CN1221659C - Waxy bacillus strain and microbial bactericide producing process with the strain - Google Patents
Waxy bacillus strain and microbial bactericide producing process with the strain Download PDFInfo
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- CN1221659C CN1221659C CN 02135876 CN02135876A CN1221659C CN 1221659 C CN1221659 C CN 1221659C CN 02135876 CN02135876 CN 02135876 CN 02135876 A CN02135876 A CN 02135876A CN 1221659 C CN1221659 C CN 1221659C
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Abstract
The present invention relates to a bacterial strain of bacillus cereus, and a production method thereof for preparing microbial germicide. The presentation of the bacterial strain of the bacillus cereus is bacillus cereus 98-I. The bacterial strain has good broad spectrum and high antibacterial activity, and has the functions of promoting the growth of various plants, promoting the germination of seeds and inducing the resistance of plants. In the production method of the microbial germicide by the bacterial strain, the process flow comprises: preparing the bacterial strain in an inclined plane, preparing the bacterial strain in a shaking table, preparing and fermenting the first level bacterial strain in a big tank, and providing the relevant parameters of each processing step. The broad application of the microbial germicide determines the universality and the broad spectrum of the popularization of the microbial germicide; the microbial germicide prevents soil-borne diseases of vegetables and melons, has an effect on controlling the soil-borne diseases of flowers and lawn, and has a function of promoting growth. The microbial germicide has good economical and social benefits, and conforms to the requirements of ecological agriculture and sustainable agriculture.
Description
Technical field
The present invention relates to a kind of bacterial isolates, be specially a kind of bacterial strain of bacillus cereus, relate to the production method for preparing microbial bactericide with this bacterial strain simultaneously.
Background technology
Bacillus cereus (Bacillus cereus) belongs to bacillus.At present, there is the different strains of wax shape (matter) genus bacillus to be used for the bibliographical information of phytopathogen, disease control and growth-promoting functions both at home and abroad, domestic as the inhibition of B.C.sp to rice warm disease bacterium, B.C.S14 is to the control and the growth-promoting functions of cotton disease, B.C.R2 is to the control of rice sheath blight disease, B.C.752 has growth-promoting functions and the germ spore is had the effect of clearing up paddy rice, watermelon, corn, B.C.CS51 has antagonistic action to the narcissus brown spot, B.C.B017 can promote the apple seedling growth, and B.C has antagonistic action etc. to gaeumannomyces graminis and cotton wilt; External as bacillus cereus B.C.uw85, B.C.S18, B.C.W35, B.C.28, B.C.B
0, bacterial strain such as B.C.DGA34, B.C.MS1-9, B.C.AS4-12 is mainly to the control of tomato, cucumber samping off (phytophthora rot).Every kind of bacterial strain of existing bacillus cereus is mainly to a kind of disease of plant, have preventive and therapeutic effect to two kinds of diseases individually, growth-promoting functions also is that each bacterial strain works to indivedual 1-2 kind of plant, therefore, its disease species that can prevent and treat is single, growth-promoting functions at plant or the kind of crop single, broad spectrum is relatively poor.
Summary of the invention
The invention provides a kind of bacterial strain of bacillus cereus, and provide the production method for preparing microbial bactericide with this bacterial strain.
The present invention adopts following technical scheme to realize: a kind of bacterial strain of bacillus cereus, this bacterial strain are expressed as bacillus cereus (Bacillus cereus) 98-I.This bacterial strain is at non-patent literature---and the 7th the 2nd phase of volume of calendar year 2001 " using and the environmental organism journal " goes up with the form of paper open; public's desire obtains this bacterial strain and can ask for to the application of Shanxi Province's agricultural chemicals key lab of Inst. of Plant Protection, Shanxi Prov. Academy of Agricaultural Sciences; the holder of this bacterial strain---Shanxi Province's agricultural chemicals key lab of Inst. of Plant Protection, Shanxi Prov. Academy of Agricaultural Sciences guarantees under the condition that meets the legal requirements, and will provide this bacterial strain to the public in 20 years applyings date.
