CN104663726A - Method for preparing bactericide for preventing and treating plant blight by fermenting lactose refined wastewater - Google Patents
Method for preparing bactericide for preventing and treating plant blight by fermenting lactose refined wastewater Download PDFInfo
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- CN104663726A CN104663726A CN201410641762.XA CN201410641762A CN104663726A CN 104663726 A CN104663726 A CN 104663726A CN 201410641762 A CN201410641762 A CN 201410641762A CN 104663726 A CN104663726 A CN 104663726A
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Abstract
The invention provides a method for preparing a bactericide for preventing and treating plant blight by fermenting lactose refined wastewater. The technological characteristics are that lactose refined wastewater acts as the main raw material; according to the proportion of adding 5g of potassium chloride and 25g of crushed bean pulp into per liter of wastewater, a proper amount of potassium chloride and the crushed soybean pulp are added, and stirred uniformly, and are adjusted until the pH value is 7.0 to act as fermentation raw materials; bacillus amyloliquefaciens H47 with the preservation number of CCTCC M2013283 preserved in China Center for Type Culture Collection in June 24th, 2013 acts as a fermentation bacteria strain; the fermentation parameters are that the ventilatory capacity in per liter of a fermentation liquid is 1liter/min; the fermentation temperature is 30 DEG C; the stirring rotation speed is 220rpm; the fermentation liquid obtained by fermenting for 32-36 hours is the biological bactericide. The biological bactericide has the functions of improving the soil fertility and promoting the synthesis and oriented transportation of photosynthetic products, and can provide plant growth adjustment substances, the synthesized antibacterial substance has an obvious effect of preventing and treating the plant blight and can enable industrial and agricultural wastes to be utilized reasonably to a certain degree; the method cannot cause environment pollution, can turn waste into wealth, is simple, convenient and feasible, and belongs to the technical field of biology, and the price is low.
Description
Technical field
The present invention with lactose refining waste water for primary raw material, with the deposit number that on June 24th, 2013 is stored in China typical culture collection center be: the bacillus amyloliquefaciens of the Bacillus amyloliquefaciens H47 of CCTCC M 2013283 is for fermentation strain, the zymotic fluid obtained for 32-36 hour that ferments is exactly a kind of bacteria agent, not only have and improve soil fertility, promote the synthesis of photosynthetic product and directed transport, plant growth regulating substance is provided, and synthesized antibacterial material has obvious effect to control plant fusarium wilt, and work can be made to a certain extent, agricultural wastes obtain Appropriate application, there is no environmental pollution, belong to biological technical field.
Background technology
Find that sickle-like bacteria can be infected banana and be caused fusarium wilt the earliest, it also causes the melon crops such as watermelon, cucumber, muskmelon, the vegetables such as eggplant, capsicum, tomato, wild cabbage, kidney bean, cowpea, and fusarium wilt occurs for banana, pomegranate, grape, apple, strawberry etc.; Agricultural production is caused and has a strong impact on.Crop rotation management and chemicals is mainly adopted to kill to the control of fusarium wilt at present, but limited efficiency.Use chemical fertilizer in a large number for a long time, make soil secondary salinization and acidification trend seriously, soil permeability and granular structure are destroyed, and preserve moisture and fertility ability reduces also makes the root system of crops and residual body throughout the year for Pathogen of Fusarium Wilt provides nutrition and raw place.Such chemical pesticide is difficult to the generation fundamentally controlling fusarium wilt, and can bring out pathogen pesticide resistance, uses agricultural chemicals in a large number for a long time, also easily infiltrates through soil and underground water and causes the degree of depth of environment to pollute and melon and fruit residue of pesticide.Apply to the microorganism of environment and bio-safety and produce metabolite and fusarium wilt prevented and treated to the main flow and Main way that become current Overseas Development.
Research finds that Biocontrol microorganism prevents and treats fusarium wilt mainly through hyperparasitism, antibiosis and competitive approach, the large class of main point fungus and bacterium two.Mycophyta comprises rasp berry, arbuscular mycorrhizal fungi and non-pathogenic sickle-like bacteria; Bacterium class mainly contains 18 genus, and wherein some mushroom effects of bacillus are better, and biological safety is also high, is more satisfactory biocontrol microorganisms.As far back as Wang Ya equality using Bacillus subtilis TG26 in 1993, have studied the preventive and therapeutic effect of its zymotic fluid to watermelon blight and tobacco fusarium wilt, find that the control efficiency of leaching root can reach 100%, field trial also has and obviously promotes production-increasing function.This quasi-microorganism can not only produce secondary metabolite to be suppressed or pathogenic microbe killing, can also produce systemic immunity, controlling plant diseases by inducing plant.
