CN105483053B - A kind of microbial composite bacteria and its preparation method and application for preventing and treating Citrus Huanglongbing pathogen - Google Patents
A kind of microbial composite bacteria and its preparation method and application for preventing and treating Citrus Huanglongbing pathogen Download PDFInfo
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- CN105483053B CN105483053B CN201610000686.3A CN201610000686A CN105483053B CN 105483053 B CN105483053 B CN 105483053B CN 201610000686 A CN201610000686 A CN 201610000686A CN 105483053 B CN105483053 B CN 105483053B
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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
- C12N1/20—Bacteria; Culture media therefor
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G13/00—Protecting plants
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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
- 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
Abstract
The present invention provides a kind of microbial composite bacterias and preparation method thereof and its application in terms of preventing and treating Citrus Huanglongbing pathogen.The biological deposits number of the microbial composite bacteria, including Ke Liben series bacillus and bacillus subtilis, the Ke Liben series bacillus is CGMCC NO.7996, and microbial composite bacteria specifically can be microbe composite bacterial liquid or complex microbial inoculum.The microbial composite bacteria preparation method, specific steps are as follows: mixed in proportion after Ke Liben series bacillus and fermentation of bacillus subtilis.The present invention from root pouring root and foliar spray mode by inhibiting the reproduction of gramnegative bacterium to multiply, " fungus treatment ", harmful bacteria of soil is killed, optimizes soil property, rhizome phloem sieve tube can be dredged, it solves nutriment and conveys problem on obstacle, restore diseased plant nutrient, promotes new root, make tree root more flourishing, tree cell tubulose is more unobstructed, turns the yellow leaves of the fruit trees into green, health is newly slightly grown.
Description
Technical field
The present invention relates to field of biological control, in particular to a kind of microbial composite bacteria and its system for preventing and treating Citrus Huanglongbing pathogen
Preparation Method and application.
Background technique
Citrus is the most fruit of world wide production, has had more than 4000 years cultivation histories in China.China mandarin orange in 2009
Tangerine cultivated area and yield are more than Brazil, the U.S., are ranked the first in the world, but while Citrus Industry rapidly develops, Huanglong
Disease, which has become, endangers destructive disease the most serious on Orange Producing.Early in the mid-eighteenth century, just there is relevant report in India.
Reinking has been reported for the first time that South China of China, since over half a century, Citrus Huanglongbing pathogen is tight the twenties in 19th century
The development in Guangdong, Guangxi, 3 provinces and regions Citrus Industry of Fujian is constrained again.Late 1970s, Sichuan Province Xichang Region and ferry
City, Ganzhou area and Yunnan Province obtain some areas and also successively find Citrus Huanglongbing pathogen, to mid-term the 1980s, mandarin orange
Tangerine yellow twig is in the citrus producing region extensive widespread in China Guangdong, Guangxi, Fujian, Hainan and Taiwan, and in Zhejiang, Guizhou, lake
Southern to occur in succession, existing 11 of 19 provinces, the area of China's citriculture is endangered, and injured area accounts for the total cultivated area of citrus
80% or more, yield accounts for 85% of total output or so.SaoPaulo State,Brazils in 2004 and Fla. Huanglong in 2005
It is sick to occur in succession, cause the fear of citrus practitioner.So far, yellow twig has been distributed widely in Asia, Africa, Oceania, South America
More than 40 countries in continent and North America.
Young sprout, tender leaf, flower and the fruit of yellow twig main harm citrus.There are a small number of young sprouts in dark green tree crown in initial stage
Yellow is commonly called as " slotting golden flower ", " chicken head is yellow ".There is three kinds of uniform yellow, mottled type yellow and the yellow of nutritional deficiency type symptom classes in blade
Type.Mottled yellow blade and " red nose fruit " are the symptoms of most allusion quotation, can be used as the main foundation of the Morphological Identification disease.?
