CN109943497B - Pseudomonas stutzeri with aerobic denitrification function and application thereof - Google Patents
Pseudomonas stutzeri with aerobic denitrification function and application thereof Download PDFInfo
- Publication number
- CN109943497B CN109943497B CN201910071406.1A CN201910071406A CN109943497B CN 109943497 B CN109943497 B CN 109943497B CN 201910071406 A CN201910071406 A CN 201910071406A CN 109943497 B CN109943497 B CN 109943497B
- Authority
- CN
- China
- Prior art keywords
- nitrogen
- pseudomonas stutzeri
- nitrite
- aerobic
- nitrate
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
- Y02W10/00—Technologies for wastewater treatment
- Y02W10/10—Biological treatment of water, waste water, or sewage
Landscapes
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Purification Treatments By Anaerobic Or Anaerobic And Aerobic Bacteria Or Animals (AREA)
Abstract
The invention belongs to the field of microorganisms, and discloses a Pseudomonas stutzeri ZH-14(Pseudomonas stutzeri ZH-14) with an aerobic denitrification function, wherein the preservation number is CCTCC NO: M2018730. The pseudomonas stutzeri can be applied to degrading ammonia nitrogen, nitrate nitrogen, nitrite nitrogen or COD in nitrogen-containing sewage. Has the advantages that: pseudomonas stutzeri ZH-14 has both aerobic denitrification function and heterotrophic nitrification function, and contains very low content of nitrite nitrogen accumulation in the degradation process; nitrite can be directly utilized for growth and metabolism; under the environment with low carbon-nitrogen ratio, the activity of periplasmic nitrate reductase of the pseudomonas stutzeri ZH-14 is still strong, the aerobic expression is less influenced by the type of the used carbon source, and the problem of the carbon source in the process of treating the nitrogen-containing sewage by using aerobic denitrifying bacteria is solved.
Description
Technical Field
The invention relates to the field of microorganisms, in particular to pseudomonas stutzeri with an aerobic denitrification function and application thereof.
Background
In recent years, with the improvement of the scale breeding degree of livestock and poultry, the breeding pollution problem becomes more and more prominent, and becomes an important bottleneck for limiting the development of animal husbandry. The biogas slurry is a product of livestock and poultry sewage treated by anaerobic biogas engineering, and because the biogas slurry contains more organic matters and pollutants such as nitrogen, phosphorus and the like, the biogas slurry cannot be directly discharged, and the influence on the environment can be avoided only by further treatment. The current common biogas slurry treatment process comprises various means such as physics, chemistry, biology and the like. The technology for treating biogas slurry by adopting microorganisms plays an important role in biogas slurry purification due to the advantages of small occupied area, low cost and the like. However, researches show that after microbial treatment, although ammonia nitrogen content in biogas slurry is greatly degraded, Total Nitrogen (TN) removal rate is still low, and particularly, biochemical treatment effluent still contains a large amount of nitrate nitrogen, so that potential environmental risks are caused.
Traditional nitrate nitrogen removal requires a strictly anaerobic environment and is more susceptible to oxygen or nitrite inhibition. With the discovery of an aerobic denitrification path, a new idea is provided for removing nitrate nitrogen under an aerobic condition. In recent years, research at home and abroad continuously screens and obtains aerobic denitrifying strains from various sources such as activated sludge of domestic sewage treatment plants, bay sediments, agricultural fertilizer soil and the like, and the aerobic denitrifying strains have good effects on removing nitrate nitrogen and TN. Although a certain research is carried out on the screening of aerobic denitrifying bacteria at present, the research on the targeted screening of strains suitable for denitrification treatment from biogas slurry and sludge in a pig farm is still less, and particularly, reports on the addition of suitable microbial inoculum when the biogas slurry is actually applied are made. The existing aerobic denitrifying bacteria are easily influenced by high-concentration organic matters or ammonia nitrogen and have no influence on OD (oxygen-to-density) in water600TN, nitrate nitrogen, nitrite nitrogen or ammonia nitrogen are not ideal. Conventional biological denitrification processes typically include autotrophic nitrification and heterotrophic nitrificationIn the first stage, ammonia nitrogen is oxidized into nitrite by ammonia oxidizing bacteria under aerobic condition, and ammonium nitrite is oxidized into nitrate by nitrifying bacteria; the second stage converts nitrite and nitrate to N2 gas by denitrifying bacteria under anaerobic conditions. Enzymes involved in the reaction during the whole process include Ammonia Monooxygenase (AMO), hydroxylamine oxidase (HAO), periplasmic nitrate reductase (NAP), nitrite reductase (NIR), Nitric Oxide Reductase (NOR), nitrous oxide reductase (NOS), and the like. Not only is this entire reaction time consuming, but there is a low nitrification rate and the complexity of the sewage treatment system separating aerobic and anoxic zones.
Disclosure of Invention
One of the purposes of the invention is to provide pseudomonas stutzeri which is screened from the pig farm biogas slurry, can efficiently degrade ammonia nitrogen, nitrate nitrogen, nitrite nitrogen and COD, has high tolerance to high organic load and simultaneously has an aerobic denitrification function and a heterotrophic nitrification function.
