CN1308439C - Biological denitrification method for ammonium-nitrogen containing waste water and microbes thereof - Google Patents
Biological denitrification method for ammonium-nitrogen containing waste water and microbes thereof Download PDFInfo
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- CN1308439C CN1308439C CNB2004100051584A CN200410005158A CN1308439C CN 1308439 C CN1308439 C CN 1308439C CN B2004100051584 A CNB2004100051584 A CN B2004100051584A CN 200410005158 A CN200410005158 A CN 200410005158A CN 1308439 C CN1308439 C CN 1308439C
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- bacillus
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- nitrification activity
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- 238000000034 method Methods 0.000 title claims abstract description 130
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- 239000002351 wastewater Substances 0.000 title claims abstract description 60
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims abstract description 187
- 241000894006 Bacteria Species 0.000 claims abstract description 165
- 229910052757 nitrogen Inorganic materials 0.000 claims abstract description 91
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- HEBKCHPVOIAQTA-SCDXWVJYSA-N xylitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)CO HEBKCHPVOIAQTA-SCDXWVJYSA-N 0.000 description 1
- 235000010447 xylitol Nutrition 0.000 description 1
- 229960002675 xylitol Drugs 0.000 description 1
Classifications
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
- Y02W10/00—Technologies for wastewater treatment
- Y02W10/10—Biological treatment of water, waste water, or sewage
Landscapes
- Purification Treatments By Anaerobic Or Anaerobic And Aerobic Bacteria Or Animals (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention public a new biotechnology treatment method of containing ammonium and nitrogen wastewater.In this method,adding nitrating activity heterotrophic bacteria or heterotrophic bacteria family into the ammonium wastewater,then removing the ammonium nitrogen of ammonium wastewater directly by the microorganism fuction at the suitable microorganism growth conditions.Also,this invention public one kind of nitrite-oxidizing bacteria which can remove the ammoniacal nitrogen effectively and protect the environment.
Description
The present invention requires the right of priority at Chinese patents CN03118600.9.Application number is to be on February 14th, 2003 CN03118600.9 patent application day.
Field that the present invention belongs to:
The invention belongs to field of waste water treatment, specifically, the present invention relates to a kind ofly contain the method for ammonium nitrogen wastewater by the microbiological treatment with nitrification activity, wherein ammonium mainly is converted to dinitrogen gas and other nitrogenous gas; The invention still further relates to employed heterotrophic organism in this method of wastewater treatment.
Background technology:
Ammonium nitrogen is one of main nitrate pollution thing that causes body eutrophication.And high density NH
4 +Existence and the difficult treatment of-N waste water in industrial production such as fermentation industry production is an important topic that needs to be resolved hurrily at present.
At present nitrated-denitrification the pattern that is applied in most of scientific research personnel's the brains is NH
4 +The pattern of nitrated-denitrification relay.That is:
NHx wherein
+→ NO
3 -Process be nitrification; And with NO
2 -Or NO
3 -→ N
2O or N
2Be denitrification.
Up to now, still fettered, be that " nitrobacteria of autotrophy " mainly finish by " chemoautotrophy theory " about nature nitrification releaser's understanding.The theoretical most important foundation of this authority is exactly: nitrobacteria can not utilize organic matter, and is organic toxic to nitrification.So nitrobacteria can not be that nutrient gelatin or nutrient agar plate separate acquisition at the Ke He flat board.
At NH
4 +The theory and technology that uses in-N wastewater biological denitrificaion the technology is still by " the nitrated theory of autotrophy ", although existing problems, but numerous investigators are managing solution, be still the new answer of searching in the notion of " autotrophy ", utilize combination membrane bioreactor for treatment high-concentration ammonia nitrogenous wastewater (Li Hongyan, the high first month of spring etc., environmental science as Li Hongyan etc., 2003,23 (5): 62-66).Recently the Crewe of He Lan Delft dimension institute utilizes a newfound notion.(Jetten M S M，Horn S J，et al，Wat.Sci.Tech.，1997，35(9)：171-180；Philips S，Wyffels S，et al，Appl.Microbiol.Biotechnol.，2002，59：557-566)。Be noted that, mentioned European patent EP-A-562466 of a utilization heterotrophic organism in the specification sheets of the treatment process of patent CN99808998.2---ammonia-containing water, " in the method; special microbial mixture; as pseudomonas, acinetobacter calcoaceticus, catarrhalis, coryneform bacteria; micrococci, Flavobacterium and genus bacillus, growth is also rendered to the sewage treatment equipment from these reactors continuously or discontinuously in the just so-called breeding device of independent reactor ".These heterotrophic bacteriums that this patent is mentioned come usefulness with it as remove organic carbon in the waste water (COD) before nitrification takes place, and to guarantee the nitrated good as far as possible generation of autotrophic bacteria, it are not worked as nitrobacteria and use.
Denitrification denitrogenation, for a long time, the notion of biological denitrificaion is with NH by nitrobacteria
4 +-N is oxidized to NO
3 --N product and a large amount of accumulation as disliking the denitrifying nitrogenous source of gas, thereby have reached the purpose of denitrogenation.SHARON method and Anammox method are improved this method, only with NH
4 +-N is oxidizing to NO
2 --N, and to make it the more traditional method of accumulation should be progressive, particularly in the shortening in reaction times, and the advantage that adopts single reaction vessel etc. to dwindle space-time,
Nitrification and denitrification, these two process occurrence conditions are considered to more:
A) nitrification: aerobic conditions can not have organism, substrate NH
4 +, autotrophic bacteria is finished;
B) denitrification: the condition of being sick of must add organism, substrate NO
2 -Or NO
3 -, heterotrophic bacterium is finished.
Under the guidance of this theoretical pattern, the industrial technology of the biological denitrificaion of most of ammonium nitrogen wastewaters is used and is all carried out according to this pattern.Engineering practice for many years finds to exist suitable problem.As: rate of propagation is slow and be difficult to keep higher biological concentration, needs to handle to reduce organic concentration through aeration earlier, and biologic activity be grown and be showed to bacterial strain could, a little less than the impact resistance; Ammonia nitrogen in high density and nitrite can suppress the growth of nitrifier again, make nitrification incomplete, cause nitrogen removal rate very low.
At present, there is the investigator that the biological denitrification process under the aerobic condition is studied abroad, utilizes the certain micro-organisms population under aerobic condition, to have denitrifying characteristic and realize synchronous nitration and denitrification.Result of study shows that general foster sulphur coccus (Thiosphera pantotroph), Alcaligenes faecalis (Alcaligenea faecalis), pseudomonas (Pseadonmonas sp), comamonas microorganisms such as (Comamonos sp.) can utilize NO under aerobic condition
x-N carries out denitrification.Nitrifier and denitrifying bacteria are placed mixed culture in same reactor such as the aeration tank, though can reach the synchronous nitration and denitrification of single reactor.But the denitrification result is unsatisfactory, and a large amount of nitrogen are converted to oxidation state nitrogen, and this oxidation state nitrogen is again the arch-criminal that atmosphere greenhouse gases effect and ozone cavity form, and causes secondary pollution easily.
Domesticly also carried out some relevant research work: (Geng Jinju, Liu Dengru etc. use and the environmental organism journal 2002,8 (1): 78-82) to utilize aerobic denitrifying bacteria group and autotrophy nitrifying bacteria community combined denitrification.Though have ammonia nitrogen removal ability preferably, impact resistance a little less than, the ammonia nitrogen that ammonia nitrogen concentration is higher than the high density of 0.3 grams per liter can suppress the growth of thalline, and ammonia nitrogen concentration is when being higher than 0.2 grams per liter, the ammonia nitrogen residual volume is more after the denitrogenation; Not anti-high organic carbon concentration of while, the organic carbon concentration of 0.5 grams per liter suppress thalli growth and also reduce denitrification effect.And all kinds of microbial culture and growth conditions in this combination flora are inconsistent, the opposing party but is in holddown during one side performance function, cause inharmonious each other, the biological denitrification process time lengthening, cost increases, the more important thing is that nitric efficiency is general, the water body after the denitrogenation also has distance from environmental requirement.
The invention technology contents:
An object of the present invention is to provide the method that a kind of new processing contains ammonium nitrogen wastewater, this method only needs a step ammonia in the waste water can be changed into free of contamination dinitrogen gas and other nitrogenous gas.
Another object of the present invention provides heterotrophic organism or its bacterial flora that a class of using has nitrification activity in this method of wastewater treatment.
A kind of treatment process that contains ammonium nitrogen wastewater, it is characterized in that in ammoniated wastewater, adding the organic carbon source that physiological is produced alkali, and add an amount of heterotrophic organism or heterotrophic organism group with nitrification activity, pH be 6 to 8 aerobic conditions and temperature 20-35 ℃ under static or little whipped state culturing bacterium 15-35 days down, directly remove the ammonium nitrogen in the ammoniated wastewater;
The heterotrophic organism that wherein has nitrification activity is to grow on the PM flat board or to separate, and the direct drop of griess reagent is positive, and when adding the good air culture of inorganic ammonium salt and support with the carbon source that physiological is produced alkali, the bacterial strain that has full nitrogen to wane.
In the present invention, the organic carbon source that adds in ammoniated wastewater refers to it is the class organic compound that physiological is produced alkali, and this class material can cause culture medium pH to rise through the metabolism of microorganism carbon.Particularly, the organic carbon source that physiological is produced alkali is meant organic acid or its esters with carboxyl, preferred pyruvic acid or Sodium.alpha.-ketopropionate or sodium acetate or citric acid or Trisodium Citrate or sodium formiate etc.
The plain different chemical form of N under non-life exists in the chemical reaction transforms (gas, liquid, oxidation-reduction attitude), and the inorganic chemistry diagram is pointed out:
Form: NH
4 +→ N
2H
5 +→ NH
2OH → N
2→ N
2O → NO → HNO
2
Valence state :-3-2-1 0+1+2+3
Form: N
2O
4→ NO
3 -
Valence state :+4+5
Go forward one by one as OR physical and chemical process order on current potential, from the highest ortho states NH that goes back
4 +-N is to the NO of highest oxidation state
3 -Can have multiple oxidation products to occur in the oxidising process of-N, specifically be to form which kind of product, depends on another substrate that participates in reaction and occurrence condition such as temperature, pressure, pH value, catalyzer etc.For example, HNO
3Mainly be reduced to following material as oxygenant:
HNO
3→NO
2→HNO
2→NO→N
2O→N
2→NH
4 +
+5 +4 +3 +2 +1 0 -3
In the chemical reaction, obtain several mixture of products usually.At least that a kind of product any more depend on oxygenant (HNO
3) concentration and the activity of reductive agent.
The chemolithotrophy biological chemistry theory of oxidation of microorganism ammonium and nitrous acid oxidation is pointed out that a kind of extreme isolated biological phenomenon, the i.e. nitrification theory of-3 valency N →+5 valency highest oxidation state N under no organism, aerobic conditions, this theory of one side is mutually disloyal with the diversity of the N oxidising process product that chemical theory is pointed out, the denitrification organic carbon adds on the other hand, under anaerobic highest oxidation state N can be reduced to zeroth order N again
2Or N
2O.The diversity of the N meta-bolites that the former the exclusive state product and the latter form presents a contrast! Constitute the focus organism of this species diversity so, certain organism type exists and may cause " can utilizing organic numerous aerophile bacterium at NH
4 +The diversity that may occur the N oxidation products in assimilation-decomposition of-N." may there be such pathways metabolism in the present invention:
And with NH
4 +Is that a class has the heterotrophic organism of nitrification activity or their mixture by this class by way of the bacterium that changes into dinitrogen gas, the heterotrophic organism that this class has nitrification activity can or separate in the dull and stereotyped growth of PM, the direct drop of griess reagent is positive, the carbon source of producing alkali with physiological adds the good air culture of inorganic ammonium salt when supporting, the bacterial strain that has full nitrogen to wane.
In the present invention, the temperature of cultivating heterotrophic organism can be remained on about 30 ℃, as 28 ℃, 29 ℃, 30 ℃, 35 ℃ etc., and the heterotrophic organism that will have a nitrification activity is the static culturing bacterium of ammonia-containing water about 25 days, as 18 days, or 20 days, or 25 days, or 28 days, all can effectively the ammonium nitrogen in the waste water be changed into dinitrogen gas to some extent.
On the other hand, add the heterotrophic organism with nitrification activity and should control certain quantity in containing ammonium nitrogen wastewater, the cell quantity that adds usually in the waste water can be 10
4-10
7Individual/milliliter.General add-on is 10
5-10
6Individual/milliliter is advisable.
In the present invention, heterotrophic organism or their mixture with nitrification activity can separate from soil, or screening obtains from the bacterial strain of DSMZ's preservation, and separation or screening process also make the PM flat band method be:
1 coats the PM flat board with detected sample;
2 picking list bacterium colonies or mixed bacterium are to PM plate streaking purifying;
3 drop Griess reagents are to flat-plate bacterial colony;
4 in certain developing time, and Griess reagent shows red person for waiting to look into positive bacterium colony
5 pickings wait to look into positive bacterium colony repeating step 2,3, the 4 apparent positive bacterium colony of red person of Griess reagent once more, are heterotrophic organism or bacterial flora with nitrification activity.
The microorganism of these addings in ammoniated wastewater is or to separate in the dull and stereotyped growth of PM, and the direct drop of griess reagent is positive, and produces the carbon source of alkali and adds inorganic ammonium salt when cultivating under aerobic conditions with physiological, the bacterial strain that has full nitrogen to wane.
In the specific examples that provides in the present invention, the genus bacillus that one class has nitrification activity is that investigator of the present invention tries soil sample from this caustic lime soil of the North China moisture soil of Fengqiu, Henan Province as confession, through PM plate isolation, purifying, active check, Griess reagent is identified and is obtained through repeated screening.Bacterial strain individual morphology feature sees Table 1:
The individual morphology feature of table 1 14 strain bacteriums
| Bacterial strain number | |
| NBB422 NBB324 NBB319 NBB295 NBB247 NBB204 NBB135 NBB112 NBB72 NBB46 NBB15 NBB58-3 NBB609 NBB19 | Bacterium colony circle on the PM flat board, ecru, translucent, moistening.Gram-positive; Cell ellipse, shaft-like is arranged in pairs; Bacterium colony circle on the gemma PM flat board is arranged, little Huang is flat, transparent, thin.Gram-positive; The shaft-like fusiformis that slightly is of cell, paliform is arranged; Gemma is arranged, bacterium colony circle on the PM flat board spherical in shape, little Huang is flat, transparent, thin.Gram-positive; The shaft-like two ends of cell are point slightly, and paliform is arranged; Gemma is arranged, and sporocyst expands bacterium colony circle on the PM flat board, and the edge is irregular, and is greyish white, opaque, surface drying.Gram-positive; Cell is shaft-like straight or slightly crooked, two terminal circle; Gemma is arranged, and it is less that sporocyst expands on the PM flat board bacterium colony, and circle is greyish white, opaque, projection, surface drying.Gramstaining is weak positive; Cell is shaft-like, the growth catenation; Gemma is arranged, and sporocyst expands bacterium colony ecru on the PM flat board, and flat, little moistening, the edge is irregular.Gram-positive; Cell is shaft-like straight or slightly crooked, becomes catenation; There is on the gemma PM flat board bacterium colony big, circle, white, opaque, moistening, the edge is irregular.Gram-positive; Cell is shaft-like, two terminal circle, and long-chain shape or splayed are arranged; Bacterium colony circle on the gemma PM flat board is arranged, and greyish white, flat, the edge is irregular, and surface drying has gauffer.Gram-positive; Cell is shaft-like, and splayed is arranged; Gemma is arranged, bacterium colony circle on the PM flat board, the edge is irregular, and is greyish white, opaque, surface drying.Gram-positive; The shaft-like two terminal circle of cell, it is inhomogeneous to dye, and the long-chain shape is arranged; Gemma is arranged, and sporocyst expands bacterium colony circle on the PM flat board, and flat, the edge is irregular, and is greyish white, opaque, surface drying.Gram-positive; The shaft-like two terminal circle of cell is arranged in pairs; Gemma sphere, sporocyst are expanded bacterium colony circle on the PM flat board, ecru, low projection, surface wettability.Gram-positive; Cell is shaft-like straight or slightly crooked, the growth catenation; Gemma sphere, sporocyst are expanded bacterium colony lark on the PM flat board, and be flat, big, the edge diffusion.Gram-positive; The shaft-like two terminal circle of cell; Bacterium colony circle on the gemma PM flat board is arranged, sallow, flat, drying has the characteristic gauffer; Gram-positive; The blunt circle in the shaft-like two ends of cell, paliform and Eight characters shape are arranged; Have that bacterium colony is interlacing shape, ecru, surface wettability on the gemma PM flat board; Gram-positive; The blunt circle in the shaft-like two ends of cell, paliform and Eight characters shape are arranged; Gemma is arranged |
Further, the inventor has the genus bacillus experiment and evaluation of heterotrophic nitrification ability to 14 strains on physio-biochemical characteristics, the results are shown in Table 2.
The physio-biochemical characteristics of table 2 14 strain bacteriums
| Bacterial strain number | Catalase | Methylene blue dyeing | Anaerobic growth | VP | VP cultivates pH | The starch hydrolysis | Glucose fermentation | The 7%Nacl growth | The indoles experiment | Nitrate reduction | Xylose utilization | Seminose utilizes |
| NBB422 NBB324 NBB319 NBB295 NBB247 NBB204 NBB135 NBB112 NBB72 NBB46 NBB15 NBB58-3 NBB609 NBB19 | + + + + + + + + + + + + + + | + - - + - + - - | - - - + - - + - + - - + - + | - - - + - - + + + - - + + + | ＜6 ＞7 ＞7 ＜6 ＜6 ＞7 ＜6 ＜6 ＜6 ＞7 ＞7 ＜6 ＜6 ＜6 | + - - + + + + + + + - + + + | + + + + + + + + + + - + + + | + - - - - + + + + - - + + + | - - - - - - - - - - - - - - | + + + + + - + - + + - + + + | + + + + + + + - | + + + - - + + + |
Annotate :+expression positive reaction maybe can utilize;-expression negative reaction maybe can not be utilized.
Therefrom as can be seen, cultivate on the indexs such as PH, starch hydrolysis, glucose fermentation, 7%Nacl growth, indoles experiment, nitrate reduction, xylose utilization and seminose utilization difference to some extent between the 14 strain bacterium at catalase, methylene blue dyeing, anaerobic growth, VP, VP.The classification evaluation that combining form and reference uncle Jie Shi Bacteria Identification handbook are the 9th edition, difference called after bacillus:
Bacillus megaterium Bacillus megaterium NBB-422, CGMCC NO.0554
Bacillus firmus Bacillus firmus NBB-324 CGMCC NO.0555
Bacillus brevis Bacillus brevis NBB-319 CGMCC NO.0556
Bacillus circulans Bacillus circulans NBB-295 CGMCC NO.0557
Bacillus coagulans Bacillus coagulans NBB-247 CGMCC NO.0558
Bacillus lentus Bacillus lentus NBB-204 CGMCC NO.0559
Cured shape bacillus cereus NBB-135 CGMCC NO.0560
Bacillus pumilus Bacillus pumilus NBB-112 CGMCC NO.0561
Bacillus licheniformis Bacillus licheniformis NBB-72 CGMCC NO.0562
Bacillus globisporus Bacillus globisporus NBB-46 CGMCC NO.0563
Bacillus sphaericus Bacillus sphaericus NBB-15 CGMCC NO.0564
Bacillus badius Bacillus badius NBB-58-3 CGMCC NO.0565
Subtilis Bacillus subtilis NBB-609 CGMCC NO.0565 and
Bacillus mycoides Bacillus mycoides NBB-19 CGMCC NO.0586.
These bacteriums are respectively at delivering by specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms's common micro-organisms center preservation on April 9 calendar year 2001 and May 31 calendar year 2001.
In the present invention, the inventor tries soil sample from this caustic lime soil of the North China moisture soil of Fengqiu, Henan Province as confession, through PM plate isolation, purifying, active check, Griess reagent identifies and obtains S-12 and S-27 bacterial strain through repeated screening that S-12 and S-27 bacterial strain individual morphology feature see Table 3.In the present invention, the microorganism that the contriver also is separated to from the black grid in Changshu, Jiangsu yellow soil native and Lianyun Harbour, Jiangsu respectively also has nitrated property, as bacterial strain NH-2, NH-14 etc., NH-2 and NH-14 bacterial strain individual morphology feature
See Table 5:
The individual morphology feature of table 3 bacterial strain S-12 and S-27
| Bacterial strain number | |
| S-12 S-27 | Bacterium colony circle on the PM flat board, lawn is thicker, and surface drying is rough, and coarse a little, the surface is creamy white, not water breakthrough dissolubility pigment secretion.Gram-positive; Cell is shaft-like, sometimes different in size, bacterium colony circle on the tangible bifurcation PM flat board is arranged, lawn is thicker, smooth surface, moistening, be orange, not water breakthrough dissolubility pigment secretion.Gram-positive; Cell is shaft-like, and is different in size sometimes, and thalline is thin slightly than S-12, and tangible bifurcation is arranged. |
Further, the inventor experimentizes to S-12, S-27, NH-2 and the NH-14 with heterotrophic nitrification ability on physio-biochemical characteristics and identifies, the results are shown in Table 4-1, table 4-2, table 4-3.
The physiological and biochemical property (1) of table 4-1 bacterial strain S-12, S-27, NH-2 and NH-14
| Project | NH-2 | NH-14 | S-12 | S-27 | |
| A1 A2 A3 A4 A5 A6 A7 A8 A9 A10 A11 A12 | Contrast cyclohexaamylose cycloheptaamylose dextrin glycogen inulin mannosan polysorbate40 Tween 80 N-acetyl-D-galactosamine N-ACETYL-D-GLUCOSAMINE amarogentin | - - +/- +/- +/- - - + - - - - | - - - - +/- - - + + - - - | + + + + + + - - - + + + | - +/- + + + - - + + - - - |
| B1 B2 B3 B4 B5 B6 B7 B8 B9 B10 B11 B12 | Arabinose D-arabitol arbutin D-cellobiose D-Fructose fucose D-galactolipin D-galacturonic acid gentiobiose maltonic acid alpha-D-glucose m-inositol | - - - - - - - - - +/- +/- - | + - + - + - - - - +/- +/- - | + + + - + - + - +/- + + - | - + - - + - - - - - + - |
The physiological and biochemical property (2) of table 4-2 bacterial strain S-12, S-27, NH-2 and NH-14
| Project | NH-2 | NH-14 | S-12 | S-27 | |
| C1 C2 C3 C4 C5 C6 C7 C8 C9 C10 C11 C12 | α-D-lactose lactulose (lactulose) maltose maltotriose PEARLITOL 25C D-MANNOSE D-melezitose D-melibiose Alpha-Methyl-D-galactoside Beta-methyl-D-galactoside 3-methyl-glucose Alpha-Methyl-D-Glucose glycosides | + - - - + +/- - - - - - - | - - - - - - - - - - - - | + +/- + + - - + - +/- +/- + + | - - - - - - - - - - +/- + |
| D1 D2 D3 D4 D5 D6 D7 D8 D9 D10 D11 D12 | Beta-methyl-D-Glucose glycosides Alpha-Methyl-D-MANNOSE glycosides palatinose D-Psicose D-gossypose L-rhamnose D-ribose salicin red-spotted stonecrop glycan in heptan D-sorbose wood (four) sugarcane sugar | - - - +/- - - - - +/- + - + | +/- - - + - + + + +/- - - - | + - - - + - - - - + - + | - - + + - - + - + + - + |
| E1 E2 E3 E4 E5 E6 E7 E8 E9 E10 E11 E12 | D-Tag D-trehalose turanose xylitol D-wood sugar acetic acid Alpha-hydroxy-butyric acid beta-hydroxy-butyric acid r-hydroxy-butyric acid p-hydroxyl phenylacetic acid KG β-ketone valeric acid | - - - - - + - - - - - - | +/- +/- - - + + + + - - + + | - + - - + + + + + - + + | - + + + + + + + + - + + |
The physiological and biochemical property (3) of table 4-3 bacterial strain S-12, S-27, NH-2 and NH-14
| Project | NH-2 | NH-14 | S-12 | S-27 | |
| F1 F2 F3 F4 F5 F6 F7 F8 F9 F10 F11 F12 | Lactamide D-ALPHA-Hydroxypropionic acid methyl esters Pfansteihl D-malic acid L MALIC ACID pyruvic acid methylsuccinic acid methylpropanoic acid pyruvic acid succinamic acid butanedioic acid N-acetic acid-L-lactamide glutamic acid | - - - - - - - - - - - - | - + + - + + + - + + + - | + + + + + + + +/- + + + - | + + + - + + + + + + + + |
| G1 G2 G3 G4 G5 G6 G7 G8 G9 G10 G11 G12 | L-propionamide D-alanine ALANINE L-alanyl-glycine altheine Pidolidone glycine-Pidolidone L-Glutimic acid Serine butanediamine 2,3-butanediol glycerine | - - - - - +/- +/- +/- - - - - | +/- +/- - - - - +/- +/- - - - + | - + + +/- + + +/- +/- - - - - | +/- - - - +/- +/- + - + + + |
| H1 H2 H3 H4 H5 H6 H7 H8 H9 H10 H11 H12 | Adenosine 2-desoxyadenossine inosine thymidine uridine 5 '-AMP 5 '-monophosphate thymus gland glycosides 5 '-monophosphate crow glycosides 6-phosphoric acid-D-Fructose 1-phosphoric acid-alpha-D-glucose 6-phosphoric acid D-Glucose D-L-alpha-phosphate glycerine | - - - - - +/- +/- - - - - - | - - +/- - - - - - - - +/- +/- | + + + +/- + - - - - - - +/- | - - - + - - - - + + + - |
The individual morphology feature of table 5 bacterial strain NH-2 and NH-14
| Bacterial strain number | |
| NH-2 NH-14 | Bacterium colony circle on the PM flat board, lawn is thicker, and smooth surface, moistening is lemon yellow, does not have water-soluble pigment secretion.Gram-positive; Cell is an elongated rod shape; As seen statospore forms, and gemma is held life partially, and cyst does not expand, the gemma ovalize that comes off.Bacterium colony circle on the PM flat board, lawn is thicker, and the surface is dry, not moistening slightly, is creamy white not water breakthrough dissolubility pigment secretion.Gram-positive; Cell is a rod-short, thick shape; As seen statospore forms, and gemma is held life partially, and cyst does not expand, and gemma is oval. |
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that the S-12 bacterial strain is branch Arthrobacter (Arthrobacter ramosus), therefore with its called after branch Arthrobacter S-12; Branch Arthrobacter S-12 on January 3rd, 2004 in specified depositary institution of Patent Office of the People's Republic of China-Chinese typical culture collection center (CCTCC) preservation, deposit number is CCTCC M203103.The S-27 bacterial strain is sulphur look Arthrobacter (Arthrobacter sulfurous), therefore with its called after sulphur look Arthrobacter S-27, sulphur look Arthrobacter S-27 on January 3rd, 2004 in specified depositary institution of Patent Office of the People's Republic of China-Chinese typical culture collection center (CCTCC) preservation, deposit number is respectively CCTCC M203104.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that the NH-2 bacterial strain is class bacillus firmus (Bacillus pseudofirmus), therefore with its called after class bacillus firmus NH-2; Class bacillus firmus NH-2 on January 3rd, 2004 in specified depositary institution of Patent Office of the People's Republic of China-Chinese typical culture collection center (CCTCC) preservation, deposit number is CCTCC M 203101.The NH-14 bacterial strain is Campinas series bacillus (Paenibacillus campinasensis), therefore with its called after Campinas series bacillus NH-14, Campinas series bacillus NH-14 on January 3rd, 2004 in specified depositary institution of Patent Office of the People's Republic of China-Chinese typical culture collection center (CCTCC) preservation, deposit number is respectively CCTCC M203102.
18 strain bacteriums provided by the invention can be carried out nitrification function separately, also can be at least a and various bacteria unite the execution nitrification function, can also finish denitrogenation together with nitrobacteria and denitrifying bacterium to water body, eliminate the pollution of nitrogen to environment.Wherein receivable nitrobacteria and denitrifying bacterium are meant autotrophy commonly used or heterotrophic nitrification and denitrification bacterium in the water body denitrification technology, as CCTCC-M202043, and CCTCC-M202044 etc.
In the present invention, from the bacterium that is deposited in preservation mechanism, also screen the heterotrophic organism that a class has nitrification activity, as Alcaligenes faecalis (CGMCC No.1.1799), Sarcina lutea (CGMCC No.1.880) etc.That is to say that separate or the bacterioid that screens all can join and contains ammonium nitrogen wastewater by the PM flat band method, after under 20-35 ℃ of static or little whipped state culturing bacterium 15-35 days, the ammonium nitrogen transformation becomes free of contamination dinitrogen gas, containing ammonium nitrogen wastewater becomes clean water.
In the present invention, the present invention also provides soil that a class has nitrification activity or active sludge also can carry out will to contain ammonium nitrogen wastewater and has become clean water.Soil or active sludge that this class has nitrification activity can obtain by the PM flat band method, and the PM flat band method is as follows:
1 coats the PM flat board with detected sample such as soil or active sludge;
2 picking list bacterium colonies or mixed bacterium are to PM plate streaking purifying;
3 drop Griess reagents are to flat-plate bacterial colony;
4 in certain developing time, and Griess reagent shows red person for waiting to look into positive bacterium colony
5 pickings wait to look into positive bacterium colony repeating step 2,3, the 4 apparent positive bacterium colony of red person of Griess reagent once more, are soil or the active sludge of the bacterium with nitrification activity.
But use bacterium provided by the invention or heterotrophic organism group or contain the soil of nitrification activity or the 80-95% of active sludge one-step removal water body ammonium nitrogen.The nitrification that the present invention has overcome the ammonia that present people generally believe is the understanding of being finished by a class autotrophy nitrobacteria and research mistaken ideas, overcome the eutrophication water that exists in the present autotrophy nitrification and denitrification biological denitrification process and needed sudden and violent gas disposal, autotrophy nitrobacteria quantity not sufficient, difficulties such as the many and complex process of biological reaction tank.
Method advantage provided by the invention is:
Energy-conservation, save time, the reaction that is taken place can be in same reactor a step carry out, need not to carry out earlier the removal of COD; The obvious accumulation of no nitrite in the process; The ammonium nitrogen wastewater that the ammonium nitrogen concentration is higher than the high density of 0.5 grams per liter can not suppress the growth of thalline, and ammonium nitrogen decreasing ratio is more than 80%, and denitrification effect is fine, and ammonium nitrogen residual concentration is very low.
In the present invention, the heterotrophic organism with nitrification activity be meant can be from natural soil, mud, waste water or these natural goodses through NH
4 +Can be in the culture system in the dull and stereotyped growth of PM or isolating; And be positive through the direct drop of griess reagent, show NO
2 -Produce; And the carbon source (as formic acid, acetate, pyruvic acid and other organic acid salt) of producing alkali with physiological, when the good air culture of inorganic ammonium salt is supported, the bacterial strain that has full nitrogen to wane.This bacterioid comprises the various bacteriums of genus bacillus, excellent bacillus, Arthrobacter, Rhodopseudomonas.
Below in conjunction with specific embodiment.Further set forth the present invention, should be appreciated that these embodiment only are used to the present invention is described and are not used in restriction the scope of protection of present invention.
There is the same employing national standard of national standard in various units of Shi Yonging in the present invention, the employing industry standard of no national standard, and nitrite concentration is 60.0mg NO
2-N L
-1, represent to contain in every liter of solution 60 milligrams of nitrite nitrogens, nitrate content is 0.18mg N L
-1Represent to contain in every liter of solution 0.18 milligram of nitric nitrogen.
Below in conjunction with specific embodiment.Further set forth the present invention, be to be understood that, these embodiment only are used to the present invention is described and are not used in restriction the scope of protection of present invention, unreceipted concrete experiment condition and method in the following example, usually compile as: soil microorganisms research association " day " according to normal condition, Ye Weiqing etc. translate, Science Press, 1983, the soil microorganisms laboratory method; Permitted volumes such as radiance, Beijing agricultural press, 1986, soil microorganisms analytical procedure handbook; And microbial room of Nanjing Soil Inst., Chinese Academy of Sciences volume, Science Press, 1985, the condition described in the books such as soil microorganisms organon, or connect the condition of advising according to manufacturer.
Embodiment
The preparation of embodiment 1 substratum, simulation ammoniated wastewater and reagent
1.1PM liquid nutrient medium (beef-protein medium) preparation
Claim 3 gram beef extracts, 5 gram peptones are dissolved in and form the PM liquid nutrient medium in 1000 ml distilled waters, add 20 gram agar in above-mentioned unpasteurized liquid PM substratum, form the PM solid medium.
Transfer medium pH to 7.1 with 1N sodium hydroxide.Be divided in the triangular flask, sterilization is fallen dull and stereotyped when substratum is cooled to 56 ℃.The PM liquid nutrient medium is through high pressure liquid chromatographic analysis, and the result is that nitrite and nitrate content are the mark amount: nitrite does not detect, and nitrate content is 0.18mg N L
-1
1.2 the preparation of Griess reagent
1.2.1 Sulphanilic Acid reagent (A liquid): 0.5 gram Sulphanilic Acid (Sulfanilic acid) is dissolved in 150 milliliter 20% the dilute acetic acid solution, stores in brown bottle, refrigerates standby.
1.2.2 alpha-naphthylamine reagent (B liquid): (α-naphthylamine) is added in the dilute acetic acid solution of 20 ml distilled waters and 150 milliliter 20% 0.5 gram alpha-naphthylamine, stores in brown bottle, refrigerates standby.
1.3 the preparation of NB liquid nutrient medium
1.3.1 the preparation of inorganic salt solution
Take by weighing (NH
4)
2SO
42.1 gram, NaH
2PO
40.25 gram, K
2HPO
40.75 gram,
MgSO
47H
2O 0.03 gram, MnSO
4H
2O 0.01 gram,
FeSO
47H
2O 0.01 gram is dissolved in 1000 ml distilled waters,
Transferring the pH value of substratum with 1M NaOH is 7.1, is divided in the triangular flask sterilization.
1.3.2 the preparation of organic carbon solution
1.3.2.1 the preparation of glucose solution: take by weighing 36 gram glucose and be dissolved in the glucose solution that forms 0.20M in 1000 ml distilled waters, sterilization.
1.3.2.2 the preparation of sodium acetate solution: take by weighing 27.2 gram sodium acetate trihydrate and be dissolved in the sodium acetate solution that forms 0.20M in 1000 ml distilled waters, sterilization.
1.3.2.3 the preparation of sodium formate solution: take by weighing 20.8 grams, two water sodium formiates and be dissolved in the sodium formate solution that forms 0.20M in 1000 ml distilled waters, sterilization.
1.3.2.4 the preparation of pyruvic acid solution: draw the 14.0ml pyruvic acid and be dissolved in the pyruvic acid solution that forms 0.20M in 1000 ml distilled waters, sterilization.
Get inorganic salt solution 90 parts (volume ratios) during use and mix for 10 parts, form the NB substratum with organic carbon solution.
1.4 the preparation of simulation ammoniated wastewater
1.4.1 the preparation of inorganic salt solution
Take by weighing (NH
4)
2SO
42.1 gram, NaH
2PO
40.25 gram, K
2HPO
40.75 gram,
MgSO
47H
2O 0.03 gram, MnSO
4H
2O 0.01 gram,
FeSO
47H
2O 0.01 gram is dissolved in 1000 ml distilled waters,
Transferring the pH value of substratum with 1M NaOH is 7.1, is divided in the triangular flask sterilization.
14.2 the preparation of organic carbon solution
1.4.2.1 the preparation of sodium acetate solution: take by weighing 27.2 gram sodium acetate trihydrate and be dissolved in the sodium acetate solution that forms 0.20M in 1000 ml distilled waters, sterilization.
1.4.2.2 the preparation of pyruvic acid solution: draw the 14.0ml pyruvic acid and be dissolved in the pyruvic acid solution that forms 0.20M in 1000 ml distilled waters, sterilization.
14.2.3 the preparation of sodium formate solution: take by weighing 20.8 grams, two water sodium formiates and be dissolved in the sodium formate solution that forms 0.20M in 1000 ml distilled waters, sterilization.
Get inorganic salt solution 90 parts (volume ratios) during use and mix for 10 parts, form the simulation ammoniated wastewater with organic carbon solution.
Embodiment: 2, isolation identification has the heterotrophic organism bacterial strain of nitrification activity
2.1 pedotheque sees Table 6.
The source of each soil sample of table 6
| Sequence number | The sampling position | Specific name | The source | PH (water is carried) |
| 1 2 3 | Lianyun Harbour, Jiangsu, Changshu, Jiangsu, Fengqiu, Henan | North China moisture soil crow grid colour of loess earth | The nonirrigated farmland, topsoil soils paddy field, topsoil soils paddy field, topsoil soils | 8.48 7.41 6.32 |
2.2 take by weighing 1.0 gram wind desiceted soils in 250 milliliters of triangular flasks that contain 50 milliliters of sterile distilled waters, 90r/min shaking table vibration 4 hours.
2.3 soil suspension is coated the PM flat board, three repetitions of each extent of dilution after diluting by 10 times of methods.Cultivate after 7 days for 28 ℃, picking list bacterium colony is to the PM flat board, the purifying of ruling.Microscopy proves purity.
2.4 primary dcreening operation (discriminating of active bacterium)
The heterotrophism bacterial strain after separation and purification that obtains is inoculated in the PM flat board, cultivated 10 days for 28 ℃, Griess reagent directly point drips to flat board, carries out nitrification activity and confirms, and make blank with the flat board that does not connect bacterium.In 1 minute, the Griess reagent colour developing takes on a red color, and showing has nitrite to generate.Repeated authentication, color reaction still is positive, and tentatively confirms as the nitrification activity bacterium.(seeing Table 7).The PM substratum is through high pressure liquid chromatographic analysis, and show that wherein nitrite and nitrate content are the mark amount: wherein nitrite does not detect, and nitrate content is lower than 0.2mg N L
-1, can not constitute and just disturb the result.
The classification of table 7 part active bacterial strain
| Plant (genus) name | Representative strain |
| ( Bacillus ) ( Bacillus megaterium ) ( Bacillus brevis ) ( Bacillus licheniformis ) ( Bacillus circulans ) ( Bacillus firmus ) ( Bacillus coagulans ) ( Bacillus lentus ) ( Bacillus cereus ) ( Bacillus pumilus ) ( Bacillus globisporus ) ( Bacillus sphaericus ) ( Bacillus badius ) ( Bacillus subtilis ) ( Bacillus mycoides ) ( Bacillus pseudofirmus ) ( Paenibacillus campinasensis ) ( Arthrobacter ramosus ) ( Arthrobacter sulfureus ) | NBB422，NBB217 NBB319，WY-2 NBB072，WX-2 NBB295，WR-8 NBB324，WR-9 NBB247，NBB300 NBB204，NBB118 NBB135，NBB310 NBB112，NBB557 NBB46，NBB534 NBB15，NBB614 NBB58-3，NBB101 NBB609，NBB617 NBB19，NBB59 NH-2，NH-19 NH-14，NH-22 S-12，S-4 S-27，S-11 |
2.5 multiple sieve
The representative bacterial strain that activity is stronger when choosing primary dcreening operation carries out.Scrape and get the pure bacterium lawn that grows in the PM flat board and go into 30 ml sterile waters, it fully is uniformly dispersed makes bacteria suspension.Inoculate 1 milliliter of bacteria suspension respectively and go into to contain in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum that different organism are carbon source every group of three repetitions.And make blank with the substratum of not inoculating.28 ℃ of static cultivations are after 21 days, 4 ℃ of centrifugal 15min of following 5000g of nutrient solution, the nitrite concentration in the Griess reagent method colorimetric estimation supernatant liquor.Bacterial classification sample cultivation supernatant colourimetric number subtracts the difference of blank supernatant colourimetric number greater than 0.3mg N L
-1The time be judged to the active bacterium (seeing Table 8) of heterotrophic nitrification.
2.6 the physiology of active bacterial strain is identified
With reference to uncle Jie Shi Bacteria Identification handbook the 9th edition.Characteristic of bacteria sees Table 1 and table 2.
The colourimetric number of each bacterial strain of table 8 (is represented NO with nitrite
2 --N mg L
-1)
Embodiment: the evaluation of 3 bacillus megaterium NBB-422
1. colony morphology characteristic: after 28 ℃ of good air cultures of constant temperature on the nutrient agar medium PM flat board are supported 4 days, the bacterium colony circle, ecru, translucent, moistening.
2. morphological features: Gram-positive; Cell ellipse, shaft-like is arranged in pairs; Gemma is arranged
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 15.0mg NO
2-N L
-1, indivedual individual plants can reach 23mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is bacillus megaterium (Bacillusmegaterium).Therefore with its called after bacillus megaterium NBB-422.Bacillus megaterium NBB-422 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0554.
Embodiment: the evaluation of 4 bacillus firmus NBB-324
1. colony morphology characteristic: after 28 ℃ of good air cultures of constant temperature on the nutrient agar medium PM flat board are supported 4 days, the bacterium colony circle, little Huang is flat, transparent, thin.