Bacillus cereus of the present invention (Bacillus cereus) 98-I[is hereinafter to be referred as B.C.98-I] research originate from " make compost control and the mechanism research of steeping fluid " to Plant diseases.Find in the research to make compost and exist the bacterial isolates that the various plants pathogenic fungi is had strong antagonistic action in the liquid, from making compost steeping fluid, soil, screen bacteria culture in the plant rhizosphere, collect 330 of active bacterial strains altogether, utilize it to producing common fungal diseases of plants as cucumber fusarium axysporum, the green pepper blight, the green pepper eqpidemic disease, tomato wilt, early blight of tomato, watermelon blight, poinsettia, 8 kinds of diseases such as Cyclamen persicum blight are carried out the screening of plate face-off antagonism bacterium, screening of greenhouse diseases prevention antagonism bacterium and control in field effect test filter out the control vegetables that wide spectrum has growth-promoting functions concurrently at last, melon, antagonism bacterium---the B.C.98-I of soil-borne diseases such as flowers.
The B.C.98-I bacterial strain has following bacteriology characteristic:
1, morphological character
(1) colony characteristics:
This bacterium can grow on PDA, NA substratum, the bacterium colony circle, and the edge is irregular, and the initial stage is glossy, and bacterium colony is yellowish pink, substratum nondiscoloration, slightly peculiar smell.
(2) thalline feature:
Bacillus, general size is 1.0-1.2 * 2.5-4.5 micron, and gemma is arranged, middle life is given birth to time end, does not generally expand positive, the purple of gramstaining.Thalline moves about, peritrichous.
2, biochemical characteristic:
The catalase positive, aerobic growth, the VP positive, producing acid in the VP liquid is 5.6, the egg yolk reaction positive is positive to the resistance of N,O-Diacetylmuramidase.Hydrolyzed starch, reduction nitrate becomes nitrite, decomposes casein, tyrosine, produces acid in the litmus milk, peptonizes.
3, physiological property
Can utilize glucose, N.F,USP MANNITOL to produce acid, utilize citric acid.Pectinose, the negative reaction of wood sugar.PH can grow in 5.7 o'clock, grew in 7% sodium-chlor, and growth temperature 10-45 ℃, high temperature is influential to it more than 60 ℃.
4, bacteriostatic activity
1% concentration (10
8Cfu/ml) inhibiting rate to cucumber fusarium axysporum is 62.6%
1% concentration (10
8Cfu/ml) inhibiting rate to withered germ of water-melon is 80.2%
1% concentration (10
8Cfu/ml) inhibiting rate to the green pepper wilt is 82.4%
1% concentration (10
8Cfu/ml) inhibiting rate to the tomato wilt bacterium is 69.5%
B.C.98-I bacterial strain principal character index and known 44 strain bacillus cereuses are compared (seeing the following form)
| Main proterties | The reaction of 44 strain bacillus cereuses | The reaction of B.C.98-1 antagonism bacterium |
| 1, microscopic morphology | Shaft-like | Shaft-like |
| 2, length | 1.0-1.2 * 3.0-5.0 micron | 1.0-1.2 * 2.5-4.5 micron |
| 3, gemma has or not | Gemma is arranged | Gemma is arranged |
| 4, growth position | The middle end of giving birth to is once given birth to | The middle end of giving birth to is once given birth to |
| 5, expand situation | Generally do not expand | Generally do not expand |
| 6, gramstaining | + | + |
| 7, flagellum growth | Peritrichous | Peritrichous |
| 8, growth temperature | 35-45℃ 10-20℃ | 35-45℃ 10-20℃ |
| 9, catalase | + | + |
| 10, anaerobic growth | + | + |
| 11、VP | + | + |
| 12, produce acid in the VP liquid | 4.3-5.6 | 5.6 |
| 13, egg yolk reaction | + | + |
| 14, to the resistance of N,O-Diacetylmuramidase | + | + |
| 15, in 7% the sodium-chlor | + | + |
| 16, PH is 5.7 o'clock | + | + |
| 17, produce acid from glucose | + | + |
| 18, starch hydrolysis | + | + |
| 19, utilize citric acid | + | + |
| 20, reduction nitrate becomes nitrous acid | + | + |
| 21, decompose casein | + | + |
| 22, tyrosine | + | + |
| 23, in the litmus milk | Produce alkali, produce acid or peptonize individually | Produce acid, peptonize |
| 24, pectinose | - | |
| 25, wood sugar | -, indivedual positive | +、- |
| 26, N.F,USP MANNITOL produces acid | - | + |
| The deoxidation in 7 days of 27 phenylalanines | - | - |
By comparing as can be seen: the principal character index of B.C.98-I bacterial strain and known bacillus cereus strain basic identical, but certain difference being arranged with known bacterial strain, its main difference is selected and is that N.F,USP MANNITOL produces the apparent length range positive, its bacterial strain of acid and slightly dwindles." general the common authentication method of the bacterium " book write according to Institute of Microorganism, Academia Sinica's division bacteria group and above-mentioned bacteria characteristic and compare the result, show that the B.C.98-I bacterial strain belongs to bacillus, the bacillus cereus kind, and this bacterial strain named be bacillus cereus (Bacillus cereus) 98-I.
