CN105176901A - Microbial preparation for treating high N, P and S polluted sewage and application thereof - Google Patents
Microbial preparation for treating high N, P and S polluted sewage and application thereof Download PDFInfo
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- CN105176901A CN105176901A CN201510605698.4A CN201510605698A CN105176901A CN 105176901 A CN105176901 A CN 105176901A CN 201510605698 A CN201510605698 A CN 201510605698A CN 105176901 A CN105176901 A CN 105176901A
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Abstract
The invention provides a microbial preparation for treating high N, P and S polluted sewage and an application thereof. The steps of pretreatment and direct microbial agent treatment are conducted. Three microorganisms are applied to be prepared into a microbial agent, sulphur, nitrogen and phosphorus in the sewage are effectively removed, and better application prospect is achieved.
Description
Technical field
The present invention relates to a kind of sewage water treatment method, belong to integrated wastewater processing technology field.
Background technology
Along with the raising of industrial development and people's living standard, industrial sewage amount and city domestic sewage amount are surged just with surprising rapidity, these sewage or or polluted the river that the mankind depend on for existence, river and lake, have formed one of reason of prestige association environment for human survival.And existing various sewage water treatment method, its processing power is extremely limited, no matter is sewage load, or wastewater treatment efficiency, can not meet the actual requirement of process industry in time and sanitary sewage all far away.Because in existing sewage water treatment method, as biochemical process, aeration oxygen replenishing method, filtration method etc., not only investment is large, and floor space is large, and working cost is high, and processing power and treatment effect undesirable; For another example simple Coagulation Method, then also exist poor processing effect, and running cost is high, and easily cause secondary pollution problems.In addition, existing sewage water treatment method, mostly also exists and needs supporting pipe network system and the how problem such as reuse water.Therefore, a kind of effective sewage water treatment method is sought in the task of top priority, to solve the sewage discharge problem be on the rise.
At present, the method adopting iron autotrophic denitrification technique to carry out synchronous denitrification dephosphorizing to sewage is open, as Chinese Patent Application No. 201110234430.6, Shen Qing Publication day is on November 28th, 2011, and invention and created name is: the method realizing sewage synchronous denitrification dephosphorizing.This invention adopts a kind of fe that can utilize as the autotrophic denitrification bacterium of electron donor, in its denitrification process, nitrate ion is reduced to nitrogen, fe is oxidized to iron ion, simultaneously, iron ion and phosphate anion are combined into precipitation, realize the method for the double effects of denitrogenation dephosphorizing.But this invention is that electron donor can produce a large amount of hydroxide radicals with fe, is unfavorable for that systems stay carries out, also can affects phosphor-removing effect, cause there is no good Tuo Dan Chu Pity effect.
Summary of the invention
The object of the invention is to the deficiency overcoming prior art, N, P, S treatment process in a kind for the treatment of effect sewage that good biochemistry combines with physics is provided.
Another object of the present invention is to provide a kind of complex microorganism preparations for freshwater aquiculture water improvement, it is characterized in that, according to weight percent, microorganism is: subtilis 25-30wt%; Bacillus amyloliquefaciens 25-30wt%; Thiobacillus denitrificans 15-20wt%.
The complex microorganism preparations of the aquaculture water improvement that the present invention is above-mentioned, wherein, microbial preparation can also comprise carrier, and described carrier can be liquid vehicle, as water; Also can be solid carrier, as the mixture of any one or a few in zeolite, calcium carbonate, kaolin, wilkinite.
The complex microorganism preparations for aquaculture water improvement that the present invention is above-mentioned, wherein, can also comprise nutritive substance, described nutritive substance can be any one or a few the mixture be selected from starch, molasses, trehalose, sucrose, maltose, isomaltose, wood sugar, fructose, stachyose, raffinose.
Third aspect of the present invention is to provide a kind of method preparing the above-mentioned complex microorganism preparations for aquaculture water improvement, said method comprising the steps of:
Step 1, provides subtilis, bacillus amyloliquefaciens and thiobacillus denitrificans seed culture fluid;
Step 2, subtilis, bacillus amyloliquefaciens and thiobacillus denitrificans seed culture fluid are inoculated in substratum successively, fermentation, then concentrate and obtain high-concentration bacterial liquid;
Step 3, gained high-concentration bacterial liquid mixes with carrier, nutritive substance, obtains described microbial preparation.