Field sick tree investigate when, the childhood tree of non-result using mottled type yellow blade as diagnosis basis, bearing-age tree using " red nose fruit " as
Diagnosis basis.
The prior art generallys use the method for integrated control to control the disease: first is that stringent plant quarantine, from nursery stock
It checks on；Second is that cultivating and promoting virus-free nursery stock；Third is that excavating diseased plant in time, reinforce orchard management；Fourth is that passing through physics and change
Prescription formula of learning to farm prevents and treats diaphorina citri, cuts off the infection sources.But these methods " palliative " do not solve to ask from cause of disease depths
Topic, currently available technology lack the feasible method that effectively can directly reduce or eliminate cause of disease in diseased plant.
Summary of the invention:
In view of this, effectively can directly reduce or eliminate Citrus Huanglongbing pathogen diseased plant the purpose of the present invention is to provide a kind of
The microbial composite bacteria and its preparation method and application of middle cause of disease.
In order to achieve the above-mentioned object of the invention, the present invention the following technical schemes are provided:
It is described the present invention provides a kind of microbial composite bacteria, including Ke Liben series bacillus and bacillus subtilis
The biological deposits number of Ke Liben series bacillus is CGMCC NO.7996, and the microbial composite bacteria is that microorganism is compound
Bacterium solution or complex microbial inoculum.
Preferably, the volume ratio of Ke Liben series bacillus and bacillus subtilis is in the microbe composite bacterial liquid
(20~60): (5~8)；
Preferably, in the microbe composite bacterial liquid Ke Liben series bacillus total viable count be 50~10,000,000,000/
Ml, the total viable count of bacillus subtilis are 50~10,000,000,000/ml.
Preferably, the weight ratio of Ke Liben series bacillus and bacillus subtilis is in the complex microbial inoculum
(5~80): (10~80).
Preferably, the total viable count of Ke Liben series bacillus is 50~20,000,000,000/g in the complex microbial inoculum,
The total viable count of bacillus subtilis is 60~15,000,000,000/g.
The present invention also provides a kind of preparation methods of microbe composite bacterial liquid, comprising the following steps:
A1) the fermentation of Ke Liben series bacillus, the Ke Liben series bacillus fermentation condition are as follows: Liquid Culture
Base, temperature is 29~30 DEG C, incubation time is 48~120 hours, is fermented after fermentation 72~96 hours every detection in 2~4 hours
Liquid total viable count, when total viable count up to 50~10,000,000,000/ml stop ferment, obtain Ke Liben series bacillus fermentation liquid；
A2) the fermentation of bacillus subtilis, the fermentation condition of the bacillus subtilis seed are as follows: fluid nutrient medium,
Temperature is 29~30 DEG C, the time 28~48 hours, and the viable bacteria for detecting fermentation liquid after fermentation 28~32 hours every 2~4 hours is total
Number stops fermentation when total viable count is up to 50~10,000,000,000/ml and obtains bacillus subtilis fermentation liquor；
Above two fermentation liquid is aseptically (20~60) by the a3) mixing of fermentation liquid by volume: (50~
80) ratio is mixed to get microbe composite bacterial liquid；
Step a1) and step a2) between without time sequencing limit.
The present invention also provides a kind of preparation methods of complex microbial inoculum, comprising the following steps:
B1) the fermentation of Ke Liben series bacillus, the Ke Liben series bacillus fermentation condition are as follows: temperature be 29~
30 DEG C, incubation time be 48~120 hours, every 2~4 hours detection fermentation liquid total viable counts after fermentation 72~96 hours, when
Total viable count stops fermenting up to 50~10,000,000,000/ml, obtains Ke Liben series bacillus fermentation liquid；
B2) the fermentation of bacillus subtilis, the fermentation condition of the bacillus subtilis seed are as follows: temperature is 29~30
DEG C, incubation time 28~48 hours, fermentation 28~32 hours after every 2~4 hours detection fermentation liquid total viable count, work as viable bacteria
Stop fermentation when sum is up to 50~10,000,000,000/ml and obtains bacillus subtilis fermentation liquor；
Above two fermentation liquid is aseptically 1: 0.5~1.5 ratio by the b3) mixing of fermentation liquid by volume
Example is mixed to get microbe composite bacterial liquid；
B4) the drying of microbe composite bacterial liquid will obtain micro- life after the drying of microbe composite bacterial liquid obtained in step b3)
Object composite bacteria agent；
The step b1) and step b2) between without time sequencing limit.
