CN112369437B - Method and medicament for treating citrus huanglongbing - Google Patents
Method and medicament for treating citrus huanglongbing Download PDFInfo
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Abstract
The invention provides a method and a medicament for treating citrus huanglongbing, belonging to the technical field of biological disease control. The medicament comprises kiwi fruit rhizome extract and plant growth hormone; the method comprises S1, preparing plant growth hormone solution; s2, taking dried kiwi fruit rootstock, and preparing an kiwi fruit rootstock extracting solution; s3, mixing the plant growth hormone solution and the kiwi fruit root and stem extract to obtain a therapeutic agent, and applying the therapeutic agent by adopting a trunk infusion method. The invention can obviously improve the tree vigor of trees which are damaged by the huanglongbing pathogenic bacteria by exploring the value of the traditional Chinese medicine and combining the modern growth regulator technology, has very high-efficiency treatment effect on the huanglongbing, ensures that plants obtain the disease resistance to the huanglongbing, and has stable and reliable treatment effect.
Description
Technical Field
The invention relates to the technical field of plant disease control, in particular to a method and a medicament for treating citrus huanglongbing.
Background
Citrus yellow shoot is a devastating disease in citrus production, caused by a gram-negative bacterium that is parasitic within the phloem, and can infect a variety of rutaceae plants including citrus, kumquat, and murraya.
After the navel orange trees are infected with yellow dragon disease, germs are parasitized in phloem and are transmitted to roots through transmission tissues, absorption of the roots to nutrition is affected, the trees are seriously declined, metabolism of the trees is inhibited, and the internal environment is reversely changed to cause physiological disorder and continuous decline until death. The Huanglongbing trees all have fatal harm, and mainly show that the leaves have mottled yellowing phenomenon, abnormal flowering (flowering in summer and autumn can occur), unripe fruits (green nose fruits can occur and can not be eaten), and the trees decline to death.
The prevention and treatment of the citrus greening disease mainly comprises the elimination of transmission vectors such as diaphorina citri, the direct killing of pathogenic bacteria by using antibiotics, the improvement of tree vigor by using plant hormones or nutrition regulation, and the like. After the Huanglongbing trees are prevented and treated, the effect is unstable, the Huanglongbing trees are easy to relapse, and the medication and treatment process is complicated.
Disclosure of Invention
The invention aims to realize effective and stable treatment of citrus huanglongbing and solve the problems of complicated medication process, easy relapse and the like.
In one aspect of the invention, a medicament for treating citrus huanglongbing is provided, which comprises a kiwi fruit root and stem extract and a plant growth hormone.
The invention firstly explores the reason that other plants do not generate the yellow dragon disease, and then starts to research the mechanism of the yellow dragon disease from two aspects of plant immunity and treatment. The content of the extract of the root and the stem of the kiwi fruit has high compatibility with the internal environment of the tree; in the content, 2 alpha, 3 alpha, 24-trihydroxy-12-ene-28-oleanolic acid, 2 alpha, 3 alpha-dihydroxy-12-ene-28-ursolic acid, 2 alpha, 3 alpha, 19 alpha-trihydroxy-12-ene-28-ursolic acid, beta-sitosterol and beta-daucosterol have the function of treating plant leaf rot. By utilizing the principle of inter-generation and inter-restriction of Chinese medicine, the growth hormone of the diseased tree is in an abnormal secretion state in the aspect of endocrine function because the diseased tree is in a growth decline state, and the growth hormone can be used for regulating the growth of plants and promoting the treatment effect of medicaments on the tree.
Preferably, the plant growth hormones include gibberellins, cytokinins, and brassinolides. Gibberellin has the function of promoting cells to divide rapidly, cytokinin has the function of promoting synthesis of chlorophyll, and brassinolide has the function of enhancing inherent disease resistance; the three components act synergistically to regulate the normal growth of the diseased tree.
