CN101921710B - Repairing agent for microbes in water bodies of excessive culture zones - Google Patents
Repairing agent for microbes in water bodies of excessive culture zones Download PDFInfo
- Publication number
- CN101921710B CN101921710B CN 201010260196 CN201010260196A CN101921710B CN 101921710 B CN101921710 B CN 101921710B CN 201010260196 CN201010260196 CN 201010260196 CN 201010260196 A CN201010260196 A CN 201010260196A CN 101921710 B CN101921710 B CN 101921710B
- Authority
- CN
- China
- Prior art keywords
- bacillus
- cgmcc
- water
- acinetobacter
- culture
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 title claims abstract description 48
- 244000052616 bacterial pathogens Species 0.000 title claims abstract description 10
- 239000003795 chemical substances by application Substances 0.000 title claims abstract description 9
- 241000894006 Bacteria Species 0.000 claims abstract description 51
- 241000589291 Acinetobacter Species 0.000 claims abstract description 10
- 241000193830 Bacillus <bacterium> Species 0.000 claims abstract description 10
- 241000194103 Bacillus pumilus Species 0.000 claims abstract description 10
- 241000589516 Pseudomonas Species 0.000 claims abstract description 10
- 244000005700 microbiome Species 0.000 claims description 18
- 241000190932 Rhodopseudomonas Species 0.000 claims description 8
- 230000012010 growth Effects 0.000 abstract description 18
- 230000000813 microbial Effects 0.000 abstract description 12
- 238000009629 microbiological culture Methods 0.000 abstract 1
- 230000001580 bacterial Effects 0.000 description 32
- 238000002360 preparation method Methods 0.000 description 23
- 239000002609 media Substances 0.000 description 15
- 239000007788 liquid Substances 0.000 description 13
- 241000251468 Actinopterygii Species 0.000 description 10
- 235000019688 fish Nutrition 0.000 description 10
- 241000276438 Gadus morhua Species 0.000 description 9
- 238000009360 aquaculture Methods 0.000 description 9
- 244000144974 aquaculture Species 0.000 description 9
- 235000019516 cod Nutrition 0.000 description 9
- 230000001488 breeding Effects 0.000 description 8
- 235000015097 nutrients Nutrition 0.000 description 8
- 230000037250 Clearance Effects 0.000 description 7
- 235000019733 Fish meal Nutrition 0.000 description 7
- 230000035512 clearance Effects 0.000 description 7
- 239000004467 fishmeal Substances 0.000 description 7
- 238000000034 method Methods 0.000 description 7
- 230000001954 sterilising Effects 0.000 description 7
- 241000196324 Embryophyta Species 0.000 description 6
- 230000015556 catabolic process Effects 0.000 description 6
- 230000004059 degradation Effects 0.000 description 6
- 238000006731 degradation reaction Methods 0.000 description 6
- 230000000694 effects Effects 0.000 description 6
- 238000000855 fermentation Methods 0.000 description 6
- 230000004151 fermentation Effects 0.000 description 6
- 239000007787 solid Substances 0.000 description 6
- 238000004659 sterilization and disinfection Methods 0.000 description 6
- CVTZKFWZDBJAHE-UHFFFAOYSA-N [N].N Chemical compound [N].N CVTZKFWZDBJAHE-UHFFFAOYSA-N 0.000 description 5
- 238000009825 accumulation Methods 0.000 description 5
- 230000003042 antagnostic Effects 0.000 description 5
- 201000010099 disease Diseases 0.000 description 5
- 229910052760 oxygen Inorganic materials 0.000 description 5
- 239000001301 oxygen Substances 0.000 description 5
- MYMOFIZGZYHOMD-UHFFFAOYSA-N oxygen Chemical compound O=O MYMOFIZGZYHOMD-UHFFFAOYSA-N 0.000 description 5
- 230000002195 synergetic Effects 0.000 description 5
- -1 this Substances 0.000 description 5
- 241000238557 Decapoda Species 0.000 description 4
- 240000005158 Phaseolus vulgaris Species 0.000 description 4
- 235000010627 Phaseolus vulgaris Nutrition 0.000 description 4
- KMUONIBRACKNSN-UHFFFAOYSA-N Potassium dichromate Chemical compound [K+].[K+].[O-][Cr](=O)(=O)O[Cr]([O-])(=O)=O KMUONIBRACKNSN-UHFFFAOYSA-N 0.000 description 4
- 229920002472 Starch Polymers 0.000 description 4
- 239000000306 component Substances 0.000 description 4
- RWSOTUBLDIXVET-UHFFFAOYSA-N dihydrogen sulfide Chemical compound S RWSOTUBLDIXVET-UHFFFAOYSA-N 0.000 description 4
- 229910000037 hydrogen sulfide Inorganic materials 0.000 description 4
- IOVCWXUNBOPUCH-UHFFFAOYSA-M nitrite anion Chemical compound [O-]N=O IOVCWXUNBOPUCH-UHFFFAOYSA-M 0.000 description 4
- IJGRMHOSHXDMSA-UHFFFAOYSA-N nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 4
- 239000006916 nutrient agar Substances 0.000 description 4
- 230000001717 pathogenic Effects 0.000 description 4
- 239000006041 probiotic Substances 0.000 description 4
- 230000000529 probiotic Effects 0.000 description 4
- 235000018291 probiotics Nutrition 0.000 description 4
- 150000003839 salts Chemical class 0.000 description 4
- 239000011780 sodium chloride Substances 0.000 description 4
- 239000002689 soil Substances 0.000 description 4
- 239000008107 starch Substances 0.000 description 4
- 235000019698 starch Nutrition 0.000 description 4
- 239000000725 suspension Substances 0.000 description 4
