CN104480045A - Efficient aerobic denitrifying strain having heterotrophic nitrification function and application of strain - Google Patents

Efficient aerobic denitrifying strain having heterotrophic nitrification function and application of strain Download PDF

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Publication number
CN104480045A
CN104480045A CN201410775663.0A CN201410775663A CN104480045A CN 104480045 A CN104480045 A CN 104480045A CN 201410775663 A CN201410775663 A CN 201410775663A CN 104480045 A CN104480045 A CN 104480045A
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strain
yxhf
heterotrophic nitrification
bacterial strain
nitrification function
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CN201410775663.0A
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CN104480045B (en
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骆灵喜
刘欢
李旭宁
王波
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PKU-HKUST SHENZHEN-HONGKONG INSTITUTION
Shenzhen Ier Environmental Protection Engineering Technology Co Ltd
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PKU-HKUST SHENZHEN-HONGKONG INSTITUTION
Shenzhen Ier Environmental Protection Engineering Technology Co Ltd
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    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12RINDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00Microorganisms ; Processes using microorganisms
    • C12R2001/01Bacteria or Actinomycetales ; using bacteria or Actinomycetales
    • C12R2001/38Pseudomonas
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20Bacteria; Culture media therefor
    • C12N1/205Bacterial isolates
    • CCHEMISTRY; METALLURGY
    • C02TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02FTREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F3/00Biological treatment of water, waste water, or sewage
    • C02F3/02Aerobic processes
    • CCHEMISTRY; METALLURGY
    • C02TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02FTREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F3/00Biological treatment of water, waste water, or sewage
    • C02F3/34Biological treatment of water, waste water, or sewage characterised by the microorganisms used
    • YGENERAL 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
    • Y02TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02WCLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
    • Y02W10/00Technologies for wastewater treatment
    • Y02W10/10Biological treatment of water, waste water, or sewage

Abstract

The invention discloses an efficient aerobic denitrifying strain (pseudomonas otitidi) YXHF-2 having a heterotrophic nitrification function, wherein the strain is preserved in China General Microbiological Culture Collection Center with the preservation number of CGMCC No. 10047. Furthermore, the invention also discloses application of the strain YXHF-2 in sewage treatment. The efficient aerobic denitrifying strain YXHF-2 having the heterotrophic nitrification function disclosed by the invention can be used for effectively reducing the contents of ammonia nitrogen, total nitrogen and chemical oxygen demand (COD) in sewage, and the strain has relatively high application value in the treatment of domestic sewage and other various polluted water bodies.

