CN116855428B - Multifunctional microorganism strain, organic pollution repair microbial agent and application thereof - Google Patents
Multifunctional microorganism strain, organic pollution repair microbial agent and application thereof Download PDFInfo
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- 241000589776 Pseudomonas putida Species 0.000 claims abstract description 36
- 238000006731 degradation reaction Methods 0.000 claims abstract description 26
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- 239000002068 microbial inoculum Substances 0.000 claims abstract description 25
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B09—DISPOSAL OF SOLID WASTE; RECLAMATION OF CONTAMINATED SOIL
- B09C—RECLAMATION OF CONTAMINATED SOIL
- B09C1/00—Reclamation of contaminated soil
- B09C1/10—Reclamation of contaminated soil microbiologically, biologically or by using enzymes
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- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
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- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
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- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F2101/00—Nature of the contaminant
- C02F2101/30—Organic compounds
- C02F2101/32—Hydrocarbons, e.g. oil
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/38—Pseudomonas
- C12R2001/40—Pseudomonas putida
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Abstract
The invention relates to a multifunctional microorganism strain and an organic pollution repair microbial agent and application thereof, and belongs to the technical field of microbial agents. The invention screens and obtains a strain Pseudomonas putida AM21 with the capability of efficiently repairing the petroleum-polluted soil from the petroleum-polluted soil, the strain can degrade various polycyclic aromatic hydrocarbons and benzene series, and the microbial inoculum prepared by the Pseudomonas putida AM21 can effectively repair the petroleum-polluted soil and coked-polluted soil. The degradation rate of the repairing microbial inoculum prepared by pseudomonas putida AM21 on petroleum with the oil content of 3-8% and petroleum hydrocarbon of coked and combined polluted soil can reach more than 60% within 60d, and the degradation rate of the repairing microbial inoculum on various polycyclic aromatic hydrocarbon and benzene series can reach more than 70%.
Description
Technical Field
The invention relates to a multifunctional microorganism strain and an organic pollution repair microbial agent and application thereof, and belongs to the technical field of microbial agents.
Background
With the continuous increase of petroleum exploitation and the increasing severity of petroleum pollution, the restoration of petroleum polluted soil becomes a hot spot of current research. The existing methods for repairing petroleum pollution mainly comprise physical repair, chemical repair and biological repair. The physical repair method and the chemical repair method have high cost, have large disturbance to the soil environment and are easy to destroy the soil ecology; the bioremediation method has the advantages of low cost, no secondary pollution and the like, and is therefore valued by people. The soil microorganism repairing technology is a repairing technology for reducing the activity of harmful pollutants in soil or degrading the harmful pollutants into harmless substances by utilizing indigenous microorganisms or microorganisms with specific functions which are artificially domesticated or screened under proper environmental conditions and through the metabolism of the microorganisms. The microbial remediation principle of organically contaminated soil mainly comprises degradation and transformation of microorganisms, which is usually realized by means of reaction modes such as oxidation, reduction, gene transfer, hydrolysis and the like. The soil microbial remediation technology has low application cost and small negative influence on soil fertility and metabolic activity, and can avoid the influence on human health and environment caused by pollutant transfer. Therefore, the research of the soil microorganism repair technology has important significance.
Ireland patent publication IES20140311A2 (application number IES 20140311) discloses a method for remediation of organic and/or heavy metal contaminated substrates for remediation of organic contaminated and/or heavy metal contaminated soil, sediment and/or sludge. The method comprises the processes of biostimulation, biological addition, rhizosphere repair, plant repair, original ecological stack and the like. The method comprises the following steps: a. designing a microbial agent comprising pollutant-degrading bacteria, plant-promoting bacteria and/or biosurfactant-producing bacteria; b. pretreating a substrate; c. building an ecological pile; d. and sowing and repairing plants at the top of the ecological stack. The above-mentioned patent realizes promoting plant growth and repairing organic pollution and heavy metal pollution's effect jointly through the cooperation of multiple microorganism. However, due to the combined action of a plurality of microorganisms, the condition for coordinating the growth characteristics of the microorganisms is complex, and the technical requirements on the types and the quantity of the matched microorganisms are very high.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides a multifunctional microorganism strain, an organic pollution repair microbial agent and application thereof, and the organic pollution repair microbial agent has good effects of degrading petroleum hydrocarbon, polycyclic aromatic hydrocarbon, benzene series and the like, thereby achieving the purpose of rapidly repairing organic pollution soil.
The technical scheme of the invention is as follows:
pseudomonas putidaPseudomonas putida) AM21, deposited at the China general microbiological culture Collection center, accession number: the institute of microbiological science, the institute of China, national academy of sciences, no. 3, north Chen West Lu, no.1, chaoyang, beijing, city, accession number: CGMCC No. 27033.
The pseudomonas putida is [ ]Pseudomonas putida) AM21, hereinafter referred to as Pseudomonas putida AM21.
The culture method of the pseudomonas putida AM21 comprises the following steps:
(1) Inoculating pseudomonas putida AM21 into a solid activating culture medium, and carrying out activating culture to obtain an activated strain;
(2) Inoculating the activated strain prepared in the step (1) into a liquid culture medium for culture to prepare seed liquid;
(3) And (3) transferring the seed liquid prepared in the step (2) into an expansion culture medium according to the volume percentage of 2-10%, and carrying out expansion culture to obtain pseudomonas putida AM21 bacterial liquid.
According to a preferred embodiment of the invention, the Pseudomonas putida AM21 of step (1) is streaked onto a solid activation medium.
According to a preferred embodiment of the present invention, the solid activation medium in step (1) comprises the following components:
10g/L peptone, 5g/L yeast extract powder, 10g/L sodium chloride, 20g/L agar, the balance of water and natural pH.
According to a preferred embodiment of the present invention, the conditions for the activation culture in step (1) are: culturing at 33-37 deg.c for 1-2 days.
According to a preferred embodiment of the present invention, the liquid medium in step (2) and the expansion medium in step (3) have the following composition:
10g/L peptone, 5g/L yeast extract powder, 10g/L sodium chloride and the balance of water, and the pH is natural.
According to a preferred embodiment of the present invention, the culturing conditions in step (2) are: culturing for 1-2 days at 33-37 deg.C and 100-200 rpm.
According to a preferred embodiment of the present invention, the conditions for the expansion culture in the step (3) are: and (3) performing expansion culture for 1-2 days under the conditions of 33-37 ℃ and dissolved oxygen of 20-30%.
An organic pollution repair microbial agent comprises the pseudomonas putida AM21.
According to the invention, the organic pollution remediation microbial inoculum also comprises an organic matter carrier.
Further preferably, the organic pollution remediation microbial inoculum is prepared by mixing pseudomonas putida AM21 bacterial liquid with an organic matter carrier.
Further preferably, the organic matter carrier is turfy soil.
According to the present invention, the viable bacteria concentration of the organic pollution repair microbial agent is preferably (2 to 5). Times.10 9 cfu/g。
In the preferred technical scheme of the invention, the organic pollution repair microbial inoculum is prepared by mixing pseudomonas putida AM21 bacterial liquid with an organic matter carrier, and the organic matter carrier can provide a certain nutrient substance for the bacterial strain, so that the bacterial strain can reproduce in the organic matter carrier and keep activity.
The organic pollution remediation microbial inoculum is applied to remediation of petroleum and/or coked polluted soil.
According to a preferred embodiment of the invention, the application comprises the following steps:
inoculating the organic pollution remediation microbial inoculum into petroleum and/or coked polluted soil with the oil content of 3-8%, wherein the mass ratio of the organic pollution remediation microbial inoculum to the polluted soil is (1-5):100, adjusting the water content to 20-30%, mixing uniformly, and stacking at the temperature of 30+/-5 ℃ for degradation.
The pseudomonas putida AM21 is applied to the restoration of petroleum-polluted soil.
The application of the pseudomonas putida AM21 in repairing coking polluted soil.
The pseudomonas putida AM21 is applied to the restoration of petroleum and coking composite polluted soil.
The beneficial effects are that:
1. the invention screens and obtains a strain Pseudomonas putida AM21 with the capability of efficiently repairing the petroleum-polluted soil from the petroleum-polluted soil, and the strain can degrade various polycyclic aromatic hydrocarbons and benzene series. The microbial inoculum prepared from pseudomonas putida AM21 can effectively repair petroleum-polluted soil and coking-polluted soil.
2. The repairing microbial inoculum prepared by pseudomonas putida AM21 has good repairing efficiency on petroleum and coking composite polluted soil, the degradation rate of petroleum hydrocarbon on petroleum and coking composite polluted soil with the oil content of 3-8% can reach more than 60% within 60d, and the degradation rate of various polycyclic aromatic hydrocarbon and benzene series can reach more than 70%. Therefore, the organic pollution remediation microbial inoculum disclosed by the invention has a good remediation effect on the soil subjected to the combined pollution of petroleum hydrocarbon and high-concentration organic matters in a coking plant, can degrade petroleum hydrocarbon, polycyclic aromatic hydrocarbon, benzene series and the like in the combined pollution soil, and has high remediation efficiency.
Drawings
FIG. 1 is a 16S rDNA agarose gel electrophoresis of Pseudomonas putida AM21.
Detailed Description
The technical scheme of the present invention will be further described with reference to examples, but the scope of the present invention is not limited thereto. The reagents and materials used in the examples were the same as those of the conventional commercial products unless otherwise specified.
Example 1
Isolation and identification of Pseudomonas putida AM 21:
collecting high-concentration petroleum-contaminated soil 5g of victory oil field, placing in 250 mL sterile triangular flask, adding sterile inorganic salt culture medium 100 mL, culturing at 30deg.C and 150 rpm for 5 d, absorbing bacterial liquid, gradient diluting with sterile water to 10 -1 ，10 -2 ，10 -3 ，10 -4 ，10 -5 The number of times of the number of times,100 mu L of each of the culture solution is coated on a sterile petroleum-inorganic salt solid culture medium, the culture solution is subjected to stationary culture at 30 ℃ for 3 d, the clone with quick growth and large colony is selected to be 100 mL petroleum-inorganic salt liquid culture medium, the culture solution is subjected to culture at 30 ℃ for 3 d at 150 rpm, 100 mu L of bacterial solution is absorbed and coated on the sterile petroleum-inorganic salt solid culture medium, and single colony is selected.
Wherein, the inorganic salt culture medium comprises the following components per liter:
KNO 3 1.5g，(NH 4 ) 2 SO 4 1.5g，K 2 HPO 4 1g，KH 2 PO 4 1g，MgSO 4 ·7H 2 O 0.5g，NaCl 130g，FeSO 4 ·7H 2 O 0.01 g，dH 2 o is fixed to 1L;
the petroleum-inorganic salt solid medium comprises the following components per liter:
KNO 3 1.5g，(NH 4 ) 2 SO 4 1.5g，K 2 HPO 4 1g，KH 2 PO 4 1g，MgSO 4 ·7H 2 O 0.5g，NaCl 130g，FeSO 4 ·7H 2 o0.01 g, petroleum 20g, agar 20g, dH 2 O is fixed to 1L;
the petroleum-inorganic salt liquid culture medium comprises the following components per liter:
KNO 3 1.5g，(NH 4 ) 2 SO 4 1.5g，K 2 HPO 4 1g，KH 2 PO 4 1g，MgSO 4 ·7H 2 O 0.5g，NaCl 130g，FeSO 4 ·7H 2 o0.01 g, petroleum 20g, dH 2 O is fixed to volume of 1L.
The single colony is sent to sequencing company for sequencing, and the 16S rDNA sequence contains 1390 and bp, and the nucleotide sequence is shown as SEQ ID NO. 1.
The strain identification process is as follows:
sample: the bacterial liquid obtained by the single colony culture is selected;
bacterial genomic DNA extraction kit: bioengineering (Shanghai) stock Co.Ltd;
TAE buffer (50×, 1L): t (T)ris 242g, glacial acetic acid 57.1mL, na 2 EDTA·2H 2 O37.2 g, water to 1L;
agarose: BIOWET, AGAROSE G-10;
2× Pfu PCR MasterMix, D2000 DNA Marker, nucleic acid dye, loading buffer, etc.: bioengineering (Shanghai) stock Co.Ltd;
DNA purification recovery kit: bioengineering (Shanghai) stock Co.Ltd;
centrifuge tube, gun head and other consumables: gene Era Biotech, USA;
primer: synthesized by Shanghai Co., ltd, ddH was added according to the synthetic order 2 O, 10. Mu.M solution was prepared.
1. Genomic DNA extraction was performed according to the bacterial genomic DNA extraction kit instructions.
2.PCR amplification
2.1 general primer information, see Table 1:
TABLE 1 general primer information
2.2 The components and the constitution of the PCR amplification system are shown in Table 2:
TABLE 2 PCR amplification System Components and constitution
2.3 PCR amplification cycle parameters
Pre-denaturation: 94 ℃ for 3min; denaturation 94 ℃,30s, annealing 55 ℃,30s, extension, 72 ℃,1.5min (total 35 cycles); extending at 72deg.C for 10min; preserving at 4 ℃.
3. Agarose gel electrophoresis detection
A1.0% agarose gel was prepared, the voltage was set at 18V/cm during electrophoresis, and the electrophoresis time was 20min. Agarose electrophoresis staining was performed using nucleic acid dyes, and photographing was performed using an ultraviolet gel imaging system. The results are shown in FIG. 1.
