouping treatment method
4) Pot culture assay to determine the inhibition of TMV by fermentation filtrate on tomatoes: the tomato variety is L402, a juice friction inoculation method is adopted, TMV is artificially inoculated to potted healthy tomato leaves to measure the inhibitory activity of fermentation filtrate on the TMV, the experiment is divided into 3 experiment groups and 1 control group, each group contains 30 plants and is repeated for 3 times, and the specific treatment method is shown in the following table 2. And respectively recording the tomato morbidity according to a grading standard, calculating the control effect, and carrying out crop safety investigation after application.
TABLE 2 potted plant test determination of TMV inhibition of fermentation filtrates on tomatoes
Grading standard: level 0: no symptoms are caused; level 1: the veins of heart and leaves are the bright veins, and 1-2 true leaves present flower and leaves; and 3, level: middle upper leaf; and 5, stage: most leaf and leaf, few leaf malformation; and 7, stage: most leaves are heavy leaves, deformed and shriveled; and 9, stage: almost all leaves are heavy leaves, deformed and shriveled, and the plant dwarfing is obvious.
5) The pot experiment determines the prevention effect of the fermentation filtrate on the tomato mosaic virus disease: the tomato variety is L402, 5 groups are provided, and 3 experimental groups and 2 control groups are included. Experimental group 1: spraying 20 times diluted fermentation filtrate at 3d for temporary planting of tomato, and spraying for 3 times at 5d intervals; experimental group 2: soaking the seeds for 5 hours by using fermentation filtrate diluted by 20 times, and normally planting; experimental group 3: after the tomatoes are temporarily planted for 3 days, irrigating roots with 20-time diluted fermentation filtrate, 10mL of each plant, and irrigating for 1 time at intervals of 5 days for 3 times; control group 1: spraying clear water; control group 2: spraying amino-oligosaccharin (sea island essence). Adopting a sap rubbing inoculation method, rubbing and inoculating the TMV at the true leaf stage of 5 tomato slices, investigating the disease occurrence condition of the plant 20d after inoculation according to the grading standard of the TMV disease symptoms of the tomato, and calculating each treatment disease index and the prevention and treatment effect.
Tomato TMV disease symptom grading criteria: level 0: no symptoms are caused; level 1: the veins of heart and leaves are the bright veins, and 1-2 true leaves present flower and leaves; and 3, level: middle upper leaf; and 5, stage: most leaf and leaf, few leaf malformation; and 7, stage: most leaves are heavy leaves, deformed and shriveled; and 9, stage: almost all leaves are heavy leaves, deformed and shriveled, and the plant dwarfing is obvious.
6) The potted plant test determines the prevention effect of the fermentation filtrate on tomato yellow leaf curl virus diseases: the tomato variety is L402, and 3 groups are provided, including 1 experimental group and 2 control groups. Experimental group 1: spraying 20 times diluted fermentation filtrate at 3d after temporary planting of tomato, and spraying for 3 times at an interval of 5 d; control group 1: spraying clear water; control group 2: spraying amino-oligosaccharin (sea island essence). And (3) after the 3 rd application, uniformly releasing the whiteflies with the tomato yellow leaf curl viruses fed in the poison source chamber onto test tomato plants for virus transmission, ensuring that more than 10 heads of each tomato plant are transmitted, investigating the disease incidence of the diseased plants 15d after the disease according to the tomato yellow leaf curl virus disease symptom grading standard, and calculating the disease incidence, disease index and prevention and treatment effect.
TABLE 3 Classification criteria for tomato yellow leaf curl virus symptoms
7) The field plot experiment determines the prevention effect of the fermentation filtrate on tomato yellow leaf curl virus diseases: the tomato variety is L402, and the insect-proof net barrier is arranged, 5 groups are arranged in total, the tomato variety comprises 3 experimental groups and 2 control groups, and each group has 3 repetitions. Experimental group 1: spraying 20 times diluted fermentation filtrate after planting tomatoes for 3 days, and spraying for 3 times at an interval of 5 days; experimental group 2: soaking the seeds for 5 hours by using fermentation filtrate diluted by 20 times, and normally cultivating; experimental group 3: irrigating roots of tomatoes with 20-time diluted fermentation filtrate after planting for 3 days, wherein each plant is 10mL, irrigating for 1 time at intervals of 5 days, and irrigating for 3 times; control group 1: spraying clear water; control group 2: spraying amino-oligosaccharin (sea island essence). And (3) after 10 days of treatment, the whiteflies with TYLCV are released into the shed, no insecticide is sprayed in the whole process, and after 15 days of disease occurrence, the disease index and the prevention and treatment effect are calculated according to the grading standard of tomato yellow leaf curl virus symptoms.
