CN102337232A - Aerobic denitrification psychrotolerant bacterium and preparation method thereof - Google Patents
Aerobic denitrification psychrotolerant bacterium and preparation method thereof Download PDFInfo
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Abstract
The invention provides an aerobic denitrification psychrotolerant bacterium and a preparation method thereof. The collection number of the aerobic denitrification psychrotolerant bacterium is CGMCC No.4753. A strain can undergo denitrification at high dissolved oxygen level and low dissolved oxygen level, and has a wider appropriate growing temperature range. After the aerobic denitrification psychrotolerant bacterium is inoculated into lake water polluted by agricultural runoff, and is cultured at 10 DEG C for 7 days, the removing rate of nitrate ions is over 80 percent. The aerobic denitrification psychrotolerant bacterium has a higher treatment value on low-temperature eutrophic water bodies.
Description
Technical field
The present invention discloses a kind of aerobic denitrification psychrotroph, and the preparation method of this psychrotroph also is provided simultaneously, belongs to water environment pollution bioremediation technology field.
Background technology
China's water resources per capita is poor, and a large amount of nitrogenous nutritive salt is prone to cause body eutrophication after getting into lake and reservoir, has further aggravated the situation of China's shortage of water resources, serious threat drinking water safety and ecosystem health.22 that reach V class and following water quality in the emphasis lakes (storehouse) are controlled in 28 states of China in 2008, account for 78.6%, and wherein total nitrogen is one of main contamination index.In essence, the control body eutrophication mainly is to solve the nitrogen and phosphorus pollution problem, promptly reduces or block the input and the removal endogenous pollution load of exogenous nutrition salt.At present, main method has the combined action of physical method (cutting dirty and bed mud dredging etc.), chemical process (adding flocculation agent and chemical agent etc.), biological method (utilizing the life Metabolic activity of unicellular algae, plant and mikrobe etc.) and different methods.Compare with chemical method with the physics method, that biological process has is easy and simple to handle, environmental perturbation is little, facility and working cost is low, non-secondary pollution, save energy and help setting up healthy advantages such as aquatic ecosystem.Unicellular algae is because volume is small and be difficult to collect, and the application in eutrophication control receives certain restriction.High waterplant is removed, rebuilds at nitrogen phosphorus and recovers aspect such as good aquatic ecosystem has vital role, but it uses and often receive the influence in season, and like area northeastward, have only about 4 months annual effective acting time.Mikrobe then because kind is many, breeding is fast and adapt to characteristics such as strong, is showing unique effect aspect the polluted-water biological prosthetic.
Seeing that body eutrophication phenomenon in various degree all took place in some main Hu Ku of north China in recent years; Therefore develop and a kind ofly can effectively remove the nutritive salt in the eutrophication water, and then reduce the technology that nutritive salt accumulates have vital role in water body fundamentally administering eutrophication water at hypothermic phase.
Research shows that psychrotroph can keep higher growth and metabolic activity in low temperature environment, can remove organism and total nitrogen in the sewage effectively.Therefore, research hypothermic phase psychrotroph is to effective removal effect of nitrogen in the eutrophication water, and is significant to the body eutrophication problem of some main Hu Ku (especially northern area) of fundamentally administering China.
In the low temperature eutrophication water; Except that mikrobe, other biological life Metabolic activity stops basically, and the organism decomposition rate in the water body is also slack-off; Cause dissolved oxygen levels in the water body than higher; Therefore aerobic denitrifying bacteria can become dominant bacteria in this environment, and can effectively remove the nitrate ion in the water body through denitrification.
The isolating aerobic denitrifying bacteria of past people can utilize oxygen molecule and nitrate ion to carry out denitrification process as electron acceptor(EA) simultaneously; But denitrification rate is slow, and the existence of oxygen molecule utilizes nitrate ion to have certain disadvantageous effect to bacterial strain.There are some researches show also that simultaneously under anaerobic mikrobe can be that unique electron acceptor(EA) carries out anti-nitre process with nitrate ion, but process be slow under the catalysis of anaerobic denitrifying enzyme, denitrification efficient is low.
Summary of the invention
The present invention discloses a kind of aerobic denitrification psychrotroph, for a kind of new bacterial strain, through denitrification, effectively removes the nitrate ion in the water body.
The present invention also provides the preparation method of above-mentioned bacterial strains, uses suitability for industrialized production.
The present invention further discloses this bacterial strain in the purposes that effectively reduces the safety non-pollution of nitrate ion concentration in the agricultural run-off polluted river water.