Some microorganisms of bacillus are a class very important rhizosphere growth-promoting microorganisms, be conducive to improving soil fertility, plant growth regulating substance is provided, many bacillus also can synthetic antimicrobial material, as Bacillus subtilis, Bacillus licheniformis, Bacillus pumilus, B.amyloliquefaciens B128, B.circulans ATCC 4513, Bacillus amyloliquefaciens etc. can produce fat peptide matters, Sabate ' waits report, Bacillus subtilis Surfactin that C4 produces energy antagonism series bacillus, control bee larva is downright bad sick.The discoveries such as Tao, Surfactin alleviates Fengycin can suppress soft rot nosophyte Rhizopus stolonifer.The reports such as Hassan, several bacillus that can be used for preventing and treating sugarcane red rot also can synthesize Surfactin.Fat peptide matters can also produce defense reaction by inducing plant, thus is conducive to improving disease resistance of plant.The discoveries such as Huang, Surfactin and Iturin can kill the Penicillium notatum in peach juice.Can be used for that Salmonella killed by meat, Escherichia coli killed by milk and bacillus cereus.The reports such as Huang, Bacillus cereus C1L can alleviate maize leaves rot; The discoveries such as Ye, the Iturin A2 that Bacillus subtilis produces can effective control of maize leaf blight.The microorganism of bacillus also can be used for control by the microbial wheat diseases of reaping hook.The reports such as Dunlap, Bacillus amyloliquefaciens AS 43.3 can be used for the microbial fringe rot of control reaping hook.The reports such as Kumar, Bacillus subtilis MBI 600 can the water prevention rice sheath blight.Bacillus micro-organism also has good preventive and therapeutic effect to bacterialo wilt disease.Abd-Allah reports, Bacillus subtilis can be used for preventing and treating peanut sclerotium rolfsii.In sum, bacillus micro-organism not only has the microbial fusarium wilt of antagonism reaping hook, also has good control efficiency to other fungus-caused disease and bacterialo wilt disease.But fermentation of bacillus technology of preparing cost is higher at present, and microbial inoculum function is single, should not apply at agriculture field; Therefore studying and setting up with agriculture and industry waste is that fermenting raw materials prepares fusarium wilt multifunctional bio control agent, popularization for fusarium wilt biocontrol agent is significant with further application, the present invention is exactly under such guiding theory, succeeds through overtesting.
Summary of the invention
First the present invention invents with lactose refining waste water as primary raw material, the ratio of 5 grams, potassium chloride and the dregs of beans 25 grams through pulverizing is added in often liter of waste water, stir after adding the dregs of beans of appropriate potassium chloride and pulverizing respectively and adjust PH=7.0, with the deposit number that on June 24th, 2013 is stored in China typical culture collection center be: the bacillus amyloliquefaciens of the Bacillus amyloliquefaciens H47 of CCTCC M 2013283 is for fermentation strain, be exactly a kind of bacteria agent at certain fermentation condition bottom fermentation 32-36 hour preparation-obtained zymotic fluid, not only have and improve soil fertility, promote the synthesis of photosynthetic product and directed transport, plant growth regulating substance is provided, and synthesized antibacterial material has obvious effect to control plant fusarium wilt, and work can be made to a certain extent, agricultural wastes obtain Appropriate application, there is no environmental pollution, cheap and can turn waste into wealth, simple and easy to do.
Technical scheme of the present invention:
With the deposit number that on June 24th, 2013 is stored in the China typical culture collection center of Luo Jia Shan Wuhan University of wuchang, wuhan district be: the bacillus amyloliquefaciens of the Bacillus amyloliquefaciens H47 of CCTCC M 2013283 is fermenting and producing starting strain, add the ratio of 5 grams, potassium chloride and the dregs of beans 25 grams through pulverizing in often liter of waste water in lactose refining waste water, stir after adding the dregs of beans of appropriate potassium chloride and pulverizing respectively and adjust PH=7.0 as fermentation raw material, control plant fusarium wilt microbial inoculum is at the zymotic fluid of certain fermentation condition bottom fermentation 32-36 hour manufacture, technique is:
1) get the waste water 4 liters discharged in lactose refining industrial production, add dregs of beans 100 grams, 20 grams, potassium chloride, stirs evenly, and adjusts PH=7.0, puts into 5 liters of fermentation tanks, and 115 DEG C of sterilizings 30 minutes, are cooled to 30 DEG C, stand-by.