8) And (3) detecting the influence of the fermentation filtrate on the growth of tomato seeds and seedlings: the experiment is provided with 6 groups comprising 3 experimental groups and 3 control groups, and each group has 3 replicates; wherein the experimental group 1 and the experimental group 2, the control group 1 and the control group 2 are used for detecting the influence on the growth of the seeds; the experimental group 3 and the control group 3 were used for the detection of the effect on the growth of seedlings.
Detection of influence on seed growth
Experimental group 1: soaking tomato seeds in the fermentation filtrate for 5 hours, then putting the seeds into a sterile culture dish paved with wet filter paper, carrying out heat preservation culture at 28 ℃, and observing the germination condition of the seeds; experimental group 2: soaking tomato seeds in the fermentation filtrate diluted by 20 times for 5 hours, then putting the seeds into a sterile culture dish paved with wet filter paper, carrying out heat preservation culture at 28 ℃, and observing the germination condition of the seeds; control group 1: soaking tomato seeds in the fermentation medium for 5 hours, then putting the seeds into a sterile culture dish paved with wet filter paper, carrying out heat preservation culture at 28 ℃, and observing the germination condition of the seeds; control group 2: and (3) soaking the tomato seeds in the fermentation culture medium diluted by 20 times for 5 hours, then putting the seeds into a sterile culture dish paved with wet filter paper, carrying out heat preservation culture at 28 ℃, and observing the germination condition of the seeds.
② detection of influence on growth of seedling
Experimental group 3: soaking tomato seeds in fermentation filtrate diluted by 20 times for 5 hours, accelerating germination in a constant temperature box at 28 ℃, sowing the seeds in a culture pot after the seeds germinate, and placing the seeds in a greenhouse for normal management; control group 3: and (3) soaking the tomato seeds in a fermentation medium diluted by 20 times for 5 hours, accelerating germination in a constant temperature box at 28 ℃, sowing the seeds in a culture pot after the seeds germinate, and placing the seeds in a greenhouse for normal management. Pulling out seedlings after one month, cleaning with clear water, drying with absorbent paper, and measuring plant height, root, fresh weight of overground part and fresh weight of underground part; weighing fresh weight, deactivating enzyme in a ventilation drying oven at 105 deg.C for 30min, drying at 75 deg.C to constant weight, and weighing the dry weight of aerial part and underground part.
9) Broad spectrum bacteriostasis test of SNB 55: taking a rice sheath blight bacterium, cucumber wilt bacterium, tobacco leaf blight bacterium and phytophthora capsici dish with the diameter of 5mm, reversely buckling the center of a PDA (personal digital assistant) flat plate, marking two parallel straight lines at a position about 2cm away from the upper part and the lower part of the plate by using an inoculating loop stained with Bacillus beleisis SNB55, culturing for 3d at 28 ℃, and observing the bacteriostasis condition.
10) Determination of SNB55 secretion of Indole Acetic Acid (IAA): preparation of group samples Bacillus beilesiensis SNB55 was inoculated into a sample containing L-tryptophan (200 mg. L.)-1) The LB liquid medium of (5) was subjected to shaking culture at 28 ℃ and 180rpm for 4 d. 50 μ L of the bacterial suspension was dropped on a white ceramic plate, and 50 μ L of a Salkowski colorimetric solution (50mL of 35% HCLO) was added simultaneously4+1mL 0.5M FeCL3)(Libbert&Risch, 1969). 50 μ L (50 mg. L) of IAA was added-1Shanghai biological engineering Co., Ltd., purity > 99%) as a control group. The white ceramic plates of the experimental group and the control group are placed at room temperature in the dark for 30min and observed, and the IAA can be secreted when the color of the white ceramic plates turns red.
(2) The result of the detection
1) And (3) colony morphology detection results: bacterial colonies of the strain SNB55 on the plate culture medium are white to milky, nearly circular, rough, irregular in edge, opaque, wrinkled in surface and sticky in texture, and are shown in figure 1; after liquid culture, standing to form a white film on the liquid surface, wherein the liquid is turbid and uniform and does not cake, and the strain is characterized by being rod-shaped, with the size of (0.5-0.6) Mumx (1.5-3.2) Mum, polymerized together and arranged in short chains or moniliforme shape, and provided with flagella and capable of moving; gram positive staining; the spore is oval, the mesogenic cyst and the cyst are expanded, and no parasporal crystal exists.
2) The result of the detection of the activity of the strain SNB55 for inhibiting the bacillus cereus is as follows: the fermentation liquor and the sterile fermentation filtrate of the strain SNB55 have strong inhibition effect on the bacillus cereus, the specific results are shown in Table 4, and the inhibition zone condition is shown in FIG. 2.
TABLE 4 inhibitory Effect of the SNB55 Strain fermentation broth and the sterile fermentation filtrate on Bacillus cereus
As can be seen from the table, the diameter of the treated inhibition zone of the fermentation liquor reaches 22mm, the inhibition zone is clear and transparent, and the inhibition zone is still clear and transparent and has no change in size after being observed after being placed at room temperature for 15 days, which indicates that the fermentation liquor of the strain SNB55 has strong inhibition effect on the bacillus cereus and has lasting inhibition effect. The diameters of the bacteriostatic circle of the fermentation liquor and the sterile fermentation filtrate on the bacillus cereus have no obvious difference, which indicates that the bacteriostatic active substance is mainly a fermentation product of the strain SNB 55.
3) Pot culture test determination of inhibition of fermentation filtrate on cigarette snuff on TMV test results: the strain SNB55 fermentation filtrate can inhibit infection of TMV and reduce the number of scorched spots on the heart-leaf tobacco by various treatments, the specific results are shown in Table 5, and the passivation effect is shown in figure 3 and figure 4.
TABLE 5 inhibitory Effect of the Strain SNB55 fermentation filtrate on TMV on cigarette Heart tobacco
As can be seen from Table 5, the treatment of mixing the fermentation filtrate with TMV has the strongest passivation effect on TMV, and the inhibition rate of scorched spots is as high as 94.75%; the inhibition rate of spraying 48h before inoculation is 84.99%, and the inhibition rate of spraying 24h before inoculation is 80.70%; the inhibition rate of spraying 24h after inoculation is 70.18%. The inhibition rate of the withered spots sprayed before inoculation is obviously higher than that of the spray after inoculation on TMV, which shows that the prevention effect is better than the treatment effect, and the spraying effect is better 48 hours in advance.
4) Pot culture test determination of inhibition of fermentation filtrate on tomato to TMV test results: various treatments of the strain SNB55 fermentation filtrate have certain inhibition effect on TMV, and specific results are shown in Table 6.
TABLE 6 inhibitory Effect of SNB55 fermentation filtrate on TMV on tomato
As can be seen from the table, the most effective for inhibiting the virus activity is the treatment of directly mixing the fermentation filtrate with TMV, and the prevention effect of the 20 times diluted fermentation filtrate on TMV reaches 80%; the preventive effect of spraying before inoculation is 62.40%, which is obviously higher than the curative effect (50.71%) of spraying after inoculation, and the growth rate of the disease rate of fermentation filtrate treatment is relatively slow, the tomatoes have good growth vigor and green color, and the leaves are stretched and thickened.
5) The potted plant test determines the control effect test result of the fermentation filtrate on the tomato mosaic virus disease: the disease indexes of the tomatoes in the experimental group 1, the experimental group 2 and the experimental group 3 are obviously reduced, the control effects are 63.25%, 61.80% and 66.51% respectively, the control effects are higher than those of the control group 2, the control effect of root irrigation treatment is greater than that of leaf surface spraying, and the control effect of seed coating is greater than that of the specific results shown in table 7. And the tomatoes treated by the fermentation filtrate have good growth vigor and green color, the stems are thick and strong, the tomato plants of the control group are short and small, and the leaves are yellow.
TABLE 7 preventive effect of the fermentation filtrate of SNB55 strain on tomato mosaic Virus
3) Pot experiment determination of the control effect test result of the fermentation filtrate on tomato yellow leaf curl virus disease: the SNB55 fermentation filtrate can inhibit tomato yellow leaf curl virus infection, thereby reducing tomato morbidity, and specific results are shown in Table 8.
TABLE 8 preventive effect of SNB55 fermentation filtrate on tomato yellow leaf curl virus
As can be seen from the table, the control effect of the fermentation filtrate diluted by 20 times before the TYLCV is 56.30 percent on tomato yellow leaf curl virus, the control effect of the control group 2 is 43.20 percent, and the morbidity can be obviously reduced.
