CN108684710B - Growth promoting preparation for preventing ginkgo stem rot and preparation method thereof - Google Patents
Growth promoting preparation for preventing ginkgo stem rot and preparation method thereof Download PDFInfo
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- CN108684710B CN108684710B CN201810452044.6A CN201810452044A CN108684710B CN 108684710 B CN108684710 B CN 108684710B CN 201810452044 A CN201810452044 A CN 201810452044A CN 108684710 B CN108684710 B CN 108684710B
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- A—HUMAN NECESSITIES
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- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
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Abstract
The invention relates to the technical field of biology, in particular to a growth promoting preparation for preventing ginkgo stem rot, which comprises the following raw materials in parts by weight: 1.5 to 3.5 parts of pseudomonas chlororaphis orange subspecies, 5.6 to 7.8 parts of bacillus methylotrophicus, 4.3 to 6.5 parts of pseudomonas fluorescens and 15.0 to 20.0 parts of distilled water. The invention also relates to a preparation method and application of the growth-promoting preparation for preventing the stem rot of the ginkgo. By adopting the growth promoting preparation provided by the invention, the ginkgo stem rot is not simply prevented, but is prevented by using the growth promoting preparation provided by the invention, so that the morbidity is reduced to below 10% from 40-70%, the survival rate of ginkgo seedlings is greatly improved, the planting cost is reduced, and the economic value is improved.
Description
Technical Field
The invention relates to the technical field of biology, in particular to a growth promoting preparation for preventing ginkgo stem rot.
The invention also relates to a preparation method and application of the growth-promoting preparation for preventing the stem rot of the ginkgo.
Background
The pathogenic bacteria of ginkgo stalk rot are the charcoal rot germs of large-stalk genus of Chaetoceros, Chaetocercosporaceae and Escholzia. The bacterium prefers high temperature, and the suitable growth temperature is 30-32 ℃; but has low requirement on pH value, and can well grow between the pH value of 4-9. At the early stage of the disease, the base of the seedling turns brown, and the leaves lose normal green color and slightly droop downward but do not fall off. The affected part expands rapidly, so that the whole plant dies. The cortex at the basal part of the diseased seedling is shriveled, and the intradermal tissue is decayed to be spongy or powdery and is grey white, and a plurality of tiny black sclerotia are clamped. The disease pathogen can also invade the xylem of seedlings, so that the brown hollow medulla is also produced by sclerotia microscopically. Thereafter, the germs gradually spread to the roots, causing the root cortex to decay. If the diseased seedling is pulled out by hand, only the xylem can be pulled out, and the root cortex is remained in the soil. The stem rot of the ginkgo cutting seedlings can also occur under the condition of high temperature or low temperature, the surface of the cutting slips can be sleeved on the xylem in a cylindrical shape, and the thin pendulous tissues of the phloem are completely blackened and rotten.
The phomopsis is usually decomposed and lives in soil, and belongs to weak parasitic fungi. And (3) invading from the wound of the seedling under proper conditions. Therefore, disease development is related to host and site environmental conditions. The root cause of seedling damage is germ invasion caused by high-temperature burn of the base of the seedling due to overhigh ground surface temperature. The lower the degree of lignification of the nursery stock, the higher the incidence rate of the disease. The incidence of water accumulation is also obviously increased in the low-lying part of the seedbed. When the high temperature of the seedbed reaches more than 30 ℃ in 6-8 months, the ginkgo cutting seedlings start to cause diseases after 10-l 5d are cut, and large-area scions are blackened and die when the seedlings are serious.
