CN114606131A

CN114606131A - Green alga strain and application thereof in rare earth ammonia nitrogen wastewater treatment

Info

Publication number: CN114606131A
Application number: CN202210333021.XA
Authority: CN
Inventors: 王明兹; 周有彩; 肖雪花; 何勇锦; 陈必链
Original assignee: Fujian Normal University
Current assignee: Fuzhou Wenze Biotechnology Co ltd
Priority date: 2022-03-30
Filing date: 2022-03-30
Publication date: 2022-06-10
Anticipated expiration: 2042-03-30
Also published as: CN114606131B

Abstract

The invention discloses a green alga strain and application thereof in rare earth ammonia nitrogen wastewater treatment, wherein the green alga strain is classified and named as Chlamydomonas YC with the scientific name ofChlamydomonasYc, deposited in the chinese culture collection center in wuhan at 2021, 5 months and 13 days, with the deposition number of CCTCC NO: m2021529. The green alga strain can be used for treating high-concentration ammonia nitrogen wastewater.

Description

Green alga strain and application thereof in rare earth ammonia nitrogen wastewater treatment

Technical Field

The invention belongs to the technical field of microorganisms, and particularly relates to a green alga strain and application thereof in treatment of rare earth ammonia nitrogen wastewater.

Background

The current rare earth mining process mainly adopts an in-situ mineral leaching process, wherein a liquid injection well is distributed on the surface of a mountain body, an ammonium sulfate solution with a certain concentration is injected into the ore body as a leaching agent to replace rare earth in the rare earth ore, leachate is collected, and carbonate rare earth products are enriched through ammonium bicarbonate precipitation. In-situ ore leaching does not need to strip surface soil and excavate a mountain, has small damage to vegetation of the mountain and does not generate a large amount of tailings. Because a large amount of ammonium sulfate is put into the leaching process, the soil and tail water in the mountain body form a huge amount of high-concentration ammonia nitrogen pollution after the ore is closed. The rare earth industry reportedly produces more than 20 million tons per year, with ammonia nitrogen concentrations mostly in the range of 300-5000 mg/L, and cannot be properly handled. The water flows are merged into the stream through surface leakage, rain leaching and the like, so that serious ecological and social problems around the mining area are caused. Therefore, such sewage must be treated and utilized. The pollutant discharge standard in the rare earth industry requires that the direct discharge concentration is less than or equal to 15 mg/L (GB 26451-2011), and if the pollutant is not treated properly, serious environmental pollution is caused.

So far, although the ammonia nitrogen wastewater treatment process has more methods and mature processes, the process is mainly suitable for domestic sewage and general industrial wastewater. Because of the specificity of the rare earth ammonia nitrogen sewage, such as high ammonia nitrogen concentration, low COD (chemical oxygen demand), extreme unbalance of nutrition and combination of objective comparison of input cost, an economic and easily-controlled mature process suitable for treating the rare earth mine high ammonia nitrogen sewage does not exist so far. The method for treating ammonia nitrogen wastewater by using microorganisms has good application prospect, and the biological denitrification method mainly utilizes specific microorganisms to degrade ammonia nitrogen in the wastewater so as to achieve the aim of purifying water. The method has the advantages of lower operation cost, small secondary pollution, resource utilization of valuable biomass and the like. The span range of the ammonia nitrogen concentration value of the wastewater of the south ionic rare earth mine is large, and the inorganic or organic carbon source in the wastewater is very little, so that the requirements of microorganisms on the carbon source and the energy source are difficult to meet, and the biological denitrification process is difficult to establish. For the problem, a large amount of organic carbon sources are usually supplemented, for example, a sugar supplementing process is adopted, and biological denitrification is realized through intensive input of energy and substances, but the process has high cost and does not have popularization and application values. In addition, the ammonia nitrogen content of the rare earth sewage is high, and only a few microbial flora can tolerate high-concentration ammonia nitrogen, so that the whole denitrification process is blocked, and the denitrification effect is also influenced.

Disclosure of Invention

In order to solve the problems, the invention aims to separate and screen out greenhouse gas CO which has strong tolerance and can be cheap and conveniently obtained from high-concentration rare earth ammonia nitrogen sewage₂The microalgae which is used as a carbon source and takes clean energy sunlight as main energy is expected to establish a stable and efficient rare earth sewage denitrification process through the microorganisms, so that the running power consumption and energy consumption are reduced, and the problem of ammonia nitrogen pollution which puzzles the surrounding environment of the rare earth industry is solved.

To achieve the above objects, the present invention provides a green algae strain, which is classified and named as Chlamydomonas YC, with the scientific nameChlamydomonassp, YC, deposited in the chinese type culture collection located in wuhan at 2021, 5 months and 13 days, with the deposit number CCTCC NO: m2021529.

The green alga strain can be used for treating high-concentration ammonia nitrogen wastewater.

Drawings

FIG. 1 shows a strainChlamydomonasColony plate of sp, YC.

Detailed Description

Example 1 screening, isolation and identification of strains

1) Collecting high-concentration ammonia Nitrogen (NH) of rare earth₄ ⁺Not less than 2000 mg/L), adding a proper amount of phosphorus element (total phosphorus: 5-50 mg/L), culturing the culture in 2500-7000 lux illumination intensity in an open manner, and shaking by hand 6 times every day to obtain a culture which can survive in sewage with ammonia nitrogen concentration of more than 2000 mg/L.

2) The microalgae without ammonia nitrogen tolerance ability can not grow under the inhibition of the high-concentration ammonia nitrogen wastewater, and the microalgae with tolerance ability can simultaneously grow by synthesizing a cell matrix of the microalgae by using the ammonia nitrogen of the high-concentration ammonia nitrogen wastewater. And (4) enriching and screening microalgae with high ammonia nitrogen tolerance.

3) Screening and separation of microalgae

And dripping the obtained culture onto a sterilized glass slide by using a micropipette, dripping 2-4 drops of sterile water onto the glass slide, and linearly arranging the culture and the sterile water from left to right at certain intervals. Under inverted microscope observation, the smallest possible number of cells was pipetted with a sterilized micropipette onto a drop of sterile water on the right. Carefully checking the dropping water, if only one or a plurality of cells with the same morphology need to be separated in the dropping water and no other organisms are mixed, sucking the dropping water by using a pipette and pumping the dropping water together with a pipette tip into a sterilized rare earth sewage test tube added with phosphorus elements. If the drop is observed under a microscope to contain cells of different morphology, another micropipette is taken to aspirate as few cells as possible into another drop. Repeating the steps until a certain drop of sterile water only contains one or more cells of the same morphology to be separated, sucking the water drop by using the pipette, pumping the water drop and the pipette head into a sterilized rare earth sewage test tube added with phosphorus elements, and sealing the test tube by using 8 layers of gauze. Cultivate under 2500~7000 lux illumination intensity, hand the test tube 6 times with appropriate dynamics daily, liquid becomes green in the test tube.

4) And (5) purifying the microalgae. Penicillin and gentamicin are added into the green culture in the test tube to enable the final concentration of the two antibiotics in the test tube to be 20-100 mu g/mL and 15-50 mu g/mL, and the culture is continued for one day. And (3) performing concentration gradient dilution on the green culture of the test tube to a proper multiple by using sterile water, taking 100 mu L of the diluted green culture, coating the diluted green culture on a solid plate containing solid phosphorus-containing rare earth sewage, and culturing under the illumination intensity of about 2500-7000 lux until a green colony grows out. And selecting a single colony to streak and culture on a solid plate containing phosphorus and rare earth sewage. Repeating the steps for a plurality of times to obtain pure microalgae.

5) And (5) identifying microalgae. Measuring the 18SrDNA and ITS sequences of the microalgae, comparing and identifying the 18SrDNA and the ITS sequences, and obtaining the separated microalgae which is a green alga and is named asChlamydomonassp, YC. The strain is preserved and registered in a book by China center for type culture Collection (Wuhan, China) at 2021, 5 months and 13 days, and the preservation numbers are as follows: CCTCC M2021529 YC, and the center has been tested for viability at 20/5/2021, and concluded to be viable.

Example 2 Ammonia-Nitrogen tolerance test of the Chlamydomonas sp. YC Strain

Separating and purifying the obtained microalgaeChlamydomonasThe sp and YC strains are inoculated into the phosphorus-containing rare earth ammonia nitrogen sewage liquid, and are subjected to illumination shaking culture at the constant temperature of 150 r/min and 2500lux for 5 days at the temperature of 25 ℃ to prepare microalgae YC seed liquid. Respectively taking 100 mL of microalgae YC seed liquid, centrifuging, inoculating the precipitate into 5 1L BG11 culture media containing ammonia nitrogen concentrations of 1000 mg/L, 2000 mg/L, 3000 mg/L, 4000 mg/L and 5000 mg/L, and performing shaking culture at 25 deg.C, 150 r/min and 2500lux under constant temperature illumination for 14 days. The biomass of the microalgae YC grows from 0.1 g/L to 0.4 g/L, 0.35 g/L, 0.25g/L, 0.2g/L and 0.07g/L respectively in culture media with different ammonia nitrogen concentrations.

Application example 1

1) Separating and purifying the obtained microalgaeChlamydomonasThe sp and YC strains are inoculated into the phosphorus-containing rare earth ammonia nitrogen sewage liquid, and are subjected to illumination shaking culture at the constant temperature of 150 r/min and 2500lux for 5 days at the temperature of 25 ℃ to prepare microalgae YC seed liquid. Taking 100 mL of microalgae YC seed liquid, centrifuging, inoculating the precipitate into 1L of rare earth-containing sewage (ammonia nitrogen content is 200 mg/L) diluted by 10 times, and performing aeration culture at 25 ℃ and 1000 mL/min. The ammonia nitrogen of 200 mg/L is basically completely consumed within 10 days, and the consumption rate of the ammonia nitrogen reaches 99.5 percent.

Application example 2

2) Separating and purifying the obtained microalgaeChlamydomonasThe sp and YC strains are inoculated into the phosphorus-containing rare earth ammonia nitrogen sewage liquid, and are subjected to illumination shaking culture at the constant temperature of 150 r/min and 2500lux for 5 days at the temperature of 25 ℃ to prepare microalgae YC seed liquid. Taking 100 mL of microalgae YC seed liquid, centrifuging, inoculating the precipitate into 1L of original rare earth-containing sewage (ammonia nitrogen content of 2000 mg/L), and culturing at 25 deg.C and 700 mL/min with aeration. The ammonia nitrogen content in the rare earth sewage of 2000 mg/L is reduced to 350 mg/L within 50 days, and the consumption rate of the ammonia nitrogen reaches 82.5 percent.

Sequence listing

<110> university of Fujian profession

<120> green alga strain and application thereof in rare earth ammonia nitrogen wastewater treatment

<130> 2022

<160> 1

<170> SIPOSequenceListing 1.0

<210> 1

<211> 555

<212> DNA

<213> Artificial sequences (artificial series)

<400> 1

gctcaggtcg agataatgag gggttaaagc tctcgcacag ggcaagagtt cctggttcgg 60

attgaccagc acgaatccag accaccaact aaagcattag caacgaattg cctacccagt 120

tgaggccctt agcgggtcca tgcatcaatc tctacacttc agcagacccg gctaagctct 180

cagtgaaaac ttagccgaga cggccaggtc cgtttcacac tccccagagg gagcatgaga 240

cgagtattaa cccgacgctg aggcagacat gctcttggcc gaagcctcga gcgcaatatg 300

cgttcaaaga cttgatgatt cacgtatttc tgcaattcac actaagtatc gcatttcgct 360

gcgttcttca tcattcgagg agccaagata tccgttgttg agagttgtct tggttggtga 420

gagtcagctc taagccgagc cctcgatttt tcattaggtt tggtttggtg ttgttggtta 480

gataaagacc gacgccccac atgcccgaag gcatgcacag cgacctgagc ggcctgaatt 540

acgagaaccc agaca 555

Claims

1. A green algae strain, characterized by: it is classified and named as Chlamydomonas YC with the academic nameChlamydomonassp, YC, deposited in the chinese type culture collection located in wuhan at 2021, 5 months and 13 days, with the deposit number CCTCC NO: m2021529.

2. The use of the green algae strain of claim 1 in ammonia nitrogen wastewater treatment.