CN114369544B - Denitrifying bacteria and application thereof in nitrogen-containing sewage treatment - Google Patents
Denitrifying bacteria and application thereof in nitrogen-containing sewage treatment Download PDFInfo
- Publication number
- CN114369544B CN114369544B CN202111561362.4A CN202111561362A CN114369544B CN 114369544 B CN114369544 B CN 114369544B CN 202111561362 A CN202111561362 A CN 202111561362A CN 114369544 B CN114369544 B CN 114369544B
- Authority
- CN
- China
- Prior art keywords
- nitrite
- nitrogen
- nitrate
- strain
- hydroxylamine
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/30—Aerobic and anaerobic processes
- C02F3/302—Nitrification and denitrification treatment
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, 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
- C12N1/02—Separating microorganisms from their culture media
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, 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
- C12N1/20—Bacteria; Culture media therefor
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F2101/00—Nature of the contaminant
- C02F2101/10—Inorganic compounds
- C02F2101/16—Nitrogen compounds, e.g. ammonia
-
- 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
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Biotechnology (AREA)
- Zoology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Wood Science & Technology (AREA)
- Genetics & Genomics (AREA)
- Tropical Medicine & Parasitology (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Biodiversity & Conservation Biology (AREA)
- Hydrology & Water Resources (AREA)
- Environmental & Geological Engineering (AREA)
- Water Supply & Treatment (AREA)
- Purification Treatments By Anaerobic Or Anaerobic And Aerobic Bacteria Or Animals (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention belongs to the technical field of wastewater treatment, and particularly relates to denitrifying bacteria and application thereof in nitrogen-containing wastewater treatment. The strain isPseudomonas taiwanensisEN-F2, the preservation number is CCTCC NO: M20211515. The strain of the invention has excellent denitrification capability in heterotrophic nitrification and aerobic denitrification processes, and has good application prospects in nitrogen-containing wastewater treatment.
Description
Technical Field
The invention belongs to the technical field of wastewater treatment, and particularly relates to denitrifying bacteria and application thereof in nitrogen-containing sewage treatment.
Background
Nitrogen compounds play an important role in the metabolic process of life. However, inorganic nitrogen discharged from agricultural production and rural life enters rivers and lakes, threatens the survival of aquatic organisms, initiates the reproduction of a large amount of bacteria, further worsens water resources, emits unpleasant odors, and finally threatens human health (Zheng, z., li, w., zhang, d., qin, w., zhao, y., lv, l.2019.effect of iron and manganese on ammonium removal from micro-polluted source water by immobilized HITLi (T) at 2degrees C.Bioresour Technol.285,121367). Studies have shown that when the concentration of ammonium exceeds 0.5mg/L, the treatment efficiency of drinking water is lowered. However, some contaminated surface waters in China have ammonium contents exceeding 2mg/L (Chu, H.Q., cao, D.W., dong, B.Z., qiang, Z.M.2010.Bio-diatomite dynamic membrane reactor for micro-poled surface water treatment Module. Water Res.44 (5), 1573-1579; stef.n, D., erd lyi, N., izs. A.k., B., Z.ray, G.and Vargha, M.2019.formation of chlorination by-products in drinking water treatment plants using breakpoint chloride, microchemical journal.149, 104008). Meanwhile, ammonium is used as a substrate for nitrification, and is converted into hydroxylamine and nitrite by nitrifying bacteria under aerobic or anaerobic conditions, and then is converted into nitrate. Previous reports indicate that hydroxylamine is toxic to microorganisms or has a great mutagenic effect on bacteria, plants and viruses as an intermediate in the nitration process (Gross, p.1985. Biological activity of hydroxylamine: a review.critical reviews in toxicology (1), 87-99), and also inhibits the function of denitrifying bacteria by disrupting the ribosome translation process (Xing, c.y., fang, y.c., chen, x, guo, j.s., shen, y., yan, p., fang, chen, y.2020. Effect of hydroxylamine on community of ANAMMOX SLage.environ.41 (7), 3365-3372). To date, only a few nitrifying bacteria have the ability to survive in the presence of hydroxylamine. Nitrite is an intermediate product of nitrification or denitrification. Nitrite accumulation can prevent normal blood oxygen transport, cause hypoxia and even asphyxia, and when water contains large amounts of nitrite, aquatic organisms can be harmed (Jia, W.L., wang, Q., zhang, J., yang, W.H., zhou, X.W.2016.Nutrients removal and nitrous oxide emission during simultaneous nitrification, denitrication, and phosphorus removal process: effect of iron. Environmental Science and Pollution research.23 (15), 15657-15664; lewis, W.M., morris, D.P.2011.Toxicity of Nitrite to Fish: A review. Transactions of the American Fisheries society.115 (2), 183-195). The accumulation of nitrite also inhibits the nitration process. In addition, nitrite can be oxidized to nitrate under the action of certain microorganisms (Zeng, j., liao, s., qiu, m., chen, m., ye, j., zeng, j.and Wang, a.2020.effects of carbon sources on the removal of ammonium, nitrite and nitrate nitrogen by the red yeast Sporidiobolus pararoseus y1. Bioresource technology.312, 123593). The nitrate has the following characteristics: the water solubility is high, and the relative stability in water is strong; nitrate, when converted to nitrosamine, can cause not only eutrophication of water but also carcinogenesis in humans (Devlin, j.f., efficiency, r.i., butler, b.j.2000.the effects of electron donor and granular iron on nitrate transformation rates in sediments from a municipal water supply aquifer.journal of contact water.46 (1), 81-97; zhao, y.x., feng, c.p., wang, q.h., yang, y.n., zhang, z.y., sugiura, n.2011.nitate removal from groundwater by cooperating heterotrophic with autotrophic denitrification in a biofilm-electric reactor j Hazard mater.192 (3), 1033-1039). Therefore, how to efficiently and simultaneously remove ammonium and hydroxylamine and control the accumulation of nitrite and nitrate has become a problem to be solved.
Bacteria with heterotrophic nitrification-aerobic denitrification capability have good application prospects in wastewater treatment and are widely paid attention to. The discovery of the aerobic denitrifying bacteria breaks through the traditional denitrification theory and has the advantages of high efficiency, high growth speed, low consumption, environmental protection and the like (Yao, S., ni, J., chen, Q.and Borthwick, A.G.2013.energy and characterization of a bacteria consortiumcapable of heterotrophic nitrification and aerobic denitrification at low temperature.Bioresource technology 127, 151-157). More importantly, the aerobic denitrifying bacteria can effectively reduce the accumulation of hydroxylamine and nitrite. For example, pseudomonas taiwanensis J488 can remove ammonium or hydroxylamine at low temperature and no accumulation of nitrite was observed with ammonium as the sole nitrogen source (He, t., xie, d., ni, j., li, z., li, z.2020.Nitrous oxide produced directly from ammonium, nitrate and nitrite during nitrification and Density determination. Journal of hazard materials.388, 122114).
However, in the case where a plurality of nitrogen sources coexist, few bacteria are able to efficiently and simultaneously remove hydroxylamine or ammonium of high strength. For example, under optimized conditions, pseudomonas sp.dm02 can only remove 10mg/L of ammonium in 12 hours (Deng, m., zhao, x., senbat, y., song, k., he, x.2021.nitogen removal by heterotrophic nitrifying and aerobic denitrifying bacterium Pseudomonas sp.dm02: removal performance, mechanism and immobilized application for real aquaculture wastewater process.bioresource technology 322, 124555). The Zobellella taiwanensis DN-7 and Pseudomonas mendocina X strains, although highly effective in removing ammonium, do not have the ability to remove hydroxylamine (Joo, H.S., hirai, M., shoda, M.2005. Characics of ammonium removal by heterotrophic nitrification-aerobic denitrification by Alcaligenes faecalis No.4.J Biosci Bioeng 100 (2), 184-191; xie, F., thiri, M., wang, H.2021.Simultaneous heterotrophic nitrification and aerobic denitrification by a novel isolated Pseudomonas mendocina X49.Bioresource technology 319, 124198). Pseudomonas stutzeri C3 has proven to be highly effective in nitrate removal but lacks the amoA gene and fails to undergo heterotrophic nitrification (Ji, B., yang, K., wang, H.Y., zhou, J., zhang, H.N.2014.Aerobic denitrification by Pseudomonas stutzeri C3 incapable of heterotrophic identification.bioprocess and Biosystems engineering.38 (2), 407-409). Pseudomonas stutzeri GEP-01 has heterotrophic nitrification capacity but relatively low removal rates for ammonium (3.73 mg/L/h) (Gao, J., zhu, T., liu, C., zhang, J., gao, J., zhang, J., cai, M.and Li, Y.2020.Ammonium removal characteristics of heterotrophic nitrifying bacterium Pseudomonas stutzeri GEP-01with potential for treatment of ammonium-rich water Bioprocess Eng 43 (6), 959-969). In addition, even though both strains Photobacterium sp.NNA4 and alligenes faecalisNo.4 were able to remove hydroxylamine, cells could not grow under this condition (Joo, H.S., hirai, M., sheda, M.2005. Characics of ammonium removal by heterotrophic nitrification-aerobic denitrification by Alcaligenes faecalis No.4.JBiosci Bioeng 100 (2), 184-191; liu, Y., ai, G.M., wu, M.R., li, S.S., miao, L.L., liu, Z.P.2019.Photobacterium sp.NNA4, an efficient hydroxylamine-transforming heterotrophic nitrifier/aerobic denier.J bioscien 128 (1), 64-71).