Preferably, step b4) described in drying be spray drying, the outlet temperature of spray drying is 90~95 DEG C, is entered
Mouth temperature is 145~150 DEG C,
Preferably, 1000~1500ml/h of pan feeding speed of the spray drying.
Preferably, spray drying described in step 4) need to be added protective agent, and the protective agent is skimmed milk power or sucrose,
Protectant concentration is 15~25%.
The present invention also provides application of the microbial composite bacteria in terms of preventing and treating Citrus Huanglongbing pathogen, micro- lifes
The application method of object compound bacteria is root filling or spraying；Described filling specifically: every plant of tree uses 0.5~1kg compound bacteria, pours in root
At portion 15-25 centimeters；It is described spraying specifically: every plant of tree uses 0.5~1kg of compound bacteria, after being watered 3~5kg, spraying blade.
Technical principle of the invention: Ke Liben series bacillus generates the antibacterials such as purine analogue during the fermentation
Matter, shape dominant microflora, inhibits the growth of harmful bacteria in the soil；Generate a large amount of exocellular polysaccharide and small during the growth process simultaneously
Molecule protein can be colonized in the fruit tree tip of a root and be formed " biomembrane ", and the intercellular spaces accumulation outside the vascular cylinder of root, produce
Raw " biomembrane " can greatly accelerate that fruit tree absorbs the process of nutriment.Bacillus subtilis is given birth to by nutrient competition
Long breeding can quickly form one layer of height in plant surface to occupy the mode of living space to prevent the growth of phytopathogen
Privacy protection film, makes pathogen cannot get living space, so that cover crop endangers from pathogen.
Beneficial effects of the present invention: microbial composite bacteria of the present invention can effectively prevent Citrus Huanglongbing pathogen, lead to
The reproduction procreation for inhibiting the gramnegative bacterium of citrus diseased plant root and leaf portion is crossed, reaches " fungus treatment ", kills harmful bacteria
Purpose, using microbial composite bacteria of the present invention can make mandarin tree yellow leaf turn it is green, yellow symptom disappear, the mandarin orange of test group
Tangerine yellow twig disease incidence is significantly lower than conventional treatment, is reduced to 35.6%~67.5% by the disease incidence of conventional treatment 75.1%,
Disease incidence highest reduces 52% compared with conventional treatment, has the effect of significantly preventing and treating yellow twig, the yield of test group compared with
Conventional treatment highest increases by 90 kgs/acre, and rate of growth is up to 30%, has significant effect of increasing production.
Detailed description of the invention
Fig. 1 is the control efficiency histogram for the trees disease incidence that the embodiment of the present invention 3 obtains；
Fig. 2 is that the tree yield that the embodiment of the present invention 3 obtains counts histogram.
Specific embodiment:
It is described the present invention provides a kind of microbial composite bacteria, including Ke Liben series bacillus and bacillus subtilis
The biological deposits number of Ke Liben series bacillus is CGMCC NO.7996.
Microbial composite bacteria provided by the invention includes bacillus subtilis, and the present invention carrys out the bacillus subtilis
Source is not particularly limited, using the common commercial goods of bacillus subtilis well known to those skilled in the art.
In the present invention, the microbial composite bacteria is preferably microbe composite bacterial liquid.When provided by the invention to be micro-
When biological compound fungi liquid, the volume ratio of Ke Liben series bacillus and bacillus subtilis is excellent in the microbe composite bacterial liquid
It is selected as (20~60): (50~80), more preferably (30~50)；(55~75), most preferably (35~45): (60~70).
The total viable count of Ke Liben series bacillus is preferably 50~10,000,000,000/ml in the microbe composite bacterial liquid, more preferably
80~10,000,000,000/ml, most preferably 85~9,500,000,000/ml；The total viable count of bacillus subtilis in the microbe composite bacterial liquid
Preferably 50~10,000,000,000/ml, more preferably 70~9,500,000,000/ml, most preferably 80~9,000,000,000/ml.
In the present invention, the microbial composite bacteria is preferably complex microbial inoculum.When provided by the invention for micro- life
When object composite bacteria agent, the weight ratio of Ke Liben series bacillus and bacillus subtilis is preferred in the complex microbial inoculum
For (5~80): (10~80), more preferably (40~60): (45~65), most preferably 50: 55.The microbial composite bacteria
The total viable count of Ke Liben series bacillus is preferably 50~20,000,000,000/g, more preferably 100~15,000,000,000/g in agent, most preferably
For 12,500,000,000/g；The total viable count of bacillus subtilis is preferably 60~15,000,000,000/g in the complex microbial inoculum, more excellent
It is selected as 80~13,000,000,000/g, most preferably 10,000,000,000/g.
When it is provided by the invention be microbe composite bacterial liquid when, the present invention provides a kind of preparations of microbe composite bacterial liquid
Method, comprising the following steps:
A1) the fermentation of Ke Liben series bacillus；
A2) the fermentation of bacillus subtilis；
A3) the mixing of fermentation liquid；
Step a1) and step a2) between without time sequencing limit.
The present invention ferments Ke Liben series bacillus, obtains Ke Liben series bacillus fermentation liquid.In the present invention
In, the Ke Liben series bacillus fermentation is specific are as follows:
Ke Liben series bacillus strain is subjected to activation culture；
Strain after the activation culture is inoculated in fluid nutrient medium and is carried out liquid fermentation, Ke Liben class gemma is obtained
Bacillus fermentation liquid.
The Ke Liben series bacillus strain of preservation is preferably inoculated in nutrient agar and activates by the present invention
Culture.The present invention does not have special limitation to the container of the activation culture, is trained using activation well known to those skilled in the art
Container is supported, eggplant type bottle can be such as used.In the present invention, the temperature of the activation culture is preferably 29~30 DEG C；It is described
The time of activation culture is preferably 8~12h, more preferably 10h.
In the present invention, the nutrient agar preferably includes following components: 10 parts of peptone, 3 parts of beef extract powder,
5 parts of sodium chloride, 15 parts of agar.The nutrient agar final pH is 7.1~7.5.
After completing the activation culture, the Ke Liben series bacillus after activation culture is inoculated in Liquid Culture by the present invention
Liquid Culture is carried out in base.In the present invention, the fluid nutrient medium inoculates Ke Liben series bacillus after preferably sterilizing.?
In the present invention, the sterilizing is preferably high-temperature sterilization, and the temperature of the sterilizing is preferably 120~130 DEG C, and more preferably 121 DEG C；
The time of the sterilizing is preferably 30-45min, more preferably 38min.
The present invention does not have special restriction to the container of the liquid fermentation, using fermentation well known to those skilled in the art
Equipment, such as can be fermentor.In the present invention, the temperature of the liquid fermentation is preferably 29~30 DEG C, more preferably
It is 30 DEG C；The pH value of the liquid fermentation is preferably 6.8-7.0, and more preferably 6.9；The pressure of the liquid fermentation is preferred
For 0.08~0.12MPa, more preferably 0.09~0.11MpPa；Ventilatory capacity during the liquid fermentation is preferably 0.8
~1.2V/Vmin, more preferably 1V/Vmin；The liquid fermentation time is preferably 48-120 hours, more preferably
72~84 hours；
The present invention preferably after the liquid fermentation 72~96 hours every detection in 2~4 hours ferment total viable count, with true
The fixed time point for stopping fermentation；Preferably, when total viable count stops fermenting up to 50~10,000,000,000/ml, it is furthermore preferred that when viable bacteria is total
Number stops fermenting up to 80~10,000,000,000/ml, obtains Ke Liben series bacillus fermentation liquid.The present invention is to the fermentation total viable count
The method of detection does not have special limitation, using conventional method of counting well known to those skilled in the art, such as micro- meter
Number, plate count, measurement cell weight etc..
In the present invention, the fluid nutrient medium in the Ke Liben series bacillus fermentation process is preferably included with the following group
Point: sucrose 1-50 parts by weight, sodium nitrate 0.1-5.0 parts by weight, potassium dihydrogen phosphate 1.0-10 parts by weight, magnesium sulfate 0.1-1.0 weight
Measure part, potassium chloride 0.1-2.0 parts by weight, ferric sulfate 0.01-1.0 parts by weight.
Fermentation of bacillus subtilis of the present invention specifically:
The activation of strain；
Strain after activation is inoculated in fluid nutrient medium and is carried out liquid fermentation, the fermentation of Ke Liben series bacillus is obtained
Liquid.
The Bacillus subtilis strain bought preferably is inoculated in nutrient agar and carries out activation culture by the present invention.
The present invention does not have special limitation to the container of the activation culture, using activation culture container well known to those skilled in the art
, can such as use eggplant type bottle.In the present invention, the temperature of the activation culture is preferably 29~30 DEG C；The activation training
The feeding time is preferably 8~12h, more preferably 10h.
In the present invention, the nutrient agar preferably includes following components: 10 parts of peptone, 3 parts of beef extract powder,
5 parts of sodium chloride, 15 parts of agar；The nutrient agar final pH is 7.1~7.5.
After completing the activation culture, the bacillus subtilis after activation culture is inoculated in fluid nutrient medium by the present invention
Carry out Liquid Culture.In the present invention, the fluid nutrient medium inoculates Ke Liben series bacillus after preferably sterilizing；It is described to go out
Bacterium is preferably high-temperature sterilization, and the temperature of the sterilizing is preferably 120~130 DEG C, and more preferably 125 DEG C；The time of the sterilizing
Preferably 30-45min, more preferably 40min.
The present invention does not have special restriction to the container of the liquid fermentation, using fermentation well known to those skilled in the art
Equipment, such as can be fermentor.In the present invention, the temperature of the liquid fermentation is preferably 29~30 DEG C, more preferably
It is 30 DEG C；The pH value of the liquid fermentation is preferably 6.8-7.0, and more preferably 6.9；The pressure of the liquid fermentation is preferred
For 0.05~0.1MPa, more preferably 0.07~0.08MpPa；Ventilatory capacity during the liquid fermentation is preferably 0.8~
1.2V/Vmin, more preferably 1V/Vmin；The liquid fermentation time is preferably 28~48 hours, more preferably 36
~44 hours；
The present invention preferably after the liquid fermentation 28~32 hours every detection in 2~4 hours ferment total viable count, preferably
, when total viable count up to 50~10,000,000,000/ml stop ferment, it is furthermore preferred that when total viable count up to 85~9,500,000,000/ml stop ferment,
Obtain bacillus subtilis fermentation liquor.The present invention does not have special limitation to the method for the fermentation total viable count detection, uses
Routine method of counting well known to those skilled in the art, such as microscopic count, plate count, measurement cell weight
Deng.
In the present invention, the fluid nutrient medium during the fermentation of bacillus subtilis preferably includes following components: egg
White peptone 0.01-0.1 parts by weight, yeast powder 0.01-0.1 parts by weight, sucrose 0.001-0.050 parts by weight, ammonium sulfate 0.001-
0.007 parts by weight, sodium citrate 0.001-0.01 parts by weight, potassium dihydrogen phosphate 0.001-0.01 parts by weight, magnesium sulfate 0.0001-
0.001 parts by weight, ferric sulfate 0.00001-0.0001 parts by weight, manganese sulfate 0.01-0.1 parts by weight.
It is limited between the fermentation of Ke Liben series bacillus and fermentation of bacillus subtilis step without time sequencing in the present invention,
The fermentation for first carrying out Ke Liben series bacillus, then carries out the fermentation of bacillus subtilis again or can first carry out withered grass gemma
The fermentation of bacillus, then carry out the fermentation of Ke Liben series bacillus, then Ke Liben series bacillus and bacillus subtilis
Fermentation simultaneously carry out.
After completing Ke Liben series bacillus fermentation and fermentation of bacillus subtilis, the present invention aseptically,
Preferably it is by volume by the Ke Liben series bacillus fermentation liquid and bacillus subtilis fermentation liquor (20~60): (50
~80) ratio mixing, be more preferably by volume (35~45): the ratio of (60~70) is mixed to get microbial composite bacteria
Liquid；The aseptic condition is realized by ultraviolet superclean bench or sterile workplace.
When the present invention is to provide a kind of complex microbial inoculum, the present invention also provides a kind of complex microbial inoculums
Preparation method, comprising the following steps:
Microbe composite bacterial liquid is prepared according to above-mentioned technical proposal；
The microbe composite bacterial liquid is dry, obtain complex microbial inoculum.
Microbe composite bacterial liquid is prepared according to above-mentioned technical proposal in the present invention, and details are not described herein.
In the present invention, the drying is preferably spray-dried；The outlet temperature of spray drying is preferably 90~95 DEG C, more
Preferably 93 DEG C；Inlet temperature is preferably 145~150 DEG C, and more preferably 148 DEG C；The pan feeding speed of the spray drying
Preferably 1000~1500ml/h, more preferably 1200ml/h；
Preferably protective agent is added during the spray drying in the present invention, to guarantee during spray drying
The higher survival rate of microbial composite bacteria.In the present invention, the protective agent is preferably skimmed milk power or sucrose.The present invention is preferred
Using the protective agent of solution state, the mass concentration of the protection agent solution is preferably 15~25%, and more preferably 20%.
The present invention also provides application of the microbial composite bacteria in terms of preventing and treating Citrus Huanglongbing pathogen, micro- lifes
The application method of object compound bacteria is root filling or spraying；It is preferably specially to pour the microbial composite bacteria in plant root that described, which fills,
At portion 15-25 centimeters, more preferably pour at 18~22 centimeters of root；Every plant of tree preferably uses 0.5~1kg compound bacteria, more
Preferred every plant of tree uses 0.8~0.9kg；Described spraying, every plant of tree preferably uses 0.5~1kg of compound bacteria, is watered 3~5kg
Afterwards, spraying blade, it is furthermore preferred that every plant of tree uses 0.8~0.9kg of compound bacteria, after being watered 4~4.5kg, spraying blade.
Microbial composite bacteria provided by the invention is described in detail below with reference to embodiment, but cannot be them
It is interpreted as limiting the scope of the present invention.
Embodiment 1
The preparation of Ke Liben series bacillus fermentation liquid is seeded in 121 DEG C after Ke Liben series bacillus is expanded culture
In Liquid Culture after sterilizing 35min, the Liquid Culture based component is 30 parts by weight of sucrose, 1.5 parts by weight of sodium nitrate, phosphoric acid
1.0 parts by weight of potassium dihydrogen, 0.5 parts by weight of magnesium sulfate, 0.5 parts by weight of potassium chloride, 0.01 parts by weight of ferric sulfate.It sends out in the fermenter
Ferment, fermentation condition: temperature is 30 DEG C, pH value 6.9, pressure inside the tank 0.1MPa, ferment ventilatory capacity maintenance 1v/vmin, time
Every the total viable count of 2 hours method detection fermentation liquids with micro- detection after 76 hours, fermentation 72 hours, when total viable count reaches
9600000000/ml stops fermentation and obtains Ke Liben series bacillus fermentation liquid.
