CN110283744B - Achromobacter strain capable of tolerating toxicity of zinc ions and application thereof - Google Patents
Achromobacter strain capable of tolerating toxicity of zinc ions and application thereof Download PDFInfo
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Abstract
The invention relates to a strain of Achromobacter sp which is resistant to zinc ion toxicity and application thereof. The invention provides a strain of Achromobacter sp with zinc ion tolerance, and the preservation number is CGMCC No.2964. When the concentration of zinc ions in sewage is in the range of 0-50 mg/L, the strain can take nitrate nitrogen as the sole nitrogen source, the removal rate of nitrate nitrogen can reach 100% under the aerobic condition, and the removal rate of total nitrogen can reach 100%. The method has the advantages of convenience in operation, high denitrification efficiency and the like, and has important significance in optimizing a sewage biological treatment system and weakening the influence of zinc ions on the sewage denitrification system.
Description
Technical Field
The invention belongs to the field of sewage biological treatment, and particularly relates to Achromobacter capable of tolerating the toxicity of zinc ions under an aerobic condition and synchronously removing nitrate nitrogen in a water body and application thereof in sewage treatment.
Background
Along with the acceleration of urban and industrialized production in China, industrial production is continuously developed, a large amount of industrial wastewater is generated, and great threat is generated to human life and health. Since the industrial wastewater treatment capacity of China cannot keep up with the scale of industrial water, part of industrial wastewater is still directly discharged into a town sewage pipe network without being treated. Zinc ions are widely used as an important industrial material in chemical, metallurgical, electroplating, leather and paint industries, which produce a large amount of zinc-containing wastewater which inevitably enters town sewage treatment plants. Research shows that Zn (II) concentration exceeding 10mg/L can result in N in traditional denitrification 2 The reduction process, denitrification rate and denitrification microbial activity of O are obviously inhibited, and meanwhile, the microbial community structure is changed, so that the output of the town sewage treatment system is influencedWater quality.
In recent years, the discovery of aerobic denitrification (i.e., the process of reducing nitrate nitrogen to a nitrogen-containing gas under aerobic conditions) provides a theoretical basis for achieving simultaneous nitrification and denitrification in a single aerobic reactor. Since the successful separation of the first aerobic denitrifying bacteria T.pantotripha, more efficient aerobic denitrifying strains have been separated in recent years and are applied to practical sewage treatment systems. The aerobic denitrification is used as a novel biological denitrification technology, and has been rapidly developed in recent years due to the advantages of simple process, good denitrification effect, no need of adding acid or alkali and the like.
Aiming at the problem that zinc-containing wastewater enters domestic sewage and Zn (II) and nitrate nitrogen coexist, how to effectively avoid the influence of zinc ions on nitrate nitrogen removal becomes a focus of attention, so that aerobic denitrifying bacteria with resistance to zinc ion toxicity are discovered and applied to sewage treatment, the potential risk of zinc ions on the sewage biological denitrification process can be effectively reduced, and the method has important significance for guaranteeing the normal operation of urban sewage treatment systems.
Disclosure of Invention
The invention aims to provide an efficient aerobic denitrification bacterial strain resistant to zinc ion toxicity, which strengthens the biological denitrification effect of a sewage treatment system to reduce the influence of the existence of high-concentration zinc ions on the biological denitrification process of sewage treatment.
The Achromobacter sp provided by the invention is an aerobic denitrifying bacterium with the capability of removing nitrate nitrogen in a single aerobic environment.
The invention is realized by the following technical scheme:
the Achromobacter sp with zinc ion tolerance provided by the invention has a collection number of CGMCC No.2964.
Achromobacter sp., as described above, characterized in that: under the condition of different zinc ion concentration levels, the strain Achromobacter sp can still maintain the complete cell structure, and the cell membrane is not destroyed.
Achromobacter sp., as described above, characterized in that: the strain can perform aerobic denitrification by taking nitrate nitrogen as a nitrogen source and taking organic matters as a carbon source under the condition of high-concentration zinc ions, so that the nitrate nitrogen is removed.
Use of the above-described Achromobacter sp for sewage treatment, characterized in that: achromobacter sp is added into the zinc-containing sewage, and a proper amount of carbon source is added for aeration, so that nitrate nitrogen in the sewage can be removed.
The method as described above, characterized by: the temperature in the zinc-containing sewage is controlled to be 30 ℃, the pH value is controlled to be 7.5, the dissolved oxygen is controlled to be 6mg/L, and the initial C/N ratio is controlled to be 4.
The method as described above, characterized by: when the concentration of zinc ions in the sewage is in the range of 0-50 mg/L, the removal rate of nitrate nitrogen can reach 100%, and the removal rate of total nitrogen can also reach 100%.
The beneficial effects of the invention are as follows:
(1) According to the invention, the Achromobacter sp can perform aerobic denitrification by taking nitrate nitrogen as a nitrogen source and organic matters as a carbon source under the condition of high concentration zinc ions, so that the nitrate nitrogen can be efficiently removed; solves the problem that the biological denitrification in the traditional wastewater treatment needs to adopt anoxic denitrification and aerobic nitrification sectional treatment; in addition, the process flow is simplified, and the equipment and investment cost is saved, so that the method has great economic benefit and environmental protection benefit;
(2) The bacterial strain is inoculated into biological treatment of zinc-containing sewage, and preferably, the removal rate of nitrate nitrogen in 25 hours can reach 100% in the presence of 0-50 mg/L Zn (II), which indicates that the bacterial strain has stronger zinc ion toxicity tolerance and aerobic denitrification capability. The characteristics greatly enhance the practicability of the strain in town sewage treatment systems.
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FIG. 1 is a scanning electron microscope image of Achromobacter in the presence of zinc ions.
FIG. 2 shows the aerobic denitrification profile of Achromobacter in the presence of 10mg/L Zn (II).
FIG. 3 shows aerobic denitrification characteristics of Achromobacter in the presence of 25mg/L Zn (II).
FIG. 4 shows aerobic denitrification characteristics of Achromobacter in the presence of 50mg/L Zn (II).
Detailed Description
The present invention will be further illustrated with reference to the following specific examples, but the present invention is not limited to the following examples.
In the following examples, conventional methods are used unless otherwise specified.
In the following examples, the percentages are mass percentages unless otherwise indicated.
Example 1 observation of Achromobacter apparent morphology in the Presence of Zinc ions Using scanning Electron microscopy
The highly efficient aerobic denitrifying strain used in this experiment was Achromobacter sp, which was isolated from landfill leachate treatment systems.
Achromobacter sp was inoculated into 1L of LB medium (5 g/liter of NaCl, 10 g/liter of tryptone, 5 g/liter of yeast extract), and enrichment culture was performed while preventing invasion of infectious microbes and maintaining the growth activity of the bacterial cells. Centrifuging the cultured bacterial liquid, washing with 0.5% NaCl three times to obtain optical density OD 600 1-2. The bacterial suspensions were then inoculated in 10% inoculum size respectively into LB medium containing 25mg/L Zn (II), sampled after 8h, centrifuged at 8000rpm and 4℃for 5min to remove the supernatant, and washed twice with PBS (phosphate buffer). Fixing with 2.5% glutaraldehyde for 1 hr, gradient eluting with ethanol, dispersing in absolute ethanol, air drying with small amount of dropwise onto silicon wafer, and observing cell morphology under Scanning Electron Microscope (SEM).
As shown in FIG. 1, when the concentration of zinc ions was 25mg/L, achromobacter sp exhibited a complete and smooth surface, indicating that the cell morphology of the strain was highly resistant to zinc ions.
EXAMPLE 2 aerobic denitrification Property of Achromobacter in the presence of 10mg/L Zn (II)
Denitrification Performance test Medium (BM) formulation was per liter of water: 8.45g CH 3 COONa,0.63g NH 4 Cl,0.61g NaNO 3 ,1.76g K 2 HPO 4 ·3H 2 O,0.20g MgSO 4 ·7H 2 O,0.02g CaCl 2 ,0.005gFeSO 4 ·7H 2 O,0.1mL trace element solution. The pH of the medium was adjusted to 7.5 and sterilized at 121℃for 30min.
Achromobacter sp was inoculated into BM medium containing 10mg/L Zn (II), shake-cultured at 30℃and 150rpm, and 100. Mu.L of headspace gas was extracted with a valve needle every 5 hours for N determination 2 O, 2mL of gas was drawn with a sterile syringe and injected into a 2L pure helium bag for NO determination. Meanwhile, 2mL of bacterial suspension is extracted by a sterile syringe, the bacterial solution is centrifuged at 8000rpm for 5min at 4 ℃, and the supernatant is taken for analyzing the concentration of ammonia nitrogen, nitrate nitrogen and nitrite nitrogen.