Bacillus cereus (Bacillus cereus) 98-I has the following advantages:
1, broad spectrum is good.Diseases such as cucumber fusarium axysporum, cucumber downy mildew, green pepper blight, green pepper eqpidemic disease, tomato wilt, watermelon blight, annual bluegrass blight there is tangible prevention effect.Especially aspect the preventing and treating of withered disease, outstanding effect (seeing the following form) is arranged
B.C.98-I antimicrobial range of application short of money and prevention effect
| The disease title | Plate inhibiting rate % | Greenhouse prevention effect % | Control in field effect % |
| Cucumber fusarium axysporum | 71.4 | 80.9 | 86.0 |
| The green pepper blight | 77.1 | 69.2 | 68.2 |
| Tomato wilt | 53.6 | 55.9 | - |
| Watermelon blight | 71.3 | 67.5 | 74.4 |
| The poinsettia blight | 73.2 | - | 79.9 |
2, bacteriostatic activity height.Connecing the bacterium amount is 10
8During cfu/ml, the plate inhibiting rate of cucumber Fusarium oxysporum, green pepper Fusarium oxysporum, tomato wilt and watermelon Fusarium oxysporum was reached more than 60% in 24 hours, reach more than 90% after 72 hours.
3, various plants there is the short effect of giving birth to and promoting seed germination.7 kind of plant are had growth-promoting functions, percentage of germination increase from green pepper 8.9%, cucumber, watermelon 20%, the 50-80% of Chinese cabbage, tomato, to 350% of annual bluegrass.Other antagonism bacterium only has growth-promoting functions to indivedual plants.
4, plant there is the induction of resistance effect.Warp is to watermelon, cucumber, 7 resistance index determinings of green pepper 3 kind of plant, and wherein 4 kinds of enzymes (catalase, phenylalanine transaminase, polyphenoloxidase, β-1,3 dextranase) activity has raising in various degree.
5, wide accommodation.In-18 ℃ and 60 ℃ of scopes, all can survive.Anti-certain salt cash.
6, fermentation period is short.
Another object of the present invention is to provide the production method for preparing microbial bactericide with the B.C.98-I bacterial strain, makes this antimicrobial production short of money send out mass-producing pure, and it is used in biological control on a large scale.
The production method for preparing microbial bactericide with the B.C.98-I bacterial strain, its technological process of production comprises slant strains preparation, shaking table strain preparation, first class inoculum preparation and a big jar of alcohol, and wherein, the processing condition of slant strains preparation are: the PDA substratum, 28 ℃, 48 hours; The processing condition of shaking table strain preparation are: 1. number substratum, 28-42 ℃, shaking speed 150-200 rev/min, 48 hours, 1. number substratum was: potato 20%, glucose 1.5-3.0%, sodium-chlor 0.2-0.5%, potassium primary phosphate 0.1-0.6%, all the other are water; The processing condition of first class inoculum preparation are: 2. number substratum, 28-42 ℃, 150-280 rev/min of fermentor tank rotating speed, 48 hours, pH value 4-10, air flow 1: 1,2. number substratum is: Semen Maydis powder 0.2-0.5%, bean powder 0.2-0.5%, glucose 3-6%, sodium chloride 0.1-0.8%, potassium primary phosphate 0.1-0.8%, ammonium sulfate 0.1-0.8%, peptone 0.1-0.8%, sal epsom 0.01-0.05%, zinc sulfate 0.01-0.05%, all the other are water; The processing condition of bulk fermentation are: 2. number substratum, 28-42 ℃, 150-280 rev/min of fermentor tank rotating speed, 48 hours, pH value 4-10, air flow 1: 1.Big jar is sent out the liquid that obtains behind the alcohol is exactly required product---microbial bactericide.
This production method fermentation period is short.The fermentation flow process was 4 steps, and bacterial strain concentration is more than or equal to 10 in the sterilant
10Cfu/ml can directly apply to control in field.
