CN110846246A - Preliminary screening method for bacteria with ammonia nitrogen degradation function - Google Patents
Preliminary screening method for bacteria with ammonia nitrogen degradation function Download PDFInfo
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- CN110846246A CN110846246A CN201911074985.1A CN201911074985A CN110846246A CN 110846246 A CN110846246 A CN 110846246A CN 201911074985 A CN201911074985 A CN 201911074985A CN 110846246 A CN110846246 A CN 110846246A
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Abstract
The invention discloses a preliminary screening method of bacteria with an ammonia nitrogen degradation function, which comprises the steps of preparing an ammonia nitrogen selection culture medium, preparing an ammonia nitrogen standard storage solution, preparing an ammonia nitrogen standard cone working solution, measuring absorbance, inoculating a strain, and detecting the ammonia nitrogen content in the ammonia nitrogen selection culture medium by adopting a nano reagent spectrophotometry, wherein the ammonia nitrogen degradation rate is (the ammonia nitrogen content of a control group-the ammonia nitrogen content of an experimental group)/the ammonia nitrogen content of the control group. The method can screen out bacteria with different ammonia nitrogen degradation functions in the microbial bacteria, which is absent in the existing experimental method, thereby providing favorable guarantee for the subsequent bacteria distinguishing experiment, filling the blank of the existing experimental screening of the bacteria with the ammonia nitrogen degradation functions, and further improving the ammonia nitrogen degradation experimental method.
Description
Technical Field
The invention relates to the technical field of microbial bacteria screening, in particular to a preliminary screening method of bacteria with an ammonia nitrogen degradation function.
Background
In the field of microbial bacteria, bacillus is widely distributed and has strong adaptability, wherein bacillus subtilis and bacillus licheniformis are common probiotics and have multiple biological functions. The closest prior art to the present invention includes:
(1) research on water purification effect of bacillus subtilis HAINUP40 [ J ] aquatic science, 2018,37(02): 159-.
(2) The separation and identification of a bacillus for degrading ammonia nitrogen in a prawn culture water body and the optimization of a fermentation formula [ J ]. fishery research, 2019,41(03): 175-.
(3) Bacillus megaterium has characteristics of degrading ammonia nitrogen in aquaculture water [ J ] Proc. Shenyang university of agriculture, 2006(04): 607-.
(4) The bacillus megaterium JSSW-JD has strong capability of degrading nitrite in the aquaculture water, and the degradation rates of the nitrite respectively reach 98.1%, 94.9% and 67.8% after the water in a fishpond, reservoir water and crab pond are treated for 8 days respectively. The bacillus megaterium JSSW-JD has no obvious function of degrading ammonia nitrogen.
(5) Zhangchao, Shanghan, Liuyu, Yang Qing, Liluo, Shiyinyuan, strain SDB1-2 and its application in landfill leachate treatment [ J ]. Purchase of Beijing academy of agriculture, 2019,34(02):1-4. Klebsiella pneumoniae SDB1-2 separated in landfill leachate screening, research shows that it can degrade artificial wastewater with initial ammonia nitrogen concentration up to 2000mg/L, ammonia nitrogen, COD and highest removal rate are respectively 51.15%, 35.03% and 57.45% [5 ].
In the prior art, although the ammonia nitrogen degradation function of the microbial bacteria can be realized, no matter which operation method is adopted, different bacteria in the bacterial strains cannot be screened, so that the experimental result can only be embodied in the flora, and the specific bacterial strains cannot be verified, so that the experimental result has certain accuracy.
Disclosure of Invention
The invention aims to provide a primary screening method for bacteria with an ammonia nitrogen degradation function, which can distinguish different strains through ammonia nitrogen degradation rates in an experiment, and provide reliable guarantee for the ammonia nitrogen degradation experiment result of subsequent bacteria so as to solve the problems in the background art.