According to the cause of the stem rot of ginkgo, the prevention is mainly used nowadays. A. The early sowing is carried out when the soil is thawed, the measure is favorable for early lignification of the nursery stock, and the resistance to the high temperature of the soil surface is enhanced. B. The reasonable dense sowing and dense sowing are beneficial to exerting the group effect of the nursery stock and enhancing the resistance to the external adverse environment. Tests prove that the smaller the seedling density, the higher the incidence rate and the larger the density, the lower the incidence rate. Practices also prove that after the seeds are sown by 25-40 kg per mu in the past, if the sowing amount of the seeds is changed into 80-l 00kg per mu, the morbidity is reduced, the unit seedling yield is increased, the land is saved, and the morbidity is reduced. C. After the nursery stocks for preventing and controlling the underground pests are damaged by the underground pests; is very easy to be infected by the stem rot germs, so that the underground pests need to be killed at certain time before and after sowing. D. The mechanical damage of the seedlings to annual sowing seedlings or annual transplanting seedlings is prevented, the roots and stems of the seedlings are not required to be damaged in the soil loosening weeding or seedling lifting transplanting process, and otherwise, the occurrence of stem rot is easily caused. E. Shading and cooling to prevent the ground temperature of solar radiation from increasing, and measures such as shading shed building, inter-row grass covering, corn planting, cutting and shading are adopted in the seedling culture ground to reduce the harm to seedlings. F. Irrigation and water spraying are timely performed in high-temperature seasons to reduce the surface temperature, and irrigation can be adopted in places with conditions, so that the disease is reduced.
Experiments prove that the antagonistic actinomycetes can effectively inhibit the spread of the pathogenic bacteria.
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provides a growth promoting preparation for preventing the stem rot of ginkgo.
In order to solve the above technical problems, the present invention now proposes the following technical solutions: a growth promoting preparation for preventing ginkgo stem rot is composed of the following raw materials in parts by weight: 1.5 to 3.5 parts of pseudomonas chlororaphis orange subspecies, 5.6 to 7.8 parts of bacillus methylotrophicus, 4.3 to 6.5 parts of pseudomonas fluorescens and 15.0 to 20.0 parts of distilled water.
Further, the feed additive is composed of the following raw materials in parts by weight: 2.5 parts of pseudomonas chlororaphis orange subspecies, 6.6 parts of bacillus methylotrophicus, 5.4 parts of pseudomonas fluorescens and 18 parts of distilled water.
Further, the culture method of the pseudomonas aeruginosa orange subspecies comprises the following steps: streak inoculation is carried out on a KB culture medium, the diameter of a bacterial colony reaches 2mm after 24 hours of culture at 28 ℃ in a non-anaerobic environment, the bacterial colony is round and slightly raised, is yellowish, has smooth edges, is wet in surface and is easy to pick up, the thalli generate orange pigments after 48 hours of culture and cover the surface of the bacterial colony, and meanwhile, orange pigments are generated around the bacterial colony.
Further, the formula of the KB medium is as follows: the pH of the water per 1000 ml contains peptone 20 g, glycerin 15 ml, K2HPO40.392 g and MgSO40.732 g, and is 6.5-7.5.
Further, the method for culturing the bacillus methylotrophicus comprises the following steps: firstly culturing the bacillus methylotrophicus in a seed culture medium until the density OD600 value is 0.6-0.8 to obtain a bacterial liquid, then inoculating 2-5 parts by weight of the bacterial liquid of the bacillus methylotrophicus in 100 parts by weight of the culture medium, and culturing at 37 ℃.
Further, the seed culture medium comprises the following components: 10g/L of tryptone, 5g/L of yeast extract and 10g/L of NaCl; the 100 parts by weight of culture medium comprises the following components: 15g of glucose, 1g of starch, 25g of bean cake powder, 1g of manganese sulfate, 1.5g of monopotassium phosphate, 0.5g of magnesium sulfate, 0.2g of yeast extract, 0.1g of ferric chloride, 0.1g of calcium carbonate and pH 7.0-7.2.
Further, the culture method of the pseudomonas fluorescens comprises the following steps: streak inoculation is carried out on a KB culture medium, the KB culture medium is cultured for 48h at the temperature of 28 ℃ in a non-anaerobic environment, the KB culture medium grows well, the bacterial colony is raised, is faint yellow, has smooth edges, has a wet surface and is easy to pick up; generating water-soluble yellow-green pigment, wherein strong yellow-green fluorescence can be generated under the irradiation of 254nm ultraviolet light, and the water-soluble pigment can be diffused into the culture medium after the culture for 3-4 days, and the yellow-green fluorescence is gradually weakened to disappear.
Further, the formula of the KB medium is as follows: the pH of the water per 1000 ml contains peptone 20 g, glycerin 15 ml, K2HPO40.392 g and MgSO40.732 g, and is 6.5-7.5.
The invention also provides a preparation method of the growth promoting preparation for preventing the ginkgo stem rot, which comprises the following steps: (1) inoculating Pseudomonas chlororaphis on KB culture medium by streaking, and culturing for 24-48 h at 28 ℃ in a non-anaerobic environment; (2) culturing bacillus methylotrophicus in a seed culture medium until the density OD600 value is 0.6-0.8 to obtain a bacterial liquid, then inoculating 2-5 parts by weight of the bacterial liquid of the bacillus methylotrophicus in 100 parts by weight of the culture medium, and culturing at 37 ℃; (3) inoculating fluorescent pseudomonas on a KB culture medium by streaking, culturing for 48h at 28 ℃ in a non-anaerobic environment, and culturing for 3-4d when strong yellow-green fluorescence is visible under the irradiation of 254nm ultraviolet light; (4) adding distilled water into the container according to the proportion, and sealing for a period of time at room temperature; (4) and at room temperature, sequentially adding the orange subspecies of the pseudomonas aeruginosa, the methylotrophic bacillus and the pseudomonas fluorescens according to the proportion, and uniformly shaking to obtain the bacillus fluorescens.
The invention also aims to provide the application of the growth-promoting preparation in improving the content of the lactone B in the ginkgo leaves.
Compared with the prior art, the invention has the beneficial effects that: 1. by adopting the growth promoting preparation, the ginkgo stem rot is not simply prevented, but is prevented by using the growth promoting preparation, so that the morbidity is reduced to below 10 percent from 40-70 percent, the survival rate of ginkgo seedlings is greatly improved, the planting cost is reduced, and the economic value is improved;
2. the growth promoting preparation of the invention has simple preparation method and strong practicability.
Detailed Description
For a better understanding and appreciation of the structural features and advantages achieved by the present invention, the following detailed description of the preferred embodiments is provided:
example 1
A growth promoting preparation for preventing ginkgo stem rot is composed of the following raw materials in parts by weight: 1.5 parts of pseudomonas chlororaphis orange subspecies, 5.6 parts of bacillus methylotrophicus, 4.3 parts of pseudomonas fluorescens and 15.0 parts of distilled water.
Example 2
A growth promoting preparation for preventing ginkgo stem rot is composed of the following raw materials in parts by weight: 2.5 parts of pseudomonas chlororaphis orange subspecies, 6.6 parts of bacillus methylotrophicus, 5.4 parts of pseudomonas fluorescens and 18.0 parts of distilled water.
Example 3
A growth promoting preparation for preventing ginkgo stem rot is composed of the following raw materials in parts by weight: 3.5 parts of pseudomonas chlororaphis orange subspecies, 7.8 parts of bacillus methylotrophicus, 6.5 parts of pseudomonas fluorescens and 20.0 parts of distilled water.
Example 4
A preparation method of a growth promoting preparation for preventing ginkgo stem rot comprises the following steps: (1) inoculating Pseudomonas chlororaphis on KB culture medium by streaking, and culturing for 24-48 h at 28 ℃ in a non-anaerobic environment; (2) culturing bacillus methylotrophicus in a seed culture medium until the density OD600 value is 0.6-0.8 to obtain a bacterial liquid, then inoculating 2-5 parts by weight of the bacterial liquid of the bacillus methylotrophicus in 100 parts by weight of the culture medium, and culturing at 37 ℃; (3) inoculating fluorescent pseudomonas on a KB culture medium by streaking, culturing for 48h at 28 ℃ in a non-anaerobic environment, and culturing for 3-4d when strong yellow-green fluorescence is visible under the irradiation of 254nm ultraviolet light; (4) adding distilled water into the container according to the proportion, and sealing for a period of time at room temperature; (4) and at room temperature, sequentially adding the orange subspecies of the pseudomonas aeruginosa, the methylotrophic bacillus and the pseudomonas fluorescens according to the proportion, and uniformly shaking to obtain the bacillus fluorescens.
