CN109385388A - Thermophilic salt denitrifying bacterium YL5-2 and its application - Google Patents
Thermophilic salt denitrifying bacterium YL5-2 and its application Download PDFInfo
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Abstract
The invention discloses thermophilic salt denitrifying bacterium YL5-2 and its applications.One plant of thermophilic salt denitrifying bacterium YL5-2 is preserved in China General Microbiological culture presevation administrative center for the new species that Halovibrio belongs to, and deposit number is CGMCC NO.16315, and the deposit date is on August 20th, 2018.Bacterial strain YL5-2 is a kind of facultative or aerobic denitrifying bacteria, and salt tolerant range is 3%~32%, can be in 5%~25% range of salinity with NO3- N and NO2Denitrification is carried out as electron acceptor.YL5-2 can be used for the biological denitrificaion processing of high-salt wastewater of the salt content greater than 10%, reduce the denitrogenation processing cost of such waste water.
Description
Technical field
The present invention relates to environmentally friendly microorganism fields, more particularly to thermophilic salt denitrifying bacterium YL5-2 and its application.
Background technique
Polluted by nitrogen is the one of the major reasons for causing water eutrophication.The denitrogenation processing of waste water is for maintaining water environment matter
It measures and prevents water eutrophication from playing a significant role.The nitrogenous effluent of the industries such as petroleum, chemical industry, food processing, chemical fertilizer discharge
Have the characteristics that with high salt.Certain waste water that chemical industry, food processing generate, salt content are even more than seawater.At traditional biological method
Managing Low-salinity nitrogenous effluent has advantage, but when waste strength is excessively high, can inhibit the metabolism of denitrification microorganism.Salt tolerant and
The presence of the denitrifier of thermophilic salt provides inner theoretic possibility for high-salt wastewater biological denitrificaion.
Biological denitrificaion bacterium bag includes nitrifier and denitrifying bacterium.Biological denitrificaion bacterium under the conditions of high-salt wastewater can also be divided into nitre
Change bacterium and denitrifying bacterium.It can be the bacterium of gaseous nitrogen compound by nitrate or pressure nitrate reduction that denitrifying bacterium, which is a kind of,.
Therefore, screening separates and cultivates nitrifier and the denitrifying bacterium of salt tolerant and thermophilic salt from environment, and it is raw just to become solution high-salt wastewater
The key of object Denitrogenation.
Dalian Ocean University's focus et al. (the separation identification of the thermophilic salt denitrifying bacterium of mono- plant of moderate of such as focus, flower bud and its generation
It thanks characteristic research " aquatic science and technology information ", 2018,45 (3): 149~154) being separated from mariculture purification of waste water unit
To the thermophilic salt denitrifying bacterium of one plant of moderate, belong to Halomonas category, best metabolism growth condition is 30 DEG C of temperature, salinity 100g/
L, pH7.5~8.5, C/N ratio are 4:1.
University Of Qingdao Guo gorgeous equal (the separation identification of mono- plant of the such as Guo Yanli, Zhang Peiyu slight thermophilic salt denitrifying bacterium and spy
Property " application and environmental organism journal " 2010,16 (3): 394~398) separated from the mature activated sludge of processing high-salt wastewater
Obtain one plant of slight thermophilic salt aerobic denitrifying bacteria YL-1, belong to enlightening thatch Salmonella (Dietzia sp.), the bacterium can 0%~
10%, pH 7.5~8.5, utilize acetic acid, sucrose, glucose, sodium citrate, sodium succinate carry out denitrification.
Nanjing University of Technology slanders minor benefit and (slanders separation identification and its Denitrification Characteristics research of the thermophilic salt denitrifying bacteria of minor benefit
Nanjing University of Technology's master's thesis in 2013) by only nitrogen source of sodium nitrate and the training of heterotrophic denitrification that salinity is 8%
Support base, be enriched with, separate and screening has obtained 6 plants of thermophilic salt denitrifying bacteriums from the pedotheque in Yancheng saltern: NY-1, NY-11 with
NY-13 is branch bacillus, and (Virgibacillus sp.), NY-8 and NY-10 are Halomonas (Hal-monas
Sp.), NY-4 is marinobacter (Marinobacter sp.), and salinity growth scope is 0%~12%, the most suitable growth salt
Concentration is 3%~8%.Wherein the denitrifying capacity of NY-4 is most strong, using trisodium citrate as carbon source, salinity 8%, C/N 5,
When pH is 8, NO3The removal rate of-N is 95%.
