CN114686391B - High-salt-tolerance bacterium and application thereof - Google Patents
High-salt-tolerance bacterium and application thereof Download PDFInfo
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- 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
- C02F3/301—Aerobic and anaerobic treatment in the same reactor
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- 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
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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
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- 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
- C02F2101/101—Sulfur compounds
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- Water Supply & Treatment (AREA)
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Abstract
The invention discloses a strain of high salt tolerance bacteriaHalomonas nigrificans) GXNYJ-DL-1 is preserved in China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC No.20350 in the year 7 and 13 of 2020. The high salt tolerant bacteria GXNYJ-DL-1 of the invention has the following forms: the bacterial colony is light yellow, round, moist in surface, opaque and neat in edge; the microscopic morphology under the microscope is: the bacterial cells are in the shape of rod, 0.5-0.7 μm×1.2-3.6 μm, and are arranged singly or in pairs, and gram negative. Compared with the high-salt-tolerance strain in the prior art, the high-salt-tolerance strain GXNYJ-DL-1 has excellent salt tolerance capability, can especially tolerate high sulfate, and is more suitable for COD removal of high-salt wastewater, especially high-sulfate wastewater.
Description
Technical Field
The invention relates to a high salt tolerant bacterium and application thereof in removing COD in high salt wastewater, belonging to the technical field of microorganism and wastewater treatment.
Background
The industries such as chemical industry, pharmacy, papermaking, food processing, mining, fermentation and the like are easy to produce high-salt wastewater, especially high-sulfate organic wastewater, and the sulfate concentration of some wastewater can reach 30000-50000 mg/L, and the COD exceeds 10000mg/L. The sulfate ions exist in a large amount, so that the high-salt-content wastewater is not easy to biochemically treat, and the physicochemical process routes such as membrane separation, concentration crystallization and the like become neither economical nor desirable (easy to block) due to high organic matters.
For this kind of wastewater, the existing method and research direction are as follows:
(1) The chemical method is to add lime to convert sulfate into calcium sulfate precipitate. For example, in the patent CN106865880A, CN105439374a, the core of sulfate removal is lime neutralization, and chemical flocculation is adopted at the same time, if the high-COD organic wastewater is also subjected to the process, when lime and flocculant are added, calcium sulfate precipitation can be generated, a large amount of organic matters and heavy metals can be mixed into the precipitation, and finally lime mud and sludge generated can only be used as dangerous waste, and no utilization value and secondary pollution are generated.
(2) The simple biochemical method adopts a first-stage anaerobic technology. Because the volume load of the anaerobic process is higher than that of the aerobic process, anaerobic treatment of organic wastewater is mostly adopted. If the wastewater contains sulfate, the sulfate is reduced to S under the action of Sulfate Reducing Bacteria (SRB) under the anaerobic condition 2- The ion has stronger biological toxicity, has larger inhibition effect on microbial flora, especially methanogen, and seriously affects the removal of organic matters. Thus in order to reduce S 2- The effect of ions on methanogens generally requires aThe sulfate concentration of the stage anaerobic reactor is less than 2000mg/L, and is not applicable to organic wastewater with sulfate concentration exceeding 10000mg/L. For example, in patent CN103771670A, the sulfate radical concentration is more than 1000mg/L, and a single anaerobic-aerobic process is adopted, but the process is not suitable for waste water of a dibasic acid fermentation process.
(3) In order to avoid the competition between sulfate reducing bacteria and methanogens in the anaerobic process, a two-stage anaerobic process is currently used for treating high-concentration organic wastewater at home and abroad. As described in patent CN105439374A, the primary anaerobic control is in the hydrolytic acidification stage, and sulfate reducing bacteria reduce most of sulfate radical in the wastewater into S 2- Ions and with H + Hydrogen sulfide is generated by combination, and is blown off by nitrogen and enters a dry desulfurizing device for desulfurization; the second-stage anaerobic control is in the methanogenesis stage, and the anaerobic effluent enters an aerobic reaction tank for further treatment. The dry desulfurization used in the first-stage anaerobic section is a older technology, has larger defects in desulfurizing agent replacement, continuous operation, regeneration treatment and the like, and does not definitely lead to the final sulfur removal; the organic matter in the secondary anaerobic process eventually generates methane, carbon dioxide, water and a small amount of hydrogen sulfide, namely biogas, and the patent also ignores the treatment of biogas.
(4) Biological desulphurisation, i.e. reduction of sulphate to S by sulphate-reducing bacteria under anaerobic conditions 2- Or hydrogen sulfide, and then biologically oxidizing the hydrogen sulfide by sulfur oxidizing bacteria to generate elemental sulfur, such as patent CN102795739A, CN103172218A, CN103319002A. The biological desulfurization technology has the defects of difficult control of the process and strict condition requirements, and the problems of poor separation effect and low sulfur purity of the liquid-phase prepared sulfur, so that the technology has been developed for decades, but has a certain distance from industrial application at present.
(5) The special salt-tolerant bacteria biotechnology is based on the rapid development of genetic engineering technology, and the dominant bacterial colony suitable for high-salt-content wastewater is domesticated by a scientific method, and can grow in an environment with higher salinity by virtue of the unique cell structure and substance composition of the bacterial colony, such as the patents CN201610547861, CN201510626828, CN201610720403,CN201510737150. At present, the directional patent mainly uses NaCl as a main component of salt in strain screening and culturing, and the mass ratio of sulfate is relatively low because S is generated due to local anaerobism when the sulfate concentration in the culture medium/liquid is relatively high 2- Or hydrogen sulfide, brings about a certain biotoxicity, inhibits or kills the bacterial species. The salt-tolerant bacteria related to the patent only can be used in high-salt-content wastewater simply, the stability and long-period adaptability of the strain under special environments are not examined, the specific implementation details are also lacking, and the salt-containing wastewater is NaCl-containing wastewater.
Disclosure of Invention
Aiming at the defects, the invention provides a strain of high salt-tolerant bacteria and application thereof in removing COD in high salt wastewater, and the strain has the characteristics of high salt tolerance, particularly high sulfate tolerance.
The technical purpose of the invention is realized by the following technical scheme:
the technical purpose of the first aspect of the invention is to provide a strain of high salt-tolerant bacteriaHalomonas nigrificans) GXNYJ-DL-1 is preserved in China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC No.20350 in the year 7 and 13 of 2020.
The high salt tolerant bacteria GXNYJ-DL-1 provided by the invention has the following forms: the bacterial colony is light yellow, round, moist in surface, opaque and neat in edge; the microscopic morphology under the microscope is: the bacterial cells are in the shape of rod, 0.5-0.7 μm×1.2-3.6 μm, and are arranged singly or in pairs, and gram negative.
The 16S rDNA gene sequencing analysis result of the high salt tolerant bacteria GXNYJ-DL-1 provided by the invention is shown in a sequence table.
The technical purpose of the second aspect of the invention is to provide a culture method of the high salt tolerant bacteria GXNYJ-DL-1, which comprises three stages of bacterial strain activation, seed liquid culture and intermittent aeration culture, and comprises the following specific steps:
(1) Strain activation: the strain GXNYJ-DL-1 is inoculated into a broth peptone solid culture medium with the salt content of 1-5% by adopting a plate streaking method, and is cultured in an incubator at the temperature of 28-35 ℃ for 48-72 hours.
(2) Seed liquid culture: after the strain activation is finished, the activated strain in the flat plate is selected and inoculated into a conical flask containing broth peptone liquid culture medium, wherein the salt content in the liquid culture medium is 1-5%, sulfate accounts for more than 50%, shake culture is carried out on a shaking table, the temperature is 30-35 ℃, the rotating speed is 100-200 rpm, and the culture time is 24-96 hours.
(3) Intermittent aeration culture: adding prepared salt-containing wastewater or wastewater to be treated into a reactor provided with an aeration device, wherein the salt content is 1-5%, the sulfate accounts for more than 50%, inoculating liquid seed liquid according to the volume ratio of 3-20%, controlling the pH value to be 6-9, performing indirect aeration according to the relation of the aeration duration and the aeration stopping duration of 2:1-1:2, and the total culture period is 72-144 h; s production by means of sulfate reduction reactions occurring upon indirect aeration 2- Improving S resistance of strain GXNYJ-DL-1 2- Capability of preventing other miscellaneous bacteria from breeding.
The high-salt-tolerance GXNYJ-DL-1 bacterial liquid obtained by the culture method provided by the invention can be kept stand in a refrigerator at 4 ℃ for 3-5 months without inactivation, and has strong vitality and high stability.
The technical purpose of the third aspect of the invention is to provide the application of the high salt tolerant bacteria GXNYJ-DL-1 in removing COD in high salt organic wastewater.
Further, the high salt tolerant bacteria GXNYJ-DL-1 has a tolerant salt content of up to 25wt% and can tolerate S 2- The concentration reaches 300mg/L, and compared with other salt tolerant bacteria, the method is more suitable for treating high-salt wastewater, especially the salt-containing wastewater mainly comprising sulfate.
The technical purpose of the fourth aspect of the invention is to provide a treatment process for high-sulfate organic wastewater by utilizing the high-salt-tolerance bacterium GXNYJ-DL-1, which comprises a front-end treatment section, a deep treatment section, a sludge treatment section and a tail gas treatment section.
The front-end treatment section sequentially comprises pH adjustment, primary aerobic treatment, anaerobic treatment, stripping treatment and secondary aerobic treatment; the pH adjustment is to add a pH regulator into the wastewater to adjust the pH of the wastewater to 6.5-7.5, then the wastewater enters primary aerobic treatment, and high-sulfate organic wastewater is treated by high-salt-tolerance bacteria GXNYJ-DL-1 to reduce the COD of the wastewater to below 1000 mg/L; the first-stage aerobic effluent enters an anaerobic section, sulfate is reduced into hydrogen sulfide under the action of sulfate reducing bacteria, and methane is used as blowing and degassing to carry out stripping, so that the generated hydrogen sulfide is carried out a wastewater system; the anaerobic effluent enters a secondary aerobic treatment section, COD is further removed by adopting conventional sludge strains, and the secondary aerobic effluent enters a deep treatment section.
The advanced treatment section sequentially comprises advanced oxidation, BAF and post dephosphorization; the advanced oxidation is to remove refractory organic matters in the wastewater, improve the biodegradability of the wastewater, and then further remove COD in the wastewater through BAF and play a role in filtering; the post dephosphorization is carried out by adding the medicament to carry out two-stage chemical dephosphorization, and finally the wastewater meeting the discharge standard is obtained.
The sludge treatment section is used for conveying redundant sludge generated by the primary aerobic unit, the secondary aerobic unit and the BAF unit to the sludge anaerobic unit by using a pump, and converting most of activated sludge into methane gas through sludge anaerobic oxidation and recycling the methane gas to the methane storage tank.
The tail gas treatment section comprises solvent absorption, solvent regeneration and sulfuric acid preparation by a WSA wet method; the solvent absorption is to absorb hydrogen sulfide gas in the gas by an absorbent, so that the gas is purified to obtain methane, and the methane is stored in a methane storage tank; the solvent regeneration is to heat the gas absorbed by the solvent to escape, the solvent is recycled, and the escaped gas is subjected to WSA wet sulfuric acid production to obtain sulfuric acid products.
Further, the pH regulator used for the pH regulation is alkali which does not precipitate with sulfate radical or is slightly dissolved after being added into the wastewater, more specifically sodium hydroxide or potassium hydroxide, and the pH regulation is carried out in a regulating tank.
Further, the primary aerobic bacteria GXNYJ-DL-1 is preferably selected from one of the processes with high volume load such as biological contact oxidation process and MBBR process, and the volume load is 2kg (BOD) 5 ）/m 3 D is more than, the dissolved oxygen is controlled to be more than 2mg/L, and the residence time of the wastewater is 24-120 h.
Further, the dissolved oxygen in the anaerobic treated wastewater is controlled below 0.2mg/L, methane gas is adopted for stripping, the residence time of the wastewater is 12-144 h, the temperature is 25-35 ℃, and the volume ratio of methane stripping and wastewater is not less than 5:1.
It should be understood by those skilled in the art that the primary aerobic section can decompose most of organic matters in the wastewater through the high salt tolerant bacteria GXNYJ-DL-1, partially convert the organic matters into inorganic carbon (carbon dioxide), and partially transfer the inorganic carbon into activated sludge for removal by a sludge discharge mode; the high-efficiency salt-tolerant bacterium GXNYJ-DL-1 solves the problem that common strains cannot survive under the condition of high salt content, and also solves the problem that the common salt-tolerant bacterium causes S due to uneven aeration or local anaerobic oxidation of flora in the presence of a large amount of sulfate 2- Higher concentration and even incapacity of survival; the anaerobic section is a sulfate reduction section, most sulfate is reduced to hydrogen sulfide under the action of sulfate reduction bacteria, redundant hydrogen sulfide gas is carried out of the system through continuous methane stripping, and as most organic matters are removed by the first-stage aerobic section, a small amount of residual organic matters are insufficient to enable methanogens to propagate in large quantities, at the moment, the sulfate reduction bacteria are taken as main bodies, and a small amount of organic matters are consumed as carbon sources of the sulfate reduction bacteria.
Further, the secondary aerobic treatment adopts a conventional activated sludge method, and conventional biological flora is used, so that the retention time of wastewater is 12-48 hours; the secondary aerobic main treatment primary aerobic residual organic matters and the anaerobic section partially acidify and hydrolyze organic matters, and a large amount of organic matters and sulfate are removed from the unit, so that the unit has moderate salt concentration and volume load, the conventional biological flora can achieve the treatment effect in a short time, and the treatment pressure and cost of the subsequent advanced oxidation unit are reduced.
Further, the advanced oxidation is selected from one of ozone oxidation, electrocatalytic oxidation and Fenton oxidation. So as to decompose organic substances which are difficult to degrade in the wastewater and improve the biodegradability of the wastewater; the BAF can further remove COD in the wastewater and has a filtering effect, and the BAF unit can be replaced by an MBR with similar effect.
Further, the post-phosphorus removal adopts two-stage chemical phosphorus removal, wherein the chemical agent is a ferric chloride and calcium hydroxide composite agent, and mainly aims at the problem that the total phosphorus content in the dibasic acid fermentation wastewater is relatively high; the ferric chloride is acidic, the calcium hydroxide is alkaline, and the two are compounded according to a certain proportion, so that the pH value of the liquid is ensured to be neutral, and the subsequent effluent does not need to be subjected to pH adjustment; the adding amount of the primary dephosphorization ferric chloride is 34 mg/L-680 mg/L, and the adding amount of the calcium hydroxide is 10 mg/L-300 mg/L; the adding amount of the secondary dephosphorization ferric chloride is 10 mg/L-200 mg/L, and the adding amount of the calcium hydroxide is 4 mg/L-80 mg/L; the ferric phosphate and the calcium phosphate generated by the post phosphorus removal are filtered and recovered and can be used as phosphate fertilizer.
Furthermore, the sludge treatment section mainly recovers the sludge generated by the aerobic section, and converts the biological sludge into methane through sludge anaerobic oxidation, and the sludge treatment section can effectively realize the resource recovery of the organic matters because a large amount of biological sludge is generated in the process of removing the organic matters by the first-stage aerobic section.
Further, the absorbent of the tail gas treatment section is selected from one of monoethanolamine, diethanolamine, diisopropanolamine and N-methyldiethanolamine, preferably N-methyldiethanolamine. The main absorption by the absorbent is hydrogen sulfide gas, and also includes a small amount of carbon dioxide.
Further, the solvent regeneration is performed in a heated distillation column.
Furthermore, the purity of the methane in the methane storage tank after being absorbed and separated by the precursor solvent can reach more than 90%, wherein part of the methane is recycled to the outside of the blowing and degassing, and the redundant methane can be used as a product for recycling.
Further, the WSA wet sulfuric acid preparation method is to prepare sulfuric acid through the processes of incineration, conversion and condensation: incinerating the gas separated by regenerating the solvent, wherein H 2 S combustion to generate SO 2 ，SO 2 Conversion to SO under the action of a catalyst 3 ，SO 3 And condensing the water vapor in a condenser to generate sulfuric acid.
Compared with the prior art, the invention has the following advantages:
(1) The invention obtains a high salt-tolerant strain GXNYJ-DL-1 through screening culture, provides a culture method thereof, and the GXNYJ-DL-1 bacterial liquid obtained through the culture method can be stored for 3-5 months at 0-8 ℃ without inactivation, and has very strong stability. Compared with the high-salt-tolerance strain in the prior art, the high-salt-tolerance strain GXNYJ-DL-1 has excellent salt tolerance capability, can especially tolerate high sulfate, and is more suitable for COD removal of high-salt wastewater, especially high-sulfate wastewater.
(2) The invention provides a process route which takes a high salt-tolerant strain GXNYJ-DL-1 as a core and takes resource recycling as a aim, solves the problem of overhigh sulfate radical and COD in wastewater, provides a process raw material (sulfuric acid) for a source, and simultaneously realizes the resource utilization of organic matter-biological sludge-methane.
(3) The method for treating the sewage by the binary acid fermentation of the invention can maximize the recycling of the sulfur element resource, and is specifically characterized in that: sodium hydroxide, potassium hydroxide and the like are added in a pH regulating tank instead of calcium hydroxide, so that sulfur element does not enter solid hazardous waste taking calcium sulfate and flocculating agent as main bodies; maximizing sulfate reduction with independent anaerobism; methane stripping is used for strengthening the recovery of hydrogen sulfide; the recycling of sulfuric acid and sulfur recovery are realized by combining solvent absorption, regeneration and WSA wet acid preparation, and finally the high-efficiency recovery of hydrogen sulfide and the recycling of sulfur element are realized.
(4) Aiming at the characteristic of high sludge yield of the aerobic process, the process route of sludge recovery, sludge anaerobic oxidation, methane circulation stripping, methane separation and purification and methane recovery and utilization is developed, the recycling of organic matters is realized, and high-purity methane is prepared.
(5) The waste water standard emission is finally realized by the dibasic acid fermentation sewage treatment process, the total phosphorus in the sewage is removed by the rear two-stage phosphorus removal process, the phosphorus fertilizer can be recycled, and meanwhile, the secondary pollution is greatly reduced by the whole process due to the recycling of hydrogen sulfide and methane.
Drawings
FIG. 1. Microscopic morphology of highly salt tolerant strain GXNYJ-DL-1 under a microscope;
FIG. 2A photograph of colony morphology of highly salt tolerant strain GXNYJ-DL-1 on solid medium;
FIG. 3 growth curves of the strains of example 3 at different salt concentrations;
FIG. 4 removal of COD by the strain of example 3 at different salt concentrations;
FIG. 5 strain S in example 4 2- Growth curve at concentration;
FIG. 6 is a flow chart of the long-chain dibasic acid fermentation sewage treatment.
Description of biological Material preservation
The strain with high salt tolerance provided by the inventionHalomonas nigrificans) GXNYJ-DL-1 is preserved in China general microbiological culture Collection center (China Committee for culture Collection of microorganisms); address: the institute of microorganisms of national academy of sciences of China, national institute of sciences, no. 1, no. 3, north Chen West Lu, the Korean region of Beijing; preservation number: CGMCC No. 20350; preservation date: 7 months and 13 days 2020.
Detailed Description
The present invention will be described in further detail with reference to specific examples. The embodiments and specific operation procedures are given on the premise of the technical scheme of the invention, but the protection scope of the invention is not limited to the following embodiments.
Example 1
The screening process of the high salt tolerant bacteria GXNYJ-DL-1 strain provided by the invention comprises the following steps:
(1) Sampling: taking activated sludge and sewage samples of Qingjiang petrochemical diacid sewage treatment plant in Huaishu, jiangsu province, and storing the activated sludge and sewage samples in a refrigerator at 4 ℃ for later use.
(2) And (3) sludge activation: putting the standby sewage and activated sludge in a refrigerator into a reactor provided with an aeration device, and performing stuffy aeration for 48-72 h, wherein the dissolved oxygen is controlled to be more than or equal to 2mg/L.
(3) Primary domestication: putting 10mL of mixed bacterial liquid into a conical flask filled with 200mL of liquid culture medium after the stuffy exposure is finished, then putting into an oscillator for culture, wherein the temperature is 30-35 ℃, the rotating speed is 100-200 rpm, the culture period is 48-72 h, and putting 10mL of bacterial liquid into a second batch of liquid culture medium after the culture is finished, and so on; containing the first liquid mediumThe salt content is 1% (mass percentage concentration), the second time is 2%, the fifth time is 5%, and the total amount is 5 batches; the liquid culture medium is a broth peptone culture medium, and comprises the following components: beef extract 3g, peptone 10g, distilled water 1000ml, naCl 5g, pH 7, na 2 SO 4 5-45 g, and preparing liquid culture media with different mass concentrations through different sodium sulfate ratios.
(4) Domesticating again: adding 10ml of bacterial liquid of the 5 th batch into a conical flask filled with 200ml of liquid culture medium after the initial domestication is finished, adding sodium sulfide to prepare S 2- The concentration is 100mg/L, then the culture is carried out in an oscillator for 48 to 72 hours, 10ml of bacterial liquid is taken out after the culture is finished and is put in a liquid culture medium of a second batch, and the like; second liquid Medium S 2- The concentration was 150mg/L, 300% for the fifth time, for a total of 5 batches.
(5) Screening of high-efficiency salt-tolerant strains: taking salt content of 5% S 2- 1mL of bacterial liquid in a liquid culture medium with the initial concentration of 300mg/L and after the shake reaction for 48-72 hours is diluted 1000 times by sterile distilled water, inoculated into a solid culture medium by adopting a plate streak separation method, and then cultured for 48-72 hours at 30 ℃; after the culture is finished, selecting strains with different colony forms, continuously inoculating the strains into a new solid culture medium by adopting a plate streak separation method, repeating for 3-8 times until strains with consistent colony forms are screened out, and finally storing the strains in a refrigerator at 4 ℃; the solid culture medium is broth peptone culture medium, agar 20g is added, wherein Na 2 SO 4 45g.
The strain obtained by the method is in a form of a rod-shaped thallus under a microscope, and the thallus is 0.5-0.7 mu m multiplied by 1.2-3.6 mu m, arranged singly or in pairs and is gram-negative. The solid cultured colonies were pale yellow, round, moist on the surface, opaque, and clean on the edges, as shown in fig. 1 and 2.
Example 2
Identification of strains:
the physiological and biochemical identification and the 16S rDNA gene sequencing analysis show the results in Table 1 and the gene sequence determination results in the sequence table.
TABLE 1
And (3) injection: "+" indicates positive response or availability; "-" indicates a negative reaction or failure to utilize.
Example 3
Salt tolerance measurement of high salt tolerance bacterium GXNYJ-DL-1
Preparing simulated wastewater (g/L): phenol 0.4, naCl 3, feSO 4 0.02，CaCl 2 0.03，MgSO 4 1，Na 2 SO 4 3，KH 2 PO 4 0.034 ，NH 4 Cl 0.3, yeast extract 0.1, tryptone 0.05g, pH 7, salt content about 1% (mass percent). Na is added additionally on the basis of simulating the wastewater with 1 percent of salt content 2 SO 4 The prepared waste water has salt content of 5%, 9%, 13%, 17%, 21% and 25% respectively.
Taking GXNYJ-DL-1 bacterial liquid, adding the bacterial liquid into a conical flask according to the volume ratio of the bacterial liquid to the simulated wastewater of 1:20, adopting a shaking table oscillation method, controlling the temperature at 35 ℃, controlling the rotating speed at 150rpm, sampling at regular time, and measuring the bacterial density (OD) by using a spectrophotometer 600 ) Drawing a strain growth curve, wherein the strain growth curve under different salt concentrations is shown in figure 3; and the COD value of the final reaction solution was measured to determine the removal rate of COD by the strain, and the removal rate of COD of the strain at different salt concentrations after 76 hours is shown in FIG. 4.
According to the results shown in FIG. 3 and FIG. 4, the growth of the strain is relatively slowed down along with the increase of the salt concentration, but the strain can be rapidly increased after a certain adaptation period, the strain grows faster under the salt concentration of 1% -13%, and the COD removal rate (initial phenol COD is about 1247 mg/L) is also higher than 65%; at 25% salt concentration, the strain adaptation period is relatively long, about 50 hours, after which the strain starts to enter the growth phase, OD 600 The value is obviously increased, and the corresponding COD removal rate can still reach 53 percent.
The embodiment shows that the strain GXNYJ-DL-1 has stronger salt tolerance, and the COD removal rate can still reach 53% under the condition of 25% of salt concentration.
Example 4
S-tolerance of high salt tolerant bacteria GXNYJ-DL-1 2- Toxicity determination
Preparing simulated wastewater (g/L): phenol 0.4, naCl 3, feSO 4 0.02，CaCl 2 0.03，MgSO 4 1，Na 2 SO 4 43，KH 2 PO 4 0.034 ，NH 4 Cl 0.3, yeast extract 0.1, tryptone 0.05g, pH 7, salt content about 5% (mass percent). Additional addition of Na based on simulated wastewater 2 S is prepared into S 2- Waste water with mass concentration of 0mg/L, 50mg/L, 100mg/L, 150mg/L, 200mg/L, 250mg/L and 300mg/L.
Taking GXNYJ-DL-1 bacterial liquid, adding the bacterial liquid into a conical flask according to the volume ratio of the bacterial liquid to the simulated wastewater of 1:20, standing for 24 hours, oscillating by a shaking table, controlling the temperature at 35 ℃, rotating at 150rpm, sampling at regular time, and measuring the bacterial density (OD) by a spectrophotometer 600 ) Strain growth curves were plotted as shown in figure 5.
As can be seen from FIG. 5, the growth of the strain during standing is very slow, limited by the dissolved oxygen on the one hand and S on the other hand 2- Toxicity inhibition, after 24h of standing period, starting shaking table shaking reaction, wherein the concentration of the strain starts to be obviously increased, but compared with example 3, the strain grows relatively slowly; over two days of growth, overall OD 600 The value was increased from 0.25 to 0.45, indicating that the strain was not due to early S 2- Is lost, and gradually resumes viability after a relatively long adaptation period, in particular at 300mg/L of S 2- The strain concentration of the sample is steadily increasing.
As shown in the example, the strain GXNYJ-DL-1 has strong S resistance 2- Toxicity capability, which has been demonstrated to be S tolerant 2- The concentration reaches 300mg/L.
Example 5
Long chain diacid fermentation sewage treatment by utilizing high salt tolerant bacteria GXNYJ-DL-1
The process flow chart for treating the diacid fermentation sewage is shown in figure 6: the wastewater produced by the dibasic acid fermentation process has the following water quality: COD 10100mg/L, sulfate 24000mg/L, total salt content 37000mg/L, pH 3.5 and wastewater flow rate 10t/h. NaOH is added into the regulating tank until the pH value is 6.8, and the wastewater flows into a first-stage aerobic unit, wherein the unit adopts a biological contact oxidation tank process, the strain is high-efficiency salt-tolerant bacteria GXNYJ-DL-1, the dissolved oxygen is controlled to be more than 2mg/L, the retention time of the wastewater is 72h, and the COD of the final effluent is 950mg/L and flows into an anaerobic unit. Controlling the dissolved oxygen of the anaerobic unit below 0.2mg/L, keeping the wastewater at the temperature of 30 ℃ for 90 hours, adopting methane gas for blowing off, and enabling hydrogen sulfide generated by anaerobic to be separated from a water phase, enter a gas phase along with methane and enter a tail gas treatment section, wherein the volume ratio of methane blowing off to wastewater is 6:1. The effluent from the anaerobic unit enters a secondary aerobic unit, COD of the inlet water is 558mg/L, the sulfate content is 2500mg/L, the unit adopts a conventional activated sludge method, conventional biological flora is used, the residence time of wastewater is 24h, COD of the final outlet water is 150mg/L, and the outlet water enters a subsequent advanced treatment section.
Advanced oxidation in the advanced treatment section adopts ozone catalytic oxidation, effluent is further treated by BAF, and COD of the final effluent is less than or equal to 60mg/L, and total phosphorus is about 60mg/L. The post-phosphorus removal is carried out by selecting two-stage chemical phosphorus removal, wherein the chemical agents are ferric chloride and calcium hydroxide which accord with the agents, the adding amount of the first-stage phosphorus removal ferric chloride is 300mg/L, the adding amount of the calcium hydroxide is 120mg/L, the adding amount of the second-stage phosphorus removal ferric chloride is 100mg/L, the adding amount of the calcium hydroxide is 40mg/L, the generated phosphate precipitate is filtered and recycled, the COD of the post-phosphorus removal effluent is less than or equal to 60mg/L, the sulfate is about 1700mg/L, the total salt amount is 4200 mg/L, the total phosphorus is less than or equal to 0.5mg/L, and the standard emission is realized.
The sludge generated by the primary aerobic unit, the secondary aerobic unit and the BAF unit is conveyed to a sludge anaerobic oxidation unit by a pump except for normal backflow, and the sludge is converted into methane gas by sludge anaerobic oxidation and is recovered to a methane storage tank.
And (3) absorbing tail gas generated by anaerobic treatment by using lean amine solution N-methyldiethanolamine to obtain methane with the purity of 97%, storing the methane into a methane tank, recycling part of the methane tank, and utilizing part of methane as a product. After the solvent absorption section absorbs and saturates, the rich amine liquid enters a solvent regeneration tower for regeneration, the regenerated lean amine liquid returns to the solvent absorption section, the escaped gas is hydrogen sulfide with the purity of about 93 percent, and enters a WSA (wireless sensor array) acid making process, and finally sulfuric acid with the concentration of about 30 percent is prepared and is recycled as a raw material of a binary acid process.
The embodiment of the invention can effectively treat long-chain binary acid fermentation sewage, so that the sewage is discharged up to the standard and the resource recycling is realized.
Example 6
High sulfate and high organic wastewater treatment by utilizing high salt tolerant bacteria GXNYJ-DL-1
A high-salinity-tolerance bacterium GXNYJ-DL-1 is adopted to treat a certain strand of high-sulfate high-organic wastewater, and the sewage treatment flow chart is shown in figure 6.
The high sulfate high organic wastewater has the following water quality: COD 12000mg/L, sulfate 35000mg/L, total salt content 54000mg/L, pH 6 and wastewater flow 10t/h. NaOH is added into the regulating tank until the pH value is 7, and the wastewater flows into a primary aerobic unit, wherein the unit adopts a biological contact oxidation tank process, the strain is high-efficiency salt-tolerant bacteria GXNYJ-DL-1, the dissolved oxygen is controlled to be more than 2mg/L, the wastewater residence time is 80h, and the COD of the final effluent is 980mg/L, and the wastewater flows into an anaerobic unit. Controlling the dissolved oxygen of the anaerobic unit below 0.2mg/L, keeping the wastewater at the temperature of 30 ℃ for 140h, adopting methane gas for blowing off, and enabling hydrogen sulfide generated by anaerobic to be separated from a water phase, enter a gas phase along with methane and enter a tail gas treatment section, wherein the volume ratio of methane blowing off to wastewater is 7:1. The effluent from the anaerobic unit enters a secondary aerobic unit, COD of the inlet water is 718mg/L, the sulfate content is 2650mg/L, the unit adopts a conventional activated sludge method, conventional biological flora is used, the residence time of wastewater is 24h, COD of the final outlet water is 180mg/L, and the outlet water enters a subsequent advanced treatment section.
Advanced oxidation in the advanced treatment section adopts ozone catalytic oxidation, effluent is further treated by BAF, and COD of the final effluent is less than or equal to 60mg/L, and total phosphorus is about 5mg/L. The post-phosphorus removal adopts primary chemical phosphorus removal, the chemical agents are ferric chloride and calcium hydroxide which accord with the agents, the adding amount of the ferric chloride is 35mg/L, the adding amount of the calcium hydroxide is 15mg/L, COD (chemical oxygen demand) of post-phosphorus removal effluent is less than or equal to 60mg/L, sulfate is about 1750mg/L, the total salt amount is 5045 mg/L, and the total phosphorus is less than or equal to 0.5mg/L, so that the standard emission is realized.
The sludge generated by the primary aerobic unit, the secondary aerobic unit and the BAF unit is conveyed to a sludge anaerobic oxidation unit by a pump except for normal backflow, and the sludge is converted into methane gas by sludge anaerobic oxidation and is recovered to a methane storage tank.
And (3) absorbing tail gas generated by anaerobic treatment by using lean amine solution N-methyldiethanolamine to obtain methane with purity of 95%, storing the methane into a methane tank, partially recycling the methane, and partially utilizing the methane as a product. After the solvent absorption section absorbs and saturates, the rich amine liquid enters a solvent regeneration tower for regeneration, the regenerated lean amine liquid returns to the solvent absorption section, the escaped gas is hydrogen sulfide with the purity of about 94%, the hydrogen sulfide enters a WSA (wireless sensor array) acid making process, and finally sulfuric acid with the concentration of about 31% is prepared and is recycled as a raw material of the diacid process.
Sequence listing
<110> China petrochemical Co., ltd
China Petroleum & Chemical Corporation Dalian Petrochemical Research Institute
<120> a strain of highly salt tolerant bacteria and use thereof
<130> New patent application
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1386
<212> DNA
<213> Halomonas nigrificans
<400> 1
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gccacatcgg gactgagaca cggcccgaac tcctacggga ggcagcagtg gggaatattg 300
gacaatgggc gcaagcctga tccagccatg ccgcgtgtgt gaagaaggcc ttcgggttgt 360
aaagcacttt cagcgaggaa gaacgcctag tggttaatac ccattaggaa agacatcact 420
cgcagaagaa gcaccggcta actccgtgcc agcagccgcg gtaatacgga gggtgcgagc 480
gttaatcgga attactgggc gtaaagcgcg cgtaggtggc ttgataagcc ggttgtgaaa 540
gccccgggct caacctggga acggcatccg gaactgtcaa gctagagtgc aggagaggaa 600
ggtagaattc ccggtgtagc ggtgaaatgc gtagagatcg ggaggaatac cagtggcgaa 660
ggcggccttc tggactgaca ctgacactga ggtgcgaaag cgtgggtagc aaacaggatt 720
agataccctg gtagtccacg ccgtaaacga tgtcgaccag ccgttgggtg cctagcgcac 780
tttgtggcga agttaacgcg ataagtcgac cgcctgggga gtacggccgc aaggttaaaa 840
ctcaaatgaa ttgacggggg cccgcacaag cggtggagca tgtggtttaa ttcgatgcaa 900
cgcgaagaac cttacctact cttgacatct acagaagccg gaagagattc tggtgtgcct 960
tcgggaactg taagacaggt gctgcatggc tgtcgtcagc tcgtgttgtg aaatgttggg 1020
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cttgtacaca ccgcccgtca caccatggga gtggactgca ccagaagtgg ttagcctaac 1380
gcaaga 1386
Claims (14)
1. High salt-tolerant bacteriaHalomonas nigrificans) GXNYJ-DL-1 is preserved in China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC No.20350 in the year 7 and 13 of 2020.
2. The culture method of the high salt tolerant bacteria GXNYJ-DL-1, which comprises three stages of strain activation, seed liquid culture and intermittent aeration culture, and comprises the following specific steps:
(1) Strain activation: inoculating the strain GXNYJ-DL-1 into a broth peptone solid culture medium with the salt content of 1-5% by adopting a plate streaking method, and culturing in an incubator at the temperature of 28-35 ℃ for 48-72 hours;
(2) Seed liquid culture: after the strain activation is finished, the activated strain in the flat plate is selected and inoculated into a conical flask containing broth peptone liquid culture medium, wherein the salt content in the liquid culture medium is 1-5%, sulfate accounts for more than 50%, shake culture is carried out on a shaking table, the temperature is 30-35 ℃, the rotating speed is 100-200 rpm, and the culture time is 24-96 hours;
(3) Intermittent aeration culture: adding prepared salt-containing wastewater or wastewater to be treated into a reactor provided with an aeration device, wherein the salt content is 1-5%, the sulfate accounts for more than 50%, inoculating liquid seed liquid according to the volume ratio of 3-20%, controlling the pH value to be 6-9, performing indirect aeration according to the relation of the aeration duration and the aeration stopping duration of 2:1-1:2, and the total culture period is 72-144 h; s production by means of sulfate reduction reactions occurring upon indirect aeration 2- Improving S resistance of strain GXNYJ-DL-1 2- Toxicity ability, and other miscellaneous bacteria are prevented from breeding.
3. The use of the high salt tolerant bacteria GXNYJ-DL-1 according to claim 1 for removing COD from high salt organic wastewater.
4. The use according to claim 3, wherein the high salt tolerant bacteria GXNYJ-DL-1 has a salt tolerance content of up to 25wt%, tolerates S 2- The concentration is up to 300mg/L.
5. The process for treating high-sulfate organic wastewater by using the high-salt-tolerance bacterium GXNYJ-DL-1, which is characterized by comprising a front-end treatment section, a deep treatment section, a sludge treatment section and a tail gas treatment section;
wherein the front-end treatment section sequentially comprises pH adjustment, primary aerobic treatment, anaerobic treatment, stripping treatment and secondary aerobic treatment; the primary aerobic treatment is carried out on the high-sulfate organic wastewater by adopting the high-salt-tolerance bacterium GXNYJ-DL-1;
the advanced treatment section sequentially comprises advanced oxidation, BAF and post dephosphorization;
the sludge treatment section is used for conveying redundant sludge generated by the primary aerobic unit, the secondary aerobic unit and the BAF unit to a sludge anaerobic unit by a pump, and converting most of activated sludge into methane gas through sludge anaerobic oxidation and recycling the methane gas to a methane storage tank;
the tail gas treatment section comprises solvent absorption, solvent regeneration and sulfuric acid preparation by a WSA wet method.
6. The treatment process according to claim 5, wherein in the front-end treatment section, the pH adjustment is to adjust the pH of the wastewater to 6.5 to 7.5; the first-stage aerobic effluent enters an anaerobic section, sulfate is reduced into hydrogen sulfide under the action of sulfate reducing bacteria, and methane is used as blowing and degassing to carry out stripping, so that the generated hydrogen sulfide is carried out a wastewater system; the anaerobic effluent enters a secondary aerobic treatment section, COD is further removed by adopting conventional sludge strains, and the secondary aerobic effluent enters a deep treatment section.
7. The treatment process according to claim 5, wherein the advanced oxidation of the advanced treatment section is to remove refractory organics in the wastewater while improving the biodegradability of the wastewater, and then further remove COD in the wastewater via BAF and perform a filtration function; the post dephosphorization is carried out by adding the medicament to carry out two-stage chemical dephosphorization, and finally the wastewater meeting the discharge standard is obtained.
8. The process of claim 5, wherein the primary aerobic is a biological contact oxidation process or an MBBR process.
9. The treatment process according to claim 5, wherein dissolved oxygen in the anaerobic treatment wastewater is controlled below 0.2mg/L, methane gas is adopted for stripping, the residence time of the wastewater is 12-144 h, the temperature is 25-35 ℃, and the volume ratio of methane stripping gas to wastewater is not less than 5:1.
10. The process of claim 5, wherein the advanced oxidation is selected from one of ozone oxidation, electrocatalytic oxidation and Fenton oxidation.
11. The treatment process according to claim 5, wherein the post-dephosphorization is two-stage chemical dephosphorization, and the chemical agent is a compound agent of ferric chloride and calcium hydroxide; wherein the adding amount of the first-stage dephosphorization ferric chloride is 34 mg/L-680 mg/L, and the adding amount of the calcium hydroxide is 10 mg/L-300 mg/L; wherein the adding amount of the secondary dephosphorization ferric chloride is 10 mg/L-200 mg/L, and the adding amount of the calcium hydroxide is 4 mg/L-80 mg/L.
12. The process of claim 5, wherein the absorbent of the tail gas treatment section is selected from one of monoethanolamine, diethanolamine, diisopropanolamine and N-methyldiethanolamine.
13. The process of claim 12 wherein the absorbent of the tail gas treatment section is N-methyldiethanolamine.
14. The process of claim 5, wherein the WSA wet sulfuric acid is prepared by incineration, conversion and condensation processes: incinerating the gas separated by regenerating the solvent, wherein H 2 S combustion to generate SO 2 ，SO 2 Conversion to SO under the action of a catalyst 3 ，SO 3 And condensing the water vapor in a condenser to generate sulfuric acid.
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