CN102465105B - Nitrous acid-type denitrification bacteria strain and application thereof - Google Patents
Nitrous acid-type denitrification bacteria strain and application thereof Download PDFInfo
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Abstract
The invention relates to a bacteria strain for short-cut denitrification and an application thereof. The short-cut denitrification bacteria strain is arthrobacter creatinolyticus FDN-1 and is preserved in Ordinary Microorganism Center of China Committee for Culture Collection of Microorganisms on March 11, 2010, and the preservation number is CGMCC No. 3657. The bacteria strain can directly adopt nitrite nitrogen as a substrate to complete the short-cut denitrification process. When the arthrobacter creatinolyticus FDN-1 is used for processing ammonia-contained waste water, the technique is simple, high denitrification activity can be still obtained under the condition that low-concentration organic carbon source exists, and at the same time the high-concentration organic carbon source can be withstood. When the bacteria strain is in use, the system is rapid to start, and a vast application prospect can be realized in the denitrification treatment process of different types of waste water.
Description
Technical field
The invention belongs to environmental microorganism field, be specifically related to a kind of efficient nitrite type denitrification bacterial strain.Denitrogenation bacterial strain of the present invention can carry out taking nitrite nitrogen as nitrogenous source denitrification, is specially adapted to the denitrogenation in ammonia-containing water short-cut nitrification and denitrification process.
Background technology
Traditional biological denitrification process mainly comprises two processes of nitrification and denitrification, and nitrated is the prerequisite of denitrification process, and denitrification process could really remove the nitrogenous compound in waste water.Because nitrifier has strong aerobic, nitrifying process is aerobic; And denitrifying bacteria is generally facultative bacteria, under aerobic conditions, using oxygen as electron acceptor(EA), carry out aerobic repiration, while only having anaerobic state, just using nitrate or nitrite as electron acceptor(EA), obtain the energy of synthetic cell body.It is generally acknowledged under aerobic condition, nitric acid or nitrite reductase can not be expressed or oxygen molecule can produce and suppress these enzymes.As Pseudomonas nautica nitrate reductase in the time that dissolved oxygen is greater than respectively 4.05mg/L, 2.15mg/L, 0.25mg/L, nitrite reductase, reduction of nitrogen oxide enzyme complete deactivation.
Traditional denitrification process is that nitrification and denitrification is arranged in two structures and carries out classification denitrogenation mostly, exists sewage treatment process longer, and initial cost is high, need to supplement to nitrification tank the deficiencies such as basicity.So some denitrification microorganisms novel, better effects if, as allotrophic nitrobacteria, aerobic denitrifying bacteria, anaerobic ammonium oxidizing bacteria etc. are found in succession.
Isolate so far more than 60 kind of denitrifying bacteria, mainly be distributed in Pseudomonas (Pseudomonas), Alkaligenes (Alcaligeneas), general foster sulphur Coccus (Thiosphera pantotroph), Zoogloea (Zoogloeas) and bacillus (Bacillus) etc., great majority are facultative anaerobe.Lukow from the nitrated workshop section of percolate isolate belong to Zoogloeas mono-strain bacterium TL1 (Aerobic denitrification by a newly isolated heterotrophic bacterium strain TL1[J] .Biotechnology Letters, 199711 (19): 1157～1159), this bacterium can utilize oxygen and degrade nitrate simultaneously.Naokia sieves to obtain two strain denitrifying bacterias, be respectively Pseudomonas stutzeri TR2 and Pseudomonas sp.K50 (Aerobic Denitrifying Bacteria that Produce low Levels of Nitrous Oxide[J] .Applied and Environmental Microbiology, 2003,69 (6): 3152-3157).Zhang Guangya (Huaqiao University's journal, 2004,25 (1)) from soil, isolate the aerobic denitrifying bacteria HN of a strain taking SODIUMNITRATE as nitrogenous source, through being accredited as Rhodococcus sp., (the environmental science such as Wang Lin, 2009, (30) 1) taking nitrate nitrogen as electron acceptor(EA), be separated to a strain Bacillus foecalis alkaligenes (Acaligenes faecalis) taking Trisodium Citrate as carbon source, from fluvial deposit.(the ACTA Scientiae Circumstantiae such as Wang Xiaoyu, 2009, (29) 1) from active sludge, be separated to false unit cell Alcaligenes (Pseudomonas pseudoalcaligenes) and Rhodopseudomonas Mendoza bacterium (Pseudomonas mendocina).Above the research of denitrifying microorganism is mainly concentrated on to the denitrifying bacteria taking nitrate nitrogen as electron acceptor(EA).
(the microbiology circular such as Zhang little Ling, 2008,35 (10)) from sediment of pond, be separated to a strain aerobic denitrifying bacteria H2, belong to bacillus through preliminary evaluation, this bacterial strain is 0.885mg/ (Ld) to the highest degradation rate of nitrite; Li Bing etc. (aquatic ecological magazine, 2009,2 (3)) screen genus bacillus D5 from soil, and the most degradation speed of this bacterial strain is 2.22mg/ (Lh).(the Research of Environmental Sciences such as healthy, 2010,23 (1)) from prawn culturing pond, screen a bacillus coagulans (Bacillus coagulans) YX-6, this bacterial strain can be down to 0 by nitrite nitrogen by 10mg/L in 14h.The degrading nitrite nitrogen bacterial strain being more than separated to all belongs to bacillus (Bacillus), these bacterial strains can carry out taking nitrite nitrogen as electron acceptor(EA) denitrification denitrogenation, and degradation rate can reach 10mg/ (Lh), and the nitrite nitrogen concentration of the waste water of processing has also reached 100mg/L.
Arthrobacter, as a kind of environmental microorganism, plays an important role in the practical application of Industrial Wastewater Treatment.Zhu Xikun etc. (agro-environment science journal, 2009,28 (3)) are separated to the Arthrobacter AD26 bacterial strain of efficient degradation herbicide atrazine.Jiang Yongrong etc. (biotechnology, 2007,17 (1)) screening and separating goes out the genus arthrobacter LZP08X bacterial strain of efficient degradation phenol.Separate elegant equality (ACTA Scientiae Circumstantiae, 2006,26 (10)) be separated to the bacterium L4 bacterial strain that a strain can the growth taking parathion-methyl and intermediates p-NP thereof as sole carbon source, through 16SrRNA gene sequencing, this bacterial strain is accredited as genus arthrobacter (Arthrobactersp.), this bacterial strain in 5h to 50mgL
-1degraded parathion-methyl and the degradation rate of p-NP be respectively 85% and 98%.(refining of petroleum and the chemical industry such as Tian Zhiguo, 2007,38 (10)) from the pedotheque of oil pollution, filtering out a strain can be the desulfurization Arthrobacter bacterial strain T1 of 2-xenol (2-HBP) by dibenzothiophene (DBT) metabolism.The waste water of above-mentioned genus arthrobacter bacterial strain processing does not relate to degraded nitrite nitrogen.CN200910021020.7 discloses in a kind of preparation method of the water quality modifying microecological preparation that falls ammonia nitrogen and nitrous acid nitrogen and has related to a strain Arthrobacter CGMCC NO.1282; but the probiotics of this invention belongs to aquaculture technology and ecological environmental protection technical field, there is not yet the report of Arthrobacter for the treatment of trade effluent Nitrite Nitrogen.
Therefore obtain efficient degrading nitrite nitrogen bacterial strain, and make it really be applied in actual waste water treating processes, become the dominant population in reaction system and bring into play denitrification and remain the direction of research.
Summary of the invention
The object of the present invention is to provide a kind of have spontaneous growth advantage, the direct new bacterial strain Arthrobacter using nitrite as electron acceptor(EA), Arthrobacter activity of nitrous acid reductase provided by the invention is high, under aerobic and oxygen free condition, all can express.Finding that there is of new bacterial strain is beneficial to promotion short-cut nitrification and denitrification technique and the application of synchronous nitration and denitrification technique in actual sewage treating processes.
The invention provides a kind of new bacterial strain for short-cut denitrification denitrogenation: Arthrobacter (Arthrobacter creatinolyticus) FDN-1, be preserved in that " China Committee for Culture Collection of Microorganisms's common micro-organisms " center ", its preserving number is CGMCC NO.3657 on March 11st, 2010.
Arthrobacter FDN-1 Main Morphology provided by the invention is characterized as: colony colour is yellow, bacterial strain is individual is bar-shaped, without gemma, can move.
The physiological and biochemical property of described Arthrobacter FDN-1 shows as: gramstaining is positive, the catalase positive, and oxidase negative, can utilize several kinds of carbon source.
Arthrobacter FDN-1 of the present invention can carry out taking nitrite nitrogen as nitrogenous source denitrification denitrogenation, and the expression of nitrite reductase is not limited by dissolved oxygen conditions, all can carry out denitrification under aerobic and anaerobic environment.
Denitrifying bacteria 16SrRNA gene order provided by the invention is shown in sequence table.
Arthrobacter FDN-1 provided by the invention, is mainly used in short-cut denitrification denitrogenation, and this new strain growth speed is fast, cell yield is high, requires C/N than low, no matter is that fresh medium or the nutrient solution of very low temperature preservation all have higher denitrification activity.The condition of this bacterial strain short-cut denitrification denitrogenation is: 15 DEG C～35 DEG C of temperature; PH6.5～10.0; Carbon-nitrogen mass ratio 2: 1～5: 1.
Arthrobacter of the present invention is processing when nitrogenous effluent, can enduring high-concentration nitrite nitrogen, reach as high as 800mg/L, and degradation rate can be greater than 20mg/ (Lh).
Arthrobacter FDN-1 of the present invention can utilize lower concentration carbon source to carry out denitrification, saves denitrification needed carbon source, organic carbon source that simultaneously can enduring high-concentration.
Arthrobacter of the present invention is in the time processing nitrogenous effluent, and technique is simple, still has higher denitrification activity in the situation that lower concentration organic carbon source exists, simultaneously can enduring high-concentration organic carbon source.After coming into operation, system starts fast, in all kinds of denitrogenation of waste water treating processess, there is boundless application prospect, be particularly suitable for, in short-cut nitrification and denitrification or synchronous nitration and denitrification process, in the situation that denitrification effect is undesirable, can be used as bioremediation agents and adding.Can also prepare denitrogenation microbial inoculum.
biomaterial preservation explanation
Arthrobacter provided by the invention (Arthrobacter creatinolyticus) FDN-1 bacterial strain, is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center (CGMCC); Address: Datun Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica; Deposit number: CGMCC NO.3657; Preservation date: on March 11st, 2010.
Brief description of the drawings
Fig. 1 is the growth conditions of Arthrobacter creatinolyticus FDN-1 on flat board.
Fig. 2 is the denitrification effect of Arthrobacter creatinolyticus FDN-1.
Embodiment
Arthrobacter FDN-1 of the present invention screens and obtains from Fushun No.3 Petroleum Factory's sludge sewage, can carry out denitrification denitrogenation taking nitrite nitrogen as nitrogenous source, and its physiological and biochemical test the results are shown in Table 1.
The physiological and biochemical test result of table 1 Arthrobacter FDN-1
Through " China Committee for Culture Collection of Microorganisms's common micro-organisms " center " qualification, this bacterial strain belongs to Arthrobacter (Arthrobacter creatinolyticus), called after FDN-1.Fig. 1 is the growth conditions of this Arthrobacter on agar plate, and this bacterium colony is yellow, emulsus, colour purity.
Enrichment culture and the separation and purification of embodiment 1 bacterium
1. the enrichment culture of denitrifying bacterium
Get the active sludge in nitrification tank, utilize the substratum that contains nitrite under low DO condition, to carry out enrichment culture certain hour, obtain denitrifying activated sludge.In enrichment process, supplement fresh medium in good time.
Enrichment culture liquid formula: NaNO
2, methyl alcohol, also contains and comprises Fe
2+, Mg
2+, K
+, Ca
2+four kinds of metallic cations.Culture condition is: 30 DEG C of temperature; PH is that 8.2, DO (dissolved oxygen) is 0.2-1.0mgL
-1.
Mole allocation ratio of four kinds of metallic cations is 1: (4-8): (5-15): (1-5).In culturing process, add the growth stimulant that metallic cation total concn is 0.15mol/L every day, and additional amount is 0.1% of the interior volume of material of reactor.
The screening of 2 denitrifying bacteriums
Draw the above-mentioned enrichment culture liquid of 1ml, resuspended and as far as possible particle is broken up with distilled water, get after bacteria suspension dilutes different multiples and be coated with the beef extract-peptone flat board that has permeated denitrification nutrient solution, the screening of denitrifying bacterium is carried out in 30 DEG C of cultivations, the upgrowth situation of routine observation thalline in culturing process, repeats aforesaid operations until obtain single bacterium colony.Individual plant purifying is carried out in single bacterium colony line on picking flat board, transfers twice, obtains the bacterial strain FDN-1 of purifying.
Separation and Culture based formulas: extractum carnis: 5g/L, peptone: 10g/L, NaNO
2: 1g/L, flat board adds 2% agar.
Embodiment 2 bacterial strain denitrification ability of the present invention is investigated
First from flat board, picking list bacterium colony FDN-1 is inoculated in beef-protein medium 30 DEG C, 120r/min shaking table and cultivates cell concentration maximum after 42h, finishes to cultivate.After bacteria suspension is centrifugal, proceed in Sodium Nitrite-methyl alcohol nutrient solution takes the mode of batch feed supplement to carry out denitrification ability investigation under similarity condition.Table 1 is to add nitrite nitrogen and the COD concentration in reaction system after material, and denitrification effect is shown in Fig. 2.
Table 1 is added material situation
From Fig. 2 nitrite nitrogen concentration over time trend can see, front 3 batches experienced respectively 9 days, 5 days and 2 day time nitrite nitrogen remove completely, after this also improve gradually in the process of concentration in batch feed supplement, nitrite nitrogen all removes completely after 24h, the high energy of handled nitrite nitrogen concentration reaches 628mg/L, and nitrite nitrogen is removed rate 26.17mg/ (Lh).This bacterial strain has higher denitrifying capacity as can be seen here, also improves gradually within the specific limits in the process of concentration in batch feed supplement, and the denitrification ability of bacterial strain can strengthen gradually.
The application of embodiment 3 bacterial strain of the present invention in nitrogenous fertilizer denitrogenation of waste water process
Get the bacteria suspension that a certain amount of embodiment 2 obtains and inoculate according to 15% inoculum size, be 7.7 at 28 DEG C of temperature, pH, DO (dissolved oxygen) is 0.2～0.9mgL
-1under condition, process the short distance nitration reaction water outlet of the ammonia-containing water producing in nitrogen fertilizer production process, in this water: NO
2 --N average out to 400mg/L, COD average out to 800mg/L.Water outlet NO after treatment
2 --N and COD concentration are respectively 0.2mg/L～5.7mg/L and 38mg/L～49mg/L, NO
2 --N and COD clearance are greater than respectively 99% and 93%.
The application of embodiment 4 bacterial strain of the present invention in containing wastewater from catalyst denitrification process
Get the bacteria suspension that a certain amount of embodiment 2 obtains and inoculate according to 15% inoculum size, be 8.0 at 30 DEG C of temperature, pH, DO (dissolved oxygen) is 0.3mgL
-1under condition, the ammonia-containing water short distance nitration water outlet producing in Catalyst Production process is carried out to denitrification processing, in this water, COD is 800mg/L～1200mg/L, NO
2 --N is 500mg/L～780mg/L, NO
3 --N is 10mg/L.Determination of Total Nitrogen in Waste Water clearance reaches more than 98% after treatment.
Claims (7)
1. the bacterial strain for short-cut denitrification denitrogenation: Arthrobacter (Arthrobacter creatinolyticus) FDN-1, be preserved in that " China Committee for Culture Collection of Microorganisms's common micro-organisms " center ", its preserving number is CGMCC NO.3657 on March 11st, 2010.
2. according to bacterial strain claimed in claim 1, it is characterized in that the morphological specificity of Arthrobacter FDN-1 is: colony colour is: yellow, bacterial strain is individual is bar-shaped, without gemma, can move.
3. according to bacterial strain claimed in claim 1, it is characterized in that the physiological and biochemical property of Arthrobacter FDN-1 shows as: gramstaining is positive, the catalase positive, oxidase negative, can utilize several kinds of carbon source.
4. according to bacterial strain claimed in claim 1, it is characterized in that Arthrobacter FDN-1 can carry out taking nitrite nitrogen as nitrogenous source denitrification denitrogenation.
5. according to bacterial strain claimed in claim 1, it is characterized in that Arthrobacter FDN-1 can tolerate nitrite nitrogen and reach 800mg/L.
6. described in the arbitrary claim of claim 1～5, Arthrobacter FDN-1 is applied to short-cut denitrification denitrogenation.
7. according to application claimed in claim 6, it is characterized in that Arthrobacter FDN-1 is 15 DEG C～35 DEG C of temperature for the operational condition of short-cut denitrification denitrogenation; PH6.5～10.0; Carbon-nitrogen mass ratio 2: 1～5: 1.
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| CN104862260B (en) * | 2015-06-10 | 2018-09-25 | 国家海洋局第三海洋研究所 | One plant of arthrobacterium and its application with aerobic denitrification ability |
| CN108118008A (en) * | 2016-11-29 | 2018-06-05 | 中国石油化工股份有限公司 | Microbial bacterial agent of Efficient salt-tolerant and its preparation method and application |
| CN108118022B (en) | 2016-11-29 | 2020-09-11 | 中国石油化工股份有限公司 | Microbial culture promoter for completing denitrification process and application thereof |
| CN108285880B (en) * | 2018-01-04 | 2021-03-30 | 广州大学 | Bacillus with electrogenesis characteristic and denitrification activity and application thereof |
| CN112029688B (en) * | 2020-09-24 | 2022-04-26 | 新疆水处理工程技术研究中心有限公司 | Nitrifying bacteria agent for industrial wastewater and preparation method thereof |
| CN113005062B (en) * | 2021-03-24 | 2022-05-13 | 中国科学院成都生物研究所 | Facultative ammonia oxidizing bacteria and application thereof |
| CN116240138A (en) * | 2021-08-18 | 2023-06-09 | 上海交通大学 | Arthrobacter and application thereof in degradation of aristolochic acid |
| LU503114B1 (en) | 2022-11-28 | 2024-05-30 | Biosolum B V | Method for the preparation of a ammonia and nitrous acid tolerant bacterial consortium and method for removal of high ammonium, urea, nitrite, and/or nitrate concentrations in watery streams. |
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| CN101875909A (en) * | 2010-01-26 | 2010-11-03 | 中国科学院成都生物研究所 | Heterotrophic nitrification aerobic denitrifying bacteria and culture method and application thereof |
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| RU2177034C1 (en) * | 2001-04-03 | 2001-12-20 | Закрытое акционерное общество "Биоамид" | Strain of bacterium alcaligenes denitrificans as nitrilase producer |
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| CN101130760A (en) * | 2006-10-13 | 2008-02-27 | 北京工商大学 | Grass spirillum with aerobic denitrification and method for processing wastewater |
| CN101875909A (en) * | 2010-01-26 | 2010-11-03 | 中国科学院成都生物研究所 | Heterotrophic nitrification aerobic denitrifying bacteria and culture method and application thereof |
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| EP3480295A1 (en) * | 2017-11-07 | 2019-05-08 | BioSolum B.V. | Micro-organism tolerant for inorganic nitrogen compounds |
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