In conclusion, the existing heterotrophic nitrification-aerobic denitrification bacteria have limitations in the denitrification process, cannot be propagated by using hydroxylamine, cannot grow in the presence of hydroxylamine and nitrite at the same time, and the nitrification capacity needs to be further improved so as to improve the treatment effect of nitrogen-containing sewage.
Disclosure of Invention
In view of the above, the present invention aims to provide denitrifying bacteria and application thereof in nitrogen-containing wastewater treatment.
In order to achieve the above purpose, the technical scheme of the invention is as follows:
the invention aims at providing a strain which is Pseudomonas taiwanensis EN-F2 and is preserved in China center for type culture Collection, address: eight paths of Lopa nationality mountains in Wuchang district of Wuhan, hubei province are preserved with the number CCTCC NO: m20211515.
The inventor finds that the Pseudomonas taiwanensis EN-F2 strain can simultaneously and efficiently remove various inorganic nitrogen in a water body in the research process.
The invention also aims to provide a screening method of the strain, which comprises the following steps:
collecting a soil sample, and placing the soil sample in a sterilized primary screening culture medium for culture; and 5 mu L of bacterial suspension is smeared on a rescreening culture medium and streaked, and the culture is continued, and bacteria which can turn the bromothymol blue solid culture medium into blue are selected and purified as candidate bacteria.
Further, the preliminary screening medium includes: HONH (HONH) 3 Cl、NaNO 2 、CaCI 2 、K 2 HPO 4 、MgSO 4 、KH 2 PO 4 、C 6 H 5 Na 3 O 7 ·2H 2 O and Fe 2 (SO 4 ) 3 And pH of the primary screening medium=7.20.
Further, the preliminary screening medium includes: 0.0496g/L HONH 3 Cl、0.0986g/L NaNO 2 、0.014g/L CaCI 2 、3.5g/L K 2 HPO 4 、0.04g/L MgSO 4 、1.5g/L KH 2 PO 4 、3.676g/LC 6 H 5 Na 3 O 7 ·2H 2 O and 0.009g/L Fe 2 (SO 4 ) 3 。
Further, the rescreening medium comprises：(NH 4 ) 2 SO 4 、MgSO 4 、KH 2 PO 4 、Na 3 C 6 H 5 O 7 .2H 2 O、FeSO 4 ·7H 2 O,、CaCl 2 Agar and bromothymol blue reagent.
Further, the rescreening medium comprises: 0.236g (NH) 4 ) 2 SO 4 ,0.488g MgSO 4 、1gKH 2 PO 4 、8.5g Na 3 C 6 H 5 O 7 .2H 2 O、0.592g FeSO 4 .7H 2 O、0.094g CaCl 2 18g agar and bromothymol blue reagent 1mL.
Further, the concentration of bromothymol blue reagent was 1.5g/100ml ethanol.
The invention also aims to provide the application of the strain in nitrogen-containing sewage treatment.
The invention has the beneficial effects that:
the strain of the invention has excellent denitrification capability in both heterotrophic nitrification and aerobic denitrification processes.
The strain provided by the invention has a good application prospect in nitrogen-containing wastewater treatment.
Drawings
FIG. 1 shows the morphology of strain EN-F2; wherein a is colony morphology on LB medium; b is colony morphology on the rescreening medium plate; c is the morphology of gram-stained colonies under an optical microscope; d is the cell morphology under a scanning electron microscope;
FIG. 2 is a phylogenetic tree;
FIG. 3 is a graph showing the results of heterotrophic nitrification capacity, wherein A is a graph showing the results of detection using ammonium sulfate as a single nitrogen source; b is a detection result graph taking hydroxylamine as a single nitrogen source; c is a graph of the effect of hydroxylamine on ammonium; the detection results are the mean ± SD (standard error) of three replicates.
FIG. 4 is a diagram showing the detection result of denitrification capability, wherein A is a diagram showing the detection result of nitrate as a single nitrogen source; b is a detection result graph taking nitrite as a single nitrogen source; c is a graph of the effect result of nitrite on nitrate; the detection results are the mean ± SD (standard error) of three replicates.
FIG. 5 is a graph of the results of detection of synchronous nitrification-denitrification performance, wherein A is a graph of the results of detection with ammonium salt and nitrate as nitrogen sources; b is a detection result graph taking ammonium and nitrite as nitrogen sources;
fig. 6 is a graph of detection results of simultaneous nitrification and denitrification of hydroxylamine and nitrate/nitrite, wherein a is a graph of detection results using hydroxylamine and nitrate as nitrogen sources, and B is a graph of detection results using hydroxylamine and nitrite as nitrogen sources.
Detailed Description
The examples are presented for better illustration of the present invention, but are not intended to limit the scope of the present invention to the examples. Those skilled in the art will appreciate that various modifications and adaptations of the embodiments described above are possible in light of the above teachings and are intended to be within the scope of the invention. In the present invention, the term "wt%" means mass content.
Example 1
Isolation and identification
Soil samples were collected from vegetable fields (Guizhou Kogyo, china), 1g of wet soil was transferred to a 250mL Erlenmeyer flask, the conical flask is filled with 100mL of sterilized primary screening culture medium, and then shake-cultured at 25deg.C and 150r/min for 3d; the culture was continued three times under this culture condition. Wherein, the formula of the preliminary screening culture medium is as follows: 0.0496g/L HONH3Cl, 0.0986g/L sodium nitrite (sodium nitrite is used in this example, as long as the nitrogen content reaches 50mg/L, other nitrites can also be used), 0.014g/L CaCI 2 、3.5g/L K 2 HPO 4 、0.04g/L MgSO 4 、1.5g/L KH 2 PO 4 、3.676g/L C 6 H 5 Na 3 O 7 ·2H 2 O and 0.009g/L Fe 2 (SO 4 ) 3 And the pH of the medium was adjusted to 7.2;
taking 5 mu L of the bacterial suspension obtained by culture, coating streaks on a rescreening culture medium plate, and culturing for 3d at 25 ℃; selecting and purifying bacteria which can turn the rescreening culture medium into blue as candidate bacteria; efficiency of hydroxylamine and nitrite removalThe highest strain (hereinafter referred to as strain EN-F2) is kept in 30% glycerol solution for long-term storage at-20deg.C; wherein, the formula of the rescreening culture medium is as follows: 0.236g (NH) 4 ) 2 SO 4 ,0.488gMgSO 4 、1gKH 2 PO 4 、8.5g Na 3 C 6 H 5 O 7 .2H 2 O、0.592g FeSO 4 .7H 2 O、0.094g CaCl 2 18g agar and bromothymol blue reagent 1mL;
the morphology of strain EN-F2 was observed using a scanning electron microscope (SU 8100, hitachi, tokyo, japan) and an optical microscope (Olympus BX53-DIC, japan), and as a result, as shown in FIG. 1, the formulation of LB medium used during the morphology of strain EN-F2 was: 10.0g/L tryptone, 10g/LNaCl and 5.0g/L yeast extract, the pH of the culture medium is adjusted to 7.20, 18% agar is added into the liquid culture medium for solid culture medium (liquid LB culture medium is used for activating and expanding bacteria, and solid LB culture medium is used for purifying the bacteria by coating and streaking the bacteria).
Extracting DNA using a bacterial DNA extraction kit (Magen) and serving as a template for polymerase chain reaction amplification (PCR); PCR amplification (Wei, R., hui, C., zhang, Y., jiang, H., zhao, Y.and Du, L.2020.Nitrogen removal characteristics and predicted conversion pathways of a heterotrophic nitrification-aerobic denitrification bacterium, pseudomonas aeruginosa P-1.Environmental Science and Pollution Research 28 (6), 7503-7514) was performed by selecting F27 and R1492 universal primers (wherein the nucleotide sequence of the F27 universal primer is 5'-AGAGTTTGATCCTGGCTCAG-3' and the nucleotide sequence of the R1492 universal primer is 5'-GGTTACCTTGTTACGACTT-3'). PCR amplification was performed using a 25. Mu.l reaction system containing 12.5. Mu.l 2X Tap Plus Master mix ll, 1. Mu.l Primer1, 1. Mu.l Primer 2, 0.5. Mu.l DNA, 10. Mu.l sterile water; the PCR conditions are denaturation at 94 ℃ for 5min, 30 cycles per minute, annealing at 55.5 ℃ for 30s and extension at 72 ℃ for 10min;
gel electrophoresis is carried out on the amplified PCR products, and sequencing is carried out by Shanghai Biotechnology company; the gel electrophoresis comprises the following specific steps: weighing 0.4g of agar into 40ml of AE x buffer solution, melting by a microwave oven, adding 5ul of coloring agent, shaking uniformly, pouring into an electrophoresis plate, inserting 11-hole comb, extracting agar after solidification, adding 5ul of DNA into each hole, using maker as a reference, placing into an electrophoresis tank, electrophoresis under 150v and 30/40min, and observing the DNA to the middle of the gel plate.
Sequencing the full-length 16S rRNA gene sequence of the amplified product by entrusting Shanghai biological technology company, and submitting NCBI to obtain accession number;
phylogenetic tree was constructed using MEGA 7.0 program by using neighbor-joining method and 1000 repeated boottrap analysis, and the results are shown in fig. 2.
As can be seen from FIG. 1, the strain EN-F2 can rapidly remove hydroxylamine and nitrite nitrogen, and the bacterial colony of the strain on LB and BTB plates has a yellow circular shape, regular edges, raised middle, smooth and moist surface and is opaque (FIGS. 1a and b); strain EN-F2 was gram negative, short rod-shaped with flagella (fig. 1c, d), whose 16SrRNA gene sequence was submitted to the GenBank database under accession number OK638151.
As can be seen from FIG. 2, the homology search of BLAST shows that the strain EN-F2 has a similarity of 99% with Pseudomonas taiwan. Phylogenetic trees were constructed based on the 16S rRNA gene sequences of strain EN-F2 and other related strains, further demonstrating that strain EN-F2 is very close to P5 of Pseudomonas taiwanensis (FIG. 2). Although He et al reported a species of pseudomonas taiwanensis, the denitrification properties of strain EN-F2 were different from strain J488 reported previously (He et al, 2021 a). In addition, the denitrification capacity of EN-F2 is obviously stronger than that of the strain J488, and a plurality of mixed nitrogen sources can be removed at the same time, so that the EN-F2 has greater application potential in wastewater treatment.
EN-F2 strain denitrification capability detection
Inoculating the strain EN-F2 into LB liquid medium (formula is described above), and shaking-activating at 25deg.C and 150r/min for 24 hr; centrifuging at 6500rpm for 5min, washing the strain with 20ml sterilized pure water for 3 times, and inoculating into 100ml nitrification, denitrification and synchronous nitrification-denitrification culture medium respectively; wherein, the formula of the nitrifying culture medium (per liter) is as follows: 0.236g (NH) 4 ) 2 SO 4 /0.0992g HONH 3 Cl、0.04gMgSO 4 、3.5g K 2 HPO 4 、0.009g Fe 2 (SO 4 ) 3 、1.5g KH 2 PO 4 ,0.014g CaCI 2 、3.064g/1.225gC 6 H 5 Na 3 O 7 ·2H 2 O (ammonium salt or hydroxylamine, respectively, may be used as nitrogen source in the nitration process, if 0.236g (NH 4 ) 2 SO 4 As nitrogen source, 3.064g C is needed 6 H 5 Na 3 O 7 ·2H 2 O, if using 0.0992g HONH 3 Cl is a nitrogen source, 1.225gC is needed 6 H 5 Na 3 O 7 ·2H 2 O), and adjusting the pH of the medium to 7.20; the denitrification medium (per liter) has the following formula: 0.361g of Nitrate (Nitrate, the Nitrate used in this example is potassium Nitrate)/0.246 g of Nitrate (Nitrite, the Nitrite used in this example is sodium Nitrite), 0.361g of KNO 3 、0.246g NaNO 2 、3.5g K 2 HPO 4 、1.5g KH 2 PO 4 、0.014g CaCl 2 、0.009g Fe 2 (SO 4 ) 3 、0.04g MgSO 4 And 3.064g/6.127g C 6 H 5 Na 3 O 7 .2H 2 O (nitrate or nitrite is used as nitrogen source in denitrification process, 3.064gC is needed if 0.361g nitrate is used as nitrogen source 6 H 5 Na 3 O 7 .2H 2 O, if 0.246g of nitrite is used as nitrogen source, 6.127g of C is required 6 H 5 Na 3 O 7 .2H 2 O), and adjusting the PH of the medium to 7.20; the formula of the simultaneous nitrification-denitrification culture medium (per liter) is as follows: 0.236g (NH) 4 ) 2 SO 4 and 0.361g KNO 3 /0.246g NaNO 2 and 0.0496g HONH 3 Cl/0.0496g HONH 3 Cl and 0.361g KNO 3 /0.0496g HONH 3 Cl and 0.246g NaNO 2 (simultaneously, the nitrification and denitrification can respectively take ammonium salt and nitrate, ammonium salt and nitrite, hydroxylamine and nitrate, hydroxylamine and nitrite as mixed nitrogen sources), 0.361g KNO 3 、3.5g K 2 HPO 4 、1.5g KH 2 PO 4 、0.04g MgSO 4 、6.127g/3.676g C 6 H 5 Na 3 O 7 ·2H 2 O (6.127 g C if ammonium, nitrate, ammonium and nitrite are used as mixed nitrogen source to perform simultaneous nitrification and denitrification 6 H 5 Na 3 O 7 ·2H 2 O, if hydroxylamine and nitrate, hydroxylamine and nitrite are used as nitrogen sources, 3.676g C 6 H 5 Na 3 O 7 ·2H 2 O, other cultures unchanged), 0.009g Fe 2 (SO 4 ) 3 And 0.014g CaCl 2 And adjusting the pH of the culture medium to 7.20;
initial cell Optical Density (OD) of Strain EN-F2 600 ) Controlled at 0.2 to remove ammonium salts, nitrates and nitrites, and at 0.3 for hydroxylamine; samples of different nitrogen sources were collected every 6 hours to determine cell density and concentration of ammonium, hydroxylamine, nitrate, nitrite and total nitrogen;
spectrophotometers (Shanghai Metash Instruments, UV-6000, shanghai, china) were used to detect cell optical density and inorganic nitrogen;
the total nitrogen is calculated by subtracting 2 times of 275nm background absorbance from the absorbance value of the digested potassium sulfate at 220 nm; total nitrogen, ammonium, hydroxylamine, nitrate and nitrite were measured by taking the supernatant of the samples in the medium, and respectively measuring by alkaline potassium persulfate digestion-ultraviolet spectrophotometry (total nitrogen measurement), indoxyl blue ultraviolet spectrophotometry (ammonium salt measurement), 8-hydroxyquinoline ultraviolet spectrophotometry (hydroxylamine measurement), ultraviolet spectrophotometry and N- (1-naphthyl) -ethylenediamine ultraviolet spectrophotometry (nitrite measurement); the alkaline potassium persulfate digestion ultraviolet spectrophotometry specifically comprises the following steps: taking 1ml of culture, namely, diluting the culture to 10ml based on 25ml of colorimetric tube, adding 5ml of alkaline potassium persulfate (needing recrystallization), boiling at 121 ℃ for 30min, cooling, adding 1ml of diluted hydrochloric acid (hydrochloric acid: water=1:9), diluting the culture to 25ml with water, and measuring at 220nm and 275nm respectively; the indoxyl blue ultraviolet spectrophotometry comprises the following specific steps: taking 0.5ml of supernatant after centrifugation (6500 rpm, 5 min) to culture on a 50ml cuvette, diluting to 30ml with deionized water (18.2), adding 5ml of phenol solution and sodium hypochlorite solution, standing for 1h, adding 1ml of masking agent, fixing the volume to 50ml, and measuring the absorbance value at 625nm wavelength of visible light; the ultraviolet spectrophotometry of the 8-hydroxyquinoline comprises the following specific steps: taking 1ml of supernatant to culture based on 10ml of colorimetric tube, adding 1ml of phosphate buffer solution and fixing the volume to 5ml, adding 0.2ml of trichloroacetic acid, adding 1ml of 8-hydroxyquinoline, shaking, adding 1ml of sodium carbonate, shaking, boiling water for 1min (generating green), standing for 15min, and measuring absorbance at 750nm wavelength; the ultraviolet spectrophotometry and the N- (1-naphthyl) -ethylenediamine ultraviolet spectrophotometry specifically comprise the following steps: taking 1ml of supernatant to culture based on 50ml of colorimetric tube, adding 1ml of color reagent, standing for 20min, and detecting the light absorption value at 540nm wavelength; the calculation formula of the denitrification rate is ef= (R1-R2)/h, wherein Ef, R1, R2 and h respectively represent the inorganic nitrogen removal rate, the initial nitrogen concentration within a certain time, the final nitrogen concentration and the time interval.
Heterotrophic nitrification capacity detection of EN-F2 strain
The ammonium oxidation rate of the strain EN-F2 is detected by adopting ammonium sulfate as a single nitrogen source, and the specific steps are as follows: 8ml of ammonium sulfate culture was taken every 6 hours, centrifuged at 6500rpm for 5min, 0.5ml of supernatant was taken in a 50ml cuvette, diluted to 30ml with deionized water, then 5ml of phenol, 5ml of sodium hypochlorite solution were added, left standing for 1h, 1ml of masking agent was added, finally the wavelength thereof was measured at 625nm, the OD value was taken into standard curve y=1.2157x+0.0062, the obtained concentration was multiplied by 50, and the result was divided by the sampling amount, and the result is shown in FIG. 3A.
As can be seen from FIG. 3A, the EN-F2 strain was directly in log phase for the first 6 hours, without delay, the 52.90mg/L initial ammonium nitrogen was almost completely removed (98.89%), the maximum removal rate was 8.72mg/L/h 6 hours after inoculation, and the maximum removal rate of ammonium was possibly higher than 8.72mg/L/h, because the ammonium nitrogen could be removed 98.89% before 6 hours. As compared to the reported strains, the maximum removal rate of ammonium nitrogen was higher than Exiguobacterium mexicanum SND-01 (2.24 mg/L/h) (Cui, Y., cui, Y.W. and Huang, J.L.2021.A novel halophilic Exiguobacterium mexicanum strain removes nitrogen from saline wastewater via heterotrophic nitrification and aerobic density technique 333,125189), vibrio diabolicusSF (2.29 mg/L/h) (Duan, J., fang, H., su, B., chen, J.and Lin, J.2015.Characterization of a halophilic heterotrophic nitrification-aerobic denitrification bacterium and its application on treatment of saline water technique Biodensity technique 179, 421-428), pseudomonas tolaasii Y-11 (2.04 mg/L/h) (He, T., li, Z., sun, Q., xu, Y.and Ye, Q.2016.Heteropic nitrification and aerobic denitrification by Pseudomonas tolaasii Y-11without nitrite accumulation during nitrogen conversion.Bioresour Technol 200,493-499).