The broad spectrum of this microbial bactericide has determined popularity and the extensity that it is applied, and not only can prevent vegetables, melon soil-borne disease, and the soil-borne disease on flowers, lawn is also had prevention effect.This sterilant also has growth-promoting functions, is again bacterial manure so it promptly is a sterilant, can be used as the bio-bacterial manure of leaf vegetables plant.This sterilant has strong resistance and adaptability, and it can be applicable under different weather condition, different ecological condition.Have high efficiency at a low price, have good market outlook.
This microbial bactericide has good economic and social benefit.This product input-output ratio is 1: 8, can retrieve economic losses 80 yuan for every mu, if promote 1,000,000 mu every year, 2-3 promotes 3,000,000 mu, can obtain 240,000,000 yuan of economic benefits.This product not only has tangible economic benefit, and has very good social benefit and ecological benefits, meets the requirement of the ecological agriculture, sustainable agriculture.
Embodiment
Embodiment 1
The production method for preparing microbial bactericide with the B.C.98-I bacterial strain, its technological process of production comprises slant strains preparation, shaking table strain preparation, first class inoculum preparation and a big jar of alcohol, and wherein, the processing condition of slant strains preparation are: the PDA substratum, 28 ℃, 48 hours; The processing condition of shaking table strain preparation are: 1. number substratum, and 28 ℃, 150 rev/mins of shaking speed, 48 hours, 1. number substratum was: potato 20%, glucose 1.5%, sodium-chlor 0.2%, potassium primary phosphate 0.1%, all the other are water; The processing condition of first class inoculum preparation are: 2. number substratum, 28 ℃, 150 rev/mins of fermentor tank rotating speeds, 48 hours.PH value 4, air flow 1: 1,2. number substratum is: Semen Maydis powder 0.2%, bean powder 0.2%, glucose 3%, sodium chloride 0.1%, potassium primary phosphate 0.1%, ammonium sulfate 0.1%, peptone 0.1%, sal epsom 0.01%, zinc sulfate 0.01%, all the other are water, the processing condition of bulk fermentation are: 2. number substratum, 28 ℃, 150 rev/mins of fermentor tank rotating speeds, 48 hours.PH value 4, air flow 1: 1.Big jar is sent out the liquid that obtains behind the alcohol is exactly required product---microbial bactericide.
The product forms of present embodiment is an aqua.
Target level of product quality:
1, concentration is more than or equal to 10
10Cfu/ml
2, be 2 * 10 to concentration in the inhibition of cucumber Fusarium oxysporum
7Cfu/ml
To concentration in the inhibition of watermelon Fusarium oxysporum is 1 * 10
7Cfu/ml
To concentration in the inhibition of green pepper Fusarium oxysporum is 0.6 * 10
7Cfu/ml
To concentration in the inhibition of tomato Fusarium oxysporum is 2 * 10
7Cfu/ml
Field using method and concentration:
Fall ill and irritated root once in previous month, concentration is 300-800 times, decides on a preceding stubble state of an illness, and 250ml is irrigated in every strain.
Embodiment 2
The production method for preparing microbial bactericide with the B.C.98-I bacterial strain, its technological process of production comprises slant strains preparation, shaking table strain preparation, first class inoculum preparation and a big jar of alcohol, and wherein, the processing condition of slant strains preparation are: the PDA substratum, 28 ℃, 48 hours; The processing condition of shaking table strain preparation are: 1. number substratum, and 42 ℃, 200 rev/mins of shaking speed, 48 hours, 1. number substratum was: potato 20%, glucose 3.0%, sodium-chlor 0.5%, potassium primary phosphate 0.6%, all the other are water; The processing condition of first class inoculum preparation are: 2. number substratum, 42 ℃, 280 rev/mins of fermentor tank rotating speeds, 48 hours, pH value 10, air flow 1: 1,2. number substratum is: Semen Maydis powder 0.5%, bean powder 0.5%, glucose 6%, sodium chloride 0.8%, potassium primary phosphate 0.8%, ammonium sulfate 0.8%, peptone 0.8%, sal epsom 0.05%, zinc sulfate 0.05%, all the other are water, the processing condition of bulk fermentation are: 2. number substratum, 42 ℃, 280 rev/mins of fermentor tank rotating speeds, 48 hours.PH value 10, air flow 1: 1.Big jar is sent out the liquid that obtains behind the alcohol is exactly required product---microbial bactericide.