Aerobic denitrification psychrotroph disclosed by the invention,Called after DBP-3 is preserved in that " China Committee for Culture Collection of Microorganisms's common micro-organisms " center ", its preserving number are CGMCC No. 4753; Preservation date: on April 7th, 2011, preservation address: No. 3, No. 1 institute in Beichen West Rd., Beijing, classification name: Acinetobacter johnsonii, Acinetobacter johnsonii.
Of the present inventionBacterial strain has following characteristic: (1) at the colony characteristics of 10 ℃ of cultivations after 3 days on the beef extract-peptone flat board is: bacterium colony size 1-2mm, and bacterium colony is rounded, smooth surface, pale, neat in edge, transparent.(2) cell morphological characteristic: the thalline rod-short, Gram-negative, thalline is about 1.4-2.2 μ m, wide about 0.9-1.2 μ m, no bud is embraced, and nothing is moved about.(3) the 16S rRNA gene sequence characteristic of Acinetobacter johnsonii DBP-3 CGMCC No. 4753: its 16S rRNA has the nucleotide sequence shown in the sequence table, and sequence length is 1578 bp.
With reference to " content of the outstanding Bacteria Identification handbook of uncle (the 8th edition), according to its morphological specificity and physiological and biochemical property, and according to the Phylogenetic Analysis that makes up based on 16S rRNA gene order, identify bacterial strain (CGMCC No. 4753) be Acinetobacter johnsonii (
Acinetobacter johnsonii).
Aerobic denitrification psychrotroph disclosed by the inventionThe preparation method, may further comprise the steps:
Gather serious rice-nutrient water body water sample and shoaling water bed mud when (1) envrionment temperature is 10-15 ℃, mix the back as the bacterium source according to volume ratio 1:1;
(2) get in the 250ml triangular flask that fills the 80ml selective medium that 20ml bacterium source joins the bacterium of going out, under 10 ℃, shaking culture (160r/min) is cultivated 48h; It is centrifugal then that (40000r/min 15min), gets the 0.01g throw out and joins static cultivation in the iodine flask that fills with fresh culture; Centrifugal behind the 48h, get the 0.01g throw out again, join and carry out aerobic cultivation in the substratum; So alternately cultivate 10 times, all do at every turn and repeat for 3 times and the control treatment that does not add the bacterium source;
(3) last aerobic nutrient solution is coated on the solid medium cultivates, choose single bacterium colony, utilize sterilized water to be prepared into every milliliter and contain 10
8-10
9Behind the bacteria suspension of individual bacterium; Get in the 250ml triangular flask that fills the 45ml minimal medium that 5ml joins sterilization, carry out aerobic cultivation again, centrifugal after 48 hours; Measure the nitrate ion concentration in the supernatant, the clearance of choosing nitrate ion reaches the bacterial strain more than 60%.The cultivation of each bacterium colony is all done the control treatment that repeats and do not add somatic cells for 3 times;
(4) the pure bacterium of primary dcreening operation is further rule separate and mineral nutrition liquid culture experiment, finally choose and to grow fast and the nitrate ion clearance is reached the bacterial strain more than 80%.
Used substratum comprises selective medium and solid medium in bacterial strain domestication of the present invention and the culturing process,
Wherein, Selective medium is made up of 1g/L saltpetre, 2g/L sodium acetate, 0.125g/L potassium primary phosphate, 0.2g/L MAGNESIUM SULPHATE HEPTAHYDRATE 99.5,0.2/L sodium-chlor, 1.0g/L ammonium sulfate, 0.025g/L Calcium dichloride dihydrate, 3g/L sodiumazide, 0.05 mg/L boric acid, 0.1 mg/L rose vitriol, 0.2 mg/L copper sulfate, 3 mg/L ferrous sulfate, 0.02 mg/L manganous chloride, 0.1 mg/L Sodium Molybdate Dihydrate, 0.03 mg/L Zinc Sulphate Heptahydrate, and the pH value is 7.2.Above-mentioned chemical substance is dissolved in the sterilized water, utilizes the hydrochloric acid soln of 0.1mol/L and the sodium hydroxide solution of 0.1mol/L to regulate the pH value.
Solid medium is formed pH=7.2 by 0.5 g/L Carnis Bovis seu Bubali cream, 1.0g/L peptone, 0.5 g/L sodium-chlor, 18.0 g/L agar.
Positively effect of the present invention is:A kind of new aerobic denitrification psychrotroph is provided, and this bacterial strain can both carry out efficient denitrification, the wider range of suitable growth under high-solubility oxygen and LDO level.Be inoculated in the lake water of agricultural run-off pollution, cultivate after 7 days for 10 ℃, the clearance of nitrate ion reaches more than 80%, has higher value aspect the improvement of hypothermic phase eutrophication water.
The present invention attempts under coldcondition to suppress under the aerobic condition bacterial strain to the utilization of oxygen molecule through in the denitrification substratum, adding cell aerobic repiration suppressor factor sodiumazide; Thereby improve utilization ratio to nitrate ion; Suppress acting inefficient denitrification enzyme under the anaerobic condition simultaneously; And, let bacterial strain that dissolved oxygen levels and temperature variation are had higher adaptive faculty through the anoxic acclimation shaking culture.Purpose is the deficiency that overcomes in the low temperature eutrophication water denitride technology, provides a kind of aerobic denitrification psychrotroph that temperature and dissolved oxygen levels are had a higher adaptive faculty that utilizes the directed acclimatization technology domestication of low temperature and filter out to come the method for the safety non-pollution of nitrate ion concentration in the effective reduction agricultural run-off polluted river water.
Description of drawings
Fig. 1 is according to 16S rDNA gene order bacterial strain PDB-3 that makes up and the phylogenetic tree that reaches isolating aerobic denitrifying bacteria with kind; Wherein: * DBP-3: bacterial strain uses therefor of the present invention;
Fig. 2 is the growth characteristics figure of bacterial strain under the different dissolved oxygen conditions;
Fig. 3 is the denitrifying capacity figure of bacterial strain under the condition of different temperatures;
Fig. 4 is the denitrifying capacity figure of bacterial strain in eutrophication water.
Embodiment
Through following examples the present invention is described for example further; And do not limit the present invention in any way; Under the prerequisite that does not deviate from technical solution of the present invention, any change or change that those of ordinary skills that the present invention did are realized easily all will fall within the claim scope of the present invention.
She leads to river serious rice-nutrient water body water sample and shoaling water bed mud to gather the Changchun, Jilin Province when (1) envrionment temperature is 10-15 ℃, mixes the back as the bacterium source according to volume ratio 1:1;
(2) get in the 250ml triangular flask that fills the 80ml selective medium that 20ml bacterium source joins the bacterium of going out, under 10 ℃, shaking culture (160r/min) is cultivated 48h; It is centrifugal then that (40000r/min 15min), gets the 0.01g throw out and joins static cultivation in the iodine flask that fills with fresh culture; Centrifugal behind the 48h, get the 0.01g throw out again, join and carry out aerobic cultivation in the substratum; So alternately cultivate 10 times, all do at every turn and repeat for 3 times and the control treatment that does not add the bacterium source;
(3) last aerobic nutrient solution is coated on the solid medium cultivates, choose single bacterium colony, utilize sterilized water to be prepared into every milliliter and contain 10
8-10
9Behind the bacteria suspension of individual bacterium; Get in the 250ml triangular flask that fills the 45ml minimal medium that 5ml joins sterilization, carry out aerobic cultivation again, centrifugal after 48 hours; Measure the nitrate ion concentration in the supernatant, the clearance of choosing nitrate ion reaches the bacterial strain more than 60%.The cultivation of each bacterium colony is all done the control treatment that repeats and do not add somatic cells for 3 times;
(4) the pure bacterium of primary dcreening operation is further rule separate and mineral nutrition liquid culture experiment, finally choose and to grow fast and the nitrate ion clearance is reached the bacterial strain more than 80%.
Picking one ring somatic cells of the present invention adds in the aseptic redistilled water of 100 μ l from flat board, boiling water bath 5min behind the whirlpool mixing, and the centrifugal 5min of 12000r/min, supernatant is used for PCR, and primer is a pair of universal primer:
Forward primer Pf:5 '-AGAGTTTGATCCTGGCTCAG-3 ';
Reverse primer Pr:5 '-ACGGCTACCTTGTTACGACT-3 ',
Correspond respectively to 8～27 and 1495～1514 bases of colibacillary 16S rRNA gene;
PCR reaction system (100 μ l) is: 10 * Ex Taq buffer, 10 μ l, 5 U Ex Taq, 10 mmol/L dNTPs, 4 μ l, each 1 μ l of 50pmol/L primer, 1 μ l template is supplemented to 100 μ l with redistilled water.The PCR reaction conditions is: 95 ℃ of 3min; 95 ℃ of 1 min; 58 ℃ of 1min; 72 ℃ of 1min 30s, 30 circulations; 72 ℃ of 10min, 4 ℃ of preservations;
Get PCR product 5 μ l and carry out the evaluation of 1% agarose gel electrophoresis.Dna gel with Beijing ancient cooking vessel state biotech company reclaims test kit recovery purified pcr product, after the DNA/RNA quantitative instrument is quantitative, delivers to a day root biotech firm and carries out nucleotide sequencing.Submit the 16S rDNA sequence that records to GenBank; Utilize Blast to carry out the homology sequence comparison; After choosing the higher sequence of homology and becoming consistent and isometric end to end sequence with the DNAMEN software editing; Choose the 16S rDNA sequence that belongs to the higher bacterium type strain of interior homology again and carry out genetic distance calculating; Carry out sequence alignment with ClustalW 2.0 softwares, application system is grown Neighbor-Joining method among the tree analysis software MEGA 4.0, adopts two-parameter computation model constructing system tree of Kimura and sequence of calculation similarity (see figure 1).In addition, the aerobic denitrifying bacteria bacterial strain of isolation identification has been also contained in the structure of phylogenetic tree.The bacterial strain that simultaneously separation and purification is obtained carries out the observation and the biochemical reactions experiment of colony characteristics and thalli morphology structure.
With the hydrochloric acid soln of the sodium hydroxide of 0.1mol/L and 0.1mol/L the pH value of liquid nutrient medium is adjusted to 7.2; In 28 250ml triangular flasks, add liquid nutrient medium 90ml respectively; 121 ℃ of following high pressure steam sterilization 30min; After the cooling more respectively among 21 triangular flasks in add bacteria suspension 10ml of the present invention, be respectively 5,10,15,20,25,30 and 35 ℃ of following shaking culture in temperature, every kind of temperature repeats for three times; And do the control treatment that does not add bacterium respectively, the shaking table hunting speed is 160rpm.24,48,72 hours collected specimens after cultivation are respectively measured the upgrowth situation of somatic cells.
The upgrowth situation of somatic cells characterizes with the absorbancy at 600nm place.The result shows that bacterial strain begins to get into logarithmic phase after getting into and passing through of short duration adaptation in the new nutrient solution.
When temperature was 5 ℃, bacterial strain of the present invention still kept certain activity, and just the speed of growth is slow; Temperature is when 5 ℃ are increased to 20 ℃, and the speed of growth of bacterial strain increases along with the rising of temperature; No significant difference (see figure 2) when the upgrowth situation when temperature is 25 ℃ and 30 ℃ and 20 ℃.Show that bacterial strain has higher growth and metabolic activity at low temperatures, the TR of suitable growth compares broad.Therefore bacterial strain should have stronger competitive capacity and long survival time in practical application, has tangible application potential aspect the body eutrophication improvement.
Test Example 1
Change the dissolved oxygen levels of culture system through the hunting speed of regulating the water bath with thermostatic control vibrator; Hunting speed is respectively 60 and 160rpm; Through the mensuration of dissolved oxygen instrument, corresponding dissolved oxygen levels is respectively LDO level (3-4mg/L) and high dissolved oxygen levels (6-7mg/L).Every kind of dissolved oxygen levels carries out 3 times to be repeated, and does the control treatment that does not add bacterium simultaneously.The add-on of bacteria suspension, liquid medium and culture condition are with embodiment 1; Wherein adjust the add-on of nitrate ion; Making its concentration is 124mg/L; The the 12nd, 24,36,48 and 60 hour collected specimens after cultivation according to the concentration range of nitrate ion in the sample of estimation, confirmed the sample collecting amount respectively.Water sample behind 0.45 micron millipore filtration suction filtration, carry out gradient dilution according to concentration after, adopt the concentration of Pbenoldisulfonic Acid spectrophotometry nitrate ion.The result shows when bacterial strain of the present invention gets into behind the culture system 24-36h maximum to the removal speed of nitrate ion, and under high-solubility oxygen and the LDO level, bacterial strain is to the clearance no significant difference of acid ion.After cultivating 60h, the clearance of nitrate ion is respectively 87% and 84% (see figure 3) under high-solubility oxygen and the LDO level, therefore can infer that bacterial strain has higher adaptive faculty to dissolved oxygen levels.
Test Example 2
Select the collection location of Jilin Agriculture University corn experimental plot as the farmland rainwash.According to weather-forecast, plant the agricultural run-off collection device previous day of heavy rain.Dig one 60cm * 60cm * 60cm heatable adobe sleeping platform at the low-lying place in field, putting into diameter is 50cm, and height is the glass scoop of 60cm.In the scoop top cover is the center of circle with the scoop, the circular cloth for keeping out rain that radius is 1 meter.During make raining, no rainfall in the 1m around the scoop is so that nutritive substance such as dissolved nitrogen phosphorus more in the rainwash of inflow scoop.After scoop fills with rainwash, send it back to laboratory, leave standstill 20min, mix according to a certain volume with near the lake water in a small-sized lake with filtrating behind the middling speed filter paper filtering, as experimental water (concentration of nitrate ion is 5.78mg/L).
Sterilization water sample+bacterium and 2 kinds of 2 kinds of control treatment handling and only add sterilization water sample and former water sample of former water sample+bacterium are cultivated 10 ℃ of following lucifuges; The concentration of regularly gathering the water determination nitrate nitrogen is studied the denitrogenation ability of bacterial strain of the present invention; Wherein the add-on of water sample and bacteria suspension is respectively 90ml and 10ml, and other culture condition is with embodiment 1.The result shows (see figure 4); Compare with the control treatment that does not add bacterial strain; Add behind the bacterial strain nitrate nitrogen in the culture system and clearance and removal speed obviously increase, bacterial strain of the present invention is 80% and 78% to the clearance of nitrate nitrogen in sterilization water sample and the former water sample.Do not adding under the situation of bacterial strain, after former water sample is cultivated for some time, the still decrease to some degree of the concentration of nitrate nitrogen, and nitrate nitrogen changes not obvious in the sterilization water sample.Can infer; The bacterial strain of the present invention that adds plays a major role to the removal of nitrate nitrogen in the water sample; Bacterial strain can reduce the concentration of the nitrate ion in the water fast through denitrification, shows that bacterial strain has stronger application potential aspect the rich nitrogen water body of reinforcement reparation low temperature.
CAGTGCCAAGCTTGCATGCCTGCAGGTCGACGATTAGAGTTTGATCCTGGCTCAGATTGAACGCTGGCGGCAGGCTTAACACATGCAAGTCGAGCGGGGAAAGGTAGCTTGCTACCTGACCTAGCGGCGGACGGGTGAGTAATGCTTAGGAATCTGCCTATTAGTGGGGGACAACATTCCGAAAGGAATGCTAATACCGCATACGCCCTACGGGGGAAAGCAGGGGATCTTCGGACCTTGCGCTAATAGATGAGCCTAAGTCAGATTAGCTAGTTGGTGGGGTAAAGGCCTACCAAGGCGACGATCTGTAGCGGGTCTGAGAGGATGATCCGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGGACAATGGGCGAAAGCCTGATCCAGCCATGCCGCGTGTGTGAAGAAGGCCTTTTGGTTGTAAAGCACTTTAAGCGAGGAGGAGGCTACCGAGATTAATACTCTTGGATAGTGGACGTTACTCGCAGAATAAGCACCGGCTAACTCTGTGCCAGCAGCCGCGGTAATACAGAGGGTGCGAGCGTTAATCGGATTTACTGGGCGTAAAGCGTGCGTAGGCGGCTTTTTAAGTCGGATGTGAAATCCCTGAGCTTAACTTAGGAATTGCATTCGATACTGGGAAGCTAGAGTATGGGAGAGGATGGTAGAATTCCAGGTGTAGCGGTGAAATGCGTAGAGATCTGGAGGAATACCGATGGCGAAGGCAGCCATCTGGCCTAATACTGACGCTGAGGTACGAAAGCATGGGGAGCAAACAGGATTAGATACCCTGGTAGTCCATGCCGTAAACGATGTCTACTAGCCGTTGGGGCCTTTGAGGCTTTAGTGGCGCAGCTAACGCGATAAGTAGACCGCCTGGGGAGTACGGTCGCAAGACTAAAACTCAAATGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGATGCAACGCGAAGAACCTTACCTGGTCTTGACATAGTAAGAACTTTCCAGAGATGGATTGGTGCCTTCGGGAACTTACATACAGGTGCTGCATGGCTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTTTCCTTATTTGCCAGCGGGTTAAGCCGGGAACTTTAAGGATACTGCCAGTGACAAACTGGAGGAAGGCGGGGACGACGTCAAGTCATCATGGCCCTTACGACCAGGGCTACACACGTGCTACAATGGTCGGTACAAAGGGTTGCTACCTAGCGATAGGATGCTAATCTCAAAAAGCCGATCGTAGTCCGGATTGGAGTCTGCAACTCGACTCCATGAAGTCGGAATCGCTAGTAATCGCGGATCAGAATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGGGAATTTGTTGCACCAGAAGTAGGTAGTCTAACCGCAAGGAGGACGTTTACCACGGTGTGGCCGATGACTGGGGTGAAGTCGTAACAAGGTAGCCGTAATCTCTAGAGGATCCCCGGGTACCGAGCTCGAATTCGTAAT
Claims (4)
1. a strain aerobic denitrification psychrotroph bacterial strain, its preserving number is: CGMCC No. 4753.
2. obtain the method for the said aerobic denitrification psychrotroph of claim 1 bacterial strain, may further comprise the steps completion:
Gather serious rice-nutrient water body water sample and shoaling water bed mud when (1) envrionment temperature is 10-15 ℃, mix the back as the bacterium source according to volume ratio 1:1;
(2) get in the 250ml triangular flask that fills the 80ml selective medium that 20ml bacterium source joins the bacterium of going out, under 10 ℃, shaking culture (160r/min) is cultivated 48h; It is centrifugal then that (40000r/min 15min), gets the 0.01g throw out and joins static cultivation in the iodine flask that fills with fresh culture; Centrifugal behind the 48h; Get the 0.01g throw out, join again and carry out aerobic cultivation in the substratum, so alternately cultivate 10 times;
(3) last aerobic nutrient solution is coated on the solid medium cultivates, choose single bacterium colony, utilize sterilized water to be prepared into every milliliter and contain 10
8-10
9Behind the bacteria suspension of individual bacterium, get 5ml and carry out aerobic cultivation again, centrifugal after 48 hours, measure the nitrate ion concentration in the supernatant, the clearance of choosing nitrate ion reaches the bacterial strain more than 60%;
(4) the pure bacterium of primary dcreening operation is further rule separate and culture experiment, finally choose and to grow fast and the nitrate ion clearance reached 80% bacterial strain.
3. according to the said method for preparing aerobic denitrification psychrotroph bacterial strain of claim 2, it is characterized in that:
Described selective medium is made up of 1g/L saltpetre, 2g/L sodium acetate, 0.125g/L potassium primary phosphate, 0.2g/L MAGNESIUM SULPHATE HEPTAHYDRATE 99.5,0.2/L sodium-chlor, 1.0g/L ammonium sulfate, 0.025g/L Calcium dichloride dihydrate, 3g/L sodiumazide, 0.05 mg/L boric acid, 0.1 mg/L rose vitriol, 0.2 mg/L copper sulfate, 3 mg/L ferrous sulfate, 0.02 mg/L manganous chloride, 0.1 mg/L Sodium Molybdate Dihydrate, 0.03 mg/L Zinc Sulphate Heptahydrate, and utilizing the hydrochloric acid soln of 0.1mol/L and the sodium hydroxide solution adjusting pH value of 0.1mol/L is 7.2;
Solid medium is made up of 0.5 g/L Carnis Bovis seu Bubali cream, 1.0g/L peptone, 0.5 g/L sodium-chlor, 18.0 g/L agar, and utilizing the hydrochloric acid soln of 0.1mol/L and the sodium hydroxide solution adjusting pH value of 0.1mol/L is 7.2.
4. the application in eutrophication water is administered according to claim 1 is characterized in that, is 10 with concentration
8-10
9The bacteria suspension of individual bacteria/milliliters, the inoculum size by 10% join in the lake water that receives the agricultural run-off pollution that the nitrate ion starting point concentration is 5.78mg/L, and after 7 days, the clearance of nitrate ion reaches more than 80% 10 ℃ of cultivations.
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| CN102628025A (en) * | 2012-04-09 | 2012-08-08 | 天津大学 | Nutrient liquid formula for culturing high-temperature thermophilic bacterium and preparation method and application thereof |
| CN107805612A (en) * | 2017-10-20 | 2018-03-16 | 中冶华天工程技术有限公司 | The enrichment and separation method and water body treating method of low-temperature aerobic denitrifying bacterium |
| CN110627228A (en) * | 2019-11-12 | 2019-12-31 | 天津软银科技有限公司 | Composition for nitrobacteria culture |
| CN110964663A (en) * | 2019-12-04 | 2020-04-07 | 北京化工大学 | Heterotrophic nitrifying bacteria for low-temperature sewage denitrification and application thereof |
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