Wherein thiobacillus denitrificans can also build for adopting following mode the thiobacillus denitrificans obtained:
Described thiobacillus denitrificans is the thiobacillus denitrificans of the encoding gene having proceeded to coding any one protein variant following, and described protein variant is suddenly change as follows respectively relative to albumen original amino acid 70P/S, 92E/I, 129Q/R, 155A/P, 242R/M, 368A/S, 433L/R, 489A/Q, 543F/L, 609V/F, 661V/S, albumen original amino acid represent see GenBank:WP_011911646.1(70P/S replace with S amino acid at the P amino acid of original series the 70th).Aminoacid sequence after these sudden changes makes the bacterial strain of the aminoacid sequence after having imported this sudden change have to have better effect relative to the bacterial strain do not suddenlyd change.And applicant is confirmed by a large amount of experiments, not all replacement all has similar effect, has the bacillus pumilus after a large amount of replacements not have the effect of good sewage disposal together with other bacterial strain.
Utilize DNAMAN software analysis, nucleotide sequence PS gene according to the synthesis of aminoacid sequence GenBank:WP_011911646.1 full genome, object fragment is obtained by carrying out amplification, pcr amplification obtains goal gene PS(and is incorporated in gene order by multiplex PCR by corresponding mutational site simultaneously, thus obtain different mutants gene), double digestion PCR primer is carried out with restriction enzyme, be connected with the same expression vector pSDK-1 through double digestion, the recombinant plasmid be proved to be successful is transformed and enters into described thiobacillus denitrificans by the mode of the plasmid transfer commonly used by this area from intestinal bacteria, obtain degraded high content nitrogen, phosphorus, the genetic engineering bacterium of sulphur sewage.
The enlarged culturing of each bacterial classification of the present invention is the cellar culture mode of this area, and the training method also can recorded in reference literature obtains.
The main beneficial effect that the present invention obtains is as follows: the present invention adopts microbiobacterial agent to dispose of sewage, and has effectively purified sewage, and the nitrogen phosphate and sulfur eliminating the high-content in sewage pollutes, and has good application prospect.
Embodiment
Below in conjunction with specific examples, the present invention is further elaborated, but embodiment does not limit the scope of the invention.
Embodiment 1 one kinds removes the method for the nitrogen phosphate and sulfur of high-content in sewage
Adopt the method for spawn culture disclosed in embodiment 1 in CN103352018A, cultivate the preparation obtaining corresponding three kinds of bacterial classifications.
Sewage is provided, in described sewage, N starting point concentration is 250mg/L, P starting point concentration is 150mg/L, S starting point concentration is 100mg/L, and described sewage is through solid-liquid separator, carry out solid-liquid separation, remove bulk solid particulate matter, liquid enters settling tank subsequently, precipitates 4 hours, again liquid is removed solid floccules by circular hole filtering net, the Circularhole diameter of circular hole filtering net is 0.1mm.Then the sewage after process is added microbial preparation process 3 days according to the amount of 10g/L.The sewage that check processing is crossed, finds that N concentration be 50mg/L, P starting point concentration be 65mg/L, S starting point concentration is 60mg/L.There is certain treatment effect.
The acquisition of embodiment 2 transgenosis thiobacillus denitrificans
Utilize DNAMAN software analysis, nucleotide sequence PS gene according to the synthesis of aminoacid sequence GenBank:WP_011911646.1 full genome, obtain object fragment by carrying out amplification, pcr amplification obtain goal gene PS(simultaneously by multiplex PCR by corresponding mutational site 70P/S, 92E/I, 129Q/R, 155A/P, 242R/M, 368A/S, 433L/R, 489A/Q, 543F/L, 609V/F, 661V/S is incorporated in gene order, thus obtain different mutants gene), double digestion PCR primer is carried out with restriction enzyme, be connected with the same expression vector pSDK-1 through double digestion, first pSDK-1 is imported in intestinal bacteria, the mode of the plasmid commonly used by this area transfer (see " clone of intestinal bacteria phosphofructokinase gene in thiobacillus thiooxidans, express and analysis to organic carbon metabolism " described in method, do not state one by one at this) recombinant plasmid be proved to be successful transformed from intestinal bacteria enter into described thiobacillus denitrificans, obtain described genetic engineering bacterium.