2. morphological features: Gram-positive; Cell is shaft-like slightly to become fusiformis, and paliform is arranged; There is gemma spherical in shape.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 25.0mg NO
2-N L
-1, indivedual individual plants can reach 36mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is bacillus firmus (Bacillusfirmus).Therefore with its called after bacillus firmus NBB-324.Bacillus firmus NBB-324 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0555.
Embodiment: the evaluation of 5 bacillus brevis NBB-319
1. colony morphology characteristic: after 28 ℃ of good air cultures of constant temperature on the nutrient agar medium PM flat board are supported 4 days, the bacterium colony circle, little Huang is flat, transparent, thin.
2. morphological features: Gram-positive; The shaft-like two ends of cell are point slightly, and paliform is arranged; Gemma is arranged, and sporocyst expands.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 15.0mg NO
2-N L
-1, indivedual individual plants can reach 26mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is bacillus brevis (Bacillusbrevis).Therefore with its called after bacillus brevis NBB-319.Bacillus brevis NBB-319 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0556.
Embodiment: the evaluation of 6 Bacillus circulans NBB-295
1. colony morphology characteristic: go up after 28 ℃ of good air cultures of constant temperature support 4 days at nutrient agar plate (PM flat board), the bacterium colony circle, the edge is irregular, and is greyish white, opaque, surface drying.
2. morphological features: Gram-positive; Cell is shaft-like or slightly crooked, two terminal circle; Gemma is arranged, and sporocyst expands.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 15.0mgNO
2-N L
-1, indivedual individual plants can reach 22mgNO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is Bacillus circulans (Bacilluscirculans).Therefore with its called after Bacillus circulans NBB-295.Bacillus circulans NBB-295 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0557.
Embodiment: the evaluation of 7 Bacillus coagulans NBB-247
1. colony morphology characteristic: after 28 ℃ of good air cultures of constant temperature on the nutrient agar medium PM flat board were supported 4 days, bacterium colony was less, and circle is greyish white, and is opaque, projection, surface drying.
2. morphological features: Gram-positive; Cell is shaft-like, and the long-chain shape is arranged; Gemma is arranged, and sporocyst expands.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 15.0mg NO
2-N L
-1, indivedual individual plants can reach 20mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is Bacillus coagulans (Bacilluscoagulans).Therefore with its called after Bacillus coagulans NBB-247.Bacillus coagulans NBB-247 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0558.
Embodiment: the evaluation of 8 bacillus lentus NBB-204
1. colony morphology characteristic: after 28 ℃ of good air cultures of constant temperature on the nutrient agar medium PM flat board are supported 4 days, the bacterium colony ecru, flat, little moistening, the edge is irregular.
2. morphological features: Gram-positive; Cell is shaft-like straight or slightly crooked, catenation; Gemma is arranged.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 20.0mg NO
2-N L
-1, indivedual individual plants can reach 27mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is bacillus lentus (Bacilluslentus).Therefore with its called after bacillus lentus NBB-204.Bacillus lentus NBB-204 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0559.
Embodiment: the evaluation of 9 cured shape genus bacillus NBB-135
1. colony morphology characteristic: after 28 ℃ of good air cultures of constant temperature on the nutrient agar medium PM flat board were supported 4 days, bacterium colony was big, circle, and white, opaque, moistening, the edge is irregular.
2. morphological features: Gram-positive; Cell is shaft-like, two terminal circle, and long-chain shape or splayed are arranged; Gemma is arranged.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 10.0mg NO
2-N L
-1, indivedual individual plants can reach 17mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is a cured shape genus bacillus (Bacilluscereus).Therefore with the cured shape genus bacillus of its called after NBB-135.Cured shape genus bacillus NBB-135 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0560.
Embodiment: the evaluation of 10 bacillus pumilus NBB-112
1. colony morphology characteristic: after 28 ℃ of good air cultures of constant temperature on the nutrient agar medium PM flat board are supported 4 days, the bacterium colony circle, greyish white, flat, the edge is irregular, and surface drying has gauffer.
2. morphological features: Gram-positive; Cell is shaft-like, and splayed is arranged; Gemma is arranged.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 10mgNO
2-N L
-1, indivedual individual plants can reach 18mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is bacillus pumilus (Bacilluspumilus).Therefore with its called after bacillus pumilus NBB-112.Bacillus pumilus NBB-112 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0561.
Embodiment: the evaluation of 11 Bacillus licheniformis NBB-72
1. colony morphology characteristic: after 28 ℃ of good air cultures of constant temperature on the nutrient agar medium PM flat board are supported 4 days, the bacterium colony circle, greyish white, the edge is irregular, and is opaque, surface drying.
2. morphological features: Gram-positive; The shaft-like two terminal circle of cell, it is inhomogeneous to dye, and the long-chain shape is arranged; Gemma is arranged, and sporocyst expands.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 25.0mgNO
2-N L
-1, indivedual individual plants can reach 37mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is Bacillus licheniformis (Bacilluslicheniformis).Therefore with its called after Bacillus licheniformis NBB-072.Bacillus licheniformis NBB-072 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0562.
Embodiment: the evaluation of 12 bacillus globisporus NBB-46
1. colony morphology characteristic: after 28 ℃ of good air cultures of constant temperature on the nutrient agar medium PM flat board are supported 4 days, the bacterium colony circle, greyish white, the edge is irregular, and is opaque, surface drying.
2. morphological features: Gram-positive; The shaft-like two terminal circle of cell is arranged in pairs; The gemma sphere, sporocyst expands.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 25.0mg NO
2-N L
-1, indivedual individual plants can reach 34mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is bacillus globisporus (Bacillusglobisporus).Therefore with its called after bacillus globisporus NBB-046.Bacillus globisporus NBB-046 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0563.
Embodiment: the evaluation of 13 Bacillus sphaericus NBB-15
1. colony morphology characteristic: go up after 28 ℃ of good air cultures of constant temperature support 4 days bacterium colony circle, ecru, low projection, surface wettability at nutrient agar plate (PM flat board).
2. morphological features: Gram-positive; Cell is shaft-like straight or slightly crooked, and the long-chain shape is arranged; The gemma sphere, sporocyst expands.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 20.0mg NO
2-N L
-1, indivedual individual plants can be up to 28mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is Bacillus sphaericus (BacillusSphaericus).Therefore with its called after Bacillus sphaericus NBB-015.Bacillus sphaericus NBB-015 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0564.
Embodiment: the evaluation of 14 bacillus badius NBB-58-3
1. colony morphology characteristic: after 28 ℃ of good air cultures of constant temperature on the nutrient agar medium PM flat board are supported 4 days, the bacterium colony circle, lark, flat, drying.
2. morphological features: Gram-positive; The shaft-like two terminal circle of cell; Gemma is arranged.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 25.0mg NO
2-N L
-1, indivedual individual plants can be up to 32mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is bacillus badius (Bacillusbadius).Therefore with its called after bacillus badius NBB-58-3.Bacillus badius NBB-58-3 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0565.
Embodiment: the evaluation of 15 subtilis NBB-609
1. colony morphology characteristic: after 28 ℃ of good air cultures of constant temperature on the nutrient agar medium PM flat board are supported 4 days, the bacterium colony circle, sallow, flat, drying has the characteristic gauffer.
2. morphological features: Gram-positive; The blunt circle in the shaft-like two ends of cell, paliform and Eight characters shape are arranged; Gemma is arranged.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 25.0mg NO
2-N L
-1, indivedual individual plants can be up to 38mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is subtilis (Bacillussubtilis).Therefore with its called after subtilis NBB-609.Subtilis NBB-609 in April 9 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0566.
Embodiment: the evaluation of 16 bacillus mycoides NBB-19
1. colony morphology characteristic: go up after 28 ℃ of good air cultures of constant temperature support 4 days at nutrient agar plate (PM flat board), bacterium colony becomes interlacing shape, ecru, surface wettability.
2. morphological features: Gram-positive; The blunt circle in the shaft-like two ends of cell, paliform and Eight characters shape are arranged; Gemma is arranged.
3. the major physiological biochemical characteristic sees Table 2, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 25.0mg NO
2-N L-1, indivedual individual plants can be up to 32mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is bacillus mycoides (Bacillusmycoides).Therefore with its called after bacillus mycoides NBB-19.Bacillus mycoides NBB-19 in May 31 calendar year 2001 in specified depositary institution of Patent Office of the People's Republic of China-China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, deposit number is CGMCC NO.0586.
The evaluation of embodiment 17 branch Arthrobacter S-12
1. colony morphology characteristic: go up after 28 ℃ of good air cultures of constant temperature support 4 days at nutrient agar plate (PM flat board), the bacterium colony circle, lawn is thicker, and surface drying is rough, and coarse a little, the surface is creamy white, water breakthrough dissolubility pigment secretion.
2. morphological features: Gram-positive; Cell is shaft-like, and is different in size sometimes, and tangible bifurcation is arranged.
3. the major physiological biochemical characteristic sees Table 4-1, table 4-2, table 4-3, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 11.0mgNO
2-N L
-1, indivedual individual plants can be up to 20mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is branch Arthrobacter (Arthrobacter ramosus).Therefore with its called after branch Arthrobacter S-12.Branch Arthrobacter S-12 on January 3rd, 2004 in specified depositary institution of Patent Office of the People's Republic of China-Chinese typical culture collection center (CCTCC) preservation, deposit number is CCTCC M 203103.
The evaluation of embodiment 18 sulphur look Arthrobacter S-27
1. colony morphology characteristic: go up after 28 ℃ of good air cultures of constant temperature support 4 days at nutrient agar plate (PM flat board), the bacterium colony circle, lawn is thicker, and smooth surface, moistening is orange, water breakthrough dissolubility pigment secretion.
2. morphological features: Gram-positive; Cell is shaft-like, and is different in size sometimes, and thalline is thin slightly than S-12, and tangible bifurcation is arranged.
3. the major physiological biochemical characteristic sees Table 4-1, table 4-2, table 4-3, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 5.0mgNO
2-N L
-1, indivedual individual plants can be up to 14mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is sulphur look Arthrobacter (Arthrobacter sulfurous).Therefore with its called after sulphur look Arthrobacter S-27.Sulphur look Arthrobacter S-27 on January 3rd, 2004 in specified depositary institution of Patent Office of the People's Republic of China-Chinese typical culture collection center (CCTCC) preservation, deposit number is CCTCC M203104.
The evaluation of embodiment 19 Campinas series bacillus NH-14
1. colony morphology characteristic: go up after 28 ℃ of good air cultures of constant temperature support 4 days at nutrient agar plate (PM flat board), the bacterium colony circle, lawn is thicker, and the surface is dry, not moistening slightly, is creamy white water breakthrough dissolubility pigment secretion.
2. morphological features: Gram-positive; Cell is a rod-short, thick shape; As seen statospore forms, and gemma is held life partially, and cyst does not expand, and gemma is oval.
3. the major physiological biochemical characteristic sees Table 4-1, table 4-2, table 4-3, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 7.0mgNO
2-N L
-1, indivedual individual plants can be up to 12mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is Campinas series bacillus (Paenibacillus campinasensis).Therefore with its called after Campinas series bacillus NH-14.Campinas series bacillus NH-14 on January 3rd, 2004 in specified depositary institution of Patent Office of the People's Republic of China-Chinese typical culture collection center (CCTCC) preservation, deposit number is CCTCC M203102.
The evaluation of embodiment 20 class bacillus firmus NH-2
1. colony morphology characteristic: go up after 28 ℃ of good air cultures of constant temperature support 4 days at nutrient agar plate (PM flat board), the bacterium colony circle, lawn is thicker, and smooth surface, moistening is lemon yellow, does not have water-soluble pigment secretion.
2. morphological features: Gram-positive; Cell is an elongated rod shape; As seen statospore forms, and gemma is held life partially, and cyst does not expand, the gemma ovalize that comes off.
3. the major physiological biochemical characteristic sees Table 4-1, table 4-2, table 4-3, and the suitableeest culture temperature is 28-32 ℃, and pH7.0-7.2 is aerobic.
4. ammoxidation activity: the nitrite concentration of cultivating accumulation in 14 days at the 90-100r/min shaking table is higher than 27.0mgNO
2-N L
-1, indivedual individual plants can be up to 35mg NO
2-N L
-1More than.
Classification with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition identifies that this bacterial strain is class bacillus firmus (Bacillus pseudofirmus).Therefore with its called after class bacillus firmus NH-2.Class bacillus firmus NH-2 on January 3rd, 2004 in specified depositary institution of Patent Office of the People's Republic of China-Chinese typical culture collection center (CCTCC) preservation, deposit number is CCTCC M203101.
The denitrogenation of embodiment 21 bacillus megaterium NBB-422
21.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
21.2 pre-the cultivation:
Go into to be equipped with L 21.2.1 connect the lawn of a ring NBB-422 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
21.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
21.3 result: mixed culture has removed 74% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 93.5%, and nitrite concentration only is 0.15mg N L
-1
The denitrogenation of embodiment 22 bacillus firmus NBB-324
22.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
22.2 pre-the cultivation:
Go into to be equipped with L 22.2.1 connect the lawn of a ring NBB-324 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
22.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
22.3 result: mixed culture has removed 67.4% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 83.7%, and nitrite concentration only is 0.18mg N L
-1
The denitrogenation of embodiment 23 bacillus brevis NBB-319
23.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
23.2 pre-the cultivation:
Go into to be equipped with L 23.2.1 connect the lawn of a ring NBB-319 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
23.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
23.3 result: mixed culture has removed 71.2% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 85.1%, and nitrite concentration only is 0.22mg N L
-1
The denitrogenation of embodiment 24 Bacillus circulans NBB-295
24.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
24.2 pre-the cultivation:
Go into to be equipped with L 24.2.1 connect the lawn of a ring NBB-295 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
24.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
24.3 result: mixed culture has removed 62.8% of the full nitrogen of system after cultivating 21 days, the ammonium nitrogen removal efficiency is 74.3%, and nitrite concentration only is 0.18mg N L
-1
The denitrogenation of embodiment 25 Bacillus coagulans NBB-247
25.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
25.2 pre-the cultivation:
Go into to be equipped with L 25.2.1 connect the lawn of a ring NBB-247 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
25.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
25.3 result: mixed culture has removed 57.2% of the full nitrogen of system after cultivating 21 days, the ammonium nitrogen removal efficiency is 73.5%, and nitrite concentration only is 0.15mg N L
-1
The denitrogenation of embodiment 26 bacillus lentus NBB-204
26.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
26.2 pre-the cultivation:
Go into to be equipped with L 26.2.1 connect the lawn of a ring NBB-204 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
26.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
26.3 result: mixed culture has removed 69.7% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 84.1%, and nitrite concentration only is 0.16mg N L
-1
The denitrogenation of embodiment 27 bacillus cereus NBB-135
27.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
27.2 pre-the cultivation:
Go into to be equipped with L 27.2.1 connect the lawn of a ring NBB-135 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
27.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
27.3 result: mixed culture has removed 60.5% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 72.7%, and nitrite concentration only is 0.45mg N L
-1
The denitrogenation of embodiment 28 bacillus pumilus NBB-112
28.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
28.2 pre-the cultivation:
Go into to be equipped with L 28.2.1 connect the lawn of a ring NBB-112 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
28.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
28.3 result: mixed culture has removed 72.5% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 87.7%, and nitrite concentration only is 0.20mg N L
-1
The denitrogenation of embodiment 29 Bacillus licheniformis NBB-72
29.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
29.2 pre-the cultivation:
Go into to be equipped with L 29.2.1 connect the lawn of a ring NBB-72 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
29.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 14 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
29.3 result: mixed culture has removed 60.3% of the full nitrogen of system after cultivating 14 days, the ammonium nitrogen removal efficiency is 73.5%, and nitrite concentration only is 0.15mg N L
-1。
The denitrogenation of embodiment 30 bacillus globisporus NBB-46 bacterial strains
30.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
30.2 pre-the cultivation:
Go into to be equipped with L 30.2.1 connect the lawn of a ring NBB-46 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
30.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
30.3 result: mixed culture has removed 73.3% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 92.7%, and nitrite concentration only is 0.15mg N L
-1
The denitrogenation of embodiment 31 Bacillus sphaericus NBB-15
31.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
31.2 pre-the cultivation:
Go into to be equipped with L 31.2.1 connect the lawn of a ring NBB-15 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
31.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
31.3 result: mixed culture has removed 55.7% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 73.2%, and nitrite concentration only is 0.18mg N L
-1
The denitrogenation of embodiment 32 bacillus badius NBB-58-3
32.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
32.2 pre-the cultivation:
Go into to be equipped with L 32.2.1 connect the lawn of a ring NBB-58-3 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
32.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
32.3 result: mixed culture has removed 64.2% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 85.7%, and nitrite concentration only is 0.15mg N L
-1
The denitrogenation of embodiment 33 subtilis NBB-609
33.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
33.2 pre-the cultivation:
Go into to be equipped with L 33.2.1 connect the lawn of a ring NBB-609 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
33.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
33.3 result: mixed culture has removed 68.4% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 87.1%, and nitrite concentration only is 0.18mg N L
-1
The denitrogenation of embodiment 34 bacillus mycoides NBB-19
34.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
34.2 pre-the cultivation:
Go into to be equipped with L 34.2.1 connect the lawn of a ring NBB-19 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
34.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
34.3 result: mixed culture has removed 62.7% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 76.3%, and nitrite concentration only is 0.18mgN L
-1
The denitrogenation of embodiment 35 branch Arthrobacter S-12
35.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
35.2 pre-the cultivation:
Go into to be equipped with L 35.2.1 connect the lawn of a ring S-12 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
35.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
35.3 result: mixed culture has removed 80% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 96.5%, and nitrite concentration only is 0.15mg N L
-1
The denitrogenation of embodiment 36 sulphur look Arthrobacter S-27
36.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
36.2 pre-the cultivation:
Go into to be equipped with L 36.2.1 connect the lawn of a ring S-27 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
36.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
36.3 result: mixed culture has removed 45% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 56.5%, and nitrite concentration only is 0.15mg N L
-1
The denitrogenation of embodiment 37 Campinas series bacillus NH-14
37.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
37.2 pre-the cultivation:
Go into to be equipped with L 37.2.1 connect the lawn of a ring NH-14 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
37.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 21 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
37.3 result: mixed culture has removed 77% of the full nitrogen of system after cultivating 21 days, the ammonium nitrogen removal efficiency is 88.2%, and nitrite concentration only is 0.15mg N L
-1
The denitrogenation of embodiment 38 class bacillus firmus NH-2
38.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
38.2 pre-the cultivation:
Go into to be equipped with L 38.2.1 connect the lawn of a ring NH-2 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
38.2.2 by 2% inoculum size, the bacterium liquid that will cultivate in advance is connected to for 1 milliliter and contains 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 14 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
38.3 result: mixed culture has removed 73% of the full nitrogen of system after cultivating 14 days, the ammonium nitrogen removal efficiency is 88.5%, and nitrite concentration only is 0.15mg N L
-1
Embodiment: the combined denitrification of 39 bacillus globisporus NBB-46 and bacillus megaterium NBB-422
39.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
39.2 pre-the cultivation:
Go into to be equipped with L 39.2.1 connect the lawn of a ring bacillus globisporus NBB-46 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
Go into to be equipped with 50ml NB substratum (0.015mol L 37.2.2 connect a ring bacillus megaterium NBB-422 lawn from the PM inclined-plane
-1Sodium acetate is a carbon source) the 250ml Erlenmeyer flask in, at 30 ℃, 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
39.3 cultivate: NBB-422 (culture of 37.2.2) and NBB-46 (culture of 37.2.1) mix with volume ratio at 1: 1, again with 2% 1 milliliter of access of inoculation mixed bacteria liquid of measuring with 0.060mol L
-1Acetate be the simulation ammonium nitrogen wastewater (compound method with 1.4) of carbon source at 30 ℃, static cultivation 28 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
39.4 result: mixed culture has removed 77.5% of the full nitrogen of system after cultivating 28 days, the ammonium nitrogen removal efficiency is 94.7%, and nitrite concentration only is 0.25mg N L
-1
Embodiment: the combined denitrification of 40 bacillus cereus NBB-135 and Bacillus circulans NBB-295
40.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
40.2 pre-the cultivation:
Go into to be equipped with L 40.2.1 connect the lawn of a ring bacillus cereus NBB-135 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
Go into to be equipped with 50ml NB substratum (0.015mol L 40.2.2 connect the lawn of a ring Bacillus circulans NBB-295 from the PM inclined-plane
-1Sodium acetate is a carbon source) the 250ml Erlenmeyer flask in, at 30 ℃, 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
40.3 cultivate: NBB-295 (culture of 38.2.2) and NBB-135 (culture of 38.2.1) mix by 3: 7 (volume ratio), again with 2% 1 milliliter of access of inoculation mixed bacteria liquid of measuring with 0.060mol L
-1Acetate be the simulation ammonium nitrogen wastewater (prescription with 1.4) of carbon source at 30 ℃, static cultivation 21 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
37.4 result: mixed culture has removed 61.3% of the full nitrogen of system after cultivating 21 days, the ammonium nitrogen removal efficiency is 75.1%, and nitrite concentration only is 0.32mg N L
-1
Embodiment: 41 genus bacillus (bacillus cereus NBB-135, bacillus pumilus NBB-112, Bacillus circulans NBB-295, Bacillus licheniformis NBB-72) and other nitrifier: the combined denitrification effect of Arthrobacter globiformis WR-2 (CCTCC M202043)
41.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
39.2 pre-the cultivation:
Go into to be equipped with 50ml NB substratum (0.015mol L 41.2.1 respectively connect the lawn of a ring bacillus cereus NBB-135, bacillus pumilus NBB-112, Bacillus circulans NBB-295, Bacillus licheniformis NBB-072 from the PM inclined-plane
-1Sodium acetate is a carbon source) the 250ml Erlenmeyer flask in, at 30 ℃, 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
Go into to be equipped with 250 milliliters of Erlenmeyer flasks of 70mlPM liquid nutrient medium 41.2.2 connect the lawn of a global shape Arthrobacter WR-2 from the PM inclined-plane, at 30 ℃, 120-130r/min shaking table shaking culture 28 hours, it is thalline OD that bacterium liquid is transferred the OD value
600=0.45.
41.3 cultivate: the mixed culture (culture of 39.2.1) of nitrifiers such as the culture of Arthrobacter globiformis WR-2 (culture of 39.2.2) and bacillus cereus NBB-135 mixes by 1: 1 (volume ratio), connects the 1ml mixed bacteria liquid and goes into the L with 0.075mol
-1Acetate be the simulation ammonium nitrogen wastewater (prescription with 1.4) of carbon source at 30 ℃, static cultivation 21 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
41.4 result: mixed culture has removed 76.3% of the full nitrogen of system after cultivating 21 days, the ammonium nitrogen removal efficiency is 87.9%, and nitrite concentration only is 0.10mg N L
-1
The separation of the heterotrophic nitrification active bacterial strain of embodiment 42 different areas Different Soil samples counting 42.1, separates and has differentiated a large amount of representative heterotrophic nitrification active bacterial strains (table 10) from China's 9 kinds of dissimilar soil samples in 4 areas (table 9) by the identical program of embodiment 2 and method.
The source of each soil sample of table 9
| Numbering | The sampling position | Specific name | The source | pH |
| 1 2 3 4 5 6 7 8 9 | Yingtan Jiangxi, Yingtan Jiangxi, Yangling Shaanxi Fengqiu Fengqiu Jiangxi Yingtan Changshu, Jiangsu Province Changshu, Jiangsu Province Lianyungang of Jiangsu | The black grid colour of loess earth of moisture soil North China, chilli oil soil North China moisture soil Quaternary Red Clays Quaternary Red Clays Quaternary Red Clays crow grid soil | The nonirrigated farmland, the topsoil soils nonirrigated farmland, the topsoil soils paddy field, the topsoil soils nonirrigated farmland, the topsoil soils paddy field, the topsoil soils waste hillside, the topsoil paddy field, the topsoil soils nonirrigated farmland, the topsoil soils paddy field, topsoil soils | 8.04 8.48 8.24 5.46 6.13 4.88 7.41 6.41 6.32 |
The nitrification activity bacterium number statistics of each soil sample of table 10
| Numbering | Soil sample | Fresh soil (Cell/g dry ground) | Domestication back (Cell/g dry ground) |
| 1 2 3 4 5 6 7 8 9 | Chilli oil soil, North China, nonirrigated farmland moisture soil, North China, nonirrigated farmland moisture soil, paddy field red clay in the quaternary period, nonirrigated farmland red clay in the quaternary period, paddy field red clay in the quaternary period, waste hillside crow grid soil, paddy field crow grid soil, the nonirrigated farmland yellow soil, the paddy field | 5.55×10 6 2.71×10 6 1.99×10 6 1.16×10 7 1.88×10 6 3.23×10 5 3.11×10 7 7.06×10 7 1.06×10 8 | 4.03×10 12 5.95×10 10 2.23×10 12 1.72×10 11 4.02×10 10 7.95×10 7 3.05×10 10 1.72×10 11 |
42.2, belong to a plurality of genus kinds such as genus arthrobacter, erwinia, corynebacterium, micromonospora, bacillus, Zoogloea, Rhod, Alkaligenes through identifying these active bacterial strains with reference to uncle Jie Shi Bacteria Identification handbook the 9th edition.
The denitrogenation of embodiment 43 different soils samples
43.1 the source of each soil sample sees Table 11.
The source of each soil sample of table 11
| Sequence number | The sampling position | Specific name | The source | PH (water is carried) |
| 1 2 3 4 5 6 | Lianyun Harbour, Jiangsu, Changshu, Yingtan Jiangsu, Yingtan Jiangxi, Yingtan Jiangxi, Jiangxi, Fengqiu, Henan | North China moisture soil red clay in quaternary period red clay in quaternary period red clay in quaternary period crow grid colour of loess earth | The nonirrigated farmland, the topsoil soils nonirrigated farmland, the old paddy field of topsoil soils, the topsoil soils waste hillside, the topsoil paddy field, the topsoil soils paddy field, topsoil soils | 8.48 5.43 4.81 5.69 7.41 6.32 |
43.2 take by weighing 10 gram wind desiceted soils in 500 milliliters of triangular flasks that contain 100 milliliters of sterile distilled waters, 90r/min shaking table vibration 4 hours.
43.3 10 milliliters of soil suspensions of shaking culture are connected to contain 0.060mol L
-1The simulation ammoniated wastewater of pyruvic acid (compound method with 1.4) is at 30 ℃, static cultivation 21 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
43.4 result: the soil suspension culture has removed the 76.5-89.8% of the full nitrogen of system after cultivating 21 days, the ammonium nitrogen removal efficiency is 97.3-98.9%, and nitrite concentration only is 0.02-0.73mg NL
-1
The denitrification effect of each soil sample of table 12
| Handle | The red soil forest land | Red soil peanut ground | The red soil paddy field | The moisture soil nonirrigated farmland | The yellow soil paddy field | Crow grid soil paddy field |
| Full nitrogen removal efficiency nitrite (the mg NL of ammonia nitrogen removal frank -1) | 98.1％ 76.5％ 0.09 | 97.3％ 84.7％ 0.07 | 96.8％ 79.1％ 0.04 | 98.9％ 89.8％ 0.73 | 98.8％ 85.8％ 0.09 | 97.4％ 84.0％ 0.02 |
Embodiment: the denitrogenation of 44 immobilization bacillus megaterium NBB, 422 bacterial strains
44.1 the preparation of the fixed film of bacillus megaterium NBB422
44.1.1 bacillus megaterium NBB422 concentrates the preparation of thalline
Connect a ring lawn from the PM inclined-plane and go into to be equipped with L with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours.By 1% amount inoculation bacterium liquid is connected to 0.015mol L is housed
-1Sodium acetate is in 500 milliliters of Erlenmeyer flasks of 150 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 72 hours.At 5000r/min, 4 ℃ centrifugal 15 minutes down, wash centrifugal twice with physiological saline after, be suspended in 15ml physiological saline.
44.1.2 bacillus megaterium NBB422WR-2 fixes
Thalline be will concentrate and 20%PVA and 1.0mol L joined
-1CaCl
2Mixing solutions in, be tiled on the poly (methyl methacrylate) plate after stirring evenly, place refrigerator ,-20 ℃ of freeze overnight are at room temperature thawed again.Repeated freezing thaws 3-4 time, has the distilled water thorough washing to show, promptly gets tabular fixation cell after birth.
44.1.3 fixed film reactor and denitrogenation experiment
With the blue fixing and assembling biological denitrification reactor of the fixation cell after birth usage of gained.It is 1600 milliliters that reactor is contained liquid measure, with 0.015mol L
-1Acetate is the simulation ammonium nitrogen wastewater (compound method is with 1.4) of carbon source, and wherein ammonium nitrogen concentration is kept to 100mg N L
-1, place 28 ℃ of constant incubators to activate, treat that cytoactive is stable after, carry out the denitrogenation experiment.In the experimentation, the control dissolved oxygen concentration is the 5-8 mg/litre.Sample analysis ammonium nitrogen, nitrite nitrogen and nitric nitrogen concentration wherein at regular intervals takes a morsel.The result is presented at and cultivates after 16 days, and the ammonium nitrogen removal efficiency is 87.7%, and nitrite concentration only is 0.22mgN L
-1
The combined denitrification of embodiment 45 bacillus pumilus NBB-112 and plant bacillus pumilis WO-8 (CCTCC M202044)
45.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
45.2 pre-the cultivation:
Go into to be equipped with L 45.2.1 connect the lawn of a ring bacillus pumilus NBB-112 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
Go into to be equipped with 50ml NB substratum (0.015mol L 45.2.2 connect the lawn of a ring plant bacillus pumilis WO-8 from the PM inclined-plane
-1Sodium acetate is a carbon source) the 250ml Erlenmeyer flask in, at 30 ℃, 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
45.3 cultivate: WO-8 (culture of 45.2.2) and NBB-112 (culture of 45.2.1) mix with volume ratio at 1: 1, again with 2% 1 milliliter of access of inoculation mixed bacteria liquid of measuring with 0.075mol L
-1Acetate be the simulation ammonium nitrogen wastewater (compound method with 1.4) of carbon source at 30 ℃, static cultivation 21 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
45.4 result: mixed culture has removed 57.5% of the full nitrogen of system after cultivating 21 days, the ammonium nitrogen removal efficiency is 72.7%, and nitrite concentration only is 0.45mg N L
-1
The combined denitrification of embodiment 46 bacillus cereus NBB-135 and plant bacillus pumilis WO-8 (CCTCCM202044)
46.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
46.2 pre-the cultivation:
Go into to be equipped with L 46.2.1 connect the lawn of a ring bacillus cereus NBB-135 from the PM inclined-plane with 0.015mol
-1Sodium acetate is in 250 milliliters of Erlenmeyer flasks of 50 milliliters of NB substratum of carbon source, at 30 ℃, and 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
Go into to be equipped with 50ml NB substratum (0.015mol L 46.2.2 connect the lawn of a ring plant bacillus pumilis WO-8 from the PM inclined-plane
-1Sodium acetate is a carbon source) the 250ml Erlenmeyer flask in, at 30 ℃, 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
46.3 cultivate: WO-8 (culture of 46.2.2) and NBB-135 (culture of 46.2.1) mix by 3: 7 (volume ratio), again with 2% 1 milliliter of access of inoculation mixed bacteria liquid of measuring with 0.075mol L
-1Acetate be the simulation ammonium nitrogen wastewater (compound method with 1.4) of carbon source at 30 ℃, static cultivation 21 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
46.4 result: mixed culture has removed 61.3% of the full nitrogen of system after cultivating 21 days, the ammonium nitrogen removal efficiency is 75.1%, and nitrite concentration only is 0.32mg N L
-1
Embodiment 47 genus bacillus (bacillus cereus NBB-135, bacillus pumilus NBB-112, Bacillus circulans NBB-295, Bacillus licheniformis NBB-072) and other nitrifier: the mixed culture of branch Arthrobacter S-12, sulphur look Arthrobacter S-27 and the combined denitrification effect of denitrifying bacteria plant bacillus pumilis WO-8 (CCTCCM202044)
47.1 substratum is with 0.015mol L
-1Sodium acetate is the NB substratum of carbon source, and compound method is with 1.3.
47.2 pre-the cultivation:
Go into to be equipped with 50ml NB substratum (0.015mol L 47.2.1 respectively connect the lawn of a ring bacillus cereus NBB-135, bacillus pumilus NBB-112, Bacillus circulans NBB-295, Bacillus licheniformis NBB-072, branch Arthrobacter S-12 and sulphur look Arthrobacter S-27 from the PM inclined-plane
-1Sodium acetate is a carbon source) the 250ml Erlenmeyer flask in, at 30 ℃, 120-130r/min shaking table shaking culture 24 hours, it is OD that bacterium liquid is transferred the OD value
600=0.45.
Go into to be equipped with 250 milliliters of Erlenmeyer flasks of 70mlPM liquid nutrient medium 47.2.2 connect ring plant bacillus pumilis WO-8 (CCTCC M202044) lawn from the PM inclined-plane, at 30 ℃, 120-130r/min shaking table shaking culture 28 hours, it is thalline OD that bacterium liquid is transferred the OD value
600=0.45.
47.3 cultivate: the mixed culture (culture of 47.2.1) of nitrifiers such as the culture of plant bacillus pumilis WO-8 (culture of 47.2.2) and bacillus cereus NBB-135 mixes by 1: 1 (volume ratio), connects the 1ml mixed bacteria liquid and goes into the L with 0.075mol
-1Acetate be the simulation ammonium nitrogen wastewater (prescription with 1.4) of carbon source at 30 ℃, static cultivation 21 days.Survey ammonia-state nitrogen with the indophenol blue colorimetry, Griess reagent colorimetric method for determining nitrite nitrogen, the Kelvin method is measured total nitrogen.
47.4 result: mixed culture has removed 76.3% of the full nitrogen of system after cultivating 21 days, the ammonium nitrogen removal efficiency is 87.9%, and nitrite concentration only is 0.10mg N L
-1
Bacterium protects catalogue
| Bacterial strain number | |||
| WO-8 WR-2 NBB422 NBB324 NBB319 NBB295 NBB247 NBB204 NBB135 NBB112 NBB72 NBB46 NBB15 NBB58-3 NBB609 NBB19 NH-2 NH-14 S-12 S-27 | CCTCC M202044 CCTCC M202043 CGMCC NO.0554 CGMCC NO.0555 CGMCC NO.0556 CGMCC NO.0557 CGMCC NO.0558 CGMCC NO.0559 CGMCC NO.0560 CGMCC NO.0561 CGMCC NO.0562 CGMCC NO.0563 CGMCC NO.0564 CGMCC NO.0565 CGMCC NO.0566 CGMCC NO.0586 CCTCC M203101 CCTCC M203102 CCTCC M203103 CCTCC M203104 | Curtoacterium plantarum WO-8 Arthrobacter globiformis WR-2 Bacillus megaterium NBB-422 Bacillus firmus NBB-324 Bacillus brevis NBB-319 Bacillus circulans NBB-295 Bacilius coagulans NBB-247 Bacillus lentus NBB-204 Bacillus cereus NBB-135 Bacillus pumilus NBB-112 Bacilllus licheniformis NBB-072 Bacillus globisporus NBB-046 Bacillus sphaericus NBB-015 Bacillus badius NBB-58-3 Bacillus subtilis NBB-609 Bacillus mycoides NBB-019 Bacillus pseudofirmus NH-2 Paenibacillus campinasensis NH-14 Arthrobacter ramosus S-12 Arthrobacter sulfurous S-27 | The cured shape bacillus of plant bacillus pumilis Arthrobacter globiformis bacillus megaterium bacillus firmus bacillus brevis Bacillus circulans bacillus coagulans bacillus lentus bacillus pumilus bacillus licheniformis bacillus globisporus Bacillus sphaericus bacillus badius bacillus subtilis bacillus mycoides class bacillus firmus Campinas series bacillus branch arthrobacterium sulphur look arthrobacterium |
Claims (23)
1. treatment process that contains ammonium nitrogen wastewater, it is characterized in that in ammoniated wastewater, adding and have the organic acid of carboxyl or the organic carbon source of its esters, and add an amount of heterotrophic organism or heterotrophic organism group with nitrification activity, pH be 6 to 8 aerobic conditions and temperature 20-35 ℃ under static or little whipped state culturing bacterium 15-35 days down, directly remove the ammonium nitrogen in the ammoniated wastewater;
The heterotrophic organism that wherein has nitrification activity is to grow on the PM flat board or to separate, and the direct drop of griess reagent is positive in 1 minute, and when adding the good air culture of inorganic ammonium salt and support with the carbon source that physiological is produced alkali, the bacterial strain that has full nitrogen to wane.
2. according to the method described in the claim 1, it is characterized in that the heterotrophic organism that will have nitrification activity is containing the static culturing bacterium of ammonium nitrogen wastewater 28 days when 30 ℃ of temperature.
3. according to the method described in the claim 1, it is characterized in that containing have nitrification activity in the ammonium nitrogen wastewater heterotrophic organism 10
5-10
6Individual/milliliter.
4. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is bacillus megaterium (Bacillus megaterium) NBB-422, CGMCC NO.0554.
5. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is bacillus firmus (Bacillus firmus) NBB-324, CGMCC NO.0555.
6. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is bacillus brevis (Bacillus brevis) NBB-319, CGMCC NO.0556.
7. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is Bacillus circulans (Bacillus circulans) NBB-295, CGMCC NO.0557.
8. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is Bacillus coagulans (Bacillus coagulans) NBB-247, CGMCC NO.0558.
9. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is bacillus lentus (Bacillus lentus) NBB-204, CGMCC NO.0559.
10. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is cured shape genus bacillus (Bacillus cereus) NBB-135, CGMCC NO.0560.
11. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is bacillus pumilus (Bacillus pumilus) NBB-112, CGMCC NO.0561.
12. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is Bacillus licheniformis (Bacillus licheniformis) NBB-72, CGMCC NO.0562.
13. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is bacillus globisporus (Bacillus globisporus) NBB-46, CGMCC NO.0563.
14. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is Bacillus sphaericus (Bacillus sphaericus) NBB-15, CGMCC NO.0564.
15. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is bacillus badius (Bacillus badius) NBB-58-3, CGMCC NO.0565.
16. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is subtilis (Bacillus subtilis) NBB-609, CGMCC NO.0566.
17. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is bacillus mycoides (Bacillus mycoides) NBB-19, CGMCC NO.0586.
18. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is class bacillus firmus (Bacillus pseudofirmus) NH-2, CCTCC M203101.
19. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is Campinas series bacillus (Paenibacillus campinasensis) NH-14, CCTCC M203102.
20. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is branch Arthrobacter (Arthrobacter ramosus) S-12, CCTCC M203103.
21. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is sulphur look Arthrobacter (Arthrobacter sulfurous) S-27, CCTCC M 203104.
22. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is the mixed bacterial of the two arbitrarily in the claim 4 to 21.
23. according to the method described in the claim 1, the heterotrophic organism that it is characterized in that having nitrification activity is to contain to grow on the PM flat board, the direct drop of griess reagent is positive, and when adding the good air culture of inorganic ammonium salt and support, the heterotrophic organism with nitrification activity that full nitrogen wanes or the soil sample or the active sludge of mixed bacterial are arranged with the carbon source that physiological is produced alkali.
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| CN101767867B (en) * | 2010-01-06 | 2011-12-21 | 华东理工大学 | Activated sludge and processing method of alkaline sewage and high ammonia-nitrogen wastewater using the activated sludge |
| CN103937712B (en) * | 2014-04-04 | 2016-10-05 | 华南理工大学 | A kind of Bacillus licheniformis and application thereof |
| CN104164388B (en) * | 2014-07-09 | 2016-09-07 | 青岛蔚蓝生物集团有限公司 | One strain bacillus megaterium and the application in aquaculture thereof |
| CN104164392B (en) * | 2014-07-09 | 2016-11-09 | 青岛玛斯特生物技术有限公司 | One bacillus licheniformis and the application in aquaculture thereof |
| CN104152377B (en) * | 2014-07-31 | 2016-04-27 | 上海交通大学 | The aerobic denitrifying bacteria of heavy metal tolerance and application thereof |
| CN105441348B (en) * | 2014-09-11 | 2019-05-21 | 北京大北农科技集团股份有限公司 | One bacillus subtilis novel bacterial strain, probiotics and application |
| CN105621626B (en) * | 2014-11-05 | 2018-09-28 | 江苏元捷环境科技有限公司 | A kind of high-concentration chemical industry sewage composite bacteria agent and its application |
| CN109234214A (en) * | 2018-07-03 | 2019-01-18 | 宜兴市永洁环保设备有限公司 | A kind of production method of composite bacteria preparation and its application in sewage treatment |
| CN111004749B (en) * | 2019-12-26 | 2020-07-28 | 青岛尚德生物技术有限公司 | Salt-tolerant bacillus lentus GBW-HB1902 and application thereof |
| CN115353987B (en) * | 2022-01-28 | 2023-07-07 | 齐齐哈尔大学 | Bacillus subtilis strain SC strain for treating pig raising wastewater and application thereof |
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