CN106834182A - One plant of secondary meningitidis strains apt to change and its application - Google Patents
One plant of secondary meningitidis strains apt to change and its application Download PDFInfo
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- CN106834182A CN106834182A CN201710107505.1A CN201710107505A CN106834182A CN 106834182 A CN106834182 A CN 106834182A CN 201710107505 A CN201710107505 A CN 201710107505A CN 106834182 A CN106834182 A CN 106834182A
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- 230000008859 change Effects 0.000 title claims abstract description 41
- RWSOTUBLDIXVET-UHFFFAOYSA-N Dihydrogen sulfide Chemical compound S RWSOTUBLDIXVET-UHFFFAOYSA-N 0.000 claims abstract description 30
- 229910000037 hydrogen sulfide Inorganic materials 0.000 claims abstract description 30
- 230000001580 bacterial effect Effects 0.000 claims abstract description 24
- 241000894006 Bacteria Species 0.000 claims abstract description 22
- 238000006477 desulfuration reaction Methods 0.000 claims abstract description 22
- 229910002651 NO3 Inorganic materials 0.000 claims abstract description 19
- NHNBFGGVMKEFGY-UHFFFAOYSA-N Nitrate Chemical compound [O-][N+]([O-])=O NHNBFGGVMKEFGY-UHFFFAOYSA-N 0.000 claims abstract description 19
- 230000023556 desulfurization Effects 0.000 claims abstract description 19
- MWUXSHHQAYIFBG-UHFFFAOYSA-N Nitric oxide Chemical compound O=[N] MWUXSHHQAYIFBG-UHFFFAOYSA-N 0.000 claims abstract description 12
- 241001478240 Coccus Species 0.000 claims abstract description 10
- 241001478304 Paracoccus versutus Species 0.000 claims abstract description 6
- KRKNYBCHXYNGOX-UHFFFAOYSA-K Citrate Chemical compound [O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O KRKNYBCHXYNGOX-UHFFFAOYSA-K 0.000 claims abstract description 4
- IOVCWXUNBOPUCH-UHFFFAOYSA-M Nitrite anion Chemical compound [O-]N=O IOVCWXUNBOPUCH-UHFFFAOYSA-M 0.000 claims abstract description 4
- 230000009604 anaerobic growth Effects 0.000 claims abstract description 4
- 150000003839 salts Chemical class 0.000 claims abstract description 4
- 238000010186 staining Methods 0.000 claims abstract 2
- FGIUAXJPYTZDNR-UHFFFAOYSA-N potassium nitrate Chemical compound [K+].[O-][N+]([O-])=O FGIUAXJPYTZDNR-UHFFFAOYSA-N 0.000 claims description 26
- 230000001954 sterilising effect Effects 0.000 claims description 21
- 238000004659 sterilization and disinfection Methods 0.000 claims description 21
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 claims description 20
- 239000002609 medium Substances 0.000 claims description 19
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 claims description 14
- 239000007836 KH2PO4 Substances 0.000 claims description 13
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 13
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 claims description 13
- 229910000030 sodium bicarbonate Inorganic materials 0.000 claims description 13
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 13
- 238000000855 fermentation Methods 0.000 claims description 12
- 230000004913 activation Effects 0.000 claims description 10
- AKHNMLFCWUSKQB-UHFFFAOYSA-L sodium thiosulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=S AKHNMLFCWUSKQB-UHFFFAOYSA-L 0.000 claims description 10
- 230000004151 fermentation Effects 0.000 claims description 9
- 239000007789 gas Substances 0.000 claims description 8
- 239000001963 growth medium Substances 0.000 claims description 8
- 239000007788 liquid Substances 0.000 claims description 8
- 229910001629 magnesium chloride Inorganic materials 0.000 claims description 7
- 229910052603 melanterite Inorganic materials 0.000 claims description 7
- 239000007787 solid Substances 0.000 claims description 6
- 229920001817 Agar Polymers 0.000 claims description 5
- 239000008272 agar Substances 0.000 claims description 5
- 239000012533 medium component Substances 0.000 claims description 4
- 239000012153 distilled water Substances 0.000 claims description 3
- 230000003068 static effect Effects 0.000 claims description 3
- 239000012092 media component Substances 0.000 claims description 2
- 235000015097 nutrients Nutrition 0.000 claims description 2
- VNWKTOKETHGBQD-UHFFFAOYSA-N methane Chemical compound C VNWKTOKETHGBQD-UHFFFAOYSA-N 0.000 abstract description 15
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 abstract description 11
- 239000001301 oxygen Substances 0.000 abstract description 11
- 229910052760 oxygen Inorganic materials 0.000 abstract description 11
- 230000012010 growth Effects 0.000 abstract description 8
- 230000003647 oxidation Effects 0.000 abstract description 5
- 238000007254 oxidation reaction Methods 0.000 abstract description 5
- 238000009395 breeding Methods 0.000 abstract description 2
- 230000001488 breeding effect Effects 0.000 abstract description 2
- 239000000370 acceptor Substances 0.000 description 16
- 244000005700 microbiome Species 0.000 description 10
- 239000000243 solution Substances 0.000 description 9
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 8
- 238000000034 method Methods 0.000 description 7
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 6
- 239000010865 sewage Substances 0.000 description 6
- 230000003321 amplification Effects 0.000 description 5
- 238000003199 nucleic acid amplification method Methods 0.000 description 5
- 239000010802 sludge Substances 0.000 description 5
- 229910052717 sulfur Inorganic materials 0.000 description 5
- 239000011593 sulfur Substances 0.000 description 5
- 238000010790 dilution Methods 0.000 description 4
- 239000012895 dilution Substances 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 238000005259 measurement Methods 0.000 description 4
- 238000004321 preservation Methods 0.000 description 4
- 241001052560 Thallis Species 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 3
- 150000001875 compounds Chemical class 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 230000007613 environmental effect Effects 0.000 description 3
- 238000011081 inoculation Methods 0.000 description 3
- 238000002955 isolation Methods 0.000 description 3
- 244000144972 livestock Species 0.000 description 3
- 229910052757 nitrogen Inorganic materials 0.000 description 3
- 238000007747 plating Methods 0.000 description 3
- 244000144977 poultry Species 0.000 description 3
- 239000002994 raw material Substances 0.000 description 3
- 108020004465 16S ribosomal RNA Proteins 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-L Sulfate Chemical compound [O-]S([O-])(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-L 0.000 description 2
- 239000005864 Sulphur Substances 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 238000001311 chemical methods and process Methods 0.000 description 2
- 230000005611 electricity Effects 0.000 description 2
- 210000003608 fece Anatomy 0.000 description 2
- 231100000518 lethal Toxicity 0.000 description 2
- 230000001665 lethal effect Effects 0.000 description 2
- 230000035479 physiological effects, processes and functions Effects 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 241001057811 Paracoccus <mealybug> Species 0.000 description 1
- UCKMPCXJQFINFW-UHFFFAOYSA-N Sulphide Chemical compound [S-2] UCKMPCXJQFINFW-UHFFFAOYSA-N 0.000 description 1
- 239000002250 absorbent Substances 0.000 description 1
- 230000002745 absorbent Effects 0.000 description 1
- 230000007059 acute toxicity Effects 0.000 description 1
- 231100000403 acute toxicity Toxicity 0.000 description 1
- 239000003463 adsorbent Substances 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 230000001651 autotrophic effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000011953 bioanalysis Methods 0.000 description 1
- 239000011942 biocatalyst Substances 0.000 description 1
- 239000002551 biofuel Substances 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 238000002485 combustion reaction Methods 0.000 description 1
- 230000007797 corrosion Effects 0.000 description 1
- 238000005260 corrosion Methods 0.000 description 1
- TXKMVPPZCYKFAC-UHFFFAOYSA-N disulfur monoxide Inorganic materials O=S=S TXKMVPPZCYKFAC-UHFFFAOYSA-N 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000005265 energy consumption Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 239000000499 gel Substances 0.000 description 1
- 239000011261 inert gas Substances 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 239000003958 nerve gas Substances 0.000 description 1
- 239000005416 organic matter Substances 0.000 description 1
- 238000006213 oxygenation reaction Methods 0.000 description 1
- 230000000243 photosynthetic effect Effects 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 230000005855 radiation Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- XTQHKBHJIVJGKJ-UHFFFAOYSA-N sulfur monoxide Chemical compound S=O XTQHKBHJIVJGKJ-UHFFFAOYSA-N 0.000 description 1
- 230000009182 swimming Effects 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
- 239000002351 wastewater Substances 0.000 description 1
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- 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
- C12N1/205—Bacterial isolates
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
-
- C—CHEMISTRY; METALLURGY
- C10—PETROLEUM, GAS OR COKE INDUSTRIES; TECHNICAL GASES CONTAINING CARBON MONOXIDE; FUELS; LUBRICANTS; PEAT
- C10L—FUELS NOT OTHERWISE PROVIDED FOR; NATURAL GAS; SYNTHETIC NATURAL GAS OBTAINED BY PROCESSES NOT COVERED BY SUBCLASSES C10G, C10K; LIQUEFIED PETROLEUM GAS; ADDING MATERIALS TO FUELS OR FIRES TO REDUCE SMOKE OR UNDESIRABLE DEPOSITS OR TO FACILITATE SOOT REMOVAL; FIRELIGHTERS
- C10L3/00—Gaseous fuels; Natural gas; Synthetic natural gas obtained by processes not covered by subclass C10G, C10K; Liquefied petroleum gas
- C10L3/06—Natural gas; Synthetic natural gas obtained by processes not covered by C10G, C10K3/02 or C10K3/04
- C10L3/10—Working-up natural gas or synthetic natural gas
- C10L3/101—Removal of contaminants
- C10L3/102—Removal of contaminants of acid contaminants
- C10L3/103—Sulfur containing contaminants
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- 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
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- 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
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/30—Fuel from waste, e.g. synthetic alcohol or diesel
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- Tropical Medicine & Parasitology (AREA)
- Medicinal Chemistry (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Purification Treatments By Anaerobic Or Anaerobic And Aerobic Bacteria Or Animals (AREA)
Abstract
The invention discloses one plant of secondary coccus (Paracoccus versutus) 2#CGMCC No.13321 apt to change and its under anoxic conditions by the use of nitrate as electron acceptor oxidation removal hydrogen sulfide in methane (H2S application).The bacterial strain Gram's staining is feminine gender, and strain cell rod-short is smaller；Bacteria colony white is raised, and neat in edge is smooth, surface wettability；Bacterial strain not salt tolerant, it is impossible to utilize citrate；In the presence of nitrate, nitrite or nitrogen oxide, anaerobic growth can be sought with them as electron acceptor.After secondary meningitidis strains CGMCC No.13321 apt to change are seeded into desulfurization reactor, can be in the reactor using nitrate as electron acceptor to H2S carries out biological oxidation, and carries out growth and breeding, reaches H in biogas2The purpose of S removings, effectively prevent tradition carries out biogas H using oxygen as electron acceptor2The potential safety hazard that methane is brought with oxygen mix is introduced when S is removed, to H in biogas2The clearance of S is up to 93.8%.
Description
Technical field
The present invention relates to a Paracoccus bacterial strain, more particularly to one kind can under anoxic conditions by the use of nitrate as electricity
Sub- acceptor oxidation removal hydrogen sulfide in methane (H2S apt to change secondary meningitidis strains).
Background technology
Biogas is a kind of emerging clean reproducible energy, and the anaerobic fermentation technology with biogas as target product is good because of its
The energy, environmental benefit be increasingly widely applied, will be to alleviating China energy crisis, environmental protection have important meaning
Justice.Biogas is a kind of bio-fuel that a kind of biological raw material is produced through anaerobic organism microbe conversion, for biogas anaerobic fermentation
Biological raw material mainly includes feces of livestock and poultry, stalk, energy-source plant, house refuse, organic wastewater the like waste.At present, China natural pond
Gas fermentation primary raw material be livestock and poultry farm excrement and sewage, due in feces of livestock and poultry and sewage containing a large amount of protein and other
Sulfur-containing compound, is converted through anaerobic fermentation, and H is contained in the biogas of generation2S gas ingredients.H2S is a kind of very strong change of corrosivity
Compound, heavy corrosion pipe-line equipment, H2S also has extremely strong acute toxicity, is a kind of nerve gas, H2S contents 0.6mg/L
When can make one lethal in 0.5-1h, content can make one lethal immediately in 1.2-2.8mg/L.Meanwhile, it is mixed with H2The biogas combustion of S
After burning, H2S can be converted into sulfur oxide and be discharged into air, cause atmosphere pollution.Therefore, in order to reach safe utilization biogas
The target of the energy and environmental protection, it is necessary to desulfurization process are being carried out to biogas using preceding.
At present, biogas removing H2The method of S mainly includes physico-chemical process and bioanalysis.Physico-chemical process removes H2S needs substantial amounts ofization
Medicament and energy consumption higher are learned, also adsorbent or absorbent is disposed, it is costly, there is secondary pollution.In biology
In doctor treatment, H is aoxidized with high-performance bio2The functional microorganism strain of S is the key that can desulfurization be realized.According to microorganism
Metabolic type, can be by H2S (or sulfide) is converted into elemental sulfur or the microorganism of sulfate can be divided into two major classes：1. luminous energy from
The type of supporting sulfur bacteria；2. chemosynthetic autotroph colorless sulfur bacteria.Photosynthetic autotrophs microorganism is in removal H2During S, though have higher
H2The advantages of under S clearances and anaerobic condition without potential safety hazard, but substantial amounts of radiation energy is needed, its financial cost is higher, and
Easily limited by light source.Therefore chemosynthetic autotroph microorganism turns into biological removal biogas H2, there is bar aerobic in the preferable selection of S
Under part, chemautotrophy organisms use oxygen is used as electron acceptor, but the regulation and control of oxygen are relatively difficult, exists when oxygen supply is excessive quick-fried
Fried hidden danger, while can introduce other inert gases such as nitrogen while oxygen supply, reduces the concentration of methane in sewage gas, reduces natural pond
The energy efficiency of gas.And under anoxic conditions, using other Alternative electronic acceptors such as nitrate beyond oxygen, be capable of achieving gentle
Under the conditions of H2S is removed.Therefore, under anoxic conditions, using nitrate as electron acceptor, oxygen is substituted as electron acceptor
Methane bio-desulfurization research turns into focus, can be prevented effectively from oxygen and the biogas potential safety hazard that causes of mixing and oxygenation and introduce
Reduction of the nitrogen to biogas energy efficiency, for China's biogas desulfurization provides a new technological approaches.
It is efficient functional microorganism bacterial strain using the core that nitrate carries out methane bio-desulfurization as electron acceptor, therefore,
Can the acquisition of efficient obligate function stem is directly connected to desulphurization system successfully construct, by the efficient obligate function of separation screening
Bacterial strain is inoculated in desulfurization reactor, can be built with obligate function stem as biocatalyst, nitrate is as electron acceptor
Methane bio-desulfurization system.
The content of the invention
H in can removing biogas as electron acceptor using nitrate it is an object of the invention to provide one kind2The secondary coccus bacterium of S
Strain, the bacterial strain in desulfurization reactor and can have a good desulfurized effect as desulfurization microbial inoculant.
To achieve the above object, the present invention provides secondary coccus (Paracoccus versutus) 2#CGMCC apt to change
No.13321, the bacterial strain is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, preservation address：Beijing
The institute 3 of city Chaoyang District North Star West Road 1, Institute of Microorganism, Academia Sinica, postcode 100101；Preservation date in November, 2016
18 days, deposit number CGMCC No.13321.
Secondary meningitidis strains CGMCC No.13321 apt to change pick up from Sichuan sewage treatment plant anaerobic sludge.The sludge that will be gathered
Tamed with the enriched medium of sulfur-containing compound under anaerobic, gradient dilution is carried out after 7d, from 10-2、10-4、10-6、
10-8、10-10Draw 0.1ml under dilution factor to be coated with isolation medium flat board, 30 DEG C of anaerobism are incubated, obtained after separating for several times
The bacterial strain for obtaining.Wherein enriched medium：Na2S2O3·5H2O 5g、KNO3 4g、KH2PO4 2g、NaHCO3 1g、MgCl2·
6H2O 0.5g、FeSO4·7H2O 0.01g, pH value 7.0, running water 1000mL.121 DEG C, the treatment of 30min high-temperature sterilizations.Separate
Culture medium：Na2S2O3·5H2O 10g、KNO3 4g、KH2PO4 2g、NaHCO3 1g、MgCl2·6H2O 0.5g、FeSO4·7H2O
0.01g, agar 20g, running water 1000mL, pH7.0-7.5.121 DEG C, the treatment of 30min high-temperature sterilizations.
Bacterial strain of the invention is examined under a microscope, strain cell rod-short, smaller；Bacteria colony white is raised, neat in edge
Smooth, surface wettability, plating medium colonial morphology is shown in Fig. 1, and the thalli morphology that microscope amplifies 1000 times is shown in Fig. 2.Through physiology
Biochemical identification is Gram-negative bacteria.Bacterial strain not salt tolerant, it is impossible to utilize citrate；When nitrate, nitrite or nitrogen oxide
In the presence of, anaerobic growth can be sought with them as electron acceptor.Bacterial strain can be in 15-45 DEG C of range growth, optimum growth temperature 30-35
℃；Can be grown in the range of pH7.0-7.5, the most suitable growth pH7.0.
The solid activation medium component of secondary meningitidis strains CGMCC No.13321 apt to change is：Na2S2O3·5H2O 10g、
KNO3 4g、KH2PO4 2g、NaHCO3 1g、MgCl2·6H2O 0.5g, FeSO4·7H2O 0.01g, agar 20g, distilled water
1000mL, pH7.0-7.5,121 DEG C, the treatment of 30min high-temperature sterilizations.
The liquid fermentation medium component of secondary meningitidis strains CGMCC No.13321 apt to change is：Na2S2O3·5H2O 10g、
KNO3 4g、KH2PO4 2g、NaHCO31g, running water 1000ml, pH7.0-7.5,121 DEG C, the treatment of 30min high-temperature sterilizations.
The desulfurization nutrient media components of secondary meningitidis strains CGMCC No.13321 apt to change is：KNO3 4g、KH2PO4 1g、
NaHCO31g, running water 1000ml, pH7.0-7.5,121 DEG C, the treatment of 30min high-temperature sterilizations.
It is de- in biogas by electron acceptor of nitrate present invention also offers secondary meningitidis strains CGMCC No.13321 apt to change
Application in sulphur.Activated culture is inoculated into the bacterium solution of the apt to change secondary meningitidis strains CGMCC No.13321 of Amplification Culture and is contained
In the desulfurization reactor of nitrate, biogas is continuously passed through, the H in biogas can be significantly reduced2S concentration.
The know-why that secondary meningitidis strains CGMCC No.13321 apt to change are applied to biogas desulfurization is secondary coccus bacterium apt to change
After strain CGMCC No.13321 are seeded to desulfurization reactor, can be in the reactor using nitrate as electron acceptor to H2S enters
Row biological oxidation, and growth and breeding is carried out, reach H in biogas2The purpose of S removings.
The application method of secondary meningitidis strains CGMCC No.13321 apt to change in the present invention：
(1) activated with solid activation medium using preceding；
(2) liquid fermentation medium for crossing sterilization treatment is quantitative standby；
(3) strain after activation is seeded to the liquid fermentation medium culture that sterilization treatment is crossed, static gas wave refrigerator 1d- respectively
3d；
(4) bacterium solution is inoculated in the reactor of the desulfurization culture medium crossed equipped with sterilization treatment in 10% ratio；
(5) biogas is passed through, beginning is passed through with low discharge, and flow gradually increases later, proceeds by biogas H2At S removings
Reason.
Compared with prior art, the beneficial effects of the invention are as follows：
(1) there is provided new biogas desulfurization bacterial strain：Secondary meningitidis strains CGMCC No.13321 apt to change；
(2) secondary meningitidis strains CGMCC No.13321 apt to change are screened from sewage treatment plant of Sichuan Province anaerobic sludge
Arrive, surrounding environment and the ecological balance will not be caused harm, meet ecological safety regulation；
(3) secondary meningitidis strains CGMCC No.13321 apt to change have autotrophic denitrification ability, can be deposited in no organic matter
Fast-growth, is sulphur simple substance or sulfate by Oxidation of Hydrogen Sulfide by nitrate reduction into nitrogen under the conditions.This characteristic profit
In by being used to strengthen the removal efficiency to biogas hydrogen sulfide after external source culture；
(4) secondary meningitidis strains CGMCC No.13321 apt to change can be removed using nitrate in anaerobic condition as electron acceptor
H in biogas2S, effectively prevent tradition carries out biogas H using oxygen as electron acceptor2Methane is introduced when S is removed to be mixed with oxygen
Close brought potential safety hazard；
(5) secondary meningitidis strains CGMCC No.13321 apt to change are using nitrate as electron acceptor, to H in biogas2The removal of S
Rate is up to 93.8%.
Figure of description
Fig. 1 is colonial morphologies of the secondary meningitidis strains CGMCC No.13321 apt to change in plating medium；
Fig. 2 is the thalli morphology that secondary meningitidis strains CGMCC No.13321 apt to change amplify 1000 times through microscope.
Specific embodiment
Technology contents of the invention are further described with reference to embodiment.
Embodiment one
1st, strain isolation purifying
Take the sewage treatment plant's anaerobic sludge of Sichuan Chengdu the 3rd.The sludge for gathering is used into sulfur-bearing chemical combination under anaerobic
The enriched medium of thing is tamed, and gradient dilution is carried out after 7d, from 10-2、10-4、10-6、10-8、10-10Drawn under dilution factor
0.1ml is coated with isolation medium flat board, scattered single bacterium colony, picking single bacterium colony, more than repetition occurs in planar surface
Operation obtains H in can removing biogas as electron acceptor using nitrate for several times, finally2The purifying bacterial strain of S.
Wherein enriched medium is：Na2S2O3·5H2O 5g、KNO3 4g、KH2PO4 2g、NaHCO3 1g、MgCl2·
6H2O 0.5g、FeSO4·7H2O 0.01g, pH value 7.0, running water 1000mL.121 DEG C, the treatment of 30min high-temperature sterilizations.Separate
Culture medium：Na2S2O3·5H2O 10g、KNO3 4g、KH2PO4 2g、NaHCO3 1g、MgCl2·6H2O 0.5g、FeSO4·7H2O
0.01g, agar 20g, running water 1000mL, pH7.0-7.5.121 DEG C, the treatment of 30min high-temperature sterilizations.
2nd, the identification of strain morphology feature and physio-biochemical characteristics
Bacterial strain of the present invention is examined under a microscope, strain cell rod-short, smaller；Bacteria colony white is raised, neat in edge light
Sliding, surface wettability, plating medium colonial morphology is shown in Fig. 1, and the thalli morphology that microscope amplifies 1000 times is shown in Fig. 2.Given birth to through physiology
Change is accredited as Gram-negative bacteria.Bacterial strain not salt tolerant, it is impossible to utilize citrate；When nitrate, nitrite or nitrogen oxide are deposited
When, can with they be electron acceptor battalion anaerobic growth.Bacterial strain can be in 15-45 DEG C of range growth, optimum growth temperature 30-35
℃；Can be grown in the range of pH7.0-7.5, the most suitable growth pH7.0.
3rd, 16SrDNA sequence analyses identification
1. first with the DP302 antibacterial agents box extraction purification bacterial strains bought in TIANGEN Biotech (Beijing) Co., Ltd.
STb gene.
2. 16SrDNA sequence amplifications, the PCR primer Ago-Gel electricity of amplification are carried out with universal primer 27F/1492R
Swimming and UV spectrophotometer measuring concentration and purity are clear with marker opposite positions when being observed under ultraviolet projecting lamp
During band, the preliminary PCR primer concentration for judging amplification is qualified；When OD260/OD280 ratios should be 1.7-1.9, show amplification
PCR primer purity it is qualified.
3. under conditions of concentration and purity are all qualified, the PCR primer to expanding send raw work bioengineering (Shanghai) share to have
Limit company specialty sequencing company carries out sequencing analysis.The 16S rDNA sequences for obtaining will be sequenced carries out Blast comparisons on NCBI,
Know and be up to more than 98% with the sequence homology of secondary coccus (Paracoccus versutus) apt to change, it is determined that purifying bacterial strain
Taxonomy.Its 16S rDNA sequence is as follows：
GCGCACCTTCGGGTGAGCGGCGGACGGGTGAGTAACGCGTGGGAATATGCCCTTTGGTACGGAATAGTC
CTGGGAAACTGGGGGTAATACCGTATGCGCCCTTCGGGGGAAAGATTTATCGCCAAAGGATTAGCCCGCGTTGGATT
AGGTAGTTGGTGGGGTAATGGCCTACCAAGCCGACGATCCATAGCTGGTTTGAGAGGATGATCAGCCACACTGGGAC
TGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTGGGGAATCTTAGACAATGGGGGCAACCCTGATCTAGCCATG
CCGCGTGAGTGATGAAGGCCCTAGGGTTGTAAAGCTCTTTCAGCTGGGAAGATAATGACGGTACCAGCAGAAGAAGC
CCCGGCTAACTCCGTGCCAGCAGCCGCGGTAATACGGAGGGGGCTAGCGTTGTTCGGAATTACTGGGCGTAAAGCGC
ACGTAGGCGGACCGGAAAGTTGGGGGTGAAATCCCGGGGCTCAACCCCGGAACTGCCTTCAAAACTATCGGTCTGGA
GTTCGAGAGAGGTGAGTGGAATTCCGAGTGTAGAGGTGAATGCTGGTTAGCGCACGGCCGTCGGGTAGACCCAACTC
CCATGGTGTGACGGGCGGTGTGTACAAGGCCCGGGAACGTATTCACCGCGGCATGCTGTTCCGCGATTACTAGCGAT
TCCAACTTCATGGGGTCGAGTTGCAGACCCCAATCCGAACTGAGATGGCTTTTGGGGATTAACCCACTGTCACCACC
ATTGTAGCACGTGTGTAGCCCAACCCGTAAGGGCCATGAGGACTTGACGTCATCCACACCTTCCTCCGACTTATCAT
CGGCAGTTCTTCCAGAGTGCCCAACCAAATGATGGCAACTGGAAGTGTGGGTTGCGCTCGTTGCCGGACTTAACCGA
ACATCTCACGACACGAGCTGACGACAGCCATGCAGCACCTGTCTCCAGGTCACCGAAGTGAAAGACCCGTCTCCGGG
CCGGTCCTGGGATGTCAAGGGTTGGTAAGGTTCTGCGCGTTGCTTCGAATTAAACCACATGCTCCACCGCTTGTGCG
GGCCCCCGTCAATTCCTTTGAGTTTTAATCTTGCGACCGTACTCCCCAGGCGGAATGCTTAATCCGTTAGGT
4th, the kind name of purifying bacterial strain determines
Comprehensive purification strain morphology feature, physio-biochemical characteristics and 16SrDNA sequence analyses, determine that the purifying bacterial strain does good
Become secondary coccus (Paracoccus versutus).
Bacterial strain is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, preservation address：Beijing
The institute 3 of Chaoyang District North Star West Road 1, Institute of Microorganism, Academia Sinica, postcode 100101；Preservation date November 18 in 2016
Day, deposit number CGMCC No.13321.
Embodiment two
The present invention secondary meningitidis strains CGMCC No.13321 apt to change are applied to H in biogas2S is removed.
1st, prepared by culture medium and inoculation bacterium solution
Solid activation medium：Na2S2O3·5H2O 10g、KNO3 4g、KH2PO4 2g、NaHCO3 1g、MgCl2·6H2O
0.5g, FeSO4·7H2O 0.01g, agar 20g, distilled water 1000mL, pH7.0-7.5,121 DEG C, the treatment of 30min high-temperature sterilizations.
Liquid fermentation medium：Na2S2O3·5H2O 10g、KNO3 4g、KH2PO4 2g、NaHCO31g, running water 1000ml, pH7.0-
7.5,121 DEG C, the treatment of 30min high-temperature sterilizations.Desulfurization culture medium：KNO3 4g、KH2PO4 1g、NaHCO31g, running water
1000ml, pH7.0-7.5,121 DEG C, the treatment of 30min high-temperature sterilizations.
Secondary meningitidis strains CGMCC No.13321 apt to change are inoculated into solid activation medium, activation culture 1d must be activated
Strain, it is stand-by；The strain that will be activated is accessed in liquid fermentation medium, and static gas wave refrigerator 1d-3d must be inoculated with bacterium solution (bacterium solution at 30 DEG C
Concentration 107-108CFU/ml), it is stand-by.
2nd, H in biogas2S removings are processed
Inoculation bacterium solution is seeded in the reactor of the desulfurization culture medium equipped with 250mL in 10% ratio；Do not simultaneously
The control of adjunction kind bacterium solution, three repetitions of every group of Setup Experiments, 30 DEG C of constant temperature is continuously passed through biogas, wherein H in biogas2S's is dense
It is 2000ppm to spend.H in METHOD FOR CONTINUOUS DETERMINATION biogas2The clearance of S, measurement result is shown in Table 1.
The biogas H of table 12The clearance of S
As seen from the above table, temperature is 30 DEG C, is inoculated with the biogas in secondary meningitidis strains CGMCC No.13321 experimental groups apt to change
H2S concentration ratio control groups are substantially reduced, wherein control group biogas H2The clearance of S is only 17.4%, and experimental group biogas H2S's
Clearance is up to 92.2%.
Embodiment three
Secondary meningitidis strains CGMCC No.13321 apt to change are used for biogas H2S is removed, and it is with the something in common of embodiment three no longer
Repeat, its difference is the treatment conditions for being inoculated with bacterium solution to desulfurization reactor.
20 DEG C for the treatment of temperature, H in METHOD FOR CONTINUOUS DETERMINATION biogas2The clearance of S, measurement result is shown in Table 2.
The biogas H of table 22The clearance of S
As seen from the above table, temperature is 20 DEG C, is inoculated with the biogas in secondary meningitidis strains CGMCC No.13321 experimental groups apt to change
H2S concentration ratio control groups are substantially reduced, wherein control group biogas H2The clearance of S is only 6.2%, and experimental group biogas H2S's goes
Except rate is up to 77.1%.Compared with 30 DEG C of temperature conditionss, although secondary meningitidis strains CGMCC No.13321 apt to change are to biogas H2S's
Clearance declines, but still shows preferable H2S removal effects.
Example IV
Secondary meningitidis strains CGMCC No.13321 apt to change are used for biogas H2S is removed, and it is with the something in common of embodiment three no longer
Repeat, its difference is the H being passed through in biogas2S concentration.
30 DEG C of constant temperature is continuously passed through biogas, wherein H in biogas2The concentration of S is 3000ppm.H in METHOD FOR CONTINUOUS DETERMINATION biogas2S's
Clearance, measurement result is shown in Table 2.
The biogas H of table 22The clearance of S
As seen from the above table, it is inoculated with the biogas H in secondary meningitidis strains CGMCC No.13321 experimental groups apt to change2S concentration ratios pair
Substantially reduced according to group, wherein control group biogas H2The clearance of S is only 11.6%, and experimental group biogas H2The clearance of S is up to
88.6%.Be passed through biogas H2S concentration is compared for 2000ppm, and secondary meningitidis strains CGMCC No.13321 apt to change are to biogas H2S's
Clearance only slightly declines, and still shows preferable H2S removal effects.
Embodiment five
Secondary meningitidis strains CGMCC No.13321 apt to change are used for biogas H2S is removed, and it is with the something in common of embodiment three no longer
Repeat, its difference is H in biogas2S treatment scales.
Inoculation bacterium solution is seeded in the reactor of the desulfurization culture medium equipped with 10L in 10% ratio, METHOD FOR CONTINUOUS DETERMINATION natural pond
H in gas2The clearance of S, measurement result is shown in Table 4.
The biogas H of table 42The clearance of S
As seen from the above table, it is inoculated with the biogas H in secondary meningitidis strains CGMCC No.13321 experimental groups apt to change2S concentration ratios pair
Substantially reduced according to group, wherein control group biogas H2The clearance of S is only 21.6%, and experimental group biogas H2The clearance of S is up to
93.8%.After illustrating that treatment scale amplifies, secondary meningitidis strains CGMCC No.13321 apt to change still show preferable H2S is removed
Effect.
Sequence table
<110>Chengdu Inst. of Biology, Chinese Academy of Sciences
<120>One plant of secondary meningitidis strains apt to change and its application
<160>1
<210>1
<211>1142
<212>DNA
<213>Secondary coccus (Paracoccus versutus) apt to change
<400>1
gcgcaccttc gggtgagcgg cggacgggtg agtaacgcgt gggaatatgc cctttggtac 60
ggaatagtcc tgggaaactg ggggtaatac cgtatgcgcc cttcggggga aagatttatc 120
gccaaaggat tagcccgcgt tggattaggt agttggtggg gtaatggcct accaagccga 180
cgatccatag ctggtttgag aggatgatca gccacactgg gactgagaca cggcccagac 240
tcctacggga ggcagcagtg gggaatctta gacaatgggg gcaaccctga tctagccatg 300
ccgcgtgagt gatgaaggcc ctagggttgt aaagctcttt cagctgggaa gataatgacg 360
gtaccagcag aagaagcccc ggctaactcc gtgccagcag ccgcggtaat acggaggggg 420
ctagcgttgt tcggaattac tgggcgtaaa gcgcacgtag gcggaccgga aagttggggg 480
tgaaatcccg gggctcaacc ccggaactgc cttcaaaact atcggtctgg agttcgagag 540
aggtgagtgg aattccgagt gtagaggtga atgctggtta gcgcacggcc gtcgggtaga 600
cccaactccc atggtgtgac gggcggtgtg tacaaggccc gggaacgtat tcaccgcggc 660
atgctgttcc gcgattacta gcgattccaa cttcatgggg tcgagttgca gaccccaatc 720
cgaactgaga tggcttttgg ggattaaccc actgtcacca ccattgtagc acgtgtgtag 780
cccaacccgt aagggccatg aggacttgac gtcatccaca ccttcctccg acttatcatc 840
ggcagttctt ccagagtgcc caaccaaatg atggcaactg gaagtgtggg ttgcgctcgt 900
tgccggactt aaccgaacat ctcacgacac gagctgacga cagccatgca gcacctgtct 960
ccaggtcacc gaagtgaaag acccgtctcc gggccggtcc tgggatgtca agggttggta 1020
aggttctgcg cgttgcttcg aattaaacca catgctccac cgcttgtgcg ggcccccgtc 1080
aattcctttg agttttaatc ttgcgaccgt actccccagg cggaatgctt aatccgttag 1140
gt 1142
Claims (6)
1. one plant of secondary meningitidis strains apt to change, Paracoccus versutus, it is characterised in that the bacterial strain is secondary coccus apt to change
2#, its deposit number is：CGMCC No.13321；Gram's staining is feminine gender, and strain cell rod-short is smaller；Bacteria colony white
Projection, neat in edge is smooth, surface wettability；Bacterial strain not salt tolerant, it is impossible to utilize citrate；Bacterial strain can be in 15-45 DEG C, pH7.0-
Grown in the range of 7.5；In the presence of nitrate, nitrite or nitrogen oxide, anaerobic growth can be sought with them as electron acceptor.
2. application of the apt to change secondary coccus described in claim 1 in hydrogen sulfide is removed.
3. application as claimed in claim 2, it is characterised in that comprise the following steps：
(1) secondary coccus CGMCC No.13321 apt to change are activated with solid activation medium using preceding；
(2) liquid fermentation medium for crossing sterilization treatment is quantitative standby；
(3) strain after activation is seeded to the liquid fermentation medium culture that sterilization treatment is crossed, static gas wave refrigerator at 30 DEG C respectively
1d-3d；
(4) bacterium solution is inoculated in the reactor of the desulfurization culture medium crossed equipped with sterilization treatment in 10% ratio；
(5) biogas is passed through, beginning is passed through with low discharge, and flow gradually increases later, proceeds by biogas H2S removings are processed.
4. application as claimed in claim 3, it is characterised in that the solid activation medium component in the step (1) is：
Na2S2O3·5H2O 10g、KNO3 4g、KH2PO4 2g、NaHCO3 1g、MgCl2·6H2O 0.5g, FeSO4·7H2O 0.01g、
Agar 20g, distilled water 1000mL, pH7.0-7.5；Activation condition is：121 DEG C, the treatment of 30min high-temperature sterilizations.
5. application as claimed in claim 3, it is characterised in that the liquid fermentation medium component in the step (2) is：
Na2S2O3·5H2O 10g、KNO3 4g、KH2PO4 2g、NaHCO31g, running water 1000ml, pH7.0-7.5；Sterilization treatment bar
Part is：121 DEG C, the treatment of 30min high-temperature sterilizations.
6. application as claimed in claim 3, it is characterised in that desulfurization nutrient media components is in the step (4)：KNO3 4g、
KH2PO4 1g、NaHCO31g, running water 1000ml, pH7.0-7.5；Sterilization treatment condition is：121 DEG C, 30min high-temperature sterilizations
Treatment.
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| CN110656057A (en) * | 2018-06-29 | 2020-01-07 | 龙岩学院 | Heterotrophic nitrification-aerobic denitrification paracoccus strain, seed liquid, preparation method and application thereof |
| CN114958659A (en) * | 2022-05-11 | 2022-08-30 | 江苏科技大学 | Paracoccus capable of being changed well and having aerobic nitrification, denitrification, nitrogen and phosphorus removal performance |
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