- 231100000041 toxicology testing Toxicity 0.000 description 4
- 239000001888 Peptone Substances 0.000 description 3
- 235000015278 beef Nutrition 0.000 description 3
- 238000000354 decomposition reaction Methods 0.000 description 3
- 239000012153 distilled water Substances 0.000 description 3
- 238000011081 inoculation Methods 0.000 description 3
- 239000002054 inoculum Substances 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 238000009781 safety test method Methods 0.000 description 3
- 238000002798 spectrophotometry method Methods 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 108010046845 tryptones Proteins 0.000 description 3
- 235000015099 wheat brans Nutrition 0.000 description 3
- 241000195493 Cryptophyta Species 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N D-Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- ZPWVASYFFYYZEW-UHFFFAOYSA-L Dipotassium phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N HCl Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- 241000237502 Ostreidae Species 0.000 description 2
- 229940069002 Potassium Dichromate Drugs 0.000 description 2
- IPQVRLSXWJPESU-UHFFFAOYSA-N [N].ON=O Chemical compound [N].ON=O IPQVRLSXWJPESU-UHFFFAOYSA-N 0.000 description 2
- 239000003463 adsorbent Substances 0.000 description 2
- QGZKDVFQNNGYKY-UHFFFAOYSA-N ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 2
- 230000003115 biocidal Effects 0.000 description 2
- 239000011248 coating agent Substances 0.000 description 2
- 238000000576 coating method Methods 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 239000003640 drug residue Substances 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- 239000012535 impurity Substances 0.000 description 2
- 239000004615 ingredient Substances 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 239000002068 microbial inoculum Substances 0.000 description 2
- 229910052757 nitrogen Inorganic materials 0.000 description 2
- 230000003000 nontoxic Effects 0.000 description 2
- 231100000252 nontoxic Toxicity 0.000 description 2
- 239000005416 organic matter Substances 0.000 description 2
- 235000020636 oyster Nutrition 0.000 description 2
- 230000029553 photosynthesis Effects 0.000 description 2
- 238000010672 photosynthesis Methods 0.000 description 2
- 239000004033 plastic Substances 0.000 description 2
- 229920003023 plastic Polymers 0.000 description 2
- 230000001105 regulatory Effects 0.000 description 2
- 238000009418 renovation Methods 0.000 description 2
- 230000029058 respiratory gaseous exchange Effects 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 108020004465 16S Ribosomal RNA Proteins 0.000 description 1
- 241000252230 Ctenopharyngodon idella Species 0.000 description 1
- 241000252210 Cyprinidae Species 0.000 description 1
- 241000252234 Hypophthalmichthys nobilis Species 0.000 description 1
- 238000010629 Molecular evolutionary genetics analysis Methods 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 230000003044 adaptive Effects 0.000 description 1
- 238000004887 air purification Methods 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 238000009835 boiling Methods 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 239000004927 clay Substances 0.000 description 1
- 229910052570 clay Inorganic materials 0.000 description 1
- 238000007906 compression Methods 0.000 description 1
- 230000000875 corresponding Effects 0.000 description 1
- 230000000593 degrading Effects 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 238000007599 discharging Methods 0.000 description 1
- 229940079593 drugs Drugs 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 230000003203 everyday Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000005755 formation reaction Methods 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 239000001963 growth media Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 238000011065 in-situ storage Methods 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 230000001788 irregular Effects 0.000 description 1
- 239000012092 media component Substances 0.000 description 1
- 239000012533 medium component Substances 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- QJGQUHMNIGDVPM-UHFFFAOYSA-N nitrogen group Chemical group [N] QJGQUHMNIGDVPM-UHFFFAOYSA-N 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 230000008506 pathogenesis Effects 0.000 description 1
- 239000002957 persistent organic pollutant Substances 0.000 description 1
- OAICVXFJPJFONN-UHFFFAOYSA-N phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 239000002504 physiological saline solution Substances 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000011076 safety test Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000010865 sewage Substances 0.000 description 1
- 231100000486 side effect Toxicity 0.000 description 1
- HEMHJVSKTPXQMS-UHFFFAOYSA-M sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000004083 survival Effects 0.000 description 1
- 231100000167 toxic agent Toxicity 0.000 description 1
