CN102864086B - Strain capable of degrading polycyclic aromatic hydrocarbons (PAHs) and application thereof in soil remediation - Google Patents
Strain capable of degrading polycyclic aromatic hydrocarbons (PAHs) and application thereof in soil remediation Download PDFInfo
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- CN102864086B CN102864086B CN201210385473.9A CN201210385473A CN102864086B CN 102864086 B CN102864086 B CN 102864086B CN 201210385473 A CN201210385473 A CN 201210385473A CN 102864086 B CN102864086 B CN 102864086B
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- polycyclic aromatic
- aromatic hydrocarbons
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Abstract
The invention belongs to the technical field of environmental engineering and microbiology engineering, and particularly relates to a strain capable of degrading PAHs and an application thereof in soil remediation. The strain is a yeast (Candidainfanticola) tzyj with the collection number of CCTCCNO: M2012242 in the China Center for Type Culture Collection (CCTCC). The strain is capable of degrading the PAHs by using the PAHs in the soil as carbon sources, and can be used for remediation of soil polluted by the PAHs.
Description
Technical field
The invention belongs to environmental engineering and microbial engineering field, relate in particular to a strain degrading polycyclic aromatic hydrocarbons yeast (
Candida infanticola) tzyj and the application in soil remediation thereof.
Background technology
Polycyclic aromatic hydrocarbons (PAHs) is a class persistence organic pollutant (POPs).The PAHs source of pollution mainly come from the lubricating oil of waste and old lubricant, electric equipment products and disassemble from waste metal in the material of the hydrocarbon-containiproducts such as timber, organic high molecular compound, paper generate through thermolysis through incomplete combustion or in reducing atmosphere.Because Polycyclic aromatic hydrocarbons and Derivatives has carinogenicity, mutagenicity and teratogenecity, and acute toxicity and inferior lethal effect, people's very big concern caused at present.
The reparation of polycyclic aromatic hydrocarbons (PAHs) contaminated soil is the focus of research during current environment is repaired.Biological degradation is one of important channel that the PAHs contaminated soil is repaired.For polycyclic aromatic hydrocarbons contaminated, carried out the screening operation of a large amount of degrading polycyclic aromatic hydrocarbons microorganisms both at home and abroad.Up to the present, isolated a plurality of bacterial strains with degrading polycyclic aromatic hydrocarbons ability, as Rhodopseudomonas, Mycobacterium, bacillus etc., Cryptococcaceae is not used for the report that polycyclic aromatic hydrocarbons is repaired.
But the bioavailability of the PAHs in soil is generally lower, it has been generally acknowledged that, the PAHs or other the substrate that are dissolved in liquid phase can be utilized by microorganism, otherwise if be adsorbed on soil, is more difficultly utilized by microorganism.
Summary of the invention
The bioavailability that the present invention is directed to P in soil AHs in prior art is generally lower, the difficult weak point of being utilized by microorganism, a kind of degrading polycyclic aromatic hydrocarbons bacterial strain is provided, can utilizes the polycyclic aromatic hydrocarbons in soil to make carbon source and degrading polycyclic aromatic hydrocarbons, can be used for the reparation of polycyclic aromatic hydrocarbon pollution.
For achieving the above object, the present invention is achieved by the following technical programs:
The bacterial strain of one strain degrading polycyclic aromatic hydrocarbons is characterized in that, this bacterial strain be yeast (
Candida infanticola) tzyj, the preserving number at Chinese Typical Representative culture collection center is CCTCC NO:M 2012242, possesses the ability of degrading polycyclic aromatic hydrocarbons.
The ability that described bacterial strain has take Polycyclic aromatic hydrocarbons as sole carbon source and the energy is grown.
Described bacterial strain has the ability of good degrading polycyclic aromatic hydrocarbons in polycyclic aromatic hydrocarbon pollution.
Described polycyclic aromatic hydrocarbons is selected from one or more mixture of naphthalene, phenanthrene, fluoranthene, pyrene, benzanthrene, benzopyrene.
The ITS sequence of described bacterial strain is as shown in SEQ NO.1.
The application of the bacterial strain of described degrading polycyclic aromatic hydrocarbons in the biological restoration of polycyclic aromatic hydrocarbons contaminated soil.
Utilize the bacterial strain of described degrading polycyclic aromatic hydrocarbons to carry out the method that soil organisms is repaired, it is characterized in that the method is comprised of following steps: choose polycyclic aromatic hydrocarbon pollution 100g, with yeast
Candida infanticolaTzyj by mass percentage 2.0% consumption and mixing with soil even, add water and make soil moisture content remain on 15%, put into constant temperature oscillator, control culture temperature under the constant temperature of 25 ℃, shaking culture was repaired 120 hours, carried out liquid-phase chromatographic analysis after the extraction purification process.
Tzyj disassembles to separate and screen certain contaminated soil from the City of Taizhou solid waste to obtain.Bacterial strain can be at 15～35 ℃, better growth under pH value 6.0～8.5 culture condition; Bacterial strain is cultivated three days colony characteristicses on the YEPD flat board be circular, white milk cream shape, smooth surface, neat in edge; The cell oval carries out gemmation, cultivates to occur afterwards not too flourishing having every pseudohypha in seven days.
As preferably, bacterial strain of the present invention can be take polycyclic aromatic hydrocarbons as sole carbon source and energy growth.
The screening method of above-mentioned efficient polycyclic aromatic hydrocarbon-degrading bacteria, can implement successively as follows:
(1) disassemble from solid waste and gather 5～20cm topsoil sample under the earth's surface, as the bacterium source carrying out the nature enrichment culture; Described enrichment medium contains: Tryptones, glucose and yeast extract paste;
(2) get enrichment culture liquid described in (1) and be inoculated in inorganic salt and select to select in substratum to cultivate, institute's inorganic salt select substratum to contain: K
2HPO
4, MgSO
47H
2O, (NH
4)
2SO
4, (concentration of polycyclic aromatic hydrocarbons naphthalene, phenanthrene, fluoranthene, pyrene, benzanthrene, benzopyrene is 150mg/L～250mg/L) to the polycyclic aromatic hydrocarbons such as naphthalene, phenanthrene, fluoranthene, pyrene, benzanthrene, benzopyrene;
(3) get and select the culture dilution spread to select to separate namely obtaining polycyclic aromatic hydrocarbon-degrading bacteria tzyj on substratum in solid inorganic salt described in (2).
Above-mentioned efficient polycyclic aromatic hydrocarbon-degrading bacteria tzyj can be used for the biological restoration of polycyclic aromatic hydrocarbons soil pollution.
Tzyj of the present invention can be at 15～35 ℃, better growth under pH value 6.0～8.5 culture condition; Bacterial strain is cultivated three days colony characteristicses on the YEPD flat board be circular, white milk cream shape, smooth surface, neat in edge; The cell oval carries out gemmation, cultivates to occur afterwards not too flourishing having every pseudohypha in seven days.
Tzyj is at YEPD(glucose: 2.0 %; Tryptones: 2.0; Yeast powder: it is circular 1.0%) cultivating three days colony characteristicses, white milk cream shape, smooth surface, neat in edge; The cell oval carries out gemmation, cultivates to occur afterwards not too flourishing having every pseudohypha in seven days.
The physiological and biochemical property of tzyj sees Table 1.
The physiological and biochemical property of table 1 tzyj
The pcr amplification of the ITS gene of tzyj, order-checking are completed by Beijing three rich polygala root company, and having obtained length is the ITS sequence of 1109bp.Select homology to carry out Phylogenetic Analysis greater than 97% gene order, show bacterial strain with
Candida infanticolaHomology is the highest, in conjunction with strain morphology, colony characteristics and physio-biochemical characteristics, identifies that bacterial strain belongs to
Candida infanticola
Tzyj of the present invention both can be at nutritional medium, as: grow in common czapek's solution, common YEPD, can grow take polycyclic aromatic hydrocarbons as sole carbon source, in the inorganic salt basic medium of the energy again.
In sum, the present invention has the following advantages: tzyj provided by the invention can utilize the polycyclic aromatic hydrocarbons in soil to make carbon source, and the polycyclic aromatic hydrocarbons in contaminated soil is had the fast degradation ability, and degraded substrate scope is wide, and degradation rate is high.Make it firmly in repairing, wide application potential be arranged in the polycyclic aromatic hydrocarbons contaminated field of absorption at contaminated soil.
The Latin literary fame:
Candida infanticolaTzyj
Depositary institution: Chinese Typical Representative culture collection center
Address: Wuhan, China Wuhan University
Preservation date: on June 24th, 2012
Deposit number: CCTCC NO:M 2012242.
Embodiment
Below by specific embodiment, technical scheme of the present invention is described in further detail, but the present invention is not limited to these embodiment.
Embodiment 1
The separating screening method of Tzyj provided by the invention.
disassemble from solid waste and gather 5～20cm topsoil sample under the earth's surface, as the bacterium source carrying out the nature enrichment culture, with glucose 10g, Tryptones 10g, yeast powder 5g joins in a certain amount of distilled water, the nutrient solution for preparing is sterilising treatment 20min under the temperature condition of 121 ℃, bacterium liquid was cultivated three days on this substratum, prepare again the inorganic salt basic medium, add naphthalene, luxuriant and rich with fragrance, fluoranthene, pyrene, benzanthrene, the polycyclic aromatic hydrocarbon compounds of benzopyrene, the above-mentioned naphthalene that adds, luxuriant and rich with fragrance, fluoranthene, pyrene, benzanthrene, the add-on of benzopyrene is 150mg/mL, cultivated three days on this substratum, dilution spread is isolated a strain bacterium called after tzyj, this bacterium is preserved in " its preserving number of Chinese Typical Representative culture collection " center " CCTCC NO:M 2012242 on June 24th, 2012.
The pcr amplification of the ITS gene of tzyj, order-checking are completed by Beijing three rich polygala root company.According to the sequencing result phylogenetic tree construction.Phylogeny show bacterial strain with
Candida infanticolaEvolutionary distance nearest, combining form, colony characteristics and physio-biochemical characteristics identify that bacterial strain belongs to
Candida infanticola
Embodiment 2
Polycyclic aromatic hydrocarbons in tzyj pollution degradation soil.
Choose polycyclic aromatic hydrocarbon pollution 100g, tzyj presses the 2.0%(mass percent) consumption and mixing with soil even, adding a small amount of water makes soil moisture content remain on 15% left and right, put into constant temperature oscillator, control culture temperature under the constant temperature of 25 ℃, shaking culture was repaired 120 hours, carried out liquid-phase chromatographic analysis after the extraction purification process, and the degradation rate of contained naphthalene, phenanthrene, fluoranthene, pyrene, benzanthrene, benzopyrene is more than 90%.
Specific embodiment described in the present invention is only to the explanation for example of the present invention's spirit.Those skilled in the art can make various modifications or replenish or adopt similar mode to substitute described specific embodiment; but can't depart from spirit of the present invention or surmount the defined scope of appended claims; within the spirit and principles in the present invention all; any modification of doing, be equal to replacement, improvement etc., within protection scope of the present invention all should be included in.
SEQUENCE LISTING
＜110〉Taizhou Vocational ﹠ Technical College
＜120〉bacterial strain of a strain degrading polycyclic aromatic hydrocarbons and the application in soil remediation thereof
<130>
<160> 1
<170> PatentIn version 3.3
<210> 1
<211> 1109
<212> DNA
<213> Candida infanticola
<400> 1
ttgtacacac gcccgtcgtt actaccgatt gaatggctta gtgaggcctc cggatgcgat 60
tcacttgtgg gcaaccgcaa cgaagtcgca gaagctggac aaacttggtc atttagagga 120
agtacaagtc gtaacaaggt ttccgtaggt gaacctggga aggatcatta ttgagattca 180
tattacacct gtgaaacaac tgaattgctt ggccgcaagg ccaatgtaac aaaaactatt 240
ttacmtatta tatctgaaaa acgaaatcaa aagtttcaac aacggatctc ttggttctcg 300
catcgagaag aacgcagcaa agcgcgatag ttagtgtgaa ttgcagacgt gaatcattga 360
gtttttgaac gcacattgca ccttctggta ttccgggagg tatacttgtg cgagcgtcat 420
ttcatcttca taaagcaatt tatgtgttgg ggctgtagcc agccttgaaa aagatgatag 480
agtacatgtt agacacaatg tgcttttcta tatttttgac ctcgtatcaa gcaagattac 540
ccgctgaact taagcatatc aataagcgga ggaaaagaaa ccaacaggga ttgccttagt 600
agcggcgagt gaagcggcaa tagctcaaat ttgaaatctg ccaacaggcc gagttgtaat 660
ttgaagaaag caattctgga tatgtctttg ctcaagttcc ttggaacagg acatcatgga 720
gggtgagaat cccgtgatgc gaaggtagca tatctatgta gaatgttttc gacgagtcga 780
gttgtttggg aatgcagctc aaagtgggtg gtaaattcca tctaaagcta aatattggcg 840
agagaccgat agcgaacaag tactgtgaag gaaagatgaa aagaactttg aaaagagagt 900
gaaaaagtac gtgaaattgt tgaaatggaa gggcttgaga ttagacatgg tgtttagtgt 960
tccactttct tttgggaatg tgtactcgct ttwcaccggg ccagcatcag ttttggtggg 1020
aggataaatg gtattgaatg tggctgcttg cagtgttata gctttatcaa atacttccaa 1080
ccgggactga ggaccgcgca ttgctagga 1109
Claims (5)
1. the bacterial strain of a strain degrading polycyclic aromatic hydrocarbons, is characterized in that, this bacterial strain be yeast (
Candida infanticola) tzyj, the preserving number at Chinese Typical Representative culture collection center is CCTCC NO:M 2012242, possesses the ability of degrading polycyclic aromatic hydrocarbons.
2. the bacterial strain of degrading polycyclic aromatic hydrocarbons according to claim 1, is characterized in that the ability that described bacterial strain has take polycyclic aromatic hydrocarbons as sole carbon source and the energy is grown.
3. polycyclic aromatic hydrocarbon-degrading bacteria strain according to claim 1 is characterized in that described bacterial strain has the ability of good degrading polycyclic aromatic hydrocarbons in polycyclic aromatic hydrocarbon pollution.
4. polycyclic aromatic hydrocarbon-degrading bacteria strain according to claim 1 is characterized in that described polycyclic aromatic hydrocarbons is selected from one or more mixture of naphthalene, phenanthrene, fluoranthene, pyrene, benzanthrene, benzopyrene.
5. the bacterial strain of degrading polycyclic aromatic hydrocarbons according to claim 1, is characterized in that the ITS sequence of described bacterial strain is as shown in SEQ NO.1.
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| CN104673683B (en) * | 2015-02-27 | 2017-10-31 | 台州职业技术学院 | One plant of salt tolerant polycyclic aromatic hydrocarbon-degrading bacteria Tzyjm and its application in marine pollution environment |
| CN104877934B (en) * | 2015-02-27 | 2018-01-09 | 台州职业技术学院 | One plant of salt tolerant polycyclic aromatic hydrocarbon-degrading bacteria Tzymx and its application in marine pollution environment |
| CN110656079A (en) * | 2019-11-02 | 2020-01-07 | 东华大学 | Preparation method of naphthalene restoration and degradation microbial inoculum in petroleum hydrocarbon polluted soil |
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| Unraveling the decolourizing ability of yeast isolates from dye-polluted and virgin environments: an ecological and taxonomical overview;Hipolito F. Pajot et al;《Antonie van Leeuwenhoek》;20100822;第99卷;443-456 * |
| 土壤/沉积物中石油烃微生物降解研究综述;郭伟等;《矿物岩石地球化学通报》;20070731;第26卷(第3去期);276-283 * |
| 绿色化学理念在化学化工过程中的实施;马俊林等;《十堰职业技术学院学报》;20061031;第19卷(第5期);64-66 * |
| 郭伟等.土壤/沉积物中石油烃微生物降解研究综述.《矿物岩石地球化学通报》.2007,第26卷(第3去期),276-283. |
| 马俊林等.绿色化学理念在化学化工过程中的实施.《十堰职业技术学院学报》.2006,第19卷(第5期),64-66. |
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Effective date of registration: 20151223 Address after: Hangzhou City, Zhejiang province 310053 Binjiang District Chunxiao Road No. 529 Jiangnan constellation A-2105 Patentee after: HANGZHOU WELLMOTO ENVIRONMENTAL TECHNOLOGY Co.,Ltd. Address before: 318000 Xueyuan Road, Jiaojiang, Zhejiang, No. 788, Patentee before: Taizhou Vocational & Technical College |
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