Ideonella sp.TH17 (1.4 mg/L/h) (Zhang, L.J., xie, Y., ding, L.Y., qiao, X.J.and Tao, H.C.2020b.Highly efficient ammonium removal through nitrogen assimilation by a hydrogen-oxidizing bacterium, ideonella sp.TH17.environ Res 191,110059) and Ochrobactrum anthropic LJ (3.85 mg/L/h) (Lei, X., jia, Y., chen, Y.and Hu, Y.2019.Simultaneous nitrification and denitrification without nitrite accumulation by a novel isolated Ochrobactrum anthropic LJ 81.Bioresource technologies 272, 442-450), when OD 600 100% of the ammonium was removed at 12h as the value increased from 0.25 to 0.98. Ammonium is released by dead bacteria after 12h inoculation resulting in an increase in ammonium nitrogen concentration (Li, c., yang, j., wang, x., wang, e., li, b., he, r.and Yuan, h.2015.removal of nitrogen by heterotrophic nitrification-aerobic denitrification of a phosphate accumulating bacterium Pseudomonas stutzeri YG-24.Bioresour Technol 182,18-25), while total nitrogen concentration decreases from 51.55 to 3.68mg/L with removal efficiency and maximum removal rates of 92.86% and 7.98mg/L/h, respectively. Furthermore, it was observed that the system pH increased from 7.19 to 9.00 (not shown in FIG. 3A) over time during the entire ammonium removal, similar to Zobellella taiwanensisDN-7 (Lei, Y., wang, Y., liu, H., xi, C.and Song, L.2016.A novel heterotrophic nitrifying and aerobic denitrifying bacterium, zobellella taiwanensis DN-7,can remove high-strangth ammonia.applMicrobiol Biotechnol 100 (9), 4219-4229), indicating that EN-F2 has denitrification capacity (Zhu, L., ding, W., feng, L.J., kong, Y., xu, J.and Xu, X.Y.2012.isolation of aerobic denitrifiers and characterization for their potential application in the bioremediation of oligotrophic ecosystem.Bioresource technology 108,1-7). Only 0.11mg/L nitrite was detected after 6h of inoculation, after 12h Disappearance indicates that the EN-F2 strain can perform denitrification during the nitrification process. Hydroxylamine was not detected in this procedure, but nitrate was observed at 0.61mg/L, which was consistent with the detection of nitrate at 12.95mg/L by strain Ochrobactrum anthropic LJ (Lei, X., jia, Y., chen, Y., and Hu, Y.2019.Simultaneous nitrification and denitrification without nitrite accumulation by a novel isolated Ochrobactrum anthropic LJ81. Bioresource technologies 272, 442-450). The results show that the strain EN-F2 can efficiently convert most of ammonium salt into nitrogen-containing gas (N 2 O and N 2 )。
To confirm the ability of strain EN-F2 to remove hydroxylamine and to further explore the nitration pathway of strain EN-F2, hydroxylamine was used as a single nitrogen source test, with the specific steps of: 8ml of hydroxylamine culture was centrifuged at 6500rpm for 5min every 6h, 0.5ml of supernatant was diluted to 30ml with deionized water in a 50ml cuvette, then 5ml of phenol and 5ml of sodium hypochlorite solution were added, and the mixture was left standing for 1h, 1ml of masking agent was added, and finally the wavelength thereof was measured at 625nm, and the OD value was taken into standard curve y=1.2157x+0.0062, and the result was obtained by dividing the value obtained by 10 and the sampling amount, and the result was shown in FIG. 3B.
As can be seen from FIG. 3B, after 18h of cultivation, the OD of the strain EN-F2 600 The value increased from 0.35 to 0.58. Meanwhile, the hydroxylamine concentration is obviously reduced from 23.32 to 1.48mg/L, and the removal efficiency and the maximum removal rate are 93.69% and 2.12mg/L/h respectively. Hydroxylamine removal rates were significantly higher than Pseudomonas taiwanensisJ488 (0.28 mg/L/H) (He, T., xie, D., ni, J., li, Z.and Li, Z.2020.Nitrous oxide produced directly from ammonium, nitrate and nitrite during nitrification and, density, journal of Hazardous Materials 388), glutamicibacter arilaitensis EM-H8 (0.21 mg/L/H) (Liu, Y., hu, T., song, Y., chen, H.and Lv, Y.2015, heteropht nitrogen removal by Acinetobacter sp.Y1 isolated from coke plant water. Journal of Bioscience and Bioengineering (5), 549-554), and Photobacterium sp NNA (0.7 mmol/L/H) (Liu, Y., ai, G.M., wu, M.R., li, S.S., miao, L.L.liu, Z.P.2019.Photobacteria sp.NNA4, an efficient hydroxylamine-transforming heterotrophic nitrifier/density, and Biob.128). When the culture time is prolonged toAt 30h, 100% of the hydroxylamine was removed. Furthermore, 62.95% of the total nitrogen is converted to other inorganic or gaseous nitrogen with a maximum removal rate of 1.82mg/L/h, which is also significantly higher than the strain described above. After 18 hours of cultivation, the nitrite accumulation reached the peak value of 14.81mg/L, and after 30 hours of cultivation, the nitrite accumulation decreased to 0.53mg/L. The pH was increased from 7.15 to 8.05. Notably, nitrite and nitrate can be detected and reduced during hydroxylamine oxidation, meaning that denitrification occurs simultaneously. In addition, cell growth and accumulation of nitrite by strain EN-F2 were different from those by strains Photobacterium sp.NNA4 (Liu, Y., ai, G.M., wu, M.R., li, S.S., miao, L.L. and Liu, Z.P.2019.Photobacterium sp.NNA4, an efficient hydroxylamine-transforming heterotrophic nitrifier/aerobic Denitrifier.J Biosci Bioeng 128 (1), 64-71) and Alcaligenes faecalis No.4 (Joo, H.S., hirai, M.and Shoda, M.2005.Characteristics No.4.J Biosci Bioeng 100 (2), 184-191), in which nitrite accumulation and cell growth were not observed even if hydroxylamine was removed from both. In summary, EN-F2 can be grown in the presence of hydroxylamine and completely removed.
To further investigate the effect of hydroxylamine on the ammoxidation process, 10mg/L of hydroxylamine was added to the above-mentioned nitrifying medium, and the ammonium nitrogen and hydroxylamine as well as nitrifying intermediates were measured every 6 hours, and the results are shown in FIG. 3C; wherein, the detection step of ammonium nitrogen is (the same as the nitrification step using ammonium sulfate as nitrogen source); the detection step of hydroxylamine is (as in the above nitrification detection step using hydroxylamine as a nitrogen source); the detection step of the nitrifying intermediate product is that (the nitrifying intermediate product has nitrate and nitrite produced, and the step is the same as the denitrification step which is carried out by taking nitrate and nitrite as nitrogen sources respectively).
As can be seen from FIG. 3C, hydroxylamine was completely converted, with an average removal rate of 1.59mg/L/h; the ammonium nitrogen removal efficiency was 85.46%. The addition of 10mg/L hydroxylamine to the ammonium salt medium resulted in an OD which was comparable to that obtained when ammonium was the sole nitrogen source 600 The improvement is 0.40 percent. After 12h of cultivation, the ammonium nitrogen concentration and the total nitrogen concentration are obviously reduced from 48.13 mg/L and 57.17mg/L to 0.48 mg/L and 11.01mg/L respectivelyThe removal efficiency was 99.00% and 80.74%. The system pH increased from 7.19 to 9.05. At the same time, the peak of nitrite (6.65 mg/L) appeared at 6h, and then remained relatively unchanged (about 2.40 mg/L). In this procedure, low concentrations (1.18 mg/L) of nitrate were detected, in contrast to the results obtained for Enterobacter cloacae CF-S27, and the EN-F2 strain did not accumulate nitrate and nitrite even in the presence of hydroxylamine (Padhi, S.K., tripath, S., mohanty, S.and Maiti, N.K.2017.Aerobic and heterotrophic nitrogen removal by Enterobacter cloacae CF-S27 with efficient utilization of hydroxyamine. Bioresource technology 232, 285-296). In addition, the maximum removal rate of ammonium after 30h of culture was reduced from 8.72 to 6.8mg/L/h due to the addition of hydroxylamine, indicating that hydroxylamine was able to retard oxidation of ammonium nitrogen. Likewise, the total nitrogen removal efficiency (80.74%) was lower than the removal efficiency (92.84%) with ammonium as the sole nitrogen source. In summary, the EN-F2 strain was able to completely remove hydroxylamine and ammonium nitrogen by prolonged culture.
Aerobic denitrification capability detection
The denitrification capability of the strain EN-F2 under the aerobic condition is detected by taking nitrate as the sole nitrogen source, and the result is shown in figure 4A; wherein, the detection steps are as follows: taking 8ml of potassium nitrate culture medium for centrifugation (6500 rpm, 5 min), taking 1ml of supernatant in a 25ml colorimetric tube, directly measuring an OD value under the conditions of 220 and 275 wavelengths, subtracting the OD value of 275 times from the OD value measured by 220, multiplying the value obtained by labeling with the curve y= 0.2396x-0.0038 by 25, and dividing the value by the sampling amount. As can be seen from FIG. 4A, after 12 hours of cultivation, the nitrate concentration of the strain EN-F2 was rapidly decreased and 88.95% was removed, and the maximum removal rate was 5.80mg/L/h, and thereafter, the strain EN-F2 was maintained at about 6.01 mg/L. The maximum nitrate removal rate was much higher than 1.90mg/L/h (He, T., wu, Q., ding, C., chen, M.and Zhang, M.2021b.hydrooxamine and nitrite are removed effectively by Streptomyces mediolani strain EM-B2. Ecotoxaol environment Saf 224,112693), 1.04mg/L/h (Ouyang, L., wang, K, liu, X., wong, M.H., hu, Z., chen, H., yang, X.and Li, S.2020.A study on the nitrogen removal efficacy of bacterium Acinetobacter tandoii MZ-5from a contaminated river of Shenzhen,Guangdong Province,China.Bioresour Technol 315,123888) of Pseudomonas mendocina X49 (Xie, F., thiri, M.and Wang, H.2021.Simuline heterotrophic nitrification and aerobic denitrification by a novel isolated Pseudomonas mendocina X49. Biol 319,124198).
To further clarify whether the strain EN-F2 can remove low concentrations of nitrate, 10mg/L of nitrate was used as the sole nitrogen source (to examine the denitrification capacity of the strain EN-F2 under aerobic conditions, the test step was to culture the strain EN-F2 in a 10mg/L potassium nitrate-containing medium, take 2ml of medium supernatant every 6 hours, and then perform the denitrification step using potassium nitrate as the nitrogen source as described above.
Only 10mg/L nitrate consumption was detected in the experiment, no nitrate was detected at 6h, indicating that all consumption was occurring and no other inorganic nitrogen detection was performed.
As shown by the test results, the strain EN-F2 can completely remove 10mg/L of nitrate. Thus, it can be inferred that strain EN-F2 requires more nutrients such as Ca 2+ Or Fe2 + To remove high concentrations of nitrate. The system pH increased from 7.21 to 9.11. In addition, total nitrogen was reduced from 58.29 to 13.10mg/L with a removal efficiency of 77.53% and a maximum removal rate of 4.42mg/L/h significantly higher than Acinetobacter tandoii MZ-5 (1.06 mg/L/h) (Ouyang, L., wang, K., liu, X., wong, M.H., hu, Z., chen, H., yang, X.and Li, S.2020.A study on the nitrogen removal efficacy of bacterium Acinetobacter tandoii MZ-5from a contaminated river of Shenzhen,Guangdong Province,China.Bioresour Technol 315,123888) and Streptomyces mediolani EN-B2 (1.60 mg/L/h) (He, T., wu, Q, ding, C., chen, M.and Zhang, M.2021 b.hydroylamine and nitrite are removed effectively by Streptomyces mediolani strain EM-B2.Ecotoxol Environ Saf 224,112693). Nitrite accumulated to a peak of 6.79mg/L after 6h of incubation and then dropped to 2.40mg/L, which was compared with Paracoccus denitrificans Z195 (Zhang, H, li, S, ma, B, huang, T, qiau, H, zhao, Z, huang, X.and Liu, K.2020a.Nitrate removal characteristics and (13) C metabolic pathways of aerobic denitrifying bacterium Paracoccus denitrificans Z195. Bioresource technology 307, 123230) and F.solani RADF- 77 (Cheng, h. -y., xu, a. -a., kumar Awasthi, m., kong, d. -d., chen, j. -s, wang, y. -f.and Xu, p.2020.aerobic denitrification performance and nitrate removal pathway analysis of a novel fungus Fusarium solani RADF-77.Bioresource Technology 295), but similar to p.dentriforms ISTOD1 conflict (Medhi, k., singhal, a., chauhan, d.k.and Thakur, i.s.2017.invest gaming the nitrification and denitrification kinetics under aerobic and anaerobic conditions by Paracoccus denitrificans istogd 1.bioresource Technology 242, 334-343), the concentration of ammonium increased from 0 to 5.34mg/L, possibly resulting from dead cell lysis. This phenomenon is consistent with the previously reported phenomenon for strain EN-B2 (He, t., wu, q., ding, c., chen, m.and Zhang, m.2021b.hydroxylamine and nitrite are removed effectively by Streptomyces mediolani strain EM-B2.Ecotoxicol Environ Saf 224,112693).
In order to further study denitrification characteristics, the strain EN-F2 is cultured under aerobic conditions by taking nitrite as a unique nitrogen source, and the denitrification capacity is detected by the following detection steps: centrifuging 8ml of nitrite culture medium, taking 0.2ml of supernatant after centrifugation, diluting to 50ml in a colorimetric tube of 50ml with deionized water, adding 1ml of color-developing agent, standing for 20min, measuring an OD value at a wavelength of 540nm, and obtaining the concentration of nitrite when the OD value is y=0.0613x+0.0024. The results are shown in FIG. 4B.
As can be seen from FIG. 4B, the growth of strain EN-F2 is positively correlated with the removal of nitrite and total nitrogen. As bacteria grew from 0.18 to 0.67, nitrite nitrogen and total nitrogen were removed 86.31% and 68.15%, respectively, after 12h of incubation, with maximum removal rates of 4.55 and 3.31mg/L/h, respectively. After 18h of cultivation, the removal efficiencies of nitrite and total nitrogen are 87.47% and 73.62%, respectively, which are similar to 89.80% when nitrate is the only nitrogen source, but lower than the removal efficiency (100%) of ammonia or hydroxylamine, which indicates that the heterotrophic nitrification capacity of the strain EN-F2 is stronger than that of aerobic denitrification. Nevertheless, the nitrite removal rate of strain EN-F2 was also significantly higher than that of most strains, such as P.putida Y-12 (1.60 mg/L/h) (Ye, Q., li, K, li, Z., xu, Y, he, T., tang, W.and Xiang, S.2017.therapic identification-Aerobic Denitrification Performance of Strain Y-12under Low Temperature and High Concentration of Inorganic Nitrogen Conditions.Water 9 (11)), P.putida NP5 (1.33 mg/L/h) (Yang, L., wang, X.H., cui, S., ren, Y.X., yu, J., chen, N., xiao, Q., guo, L.K. and Wang, R.H.2019.Simultaneous removal of nitrogen and phosphorous by heterotrophic Nitrification-aerobic denitrification of a metal resistant bacteriumPseudomonas putida strain NP5.Bioresource technology 285,121360), F.solani RADF-77 (3.40 mg/L/h) (Cheng, H.—Y., xu, A.—A., kumar Awasthi, M..Kong, D.—D..Chen, J.—S., wang, Y.—F.and Xu, P.2020.aerobic denitrification performance and nitrate removal pathway analysis of a novel fungus Fusarium solani RADF-77.Bioresource Technology 295) and Ochrobactrum anthropic LJ (4.12 mg/L/h) (Lei, X..Jia, Y..2019.Simultaneous Nitrification and denitrification without nitrite accumulation by a novel isolated Ochrobactrum anthropic LJ 81.Bioresource technology 272, 442-450). In addition, 2.12mg/L of nitrate was detected during nitrite removal and then almost completely removed at 30h, which is inconsistent with strains Arthrobacter arilaitensis Y-10 (He, t., xie, d., li, z., ni, j.and Sun, q.2017.am, stimulates nitrate reduction during simultaneous nitrification and denitrification process by Arthrobacter arilaitensis Y-10.Bioresour Technol 239,66-73) and Sporidiobolus pararoseus y1 (Zeng, j., liao, s, qiu, m., chen, m., ye, j., zeng, j.and Wang, a.2020.effects of carbon sources on the removal of ammonium, nitrite and nitrate nitrogen by the red yeast Sporidiobolus pararoseus y 1.bioresource technology 312,123593). The pH of the system increased from 7.20 to 9.04. The above results further indicate that strain EN-F2 can be denitrified under aerobic conditions.
With 50mg/L nitrite and 50mg/L nitrate (0.361 g KNO) 3 ,0.246g NaNO 2 ,3.5gK 2 HPO 4 ,1.5g KH 2 PO 4 ,0.014g CaCl 2 ,0.009g Fe 2 (SO 4 ) 3 ,0.04g MgSO 4 ,6.127gC 6 H 5 Na 3 O 7 .2H 2 O, ph=7.20.) as a mixed nitrogen source, nitrite was detectedThe influence on nitrate is added, and the detection steps are as follows: culturing the strain in LB liquid for 24h, centrifugally washing, adding the strain into a mixed culture medium containing nitrate and nitrite, taking culture medium supernatant every 6h, and measuring the contents of nitrate, nitrite, total nitrogen and ammonium salt, wherein the measuring method is the same as that when nitrate or nitrite is used as a single nitrogen source. The detection result is shown in FIG. 4C.
As can be seen from FIG. 4C, EN-F2 rapidly proliferates over a period of 0-18h, OD 600 The value increased from 0.20 to 1.24 without a lag phase and then reached 1.43 at 30 h. The system pH increased from 7.18 to 9.25. Notably, strain EN-F2 grew better in nitrite and nitrate mixed media than when nitrate was the sole nitrogen source, indicating that supplementation of nitrite in the nitrate removal system did not affect cell growth of EN-F2. In addition, the nitrate and 57.28mg/L nitrite of 59.50 dropped sharply to 25.06 and 8.34mg/L after 18h of incubation, corresponding to maximum removal rates of 3.94 and 4.34mg/L/h, respectively. The total nitrogen removal efficiency was 61.60% and the maximum removal rate was 7.07mg/L/h, which was significantly higher than the total nitrogen with nitrate (4.42 mg/L/h) or nitrite (3.31 mg/L/h) as the sole nitrogen source. After adding nitrite to the nitrate medium, the nitrate removal efficiency and maximum removal rate were reduced from 89.82% and 5.80mg/L/h to 57.88% and 3.94mg/L/h. Nevertheless, the maximum nitrite removal rate in the mixed nitrogen system was 4.34mg/L/h, similar to the 4.55mg/L/h value for a single nitrite source. The above results indicate that nitrate has no effect on the reduction of nitrite, which can be preferentially reduced during denitrification.
Synchronous nitrification-denitrification performance detection
In order to further study the synchronous nitrification-denitrification capacity of the strain EN-F2, 50mg/L of ammonium salt and nitrate are used as mixed nitrogen sources, and the detection steps are as follows: strains activated in LB liquid medium for 24 hours are centrifuged and inoculated into a mixed medium containing ammonium salt and nitrate, and the supernatant medium is taken every 6 hours for measurement, and the measurement method is the same as the method which takes ammonium salt or nitrate as a single nitrogen source respectively. The results are shown in FIG. 5A;
as can be seen from FIG. 5A, when ammonium and nitrateOD when acid salt is used as the mixed nitrogen source 600 The maximum value can reach 1.39, which is higher than 0.94 of single ammonium nitrogen and 0.65 of single nitrate; after 12h of incubation, 100% of the ammonium was removed and the average removal rate was 8.11mg/L/h, indicating that the addition of nitrate did not affect the removal rate of EN-F2 for ammonium. At the same time, the nitrate concentration gradually decreased from 53.18 to 21.62mg/L, with a maximum removal rate of 4.64mg/L/h, and then remained relatively stable. The addition of ammonium did not increase the nitrate removal rate, contrary to the Arthrobacter arilaitensis Y-10 conclusion (He, t., xie, d., li, z., ni, j.and Sun, q.2017.am monitor stimulates nitrate reduction during simultaneous nitrification and denitrification process by Arthrobacter arilaitensis Y-10.Bioresour Technol 239,66-73). After 30h of incubation, the pH of the system increased significantly from 7.18 to 9.36. In addition, when ammonium and nitrate were used as mixed nitrogen sources, nitrification was preferentially performed, and the result was consistent with Pseudomonas tolaasii Y-11 (He, t., li, z., xie, d., sun, q., xu, y., ye, q.and Ni, j.2018.simultanei nitrification and denitrification with different mixed nitrogen loads by a hypothermia aerobic bacteria. Biological degradation 29 (2), 159-170) and Rhodococcus sp HY-1 (Li, w.2013.student on Characteristics in the Removal Process of Ammonia Nitrogen and Nitrate Nitrogen by an Isolated Heterotrophic Nitrification-Aerobic Denitrification Strain Rhodococcus sp.journal of Environmental Protection (01), 74-79) strains, but different from strain CF-S9 (Padhi, s.k., tripath, s., sen, r., mahapatra, a.s., mohanty, s.and Maiti, n.k.2013.Characterisation of heterotrophic nitrifying and aerobic denitrifying Klebsiella pneumoniae CF-S9. 9 strain for bioremediation of waves international biological degradation &Biodegradation 78,67-73). Furthermore, only 0.28mg/L of nitrite was detected in the present study, which is almost equivalent to that in the nitrate removal system. The maximum removal rate of total nitrogen was 6.72mg/L/h, which is lower than total nitrogen with ammonium as the sole nitrogen source (7.98 mg/L/h), but higher than total nitrogen with nitrate as the sole nitrogen source (4.42 mg/L/h). The above results indicate that the strain EN-F2 can simultaneously perform nitrification and denitrification in the presence of mixed ammonium and nitrate.
Taking 50mg/L ammonium and 50mg/L nitrite as mixed nitrogen sources, and detecting the synchronous nitrification-denitrification performance of the strain EN-F2, wherein the detection steps are as follows: strains which are respectively activated and expanded in LB liquid medium for 24 hours are inoculated into a mixed medium containing ammonium and nitrite, the supernatant medium is taken every 6 hours for detection, the measurement method is the same as the method which takes ammonium or nitrite as a single nitrogen source respectively, and the result is shown in figure 5B.
As can be seen from fig. 5B, the removal efficiency of ammonium nitrogen reached 94.85% at 12h as the cell growth increased from 0.17 to 1.20, and was then completely removed after 18 hours of incubation. The maximum removal rate achieved by ammonium in the range of 0-6 hours was calculated to be 6.24mg/L/h, which is significantly lower than 8.72mg/L/h for a single ammonium nitrogen removal system. The results indicate that the addition of nitrite has a negative effect on the removal of ammonium, similar to those reported for strain Pseudomonas stutzeri D (Yang, x., wang, s.and methou, l.2012.effect of carbon source, C/N ratio, nitrate and dissolved oxygen concentration on nitrite and ammonium production from denitrification process by Pseudomonas stutzeri d6. Bioresource technology 104,65-72) and Paracoccus versutus LYM (Shi, z., zhang, y., zhou, j., chen, m.and Wang, x.2013.biological removal of nitrate and ammonium under aerobic atmosphere by Paracoccus versutus lym.bioresource technology 148, 144-148) under carbon source deficient conditions. Notably, at the first 6 hours, the nitrite concentration increased slightly, which may result from the oxidation of ammonium. Accumulation of nitrite suggests that ammonium salt is preferentially removed, which is consistent with the case in mixed nitrogen sources of ammonium and nitrate, but inconsistent with the contradiction between simultaneous removal of ammonium and nitrite by pseudomonas tolaasii Y-11 (He, t., li, z., xie, d., sun, q., xu, y., ye, q.and Ni, j.2018.simultane nitrification and denitrification with different mixed nitrogen loads by a hypothermia aerobic bacteria. Biodegration 29 (2), 159-170. Maximum removal rate of nitrite as the sole nitrogen source (4.55 mg/L/h) being substantially equal to 4.80mg/L/h, which is inconsistent with the result that strain Ochrobactrum anthropic LJ81 can increase nitrite removal rate with supplemental ammonium salt (Lei, x., jia, y., chen, y.and Hu, y.2019.simltane nitrification and denitrification without nitrite accumulation by a novel isolated Ochrobactrum anthropic lj81. Bioresource technologies 272, 442-450).
Furthermore, accumulation of nitrate was observed during the 24h strain culture, and then was completely removed as the culture time was prolonged to 30 hours. The pH of the system increased from 7.20 to 9.32, and these results indicated that denitrification occurred simultaneously with nitrate as the nitrogen source. The maximum removal rate of total nitrogen is 5.09mg/L/h, which is significantly lower than 7.98mg/L/h for ammonium salts as the sole nitrogen source, but higher than 3.31mg/L/h for nitrite. These results further indicate that EN-F2 can be nitrified and denitrified simultaneously and that ammonium can be preferentially removed.
In conclusion, even if high-concentration nitrate or nitrite coexist in the wastewater, the EN-F2 strain can completely remove ammonium salt. Nitrate and nitrite removal may not be complete as part of the nutrients in the medium other than inorganic nitrogen have been consumed.
Simultaneous nitrification and denitrification performance detection of hydroxylamine and nitrate/nitrite
The ability of the strain EN-F2 to synchronously remove the mixed nitrogen source of hydroxylamine (10 mg/L) and nitrate (50 mg/L) is detected, and the detection steps are as follows: the strain which is activated by LB liquid medium and is amplified for 24 hours is inoculated into a mixed medium containing hydroxylamine and nitrate, and detection is carried out every 6 hours, wherein the detection method is the same as that of taking hydroxylamine and nitrate as nitrogen sources respectively; the results are shown in FIG. 6A.
As can be seen from FIG. 6A, 10mg/L hydroxylamine, OD, was added to the nitrate nitrogen removal system 600 The value was increased by 0.44. 11.15mg/L of hydroxylamine was completely consumed, with an average removal rate of 1.86mg/L/h, indicating that low concentrations of hydroxylamine did not hinder the cell growth and denitrification capacity of strain EN-F2. Nitrate was used as an intermediate for hydroxylamine oxidation, increasing by 2.78mg/L with hydroxylamine consumption during the first 6h incubation. After 18h of culture, the nitrate removal efficiency is 77.40%, the maximum removal rate is 5.07mg/L/h, which is obviously lower than 5.80mg/L/h taking nitrate as the sole nitrogen source, and the phenomenon is similar to that of low-concentration hydroxylamine<35 mg/L) of the incongruity in promoting nitrate removal (Zhang, x, xia, y, wang, c, li, j, wu, p, ma, L, wang, y, da, f, liu, w.and Xu, l.2020c.en)hancement of nitrite production via addition of hydroxylamine to Partial Denitrification (PD) biological technologies: functional genes dynamics and enzymatic activites.Bioresource technology 318,124274). Nitrite accumulation (0.48.fwdarw.7.91 mg/L) was observed between 0 and 6h, followed by complete removal within 18 h. The maximum removal rate of total nitrogen is 5.09mg/L/h, which is significantly higher than that of a single nitrate removal system. The system pH increased from 7.15 to 9.11. In summary, the strain EN-F2 can perform simultaneous nitrification and denitrification with hydroxylamine and nitrate as mixed nitrogen sources, and can preferentially and completely remove hydroxylamine.
The hydroxylamine and nitrite removal characteristics of strain EN-F2 were tested, as follows: the strain activated by LB liquid medium and amplified for 24h is inoculated into mixed medium containing hydroxylamine and nitrate, and detection is carried out every 6h, wherein the detection method is the same as that of taking hydroxylamine and nitrite as nitrogen sources respectively. The results are shown in FIG. 6B.
As can be seen from FIG. 6B, the OD600 increased by 0.45 after adding 10mg/L hydroxylamine to the nitrite removal system, as compared to nitrite alone. The system pH increased from 7.18 to 9.22. Along with the propagation of the strain EN-F2, hydroxylamine at a concentration of 13.24mg/L was reduced to 2.85mg/L at the first 6 hours, and was completely removed after 12 hours of cultivation. The maximum removal rate of hydroxylamine was calculated to be 1.73mg/L/h, which is less than 2.21mg/L/h where hydroxylamine was the sole nitrogen source. Nevertheless, hydroxylamine can be efficiently removed in the presence of nitrite. After consumption of hydroxylamine, nitrite was significantly reduced from 53.50 to 10.71mg/L over 18h and then kept constant. The maximum removal rate of nitrite is 6.65mg/L/h, which is significantly higher than 4.55mg/L/h of nitrite as the sole nitrogen source, and the result shows that the removal capacity of the strain EN-F2 for nitrite can be enhanced by supplementing hydroxylamine, which contradicts the report that hydroxylamine is a chemical inhibitor of nitrite oxidizing bacteria (it is worth mentioning that the maximum removal rate (6.90 mg/L/h) of total nitrogen in the case of mixed nitrogen sources is significantly higher than 3.31mg/L/h of total nitrogen with single nitrite as a nitrogen source), which further proves that the addition of hydroxylamine can promote the removal of nitrite.
The above results indicate that hydroxylamine can be completely and preferentially removed in the hydroxylamine removal system, whether it be nitrate-supplemented or nitrite-supplemented. In particular, the removal rate of both nitrite and total nitrogen is significantly increased due to the addition of hydroxylamine, but the addition of hydroxylamine delays the removal rate of nitrate.
Enzyme Activity assay
To further understand the denitrification mechanism of strain EN-F2 during the nitrification and denitrification process, the specific activity of Ammonia Monooxygenase (AMO) of strain EN-F2, hydroxylamine oxidoreductase (HAO) of strain EN-F2, nitrate Reductase (NR) of strain EN-F2 and nitrite reductase (NIR) of strain EN-F2 were examined, and the results are shown in table 1; wherein, the specific activity of Ammonia Monooxygenase (AMO) of the strain is detected by company (Wela, guiyang, china);
nitrite reductase (NIR) specific activity of the strain is detected by using a nitrite reductase activity assay kit (COMIN), and the enzyme activity of the NIR is reflected by the reduction of nitrite; the specific enzyme activity analysis method of the hydroxylamine oxidoreductase (HAO) of the strain comprises the following steps: breaking strain EN-F2 under nitrite reductase kit, and reacting with enzyme extract (K) containing 20ml 3 [Fe(CN) 6 ](0.01 mol/L), EDTA (0.04 mmol/L) (Xu, N., liao, M., liang, Y., guo, J., zhang, Y., xie, X., fan, Q.and Zhu, Y.2021.biological nitrogen removal capability and pathways analysis of a novel low C/N ratio heterotrophic nitrifying and aerobic denitrifying bacterium (Bacillus thuringiensis strain WXN-23). Environ Res 195,110797), tris-HCl (10 mmol/L)) and HONH 3 Cl-N (15 mg/L), after 45min of reaction at 25℃HAO enzyme activity was reacted by hydroxylamine reduction; a reaction system (20 ml) of Nitrate Reductase (NR) contained an enzyme extract, tris-HCl (10 mmol/L), nitrate (20 mg/L), NADH (0.2 mmol/L), and the decrease in nitrate was used to evaluate the enzyme activity of NR.
TABLE 1 specific Activity of strains EN-F2
| Enzyme | Specific activity(U/mg) |
| AMO | 0.95±0.00 |
| HAO | 0.31±0.01 |
| NR | 0.42±0.13 |
| NIR | 1.54±0.02 |
As can be seen from Table 1, the AMO specific activity of EN-F2 is as high as 0.95U/mg protein, which is significantly higher than Pseudomonas taiwanensis J488 (0.65U/mg protein) (He, T., xie, D., ni, J., li, Z.and Li, Z.2020.Nitrous oxide produced directly from ammonium, nitrate and nitrite during nitrification and Density, journal of Hazardous Materials 388), bacillus thuringiensis WXN-23 (0.11U/mg protein) (Xu, N., liao, M., liang, Y., guo, J., zhang, Y., xie, X, fan, Q.and Zhu, Y.2021.biological nitrogen removal capability and pathways analysis of a novel low C/N ratio heterotrophic nitrifying and aerobic denitrifying bacterium (Bacillus thuringiensis strain WXN-23), environ Res 195,110797) and Paracoccus denitrificans ISTOD (0.07U/mg protein) (Medhi, K., singhal, A, chaun, D.K.82, U/mg protein) (I.334-B.5, K., X.E., X, fan, Q.and Zhu.2021.biological nitrogen removal capability and pathways analysis of a novel low C/N ratio heterotrophic nitrifying and aerobic denitrifying bacterium (U/mg protein). The AMO specific activity of EN-F2 was much higher than that of Glutamicibacter arilaitensis EM-H8 (0.07U/mg protein) (Chen, M., ding, C., he, T., zhang, M.and Wu, Q.2021. Effect hydroxylamine removal through heterotrophic nitrification by novel bacterium Glutamicibacter arilaitensis EM-H8.Chemosphere 288 (Pt 1), 132475), acinetobacter calcoaceticus HNR (0.05U/mg protein) (Zhao, B., he, Y.L., hughes, J.and Zhang, X.F.2010b.hetrotrophic nitrogen removal by a newly isolated Acinetobacter calcoaceticus HNR.bioresource 101 (14), 5194-5200) and Alcaligenes faecalisNR (0.02U/mg protein) (Joo, H.S., hirai, M.and Shelpha, M.2005. Charactrics of ammonium removal by heterotrophic nitrification-aerobic denitrification by Alcaligenes faecalis. 4.J Biosense (2-184)), and further demonstrated by the hydroxylamines removal of these strains in the F2. The NR (0.42U/mg protein) specific activity of the EN-F2 strain was slightly lower than Pseudomonas versutus LYM (0.47U/mg protein) (Shi, Z., zhang, Y., zhou, J., chen, M.and Wang, X.2013.biological removal of nitrate and ammonium under aerobic atmosphere by Paracoccus versutus LYM.biological Technology 148, 144-148.), but significantly higher than 0.02U/mg protein (Yang, L., wang, X.H., cui, S., ren, Y.X., yu, J., chen, N., xiao, Q., guo, L.K.and Wang, R.H.2019.Simulines removal of nitrogen and phosphorous by heterotrophic nitrification-aerobic denitrification of a metal resistant bacterium Pseudomonas putida strain NP5.biological Technology 285,121360 and P.dental ISD 1 were found in 0.02U/mg protein (Yang, L., wang, X.H., cui, S., ren, Y.X., yu, J., chen, N., xiao, Q., guo, L.K.and Wang, R.H.2019.Simulines removal of nitrogen and phosphorous by heterotrophic nitrification-aerobic denitrification of a metal resistant bacterium Pseudomonas putida strain NP5.biological Technology 285,121360 and P.dental ISD 1, i.s.2017.invest-ing the nitrification and denitrification kinetics under aerobic and anaerobic conditions by Paracoccus denitrificans ISTOD1.bioresource Technology 242, 334-343) furthermore, the NIR specific activity of EN-F2 is highest (1.54U/mg protein), much higher than 0.04U/mg protein (Zhao, b., he, y.l., huang, j., taylor, s.and Hughes, j.2010a.elevating nitrogen removal by Providencia rettgeri strain yl.j Ind Microbiol Biotechnol (6), 609-616), pseudomonas taiwanensis J488 (He, t., xie, d., ni, j., li, z.and Li, z.2020.nitrous oxide produced directly from ammonium, nitrate and nitrite during nitrification and density, journal ofHazardous Materials 388) and k.02U/mg protein of k.p.neumia CF-S9 (heat, etc.), respectively, N.K.2013. Charabacteria of heterotrophic nitrifying and aerobic denitrifying Klebsiella pneumoniae CF-S9 strain for bioremediation of Watersaid. International Biodegradation & Biodegradation 78,67-73). The results show that the four specific activities of the strain EN-F2 are higher than those of most reported aerobic denitrifying bacteria. NIR exhibits higher specific activity and lower nitrite removal capacity than AMO, HAO and NR, probably due to suboptimal culture or detection conditions of AMO, HAO and NR. In summary, the successful expression of all enzymes further suggests that strain EN-F2 is a heterotrophic nitrification-aerobic denitrification bacterium.
Furthermore, it should be understood that although the present disclosure describes embodiments, not every embodiment is provided with a separate embodiment, and that this description is provided for clarity only, and that the disclosure is not limited to the embodiments described in detail below, and that the embodiments described in the examples may be combined as appropriate to form other embodiments that will be apparent to those skilled in the art.
Sequence listing
<110> university of Guizhou
<120> a denitrifying bacterium and application thereof in treatment of nitrogenous wastewater
<160> 2
<170> SIPOSequenceListing 1.0
<210> 1
<211> 19
<212> DNA
<213> artificial seqnence
<220>
Claims (2)
1. A strain, characterized in that the strain is a strainPseudomonas taiwanensisEN-F2, the preservation number CCTCC NO: m20211515.
2. Use of the strain of claim 1 in the treatment of nitrogen-containing wastewater.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111561362.4A CN114369544B (en) | 2021-12-18 | 2021-12-18 | Denitrifying bacteria and application thereof in nitrogen-containing sewage treatment |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111561362.4A CN114369544B (en) | 2021-12-18 | 2021-12-18 | Denitrifying bacteria and application thereof in nitrogen-containing sewage treatment |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN114369544A CN114369544A (en) | 2022-04-19 |
| CN114369544B true CN114369544B (en) | 2023-07-21 |
Family
ID=81139304
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202111561362.4A Active CN114369544B (en) | 2021-12-18 | 2021-12-18 | Denitrifying bacteria and application thereof in nitrogen-containing sewage treatment |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN114369544B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115011522B (en) * | 2022-06-17 | 2023-05-12 | 贵州大学 | EN-J1 denitrifying bacteria and application thereof in nitrogen-containing sewage treatment |
Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104254611A (en) * | 2012-02-28 | 2014-12-31 | 马罗内生物创新公司 | Control of phytopathogenic microorganisms with pseudomonas sp. and substances and compositions derived therefrom |
| WO2015191682A1 (en) * | 2014-06-11 | 2015-12-17 | Academia Sinica | Composition and method for controlling insects and microorganisms using pseudomonas taiwanensis |
| WO2016162661A1 (en) * | 2015-04-09 | 2016-10-13 | University Of Essex Enterprises Limited | Bioremediation |
| KR20170045845A (en) * | 2015-10-20 | 2017-04-28 | 이상섭 | Oil-degradading mixed strain and oil degradation method using the same |
| CN107012111A (en) * | 2017-04-22 | 2017-08-04 | 陕西博秦生物工程有限公司 | One plant of Taiwan pseudomonad and its application |
| CN110004082A (en) * | 2019-04-01 | 2019-07-12 | 山东省科学院生态研究所(山东省科学院中日友好生物技术研究中心) | A kind of bacterium bacterial strain and application suitable for the regulation of industrialized agriculture nitrogen and phosphorus pollution |
| CN110438025A (en) * | 2019-05-24 | 2019-11-12 | 博瑞德环境集团股份有限公司 | A kind of denitrogenation bacterial strain and its application |
| CN110607264A (en) * | 2019-09-26 | 2019-12-24 | 中国科学院上海高等研究院 | Pseudomonas taiwanensis and application thereof |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US10801046B2 (en) * | 2016-07-25 | 2020-10-13 | Invista North America S.A.R.L. | Methods and materials for biosynthesizing multifunctional, multivariate molecules via carbon chain modification |
-
2021
- 2021-12-18 CN CN202111561362.4A patent/CN114369544B/en active Active
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104254611A (en) * | 2012-02-28 | 2014-12-31 | 马罗内生物创新公司 | Control of phytopathogenic microorganisms with pseudomonas sp. and substances and compositions derived therefrom |
| WO2015191682A1 (en) * | 2014-06-11 | 2015-12-17 | Academia Sinica | Composition and method for controlling insects and microorganisms using pseudomonas taiwanensis |
| WO2016162661A1 (en) * | 2015-04-09 | 2016-10-13 | University Of Essex Enterprises Limited | Bioremediation |
| KR20170045845A (en) * | 2015-10-20 | 2017-04-28 | 이상섭 | Oil-degradading mixed strain and oil degradation method using the same |
| CN107012111A (en) * | 2017-04-22 | 2017-08-04 | 陕西博秦生物工程有限公司 | One plant of Taiwan pseudomonad and its application |
| CN110004082A (en) * | 2019-04-01 | 2019-07-12 | 山东省科学院生态研究所(山东省科学院中日友好生物技术研究中心) | A kind of bacterium bacterial strain and application suitable for the regulation of industrialized agriculture nitrogen and phosphorus pollution |
| CN110438025A (en) * | 2019-05-24 | 2019-11-12 | 博瑞德环境集团股份有限公司 | A kind of denitrogenation bacterial strain and its application |
| CN110607264A (en) * | 2019-09-26 | 2019-12-24 | 中国科学院上海高等研究院 | Pseudomonas taiwanensis and application thereof |
Non-Patent Citations (3)
| Title |
|---|
| Ammonium and hydroxylamine can be preferentially remived during simultaneous nitrification and denitrification by pseudomonas taiwanensis EN-F2;Manman Zhang等;Bioresour Technol;全文 * |
| Hypothermia pesudomonas taiwanensis J488 exhibited strong tolerance capacity to high dosages of divalent metal ions during nitrogen removal process;Tengxia He等;Bioresour Technol;全文 * |
| 设施土壤中解磷-反硝化复合功能细菌筛选及其活性研究;张广智等;土壤通报;全文 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN114369544A (en) | 2022-04-19 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Deng et al. | Nitrogen removal by heterotrophic nitrifying and aerobic denitrifying bacterium Pseudomonas sp. DM02: Removal performance, mechanism and immobilized application for real aquaculture wastewater treatment | |
| Cui et al. | A novel halophilic Exiguobacterium mexicanum strain removes nitrogen from saline wastewater via heterotrophic nitrification and aerobic denitrification | |
| Yang et al. | Simultaneous removal of nitrogen and phosphorous by heterotrophic nitrification-aerobic denitrification of a metal resistant bacterium Pseudomonas putida strain NP5 | |
| Song et al. | A review of research progress of heterotrophic nitrification and aerobic denitrification microorganisms (HNADMs) | |
| Qiao et al. | Characteristics and metabolic pathway of the bacteria for heterotrophic nitrification and aerobic denitrification in aquatic ecosystems | |
| Yang et al. | Ammonium removal characteristics of an acid-resistant bacterium Acinetobacter sp. JR1 from pharmaceutical wastewater capable of heterotrophic nitrification-aerobic denitrification | |
| Yang et al. | Highly efficient nitrogen removal of a coldness-resistant and low nutrient needed bacterium, Janthinobacterium sp. M-11 | |
| Huang et al. | Characterization of novel Bacillus strain N31 from mariculture water capable of halophilic heterotrophic nitrification–aerobic denitrification | |
| Chen et al. | Biological nitrogen and phosphorus removal by a phosphorus-accumulating bacteria Acinetobacter sp. strain C-13 with the ability of heterotrophic nitrification–aerobic denitrification | |
| feng Su et al. | Enhancement of the denitrification in low C/N condition and its mechanism by a novel isolated Comamonas sp. YSF15 | |
| Liu et al. | Photobacterium sp. NNA4, an efficient hydroxylamine-transforming heterotrophic nitrifier/aerobic denitrifier | |
| Ke et al. | Characterization of Acinetobacter indicus ZJB20129 for heterotrophic nitrification and aerobic denitrification isolated from an urban sewage treatment plant | |
| White et al. | Conversion of cyanide to formate and ammonia by a pseudomonad obtained from industrial wastewater | |
| Xu et al. | Biological nitrogen removal capability and pathways analysis of a novel low C/N ratio heterotrophic nitrifying and aerobic denitrifying bacterium (Bacillus thuringiensis strain WXN-23) | |
| Xiujie et al. | Dominance of Candidatus saccharibacteria in SBRs achieving partial denitrification: effects of sludge acclimating methods on microbial communities and nitrite accumulation | |
| Zhang et al. | Ammonium and hydroxylamine can be preferentially removed during simultaneous nitrification and denitrification by Pseudomonas taiwanensis EN-F2 | |
| Ren et al. | Simultaneous nitrification and aerobic denitrification by a novel isolated Ochrobactrum anthropi HND19 | |
| Norton | Diversity and environmental distribution of ammonia‐oxidizing bacteria | |
| Wang et al. | Nitrate stimulation of N-Methylpyrrolidone biodegradation by Paracoccus pantotrophus: Metabolite mechanism and Genomic characterization | |
| Wei et al. | Biological nitrogen removal and metabolic characteristics of a novel cold-resistant heterotrophic nitrification and aerobic denitrification Rhizobium sp. WS7 | |
| Huang et al. | Removal of ammonium and nitrate by the hypothermia bacterium Pseudomonas putida Y-9 mainly through assimilation | |
| CN112625942B (en) | Aerobic denitrifying bacterium and application thereof | |
| Zhang et al. | Simultaneous ammonia and Cr (VI) removal by Pseudomonas aeruginosa LX in wastewater | |
| Yang et al. | The removal of ammonia nitrogen via heterotrophic assimilation by a novel Paracoccus sp. FDN-02 under anoxic condition | |
| CN114369544B (en) | Denitrifying bacteria and application thereof in nitrogen-containing sewage treatment |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |