CN106106721B - Method for controlling diseases by inducing fruit resistance and used preparation - Google Patents
Method for controlling diseases by inducing fruit resistance and used preparation Download PDFInfo
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- CN106106721B CN106106721B CN201610516655.3A CN201610516655A CN106106721B CN 106106721 B CN106106721 B CN 106106721B CN 201610516655 A CN201610516655 A CN 201610516655A CN 106106721 B CN106106721 B CN 106106721B
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- 239000000661 sodium alginate Substances 0.000 description 1
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- YFNCATAIYKQPOO-UHFFFAOYSA-N thiophanate Chemical compound CCOC(=O)NC(=S)NC1=CC=CC=C1NC(=S)NC(=O)OCC YFNCATAIYKQPOO-UHFFFAOYSA-N 0.000 description 1
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23B—PRESERVING, e.g. BY CANNING, MEAT, FISH, EGGS, FRUIT, VEGETABLES, EDIBLE SEEDS; CHEMICAL RIPENING OF FRUIT OR VEGETABLES; THE PRESERVED, RIPENED, OR CANNED PRODUCTS
- A23B7/00—Preservation or chemical ripening of fruit or vegetables
- A23B7/14—Preserving or ripening with chemicals not covered by groups A23B7/08 or A23B7/10
- A23B7/153—Preserving or ripening with chemicals not covered by groups A23B7/08 or A23B7/10 in the form of liquids or solids
- A23B7/154—Organic compounds; Microorganisms; Enzymes
- A23B7/155—Microorganisms; Enzymes; Antibiotics
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- 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
- C12N1/14—Fungi; Culture media therefor
- C12N1/16—Yeasts; Culture media therefor
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- Engineering & Computer Science (AREA)
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- Bioinformatics & Cheminformatics (AREA)
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- Tropical Medicine & Parasitology (AREA)
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- General Engineering & Computer Science (AREA)
- Virology (AREA)
- Medicinal Chemistry (AREA)
- Food Science & Technology (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Polymers & Plastics (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
- Pretreatment Of Seeds And Plants (AREA)
Abstract
The invention discloses a preparation for inducing fruit resistance to control diseases, 1L of the preparation contains 1 multiplied by 109～1×1010Cryptococcus laurentii inactivates thallus cells, and the balance is water. The invention also provides a method for controlling diseases by inducing fruit resistance by using the preparation, which comprises the following steps of pretreating the fruits in any mode before the fruits are packed and stored: firstly, soaking fruits in the preparation, draining, placing the soaked fruits in a container, and placing the container in a sealed state for 23-25 hours; secondly, in the pretreatment mode, before picking the fruits, spraying the preparation on the surfaces of the fruits, and then placing the fruits in a container to stand for 23-25 hours in a sealed state; and taking the pretreated fruits out of the container and then boxing the fruits. The invention can effectively control the postharvest diseases of the fruits on the premise of not using chemical bactericides.
Description
Technical Field
The invention relates to the technical field of fruit postharvest disease control, in particular to a biological preservation technology for inhibiting fruit postharvest diseases by inducing fruit resistance.
Background
The fruit is rotten after being picked, so that the loss is huge, and the data shows that the rotting of fruits and vegetables exceeds 8000 million tons every year in China, so that the economic loss reaches billions yuan. Pathogenic fungi are the main cause of fruit postharvest rot, and common pathogenic fungi mainly comprise penicillium spp, Botrytis spp, Alternaria spp and the like.
Currently, chemical fungicides are still the main means of controlling postharvest disease in fruits. Taking citrus as an example, the Chinese patent of invention (application No. 200910028802.2, citrus antistaling agent) provides a plurality of chemical bactericides for controlling citrus diseases, including thiophanate methyl and carbendazim. The Chinese patent of invention (application No. 200910028804.1, orange long-acting antistaling agent) provides a chemical bactericide for controlling orange diseases, which comprises 2,4-D sodium salt and thiophanate. The Chinese invention patent (application number 201110209305.X, a citrus preservative and a preparation method and application thereof) provides a plurality of chemical bactericides capable of controlling citrus diseases, including kresoxim-methyl, pyraclostrobin, trifloxystrobin, enestroburin and the like. However, the residue of the chemical bactericide is easy to cause serious harm to human health and environmental safety, and meanwhile, pathogenic microorganisms can generate drug resistance to influence the disease control effect. Therefore, the search for new, safe and effective disease control methods to replace germicides is becoming a focus of increasing attention.