The invention also aims to provide a method for applying the screened aerobic denitrifying bacteria to pig farm biogas slurry treatment, wherein in the ammonia nitrogen degradation process, the obvious accumulation of nitrite nitrogen and nitrate nitrogen is not detected, the synchronous removal of COD and ammonia nitrogen is realized, and the defect that the traditional biochemical treatment cannot tolerate high organic load during the oxidation of ammonia nitrogen is overcome.
In order to achieve the purpose, the invention adopts the technical scheme that: the Pseudomonas stutzeri ZH-14(Pseudomonas stutzeri ZH-14) with the aerobic denitrification function has the preservation number of M2018730 (CCTCC NO), the preservation time of 2018, 10 and 31 days, the preservation unit is China Center for Type Culture Collection (CCTCC), and the preservation unit address is China, Wuhan university. The 16srDNA of Pseudomonas stutzeri ZH-14(Pseudomonas stutzeri ZH-14) is shown below:
AAAATGCGCAGCTAACACATGCAAGTCGAGCGGATGAGTGGAGCTTGCTCCATGATTCAGCGGCGGACGGGTGAGTAATGCCTAGGA ATCTGCCTGGTAGTGGGGGACAACGTTTCGAAAGGAACGCTAATACCGCATACGTCCTACGGGAGAAAGTGGGGGATCTTCGGACCTCACG CTATCAGATGAGCCTAGGTCGGATTAGCTAGTTGGTGAGGTAAAGGCTCACCAAGGCGACGATCCGTAACTGGTCTGAGAGGATGATCAGT CACACTGGAACTGAGACACGGTCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGGACAATGGGCGAAAGCCTGATCCAGCCATGCCG CGTGTGTGAAGAAGGTCTTCGGATTGTAAAGCACTTTAAGTTGGGAGGAAGGGCAGTAAGTTAATACCTTGCTGTTTTGACGTTACCAACA GAATAAGCACCGGCTAACTTCGTGCCAGCAGCCGCGGTAATACGAAGGGTGCAAGCGTTAATCGGAATTACTGGGCGTAAAGCGCGCGTAG GTGGTTCGTTAAGTTGGATGTGAAAGCCCCCGGGCTCAACCTGGGAACTGCATCCAAAACTGGCGAGCTAGAGTATGGCAGAGGGTGGTGG AAATTTCCTGTGTAGCGGTGAAATGCGTAGATAATAGGAAGGAACACCAGTGGCCGAAAGGCGACCACCCTGGCCTAATACTGACCACTGA AGGTGCGAAAGCGTGGGGAGCAAACAGGATTAGATACCTGGTAGTCACGCCGTAAACGATGTCGACTAGCCGTTGGGATCCTTGAGATCTT TAGTGGCGCAGCTAACGCATTAAGTCGACCGCCTGGGGGAGTACGGCCGCAAGGTTAAAACTCAAATGAATTGACGGGGGGCCCGCACAAG CGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGCCTTGACATGCAGAGAACTTTCCAGAGATGGATTGGTGCCTT CGGGAACTCTGACACAGGTGCTGCATGGCTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGTAACGAGCGCAACCCTTGTCCT TAGTTACCAGCACGTTAAGGTGGGCACTCTAAGGAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAGTCATCATGGCCCT TACGGCCTGGGCTACACACGTGCTACAATGGTCGGTACAAAGGGTTGCCAAGCCGCGAGGTGGAGCTAATCCCATAAAACCGATCGTAGTC CGGATCGCAGTCTGCAACTCGACTGCGTGAAGTCGGAATCGCTAGTAATCGTGAATCAGAATGTCACGGTGAATACGTTCCCGGGCCTTGT ACACACCGCCCGTCACACCATGGGAGTGGGTTGCTCCAGAAGTAGCTAGTCTAACCTTCGGGGGGACGGTACCACGAGATCTGG。
the application of pseudomonas stutzeri with an aerobic denitrification function in degrading ammonia nitrogen or nitrate nitrogen or nitrite nitrogen or COD in nitrogen-containing sewage.
Compared with the prior art, the invention has the beneficial effects that:
1. the pseudomonas stutzeri ZH-14 has an aerobic denitrification function and a heterotrophic nitrification function at the same time, and the degradation process contains the accumulation of nitrite nitrogen with extremely low content;
2. the pseudomonas stutzeri ZH-14 mainly has assimilation effect on the removal of ammonia nitrogen, and a small amount of the pseudomonas stutzeri ZH-14 realizes the removal of TN through heterotrophic nitrification-aerobic denitrification;
3. pseudomonas stutzeri ZH-14 can directly utilize nitrite for growth and metabolism;
4. the "bottleneck" phenomenon in the electron transport chain between cytochrome c and cytochrome aa3 in Pseudomonas stutzeri ZH-14 is overcome and electron flow can be transported to both the denitrifying enzyme and oxygen. Under the environment with low carbon-nitrogen ratio (less than 1), the activity of the periplasmic nitrate reductase in the Pseudomonas stutzeri ZH-14 is still strong, the aerobic expression is less influenced by the type of the used carbon source, and the problem of the carbon source in the process of treating the nitrogen-containing sewage by using the aerobic denitrifying bacteria is solved;
5. the activities of catalase, peroxidase and three phosphatases in the pseudomonas stutzeri ZH-14 are stronger, the metabolism is fast, and the growth and film formation time is short;
6. pseudomonas stutzeri ZH-14 has strong nitrite nitrogen tolerance and practical application potential;
7. the synchronous removal of COD and ammonia nitrogen can be realized after the pseudomonas stutzeri ZH-14 is inoculated, the defect that the traditional biochemical treatment cannot tolerate high organic load when ammonia nitrogen is oxidized is overcome, and the removal of biogas slurry pollutants is facilitated.
Drawings
FIG. 1 is a colony morphology of Pseudomonas stutzeri ZH-14 on a BTB identification plate;
FIG. 2 is a graph showing the growth curves of Pseudomonas stutzeri ZH-14 in different nitrogen media and the effects on TN, nitrate nitrogen, nitrite nitrogen and ammonia nitrogen contents;
FIG. 3 is a graph showing the effect of adding different final concentrations of Pseudomonas stutzeri ZH-14 on the index of pollutants in biogas slurry.
Detailed Description
The invention is explained in more detail below with reference to the figures and examples:
example 1: method for screening strains
Culture medium
Denitrifying enrichment medium: na (Na)3C6H5O7·2H2O 1g/L、KNO3 1g/L、KH2PO4 1.5g/L、Na2HPO40.42g/L、 MgSO4·7H2O 1.0g/L、CuSO4·5H2O 4.0mg/L、FeSO4·7H2O 0.7mg/L、FeCl3·6H2O 7.0mg/L、 CoCl3·6H2O 0.2mg/L、Na2MO4·2H2O 3.4mg/L、CaCl2·2H2O 2.0g/L，pH 7.2。
Bromothymol blue (BTB) identification medium: na (Na)3C6H5O7·2H2O 1g/L、KNO3 1.0g/L、FeSO4·7H2O 0.05 g/L、CaCl2·2H2O 0.2g/L、MgSO4·7H2O 1.0g/L、1％BTB1.0mL/L，pH 6.8。
Denitrifying performance determination culture medium: na (Na)3C6H5O7·2H2O 1.0g/L、KNO3 1.0g/L、KH2PO4 0.75g/L、 MgSO4·7H2O 0.4g/L，pH 7.2。
Ammonia nitrogen degradation performance determination culture medium: NH (NH)4Cl 0.382g/L、CH3COONa 0.7g/L、MgSO4·7H2O 0.05g/L、K2HPO4 0.2g/L、NaCl 0.12g/L、MnSO4·4H2O 0.01g/L、FeSO4 0.01g/L，pH 7.2。
Nitrite nitrogen degradation performance determination culture medium: NaNO2 0.15g/L、CH3COONa 0.3g/L、MgSO4·7H2O 0.05 g/L、K2HPO4 0.2g/L、NaCl 0.12g/L、MnSO4·4H2O 0.01g/L、FeSO4 0.01g/L，pH 7.2。
When the plate culture medium is prepared, 1.5-2.0% of agar powder is required to be added.
Water quality index detection method
The water quality detection adopts national standard method, wherein the ammonia nitrogen detection method adopts 'determination of ammonia nitrogen in water-salicylic acid spectrophotometry' (GB 7481-87); the nitrate nitrogen content is detected by adopting a water quality nitrate nitrogen determination-ultraviolet spectrophotometry (HZ-HJ-SZ-0138); the TN detection method adopts 'determination of total nitrogen in water-alkaline potassium persulfate digestion ultraviolet spectrophotometry' (GB-11894-89); the determination of nitrite nitrogen adopts 'determination of nitrite nitrogen in water-spectrophotometry' (GB 7493-87); the COD detection method adopts a bichromate method (GB11914-89) for determining the chemical oxygen demand of water quality.
1. The source of the strain
The sludge comes from an aeration tank of an anoxic-aerobic sewage treatment system in the livestock farm of Zhejiang Anji adult husbandry with stable and good operation. The biogas liquid is taken from the tail end of a black film anaerobic fermentation tank of Jinfan ecological culture Limited company in Zhejiang, and the water quality indexes such as Chemical Oxygen Demand (COD), ammonia nitrogen, nitrate nitrogen, nitrite nitrogen, TN, pH and the like are shown in Table 1.
TABLE 1 index of biogas slurry pollutants in pig farm
2. Primary screen for aerobic denitrifying bacteria
1g of fresh activated sludge is taken and centrifuged and resuspended for 2 times by adopting 5mL of sterile 0.9% physiological saline, 1mL of the resuspended solution is taken and inoculated into 100mL of denitrification enrichment medium (300mL of triangular flask), and the culture is carried out under constant temperature shaking at 30 ℃ and 160 r/min. Every 1d of the culture medium is transferred into a new denitrification enrichment medium according to the inoculation amount of 1% (v/v), and after 4 times of continuous enrichment, the culture medium is coated on a BTB identification plate by adopting 10-fold gradient dilution. And (3) selecting single colonies with blue halos on the plate, carrying out continuous streak purification, and storing the purified single colonies on an LB inclined plane at 4 ℃.
3. Double screen for aerobic denitrifying bacteria
Activating the primary screening strain, inoculating to a denitrification performance determination culture medium, culturing in a constant temperature shaking incubator at 30 ℃ and 160r/min for 2d, and detecting indexes such as TN, nitrate nitrogen, nitrite nitrogen and the like. And screening out the optimal strain according to the TN and the nitrate nitrogen removal rate.
4. And (3) induction culture: after enrichment culture, 16 strains with obvious differences in colony morphology and blue halos around the colonies were selected from the BTB identification plate and named as ZH-1 to ZH-16, respectively. And (3) carrying out magnetization treatment on the screened strains, carrying out magnetization culture on the strains for 24-36 h under the field intensity of 250-300 mT, wherein the shake flask re-screening result of the denitrification performance identification culture medium is shown in Table 2. As can be seen from Table 2, compared to the remaining strains, TN and nitrate nitrogen removal effect of the strain ZH-14 was the best, reaching 50.73% and 99.99%, respectively, and only 0.031mg/L nitrite nitrogen accumulated after cultivation. Therefore, the strain ZH-14 is selected for subsequent identification and denitrification capability detection.
TABLE 2 preliminary screening of aerobic denitrifying bacteria
Example 2: strain morphology observation
As shown in FIG. 1, the purified preferred strain was applied to BTB solid medium, and the colony morphology of the strain was observed. The BTB plate identification result shows that ZH-14 can perform denitrification alkali production on a solid plate with potassium nitrate as a unique nitrogen source, and the pH value in a culture medium is increased, so that BTB is changed from green to blue. The strain ZH-14 is round, faint yellow, smooth in surface, non-transparent, convex and easy to pick up at the early stage of the single colony form on a BTB plate; the surface of the bacterial colony is yellowish and wrinkled in the later growth stage, and the surface area of the bacterial colony is increased.
Example 3: denitrification Performance test
1. Test of denitrification Performance of aerobic denitrifying bacteria
Selecting single colony, respectively inoculating to denitrifying performance determination culture medium, nitrite degradation performance determination culture medium and ammonia nitrogen degradation performance determination culture medium, and performing constant temperature shaking culture at 30 deg.C and 160r/min for 1d to obtain corresponding seed liquid.
Inoculating the bacterial liquid into 250mL triangular flasks filled with 100mL of denitrification performance determination culture medium, nitrite degradation performance determination culture medium and ammonia nitrogen degradation performance determination culture medium according to 1% (v/v), repeating the treatment for 3 times, and performing constant-temperature shaking culture at 30 ℃ and 160 r/min. Wherein the denitrification performance determination culture medium and the nitrite degradation performance determination culture medium are sampled every 6h, the ammonia nitrogen degradation performance determination culture medium is sampled every 4h, and OD is respectively detected600TN, nitrate nitrogen, nitrite nitrogen or ammonia nitrogen, etc. until each index tends to be stable. Recording denitrification performance determination culture medium and nitriteDegradation performance determination culture medium and ammonia nitrogen degradation performance determination culture medium in each time period D600TN, nitrate nitrogen, nitrite nitrogen and ammonia nitrogen content and record, the statistical results are shown in FIGS. 2A, 2B and 2C.
As can be seen from FIG. 2A, when Pseudomonas stutzeri ZH-14 was grown with potassium nitrate as the sole nitrogen source, the growth period entered logarithmic growth period at 24 hours and the cell concentration OD at 48 hours600Is 0.39. After the culture is carried out for 36h, the nitrate nitrogen is basically degraded, the degradation rates of the nitrate nitrogen and TN are respectively 100% and 41.76% in 48h, and the theoretical degradation rates respectively reach 2.85mg of nitrate nitrogen/(L.h) and 0.95mg of TN/(L.h). In the aspect of nitrite nitrogen, the nitrite nitrogen is slightly accumulated when being cultured for 30 hours, reaches 12.47mg/L and is then completely degraded; in the aspect of ammonia nitrogen, the content of the whole process is lower than 1.0mg/L, and no obvious accumulation exists.
As can be seen from FIG. 2B, when Pseudomonas stutzeri ZH-14 was cultured with sodium nitrite as the sole nitrogen source, the strain grew for 12h and entered the logarithmic phase, with the highest OD600Up to 0.23. The nitrite nitrogen is basically degraded within 24 hours, the degradation rate is only 0.21mg/L, and the degradation rate is 99.24%. The TN content is firstly reduced and then increased, and finally reaches 21.61mg/L, and the degradation rate is 19.0 percent; meanwhile, a small amount of ammonia nitrogen is accumulated in the culture process, and the highest concentration is 1.63mg/L when the culture is carried out for 30 hours.