Description

One strain has high-efficiency aerobic denitrification bacterial strain and the application thereof of heterotrophic nitrification function concurrently
Technical field
The invention belongs to technical field of environmental microorganism, relate to a strain particularly and have the high-efficiency aerobic denitrification bacterial strain of heterotrophic nitrification function and the application in waste water control thereof concurrently.
Background technology
Nitrogen causes one of body eutrophication and environmental pollution very important pollution factor, bio-denitrification technology is current most widely used polluted by nitrogen control method, traditional theory thinks that biological denitrificaion must be realized by nitrification and denitrification 2 self-contained process, nitrification and denitrification can not occur simultaneously, nitration reaction is carried out under the condition of aerobic, and anti-nitration reaction needs to carry out under the condition of strict anaerobism or anoxic, traditional two-part biological denitrification process ubiquity initial cost and working cost high, the deficiencies such as energy consumption is large, and operation control condition is complicated.
Traditional biological denitrificaion theory has been broken in the discovery of aerobic denitrification and heterotrophic nitrification, simultaneous nitrification and denitrification is made to become possibility, nitrification and denitrification not only can be made to complete in same reactor, accelerate whole reaction process, shorten hydraulic detention time, reduce reactor volume, reduce running cost, the water quality scope of carrying out denitrogenation processing can also be expanded.
By being separated the aerobic denitrifying bacteria having heterotrophic nitrification function concurrently, furtheing investigate the application in the growth characteristics of this bacterium, Denitrification Characteristics and sewage water denitrification thereof, having important theory value and practical significance to the denitrogenation processing efficiency and economy improving sewage.
Summary of the invention
An object of the present invention is to provide the aerobic denitrifying bacteria that a strain has heterotrophic nitrification function concurrently.
Two of object of the present invention is to provide the above-mentioned application of aerobic denitrifying bacteria in waste water control having heterotrophic nitrification function concurrently.
For realizing above object, the different aerobic denitrifying bacteria YXHF-2 having heterotrophic nitrification function concurrently disclosed by the invention, through being accredited as pseudomonas otitidi(NCBI accession number: KF159806.1), belong to Rhodopseudomonas ( psedomonassp.) disclosed by the invention have heterotrophic nitrification function concurrently aerobic denitrifying bacteria ( pseudomonas otitidi) YXHF-2 is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center on November 21st, 2014, deposit number is CGMCC No:10047.
The invention also discloses the above-mentioned high-efficiency aerobic denitrification bacterial strain YXHF-2 having heterotrophic nitrification function concurrently to apply in waste water control.
The process form that the above-mentioned high-efficiency aerobic denitrification bacterial strain YXHF-2 having heterotrophic nitrification function concurrently applies is membrane bioreactor.
The described high-efficiency aerobic denitrification bacterial strain YXHF-2 dosage having heterotrophic nitrification function concurrently is the 1-4% of membrane bioreactor volume.
The high-efficiency aerobic denitrification bacterial strain YXHF-2 having heterotrophic nitrification function concurrently of the present invention effectively can reduce the content of ammonia nitrogen, total nitrogen and COD in sewage, in sanitary sewage and other various polluted-waters are administered, have higher using value.Apply in membrane bioreactor, effectively can reduce the content of ammonia nitrogen, total nitrogen and the COD in membrane bioreactor sanitary sewage, exploitation denitrification microorganism preparation or sewage-treating agent are had broad application prospects.
preservation information
Provided by the invention have heterotrophic nitrification function concurrently high-efficiency aerobic denitrification bacterial strain ( pseudomonas otitidi) YXHF-2, be preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center (CGMCC), address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, deposit number is CGMCC No:10047, and preservation date is on November 21st, 2014.
Accompanying drawing explanation
Fig. 1 is membrane bioreactor structural representation in embodiment;
Fig. 2 is bacterial strain YXHF-2 to the removal effect figure of nitrate and total nitrogen;
Fig. 3 is the removal effect figure of bacterial strain YXHF-2 to ammonia nitrogen;
Fig. 4 is the changing conditions that bacterial strain YXHF-2 adds pollutants removal rate to membrane bioreactor.
Embodiment
Below in conjunction with specific embodiment, the present invention is described in detail.
Experimental technique in following embodiment, if no special instructions, is ordinary method.Test materials used in following embodiment, if no special instructions, is and purchases available from routine biochemistry reagent shop.
1, the acquisition of bacterial strain and Performance Detection
Enrichment culture: the sample of strains separation is from the mud suspension of certain membrane bioreactor in Guanlan town, Shenzhen, and this device adopts " sloping plate deposition+membrane bioreactor " combination process, and treatment capacity is 25m 3/ d, the residence time is 4h, gas-water ratio 30:1.Membrane bioreactor mud suspension sample 5% is seeded to enriched liquid substratum (KNO by volume 32g, KH 2pO 41g, K 2hPO 41g, MgSO 47H 2o 0.2g, glucose 5g, distilled water 1000mL), at 30 DEG C of rotating speeds be 150 r/min constant-temperature table on shaking culture 48 hours.
Wherein, certain membrane bioreactor structure is shown in accompanying drawing 1.It is the film bioreactor device of a strain aerobic synchronous nitration and denitrification, adopt " sloping plate deposition+membrane bioreactor " combination process, primarily of settling region (11), film biological treatment zone (21) and power control district (31) three part composition, main equipment material is steel construction, all makes the degree of depth anticorrosion.Settling region (11) arranges one or more swash plate (12), mainly carries out precipitation pre-treatment to oarse-grained suspended substance.Film biological treatment zone (21) mainly comprises membrane module (23), micropore aeration pipe (22) etc., wherein membrane module (23) is made up of membrane element (231) and header (232), described membrane module (23) is made up of one or more membrane elements (231) and header (232), and described header (232) is connected with membrane element (231).Wherein membrane element (231) is PVDF material flat sheet membrane, and membrane pore size is 0.2 μm, membrane flux Q=400 ~ 550L/ sheet d), header (232) is UPVC material; Micropore aeration pipe (22) is distributed in the bottom of film biological treatment zone (21) and connects on 1/3 wall of bottom, and aeration tube aperture is 6mm, and axially spaced-apart is 60 ~ 80mm, and radial aperture angle is 140 degree.Power and control region are primarily of compositions such as water pump (32), air pump (33) and PLC two-position controllers (34), and to the hydraulic detention time of film bioreactor device, the parameter such as gas-water ratio and water-out manners controls automatically.
Primary dcreening operation: get the enrichment medium stepwise dilution 10 respectively after concussion cultivation 3, 10 4with 10 5doubly, BTB solid selection medium (KNO is coated 31g, KH 2pO4 1g, CaCl 27H 2o 0.2g, FeCl 26H 2o 0.5g, glucose 5g, volume ratio is the BTB ethanolic soln 1mL of 1%, agar 20g; Distilled water 1000mL, regulates pH to be 7.0 ~ 7.3), be placed in 30 DEG C of constant incubators and cultivate 3 days.Picking makes BTB substratum present blue single bacterium colony purifying of ruling on rich solids substratum and obtains primary dcreening operation bacterial strains three times.
Multiple sieve: be seeded in enriched liquid substratum respectively by above-mentioned primary dcreening operation bacterial strain, cultivates 2 days under 150 r/min, 30 ° of C conditions.Respectively nutrient solution 5% is added to denitrification detection substratum (glucose 0.5 g, sodium acetate 0.35g, KH by volume 2pO 40.105g, KNO 30.3 g, distilled water 1000mL), cultivate under 150 r/min, 30 ° of C conditions, timing detects the concentration of nitrate and total nitrogen, acquisition has the good bacterial strain of denitrification, and by its called after bacterial strain YXHF-2, this bacterial strain YXHF-2 is shown in Fig. 2 to nitrate and total nitrogen removal effect; Bacterial strain YXHF-2 after screening adds to nitrification detection substratum (NH4Cl 0.344 g/L again, glucose 1 g/L, MgSO47H2O 0.2g/L, K2HPO4 0.2g/L, NaCl 0.12g/L, MnSO44H2O 0.01g/L, FeSO4 0.01g/L, pH7.0 ~ 7.2), the change in concentration of periodic detection ammonia nitrogen, as shown in Figure 3.
As seen from Figure 2, the removal effect of bacterial strain YXHF-2 to nitrate and total nitrogen is all higher, and 24h can remove nitrate and the total nitrogen of 91.1% and 73.6% respectively; As seen from Figure 3, bacterial strain YXHF-2 also has good removal effect to ammonia nitrogen, and 24h can remove the ammonia nitrogen of about 87.2%.Visible aerobic denitrifying bacteria YXHF-2 all has good removal effect to nitrate, total nitrogen, ammonia nitrogen, can be applied in sewage disposal.
2, the molecular biology identification of bacterial strain
" MiniBEST Bacterial Genomic DNA Extraction Kit " test kit of precious biotechnology company limited is adopted to extract the genomic dna of bacterial strain YXHF-2, and as template amplification 16SrDNA, primer: 16S rDNA-27f:5 '-AGAGTTTGATCCTGGCTCAG-3 ', 1492 r (5 '-GGTTACCTTGTTACGACTT-3 '.PCR reaction system (50 μ L): template DNA 50-100ng, PCR Premix 25 μ L, each 0.5 μ L of upstream and downstream primer, adding ddH2O to reaction system is 50 μ L.PCR program: 94 DEG C of 10min, 94 DEG C of 1min, 50 DEG C of 1min, 72 DEG C of 2min, 72 DEG C of 5min; 72 DEG C of 10min; 30 circulations.Purifying and the order-checking of PCR primer are completed by Hua Da genome company, and sequencing sequence logs in GenBank and carries out homology sequence compare of analysis.
The sequence length of bacterial strain YXHF-2 is 1497bp, sees gene order table.Through comparison, this sequence and ncbi database pseudomonas otitidis(NCBI accession number: KF159806.1) homology reaches 99%, belong to Rhodopseudomonas ( psedomonassp.).Its called after is had concurrently heterotrophic nitrification function high-efficiency aerobic denitrification bacterial strain ( pseudomonas otitidi) YXHF-2.
3, the effect of high-efficiency aerobic denitrification bacterial strain YXHF-2 in membrane bioreactor of heterotrophic nitrification function is had concurrently
Adopt above-mentioned enriched liquid substratum enlarged culturing YXHF-2 in constant-temperature table, under shaking speed 150 r/min, 30 ° of C conditions, cultivate 3d.Nutrient solution is added centrifuge tube and with the centrifugal 10min of 6000r/min, abandons supernatant and use the resuspended thalline of physiological saline, suspension OD 600be about 1.6.
Add in membrane bioreactor by bacteria suspension according to 2% volume ratio, this membrane bioreactor is primarily of High-rate sedimentation district and film biological treatment zone composition, and intake as the former water of sanitary sewage, treatment scale is 1m 3/ d, hydraulic detention time is 6h, and aeration mode is continuous aeration, and gas-water ratio is 30:1, and water temperature is 25 ~ 30 DEG C.Monitor the COD, ammonia nitrogen and the nitrogen removal rate changing conditions that add membrane bioreactor before and after bacterial strain YXHF-2 suspension and see Fig. 4.
As seen from Figure 4, YXHF-2 bacteria suspension adds to membrane bioreactor, and the clearance of COD, ammonia nitrogen and total nitrogen all has raising in various degree, and wherein the clearance increase rate of total nitrogen is maximum, improves 34.2% when the nitrogen removal rate ratio adding latter 10 days adds; Add when the ammonia nitrogen removal frank ratio after latter 10 days adds and improve 19.0%, COD clearance raising 10.2%.
As can be seen from the above-described embodiment, bacterial strain YXHF-2 not only possesses excellent aerobic denitrification capability, also has the effect of heterotrophic nitrification and removal COD simultaneously, in sanitary sewage and other various polluted-waters are administered, have higher using value.Apply in membrane bioreactor, effectively can reduce the content of ammonia nitrogen, nitric nitrogen and the COD in membrane bioreactor sanitary sewage, exploitation denitrification microorganism preparation or sewage-treating agent are had broad application prospects.
In sum, but the present invention is not limited to above-mentioned embodiment, and persons skilled in the art are in the technical scope disclosed by the present invention, and the change that can expect easily, all within protection scope of the present invention.
SEQUENCE LISTING
 
<110> Shengang CEEUSRO Environmental Protection Engineering Technology Co., Ltd., Shenzhen
Shenzhen-Hong Kong Production & Research Base
 
<120> mono-kind has high-efficiency aerobic denitrification bacterial strain and the application thereof of heterotrophic nitrification function concurrently
 
<130>
 
<160> 1
 
<170> PatentIn version 3.3
 
<210> 1
<211> 1497
<212> DNA
<213> Pseudomonas otitidi
 
<400> 1
agagtttgat cctggctcag attgaacgct ggcggcaggc ctaacacatg caagtcgagc 60
 
ggatgagtgg agcttgctcc atgattcagc ggcggacggg tgagtaatgc ctaggaatct 120
 
gcctggtagt gggggataac gtttcgaaag gaacgctaat accgcatacg tcctacggga 180
 
gaaagtgggg gatcttcgga cctcacgcta tcagatgagc ctaggtcgga ttagctagtt 240
 
ggtggggtaa tggcccacca aggcgacgat ccgtaactgg tctgagagga tgatcagtca 300
 
cactggaact gagacacggt ccagactcct acgggaggca gcagtgggga atattggaca 360
 
atgggcgaaa gcctgatcca gccatgccgc gtgtgtgaag aaggtcttcg gattgtaaag 420
 
cactttaagt tgggaggaag ggcagtaagt taataccttg ctgttttgac gttaccaaca 480
 
gaataagcac cggctaactt cgtgccagca gccgcggtaa tacgaagggt gcaagcgtta 540
 
atcggaatta ctgggcgtaa agcgcgcgta ggtggttcag caagttggat gtgaaagccc 600
 
cgggctcaac ctgggaattg catccaaaac tactgggcta gagtacggta gagggtggtg 660
 
gaatttcctg tgtagcggtg aaatgcgtag atataggaag gaacaccagt ggcgaaggcg 720
 
accacctgga ctgatactga cactgaggtg cgaaagcgtg gggagcaaac aggattagat 780
 
accctggtag tccacgccgt aaacgatgtc gactagccgt tgggatcctt gagatcttag 840
 
tggcgcagct aacgcgataa gtcgaccgcc tggggagtac ggccgcaagg ttaaaactca 900
 
aatgaattga cgggggcccg cacaagcggt ggagcatgtg gtttaattcg aagcaacgcg 960
 
aagaacctta cctggccttg acatgcagag aactttccag agatggattg gtgccttcgg 1020
 
gaactctgac acaggtgctg catggctgtc gtcagctcgt gtcgtgagat gttgggttaa 1080
 
gtcccgtaac gagcgcaacc cttgtcctta gttaccagca cctcgggtgg gcactctaag 1140
 
gagactgccg gtgacaaacc ggaggaaggt ggggatgacg tcaagtcatc atggccctta 1200
 
cggccagggc tacacacgtg ctacaatggt cggtacaaag ggttgccaag ccgcgaggtg 1260
 
gagctaatcc cataaaaccg atcgtagtcc ggatcgcagt ctggaactcg actgcgtgaa 1320
 
gtcggaatcg ctagtaatcg tgaatcagaa tgtcacggtg aatacgttcc cgggccttgt 1380
 
acacaccgcc cgtcacacca tgggagaggg ttgctccaga agtagctagt ctaaccgcaa 1440
 
ggaggacggt taccacggag tgattcatga ctggggtgaa gtcgtaacaa ggtaacc 1497

Claims (4)

1. a strain have concurrently heterotrophic nitrification function high-efficiency aerobic denitrification bacterial strain ( pseudomonas otitidi) YXHF-2, be preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, deposit number is CGMCC No:10047.
2. the high-efficiency aerobic denitrification bacterial strain YXHF-2 having heterotrophic nitrification function as claimed in claim 1 concurrently applies in waste water control.
3. apply as claimed in claim 2, it is characterized in that: described in have concurrently heterotrophic nitrification function high-efficiency aerobic denitrification bacterial strain YXHF-2 apply process form be membrane bioreactor.
4. apply as claimed in claim 3, it is characterized in that: described in have heterotrophic nitrification function concurrently high-efficiency aerobic denitrification bacterial strain YXHF-2 dosage be the 1-4% of membrane bioreactor volume.
CN201410775663.0A 2014-12-17 2014-12-17 One plant of otitis pseudomonas strains and its application Active CN104480045B (en)

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN109055252A (en) * 2017-11-13 2018-12-21 鼎正新兴生物技术(天津)有限公司 A kind of heterotrophic nitrification-aerobic denitrification complex microorganism preparations and preparation method thereof
CN110217895A (en) * 2019-05-31 2019-09-10 中国科学院微生物研究所 A kind of complex micro organism fungicide and its application for water environment treatment
CN110697883A (en) * 2018-07-09 2020-01-17 中国科学院生态环境研究中心 Method for strengthening denitrification function of membrane bioreactor and slowing down membrane pollution

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102061276B (en) * 2010-11-15 2012-06-27 北京大学 Pseudomonas sp. strain for biological denitrification under low temperature and application thereof
CN103497909B (en) * 2013-08-27 2014-12-24 中国科学院成都生物研究所 Bacterial strain JY9 and use thereof
CN103525730B (en) * 2013-10-14 2015-12-02 青岛蔚蓝生物集团有限公司 A kind of otitis pseudomonas strains and application thereof
CN103849588B (en) * 2014-03-05 2016-04-06 北京市环境保护科学研究院 One strain aerobic denitrifying bacteria and the application in sewage water denitrification thereof
CN104046582A (en) * 2014-06-04 2014-09-17 辽宁大学 Medium rich in Pseudomonas otitidis

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN109055252A (en) * 2017-11-13 2018-12-21 鼎正新兴生物技术(天津)有限公司 A kind of heterotrophic nitrification-aerobic denitrification complex microorganism preparations and preparation method thereof
CN110697883A (en) * 2018-07-09 2020-01-17 中国科学院生态环境研究中心 Method for strengthening denitrification function of membrane bioreactor and slowing down membrane pollution
CN110697883B (en) * 2018-07-09 2020-12-15 中国科学院生态环境研究中心 Method for strengthening denitrification function of membrane bioreactor and slowing down membrane pollution
CN110217895A (en) * 2019-05-31 2019-09-10 中国科学院微生物研究所 A kind of complex micro organism fungicide and its application for water environment treatment

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