Preferably, the cytokinin is one or more of 6-benzylaminopurine, kinetin, zeatin, dihydrozeatin, isopentenyladenine, zeatin nucleoside, isopentenyladenosine.
Preferably, the gibberellin is GA1、GA3、GA4、GA9、GA20One or more of (a).
Preferably, the concentration of the gibberellin is 5-80 ppm.
Preferably, the concentration of the cytokinin is 5-80 ppm.
Preferably, the concentration of the brassinolide is 0.01-1 ppm.
In another aspect of the invention, a method for treating citrus huanglongbing is provided, comprising the steps of:
s1, preparing a plant growth hormone solution;
s2, taking dried kiwi fruit rootstock, and preparing an kiwi fruit rootstock extracting solution;
s3, mixing the plant growth hormone solution and the kiwi fruit root and stem extract to obtain a therapeutic agent, and applying the therapeutic agent by adopting a trunk infusion method.
The principle of the method of the invention is as follows: plants, like animals and humans, have an autoimmune system, which is also responsible for the disease resistance of certain plants to certain pathogenic bacteria that occur naturally. The introduction of foreign matters can activate the immune system of the plant and regulate the metabolism of the plant, thereby enhancing the disease resistance and stress resistance of the plant. Utilizes the characteristics of the Chinese gooseberry rhizome such as bacteria resistance, virus resistance and disease and insect resistance, the function of treating the plant leaf rot, and utilizes the plant growth regulator to regulate the plant growth and promote the treatment effect of the medicine on the tree body. The invention can obviously improve the tree vigor of trees which are damaged by the yellow dragon germs, has very high-efficiency treatment effect on the yellow dragon diseases, enables the plants to obtain the disease resistance on the yellow dragon diseases, and has stable and reliable treatment effect on the yellow dragon diseases.
Preferably, the preparation method of the plant growth hormone solution in step S1 is: firstly, dissolving plant growth hormone, and preparing a solution with a certain concentration;
preferably, the preparation method of the kiwi fruit rhizome extracting solution in the step S2 includes: taking dry kiwifruit rhizome materials, adding ethanol for extraction or adding water for decoction.
Preferably, in the step S3, the kiwi fruit root and stem extract prepared from 3-18 g of kiwi fruit root and stem dry material is mixed with a plant growth hormone solution, and then diluted to a desired concentration to obtain the therapeutic agent.
Preferably, the administration method specifically comprises: and drilling a hole at the height of 30-100 cm of the trunk of the tree, then inserting an infusion needle, and inputting the treatment medicament into the trunk of the tree for treatment.
The invention can obtain at least one of the following beneficial effects:
1. the invention obtains the medicament formula for preventing and treating citrus diseases by exploring the value of the traditional Chinese medicine, utilizing the characteristics of bacteria resistance, virus resistance and disease and insect resistance of the rootstocks of the kiwi fruits and the function of treating the plant leaf rot and combining the modern growth regulator technology, and the formula can obviously improve the tree vigor of the trees which are damaged by the yellow dragon bacteria, has very high-efficient treatment effect on the yellow dragon diseases and ensures that the plants have the disease resistance to the yellow dragon diseases. The plants treated by the medicament have no yellow dragon disease germs through long-term molecular monitoring for many years, which shows that the medicament formula has stable and reliable treatment effect on the yellow dragon disease.
2. According to the invention, immunotherapy is focused, specific medicines for treating the huanglongbing and immunity are selected through research, plant growth hormone is utilized to regulate disease trees, and the medicines and the hormone are matched to play a high-efficiency treatment role; the growth function of the tree body is recovered by adding the plant growth hormone, the absorption of the medicine is promoted, the metabolism function of the organism is enhanced, and the effect of inhibiting and killing the yellow dragon disease germs is achieved.
3. The treatment method of the invention has simple medication and treatment process, stable treatment effect and no relapse, and can be popularized and applied in the treatment of huanglongbing.
Detailed Description
The technical solutions in the embodiments of the present invention are clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments. 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.