The invention uses ammonium chloride as a unique nitrogen source to determine the growth condition of Pseudomonas stutzeri ZH-14 and the degradation condition of different nitrogen elements in different culture times. As can be seen from FIG. 2C, Pseudomonas stutzeri ZH-14 reached maximum absorbance (OD) at 8h in logarithmic growth phase and 20h in the case of culturing with ammonia nitrogen as the sole nitrogen source600)0.881, then enter the stationary phase. In the growth process of the strain, the degradation of ammonia nitrogen tends to be stable after being cultured for 24 hours, the residual amount is 6.64mg/L, and the degradation rate reaches 94.1%; the TN content is reduced from the initial 102.4mg/L to 97.2mg/L, and the degradation rate is 5.08%; meanwhile, the content of nitrate nitrogen and nitrite nitrogen is lower than 0.2mg/L in the whole process, and no obvious accumulation exists.
The invention shows that when the Pseudomonas stutzeri ZH-14 takes nitrate as the only nitrogen source, the nitrate nitrogen and TN are both obviously reduced along with the growth of the bacteria. Since TN measured in this test contains intracellular nitrogen content of the bacterial cells, it is presumed that this strain realizes denitrification mainly by aerobic denitrification. And the Pseudomonas stutzeri ZH-14 denitrification process contains extremely low content of nitrite nitrogen accumulation, and accords with the process that nitrate is firstly reduced into nitrite and then the nitrite concentration is reduced in the traditional denitrification process. The invention uses ammonia nitrogen as the only nitrogen source to discuss whether the Pseudomonas stutzeri ZH-14 has heterotrophic nitrification. Researches find that the ammonia nitrogen removal rate can reach 94.1 percent at most, and in the ammonia nitrogen degradation process, the invention does not detect obvious accumulation of nitrite nitrogen and nitrate nitrogen. The pseudomonas stutzeri ZH-14 mainly has assimilation effect on the removal of ammonia nitrogen, and a small amount of the pseudomonas stutzeri ZH-14 realizes the removal of TN through heterotrophic nitrification-aerobic denitrification. The result of the invention shows that Pseudomonas stutzeri ZH-14 can directly utilize nitrite to carry out growth metabolism, the degradation rate of TN in the research is 19% and shows the trend of rising first and then falling, which is probably related to the endogenous respiration of the bacterial strain in the later growth stage to decompose the endogenous nitrogen of the bacterial strain into ammonia nitrogen. The accumulation of ammonia nitrogen content at the later stage of culture can also be influenced by assimilation reduction in nitrite metabolism. The accumulation of nitrite in water can affect the growth and metabolic activity of microorganisms and cause damage to aquatic animals, the Pseudomonas stutzeri ZH-14 has no discovery of large accumulation of nitrite when ammonia nitrogen, nitrite nitrogen or nitrate nitrogen is used as a unique nitrogen source, and the nitrite nitrogen-containing compound grows well in a system with nitrite as a unique nitrogen source, so that the nitrite nitrogen-containing compound has nitrite nitrogen tolerance capability and practical application potential.
As can be seen from FIG. 2, when potassium nitrate was used as the sole nitrogen source, sodium nitrite was used as the sole nitrogen source, or ammonium chloride was used as the sole nitrogen source, and COD/TN was 1:1, Pseudomonas stutzeri ZH-14 was cultured for 48 hours, and almost no nitrite nitrogen was accumulated in the solution. Research shows that the carbon source is a limiting factor for the normal operation of the denitrification function of the strain. When the carbon source is insufficient, insufficient energy is provided for denitrification, which may result in the accumulation of intermediates, such as nitrite. The pseudomonas bacteria in the prior art have high carbon source demand, and when the carbon-nitrogen ratio is lower than 12, the growth and denitrification capability of the bacteria are affected. In the research, COD/TN of the biogas slurry is 1:1, which belongs to the level of low carbon-nitrogen ratio, but as can be seen from figure 2, no nitrite nitrogen is accumulated in the biogas slurry. The "bottleneck" phenomenon in the electron transport chain between cytochrome c and cytochrome aa3 in Pseudomonas stutzeri ZH-14 is overcome and electron flow can be transported to both the denitrifying enzyme and oxygen. Under the environment with low carbon-nitrogen ratio (less than 1), the activity of the periplasmic nitrate reductase in the pseudomonas stutzeri ZH-14 is still strong, the aerobic expression is less influenced by the type of the used carbon source, and the problem of the carbon source in the process of treating the nitrogen-containing sewage by using the aerobic denitrifying bacteria is solved.
2. Evaluation of denitrification effect of aerobic denitrifying bacteria on biogas slurry
Inoculating the strains into 500mL triangular flasks containing 200mL of sterilized biogas slurry until the final concentration of the strains is 105、106、107And 108CFU/mL, using sterile biogas slurry without inoculation as a control group, carrying out constant temperature shaking culture at 30 ℃ and 160r/min, sampling 10mL biogas slurry every 12h, and detecting the change conditions of nitrate nitrogen, nitrite nitrogen, ammonia nitrogen and COD. Each set of experiments contained 3 replicates. The inoculum sizes were recorded as 105、106、107And 108The statistical data for the values of nitrate nitrogen, nitrite nitrogen, ammonia nitrogen and COD for each time period in 4 sets of CFU/mL experiments are shown in figure 3.
As can be seen from FIG. 3A, 10 ammonia nitrogen removal was observed compared with the control group without administration of bacteria5The ammonia nitrogen degradation rate of the CFU/mL bacterium feeding group is the slowest, 106The ammonia nitrogen degradation rate of the CFU/mL bacterium feeding group is before 24h and 10 h7CFU/mL and 108CFU/mL dosing groups were consistent but slowed down as culture time was extended. After 48h of incubation, 107CFU/mL and 108The ammonia nitrogen degradation rates of the CFU/mL bacterium feeding group are 55.4% and 54.48% respectively. In the aspect of nitrate nitrogen removal (figure 3B), the pseudomonas stutzeri ZH-14 with high concentration is added to remove the nitrate nitrogen content of the biogas slurry fastest, the degradation stable period is reached after 24h, and finally 10 percent of pseudomonas stutzeri ZH-14 is added7The removal rate of nitrate nitrogen of the bacterium feeding group with the CFU/mL final concentration is up to 97.7%. In terms of COD removal (FIG. 3C), the bacterial group was dosed at low concentration (10)5CFU/mL) has lower COD degradation efficiency in the early culture period than other groupsHowever, with the prolonging of the culture time, the COD degradation difference of 4 groups of the strain feeding groups is not obvious at 48h, and the highest degradation rate is 107CFU/mL final concentration group, reach 77.9%. In terms of nitrite nitrogen (fig. 3D), the control group had no significant change in nitrite nitrogen content during the culture. However, the 4 test groups had different degrees of nitrite nitrogen accumulation in the first 24h compared to the control group, of which 106The accumulation condition of the CFU/mL bacterium feeding group is most obvious and reaches 9.95mg/L at most. The content of nitrite nitrogen gradually decreases with the prolonging of the culture time, wherein 10 is added7The final CFU/mL concentration in the test group was the lowest, 1.49 mg/L.
As can be seen from FIG. 3, the removal rates of ammonia nitrogen, nitrate nitrogen and COD in the bacteria feeding group are up to 54.48%, 97.7% and 77.9% respectively after 2d cultivation. The synchronous removal of COD and ammonia nitrogen can be realized after the pseudomonas stutzeri ZH-14 is inoculated, the defect that the traditional biochemical treatment cannot tolerate high organic load when ammonia nitrogen is oxidized is overcome, and the removal of biogas slurry pollutants is facilitated.
The pseudomonas stutzeri with the aerobic denitrification function is applied to degrading ammonia nitrogen or nitrate nitrogen or nitrite nitrogen or COD in nitrogen-containing sewage.
While one embodiment of the present invention has been described in detail, the description is only a preferred embodiment of the present invention and should not be taken as limiting the scope of the invention. All equivalent changes and modifications made within the scope of the present invention shall fall within the scope of the present invention.
Sequence listing
<110> Zhejiang province academy of agricultural sciences
<120> pseudomonas stutzeri with aerobic denitrification function and application thereof
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1445
<212> DNA
<213> 2 Ambystoma laterale x Ambystoma jeffersonianum
<400> 1
aaaatgcgca gctaacacat gcaagtcgag cggatgagtg gagcttgctc catgattcag 60
cggcggacgg gtgagtaatg cctaggaatc tgcctggtag tgggggacaa cgtttcgaaa 120
ggaacgctaa taccgcatac gtcctacggg agaaagtggg ggatcttcgg acctcacgct 180
atcagatgag cctaggtcgg attagctagt tggtgaggta aaggctcacc aaggcgacga 240
tccgtaactg gtctgagagg atgatcagtc acactggaac tgagacacgg tccagactcc 300
tacgggaggc agcagtgggg aatattggac aatgggcgaa agcctgatcc agccatgccg 360
cgtgtgtgaa gaaggtcttc ggattgtaaa gcactttaag ttgggaggaa gggcagtaag 420
ttaatacctt gctgttttga cgttaccaac agaataagca ccggctaact tcgtgccagc 480
agccgcggta atacgaaggg tgcaagcgtt aatcggaatt actgggcgta aagcgcgcgt 540
aggtggttcg ttaagttgga tgtgaaagcc cccgggctca acctgggaac tgcatccaaa 600
actggcgagc tagagtatgg cagagggtgg tggaaatttc ctgtgtagcg gtgaaatgcg 660
tagataatag gaaggaacac cagtggccga aaggcgacca ccctggccta atactgacca 720
ctgaaggtgc gaaagcgtgg ggagcaaaca ggattagata cctggtagtc acgccgtaaa 780
cgatgtcgac tagccgttgg gatccttgag atctttagtg gcgcagctaa cgcattaagt 840
cgaccgcctg ggggagtacg gccgcaaggt taaaactcaa atgaattgac ggggggcccg 900
cacaagcggt ggagcatgtg gtttaattcg aagcaacgcg aagaacctta ccaggccttg 960
acatgcagag aactttccag agatggattg gtgccttcgg gaactctgac acaggtgctg 1020
catggctgtc gtcagctcgt gtcgtgagat gttgggttaa gtcccgtaac gagcgcaacc 1080
cttgtcctta gttaccagca cgttaaggtg ggcactctaa ggagactgcc ggtgacaaac 1140
cggaggaagg tggggatgac gtcaagtcat catggccctt acggcctggg ctacacacgt 1200
gctacaatgg tcggtacaaa gggttgccaa gccgcgaggt ggagctaatc ccataaaacc 1260
gatcgtagtc cggatcgcag tctgcaactc gactgcgtga agtcggaatc gctagtaatc 1320
gtgaatcaga atgtcacggt gaatacgttc ccgggccttg tacacaccgc ccgtcacacc 1380
atgggagtgg gttgctccag aagtagctag tctaaccttc ggggggacgg taccacgaga 1440
tctgg 1445
Claims (3)
1. Pseudomonas stutzeri (pseudomonas stutzeri) with aerobic denitrification functionPseudomonas stutzeri) ZH-14, which is characterized in that the strain is preserved in China center for type culture Collection in 2018, 10 and 31 months, and the preservation number is CCTCC NO: M2018730.
2. The Pseudomonas stutzeri (Pseudomonas stutzeri) with aerobic denitrification function according to claim 1Pseudomonas stutzeri) The ZH-14 is applied to degrading ammonia nitrogen, nitrate nitrogen, nitrite nitrogen or COD in nitrogen-containing sewage.
3. The Pseudomonas stutzeri (Pseudomonas stutzeri) with aerobic denitrification function according to claim 2Pseudomonas stutzeri) The application of ZH-14 is characterized in that the specific application method comprises the following steps: (ii) the Pseudomonas stutzeri: (Pseudomonas stutzeri) ZH-14 is inoculated into the sewage to be treated, the inoculum size of the strain is 107And (3) culturing the CFU/mL at 30 ℃, and detecting the contents of nitrate nitrogen, nitrite nitrogen, ammonia nitrogen and COD in the water until the contents meet the requirements.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910071406.1A CN109943497B (en) | 2019-01-25 | 2019-01-25 | Pseudomonas stutzeri with aerobic denitrification function and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910071406.1A CN109943497B (en) | 2019-01-25 | 2019-01-25 | Pseudomonas stutzeri with aerobic denitrification function and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN109943497A CN109943497A (en) | 2019-06-28 |
| CN109943497B true CN109943497B (en) | 2021-02-12 |
Family
ID=67007317
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201910071406.1A Active CN109943497B (en) | 2019-01-25 | 2019-01-25 | Pseudomonas stutzeri with aerobic denitrification function and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN109943497B (en) |
Families Citing this family (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110386667A (en) * | 2019-07-18 | 2019-10-29 | 曲阜师范大学 | A kind of denitrification Pseudomonas stutzeri reinforcing anoxic pond sewage denitrification method |
| CN110452820A (en) * | 2019-08-16 | 2019-11-15 | 鹭滨环保科技（上海）股份有限公司 | A kind of screening of autotrophic denitrification bacterium and identification method |
| CN110669689A (en) * | 2019-10-15 | 2020-01-10 | 郑州大学 | Heterotypic nitrification-aerobic denitrification bacterium and application thereof |
| CN110791444B (en) * | 2019-10-24 | 2020-06-30 | 南京大学 | Pseudomonas stutzeri, composite microbial inoculum prepared from pseudomonas stutzeri and application of composite microbial inoculum |
| CN111826307A (en) * | 2019-10-29 | 2020-10-27 | 浙江大学 | Biological agent for improving mark-thickness survival rate of young penaeus vannamei |
| CN110982732B (en) * | 2019-11-13 | 2022-12-30 | 重庆理工大学 | Salt-tolerant and high-ammonia-nitrogen-resistant heterotrophic nitrification-aerobic denitrification composite microbial agent and preparation and application thereof |
| CN111534448B (en) * | 2019-12-25 | 2022-12-16 | 广东石油化工学院 | Heterotrophic nitrification-aerobic denitrification pseudomonas as well as culture method and application thereof |
| CN111607543B (en) * | 2020-06-28 | 2022-06-14 | 黄河三角洲京博化工研究院有限公司 | Pseudomonas stutzeri with aerobic denitrification function and application thereof |
| CN111961609B (en) * | 2020-07-21 | 2023-04-28 | 广东石油化工学院 | Heterotrophic nitrification-aerobic denitrification microbial inoculum and application thereof |
| CN112251387B (en) * | 2020-11-05 | 2022-05-10 | 广东省微生物研究所（广东省微生物分析检测中心） | Denitrifying bacteria and application thereof |
| CN114703093B (en) * | 2022-03-18 | 2023-08-22 | 曲阜师范大学 | Facultative anaerobic complete denitrification gas producing bacterium Y23 and application thereof |
| CN114933988A (en) * | 2022-05-11 | 2022-08-23 | 江苏科技大学 | Pseudomonas with aerobic denitrification synchronous nitrogen and phosphorus removal performance |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101705202A (en) * | 2009-12-03 | 2010-05-12 | 华中农业大学 | Pseudomonas stutzeri YZN-001 for removing ammoniacal nitrogen, nitrate nitrogen and nitrous nitrogen in water body and application |
| CN106754570A (en) * | 2017-02-28 | 2017-05-31 | 重庆大学 | One plant of Pseudomonas stutzeri and its application |
| CN107201325A (en) * | 2017-05-03 | 2017-09-26 | 上田环境修复股份有限公司 | Pseudomonad strain and its cultural method and application |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104830710B (en) * | 2015-02-16 | 2017-08-25 | 南京大学 | One plant of resistant to elevated temperatures aerobic denitrifying bacteria and its application |