Embodiment 2
The preparation of bacillus subtilis fermentation liquor is seeded in 121 DEG C of sterilizing 40min after bacillus subtilis is expanded culture
In Liquid Culture afterwards, the Liquid Culture based component is 0.03 parts by weight of peptone, 0.03 parts by weight of yeast powder, sucrose
0.025 parts by weight, 0.007 parts by weight of ammonium sulfate, 0.003 parts by weight of trisodium citrate, 0.003 parts by weight of potassium dihydrogen phosphate, sulphur
Sour 0.0005 parts by weight of magnesium, 0.00005 parts by weight of ferric sulfate, 0.01 parts by weight of manganese sulfate.It ferments in the fermenter, ferment item
Part: temperature is 29 DEG C, pH value 6.8, pressure inside the tank 0.07MPa, ferment ventilatory capacity maintenance 1v/vmin, the time 36 hours, hair
Every 2 hours detection fermentation liquid total viable counts after ferment 28 hours, withered grass gemma is obtained when total viable count stops fermentation up to 9,500,000,000/ml
Bacillus fermentation liquid.
Embodiment 3
The preparation of complex microbial inoculum, by Ke Liben series bacillus fermentation liquid obtained in embodiment 1 and embodiment 2
Obtained in bacillus subtilis fermentation liquor, in sterile working room press Ke Liben series bacillus fermentation liquid and bacillus subtilis
The ratio that fermented liquid volume ratio is 1: 0.83 is mixed to get microbe composite bacterial liquid.Obtained microbe composite bacterial liquid is carried out
Spray drying, the outlet temperature of spray drying are 94 DEG C, and inlet temperature is 148 DEG C, and pan feeding speed is 1200ml/h, spray drying
The skimmed milk power of Shi Tianjia 21% obtains complex microbial inoculum as protective agent.
Embodiment 4
Microbial composite bacteria of the present invention detects the control efficiency of Citrus Huanglongbing pathogen, is tested on January 1st, 2014
It was carried out to December 31 in Ganzhou City Anyuan County high hill back navel orange fruit industry base, the morbidity of test area yellow twig is heavier, using filling
The mode of root and foliar spray carries out.It is divided into 4 groups of processing, handles 1kg/ plants of 1 pouring root, handle 1.5kg/ plants of 2 pouring root, processing 3 fills
0.5kg/ plants of 1kg/ plants+foliar spray of root, processing 4 Routine controls (ck) i.e. root fill 1.5kg/ plants of clear water, and root-irrigation method is will
The microbe composite bacterial liquid or clear water pours at every plant of trees ' root 15-25 centimetre, fills primary every 30 Tiangengs, blade face
Spraying method is that every plant of tree uses complex microbial inoculum 0.5kg, after being watered 4kg, spraying blade, and the spraying blade every 30 days
Once.
Control efficiency is shown in Fig. 1, and the Citrus Huanglongbing pathogen disease incidence of processing 1~3 is significantly lower than conventional treatment, by conventional treatment
75.1% disease incidence is reduced to 35.6%~67.5%, and disease incidence highest reduces 52% compared with conventional treatment, has aobvious
The effect of the prevention and treatment yellow twig of work.
Effect of increasing production is shown in Fig. 2, and yield increases to 390 kgs/acre by processed conventionally 300 kgs/acre, the yield of test group
Increase by 90 kgs/acre compared with conventional treatment highest, rate of growth is up to 30%, has significant effect of increasing production.
The above is only a preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also answered
It is considered as protection scope of the present invention.
Claims (1)
1. a kind of application of microbial composite bacteria in terms of preventing and treating Citrus Huanglongbing pathogen；
The application method of the microbial composite bacteria is root filling+foliar spray；
Described filling specifically: every plant of tree uses 0.5~1kg microbe composite bacterial liquid, pours at the 15-25 centimetre of root；
It is described spraying specifically: every plant of tree uses 0.5~1kg of composite bacteria agent, after being watered 3~5kg, spraying blade；
The microbial composite bacteria includes Ke Liben series bacillus and bacillus subtilis, the Ke Liben series bacillus
Biological deposits number is CGMCCNO.7996；
The volume ratio of Ke Liben series bacillus and bacillus subtilis is (20~60) in the microbe composite bacterial liquid:
(50~80)；The total viable count of Ke Liben series bacillus is 50~10,000,000,000/ml, withered grass in the microbe composite bacterial liquid
The total viable count of bacillus is 50~10,000,000,000/ml；
The weight ratio of Ke Liben series bacillus and bacillus subtilis is (5~80) in the complex microbial inoculum: (10
~80)；The total viable count of Ke Liben series bacillus is 50~20,000,000,000/g, withered grass gemma in the complex microbial inoculum
The total viable count of bacillus is 60~15,000,000,000/g.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610000686.3A CN105483053B (en) | 2016-01-04 | 2016-01-04 | A kind of microbial composite bacteria and its preparation method and application for preventing and treating Citrus Huanglongbing pathogen |
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|---|---|---|---|
| CN201610000686.3A CN105483053B (en) | 2016-01-04 | 2016-01-04 | A kind of microbial composite bacteria and its preparation method and application for preventing and treating Citrus Huanglongbing pathogen |
Publications (2)
| Publication Number | Publication Date |
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| CN106350464A (en) * | 2016-08-29 | 2017-01-25 | 佛山市艳晖生物科技有限公司 | Microbial agent for preventing and treating cumquat huanglongbing and preparation method thereof |
| CN106719633A (en) * | 2016-08-30 | 2017-05-31 | 江西天人生态股份有限公司 | A kind of Ke Liben series bacillus wettable powder |
| CN106417381A (en) * | 2016-09-23 | 2017-02-22 | 江西天祥通用航空股份有限公司 | Paenibacillus kribbensis oil suspension and preparation method thereof |
| CN108719334A (en) * | 2017-04-19 | 2018-11-02 | 柏玉兰 | The antimicrobial plant vaccine and production method of yellow twig can be prevented |
| CN108220185B (en) * | 2017-12-19 | 2021-08-17 | 华中农业大学 | Application of biological control agent in removing citrus sapling huanglongbing pathogen |
| CN108294049B (en) * | 2018-01-19 | 2019-07-19 | 江西省科学院微生物研究所 | A kind of Paenibacillus polymyxa culture solution combination drug and preparation method thereof for treating Citrus Huanglongbing pathogen |
| CN109601237A (en) * | 2019-02-20 | 2019-04-12 | 易之泰生物科技（龙岩）有限公司 | The control method of Citrus Huanglongbing pathogen |
| CN110117561B (en) * | 2019-05-05 | 2020-07-28 | 中铁十八局集团有限公司 | Compound biological agent for increasing survival rate of new trees and preparation method |
| CN111837766B (en) * | 2020-07-27 | 2021-09-28 | 华中农业大学 | Method for controlling pathogen of seedling phlobacterium phlogistii by composite microorganism treatment and application |
| CN111793498A (en) * | 2020-07-31 | 2020-10-20 | 福建三炬生物科技股份有限公司 | Microbial agent and preparation method and application thereof |
| CN116496941B (en) * | 2023-04-04 | 2024-01-16 | 华中农业大学 | Microbial fermentation compound and application thereof in preventing and treating citrus yellow dragon disease |
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