Plants themselves have a complex set of defense mechanisms to combat pathogenic bacterial invasion. Exogenous factors can cause this defense mechanism to manifest by stimulating plant tissues, thereby causing disease resistance in plants, a phenomenon known as induced resistance. A large number of researches show that heat treatment, ultraviolet ray and calcium treatment, plant hormone, natural antibacterial substances, biological preservatives and the like can induce fruits to generate resistance. The induced resistance is used as a control method by utilizing the self disease resistance mechanism of plants, has the advantages of safety, stability, environmental friendliness and the like compared with chemical bactericides, can effectively reduce the rot of fruits in the storage process after picking, and is considered as one of effective novel disease control methods. Chinese granted patent for invention mixture and method for inducing resistance in plants, publication No. 1925748, discloses a method for inhibiting pathogen infection by stimulating the natural defense system of plants and inducing resistance in plants using two or more compounds, including salicylic acid or functionally similar products thereof, accelerating compounds, regulatory compounds. The Chinese invention patent (application No. 201510015741.1, a melon disease control method) provides a safe melon disease control method, i.e. acetylsalicylic acid is sprayed in four periods of melon young fruit period, fruit rapid expansion period, reticulate formation period and 48 hours before harvest to induce the resistance of the fruit. The Chinese invention patent (application No. 201410314131.7, method for inducing fruit resistance to control diseases and used preparation) provides a method for inducing fruit resistance to control diseases by using colloidal chitin solution.
Cryptococcus laurentii is a post-harvest biocontrol yeast strain widely studied at home and abroad. The invention of China's patent of culture method of antagonistic bacteria of yeast and its special culture medium', patent No. ZL200510090063.1, discloses a culture medium formula of Cryptococcus laurentii; a vacuum freeze-dried product of antagonistic yeast C.laurentii and its preparation method (patent No. ZL200510090065.0) discloses a preparation method of a freeze-dried product of Cryptococcus laurentii; a biological antagonistic bacterium and its application (application No. 200310115336.4) disclose a method for preventing and treating the diseases of sweet cherry, winter jujube and apple by using Cryptococcus laurentii, bactericide and calcium chloride; a bioactive fruit and vegetable preservative film (application No. 200810128441.4) discloses a method for preserving fruits such as apples, cherry tomatoes and the like by utilizing Cryptococcus laurentii, sodium alginate and the like; a method for preventing and treating postharvest diseases or fruit grain shedding of grapes and a special preventing and treating agent thereof (application No. 200910078858.9) disclose a method for preventing and treating diseases or threshing of grapes by using Cryptococcus laurentii, borate and the like; a Cryptococcus capsules and a preparation method thereof (application No. 200910079774.7) disclose a preparation method of capsules for preparing Cryptococcus laurentii preserved at normal temperature.
Disclosure of Invention
The invention aims to provide a method for controlling diseases by inducing fruit resistance and a preparation used by the method.
In order to solve the above problems, the present invention provides a preparation for controlling diseases by inducing fruit resistance, wherein 1L of the preparation contains 1X 109～1×1010Cryptococcus laurentii inactivates the somatic cells, and the balance is water (sterile distilled water).
Improvement of the preparation for inducing fruit resistance to control diseases of the present invention: sterilizing the cryptococcus laurentii thallus cells at 121 ℃ for 20min to obtain the cryptococcus laurentii inactivated thallus cells.
As an improvement of the preparation for inducing fruit resistance to control diseases of the invention, the 1L preparation contains 1X 1010Cryptococcus laurentii inactivates the somatic cells, and the balance is water (sterile distilled water).
The invention also provides a method for controlling diseases by inducing fruit resistance by using the preparation, which comprises the following steps of pretreating the fruits in any mode before the fruits are packed and stored:
the first pretreatment mode,
Firstly, putting fruits into the preparation for soaking, draining, putting the fruits into a container, and standing for 23-25 hours in a sealed state;
secondly, a pretreatment mode is adopted, before fruits are picked, the preparation is sprayed on the surfaces of the fruits (only by wetting), and then the fruits are placed in a container and placed for 23-25 hours in a sealed state;
and taking the pretreated fruits out of the container and then boxing the fruits.
As an improvement of the method for controlling diseases by inducing fruit resistance of the present invention:
in the first pretreatment mode, the soaking time is 8-12 minutes (for example, 10 minutes).
As a further improvement of the method for controlling diseases by inducing fruit resistance of the present invention:
and the containers in the first pretreatment mode and the second pretreatment mode are both fresh-keeping film bags.
In the present invention, the diseases include, for example, penicilliosis and melasma.
In the invention, the Cryptococcus laurentii is Cryptococcus laurentii ZJU10 with the preservation number of CGMCC NO. 3590. This strain has been disclosed in patent application No. 2010101521084.
In the present invention, in the case of the present invention,
the first processing mode is as follows: fruits with substantially consistent appearance quality, such as no mechanical damage, no infection, size and maturity, are selected, rinsed with tap water and allowed to dry at room temperature (about 25 ℃). Soaking the cleaned fruits in the preparation of the invention for 8-12 minutes (preferably 10 minutes), taking out, drying (namely draining water), putting the fruits into a fresh-keeping film bag (the opening of the bag is tightened to seal the bag) for 23-25 hours (namely about 24 hours) at room temperature (about 25 ℃), taking out the fruits from the film bag, and boxing and storing the fruits.
The second treatment method comprises the following steps: the preparation is sprayed on the surface of the fruit (only wet) before picking, the fruit is taken down after drying (no liquid is dropped on the surface of the fruit), the fruit is placed in a fresh-keeping film bag (the bag opening is tightened to seal the bag opening) and kept for 23-25 hours (namely about 24 hours) at room temperature (about 25 ℃), and then the fruit is taken out of the film bag and boxed for storage.
The preparation method of the cryptococcus laurentii inactivated thallus suspension liquid comprises the following steps:
cryptococcus laurentii ZJU10 with the preservation number of CGMCC NO.3590 is stored in an NYDA culture medium (8 g of beef extract, 5g of yeast powder, 10g of glucose and 20g of agar at a low temperature (4 ℃), the culture medium is taken out when activated, the culture medium is cultured for 48 hours at a temperature of 28 ℃ (the culture conditions are 200rpm and 28 ℃), after repeated subculture is carried out for 2 times, the activated yeast is inoculated to the NYDB culture medium (8 g of beef extract, 5g of yeast powder and 10g of glucose and the volume is 1000mL with water, the culture medium is sterilized by high pressure steam and the temperature of 121 ℃) and cultured for 24 hours at the conditions of 200rpm and 28 ℃, culture solution is collected, the culture medium is centrifuged for 15 minutes at the temperature of 4000rpm and 4 ℃, and the culture medium is washed for 2 times by sterile distilled water to remove the culture medium. Resuspending yeast cells in sterile distilled water, determining the concentration with a blood cell counter plate (adjusted to the required concentration with sterile distilled water), and sterilizing at 121 deg.C for 20min in a high-pressure steam sterilizer to obtain suspension of inactivated Cryptococcus laurentii. Diluting the suspension of inactivated bacteria of Cryptococcus laurentii with sterile distilled water until the cells of the inactivated bacteria of Cryptococcus laurentii are 1 × 109～1×1010And (4) obtaining a preparation for inducing fruit resistance to control diseases.
The invention has the advantages that: (1) the cryptococcus laurentii in the invention has rich resources, low cost, high cost performance and biodegradability; (2) the induced resistance is a natural reaction generated after the plant is infected by diseases and insect pests, and can generate lasting and systematic broad-spectrum resistance to the diseases and insect pests; (3) the postharvest disease of the fruits can be obviously reduced, and the fruit tree seedling culture medium has no toxic action on the environment and human health, and has the characteristics of economy, practicability, safety, high efficiency, environmental friendliness and the like; (4) the preparation can effectively control postharvest diseases of fruits on the premise of not using chemical bactericides. (5) And the inactivated thallus cells are more convenient to store.
Drawings
The following describes embodiments of the present invention in further detail with reference to the accompanying drawings.
FIG. 1 is a graph comparing the induction of resistance of inactivated yeast to Penicillium disease in pear;
wherein graph (a) is the incidence of wound treatment and graph (b) is the lesion diameter of wound treatment.
Detailed Description
The invention will be further described with reference to specific examples, but the scope of the invention is not limited thereto.
The Cryptococcus laurentii is Cryptococcus laurentii ZJU10 with the preservation number of CGMCC NO. 3590.
Example 1 preparation for controlling postharvest disease of fruit
Adjusting the concentration of Cryptococcus laurentii to 1 × 108Sterilizing each cell/mL at 121 ℃ for 20min to obtain the cryptococcus laurentii inactivated thallus suspension.
Taking 1L of the cryptococcus laurentii inactivated thallus suspension, and adding sterile water for dilution. Until the obtained preparation contains 1 × 10 per 1L9～1×1010Inactivating thallus cells by Cryptococcus laurentii, and adding water (sterile distilled water) for the rest; obtaining the preparation for inducing fruit resistance to control diseases.
In order to demonstrate the effect of the practical use of the formulation of the present invention, the inventors carried out the following experiments:
experiment 1, Induction Effect of inactivated Yeast on Penicillium disease resistance of Pear fruit
1. Experimental materials:
the fruit is crystal pear.
Pathogenic bacteria: penicillium expansum (Penicillium expansum), activated at 25 ℃ for 7 days for use.
2. And (3) treatment:
selecting fruits with neat appearance, no plant diseases and insect pests and no mechanical damage, cleaning the fruits by using tap water, then soaking the fruits in 0.1 percent (mass percent) of sodium hypochlorite solution for disinfection for 2 minutes, taking the fruits out, then washing the fruits by using the tap water, removing residual sodium hypochlorite, and airing the fruits for later use. Wounds of uniform size and depth were created on the surface of each fruit with a sterile punch. 30 μ L of each wound was added at a concentration of 1X 105、1×106、1×107、1×108cells/mL of the suspension of the inactivated thalli of the Cryptococcus laurentii, and the same amount of sterile water is added as a reference; and sealed with PE plastic film. After 24 hours, the PE plastic film is opened, and 30 mu L of pathogenic spore suspension (Penicillium expansum, concentration of 1X 10) is added to each wound4spores/mL), stored at normal temperature (about 25 ℃ C.), and sealed again with a PE plastic film for moisturizing treatment. Each treatment was repeated 3 times, each for 9 fruits, and the experiment was repeated twice, subject to the same results. The results were observed on day 3.
3. As a result:
as shown in FIG. 1, the disease incidence of the sterile water control group reached 87.2% at a concentration of 1X 10 at day 3 after inoculation of the pathogenic bacteria5、1×106、1×107The morbidity of the cell/mL cryptococcus laurentii inactivated thallus suspension treatment group is 85.7 percent, 59.3 percent and 53.7 percent respectively, which are obviously lower than that of the control group, wherein the morbidity is 1 multiplied by 107The effect of cells/mL was the best, which was reduced by 33.5% compared to the control group. Similar results were obtained with respect to lesion diameter. However, 1X 108The cell/mL-treated group had no effect of suppressing the disease in terms of incidence, and although the value was smaller than that of the control group in terms of lesion diameter, the effect was apparently inferior to that of the 1X 106、1×107cells/mL treatment group.
1. Experimental materials:
the fruit is cherry tomato.
Pathogenic bacteria: alternaria alternata (Alternaria alternata) was activated for 14 days at 25 ℃ for use.
2. And (3) treatment:
selecting fruits with neat appearance, no plant diseases and insect pests and no mechanical damage, cleaning the fruits by using tap water, then soaking the fruits in 0.1% sodium hypochlorite solution for disinfection for 2 minutes, taking the fruits out, then washing the fruits by using the tap water, removing residual sodium hypochlorite, and airing the fruits for later use.
Soaking fructus Pruni Pseudocerasi in 1 × 107cell/mL Cryptococcus laurentii inactivated thallus suspension for 10min, taking out with sterile water as control, draining, and adding PEAnd (5) sealing by using a plastic film. After 24 hours, wounds of uniform size and depth were formed on the surface of each cherry tomato fruit using a sterile punch, and 20. mu.L of A.alternata spore suspension was added at a concentration of 1X 104spores/mL. The occurrence of diseases at the wounds of the fruits was observed and recorded at regular intervals every day, and the results were expressed as the average incidence (%) of disease. Each treatment was repeated 3 times, each 20 fruits, and the experiment was repeated twice, subject to the same results. The final results were recorded after 72 hours.
3. As a result:
as shown in Table 1, 1X 107The whole cherry tomato fruit is soaked by the cell/mL cryptococcus laurentii inactivated thallus suspension, and after 24 hours, pathogenic bacteria are inoculated, so that the incidence of the inactivated yeast thallus is reduced by 17.7 percent compared with that of a control group, and the obvious difference is achieved.
TABLE 1 incidence of cherry tomato fruits after 24 hours of induction (%)
| Incidence after 72 hours (%) | |
| Sterile water | 90.2±3.0 |
| 1×107cell/mL inactivated bacteria | 72.5±3.7 |
Comparing experiment 1, changing the "wounds of uniform size and depth formed on the surface of each cherry tomato fruit by using the sterilized punch after" 24 hours "described in experiment 2 to" 0 hours, 6 hours and 48 hours, the wounds of uniform size and depth formed on the surface of each cherry tomato fruit by using the sterilized punch were formedDeep wound ", the suspension of inactivated bacteria of Cryptococcus laurentii still has a concentration of 1X 107Per mL; the rest is identical to experiment 2.
The results observed at 72 hours are shown in table 2:
TABLE 2 incidence of cherry tomato fruits at different induction times (%)
Under the conditions of no induction (0 hour), short induction (6 hours) and long induction (48 hours), the inhibitory effect of the cryptococcus laurentii inactivated thalli on the black spot of the cherry tomato fruit is not the same as that of the invention (the induction time is 24 hours). Wherein the inactivated yeast does not exhibit any inhibitory effect under the conditions of no induction and short-time induction.
Comparative example (c),
The Bacillus subtilis has good inhibition effects on citrus Penicillium disease and green mold, and on brown rot (Monilinia fructicola) of fruits such as cherry, yellow peach and nectarine, black spot after Chinese chestnut harvest and the like. However, when Bacillus subtilis is inactivated, it no longer has an inhibitory effect even if fruits are induced as in the present invention (the induction time is set to 1 to 48 hours, and the concentration is set to 1X 10)6/L～1×1012L), none of them has an inhibitory effect.
Finally, it is also noted that the above-mentioned lists merely illustrate a few specific embodiments of the invention. It is obvious that the invention is not limited to the above embodiments, but that many variations are possible. All modifications which can be derived or suggested by a person skilled in the art from the disclosure of the present invention are to be considered within the scope of the invention.
Claims (2)
1. The method for controlling diseases by inducing fruit resistance by using the preparation is characterized by comprising the following steps: before the fruits are packed and stored, the following pretreatment is carried out:
firstly, putting fruits into the preparation for soaking for 8-12 minutes; draining, placing in a container, and standing in a sealed state for 23-25 hours;
taking the pretreated fruits out of the container and then boxing the fruits;
1L of the preparation contains 1 × 1010Inactivating thallus cells by Cryptococcus laurentii, and adding water for the rest;
sterilizing the cryptococcus laurentii thallus cells at 121 ℃ for 20min to obtain cryptococcus laurentii inactivated thallus cells;
the diseases are penicilliosis and black spot.
2. The method for controlling diseases by inducing fruit resistance according to claim 1, wherein:
the container is a fresh-keeping film bag.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1733898A (en) * | 2005-08-11 | 2006-02-15 | 中国科学院植物研究所 | Vacuum freeze-dried product of antagonistic yeast C.laurentii and preparation method thereof |
| CN1733897A (en) * | 2005-08-11 | 2006-02-15 | 中国科学院植物研究所 | A kind of cultural method of antagonistic yeast and special culture media thereof |
| CN101491273A (en) * | 2009-03-04 | 2009-07-29 | 中国科学院植物研究所 | Grape postharvest diseases or fruit abscission prevention and control method and special prevention and control agent |
| CN104161116A (en) * | 2014-07-04 | 2014-11-26 | 浙江大学 | Preparation for controlling diseases by inducing fruit resistibility and corresponding induction method |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1733898A (en) * | 2005-08-11 | 2006-02-15 | 中国科学院植物研究所 | Vacuum freeze-dried product of antagonistic yeast C.laurentii and preparation method thereof |
| CN1733897A (en) * | 2005-08-11 | 2006-02-15 | 中国科学院植物研究所 | A kind of cultural method of antagonistic yeast and special culture media thereof |
| CN101491273A (en) * | 2009-03-04 | 2009-07-29 | 中国科学院植物研究所 | Grape postharvest diseases or fruit abscission prevention and control method and special prevention and control agent |
| CN104161116A (en) * | 2014-07-04 | 2014-11-26 | 浙江大学 | Preparation for controlling diseases by inducing fruit resistibility and corresponding induction method |
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| 植物诱导抗病性诱导剂研究进展;张梦月，等;《中国线虫学研究》;20141231;第5卷;第326页 * |
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