The prior art is less about the report of thermophilic salt denitrifying bacterium, the anti-nitre of thermophilic salt under conditions of especially salinity > 10%
Change bacterium.It is the skill that such wastewater biological denitrificaion needs to solve that acquisition salt resistance ability, which is greater than 10% thermophilic salt denitrifying bacterium, from environment
Art problem.
Summary of the invention
Under the conditions of to solve the problems, such as the biological denitrificaion under high salt conditions, especially salinity greater than 10%
High-salt wastewater biological denitrificaion problem, a kind of thermophilic salt denitrifying bacterium is provided, can 10% or more salinity under conditions of with
NO3- N carries out denitrification as electron acceptor.
It is a further object of the present invention to provide the applications of the thermophilic salt denitrifying bacterium.
What the purpose of the present invention can be achieved through the following technical solutions:
Thermophilic salt denitrifying bacterium YL5-2 disclosed by the invention is preserved in China General Microbiological culture presevation administrative center,
Deposit number is CGMCC NO.16315, and the deposit date is on August 20th, 2018.Based on 16SrRNA Phylogenetic Analysis, genome
Sequencing, Fatty acid compositions, breathes the differences such as other quinones, Physiology and biochemistry and phenotypic characteristic at DNA hybridization test, can determine bacterial strain
YL5-2 is the new species that Halovibrio belongs to, and is named as Halovibrio salipaludis sp.nov.
The login of the GenBank/EMBL/DDBJ of the 16S rRNA sequence of thermophilic salt denitrifying bacterium YL5-2 disclosed by the invention
Number be MF782425, nucleotide sequence is as shown in SEQ ID NO.1；The GenBank/EMBL/DDBJ's of whole genome sequence steps on
Record number is NSKD00000000.1.
Thermophilic salt denitrifying bacterium YL5-2 disclosed by the invention is Gram-negative, facultative aerobic, direct rod shape or 0.5~0.8 μm
× 1.0~3.5 μm of small vibrios character, and by unipolarity flagellum movement, the bacterium colony on solid medium be it is smooth and
It is light yellow.
Halophilic vibrio Halovibrio sp.YL5-2 of the present invention can be in salinity 3%~32%, pH6.5~11.0, temperature
It is grown within the scope of 15~45 DEG C of degree；The most suitable growth salinity is 10%~25%, and the most suitable growth pH is 7.5~8.0, the most suitable growth
Temperature is 30~35 DEG C.
The main breathing quinone of halophilic vibrio Halovibrio sp.YL5-2 of the present invention is Q-9, and main fatty acid is C18:
1 ω 9c, C16:0, C19:0cyclo ω 8c and Summed Feature 8, main polar lipid are
Diphosphatidylglycerol (DPG), Phosphatidylethanolamine (PE), Phosphatidylglycerol
(PG), Phosphatidylcholine (PC).
Halophilic vibrio Halovibrio sp.YL5-2 of the present invention can use bromo- succinic acid, propionic acid and acetic acid as unique
Utilization of carbon source；But D-Maltose, D-Fructose, D- galactolipin, D- cellobiose, stachyose, D- melibiose, N- acetyl cannot be utilized
Base-D galactosamine, D- fucose, L- rhamnose, PEARLITOL 25C, D- galacturonic acid, D-Asp, D-Ser, the Portugal D-
Grape uronic acid, Pfansteihl, chinic acid, glactaric acid, D-malic acid, gamma-amino-butyric acid, formic acid, acetoacetate is as sole carbon source.
The utility model has the advantages that
Present invention finds the new species that a Halovibrio belongs to, and are named as Halovibrio salipaludis
sp.nov.The bacterial strain is a kind of facultative or aerobic denitrifying bacteria, and salt tolerant range is 3%~32%, can salinity 5%~
Multiple pollutant in 25% range in degrading waste water.Thermophilic salt denitrifying bacterium YL5-2 disclosed by the invention can be used for item with high salt
The degradation, conversion of pollutant and biological denitrification process under part, including high-salt wastewater processing, polluted seawater are administered, and salt-soda soil is repaired,
Nitrogen nutrition is consumed, algae excessive propagation, purifying water body, improvement substrate etc. are inhibited.
Detailed description of the invention:
Transmission electron microscope (TEM) photo (scale bar 2 of Fig. 1 halophilic vibrio Halovibrio sp.YL5-2 cell
μm)；
Fig. 2 is halophilic vibrio Halovibrio sp.YL5-2 (a) and Halovibrio denitrificans
The polar lipid map of DSM15503 (b), Halovibrio variabilis DSM 3050 (c).DPG:
diphosphatidylglycerol；PG:phosphatidylglycerol；AL:Aminolipid；PL:Phospholipid；
PE:Phosphatidylethanolamine；PGNL:Phosphoaminoglycolipid；F: the first dimension；S: the second dimension
Fig. 3 is halophilic vibrio Halovibrio sp.YL5-2 and Halovibrio denitrificans DSM15503 structure
The systematic growth tree graph based on 16S rRNA built.
Fig. 4 be halophilic vibrio Halovibrio sp.YL5-2 and Halovibrio variabilis DSM 3050T and its
The systematic growth tree graph of maximum parsimony method (MP) building of its strain based on 16S rRNA.
Fig. 5 is the systematic growth tree graph that halophilic vibrio Halovibrio sp.YL5-2 is constructed based on GGD matrix.
Biomaterial preservation information
YL5-2, classification naming is Halovibrio salipaludis, the deposit date is on August 20th, 2018, preservation list
Position is China General Microbiological culture presevation administrative center, and preservation address is Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3 China
Institute of microbiology, the academy of sciences, deposit number: CGMCC NO.16315.
Specific embodiment
The separation and preservation of the thermophilic salt denitrifying bacterium of embodiment 1
Thermophilic salt denitrifying bacterium YL5-2 is deposited from Qinghai Province Golmud Cha Er Han Salt Lake (36 ° of 51 ' N, 94 ° of 95 ' E)
It is isolated in soil.Cha Er Han Salt Lake lake water is saturation salinity or close saturation salinity throughout the year.
It configures NaCl concentration and is 20% LB liquid medium, and glycerol 250mg/L, glucose 250mg/L, methanol is added
50mg/L, to Cha Er Han Salt Lake sedimentary soil enrichment culture 48h under the conditions of 30 DEG C.Using YL solid medium to enrichment culture
Bacterial strain in liquid is separated.Contain following components in 1L culture medium: glucose: 0.6g, trisodium citrate 0.5g, glycerol 2mL,
Yeast extract 0.8g, peptone 1.6g, dipotassium hydrogen phosphate 0.35g, potassium dihydrogen phosphate 0.1g, ammonium sulfate 0.25g, ammonium chloride
0.25g, MgSO40.5g, CaCl20.1g, NaCl 180g；Microelement SL-4 10mL, pH 7.0-7.2；Agar 2.5%.
Bacterial strain YL5-2 is deposited in China General Microbiological culture presevation administrative center, and deposit number is CGMCC NO.16315,
The deposit date is on August 20th, 2018.
The analysis of the thermophilic salt denitrifying bacterium YL5-2 16S rRNA sequence of embodiment 2 and full gene sequencing and analysis
Bacterial strain YL5-2 extracting genome DNA uses TaKaRa kit (TaKaRa MiniBEST Bacteria
Genomic DNA Extraction 68Kit Ver.3.0)。
16S rRNA amplification uses universal bacterial primer 2 7F (5 '-AGAGTTTGATCMTGGCTCA G-3 ') and 1492R
(5'-TACGGYTACCTTGTTACGACTT-3').PCR sequencing commission Shanghai Sheng Gong Biotechnology Co., Ltd carries out.Bacterial strain
The complete 16S rRNA sequence of YL5-2 is 1518bp, and as shown in SEQ ID NO.1, GenBank accession number is MF782425.
The genome sequencing of bacterial strain YL5-2 is using Shanghai's style Sen Nuo Biotechnology Co., Ltd Illumina on Shanghai
2000 high-flux sequence platform of MiSeq.Raw sequencing data is filtered using PRINSEQ (version number v 0.20.4) software
And amendment, the base of genome is then carried out using SOAP denovo software (version number v1.05) software with default parameters
Pairing, then using the integrality of CheckM software (version 1.03) assessment genome.Protein coding open reading frame is adopted
It is predicted with Glimmer software (version number 1.2).RNA prediction uses RNAmmer software (version 1.2).Bacterial strain YL5-2 is complete
Totally 3,495,096bp, GenBank accession number is NSKD00000000.1 to genome sequence.
DNA-DNA cross experiment passes through bacterial strain YL5-2 and the immediate vibrio Halovibrio mode of its genetic development
Bacterial strain carries out.This method is equal to 1970 by De Ley to be proposed, DNA hybridization value (dDDH) uses the 2nd kind of mode (version of GGDC software
This number comparison one by one for 2.0) carrying out gene order obtains.DNA test and analysis the result shows that, bacterial strain YL5-2 and
Halovibrio variabilis DSM 3050T、Halovibrio denitrificans DSM15503TDNA-DNA it is miscellaneous
Friendship value is respectively 43.5% and 38.2%, far below 70% threshold value (species divide generally accepted threshold value).
Nucleotide average homogeneity value carries out 1000 repetition topology verifications using the base group of whole genome sequence and obtains.
This method is equal to 2007 by Goris and proposes, the software used is MUMmer (version number 3.23) and Jspecies (version number
1.2.1).Based on the ANI threshold range (95-96%) that Kim et al. and Richter et al. species proposed divide, to bacterial strain
Genome closely related therewith carries out ANI analysis (table 1) in the genome and GenBank of YL5-2.The result shows that bacterial strain
Average nucleotide identity (ANI) value highest of YL5-2 itself and Tamilnaduibacter alinus Mi 7 is 88.5%
(Supplementary Table S1), this is bacterial strain YL5-2TA kind of new species for belonging to Halovibrio category provide opinion
According to being named as Halovibrio salipaludis sp.nov.
1 bacterial strain YL5-2 of tableTAverage nucleotide identity (ANI) between genome closely related in GenBank and
DDH value.
The phenotypic characteristic and physiological and biochemical property of the thermophilic salt denitrifying bacterium YL5-2 of embodiment 3 is identified
Gram's staining characteristic is tested using BD Gram's staining kit.
Cell mobility is measured using half MA culture medium (0.5% agar, w/v).
Cellular morphology uses transmission electron microscope (TEM) analysis detection.I.e. picking is thin from the culture solution of exponential growth
Born of the same parents, with 0.5% uranyl acetate staining cell, and it is right at microscope (Tecnai Spirit, FEI, Hillsboro, OR, USA)
Cell is taken pictures.
Oxidase active uses oxidase reagent box (bioM é rieux), by by 3.0%H2O2Solution pours into bacterial clump
And it observes bubble and generates to measure catalase activity.
Temperature growth condition carries out on YL liquid agar medium, temperature is respectively 4,10,15,20,25,30,33,
37,40,45 and 50 DEG C, pH constant is 7.5, compares bacterial strain YL5-2 under different temperaturesTGrowth rate determine its optimum growh temperature
Degree.
YL agar and YL fluid nutrient medium of the salt resistance ability in 0.0-30.0%NaCl (w/v) carry out.Use buffer
(Na2HPO4/NaH2PO4(pH 5.0-7.0), Na2CO3/NaHCO3(pH 8.0-12.0)) pH is adjusted to 5.0,5.5,6.0,
7.0,8.0,9.0,10.0 and 11.0 (15.0%NaCl, 35 DEG C) are to measure the pH range for being suitble to growth.
Utilization of carbon source ability and enzymatic activity test use API 20NE, API ZYM (bioM é rieux) and Biolog
GENIII microwell plate.The cell of pregrown on all test inoculation YL culture mediums, and diluted with relevant inoculation medium.
The phenotypic characteristic and physiological and biochemical property qualification result of bacterial strain YL5-2 is as shown in table 2:
2 halophilic vibrio Halovibrio sp.YL5-2 (a) of table and Halovibrio denitrificans DSM15503T
(b), Halovibrio variabilis DSM 3050T(c) phenotype in terms of distinguishing characteristics compared with
Illustrate :+, it is positive；, negative.
Thermophilic salt denitrifying bacterium YL5-2 is that Gram-negative, alkalinity are aerobic, direct rod shape or 0.5-0.8x1.0-3.5 μm
Small vibrios shape, and pass through unipolarity flagellum movement (Fig. 1).
Bacterial strain YL5-2 is grown under aerobic condition using acetic acid, then grows (API using nitrate under anoxic conditions
20NE)。
The identification of 4 halophilic vibrio Halovibrio sp.YL5-2 cell fatty acid of embodiment
3 days YL5-2, Halovibrio is cultivated in cell fatty acid identification using 30 DEG C on YL culture medium
3050 cell of denitrificans DSM15503 and Halovibrio variabilis DSM.Key step are as follows: trained from YL
It supports and scrapes being saponified with 50% methanol containing sodium hydroxide for 100mg cell on base；Cell after saponification is freeze-dried,
Then use ratio is chloroform/methanol/0.3% (w/v) the NaCl extraction with aqueous solution cell fatty acid of 1:2:0.8 (v/v/v).
Cell fatty acid total amount is detected using phosphomolybdic acid method.
Fatty acid is qualitative and quantitative detection uses 6890N gas chromatograph (Agilent) and Sherlock microbial identification
The library standard MIS in system generate software (VERSION 6.0and Date 4, Microbial ID Inc., Newark,
DE, USA) (Sasser, 1990).
Strain idenfication based on cell fatty acid then uses German DSMZ correlation analysis tool.
Cell fatty acid qualification result is as shown in Figure 2.
Unique breathing quinone of YL5-2 is identical as Halovibrio variabilis DSM 3050, is all ubiquinone Q-9.
The main cell fatty acid of YL5-2 includes C18:1ω9c、C16:0、C19:0Cyclo ω 8c and Summed Feature 8
(table 3)；Main polar lipid is Diphosphatidylglycerol (DPG), Phosphatidylethanolamine
(PE), Phosphatidylglycerol (PG), Phosphatidylcholine (PC) and two kinds of unidentified lipids (L).This
It is a little similar to Halovibrio denitrificans to the relationship kind Halovibrio variabilis of YL5-2.
3 halophilic vibrio Halovibrio sp.YL5-2 (a) of table and Halovibrio denitrificans DSM
15503T(b), Halovibrio variabilis DSM 3050TCell fatty acid composition (%) compare.
Illustrate:*Summed features indicates two or three of the fatty acid that cannot be separated by GLC with MIDI system
Mixing.Summed feature 3 includes C16:1ω 7c and/or C16:1ω 6c, Summed feature 8 includes C18:1ω6c
And/or C18:1ω7c。
The fermented and cultured of 5 halophilic vibrio Halovibrio sp.YL5-2 of embodiment is tested
(1) medium component is glycerol 500mg/L, glucose 250mg/L, methanol 500mg/L, methylamine 200mg/L, chlorination
100~250g/L of sodium, sodium acetate 250mg/L, trisodium citrate 250mg/L, yeast powder 100mg/L, peptone 200mg/L, ox
Meat extract 200mg/L, microelement is a small amount of, and pH is 7.5~8.0.Fluid nutrient medium sterilizes, while controlling moisture evaporation.
(2) 48h is cultivated under the conditions of 35 DEG C after being inoculated in the triangular flask of 1L, the lost influence to keep the skin wet in incubation
The variation of salinity.Measured after 48h OD600 under 10%, 15%, 20%, 25% salt concentration conditions be respectively 1.72,1.65,
1.62,1.60,1.63.Switching culture twice is carried out, cultivates 48h after switching every time, OD600 is respectively after the completion of domestication culture
2.56、2.68、2.72、2.68、2.52。
(3) 1L culture solution is inoculated into the aerobic fermentation tank of 20L, and fermented and cultured is repeated, and medium component is kept not
Become.Still 48h is cultivated, cultivation temperature is 35 DEG C, mixing speed 100rpm, and dissolved oxygen is 2.0~4.0mg/L.48h post-fermentation
The OD600 of bacterium solution can reach 2.6~3.0 in tank.
(4) it examines: sampling observation daily using microscope in incubation, check whether there is miscellaneous bacteria and be mixed into growth；Simultaneously
Observe the growth and metamorphosis situation of YL5-2.
(5) result: test result shows that bacterial strain YL5-2 can be rapidly performed by fermented and cultured and amplification, this shows YL5-
2 potentiality with large-scale engineering applications.
The salt resistance ability of 6 halophilic vibrio Halovibrio sp.YL5-2 of embodiment is tested
(1) medium component is glycerol 50mg/L, glucose 25mg/L, methanol 50mg/L, methylamine 20mg/L, sodium chloride
250g/L, sodium acetate 25mg/L, trisodium citrate 25mg/L, yeast powder 10mg/L, peptone 20mg/L, beef extract 20mg/L,
Agar 20g/L.
(2) fresh above-mentioned culture medium activated spawn is used, is enriched to lawn growth within culture 3 days spare
(3) salinity gradient is arranged: according to the enrichment isolation condition of YL5-2 strain, the salinity gradient difference of culture medium is arranged
For 0%, 0.5%, 1%, 2%, 3%, 5%, 8%, 10%, 12%, 15%, 18%, 21%, 24%, 26%, 28%, 30%,
32%, 34%.
(4) it prepares solid medium: culture medium, the higher culture medium hot water of salinity is prepared according to the culture medium prescription of setting
It sterilizes after thawing, every bottle of evaporation water for adding 5ml is added after volatile medium component is subject to sterilization and shakes up, is cooled to 60
DEG C or so, plate processed, salinity height easily solidifies, and therefore, plate processed quickly will fast (32% salinity medium plate
When, have after cooled and solidified and salt out on a small quantity, 34% salinity has a large amount of salt crystals to be precipitated).
(5) inoculated and cultured: the fresh lawn of one ring of picking accesses each salinity medium under aseptic condition, from Low-salinity toward with high salt
Gradient switching is spent, as scribing line track there are a large amount of salt crystals to be precipitated after the scribing line of 34% salinity medium, inoculation finishes culture dish use
Sealed membrane sealing, 35-37 DEG C culture 3-7 days, observation growth situation.
(6) test result: YL5-2 strain salt tolerant range is 3-32%.YL5-2 bacterial strain is three in 3-30% salinity medium
Can obviously observe the lawn newly grown in it, but grow 7 days in 32% saturation salinity medium or more can just grow
Visually visible lawn.Show that YL5-2 speed of growth in 3%~30% salt concentration range is very fast.
Growing state of 4 YL5-2 of table on different salinity culture mediums
The salt tolerant denitrifying capacity of the thermophilic salt denitrifying bacterium YL5-2 of embodiment 7 is tested
(1) culture medium: acetic acid 2000mg/L, peptone 20mg/L, beef extract 20mg/L, NO3- N is 100mg/L, NaCl
Concentration 3%~30%, buffer, which is added, makes pH 7.5~8.0.
(2) experimental design: carrying out in the triangular flask of 10 500mL of denitrification test, each that culture medium 300mL, setting 8 is added
A salinity gradient, salt content is respectively 3%, 6%, 10%, 12%, 15%, 20%, 25%, 30%, wherein salt-free blank
Group is also provided with 2 in parallel.
(3) denitrification is tested: being inoculated with YL-5 culture solution about 10mL after all samples sterilizing, is trained on constant-temperature table
It supports, temperature is 30~35 DEG C；Shaking speed is respectively 10ppm；It is measured by sampling in triangular flask respectively at for 24 hours, after 48h and 72h
NO3The concentration of-N.Test result such as following table data:
5 YL5-2 of table denitrification under different salinity removes NO3The test result (unit: mg/L) of-N
| 0 | 0 | 3% | 6% | 10% | 12% | 15% | 20% | 25% | 30% | |
| 24h | 99.6 | 99.5 | 91.3 | 76.2 | 68.8 | 67.5 | 66.4 | 67.6 | 72.6 | 89.6 |
| 48h | 99.1 | 99.2 | 55.4 | 18.6 | 14.4 | 11.7 | 9.9 | 14.7 | 19.1 | 40.6 |
| 72h | 98.5 | 98.8 | 13.5 | 6.7 | 4.5 | 3.6 | 3.8 | 4.2 | 8.2 | 12.5 |
The salt tolerant denitrifying capacity of the thermophilic salt denitrifying bacterium YL5-2 of embodiment 8 is tested
(1) culture medium: acetic acid 2000mg/L, peptone 20mg/L, beef extract 20mg/L, NO2- N is 100mg/L, NaCl
Concentration 3%~30%, buffer, which is added, makes pH 7.5~8.0.
(2) experimental design: carrying out in the triangular flask of 10 500mL of denitrification test, each that culture medium 300mL, setting 8 is added
A salinity gradient, salt content is respectively 3%, 6%, 10%, 12%, 15%, 20%, 25%, 30%, wherein salt-free blank
Group is also provided with 2 in parallel.
(3) denitrification is tested: being inoculated with YL-5 culture solution about 10mL after all samples sterilizing, is trained on constant-temperature table
It supports, temperature is 30~35 DEG C；Shaking speed is respectively 10ppm；It is measured by sampling in triangular flask respectively at for 24 hours, after 48h and 72h
NO2The concentration of-N.Test result such as following table data:
6 YL5-2 of table denitrification under different salinity removes NO2The test result (unit: mg/L) of-N
| 0 | 0 | 3% | 6% | 10% | 12% | 15% | 20% | 25% | 30% | |
| 24h | 99.7 | 99.7 | 95.3 | 86.2 | 78.8 | 77.5 | 76.4 | 77.6 | 81.6 | 91.6 |
| 48h | 99.3 | 99.1 | 62.4 | 20.3 | 16.5 | 14.8 | 11.3 | 15.7 | 19.1 | 30.6 |
| 72h | 99.0 | 98.6 | 15.8 | 9.7 | 6.6 | 4.2 | 3.8 | 7.3 | 8.5 | 15.3 |
Sequence table
<110>Co., Ltd, Lianyungang Design and Research Institute (Lanai Engineering Co.)
<120>thermophilic salt denitrifying bacterium YL5-2 and its application
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1431
<212> DNA
<213>Halovibrio belongs to (Halovibrio sp.)
<400> 1
cccatggggg cagctacaca tgcagtcgag cggcagcagc tccttcggga ggctggcgag 60
cggcggacgg gtgagtaacg catgggaact tacccagtag tgggggatag cccggggaaa 120
cccggattaa taccgcatac gccctgaggg ggaaagcggg ctccggctcg cgctattgga 180
tgggcccatg tcggattagt tagttggtgg ggtaatggcc taccaaggcg acgatccgta 240
gctggtctga gaggatgatc agccacaccg ggactgagac acggcccgga ctcctacggg 300
aggcagcagt ggggaatatt ggacaatggg ggcaaccctg atccagccat gccgcgtgtg 360
tgaagaaggc cttagggttg taaagcactt tcagcaggga ggaaaagctg atcgttaata 420
ccggtcagtg ttgacgttac ctgcagaaga agcaccggct aactccgtgc cagcagccgc 480
ggtaatacgg agggtgcaag cgttaatcgg aattactggg cgtaaagggc gcgtaggcgg 540
tttggtaagc gagttgtgaa agccccgggc tcaacctggg aatggcaatt cgaactgcca 600
agctagaatg cagcagaggg cagtggaatt ccaggtgtag cggtgaaatg cgtagatatc 660
tggaggaaca ccagtggcga aggcgactgc ctgggctgac actgacgctg aggtgcgaaa 720
gcgtgggtag caaacaggat tagataccct ggtagtccac gctgtaaacg ctgagaacta 780
gtcgttgggg ctattagagc cttagtgacg cagctaacgc gataagttct ccgcctgggg 840
agtacggccg caaggttaaa actcaaatga attgacgggg gcccgcacaa gcggtggagc 900
atgtggttta attcgacgca acgcgaagaa ccttacctgg tcttgacatc ctgcgaactt 960
ggtagagata ccttggtgcc ttcgggagcg cagtgacagg tgctgcatgg ccgtcgtcag 1020
ctcgtgtcgt gagatgttgg gttaagtccc gtaacgagcg caacccttgt ccttagttgc 1080
cagcggtccg gccgggaact ctagggagac tgccggtgac aaaccggagg aaggtgggga 1140
tgacgtcagg tcatcatggc ccttacggcc agggctacac acgtgctaca atggggcgca 1200
cagagggcag caagcgcgcg agtgcaagcg aatcccttaa aacgcctcgt agtccggatc 1260
ggagtctgca actcgactcc gtgaagtcgg aatcgctagt aatcgcagat cagaatgctg 1320
cggtgaatac gttcccgggc cttgtacaca ccgcccgtca caccatggga gtggactgca 1380
ccagaagcgg ttagtctaac cttcgggagg acgatcgcca cggtgtctgt a 1431
Claims (8)
1. one plant of thermophilic salt denitrifying bacterium YL5-2 is preserved in China General Microbiological strain guarantor for the new species that Halovibrio belongs to
Administrative center is hidden, deposit number is CGMCC NO.16315, and the deposit date is on August 20th, 2018.
2. application of the thermophilic salt denitrifying bacterium YL5-2 described in claim 1 in the high-salt wastewater processing of salinity 3%~32%.
3. application according to claim 2, it is characterised in that thermophilic salt denitrifying bacterium YL5-2 described in claim 1 is in salt
Application in the biological denitrificaion processing of the high-salt wastewater of degree 3%~32%.
4. application according to claim 3, it is characterised in that the thermophilic salt denitrifying bacterium YL5-2 under anoxic conditions or
Denitrification is carried out to high-salt wastewater under aerobic condition.
5. application according to claim 4, it is characterised in that the thermophilic salt denitrifying bacterium YL5-2 under anoxic conditions or
High-salt wastewater under aerobic condition to salinity greater than 10% carries out denitrification.
6. application of the thermophilic salt denitrifying bacterium YL5-2 described in claim 1 in polluted seawater is administered, salt-soda soil is repaired.
7. thermophilic salt denitrifying bacterium YL5-2 described in claim 1 inhibits algae excessive propagation, purified water in consumption nitrogen nutrition
Application in body.
8. application of the thermophilic salt denitrifying bacterium YL5-2 described in claim 1 in improvement sediment.
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| WO2020134728A1 (en) * | 2018-12-29 | 2020-07-02 | 中蓝连海设计研究院有限公司 | Halophilic denitrifying bacteria yl5-2 and application thereof |
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| WO2020134728A1 (en) * | 2018-12-29 | 2020-07-02 | 中蓝连海设计研究院有限公司 | Halophilic denitrifying bacteria yl5-2 and application thereof |
| CN109928505A (en) * | 2019-03-15 | 2019-06-25 | 江苏大学 | A method of pickling waste water is handled using salt algae, Halophiles |
| CN115181694A (en) * | 2022-06-23 | 2022-10-14 | 北京工业大学 | A moderately halophilic bacterium with high salinity wastewater assimilation and denitrification function and its application |
| CN115181694B (en) * | 2022-06-23 | 2024-03-22 | 北京工业大学 | Moderately halophilic bacteria with high-salinity wastewater assimilation denitrification function and application thereof |
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