As a result, as shown in FIG. 2, 100mg/L of nitrate nitrogen was utilized by the strain immediately after inoculation and was completely consumed within 10 hours, and the average removal rate of nitrate nitrogen was 10.0mg/L/h. As the nitrate nitrogen is reduced, the nitrite nitrogen rapidly accumulates and reaches a maximum of 32.1mg/L at 10 hours and is then reduced at 20 hours. N (N) 2 O also gradually accumulated, reaching the highest value of 0.42mg/L at 10h, and was then fully reduced at 25 h. The highest value of the accumulation amount of the intermediate NO in the denitrification process is 82.5 mug/L, and the accumulation amount is small, which indicates that the strain can better play the role of aerobic denitrification in the presence of 10mg/L Zn (II).
EXAMPLE 3 aerobic denitrification Property of Achromobacter in the Presence of 25mg/L Zn (II)
Achromobacter sp were inoculated into BM medium containing 25mg/L Zn (II) and tested for aerobic denitrification performance. As a result, as shown in FIG. 3, 100mg/L of nitrate nitrogen was utilized by the strain immediately after inoculation and was completely consumed within 25 hours, and the average removal rate of nitrate nitrogen was 4mg/L/h. As the nitrate nitrogen is reduced, the nitrite nitrogen gradually accumulates and reaches a maximum of 33.4mg/L at 15 hours and is then reduced at 25 hours. N (N) 2 O also gradually accumulated, reaching the highest value of 1.01mg/L at 20h, and was then completely reduced at 45 h. It has to be noted that in this case the accumulation of NO as intermediate product of the denitrification processThe highest value of (2) is 83.1 mug/L, which is only 0.083% of the nitrate nitrogen removal amount. From this, it can be seen that the strain still has high-efficiency aerobic denitrification capability under 25mg/L Zn (II) condition.
EXAMPLE 4 aerobic denitrification Property of Achromobacter in the presence of 50mg/L Zn (II)
Achromobacter sp were inoculated into BM medium containing 50mg/L Zn (II) and tested for aerobic denitrification performance. As a result, as shown in FIG. 4, 100mg/L of nitrate nitrogen was utilized by the strain immediately after inoculation and was completely consumed within 25 hours, and the average removal rate of nitrate nitrogen was 4.0mg/L/h. As the nitrate nitrogen is reduced, the nitrite nitrogen gradually accumulates and reaches a maximum of 33.5mg/L at 20 hours and is then reduced at 30 hours. At the same time N 2 O also gradually accumulated, reaching the highest value of 1.23mg/L at 25h, and was then fully reduced at 35 h. It must be noted that the highest value of the accumulation of NO as an intermediate product of the denitrification process is 17.3. Mu.g/L, and only 0.017% of the removal amount of nitrate nitrogen is obtained. From this, it can be seen that the strain still has high-efficiency aerobic denitrification capability under the condition of 50mg/L Zn (II).
EXAMPLE 5 aerobic denitrification Property of Achromobacter in the presence of 100mg/L Zn (II)
Achromobacter sp were inoculated into BM medium containing 100mg/L Zn (II) and tested for aerobic denitrification performance. The growth of the strain is obviously inhibited, and nitrate nitrogen is hardly removed, so that 100mg/L Zn (II) is the upper limit concentration of the strain, and the strain does not have the capacity of aerobic denitrification under the condition. However, as can be seen from the above examples 1-4, the bacterial strain is inoculated into biological treatment of zinc-containing sewage, and the removal rate of nitrate nitrogen can reach 100% in the presence of 0-50 mg/L Zn (II), which shows that the bacterial strain has stronger zinc ion toxicity resistance and aerobic denitrification capability (see Table 1).
TABLE 1 denitrification Properties of Achromobacter in the Presence of different concentrations of Zn (II)
EXAMPLE 6 aerobic denitrification Property of Achromobacter and Pseudomonas stutzeri in the presence of 50mg/L Zn (II)
Achromobacter sp and Pseudomonas stutzeri (P.stutzeri) were inoculated into BM medium containing 50mg/LZn (II) and tested for aerobic denitrification performance. As shown in Table 2, 100mg/L of nitrate nitrogen was immediately available to Achromobacter immediately after inoculation, the maximum removal rate of nitrate nitrogen was 5.8.+ -. 0.5mg/L/h, and the final product was N 2 Indicating that the denitrification process is not inhibited. While Pseudomonas stutzeri has a maximum nitrate nitrogen removal rate of only 2.9+ -0.4 mg/L/h in the presence of 50mg/L Zn (II), the rate is only half that of Achromobacter although the final denitrification product is also nitrogen. As is well known, pseudomonas stutzeri is a typical denitrifying strain, and comparison shows that the Achromobacter provided by the invention has high tolerance to Zn (II) while performing efficient aerobic denitrification (see Table 2).
TABLE 2 comparison of aerobic denitrification Properties of Achromobacter and Pseudomonas stutzeri in the presence of 50mg/L Zn (II)
EXAMPLE 7 use of Achromobacter in Zinc-containing wastewater
Adding Achromobacter into an activated sludge system to treat zinc-containing wastewater, and continuously aerating to keep the dissolved oxygen of the system to be 6mg/L, wherein the water quality of the wastewater is as follows: the pH is 7.5, the nitrate nitrogen content is 100mg/L, and the Zn (II) content is 50mg/L. The results are shown in Table 2, and the total nitrogen removal rate is lower in the normal activated sludge system under the aerobic condition; the aerobic denitrification performance is affected by adding 50mg/L Zn (II). However, the nitrate nitrogen removal rate of the system after the fortification with Achromobacter was 96.4% for 36 hours, and the total nitrogen removal rate was 90.5% (see Table 3).
TABLE 3 biological denitrification results of Achromobacter applied in activated sludge systems
Therefore, aiming at the problem that zinc-containing wastewater enters domestic sewage and Zn (II) and nitrate nitrogen coexist, the strain can effectively avoid the influence of zinc ions on nitrate nitrogen removal, can be widely applied to sewage treatment, effectively reduces the potential risk of zinc ions on the sewage biological denitrification process, and has important significance for guaranteeing the normal operation of town sewage treatment systems.
It should be noted that the above embodiments are only for further detailed description of the present invention and are not intended to limit the present invention, and those skilled in the art can make various modifications or changes without departing from the spirit and scope of the present invention, and still fall within the protection scope of the appended claims.
Claims (1)
1. An application of Achromobacter strain tolerant to zinc ion toxicity, which is characterized in that: the biological denitrification effect of the sewage treatment system is enhanced by an aerobic denitrifying bacterium Achromobacter sp, so that the influence of the existence of excessive zinc ions on the biological denitrification process of sewage treatment can be reduced;
the strain can perform aerobic denitrification by taking nitrate nitrogen as a nitrogen source and organic matters as a carbon source under different zinc ion concentration levels, so that the nitrate nitrogen is removed;
the preservation number of the strain is CGMCC No.2964, and the strain has better tolerance to zinc ions with different concentration levels;
achromobacter sp is added into zinc-containing sewage, and a proper amount of carbon source is added for aeration, so that nitrate nitrogen in the sewage can be removed;
controlling the temperature in the zinc-containing sewage to be 30 ℃, the pH value to be 7.5 and the dissolved oxygen to be 6mg/L;
controlling the initial C/N ratio in the zinc-containing sewage to be 4;
the concentration range of zinc ions contained in the sewage is 10-50 mg/L, the removal rate of nitrate nitrogen can reach 100% in an aerobic environment, and the removal rate of total nitrogen can also reach 100%.
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| US5593592A (en) * | 1993-12-16 | 1997-01-14 | Kagawa; Haruo | Biodegradation process for treating organic wastewater |
| CN101560486A (en) * | 2009-06-03 | 2009-10-21 | 北京大学 | Achromobacter xylosoxidans strain for biological denitrificaion and application thereof |
| CN105062926A (en) * | 2015-08-26 | 2015-11-18 | 华中农业大学 | Achromobacter for heavy metal cadmium pollution treatment and application thereof |
| CN105802890A (en) * | 2016-04-27 | 2016-07-27 | 中南民族大学 | Achromobacter CZ207 strain capable of resisting heavy metal and promoting plant growth and application thereof |
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