- 239000003440 toxic substance Substances 0.000 description 1
- 238000009423 ventilation Methods 0.000 description 1
- 230000017613 viral reproduction Effects 0.000 description 1
- 230000001018 virulence Effects 0.000 description 1
- 239000010457 zeolite Substances 0.000 description 1
Images
Abstract
The invention relates to a repairing agent for microbes in water bodies of excessive culture zones, which comprises the following bacteria: Bacillus, Pseudomonas, Bacillus pumilus and Acinetobacter; the collection institution is the China General Microbiological Culture Collection Center (CGMCC); the collection numbers are respectively CGMCC No: 3737, 3739, 3752 and 3712; and the mass ratio of the Bacillus to the Pseudomonas to the Bacillus pumilus to the Acinetobacter is 1.4-1.7:1.0-1.4:0.9-1.1:1.2-1.5. The repairing agent of the invention is a microbial ecological agent which can be used for repairing excessive culture zones of water environment, is harmless to culture organisms, can inhibit propagation and growth of harmful microbes in water bodies, purifies the water quality and improves the quality and the performance of aquatic products.
Description
Technical field
The invention belongs to the Applied Biotechnology field, relate to the excessive culture zone of a kind of water surrounding microorganism renovation agent.
Background technology
Microorganism is as a kind of ecological regulator, administers cultivation water environment and can obviously improve water quality, increases dissolved oxygen, reduces ammonia nitrogen and nitrous acid nitrogen, suppresses harmful microbe and breeds, and degradation of organic substances has advantage such as cost-effective simultaneously and comes into one's own day by day.
Appearance along with the aquatic products intensive culture, aquaculture model is mainly based on bait throwing in, and cultivation density increases substantially, cause that the eutrophy factors such as residual bait, ight soil, nitrogen, phosphorus increase in the water body, impel microorganism to breed (comprising pathogenic bacterium) in a large number, COD, ammonia-nitrogen content rise, dissolved oxygen descends, what have surpasses national three class water quality standard, breeding environment constantly worsens, make culture fishery take a bath, one ton of shrimp of every production can make according to estimates increases by 0.2 ton nitrogen element and 0.05 ton phosphoric in the water body, consequently cause fish and shrimp diseases and output to descend.Therefore the water quality quality has very big influence to culturing success or failure and output size; Water quality is good, and pathogenic bacteria is few, and the hydrocoles growth nature of breed is all right, and condition of water quality is poor, the toxic substance accumulation, and pathogenic bacteria is many and virulence is strong, and the hydrocoles of breed is just encroached on easily.In the prior art, often use pharmaceutical chemicals to come sterilization, but cause pathogenic agent to develop immunity to drugs like this, and medicine is residual in a large number, the polluted water environment can be murdered beneficial bacteria in the water simultaneously, causes the imbalance of aquatic ecological.Therefore drop into the good method that microbial preparation is improvement water quality to the culture zone, after in aquaculture water, dropping into a certain amount of microbial preparation and bacterial classification, can suppress the harmful microbe growth, increase dissolved oxygen, reduce the content of ammonia nitrogen and nitrous acid nitrogen, suppress the harmful microbe breeding, degradation of organic substances has cost-effective advantage simultaneously, can set up a kind of new microecological balance relation, improvement water quality makes growing of aquaculture organism health.
Beneficial microorganism generally all under experiment condition from soil, water body bed mud separation screening come out, single microorganism only has specific degradation capability to certain organic pollutant; Though genetic engineering bacterium can strengthen the purification function of microbial preparation, Biosafety is again the ecological problem that is difficult to go beyond.
After microbial preparation drops into aquaculture water, its degrading activity is subjected to temperature, the influence of environmental factors such as DO, unfavorable temperature, humidity, environmental factors such as pH value and DO can make the validity of microbial preparation reduce or forfeiture, some microbial preparations on the market can not adapt to the in situ environment condition of excessive culture zone, what make that wherein microorganism can not be very fast becomes dominant population in the water body, repairing effect is very not obvious, and these external microorganisms are caused potential threat to the security of cultured fishes, have limited the application of these microorganisms in water surrounding culture zone microorganism reparation field.
