CN106906173A - A kind of Thiobacillus thioxidans and its application in removing heavy metals are removed - Google Patents
A kind of Thiobacillus thioxidans and its application in removing heavy metals are removed Download PDFInfo
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- CN106906173A CN106906173A CN201710312217.XA CN201710312217A CN106906173A CN 106906173 A CN106906173 A CN 106906173A CN 201710312217 A CN201710312217 A CN 201710312217A CN 106906173 A CN106906173 A CN 106906173A
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Abstract
The present invention relates to a kind of Thiobacillus thioxidans Acidithiobacillus thiooxidans ZJ 1 and its application in removing heavy metals are removed, the bacterial strain has been preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center at present, and deposit number is CGMCC No.12387.The optimum growth temperature of bacterial strain ZJ 1 is 28 DEG C, and the most suitable growth pH is 3.0；Strain of the invention can separate out the heavy metal in all kinds of discarded objects, can be used for removing the heavy metal in all kinds of wastes, and the secondary pollution for preventing heavy metal plays an important roll.
Description
Technical field
The invention belongs to biological treatment field, and in particular to a kind of microorganism that can remove removing heavy metals and application field.
Background technology
In all kinds of wastes such as municipal sludge, usually containing the nutrient needed for substantial amounts of organic matter and plant, tool
There is high agricultural value.But substantial amounts of heavy metal would generally be contained in sludge, and heavy metal is extremely difficult to be biodegradable,
Also easily it is bioaccumulation by food chain simultaneously, eats the vegetables poured by this class sludge, it will seriously threaten people
It is healthy.
Thus, we generally need heavy metal to be processed, can be safer and more effective be utilized.In reality
In processing procedure, using physico-chemical process, generally existing that time-consuming, high cost, it is easy to produce the problem of secondary pollution, because
This, increasing people tends to carry out heavy metal using biological method to be processed, and the method that biology leaches also increasingly is received
To favor.So filtering out a kind of microorganism that can efficiently remove removing heavy metals has high potential value.
The content of the invention
The purpose of invention：Mainly for containing substantial amounts of heavy metal in waste, it is impossible to be used directly, and current
Minimizing technology has that efficiency is low, high cost, and a kind of microorganism that can directly remove removing heavy metals for providing and its
Application process.
Technical scheme：
A kind of to be used to the microorganism of removing heavy metals, its Classification And Nomenclature is Acidithiobacillus thiooxidans
ZJ-1, is preserved in China Microbial Culture Preservation Commission common micro-organisms center CGMCC, and deposit number is CGMCC No:
12387, preservation date is on April 21st, 2016, and preservation address is Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3 Chinese science
Institute of microbiology of institute.
Bacterial strain ZJ-1 of the present invention, the nucleotide sequence of its 16s RNA is as shown in table 1.
The screening technique of Acidithiobacillus thiooxidans ZJ-1 of the present invention：
1. 10g pickling mud samples are weighed, is put into equipped with the training of 100ml (121 DEG C, 30min wet methods sterilized) neat liquid
In the conical flask of foster base (SM culture mediums), then (sulphur powder is used as acidophilia Thiobacillus thioxidans to be added thereto to appropriate sulphur powder
The source of nutriment and acid sulfate)；
2. when turbid liquid pH value is down to 2.0 or so in bottle, supernatant liquor 10mL is drawn after standing, be inoculated into and fill 100mL
In the 250mL conical flasks of the SM culture mediums of wet method sterilizing, and add 10g sulphur powders, be placed in culture in constant-temperature table (30 DEG C,
150rpm)；
3. cultivate 8d-10d, treat that culture medium becomes muddy milky (pH value is about 2).10mL is therefrom drawn to be inoculated into newly
In fresh SM culture mediums；
4. repeated by step 2,3, enrichment culture several times after so that acidophilia Thiobacillus thioxidans turn into it is therein
Dominant microflora；
5. pair culture medium being enriched with is diluted and coats solid SM culture mediums；
6. each concentration is repeated to be coated with three pieces of flat boards, and corresponding record is carried out on culture dish.