-
2019
- 2019-01-25 CN CN201910071406.1A patent/CN109943497B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101705202A (en) * | 2009-12-03 | 2010-05-12 | 华中农业大学 | Pseudomonas stutzeri YZN-001 for removing ammoniacal nitrogen, nitrate nitrogen and nitrous nitrogen in water body and application |
| CN106754570A (en) * | 2017-02-28 | 2017-05-31 | 重庆大学 | One plant of Pseudomonas stutzeri and its application |
| CN107201325A (en) * | 2017-05-03 | 2017-09-26 | 上田环境修复股份有限公司 | Pseudomonad strain and its cultural method and application |
Non-Patent Citations (4)
| Title |
|---|
| "Removal of nitrogen by heterotrophic nitrification-aerobic denitrification of a phosphate accumulating bacterium Pseudomonas stutzeri YG-24";Chune Li等;《Bioresources Technology》;20150131;第18-25页 * |
| "一株反硝化细菌的鉴定、功能基因检测及其反硝化特性";胡金星等;《环境科学与技术》;20121215;第35卷(第12期);第27-31页 * |
| "一株高效异养硝化-好氧反硝化菌的分离鉴定及脱氮性能";周迎芹等;《环境工程学报》;20131005;第7卷(第10期);第4127-4131页 * |
| "异养硝化-好氧反硝化菌株DN1.2的脱氮特性研究";黄钧等;《高技术通讯》;20090417;第19卷(第2期);第213-218页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN109943497A (en) | 2019-06-28 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN109943497B (en) | Pseudomonas stutzeri with aerobic denitrification function and application thereof | |
| CN102443558B (en) | Composite heterotrophic nitrifying bacterial agent and application of same in nitrogen removal treatment of waste water containing ammonia and nitrogen | |
| CN109110912A (en) | A kind of dirt collecting pit pool black and odorous water administering method | |
| CN109455828B (en) | Application method of immobilized microorganism in livestock and poultry breeding wastewater treatment | |
| CN111607543B (en) | Pseudomonas stutzeri with aerobic denitrification function and application thereof | |
| CN114703095B (en) | Pseudomonas adulthood and application thereof in field of sewage and wastewater purification | |
| CN104845920A (en) | Marine zobellella sp. and application thereof | |
| CN115353986B (en) | Bacillus bailii strain WB strain for treating pig raising wastewater and application thereof | |
| CN109052654A (en) | A method of for improving the smelly water of urban black | |
| CN101607783B (en) | Treating method of river bottom mud | |
| CN110699285B (en) | Palyalisma and application thereof in treating landfill leachate membrane concentrated solution | |
| Lu et al. | A review of ammonia-oxidizing archaea and anaerobic ammonia-oxidizing bacteria in the aquaculture pond environment in China | |
| CN101386823A (en) | Special effect anaerobic denitrifying bacterium and waste water processing method using thereof | |
| CN109081447B (en) | Method for removing nitrogen and phosphorus in culture wastewater by combining chlorella, acinetobacter and pseudomonas | |
| CN104651281A (en) | Preparation method of photosynthetic bacterium capable of improving yield and/or quality of aquatic animal | |
| Catalan-Sakairi et al. | High-rate seawater denitrification utilizing a macro-porous cellulose carrier | |
| CN115353211B (en) | Application of bacillus megatherium LZP03 in treatment of pig raising wastewater | |
| CN115353210B (en) | Application of bacillus pumilus LZP02 in treatment of pig raising wastewater | |
| CN107176678A (en) | A kind of method by functional microorganism intensive treatment livestock breeding wastewater | |
| CN115353987B (en) | Bacillus subtilis strain SC strain for treating pig raising wastewater and application thereof | |
| Wang et al. | Review of the mechanisms involved in dissimilatory nitrate reduction to ammonium and the efficacies of these mechanisms in the environment | |
| Jiang et al. | Marine aquaculture regulates dissimilatory nitrate reduction processes in a typical semi-enclosed bay of southeastern China | |
| CN110305815A (en) | A kind of aerobic denitrifying bacteria and its application | |
| Chen et al. | Nitrogen cycling process and application in different prawn culture modes | |
| CN115820466B (en) | Sulfur autotrophic denitrification strain, bacterial preparation and application thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |