CN111280181A - Powder moistening microbial inoculum for directional culture and preservation of microorganisms - Google Patents
Powder moistening microbial inoculum for directional culture and preservation of microorganisms Download PDFInfo
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- CN111280181A CN111280181A CN201910247456.0A CN201910247456A CN111280181A CN 111280181 A CN111280181 A CN 111280181A CN 201910247456 A CN201910247456 A CN 201910247456A CN 111280181 A CN111280181 A CN 111280181A
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- preservation
- microbial inoculum
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N25/00—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests
- A01N25/08—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests containing solids as carriers or diluents
Abstract
A powder moistening microbial inoculum for directional culture and preservation of microorganisms. The invention belongs to the field of biotechnology, and discloses a microbial directional culture preservation powder-moistening microbial inoculum prepared from microbial fermentation liquor, polyhydroxy compounds, amino compounds and bioactive adsorption materials.
Description
Technical Field
The present invention belongs to the field of biological pesticide and biochemical technology.
Background
Chemical pesticides play an important role in controlling agricultural pests, protecting agricultural production safety, reducing labor intensity and the like, but with the wide application of a large amount of chemical pesticides, environmental problems and video safety problems caused by chemical pesticide residues are increasingly concerned. The search for new agricultural measures meeting the agricultural sustainable development needs becomes a hot spot of the current agricultural production research. The biocontrol bacteria and metabolites thereof can not only inhibit the growth of various pathogenic bacteria, but also promote the growth of plants and improve the yield of crops, and the development of biocontrol strains and effective antibacterial active ingredients thereof become the focus of research and competition at present.
The fatal defects of microbial pesticides are sensitivity to external conditions under natural conditions, easy death and short shelf life. The problems of short shelf life, easy death in nature, easy influence by environment and the like of microbial pesticides can be solved by improving the dosage form. In order to prolong the storage period, many microbial pesticides are optimized through fermentation to culture organisms with strong stress resistance, such as spores, chlamydospores and the like, and then are prepared into wettable powder, micro-capsule granules, water dispersible granules and the like. This method increases the resistance of microbial pesticides to some extent under natural conditions, but the shelf life is still not long.
At present, in the aspect of protecting microbial bactericides from being damaged by environmental factors, some researches are carried out in the aspect of dosage forms, mainly comprising the development of non-aqueous dosage forms, the application of ultraviolet light protective agents, the development of starch and alginate capsules, the development and application of rain scour resistance agents and synergistic agents, the development of biological capsules and the like.
These previous studies suggest that the microbial strain to be fermented is hydrophilic, and if the prepared formulation contains water, the strain is easily inactivated when stored at normal temperature for a long period of time, and therefore it is generally necessary to reduce the water content of the formulation of the microbial agent to 5% or less. These methods involve drying the cultured microorganisms to form the product, which not only results in inactivation of a significant amount of the bacteria during the drying process, but also increases the manufacturing cost.
Disclosure of Invention
A powder moistening microbial inoculum for directional culture and preservation of microorganisms is characterized in that microbial fermentation mother liquor is inoculated to a culture medium containing the following components in a ratio of 2-5: 3-10, and preparing a powdering carrier prepared from polyhydroxy compound, amino compound and bioactive adsorption material, wherein the pH value of the powdering carrier is 6.1-8.2, and the water content of the powdering carrier is 5.1-25%.
Such microorganisms include, but are not limited to, Bacillus subtilis, Pseudomonas fluorescens, Bacillus pumilus, Bacillus amyloliquefaciens, Bacillus brevis, Bacillus cereus, Paenibacillus polymyxa, Streptomyces hygroscopicus, Trichoderma, Verticillium pachycem, Bacillus firmus, Bacillus thuringiensis, Beauveria bassiana, Paecilomyces lilacinus, and the like.
The polyhydroxy compound includes but is not limited to one or more of trehalose, sucrose, sorbose, melezitose, fructose, mannose, maltose, lactose, arabinose, xylose, ribose, rhamnose, glucose, mannitol, glycerol, xylitol, erythritol, threitol, sorbitol and raffinose, dextrin and dextran.
The amino compound includes, but is not limited to, one or more of lysine, arginine, glutamic acid, aspartic acid, glycine, methionine, histidine, a polypeptide, peptone and gelatin.
The bioactive adsorption material comprises but is not limited to one or more of activated carbon, diatomite, white carbon black, medical stone, kaolin, bran shells, starch, carboxymethyl starch, hydroxyethyl starch, porous starch, soluble starch and macroporous adsorption starch, and the medical stone is more preferable.
The combination ratio of the polyhydroxy compound to the amino compound is 2-5: 3-10, more preferably 2: 5 in combination.
The microbial emollient form has a pH of 6.1-8.2 and a water content of 5.1-25%, more preferably a pH of 7.2 and a water content of 12%.
The plant diseases and insect pests include, but are not limited to, plant-harmful microorganisms such as nematodes, fungi or bacteria, and insect pests caused by chewing mouthparts, piercing-sucking mouthparts, rasping-sucking mouthparts, scraping-sucking mouthparts and feeding-sucking mouthparts.
The innovation point of the invention is that the bioactive adsorption material is processed by a compound containing hydroxyl and amino to prepare a bioactive carrier which is more compatible with the microbial bacteria, so that the microbial bacteria can continuously grow and reproduce on the carrier, and the storage performance of the microbial bacteria is improved in the preservation of active life, and the microbial bacteria can be stored for a long time at normal temperature. The second innovation point is that the technical problem that the activity of the microbial agent product can be preserved for a long time only by reducing the water content to be below 5% through drying is solved, so that the microbial agent can be directly preserved on a carrier with the water content of more than 10% for a long time without drying, and the production cost is obviously reduced.
The invention solves the technical problem of directional culture and preservation of the live bacteria biopesticide, so that the biopesticide can still stably exert the effect after being preserved for a long time.
Detailed description of the preferred embodiments
The present invention is further described in terms of the following examples, which are included to provide further understanding to those skilled in the art. Any modifications and alterations made without departing from the spirit of the invention are within the scope of the invention.
Embodiment one
1. Preparation of microbial directional culture and preservation pollen-moistening bacterial agent
1.0g of trehalose, 1.5g of maltose, 8.5g of mannitol, 4.5g of aspartic acid, 15g of peptone, 1.5g of monopotassium phosphate and 5.0g of gelatin are added into 1 liter of water, medical stone is added after stirring and mixing uniformly, reaction is carried out for 2 hours at 125 ℃, and a microbial oriented culture bioactive material carrier is prepared, wherein the pH value of the carrier is 7.5, and the water content is 13%.
Inoculating bacillus subtilis to an LB culture medium, culturing at 28 ℃ for 72 hours at 150r/min, uniformly inoculating fermentation liquor to a bioactive material carrier to prepare a powder moistening microbial inoculum with the pH of 7.5 and the water content of 13%, packaging the powder moistening microbial inoculum in a sealed container, storing at normal temperature, and measuring the content (cfu/g) of the bacillus subtilis in 6 months, 12 months and 18 months respectively. The survival rates of microorganisms of different formulations were compared with commercially available 10-hundred million Bacillus subtilis wettable powder and 1-hundred million Bacillus subtilis micro-encapsulated granules, and the results are shown in Table 1.
TABLE 1 survival rates of microorganisms of different dosage forms
| Dosage forms | Powder-moistening microbial inoculum | Wettable powder formulation | Microcapsule granule |
| Initial detection of content | 5.2x108 | 9.5x108 | 1.7x108 |
| 6 months old | 3.6x109 | 7.5x107 | 1.6x108 |
| 12 months old | 7.5x108 | 3.1x106 | 1.5x108 |
| 18 months old | 5.2x108 | 5.6x105 | 1.4x108 |
The result shows that the bacterial load of the bacillus subtilis of the powder-moistening microbial agent prepared by the bioactive material carrier is slightly increased in the first 6 months, and then the bacillus subtilis is relatively stable, the microcapsule has strong protection effect on bacteria, and the wettable powder has weak protection effect.
Embodiment two
In the incidence area of pepper root rot, bacillus subtilis powder-moistening inoculants in different storage periods are taken, tests are designed according to random block groups, root irrigation treatment is carried out 24 hours after inoculation according to 100 ml of liquid medicine for each seedling, and the treatment is repeated for 4 times for each 20 plants. The experiment was divided into 3 treatments: treatment 1: the bacillus subtilis powder-moistening agent stored for 3 months is diluted by 500 times of the liquid medicine; and (3) treatment 2: the bacillus subtilis powder-moistening agent stored for 12 months is diluted by 500 times of the liquid medicine; and (3) treatment: the bacillus subtilis powder-moistening agent stored for 18 months is diluted by 500 times of the liquid medicine; and (4) treatment: clear water control; results as shown in table 2, the control plot was severely affected and the bioactive material vehicle formulation significantly reduced the risk of pepper root rot.