4) The field plot experiment determines the control effect test result of the fermentation filtrate on tomato yellow leaf curl virus disease: the strain SNB55 fermentation filtrate has better prevention and treatment effect on tomato yellow leaf curl virus, and is superior to amino-oligosaccharin, and the specific results are shown in Table 9.
TABLE 9 prevention of tomato yellow leaf curl virus in field plot by strain SNB55 fermentation filtrate
The tomatoes begin to attack about 35 days after field planting, and are investigated 15 days after the attack, as can be seen from table 9, the disease indexes of the tomatoes in the experimental group 1, the experimental group 2 and the experimental group 3 are obviously reduced, the control effects are 52.86%, 31.91% and 56.31% respectively, the control effect of the experimental group is higher than that of the medicament control group 2, the control effect of root irrigation treatment > the control effect of foliage spray > the control effect of seed coating, the disease attack of the experimental group sprayed with the fermentation filtrate of the strain SNB55 is obviously lighter, and the growth vigor of the plants is high.
5) And (3) detecting the influence of the fermentation filtrate on the growth of tomato seeds and seedlings:
influence on seed growth: the SNB55 fermentation filtrate stock solution can inhibit seed germination, the 20-fold dilution of the strain SNB55 fermentation filtrate can promote tomato seed germination and rooting, the specific results are shown in Table 10, and the rooting condition of the seeds is shown in FIGS. 5 and 6.
TABLE 10 results of the effect of fermentation filtrates on the growth of tomato seeds
As can be seen from the table, the seed treated by the strain SNB55 fermentation filtrate stock solution is 42.86% shorter than the control, while the seed treated by diluting the fermentation filtrate by 20 times is germinated earlier than the control, rooted earlier and has more root systems, and when the root length treated by the fermentation dilution solution reaches 5cm, the root length is increased by 42.86% than the control.
Influence on the growth of seedlings: the 20-fold dilution of the fermentation filtrate of the strain SNB55 has the effect of promoting the growth of seedlings, the specific results are shown in Table 11, and the growth conditions of the seedlings are shown in figures 7 and 8.
TABLE 11 growth promoting effect of fermentation broth of strain SNB55 on tomato seedlings
It can be seen from the table that the tomato seeds treated by the SNB55 fermentation filtrate diluted by 20 times grow well and have dark green color, indexes such as plant height, root length, fresh weight/dry weight of overground part, fresh weight/dry weight of underground part and the like and growth vigor are obviously superior to those of the control, and are respectively increased by 12.07%, 8.53%, 23.77%, 31.88%, 16.22% and 34.51% compared with the control.
6) Broad spectrum bacteriostasis test results of SNB 55: SNB55 has certain inhibition effect on cucumber fusarium wilt, tobacco target spot pathogen, phytophthora capsici, botrytis cinerea and rice sheath blight, wherein the inhibition effect on the tobacco target spot pathogen is strongest, and the inhibition effect on the rice sheath blight is next.
7) Assay results for SNB55 secretion of Indole Acetic Acid (IAA): after the strain SNB55 bacterial suspension is added into the SaLkowski colorimetric solution, the color turns red, which shows that the strain SNB55 can secrete IAA.
It should be understood that the above-described specific embodiments are merely illustrative of the present invention and are not intended to limit the present invention. Obvious variations or modifications which are within the spirit of the invention are possible within the scope of the invention.
Claims (2)
1. A strain of Bacillus belgii for inhibiting virus and promoting plant growth is named as Bacillus belgii (Bacillus belgii)Bacillus velezensis）SNB55, the preservation unit is China general microbiological culture Collection center, the preservation number is CGMCC NO.17315, and the preservation date is 2019, 3 and 8 days.
2. The application of the bacillus belgii for inhibiting viruses and promoting plant growth according to claim 1, wherein the bacillus belgii SNB55 is applied to control of tobacco mosaic virus diseases, tomato yellow leaf curl virus diseases, cucumber fusarium wilt, tobacco target spot, pepper blight, rice sheath blight and tomato gray mold.
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| CN114854650B (en) * | 2022-07-05 | 2022-09-30 | 烟台泓源生物肥料有限公司 | Microbial agent and preparation method thereof |
| CN115011530B (en) * | 2022-07-21 | 2023-06-16 | 东北农业大学 | Multifunctional composite bacillus beijensis microbial inoculum for tomatoes and preparation method and application thereof |
| CN116083321B (en) * | 2023-03-07 | 2023-06-23 | 潍坊科技学院 | Bacillus pumilus F1-3 and application thereof in plant virus control |
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