First, test method
1. Pharmaceutical formulation
Formula 1:
a medicament for treating citrus greening disease, comprising, on a dose per tree basis: 3g of extract of dried root and stem of Kiwi berry, 10ppm of gibberellinGA165ppm zeatin, 0.01ppm brassinolide.
And (2) formula:
a medicament for treating citrus greening disease, comprising, on a dose per tree basis: 10g of the extract of the dried root and stem of Kiwi berry, 70ppm of gibberellin GA2015ppm of isopentenyl adenosine, 0.5ppm of brassinolide.
And (3) formula:
a medicament for treating citrus greening disease, comprising, on a dose per tree basis: 5g of the extract of the dried root and stem of Kiwi berry, 25ppm of gibberellin GA325ppm of 6-benzylaminopurine, 0.1ppm of brassinolide.
And (4) formula:
a medicament for treating citrus greening disease, comprising, on a dose per tree basis: 7g of an extract of dried kiwi fruit rhizomes, 50ppm of gibberellin GA135ppm zeatin, 0.8ppm brassinolide.
And (5) formula:
a medicament for treating citrus greening disease, comprising, on a dose per tree basis: 18g of extract of dried root and stem of Kiwi berry, 40ppm of gibberellin GA155ppm of zeatin and 0.05ppm of brassinolide.
And (6) formula:
a medicament for treating citrus greening disease, comprising, on a dose per tree basis: 22g of the extract of dried kiwi fruit rhizome, 25ppm of gibberellin GA325ppm of 6-benzylaminopurine, 0.1ppm of brassinolide.
And (3) formula 7:
a medicament for treating citrus greening disease, comprising, on a dose per tree basis: 2g of the extract of dried root and stem of Kiwi berry, 25ppm of gibberellin GA325ppm of 6-benzylaminopurine, 0.1ppm of brassinolide.
And (4) formula 8:
a medicament for treating citrus greening disease, comprising, on a dose per tree basis: 10ppm of tea saponin, 25ppm of gibberellin GA325ppm of 6-benzylaminoPurine, 0.1ppm brassinolide.
Formula 9:
a medicament for treating citrus greening disease, comprising, on a dose per tree basis: 10g of extract of dried gynura segetum, 25ppm of gibberellin GA325ppm of 6-benzylaminopurine, 0.1ppm of brassinolide.
2. Method of treatment
A method for treating citrus huanglongbing comprising the steps of:
s1, preparing a plant growth hormone solution: firstly, accurately weighing gibberellin, zeatin and brassinolide, adding water, stirring to dissolve, and then respectively preparing mother liquor of 5.00 g/kg;
s2, taking dried kiwi fruit rhizome materials, grinding the dried kiwi fruit rhizome materials into fine powder, and adding ethanol for extraction for 48 hours to obtain an kiwi fruit rhizome extracting solution;
s3, accurately weighing gibberellin mother liquor, zeatin mother liquor and brassinolide mother liquor, mixing the gibberellin mother liquor, zeatin mother liquor and brassinolide mother liquor with the kiwi fruit root and stem extracting solution, and adding water to dilute to a required concentration to obtain a therapeutic agent;
s4, filling the therapeutic agent into an infusion bag, drilling a hole at the height of 50cm of the trunk of the tree, inserting an infusion needle, and inputting the therapeutic agent into the trunk of the tree for treatment.
3. Time and place of experiment
(1) Test site: classic demonstration navel orange garden in Yudu county of Jiangxi province
(2) Test time: 4 months in 2017-2020 and 5 months
(3) The test varieties are as follows: new-lotus navel orange with age of 6-8 years
(4) Classifying disease trees: firstly, dividing the Huanglongbing trees into 5 disease levels according to the following classification standard of the citrus Huanglongbing disease, wherein:
level 1: 5-6 tip leaves on the tree present mottle yellowing symptoms;
and 2, stage: partial lateral branches or main branches have mottled yellowing symptoms, which account for less than one third of the whole tree;
and 3, level: the lateral branches or main branches with mottled etiolated leaves account for 1/3-2/3 of the whole tree;
4, level: the branches with the symptom of the yellow dragon disease account for more than two thirds;
and 5, stage: the whole tree is close to death.
Second, small test
Performing a small test in a typical demonstration navel orange garden in Yudu county, Jiangxi province, selecting 10 navel orange trees (severe trees with disease in 4-5 level) with the same tree age and the same field disease degree basically similar to that of the navel orange trees, injecting solutions for radically treating citrus yellow dragon disease into 5 of the navel orange trees, wherein the numbers of the solutions are treatment 1, treatment 2, treatment 3, treatment 4 and treatment 5, and continuously treating the navel orange trees for 30 days by using the treatment agent and the treatment method of the formula 3; the remaining 5 plants served as control, 2 of which were not treated, numbered as control 1 and control 2, and the remaining 3 plants used only kiwifruit rhizome extract as therapeutic agent. And observing, photographing and recording results.
And (3) fluorescent quantitative PCR detection: and (3) dynamically detecting the pathogenic content of the yellow dragon disease of the leaf of the navel orange before, during and after the test by adopting a 96 real-time fluorescent quantitative PCR instrument. The method for selecting the navel orange tree leaves comprises the following steps: randomly taking one tip branch in the south, east and west of the navel orange tree, and ensuring that five leaves can be detected for each branch. The results of the test data are shown in table 1.
TABLE 1
(Note: when the CT value is more than 35, the Huanglong pathogen is judged to be completely cleared and negative)
The navel orange trees of the treatment group all generate callus, which indicates that no rejection reaction occurs and has good tolerability effect.
Compared with control groups 1-5, the Huanglongbing trees treated by the method of the invention are all converted into negative in PCR detection. The results show that the treatment medicament and the treatment method have obvious curative effect on the huanglongbing.
And thirdly, parallel experiment:
selecting severe trees with disease states of 4-5 grades, dividing navel orange trees into seven groups, numbering 10 diseased trees in each group, continuously treating for a period of time by using the treatment agents and treatment methods of formulas 1-9 respectively, observing, photographing and recording results (the data in each group is the average value of the detection results of 10 diseased trees), and screening out a better formula.
And (3) fluorescent quantitative PCR detection: and (3) dynamically detecting the pathogen content of the yellow dragon disease of the plant leaves before, during and after the test by using a 96 real-time fluorescent quantitative PCR instrument. The test data are shown in Table 1.
TABLE 2
(Note: when the CT value is more than 35, the Huanglong pathogen is judged to be completely cleared and negative)
The navel orange trees treated by the treatment groups 11, 13 and 14 have rejection reaction, the treatment part of the trunk generates colloidal substances, and the therapeutic agents have poor compatibility. The theasaponin and the gynura segetum have the characteristics of resisting bacteria, viruses and diseases, but have poor compatibility in the navel orange trees, and are not suitable for being used as a therapeutic agent for treating yellow dragon disease of the navel orange trees. The navel orange trees of the other treatment groups all generate callus, which indicates that no rejection reaction occurs and has good tolerability effect.
Experiments show that the therapeutic agent with the formula 1-5 has a good therapeutic effect on Huanglong disease, and the Huanglong pathogen in the navel orange tree is completely eliminated after 1 month of treatment. After 1 and 2 years of treatment, the tree grows well, and the newly born leaves of the spring shoots are all green and have no yellowing and mottle phenomena; the phenomenon that the navel oranges are infected by yellow dragon disease bacteria again is not found, the yellow dragon disease bacteria are not detected yet, and the phenomena of relapse and reinfection are not generated; the medicament formula has stable and reliable treatment effect on the huanglongbing. In addition, the Huanglongbing pathogens in the navel orange trees treated by 8 groups (formula 3) are completely eliminated after 1 month of application, and the cure effect of the Huanglongbing pathogens is the best.
Fourthly, verification experiment:
1. longitudinal experiment
Navel orange trees suffering from different degrees of huanglongbing are selected and divided into three groups: the disease states of severe trees with the disease states of 4-5 levels, moderate trees with the disease states of 2-3 levels and mild trees with the disease states of 1 level, 10 trees in each group, treatment numbers of three groups of 10 trees are respectively treatment 15, treatment 16 and treatment 17, and the formula 3 is used as a treatment medicament for treatment. The pathogen content of the yellow dragon disease of each navel orange tree is detected by fluorescent quantitative PCR, and the result is listed in Table 3.
TABLE 3
(Note: when the CT value is more than 35, the Huanglong pathogen is judged to be completely cleared and negative)
The results show that the treatment medicament and the treatment method have good treatment effect on navel orange diseased trees with different disease degrees; the disease of the huanglongbing disease trees is completely cleared after the huanglongbing disease trees are continuously treated for 1 month. Detecting yellow dragon germs on all the treated navel orange trees 1 year and 2 years after the treatment, and observing tree vigor of the tree bodies. The detection result is as follows: the tree grows well, and the newly born leaves of the spring shoots are all green and have no yellowing and mottle phenomena; the phenomenon that the navel oranges are infected by yellow dragon germs again is not found.
2. Repeat the experiment
The experiment is repeated every year from the newly diseased tree in the current year, and the treated navel orange tree is a severe tree with disease in 4-5 grades. The treatment start times were: the treatment method comprises the following steps of respectively naming treatment 18, treatment 19, treatment 20, treatment 21 and treatment 22 on 15 days in 2017, 5 and 5 days in 2018, 25 days in 7, 5 and 2 days in 2019 and 5 and 18 days in 2020. The results are shown in Table 4.
TABLE 4
(Note: when the CT value is more than 35, the Huanglong pathogen is judged to be completely cleared and negative)
The results show that the treatment agent and the treatment method have good treatment effect on the navel orange severe disease trees; the disease of the huanglongbing disease trees is completely cleared after the huanglongbing disease trees are continuously treated for 1 month. Detecting yellow dragon germs on all the treated navel orange trees 1 year and 2 years after the treatment, and observing tree vigor of the tree bodies. The detection result is as follows: the tree grows well, and the newly born leaves of the spring shoots are all green and have no yellowing and mottle phenomena; the phenomenon that the navel oranges are infected by yellow dragon germs again is not found.
Finally, it should be noted that: although the present invention has been described in detail with reference to the foregoing embodiments, it will be apparent to those skilled in the art that modifications may be made to the embodiments or portions thereof without departing from the spirit and scope of the invention.
Claims (3)
1. A method for treating citrus huanglongbing, comprising the steps of:
s1, preparing a plant growth hormone solution, wherein the plant growth hormone comprises gibberellin, cytokinin and brassinolide;
s2, taking dried kiwi fruit rootstock, adding ethanol for extraction to prepare an kiwi fruit rootstock extracting solution;
s3, mixing a kiwi root and stem extracting solution prepared by using 3-18 g of kiwi root and stem dry material for each tree with a plant growth hormone solution, diluting to a required concentration to obtain a therapeutic agent, wherein the concentration of gibberellin is 5-80 ppm, the concentration of cytokinin is 5-80 ppm, and the concentration of brassinolide is 0.01-1 ppm, applying medicine by adopting a trunk infusion method, drilling holes at the height of 30-100 cm of a trunk of the tree, inserting an infusion needle, and inputting the therapeutic agent into the trunk of the tree for treatment.
2. The method for treating citrus huanglongbing as claimed in claim 1, wherein said cytokinin is one or more of 6-benzylaminopurine, kinetin, zeatin, dihydrozeatin, isopentenyladenine, zeatin nucleoside, isopentenyladenosine.
3. The method for treating citrus huanglongbing as claimed in claim 1, wherein said gibberellin is GA1、GA3、GA4、GA9、GA20One or more of (a).
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