In order to achieve the purpose, the invention provides the following technical scheme:
a primary screening method of bacteria with ammonia nitrogen degradation function comprises the following steps:
s1: preparing an ammonia nitrogen selective culture medium, comprising: (NH)4)2SO4：1.18g/L；FeSO4·7H2O：0.04g/L；K2HPO4：1g/L；KH2PO4：0.25g/L；MgSO4·7H20: 0.5 g/L; glucose: 10 g/L; trace elements: 1 mL;
s2: preparing an ammonia nitrogen standard storage solution, wherein rho N is 1000 mu g/mL; weighing 3.819g of ammonium chloride, drying in an oven, dissolving in water, transferring into a 1000mL volumetric flask, and diluting to a marked line;
s3: preparing an ammonia nitrogen standard vertebral working solution, wherein rho N is 10 mug/mL, sucking 5.00mL of ammonia nitrogen standard stock solution into a 500mL volumetric flask, and diluting to a scale;
s4: respectively adding 0.00 mL, 0.10 mL, 0.20 mL, 0.40 mL, 0.80 mL, 1.20 mL, 1.60 mL and 2.00mL of ammonia nitrogen standard working solution into 8 10mL colorimetric tubes, adding water to a marked line, adding 200mL of potassium sodium tartrate solution, shaking up, adding 300mL of Nashi reagent, shaking up, standing for 10min, and measuring absorbance by using a 10mm cuvette and water as a reference at a wavelength of 420 nm;
s5: inoculating the strain: sucking bacteria liquid from 1mL to 1.5mL in a sterile centrifugal tube, centrifuging for 5min after 5000 revolutions, discarding the supernatant, resuspending with sterile water, repeating for three times, washing off the culture medium with thallus residue, inoculating into an ammonia nitrogen selective culture medium according to 10 percent of inoculation amount, simultaneously establishing a group of control groups for inoculating sterile water, and culturing for 3 days and 6 days at 34.5 ℃ after 180 revolutions;
s6: and detecting the ammonia nitrogen content in the ammonia nitrogen selection culture medium by adopting a nano reagent spectrophotometry, wherein the ammonia nitrogen degradation rate is (the content of the ammonia nitrogen in a control group-the content of the ammonia nitrogen in an experimental group)/the content of the ammonia nitrogen in the control group.
Further, a solution of trace elements in selective medium of ammonia nitrogen in S1 is prepared, including: 3.9g of zinc sulfate heptahydrate, 7g of calcium chloride dihydrate, 5.1g of manganese chloride tetrahydrate, 1.1g of ammonium molybdate, 1.6g of copper sulfate pentahydrate, 1.6g of cobalt chloride hexahydrate and 1000mL of distilled water.
Compared with the prior art, the invention has the beneficial effects that:
according to the preliminary screening method for bacteria with the ammonia nitrogen degradation function, provided by the invention, bacteria with different ammonia nitrogen degradation functions in microbial bacteria can be screened out, which is absent in the existing experimental method, so that a favorable guarantee is provided for a subsequent bacteria distinguishing experiment, the blank of the existing experimental screening for bacteria with the ammonia nitrogen degradation function is filled, and the ammonia nitrogen degradation experimental method is further improved.
Drawings
FIG. 1 is a standard curve diagram of the ammonia nitrogen content of the invention.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
In the embodiment of the invention: the method for primarily screening bacteria with ammonia nitrogen degradation function comprises the following steps:
the first step is as follows: preparing an ammonia nitrogen selective culture medium, comprising: (NH)4)2SO4：1.18g/L；FeSO4·7H2O：0.04g/L；K2HPO4：1g/L；KH2PO4：0.25g/L；MgSO4·7H20: 0.5 g/L; glucose: 10 g/L; trace elements: 1 mL; as shown in table 1 below:
table 1: ammonia nitrogen selective medium component
Wherein the trace element solution comprises: 3.9g of zinc sulfate heptahydrate, 7g of calcium chloride dihydrate, 5.1g of manganese chloride tetrahydrate, 1.1g of ammonium molybdate, 1.6g of copper sulfate pentahydrate, 1.6g of cobalt chloride hexahydrate and 1000mL of distilled water.
The second step is that: preparing an ammonia nitrogen standard storage solution, wherein rho N is 1000 mu g/mL; weighing 3.819g of ammonium chloride, drying in an oven, dissolving in water, transferring into a 1000mL volumetric flask, and diluting to a marked line;
the third step: preparing an ammonia nitrogen standard vertebral working solution, wherein rho N is 10 mug/mL, sucking 5.00mL of ammonia nitrogen standard stock solution into a 500mL volumetric flask, and diluting to a scale;
the fourth step: respectively adding 0.00 mL, 0.10 mL, 0.20 mL, 0.40 mL, 0.80 mL, 1.20 mL, 1.60 mL and 2.00mL of ammonia nitrogen standard working solution into 8 10mL colorimetric tubes, adding water to a marked line, adding 200mL of potassium sodium tartrate solution, shaking up, adding 300mL of Nashi reagent, shaking up, standing for 10min, and measuring absorbance by using a 10mm cuvette and water as a reference at a wavelength of 420 nm;
the fifth step: inoculating the strain: sucking bacteria liquid from 1mL to 1.5mL in a sterile centrifugal tube, centrifuging for 5min after 5000 revolutions, discarding the supernatant, resuspending with sterile water, repeating for three times, washing off the culture medium with thallus residue, inoculating into an ammonia nitrogen selective culture medium according to 10 percent of inoculation amount, simultaneously establishing a group of control groups for inoculating sterile water, and culturing for 3 days and 6 days at 34.5 ℃ after 180 revolutions;
and a sixth step: and detecting the ammonia nitrogen content in the ammonia nitrogen selection culture medium by adopting a nano reagent spectrophotometry, wherein the ammonia nitrogen degradation rate is (the content of the ammonia nitrogen in a control group-the content of the ammonia nitrogen in an experimental group)/the content of the ammonia nitrogen in the control group.
The experimental results are as follows:
referring to fig. 1, the results of the ammonia nitrogen content standard curve;
referring to the following table 2, the screening results of the ammonia nitrogen degrading bacteria are as follows:
table 2: ammonia nitrogen degradation rate index results
And (3) analyzing an experimental result:
the ammonia nitrogen degradation rate of 9 strains of bacillus subtilis (including Bacillus belief and Bacillus amyloliquefaciens) is 35.66-53.33%, the ammonia nitrogen degradation rate of 12 strains of Klebsiella is 36.66-54.65%, the ammonia nitrogen degradation rate of 13 strains of Bacillus megaterium is 2.86-14.77%, the ammonia nitrogen degradation rate of 4 strains of Bacillus cereus is 0%, the ammonia nitrogen degradation rate of 6 strains of Bacillus licheniformis is 2.48-42.65%, the ammonia nitrogen degradation rate of 18 strains of Acinetobacter is 0-60.06%, the ammonia nitrogen degradation rate of 4 strains of Enterobacter cloacae is 4.0-53.39%, and the ammonia nitrogen degradation rate of 11 strains of Proteus mirabilis is 0-55.26%.
In summary, the following steps: according to the preliminary screening method for bacteria with the ammonia nitrogen degradation function, provided by the invention, bacteria with different ammonia nitrogen degradation functions in microbial bacteria can be screened out, so that a favorable guarantee is provided for a subsequent bacteria distinguishing experiment, the blank of screening the existing bacteria with the ammonia nitrogen degradation function is filled, and the ammonia nitrogen degradation experiment method is further improved.
The above description is only for the preferred embodiment of the present invention, but the scope of the present invention is not limited thereto, and any person skilled in the art should be able to cover the technical solutions and the inventive concepts of the present invention within the technical scope of the present invention.
Claims (2)
1. A primary screening method of bacteria with ammonia nitrogen degradation function is characterized by comprising the following steps:
s1: preparing an ammonia nitrogen selective culture medium, comprising: (NH)4)2SO4：1.18g/L；FeSO4·7H2O：0.04g/L；K2HPO4：1g/L；KH2PO4：0.25g/L；MgSO4·7H20: 0.5 g/L; glucose: 10 g/L; trace elements: 1 mL;
s2: preparing an ammonia nitrogen standard storage solution, wherein rho N is 1000 mu g/mL; weighing 3.819g of ammonium chloride, drying in an oven, dissolving in water, transferring into a 1000mL volumetric flask, and diluting to a marked line;
s3: preparing an ammonia nitrogen standard vertebral working solution, wherein rho N is 10 mug/mL, sucking 5.00mL of ammonia nitrogen standard stock solution into a 500mL volumetric flask, and diluting to a scale;
s4: respectively adding 0.00 mL, 0.10 mL, 0.20 mL, 0.40 mL, 0.80 mL, 1.20 mL, 1.60 mL and 2.00mL of ammonia nitrogen standard working solution into 8 10mL colorimetric tubes, adding water to a marked line, adding 200mL of potassium sodium tartrate solution, shaking up, adding 300mL of Nashi reagent, shaking up, standing for 10min, and measuring absorbance by using a 10mm cuvette and water as a reference at a wavelength of 420 nm;
s5: inoculating the strain: sucking bacteria liquid from 1mL to 1.5mL in a sterile centrifugal tube, centrifuging for 5min after 5000 revolutions, discarding the supernatant, resuspending with sterile water, repeating for three times, washing off the culture medium with thallus residue, inoculating into an ammonia nitrogen selective culture medium according to 10 percent of inoculation amount, simultaneously establishing a group of control groups for inoculating sterile water, and culturing for 3 days and 6 days at 34.5 ℃ after 180 revolutions;
s6: and detecting the ammonia nitrogen content in the ammonia nitrogen selection culture medium by adopting a nano reagent spectrophotometry, wherein the ammonia nitrogen degradation rate is (the content of the ammonia nitrogen in a control group-the content of the ammonia nitrogen in an experimental group)/the content of the ammonia nitrogen in the control group.
2. The primary screening method of bacteria with ammonia nitrogen degradation function according to claim 1, wherein the preparation of the trace element solution in the ammonia nitrogen selective medium in S1 comprises the following steps: 3.9g of zinc sulfate heptahydrate, 7g of calcium chloride dihydrate, 5.1g of manganese chloride tetrahydrate, 1.1g of ammonium molybdate, 1.6g of copper sulfate pentahydrate, 1.6g of cobalt chloride hexahydrate and 1000mL of distilled water.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113502237A (en) * | 2021-05-08 | 2021-10-15 | 四川轻化工大学 | Enterobacter reuteri for degrading ammonia nitrogen in white spirit wastewater and application thereof |
| CN114736837A (en) * | 2022-05-25 | 2022-07-12 | 中联环股份有限公司 | Development and application of composite microbial inoculum for degrading kitchen waste |
| CN116439165A (en) * | 2023-05-04 | 2023-07-18 | 海南珊海海洋科技有限公司 | Breeding method for improving survival rate of coral larvae |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102851227A (en) * | 2012-04-17 | 2013-01-02 | 西安建筑科技大学 | Method for screening heterotrophic nitrification bacteria for treating slightly-polluted water |
| CN104388360A (en) * | 2014-12-03 | 2015-03-04 | 青岛根源生物技术集团有限公司 | Screening method of photosynthetic bacteria with capacity of reducing ammonia nitrogen |
| WO2017120495A1 (en) * | 2016-01-07 | 2017-07-13 | Ascus Biosciences, Inc. | Methods for improving milk production by administration of microbial consortia |
| CN107244742A (en) * | 2017-05-10 | 2017-10-13 | 上田环境修复股份有限公司 | A kind of method of the microbial degradation of pollutant in trade effluent |
-
2019
- 2019-11-06 CN CN201911074985.1A patent/CN110846246A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102851227A (en) * | 2012-04-17 | 2013-01-02 | 西安建筑科技大学 | Method for screening heterotrophic nitrification bacteria for treating slightly-polluted water |
| CN104388360A (en) * | 2014-12-03 | 2015-03-04 | 青岛根源生物技术集团有限公司 | Screening method of photosynthetic bacteria with capacity of reducing ammonia nitrogen |
| WO2017120495A1 (en) * | 2016-01-07 | 2017-07-13 | Ascus Biosciences, Inc. | Methods for improving milk production by administration of microbial consortia |
| CN107244742A (en) * | 2017-05-10 | 2017-10-13 | 上田环境修复股份有限公司 | A kind of method of the microbial degradation of pollutant in trade effluent |
Non-Patent Citations (3)
| Title |
|---|
| 张昊 等: ""鸡粪堆肥脱氨氮菌株的分离筛选与鉴定"", 《中国家禽》 * |
| 邹勇 等: "《实用综合化学实验》", 30 September 2017, 西南交通大学出版社 * |
| 黄婧 等: ""畜禽养殖污水中高效氨氮降解菌的筛选、鉴定及生长条件研究"", 《环境工程学报》 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113502237A (en) * | 2021-05-08 | 2021-10-15 | 四川轻化工大学 | Enterobacter reuteri for degrading ammonia nitrogen in white spirit wastewater and application thereof |
| CN113502237B (en) * | 2021-05-08 | 2022-08-30 | 四川轻化工大学 | Enterobacter reuteri for degrading ammonia nitrogen in white spirit wastewater and application thereof |
| CN114736837A (en) * | 2022-05-25 | 2022-07-12 | 中联环股份有限公司 | Development and application of composite microbial inoculum for degrading kitchen waste |
| CN116439165A (en) * | 2023-05-04 | 2023-07-18 | 海南珊海海洋科技有限公司 | Breeding method for improving survival rate of coral larvae |
| CN116439165B (en) * | 2023-05-04 | 2024-03-19 | 海南珊海海洋科技有限公司 | Breeding method for improving survival rate of coral larvae |
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