Embodiment 3: the wastewater treatment efficiency of thiobacillus denitrificans genetic engineering bacterium and all the other several microbial inoculums is verified
According to the method for embodiment 1, carry out corresponding sewage disposal experiment, the sewage of its sewage and embodiment 1 belongs to same batch, has N, P, S concentration of same concentrations.Wherein each component composition of microbial inoculum and the identical of embodiment 1.
Adopt the genetic engineering bacterium in different mutational sites to carry out removing the reaction of N, P, S respectively, found through experiments, only have 70P/S, 92E/I, 129Q/R, 155A/P, 242R/M, 368A/S, 433L/R, 489A/Q, 543F/L, this several mutant bacteria of 609V/F, 661V/S has and has relative to original bacteria the effect obviously strengthened.Its result is as follows:
Sewage is with a collection of sewage, thus guaranteed conditions is consistent.
As can be seen from the above results, the microbial inoculum that the thiobacillus denitrificans adopting genetically engineered to build and other several bacterial strains are formed is except having the original good effect removing N, P, S, also there is the effect reducing COD and lipid acid preferably, do not state one by one at this.
Claims (5)
1. a microbiobacterial agent, it is made up of three kinds of microorganisms.
2. a microbiobacterial agent, it is made up of following each component: subtilis, bacillus amyloliquefaciens and thiobacillus denitrificans.
3. microbial inoculum as claimed in claim 2, is characterized in that: described thiobacillus denitrificans is the strain of i (bacillus) pumilus of the encoding gene having proceeded to coding any one protein variant following, and described protein variant is the 70P/S that suddenlys change as follows respectively relative to albumen original amino acid, 92E/I, 129Q/R, 155A/P, 242R/M, 368A/S, 433L/R, 489A/Q, 543F/L, 609V/F, 661V/S, albumen original amino acid is see GenBank:WP_011911646.1.
4. the microbial inoculum as described in any one of claim 1-3 is processing the application of being rich in the sewage of nitrogen phosphate and sulfur.
5. a protein variant, is characterized in that: protein variant is suddenly change as follows respectively relative to albumen original amino acid 70P/S, 92E/I, 129Q/R, 155A/P, 242R/M, 368A/S, 433L/R, 489A/Q, 543F/L, 609V/F, 661V/S, albumen original amino acid is see GenBank:WP_011911646.1.
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| CN201510605698.4A CN105176901A (en) | 2015-09-22 | 2015-09-22 | Microbial preparation for treating high N, P and S polluted sewage and application thereof |
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| CN201510605698.4A CN105176901A (en) | 2015-09-22 | 2015-09-22 | Microbial preparation for treating high N, P and S polluted sewage and application thereof |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN117343881A (en) * | 2023-12-05 | 2024-01-05 | 山东省海洋资源与环境研究院(山东省海洋环境监测中心、山东省水产品质量检验中心) | Bacillus subtilis DS5 and application thereof |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103352018A (en) * | 2013-07-19 | 2013-10-16 | 上海创博生态工程有限公司 | A kind of complex microorganism preparations for aquaculture water improvement and preparation method thereof |
-
2015
- 2015-09-22 CN CN201510605698.4A patent/CN105176901A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103352018A (en) * | 2013-07-19 | 2013-10-16 | 上海创博生态工程有限公司 | A kind of complex microorganism preparations for aquaculture water improvement and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| NCBI: "WP_011911646.1", 《GENBANK》 * |
| 周晓黎等: "一株施氏假单胞菌Pseudomonas stutzeri DN-LWX19 的脱氮性能", 《环境工程学报》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN117343881A (en) * | 2023-12-05 | 2024-01-05 | 山东省海洋资源与环境研究院(山东省海洋环境监测中心、山东省水产品质量检验中心) | Bacillus subtilis DS5 and application thereof |
| CN117343881B (en) * | 2023-12-05 | 2024-03-19 | 山东省海洋资源与环境研究院(山东省海洋环境监测中心、山东省水产品质量检验中心) | Bacillus subtilis DS5 and application thereof |
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