Table 2, effect of bacillus subtilis powder-moistening agent on preventing and treating pepper root rot
| Treatment of | The incidence of disease% |
| 500 times of 3 months old powder-moistening microbial inoculum | 11.25a1 |
| 500 times of 12 months old powder-moistening microbial inoculum | 15.50ab |
| 500 times of 18 months old powder-moistening microbial inoculum | 17.50b |
| Clear water control | 52.50c |
Note that1: the trial was designed as a random panel, with 4 replicates per 20 replicates, and the different letters followed by the data in each column of the table indicate significant differences at the 5% level between treatments.
Claims (8)
1. A powder moistening microbial inoculum for directional culture and preservation of microorganisms is characterized in that microbial fermentation mother liquor is inoculated to a culture medium containing the following components in a ratio of 2-5: 3-10, and preparing a flour moistening carrier from the polyhydroxy compound, the amino compound and the bioactive adsorption material, wherein the pH value is 6.1-8.2, and the water content is 5.1-25%, and the functions of the flour moistening microbial inoculum include but are not limited to regulating the growth and development of plants and preventing and treating plant diseases and insect pests.
2. The flour moistening microbial inoculum for directional culture and preservation of microorganisms according to claim 1, which is characterized in that: such microorganisms include, but are not limited to, Bacillus subtilis, Pseudomonas fluorescens, Bacillus pumilus, Bacillus amyloliquefaciens, Bacillus brevis, Bacillus cereus, Paenibacillus polymyxa, Streptomyces hygroscopicus, Trichoderma, Verticillium pachycem, Bacillus firmus, Bacillus thuringiensis, Beauveria bassiana, Paecilomyces lilacinus, and the like.
3. The flour moistening microbial inoculum for directional culture and preservation of microorganisms according to claim 1, which is characterized in that: the polyhydroxy compound includes but is not limited to one or more of trehalose, sucrose, sorbose, melezitose, fructose, mannose, maltose, lactose, arabinose, xylose, ribose, rhamnose, glucose, mannitol, glycerol, xylitol, erythritol, threitol, sorbitol and raffinose, dextrin and dextran.
4. The flour moistening microbial inoculum for directional culture and preservation of microorganisms according to claim 1, which is characterized in that: the amino compound includes, but is not limited to, one or more of lysine, arginine, glutamic acid, aspartic acid, glycine, methionine, histidine, a polypeptide, peptone and gelatin.
5. The inoculant according to claim 1, wherein the bioactive adsorbing material comprises one or more of activated carbon, diatomaceous earth, white carbon black, medical stone, kaolin, bran shell, starch, carboxymethyl starch, hydroxyethyl starch, porous starch, soluble starch and macroporous adsorption starch, and more preferably medical stone.
6. The flour moistening microbial inoculum for directional culture and preservation of microorganisms according to claim 1, which is characterized in that: the combination ratio of the polyhydroxy compound to the amino compound is 2-5: 3-10, more preferably 2: 5 in combination.
7. The flour moistening microbial inoculum for directional culture and preservation of microorganisms according to claim 1, which is characterized in that: the microbial emollient form has a pH of 6.1-8.2 and a water content of 5.1-25%, more preferably a pH of 7.2 and a water content of 12%.
8. The flour moistening microbial inoculum for directional culture and preservation of microorganisms according to claim 1, which is characterized in that: the plant diseases and insect pests include, but are not limited to, plant-harmful microorganisms such as nematodes, fungi or bacteria, and insect pests caused by chewing mouthparts, piercing-sucking mouthparts, rasping-sucking mouthparts, scraping-sucking mouthparts and feeding-sucking mouthparts.
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| Application Number | Priority Date | Filing Date | Title |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114606156A (en) * | 2022-02-24 | 2022-06-10 | 赤天化科技集团有限公司 | Multifunctional compound microbial agent and preparation method and application thereof |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102459568A (en) * | 2009-05-26 | 2012-05-16 | 先进生物营养公司 | Stable dry powder composition comprising biologically active microorganisms and/or bioactive materials and methods of making |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102459568A (en) * | 2009-05-26 | 2012-05-16 | 先进生物营养公司 | Stable dry powder composition comprising biologically active microorganisms and/or bioactive materials and methods of making |
Non-Patent Citations (2)
| Title |
|---|
| 熊元林等: "《微生物学实验》", 31 January 2014, 华中师范大学出版社 * |
| 王 岩 等: ""海藻糖复配保护剂对三种益生菌稳定性影响效果的试验研究"", 《农业开发与装备》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114606156A (en) * | 2022-02-24 | 2022-06-10 | 赤天化科技集团有限公司 | Multifunctional compound microbial agent and preparation method and application thereof |
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Application publication date: 20200616 |