7. after coated culture dish being placed into 2h at room temperature, inversion culture in 30 DEG C of incubators is placed on, cultivates 15d
Afterwards, find to have obtained more pure flora on solid medium, (diameter 1mm is left for the rounded convex of the flora on flat board
It is right)；
8. bacterium is taken on flat board with connecing collarium, to place into and carry out enrichment culture in fluid nutrient medium, object bacteria is obtained after 10d
Strain concentration pure bacterium solution higher；
9. Physiology and biochemistry, morphology and 16SrDNA is carried out to the acidophilia Thiobacillus thioxidans on this basis to be divided
Analysis identification.
Specifically, above-described SM culture medium prescriptions are as follows：
SM culture mediums (liquid)：Ammonium sulfate 0.4g, potassium dihydrogen sulfate 3.0g, seven water and magnesium sulfate 0.5g, two water are weighed respectively
With calcium chloride 0.25g, distilled water 1000ml is added, the amount that sulphur presses 10g/L is added.3- is placed under conditions of 28 DEG C, 180r/m
Obtained after 4d.
SM culture mediums (Gu)：A liquid：Weigh ammonium sulfate 2.0g, potassium dihydrogen sulfate 0.25g, seven water and magnesium sulfate 0.25g, chlorine
Change potassium 0.1g, four water and calcium nitrate 0.01g, adjust to 3 pH after adding distilled water 50ml；B liquid：Five water and sodium thiosulfate
22.1g, adds distilled water 100ml；C liquid：Agar 30g, adds distilled water 400ml.The liquid of A, B, C tri- is distinguished into 121 DEG C, 30min
After wet method sterilizing, 60 DEG C are cooled to, then three's mixing is down flat rapidly plate.
Applications of the bacterial strain ZJ-1 in removing heavy metals are removed.
1st, the activation of bacterial strain ZJ-1 and Amplification Culture：From picking single bacterium colony on SM solid mediums, 20mL liquid is inoculated in
In SM culture mediums, in 30 DEG C, 120r/min shaking table cultures 4 days or so, it was observed that muddy.The seed liquor of culture is connect by 1%
The amount of kind is inoculated in fresh 100mL culture mediums, in 30 DEG C, 120r/min shaking table cultures 5 days or so, it can be observed that obvious
It is muddy.
2nd, the domestication of bacterial strain ZJ-1：The bacterium solution that above-mentioned activation and Amplification Culture are obtained is inoculated according to 10% inoculum concentration
In the solid waste slurries of 100mL, and 0.3% sulphur and 0.04% ammonium sulfate are added thereto, 30 DEG C, 200r/min shaking tables
Culture, and detect wherein pH changes.
3rd, removals of the bacterial strain ZJ-1 to all kinds of heavy metals is acted on：The bacterium solution for completing will be tamed, is added into by 20% amount solid
In waste pulp, using pure solid waste as blank control group, the concentration of heavy metal ion in experimental group and blank group is monitored respectively, to sentence
Whether heavy metal is separated out in the form of ion in determining solid waste.
Beneficial effect：Small investment, operating cost are low, non-secondary pollution；Treatment effeciency is high；Condition of work requirement is low.
Brief description of the drawings
Table 1 is the nucleotides sequence list of acidophilia Thiobacillus thioxidans ZJ-1 16sRNA
Fig. 1 is the chadogram of ZJ-1
Fig. 2 is the apparent figure of strain culturing
Fig. 3 is by the strain morphology after Gram's staining
Fig. 4 is the clearance of the Cu ions under ZJ-1 effects
Fig. 5 is the clearance of the Zn ions under ZJ-1 effects
Fig. 6 is the precipitation clearance of the Cd under ZJ-1 effects
Fig. 7 is the precipitation clearance of the Pb under ZJ-1 effects
Specific embodiment
Explanation and specific embodiment are described further to technical scheme below in conjunction with the accompanying drawings, but are not construed as
Limiting the scope of the invention.Experimental technique used in following embodiments is conventional method without specified otherwise.
Experiment reagent consumptive material used in following embodiments etc. can be bought without specified otherwise from commercial use.
Embodiment 1
The screening technique of acidophilia Thiobacillus thioxidans Acidithiobacillus thiooxidans ZJ-1；
10g pickling mud samples are weighed, is put into equipped with 100ml (121 DEG C, 30min wet methods sterilized) neat liquid culture
In the conical flask of base (SM culture mediums), then sulphur powder is added thereto to, when turbid liquid pH value is down to 2.0 or so in bottle, is inhaled after standing
Supernatant liquor 10mL is taken, is inoculated into the 250mL conical flasks for filling the 100mL SM culture mediums of wet method sterilizing, and add 10g sulphur
Powder, is placed in culture (30 DEG C, 150rpm) in constant-temperature table；Culture 8d-10d, treats to become muddy milky by culture medium that (pH value is about
For 2).Therefrom draw 10mL to be inoculated into fresh SM culture mediums, after repetition enrichment culture 3-4 so that acidophilia sulfur oxide
Thiobacillus turns into dominant microflora therein；Culture medium to being enriched with is diluted coats solid SM culture mediums；Will be coated
Culture dish place 2h at room temperature after, be placed on and culture be inverted in 30 DEG C of incubators, after culture 15d, find solid medium
On obtained more pure flora, the rounded convex of the flora on flat board (diameter 1mm or so)；With connecing collarium on flat board
Bacterium is taken, to be placed into and carry out enrichment culture in fluid nutrient medium, aimed strain concentration pure bacterium solution higher is obtained after 10d.Such as
Fig. 2, Fig. 3.
Embodiment 2
Heavy metal in removal polluted soil；
Take testing sample and add water and be made mixed serum, determine all kinds of heavy metal concentrations therein.In each 250mL conical flask
Middle addition 120mL slurries, and a small amount of sulphur and ammonium sulfate are added thereto, the bacterium solution for completing will be tamed, it is added into by 20% amount
In solid waste slurries, to add the solid waste slurries of inactivated bacterial liquid as blank control group, a huge sum of money in experimental group and blank group is monitored respectively
Belong to the concentration of ion, the ratio of heavy metal concentration in slurries is accounted for as clearance with precipitation concentration.As shown in figs. 4-7.
The acidophilia Thiobacillus thioxidans ZJ-1 16s RNA sequence tables of table 1：
AGGAATCTGTCTTTGAGTGGGGGACAACCCAGGGAAACTTGGGCTAATACCGCATAAGCCCTGAGGGGGAAAGCGGG
GGATCTTCGGACCTCGCGCTGGAAGAGGAGCCTACGTCTGATTAGCTAGTTGGTAGGGTAAAGGCCTACCAAGGCGA
CGATCGGTAGCTGGTCTGAGAGGACGACCAGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCA
GTGGGGAATTTTTCGCAATGGGGGCAACCCTGACGAAGCAATGCCGCGTGAATGAAGAAGGCCTTCGGGTTGTAAAG
TTCTTTCGTGGAGGACGAAAAGGTGGGTGCTAATATCGCCTGCTGTTGACGTGAATCCAAGAAGAAGCACCGGCTAA
CTCCGTGCCAGCAGCCGCGGTAATACGGGGGGTGCAAGCGTTAATCGGAATCACTGGGCGTAAAGGGTGCGTAGGCG
GTGCATTAGGTCTGTCGTGAAATCCCCGGGCTCAACCTGGGAATGGCGGTGGAAACCGGTGTACTAGAGTATGGGAG
AGGGTGGTGGAATTCCAGGTGTAGCGGTGAAATGCGTAGAGATCTGGAGGAACATCAGTGGCGAAGGCGGCCACCTG
GCCCAATACTGACGCTGAGGCACGAAAGCGTGGGGAGCAAACAGGATTAGATACCCTGGTAGTCCACGCCCTAAACG
ATGAATACTAGATGTTTGGTGCCAAGCGTACTGAGTGTCGTAGCTAACGCGATAAGTATTCCGCCTGGGAAGTACGG
CCGCAAGGTTAAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGATGCAACGC
GAAGAACCTTACCTGGGCTTGACATGTCTGGAATCCTGCAGAGATGCGGGAGTGCCCTTCGGGGAATCAGAACACAG
GTGCTGCATGGCTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGTCCTTAG
TTGCCAGCGGTTCGGCCGGGCACTCTAGGGAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAGTCC
TCATGGCCTTTATGTCCAGGGCTACACACGTGCTACAATGGCGCGTACAGAGGGAAGCCAAGCCGCGAGGTGGAGCA
GACCCCAGAAAGCGCGTCGTAGTTCGGATTGCAGTCTGCAACTCGACTGCATGAAGTCGGAATCGCTAGTAATCGCG
GATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCC。
SEQUENCE LISTING
<110> Nanjing Tech University
<120>One plant of Thiobacillus thioxidans and its application in removing heavy metals are removed
<130> 20170427
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 1281
<212> DNA
<213> Acidithiobacillus thiooxidans ZJ-1
<400> 1
aggaatctgt ctttgagtgg gggacaaccc agggaaactt gggctaatac cgcataagcc 60
ctgaggggga aagcggggga tcttcggacc tcgcgctgga agaggagcct acgtctgatt 120
agctagttgg tagggtaaag gcctaccaag gcgacgatcg gtagctggtc tgagaggacg 180
accagccaca ctgggactga gacacggccc agactcctac gggaggcagc agtggggaat 240
ttttcgcaat gggggcaacc ctgacgaagc aatgccgcgt gaatgaagaa ggccttcggg 300
ttgtaaagtt ctttcgtgga ggacgaaaag gtgggtgcta atatcgcctg ctgttgacgt 360
gaatccaaga agaagcaccg gctaactccg tgccagcagc cgcggtaata cggggggtgc 420
aagcgttaat cggaatcact gggcgtaaag ggtgcgtagg cggtgcatta ggtctgtcgt 480
gaaatccccg ggctcaacct gggaatggcg gtggaaaccg gtgtactaga gtatgggaga 540
gggtggtgga attccaggtg tagcggtgaa atgcgtagag atctggagga acatcagtgg 600
cgaaggcggc cacctggccc aatactgacg ctgaggcacg aaagcgtggg gagcaaacag 660
gattagatac cctggtagtc cacgccctaa acgatgaata ctagatgttt ggtgccaagc 720
gtactgagtg tcgtagctaa cgcgataagt attccgcctg ggaagtacgg ccgcaaggtt 780
aaaactcaaa ggaattgacg ggggcccgca caagcggtgg agcatgtggt ttaattcgat 840
gcaacgcgaa gaaccttacc tgggcttgac atgtctggaa tcctgcagag atgcgggagt 900
gcccttcggg gaatcagaac acaggtgctg catggctgtc gtcagctcgt gtcgtgagat 960
gttgggttaa gtcccgcaac gagcgcaacc cttgtcctta gttgccagcg gttcggccgg 1020
gcactctagg gagactgccg gtgacaaacc ggaggaaggt ggggatgacg tcaagtcctc 1080
atggccttta tgtccagggc tacacacgtg ctacaatggc gcgtacagag ggaagccaag 1140
ccgcgaggtg gagcagaccc cagaaagcgc gtcgtagttc ggattgcagt ctgcaactcg 1200
actgcatgaa gtcggaatcg ctagtaatcg cggatcagca tgccgcggtg aatacgttcc 1260
cgggccttgt acacaccgcc c 1281
Claims (5)
1. a kind of Thiobacillus thioxidans, it is characterised in that Classification And Nomenclature is Acidithiobacillus thiooxidans ZJ-
1, China Microbial Culture Preservation Commission's common micro-organisms center is preserved on April 21st, 2016, deposit number is
CGMCC No:12387。
2. Thiobacillus thioxidans according to claim 1, it is characterised in that the nucleotide sequence of its 16s RNA such as institute of table 1
Show.
3. the screening technique of Thiobacillus thioxidans described in claim 1, it is characterised in that first with adding the liquid SM of sulphur powder to train
Support base bacterial strain is tamed and is enriched with, after repeating to tame and be enriched with 3-4 times, the culture medium for completing will be enriched with and be diluted coating
In solid SM culture mediums, by culture, the wherein circular, flora of convex is chosen；
It is prepared by SM culture mediums (liquid)：Ammonium sulfate 0.4g, potassium dihydrogen sulfate 3.0g, seven water and magnesium sulfate 0.5g, two water are weighed respectively
With calcium chloride 0.25g, distilled water 1000ml, sulphur 10g are added；Obtained after placing 3-4d under conditions of 28 DEG C, 180r/m；
SM culture mediums (Gu) prepare：
A liquid：Weigh ammonium sulfate 2.0g, potassium dihydrogen sulfate 0.25g, seven water and magnesium sulfate 0.25g, potassium chloride 0.1g, four water and nitre
Sour calcium 0.01g, is adjusted to 3 pH after adding distilled water 50ml；
B liquid：Five water and sodium thiosulfate 22.1g, add distilled water 100ml；
C liquid：Agar 30g, adds distilled water 400ml；
After the liquid of ABC tri- is distinguished into 121 DEG C, the sterilizing of 15min wet methods, 60 DEG C are cooled to, plate is down flat rapidly after three is well mixed.
4. the application of Thiobacillus thioxidans described in claim 1,2, it is characterised in that remove removing heavy metals.
5. the application of the Thiobacillus thioxidans according to claim 4, it is characterised in that applying step is as follows：
Bacterial strain is tamed with SM culture mediums first, heavy metal amount scope, root in heavy metal solid to be removed or liquid is detected
The amount of bacterium solution is added according to the amount selection of heavy metal.
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| CN107974422A (en) * | 2017-12-22 | 2018-05-01 | 中国科学院天津工业生物技术研究所 | A kind of bacterial strain administered for heavy metal pollution of soil |
| CN110724649A (en) * | 2019-10-09 | 2020-01-24 | 四川大学 | Efficient bioleaching composite bacterial system and application thereof in sludge dechromization process |
| CN113913324A (en) * | 2021-09-01 | 2022-01-11 | 郑州轻工业大学 | Composite microbial inoculum for efficient deodorization of biological filter and preparation method thereof |
| CN114874916A (en) * | 2022-03-28 | 2022-08-09 | 中国地质大学（武汉） | Scopulariopsis fungus (Sarocladium kiliense) ZJ-1 and application thereof |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107974422A (en) * | 2017-12-22 | 2018-05-01 | 中国科学院天津工业生物技术研究所 | A kind of bacterial strain administered for heavy metal pollution of soil |
| CN110724649A (en) * | 2019-10-09 | 2020-01-24 | 四川大学 | Efficient bioleaching composite bacterial system and application thereof in sludge dechromization process |
| CN110724649B (en) * | 2019-10-09 | 2021-07-09 | 四川大学 | Efficient bioleaching composite bacterial system and application thereof in sludge dechromization process |
| CN113913324A (en) * | 2021-09-01 | 2022-01-11 | 郑州轻工业大学 | Composite microbial inoculum for efficient deodorization of biological filter and preparation method thereof |
| CN113913324B (en) * | 2021-09-01 | 2023-06-30 | 郑州轻工业大学 | Composite microbial inoculant for efficient deodorization of biological filter and preparation method thereof |
| CN114874916A (en) * | 2022-03-28 | 2022-08-09 | 中国地质大学（武汉） | Scopulariopsis fungus (Sarocladium kiliense) ZJ-1 and application thereof |
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