Summary of the invention
The present invention provides a kind of the have excessive culture zone of water surrounding of reparation and the probiotics harmless to aquaculture organism.
The purpose of this invention is to provide a kind of microbial preparation, formed by following bacterium: bacillus Bacillus, Rhodopseudomonas Pseudomonas, bacillus pumilus Bacillus pumilus and acinetobacter Acinetobacter; Depositary institution is Chinese common micro-organisms culture presevation administrative center, and deposit number is respectively CGMCC No:3737,3739,3752,3712; The mass ratio of described bacillus Bacillus CGMCCNo:3737, Rhodopseudomonas Pseudomonas CGMCC No:3739, bacillus pumilus Bacilluspumilus CGMCC No:3752 and acinetobacter Acinetobacter CGMCC No:3712 is 1.4～1.7: 1.0～1.4: 0.9～1.1: 1.2～1.5.
The present invention screens the colonial morphology of bacterial strain
The colonial morphology of bacillus Bacillus CGMCC No:3737 on nutrient agar medium: bacterium colony is irregular, and surface colour is dark, and is opaque, the late stage of culture overstrike, and bacterium colony is bigger.
The colonial morphology of Rhodopseudomonas Pseudomonas CGMCC No:3739 on nutrient agar medium: bacterium colony is rounded, and oyster white is opaque, smooth surface, and neat in edge, bacterium colony is bigger.
The colonial morphology of bacillus pumilus Bacillus pumilus CGMCC No:3752 on nutrient agar medium: bacterium colony is opaque, oyster white, and smooth surface, neat in edge, bacterium colony is less.
The colonial morphology of acinetobacter Acinetobacter CGMCC No:3712 on nutrient agar medium: colony colourless, opaque, rounded, smooth surface, neat in edge, bacterium colony is less.
Beneficial effect of the present invention
(1) this probiotics has remarkable benefit for aquaculture, can effectively reduce the concentration of materials such as nitrite in the cultivation water, ammonia nitrogen, hydrogen sulfide, suppresses harmful microorganism breeding and growth in the water body, purifies water, and improves fishery products quality and performance.
(2) this microbial preparation has little, the high efficiency of investment, advantage such as easy to use, and nontoxic, harmless, drug residue free, does not produce resistance, and life-time service can reduce antibiotic usage quantity in the breeding process, reduces disease and takes place.
(3) can degrade the rapidly remaining feed in the water body, ight soil and other organism of fish of this microbial preparation particularly removed long aquaculture water bottom, pond, as the remaining feed of bottom, old shrimp pond, seashore accumulation, drain refuse, plant and animal residues; Simultaneously, objectionable impuritiess such as ammonia in the water body, nitrite, hydrogen sulfide can also be absorbed, the accumulation of solid organic matters and objectionable impurities can be effectively avoided.These algae are the photosynthesis that master's plant plankton produces, it is again the pond zoobenthos, the breathing of aquatic animal, organic decomposition provides oxygen, thereby form the optimum ecological circulation in pond, impel beneficial microorganism to breed in a large number, in the pond, form dominant population, can suppress the breeding of pathogenic micro-organism, reduce disease and take place.
Description of drawings
Fig. 1 is the air-purification system figure of fermentation culture of the present invention;
Fig. 2 is preparation method's schema of microbial inoculum of the present invention;
Fig. 3 is the clearance change curve to fish meal COD of bacterial strain of the present invention.
Bacterial strain used in the present invention is: bacillus Bacillus CGMCC No:3737, Rhodopseudomonas Pseudomonas CGMCC No:3739, bacillus pumilus Bacillus pumilus CGMCCNo:3752, acinetobacter Acinetobacter CGMCC No:3712, the above-mentioned bacterial strains code name is respectively KB2, SL3, WZ3 and WXW1.
Embodiment
Embodiment:
Embodiment 1-4 sees the following form:
Substratum after this four strains bacterium of the present invention fermented is routinely sneaked into clay, zeolite powder adsorbent, particle is made in compression through low temperature, the mass ratio of adsorbent and substratum is 7: 3, makes the microorganism preparation for repairing, the little restoration of the ecosystem preparation of every gram bacteria containing amount about 10,000,000,000.During use, activate earlier, activation method is to contain some water to pour in the vessel that hold preparation from the water body of excessive culture zone, leaves standstill a whole night, sprinkles in the water body of excessive culture zone in second day then.
Screening, cultivation and the fermenting process of four kinds of bacterial classifications of the present invention
1, the enrichment of bacterial classification of the present invention with separate
Getting experiment mud sample places in the plastic channel, add fish meal and sterilization fresh water, add fish meal again after cultivating a week, circulated 25 days, and took out the mud sample, dilution, coating is incubated on the TSB substratum, cultivate 24h for 28 ℃, line separate bacterial strain, adopt test tube to protect kind of a method above-mentioned bacterial strains and be preserved in 4 ℃ of refrigerators.
TSB nutrient media components: Tryptones 17 grams, phytone 3 grams, sodium-chlor 2.5 grams, dipotassium hydrogen phosphate 2.5 grams, glucose 2.5 grams, 1000mL distilled water.
2, screening bacterial strain
The above-mentioned bacterial strain that is deposited in the refrigerator is activated with the TSB substratum, picking list colony inoculation is on the fish meal solid medium, cultivated 48 hours for 28 ℃, get the bigger bacterial strain of bacterium colony that growth is very fast, form, these bacterial growths comparatively fast illustrate can utilize the bait material as unique carbon source and nitrogenous source as its necessary nutrition of growing.The inoculation that these growths bacterium colony very fast, that form is bigger adopts potassium dichromate process to measure the COD value of nutrient solution on the fish meal liquid nutrient medium every day, and finishing screen is chosen the four strains of the present invention bacterium stronger to the fish meal capacity of decomposition.
3, strain classification status of the present invention determines
The 16S rRNA sequence of four kinds of bacterial strains of the present invention is grown the tree assessment by the search of BLAST homology and MEGA 4 software system of NCBI, identify the classification position of this four strains bacterium, and be deposited in Chinese common micro-organisms culture presevation administrative center, they are respectively: bacillus Bacillus CGMCC No:3737, Rhodopseudomonas Pseudomonas CGMCC No:3739, bacillus pumilus Bacillus pumilusCGMCC No:3752 and acinetobacter Acinetobacter CGMCC No:3712, numbering is respectively KB2 in the present invention, SL3, WZ3 and WXW1.
4, fermentation culture
(1), medium component
The main component of fermention medium has: dregs of beans, wheat bran, starch, inorganic salt.
(2), medium sterilization
Method: adopt moist heat sterilization.
(3), air sterilization
Adopt the method for air filtration degerming, air filtering system 1 is air inlet, 2 coarse filters, 3 air compressor, 4 main air water coolers, 5 secondary air water coolers, 6 water traps, 7 air reservoirs, 8 cyclonic separators, 9 wire mesh demisters, 10 airheaters, 11 total air filters, divides strainer as shown in Figure 1.
(4), seed preparation
The seed preparation comprises laboratory seed preparation and the preparation of production plant seed
The substratum of laboratory seed preparation is solid medium, and main component has dregs of beans, starch and inorganic salt, and culture temperature is 30 degrees centigrade,
Incubation time 2～8 days, the different strain incubation time has difference, determines according to the normal experiment method of different strains.It is 30 degree of ingesting that solid medium, main component dregs of beans, wheat bran, starch and inorganic salt, culture temperature are also adopted in the preparation of production plant seed, 2 grades of fermentations.
(5), fermentation culture
Employing is with churned mechanically barrel mast formula reactor four kinds of bacterium of the present invention to be carried out solid-state mixed fermentation, substratum main component dregs of beans, wheat bran, starch and inorganic salt; Fermentation condition: temperature is 30 degrees centigrade, and pH 7, salinity 5.
For preparation method's schema of microbial inoculum of the present invention, can be clearly seen that whole process of preparation as shown in Figure 2.
Below four kinds of bacteriums of the present invention are carried out formation condition experiment, ecological safety experiment, antagonism experiment, synergetic property and action effect test experience.
One, organic matter degradation bacteria growth conditions test
1, experimental technique
(basic ingredient comprises Tryptones 10 grams to beef extract-peptone liquid nutrient medium basic ingredient, extractum carnis 3 grams, 1000mL distilled water, pH7.4), regulating its salinity gradient with sodium-chlor is: 0,5,10,15,20,25,30,35,40, the organic matter degradation bacterium is inoculated in the inoculum of these several concentration gradients, and cultivate down for 28 ℃, every the OD600 value of 4h with spectrophotometric determination bacterium liquid, the relatively growing state of bacterial strain under these several salinity gradients.
Four kinds of microbionations of the present invention are arrived in the beef extract-peptone liquid nutrient medium.Under 8 ℃, 12 ℃, 16 ℃, 20 ℃, 22 ℃, 24 ℃, 26 ℃, 28 ℃, 30 ℃, 32 ℃, 34 ℃, 36 ℃, 40 ℃, 44 ℃ several temperature gradients, cultivate respectively, every the OD600 value of 4h with spectrophotometric determination bacterium liquid, the relatively growing state of bacterial strain under this several temperature gradient.
The beef extract-peptone liquid nutrient medium, be that 5% sodium hydroxid and mass concentration are that 10% dilute hydrochloric acid is regulated the pH value with the mass concentration of boiling, making its pH gradient is 5.4,5.6,5.8,6.0,6.2,6.4,6.6,6.8,7.0,7.2,7.4,7.6,7.8,8.0,8.2,8.4,8.6,8.8,9.0,9.2,9.4.Four kinds of microbionations of the present invention in the inoculum of these several concentration gradients, are cultivated down for 28 ℃, every the OD600 value of 4h with spectrophotometric determination bacterium liquid, to compare the growing state of bacterial strain of the present invention under these several pH gradients.
2, experimental result
As shown in table 1, the optimal temperature of four strain bacterial growths of the present invention is mostly between 28 ℃～30 ℃, but growth temperature range is at 16 ℃～36 ℃; The pH value of optimum growth is about 7.4, and the pH scope that can grow is between 6.4～8.4; The salinity of optimum growth mostly is greatly about 5～10, and the salinity range of can growing is between 0～20.
The growth conditions test of bacterial strain of the present invention shows that four strain bacteriums of the present invention are stronger to the adaptive faculty of temperature environment, and the optimum growth temperature of suitable growth temperature and water surrounding culture zone matches.
Two, degradation bacteria ecological safety test
After the plastics tank sterilization, adds 20 liters of fresh water of sterilizing, four kinds of fishes of 30 tail carps, silver carp, grass carp, bighead of putting into 5～10cm in the tank respectively, every kind of fish establish three parallel.Microorganism renovation agent of the present invention is made bacteria suspension insert in the tank, make its final concentration in tank maintain 10
6～10
7CFU/mL, with aseptic fresh water group in contrast, continuously ventilation, according to culture object throw something and feed corresponding bait sooner or later each once, record culture object survival condition and in time remove dead individual.
Experimental result is cultured after 10 days shown in table 2,3,4 and 5, and the gained data are carried out test of difference, and the surviving rate of test group and control group does not have significant difference, proves that the present invention four strain bacterium are without any side effects to the growth of test kind.
Three, the antagonism experiment of bacterial strain of the present invention
The present invention four strain bacteriums are activated at the TSB substratum, get single bacterium colony and in physiological saline, make bacteria suspension, the bacteria suspension of a kind of bacterium is coated TSB solid culture primary surface, the circular aseptic scraps of paper soak in the bacteria suspension of another kind of bacterium, be attached to the TSB media surface of coating bacterium liquid, observation has or not inhibition zone behind the 48h, checks these several bacterium to have or not antagonism.
TSB culture medium prescription: Tryptones 17 grams, phytone 3 grams, sodium-chlor 2.5 grams, dipotassium hydrogen phosphate 2.5 grams, glucose 2.5 grams, 1000mL distilled water.
Experimental result shows all do not have antagonism between these a few strain bacterium.
Four, the synergetic property of bacterial strain of the present invention and action effect experiment
Four strain bacteriums of the present invention are activated at the TSB substratum, get single colony inoculation in the TSB liquid nutrient medium, after cultivating 24h, the bacterium liquid of every kind of bacterium is inoculated in the fish meal substratum of liquid, inoculum size is 0.5～2.0% altogether, and every 24 hours sample thiefs once, 5000r/min is centrifugal, potassium dichromate process is measured the COD of supernatant liquor, METHOD FOR CONTINUOUS DETERMINATION 7 days.As shown in Figure 3, in the 7th day, the COD clearance of SL3, WZ3 and WXW1 is the highest, is about 60%, and is as shown in table 6, and the COD clearance of the mixed bacteria liquid of bacterial strain KB2 and SL3, WZ3 and WXW1 is the highest.A kind of bacterial strain of every increase, the COD clearance of mixed bacteria liquid increases successively, proves to have synergy between the four strain bacterial strains of the present invention.
Interpretation of result
1, by check, all there is not antagonism between a few strain bacterium of the present invention;
2, the COD clearance of bacterial strain KB2 is the highest, and based on bacterial strain KB2, with other a few strain strain combinations, the COD clearance of check mixed bacterium changes, and gets as shown in table 6.Mixed bacterium with experimental group KB2, SL3, WZ3 and WXW1 is the highest to the clearance of nutrient solution, is up to 87.9%, so probiotics is made with these four kinds of bacterium.
In sum, draw as drawing a conclusion by above-mentioned four experiments:
1, this probiotics has remarkable benefit for aquaculture as shown in Table 6, can effectively reduce nitrite in the cultivation water, ammonia nitrogen, hydrogen sulfide isoconcentration, suppresses harmful microorganism breeding and growth in the water body, purifies water, and improves fishery products commodity performance.
2, by table 1-6 as can be known this microbial preparation have little, the high efficiency of investment, advantage such as easy to use, and nontoxic, harmless, drug residue free, do not produce resistance, life-time service can reduce antibiotic usage quantity in the breeding process, reduce disease and take place, the sewage environmental pollution of discharging is also less.
3, can degrade the rapidly remaining feed in the water body, ight soil and other organism of fish of microbial preparation of the present invention particularly removed long aquaculture water bottom, pond, as the remaining feed of bottom, old shrimp pond, seashore accumulation, drain refuse, plant and animal residues; Simultaneously, objectionable impuritiess such as ammonia in the water body, nitrite, hydrogen sulfide can also be absorbed, the accumulation of solid organic matters and objectionable impurities can be effectively avoided.These algae are the photosynthesis that master's plant plankton produces, it is again the pond zoobenthos, the breathing of aquatic animal, organic decomposition provides oxygen, thereby form the optimum ecological circulation in pond, impel beneficial microorganism to breed in a large number, in the pond, form dominant population, can suppress the breeding of pathogenic micro-organism, reduce disease and take place.
Subordinate list
Table 1 is the growth conditions table of bacterial strain of the present invention;
Table 2 is safety testing tables of the fish of bacterial strain bacillus Bacillus CGMCC No:3737 of the present invention;
Table 3 is safety testing tables of the fish of bacterial strain Rhodopseudomonas Pseudomonas CGMCC No:3739 of the present invention;
Table 4 is safety testing tables of the fish of bacterial strain bacillus pumilus Bacillus pumilus CGMCC No:3752 of the present invention;
Table 5 is safety testings of the fish of bacterial strain acinetobacter Acinetobacter CGMCC No:3712 of the present invention;
Table 6 is synergetic property and action effect experiment tables of bacterial strain of the present invention.
Table 1
Table 2
Table 3
Table 4
Table 5
Table 6
Claims (1)
1. repairing agent for microbes in water bodies of excessive culture zones is characterized in that it is made up of following bacterium: bacillus Bacillus, Rhodopseudomonas Pseudomonas, bacillus pumilus Bacilluspumilus and acinetobacter Acinetobacter; Depositary institution is Chinese common micro-organisms culture presevation administrative center, and deposit number is respectively CGMCC No:3737,3739,3752,3712; The mass ratio of described bacillus Bacillus CGMCC No:3737, Rhodopseudomonas Pseudomonas CGMCC No:3739, bacillus pumilus Bacillus pumilus CGMCC No:3752 and acinetobacter Acinetobacter CGMCC No:3712 is 1.5:1.2:1.0:1.3.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 201010260196 CN101921710B (en) | 2010-08-24 | 2010-08-24 | Repairing agent for microbes in water bodies of excessive culture zones |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 201010260196 CN101921710B (en) | 2010-08-24 | 2010-08-24 | Repairing agent for microbes in water bodies of excessive culture zones |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN101921710A CN101921710A (en) | 2010-12-22 |
| CN101921710B true CN101921710B (en) | 2013-07-10 |
Family
ID=43336928
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN 201010260196 Expired - Fee Related CN101921710B (en) | 2010-08-24 | 2010-08-24 | Repairing agent for microbes in water bodies of excessive culture zones |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN101921710B (en) |
Families Citing this family (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102634465B (en) * | 2011-12-27 | 2014-03-26 | 西北大学 | Acinetobacter with authigenic nitrogen fixation capacity and application thereof |
| CN103074218B (en) * | 2012-09-17 | 2014-08-20 | 北京天安生物科技有限公司 | Composite flora and additive cellulose preparation method by using the same |
| CN102888340B (en) * | 2012-09-17 | 2015-09-09 | 北京天安生物科技有限公司 | A kind of composite flora and prepare the method for textile fibres with it |
| CN104030459A (en) * | 2014-06-24 | 2014-09-10 | 天津科技大学 | Microbial preparation for brackish water aquaculture, and preparation method and application thereof |
| CN105331552B (en) * | 2014-08-13 | 2018-09-28 | 广州中国科学院先进技术研究所 | One plant of efficient denitrification acinetobacter calcoaceticus novel species and its application |
| CN106673203A (en) * | 2015-11-06 | 2017-05-17 | 丹阳市尚德生物科技有限公司 | River water quality and riverbed biological microorganism repairing solution |
| WO2021033186A1 (en) * | 2019-08-22 | 2021-02-25 | The National Institute for Biotechnology in the Negev Ltd. | Compositions for inoculating aquatic animals |
| CN111635869A (en) * | 2020-05-06 | 2020-09-08 | 南京市水产科学研究所 | Multifunctional microecological preparation for freshwater shrimp culture and preparation method and application thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2008121079A1 (en) * | 2007-03-30 | 2008-10-09 | Singapore Polytechnic | Bioremediation of hydrocarbon sludge |
| CN101717752A (en) * | 2008-10-09 | 2010-06-02 | 上海市胸科医院 | Parental human lung adenocarcinoma cell line with high potential of bone metastasis |
-
2010
- 2010-08-24 CN CN 201010260196 patent/CN101921710B/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2008121079A1 (en) * | 2007-03-30 | 2008-10-09 | Singapore Polytechnic | Bioremediation of hydrocarbon sludge |
| CN101717752A (en) * | 2008-10-09 | 2010-06-02 | 上海市胸科医院 | Parental human lung adenocarcinoma cell line with high potential of bone metastasis |
Non-Patent Citations (4)
| Title |
|---|
| 利用复合微生物菌剂控制水华的治理工程试验;吴定心;《环境科学与技术》;20100731;第33卷(第7期);第151页左栏第6段-右栏第2段，第154页左栏第1段-右栏第1段 * |
| 吴定心.利用复合微生物菌剂控制水华的治理工程试验.《环境科学与技术》.2010,第33卷(第7期),第151页左栏第6段-右栏第2段，第154页左栏第1段-右栏第1段. |
| 水产养殖对水域环境的影响及污染防治对策;边蔚;《河北农业科学》;20091231;第13卷(第6期);91-93 * |
| 边蔚.水产养殖对水域环境的影响及污染防治对策.《河北农业科学》.2009,第13卷(第6期),91-93. |
Also Published As
| Publication number | Publication date |
|---|---|
| CN101921710A (en) | 2010-12-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN101921710B (en) | Repairing agent for microbes in water bodies of excessive culture zones | |
| CN100588624C (en) | Microorganism renovation agent of water environment and preparation method thereof | |
| Cole | Interactions between bacteria and algae in aquatic ecosystems | |
| CN101376877B (en) | Compound microecological preparation for purifying cultivation water, preparation formulation thereof and preparation processes of the preparation | |
| CN102965314B (en) | Bacillus subtilis and preparation and application of microbial inoculum thereof | |
| CN102719383A (en) | Bacillus amyloliquefaciens, inoculant and application thereof | |
| CN111394272B (en) | Brevibacillus laterosporus and application thereof | |
| CN106867933A (en) | The probiotics of purification of water quality and preparation and application in being cultivated to Environment of Litopenaeus vannamei Low | |
| CN106754551A (en) | A kind of bacterium amount lactobacillus preparation high and preparation method and application | |
| CN103045498B (en) | Bacillus amyloliquefaciens and application thereof | |
| CN109355223A (en) | One plant of bacillus subtilis N2 and its application with ammonia nitrogen degradation function | |
| CN101717724A (en) | Compound microecological preparation used for water body purification of aquaculture | |
| CN101624234B (en) | Microorganism water treatment agent in culturing water and preparation technique thereof | |
| CN101333499A (en) | Complex active bacterial biological water purifying a gent and method for preparing same | |
| CN104593303B (en) | A kind of liquid complex micro organism fungicide and its production method | |
| CN102146347B (en) | Acinetobacter sp. and application of composite bacterial agent of acinetobacter species | |
| CN103204589A (en) | Method for biologically treating industrial circulating water and stabilizing quality thereof | |
| CN105132307B (en) | It is a kind of applied to the composite bacillus microbial water-purifying agent of ornamental fish and its application | |
| CN104651282B (en) | A kind of preparation method of Composite Photosynthetic Bacteria preparation | |
| CN102994417B (en) | Bacillus pumilus and application thereof in prevention and control of comma bacillus in aquatic product | |
| CN104328075B (en) | Bacillus subtilis strain capable of killing algae and application thereof | |
| CN102899277A (en) | Bacillus cereus and application thereof | |
| CN103923856B (en) | One strain basophilic pseudomonas AD-28 and the application in water body denitrification thereof | |
| CN104789548B (en) | A kind of marine aquaculture carrier immobilized antibacterial type water quality cleansing agent and preparation method thereof | |
| CN107937301B (en) | Bacillus amyloliquefaciens and application thereof in aquaculture |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20130710 Termination date: 20210824 |