2) by deposit number be: the actication of culture of the Bacillus amyloliquefaciens H47 of CCTCC M 2013283 is also prepared into zymocyte liquid:
(1) get slant strains one ring and connect mycelia at the flat lining out of PDA, cultivate 48 hours for 30 DEG C in incubator.
(2) get PDA flat-plate bacterial colony one articulating to enter to be equipped with in the triangular flask of 250ml specification of 50ml seed culture fluid, prepare some parts, be placed in shaking table, condition of culture: rotating speed 200rpm, temperature 30 DEG C, constant temperature culture 2 days, is zymocyte liquid.
(3) seed culture fluid preparation: by weight percentage, peptone 10%, yeast extract 5%, potassium chloride 0.5%, pure water 84.5%, adjusts PH=7.0.
3) zymocyte liquid is added in fermentation tank by 5% of wastewater volume, namely add 200ml zymocyte liquid in above-mentioned containing in the fermentation tank of 4 liters of waste water, it is 1 liter/min that throughput often rises zymotic fluid, fermentation temperature 30 DEG C, speed of agitator 220rpm, fermentation time 32-36 hour.
4) zymotic fluid after fermenting 32-36 hour is the microbial inoculum of control plant fusarium wilt.
Beneficial effect of the present invention:
Although the discovery of plant fusarium wilt has had the history of more than 100 year, it has administered main based on field management with use chemical pesticide; The popularizing application of biopesticide have not been reported.Its key constraints is production cost problem, industry, agricultural, domestic refuse fermentation is utilized to be then workout cost problem and effective means for the microbial inoculum of antagonism fusarium wilt, about the investigation and application of this respect, there is not been reported, especially the nutriment in waste water is utilized to produce microbial inoculum, both can save waste water treatment cost, directly can also go out product, apply for agricultural extension.
The function and the quality that increase product are another important means reduced costs, corps leaf surface spray improves crop growth, especially the Chinese medicine preparation of fruit, seed growth growth, by fermentation control, beneficial microbe is produced and be conducive to leaf photosynthesis, promotion photosynthetic product to fruit transfer and the secondary metabolite stimulating fruit development, the function improving fusarium wilt has realistic meaning to the function improving biological protection agent undoubtedly, and there is not been reported in the research of this respect.
And method of the present invention, not only administer the industrial wastewater solving refining lactose, and turn waste into wealth, using it as the deposit number that on June 24th, 2013 is stored in China typical culture collection center be: the raw material of the bacillus amyloliquefaciens fermentable of the Bacillus amyloliquefaciens H47 of CCTCC M 2013283, manufacture is provided and is improved soil fertility, promote the synthesis of photosynthetic product and directed transport, plant growth regulating substance is provided, and synthesized antibacterial material has the bacteria agent of significantly effect to control plant fusarium wilt, environmental protection, there is no discarded object, greatly reduce production cost, have broad application prospects, good economic benefit and social benefit, specifically detailed Study on mechanism report is stated in thesis for the doctorate afterwards.
Embodiment
Embodiment 1
1) get the waste water 4 kilograms discharged in lactose refining industrial production, add dregs of beans 100 grams, 20 grams, potassium chloride, stirs evenly, and adjusts PH=7.0, puts into 5 liters of fermentation tanks, and 115 DEG C of sterilizings 30 minutes, are cooled to 30 DEG C, stand-by.
2) by deposit number be: the actication of culture of the Bacillus amyloliquefaciens H47 of CCTCC M 2013283 is also prepared into zymocyte liquid:
(1) get slant strains one ring and connect mycelia at the flat lining out of PDA, cultivate 48 hours for 30 DEG C in incubator.
(2) get PDA flat-plate bacterial colony one articulating to enter to be equipped with in the triangular flask of 250ml specification of 50ml seed culture fluid, prepare some parts, be placed in shaking table, condition of culture: rotating speed 200rpm, temperature 30 DEG C, constant temperature culture 2 days, is zymocyte liquid.
(3) seed culture fluid preparation: by weight percentage, peptone 10%, yeast extract 5%, potassium chloride 0.5%, pure water 84.5%, adjusts PH=7.0.
3) zymocyte liquid is added in fermentation tank by 5% of wastewater volume, namely add 200ml zymocyte liquid in above-mentioned containing in the fermentation tank of 4 liters of waste water, it is 1 liter/min that throughput often rises zymotic fluid, fermentation temperature 30 DEG C, speed of agitator 220rpm, fermentation time 32-36 hour.
4) zymotic fluid after fermenting 32-36 hour is the microbial inoculum of control plant fusarium wilt.
Claims (1)
1. prepare the method for control plant fusarium wilt microbial inoculum with lactose refining wastewater fermentation for one kind, be characterized in utilizing the waste water discharged in lactose refining industrial production to be dominant fermentation raw material, add a small amount of dregs of beans and potassium chloride, the deposit number adopting on June 24th, 2013 to be stored in China typical culture collection center is: the Bacillus amyloliquefaciens strain of the Bacillus amyloliquefaciens H47 of CCTCC M2013283 is fermentation strain, the bacteria agent of manufacture control plant fusarium wilt, its technique is:
1) waste water 4 liters discharged in lactose refining industrial production is got, the ratio of 5 grams, potassium chloride and the dregs of beans 25 grams through pulverizing is added in often liter of waste water, 20 grams, potassium chloride is added respectively in 4 liters of lactose refining waste water, the dregs of beans 100 grams pulverized, stir evenly and adjust PH=7.0, put into 5 liters of fermentation tanks, 115 DEG C of sterilizings 30 minutes, be cooled to 30 DEG C, stand-by as fermentation raw material;
2) by deposit number be CCTCC M 2013283 Bacillus amyloliquefaciens H47 actication of culture and be prepared into zymocyte liquid:
(1) get slant strains one ring and connect mycelia at the flat lining out of PDA, cultivate 48 hours for 30 DEG C in incubator;
(2) get PDA flat-plate bacterial colony one articulating to enter to be equipped with in the triangular flask of 250ml specification of 50ml seed culture fluid, prepare some parts, be placed in shaking table, condition of culture: rotating speed 200rpm, temperature 30 DEG C, constant temperature culture 2 days, is zymocyte liquid;
(3) seed culture fluid preparation: by weight percentage, peptone 10%, yeast extract 5%, potassium chloride 0.5%, pure water 84.5%, adjusts PH=7.0;
3) in 5% ratio of wastewater volume, zymocyte liquid is joined in fermentation tank, namely above-mentioned 1) in containing 4 liters of stand-by fermentation raw materials fermentation tank in add 200ml zymocyte liquid, it is 1 liter/min that throughput often rises zymotic fluid, namely throughput is 4L/min, fermentation temperature is 30 DEG C, speed of agitator 220rpm, fermentation time 32-36 hour;
4) zymotic fluid after fermenting 32-36 hour is the microbial inoculum of control plant fusarium wilt.
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Cited By (3)
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| CN105505819A (en) * | 2015-12-19 | 2016-04-20 | 佛山市艳晖生物科技有限公司 | Method for producing amylolytic spore bacterial fertilizer through rice soaking and washing wastewater in sugar industry |
| CN106867948A (en) * | 2017-04-14 | 2017-06-20 | 江南大学 | It is a kind of to prepare the technology that rare-earth tailing improves microbial inoculum by raw material culture plant-growth promoting rhizobacteria of lactose refining waste water |
| CN106947718A (en) * | 2017-04-14 | 2017-07-14 | 江南大学 | It is a kind of that the technology that rare-earth tailing improves microbial inoculum is prepared by raw material culture plant-growth promoting rhizobacteria of piggery wastewater |
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| CN106867948A (en) * | 2017-04-14 | 2017-06-20 | 江南大学 | It is a kind of to prepare the technology that rare-earth tailing improves microbial inoculum by raw material culture plant-growth promoting rhizobacteria of lactose refining waste water |
| CN106947718A (en) * | 2017-04-14 | 2017-07-14 | 江南大学 | It is a kind of that the technology that rare-earth tailing improves microbial inoculum is prepared by raw material culture plant-growth promoting rhizobacteria of piggery wastewater |
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Application publication date: 20150603 |