Embodiment 3
The production method for preparing microbial bactericide with the B.C.98-I bacterial strain, its technological process of production comprises slant strains preparation, shaking table strain preparation, first class inoculum preparation and a big jar of alcohol, and wherein, the processing condition of slant strains preparation are: the PDA substratum, 28 ℃, 48 hours; The processing condition of shaking table strain preparation are: 1. number substratum, and 35 ℃, 180 rev/mins of shaking speed, 48 hours, 1. number substratum was: potato 20%, glucose 2.0%, sodium-chlor 0.3%, potassium primary phosphate 0.4%, all the other are water; The processing condition of first class inoculum preparation are: 2. number substratum, 35 ℃, 220 rev/mins of fermentor tank rotating speeds, 48 hours, pH value 7, air flow 1: 1,2. number substratum is: Semen Maydis powder 0.4%, bean powder 0.3%, glucose 4%, sodium chloride 0.5%, potassium primary phosphate 0.5%, ammonium sulfate 0.5%, peptone 0.5%, sal epsom 0.03%, zinc sulfate 0.03%, all the other are water.The processing condition of bulk fermentation are: 2. number substratum, 35 ℃, 220 rev/mins of fermentor tank rotating speeds, 48 hours, pH value 7, air flow 1: 1.Big jar is sent out the liquid that obtains behind the alcohol is exactly required product---microbial bactericide.
Claims (1)
1, a kind ofly prepare the production method of microbial bactericide with bacillus cereus (Bacillus cereus) 98-I bacterial strain, its technological process of production comprises slant strains preparation, shaking table strain preparation, first class inoculum preparation and bulk fermentation, it is characterized by:
---the processing condition of slant strains preparation are: PDA substratum, 28 ℃, 48 hours
---the processing condition of shaking table strain preparation are: 1. number substratum, 28-42 ℃, shaking speed 150-200 rev/min, 48 hours, 1. number substratum was: potato 20%, glucose 1.5-3.0%, sodium-chlor 0.2-0.5%, potassium primary phosphate 0.1-0.6%, all the other are water
---the processing condition of first class inoculum preparation are: 2. number substratum, 28-42 ℃, 150-280 rev/min of fermentor tank rotating speed, 48 hours, pH value 4-10, air flow 1: 1,2. number substratum is: Semen Maydis powder 0.2-0.5%, bean powder 0.2-0.5%, glucose 3-6%, sodium chloride 0.1-0.8%, potassium primary phosphate 0.1-0.8%, ammonium sulfate 0.1-0.8%, peptone 0.1-0.8%, sal epsom 0.01-0.05%, zinc sulfate 0.01-0.05%, all the other are water
---the processing condition of bulk fermentation are: 2. number substratum, 28-42 ℃, 150-280 rev/min of fermentor tank rotating speed, 48 hours, pH value 4-10, air flow 1: 1.
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Cited By (1)
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| CN103555603A (en) * | 2012-09-26 | 2014-02-05 | 中国人民解放军总医院 | Space bacillus cereus strain LCT-BC25 |
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| CN101942404B (en) * | 2010-07-07 | 2012-01-04 | 长治学院 | Bacillus cereus and application thereof in promoting growth of laxus malrel |
| CN102839137B (en) * | 2012-07-27 | 2013-10-16 | 长安大学 | Solid microbial bacterium agent for petroleum degradation and preparation method thereof |
| CN102757921B (en) * | 2012-07-27 | 2013-10-16 | 长安大学 | Bacillus cereus and application thereof |
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| CN103011416A (en) * | 2012-10-24 | 2013-04-03 | 刘颖超 | Application of bacillus cereus in degrading glyphosate |
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| CN109055270A (en) * | 2018-08-31 | 2018-12-21 | 广东容大生物股份有限公司 | A kind of high-density cultivation method and culture medium for bacillus |
| CN115261283B (en) * | 2022-08-26 | 2023-09-08 | 甘肃省科学院生物研究所 | Bacillus cereus and application thereof in prevention and control of disease of dry farming potatoes |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103555603A (en) * | 2012-09-26 | 2014-02-05 | 中国人民解放军总医院 | Space bacillus cereus strain LCT-BC25 |
| CN103555603B (en) * | 2012-09-26 | 2014-09-17 | 中国人民解放军总医院 | Space bacillus cereus strain LCT-BC25 |
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