Further, in this example, the culture method of the pseudomonas aeruginosa orange subspecies is as follows: streak inoculation is carried out on a KB culture medium, the diameter of a bacterial colony reaches 2mm after the bacterial colony is cultured for 24 hours at 28 ℃ in a non-anaerobic environment, the bacterial colony is round, slightly raised, slightly yellow, smooth in edge, relatively wet in surface and easy to pick up, after the bacterial colony is cultured for 48 hours, a red-orange pigment is generated by the thallus and covers the surface of the bacterial colony, and meanwhile, an orange-yellow pigment is generated around the bacterial colony; the formula of the KB culture medium is as follows: the pH of the water per 1000 ml contains peptone 20 g, glycerin 15 ml, K2HPO40.392 g and MgSO40.732 g, and is 6.5-7.5.
Further, in this example, the method for culturing bacillus methylotrophicus was: firstly, culturing the bacillus methylotrophicus in a seed culture medium until the density OD600 value is 0.6-0.8 to obtain a bacterial liquid, then inoculating 2-5 parts by weight of the bacterial liquid of the bacillus methylotrophicus in 100 parts by weight of the culture medium, and culturing at 37 ℃; the seed culture medium comprises the following components: 10g/L of tryptone, 5g/L of yeast extract and 10g/L of NaCl; the components of 100 parts by weight of the culture medium are as follows: 15g of glucose, 1g of starch, 25g of bean cake powder, 1g of manganese sulfate, 1.5g of monopotassium phosphate, 0.5g of magnesium sulfate, 0.2g of yeast extract, 0.1g of ferric chloride, 0.1g of calcium carbonate and pH 7.0-7.2.
Further, in this embodiment, the culture method of pseudomonas fluorescens is as follows: streak inoculation is carried out on a KB culture medium, the KB culture medium is cultured for 48h at the temperature of 28 ℃ in a non-anaerobic environment, the KB culture medium grows well, the bacterial colony is raised, is faint yellow, has smooth edges, has a wet surface and is easy to pick up; generating water-soluble yellow-green pigment, wherein strong yellow-green fluorescence can be generated under the irradiation of 254nm ultraviolet light, and the water-soluble pigment can be diffused into the culture medium after the culture for 3-4 days, and the yellow-green fluorescence is gradually weakened to disappear; the formula of the KB culture medium is as follows: the pH of the water per 1000 ml contains peptone 20 g, glycerin 15 ml, K2HPO40.392 g and MgSO40.732 g, and is 6.5-7.5.
Test examples
The experimental examples show that each component of the growth promoting preparation of the invention has a promoting effect on the prevention of the ginkgo stem rot, but after the components are combined together to form the growth promoting preparation of the invention, the incidence of the ginkgo seedlings is lower, the survival rate is higher, the effect is better, the proportion of the components in the example 2 has the best promoting effect, and the incidence of the ginkgo stem rot is only 6%. Finally, it should be noted that: the above embodiments are only used to illustrate the technical solution of the present invention, and are not used to limit the same; although the present invention has been described in detail with reference to the foregoing embodiments, it will be understood by those skilled in the art that various changes may be made and equivalents may be substituted for elements thereof; and the modifications or the substitutions do not make the essence of the corresponding technical solutions depart from the spirit and scope of the technical solutions of the embodiments of the present invention.
Claims (10)
1. A growth promoting preparation for preventing ginkgo stem rot is characterized by comprising the following raw materials in parts by weight: 1.5-3.5 parts of pseudomonas chlororaphis orange subspecies, 5.6-7.8 parts of bacillus methylotrophicus, 4.3-6.5 parts of pseudomonas fluorescens and 15.0-20.0 parts of distilled water; the growth promoting preparation is used for preventing the ginkgo stem rot.
2. The growth promoting preparation for preventing the ginkgo stalk rot according to claim 1, which is characterized by comprising the following raw materials in parts by weight: 2.5 parts of pseudomonas chlororaphis orange subspecies, 6.6 parts of bacillus methylotrophicus, 5.4 parts of pseudomonas fluorescens and 18 parts of distilled water.
3. The growth promoting preparation for preventing stem rot of ginkgo biloba according to claim 1, wherein the culture method of the pseudomonas chlororaphis subsp. Streak inoculation is carried out on a KB culture medium, the diameter of a bacterial colony reaches 2mm after 24 hours of culture at 28 ℃ in a non-anaerobic environment, the bacterial colony is round and slightly raised, is yellowish, has smooth edges, is wet in surface and is easy to pick up, the thalli generate orange pigments after 48 hours of culture and cover the surface of the bacterial colony, and meanwhile, orange pigments are generated around the bacterial colony.
4. The growth-promoting preparation for preventing stem rot of ginkgo according to claim 3, wherein the formula of the KB medium is as follows: each 1000 ml of water contains 20 g of peptone, 15 ml of glycerol and K2HPO40.392 g and MgSO40.732 g and pH 6.5-7.5.
5. The growth promoting preparation for preventing stem rot of ginkgo according to claim 1, wherein the bacillus methylotrophicus is cultured by the following method: firstly, culturing the bacillus methylotrophicus in a seed culture medium until the density of bacillus methylotrophicus is OD600The value is 0.6-0.8, and a bacterial solution is obtained, and then 2-5 parts by weight of a bacterial solution of Bacillus methylotrophicus is inoculated into 100 parts by weight of a culture medium, and cultured at 37 ℃.
6. The growth-promoting preparation for preventing stem rot of ginkgo according to claim 5, wherein the seed culture medium comprises: 10g/L of tryptone, 5g/L of yeast extract and 10g/L of NaCl; the 100 parts by weight of culture medium comprises the following components: 15g of glucose, 1g of starch, 25g of bean cake powder, 1g of manganese sulfate, 1.5g of monopotassium phosphate, 0.5g of magnesium sulfate, 0.2g of yeast extract, 0.1g of ferric chloride, 0.1g of calcium carbonate and pH 7.0-7.2.
7. The growth promoting preparation for preventing stem rot of ginkgo according to claim 1, wherein the pseudomonas fluorescens is cultured by the following method: streak inoculation is carried out on a KB culture medium, the KB culture medium is cultured for 48h at the temperature of 28 ℃ in a non-anaerobic environment, the KB culture medium grows well, the bacterial colony is raised, is faint yellow, has smooth edges, has a wet surface and is easy to pick up; generating water-soluble yellow-green pigment, wherein strong yellow-green fluorescence can be generated under the irradiation of 254nm ultraviolet light, and the water-soluble pigment can be diffused into the culture medium after the culture for 3-4 days, and the yellow-green fluorescence is gradually weakened to disappear.
8. The growth-promoting preparation for preventing stem rot of ginkgo according to claim 7, wherein the formula of the KB medium is as follows: each 1000 ml of water contains 20 g of peptone, 15 ml of glycerol and K2HPO40.392 g and MgSO40.732 g and pH 6.5-7.5.
9. A method of preparing a growth promoting formulation of claim 1, comprising the steps of: (1) inoculating Pseudomonas chlororaphis on KB culture medium by streaking, and culturing for 24-48 h at 28 ℃ in a non-anaerobic environment; (2) culturing the bacillus methylotrophicus in a seed culture medium until the density OD of the bacillus methylotrophicus600Obtaining a bacterial liquid with the value of 0.6-0.8, then inoculating 2-5 parts by weight of the bacterial liquid of the bacillus methylotrophicus into 100 parts by weight of the culture medium, and culturing at 37 ℃; (3) inoculating fluorescent pseudomonas on a KB culture medium by streaking, culturing for 48h at 28 ℃ in a non-anaerobic environment, and culturing for 3-4d when strong yellow-green fluorescence is visible under the irradiation of 254nm ultraviolet light; (4) adding distilled water into the container according to the proportion, and sealing for a period of time at room temperature; (4) and at room temperature, sequentially adding the orange subspecies of the pseudomonas aeruginosa, the methylotrophic bacillus and the pseudomonas fluorescens according to the proportion, and uniformly shaking to obtain the bacillus fluorescens.
10. Use of the pro-growth preparation of claim 1 for preventing stem rot of ginkgo biloba.
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Application publication date: 20181023 Assignee: Xintian County Tianyuan Chunxiao Ecological Agriculture Co.,Ltd. Assignor: HUNAN University OF SCIENCE AND ENGINEERING Contract record no.: X2023980036440 Denomination of invention: A growth promoting agent and preparation method for preventing ginkgo stem rot disease Granted publication date: 20210625 License type: Common License Record date: 20230612 |