4. Purification recovery
The target fragment was subjected to agarose gel recovery using a general agarose gel DNA recovery kit, and the recovered product was sent to the engineering bioengineering (Shanghai) Co.Ltd for sequencing.
Sequencing splice sequences and blast comparison results are shown in Table 3:
TABLE 3 sequencing splice sequence blast comparison results
Sequence alignment was performed by sequence 16S rDNA, and the strain with the closest genetic relationship was found to bePseudomonas putida。
Through the identification of the strains, the strains screened by the invention belong toPseudomonas putidaThe product is named as Pseudomonas putida AM21 and is preserved in China general microbiological culture Collection center, with the preservation address: the institute of microbiological science, the institute of China, national academy of sciences, no. 3, north Chen West Lu, no.1, chaoyang, beijing, city, accession number: CGMCC No. 27033.
Example 2
The culture method of pseudomonas putida AM21 comprises the following steps:
(1) Taking pseudomonas putida AM21 streak to inoculate on a solid activation culture medium, and carrying out inverted activation culture at 35 ℃ for 2 days to obtain an activated strain;
wherein, the solid activation culture medium comprises the following components per liter: 10g of peptone, 5g of yeast extract powder, 10g of sodium chloride, 20g of agar, and water to a constant volume of 1L, wherein the pH is natural;
(2) Inoculating the activated strain prepared in the step (1) into a liquid culture medium, and performing shake culture for 2 days at 35 ℃ and a rotating speed of 150 rpm to obtain seed liquid;
wherein, the liquid culture medium comprises the following components per liter: 10g of peptone, 5g of yeast extract powder, 10g of sodium chloride, and water to a constant volume of 1L, wherein the pH is natural;
(3) Taking the seed liquid prepared in the step (2), and turning the seed liquid into a seed liquid with the volume percentage of 2%Placing into an expansion culture medium, and performing expansion culture at 35deg.C under 30% dissolved oxygen for 2 days to obtain Pseudomonas putida AM21 bacterial liquid with viable bacteria concentration of 4×10 9 cfu/mL；
Wherein, the expansion medium comprises the following components per liter: 10g of peptone, 5g of yeast extract powder, 10g of sodium chloride and water to a volume of 1L, and the pH is natural.
Example 3
An organic pollution repair microbial agent is prepared by mixing Pseudomonas putida AM21 bacterial liquid prepared in example 2 with turfy soil at a mass ratio of 1:10, and naturally standing for two days.
Through detection, the concentration of viable bacteria in the organic pollution remediation microbial inoculum is 3.3X10 9 cfu/g。
Example 4
The application of the organic pollution remediation microbial inoculum containing pseudomonas putida AM21 in the remediation of petroleum-polluted soil comprises the following steps:
(1) Uniformly mixing petroleum-polluted soil with oil content of 4.3% with the organic pollution remediation microbial inoculum prepared in the embodiment 3 according to the mass ratio of 50:1 to obtain a microbial soil mixture, and regulating the water content to 25% by using sterilized distilled water;
(2) The water content of the fungus soil mixture is kept to be 25%, and 30d of fungus soil mixture is piled and degraded at the temperature of 30+/-5 ℃.
After the degradation is finished, the degradation rate of petroleum hydrocarbon in the soil is measured by adopting an infrared spectrophotometry (HJ 1051-2019), and after the petroleum polluted soil with the oil content of 4.3% is degraded for 30d by using the organic pollution repair microbial inoculum prepared by pseudomonas putida AM21, the oil content of the soil is 2.05%, and the degradation rate of the petroleum hydrocarbon can reach 52.3%.
Example 5
The application of the organic pollution remediation microbial inoculum containing pseudomonas putida AM21 in remediation of coked polluted soil comprises the following steps:
(1) Uniformly mixing coked contaminated soil and the organic pollution remediation microbial inoculum prepared in the embodiment 3 according to the mass ratio of 50:1 to obtain a microbial soil mixture, and regulating the water content to 25% by using sterilized distilled water;
(2) The water content of the fungus soil mixture is kept to be 25%, and the fungus soil mixture is piled and degraded at 30+/-5 ℃ for 40 d.
After the degradation is completed, the content of main pollutants in the soil is detected, and compared with the soil before restoration, the degradation rate of the organic pollutants is calculated, and the results are shown in the following table:
TABLE 4 degradation of organic pollutants in coked contaminated soil
As shown in Table 4, the organic pollution remediation microbial inoculum of the invention has strong degradation capability to various pollutants in the coking polluted soil; wherein, the degradation rate of fluorene, phenanthrene, fluoranthene and pyrene reaches more than 90 percent, the degradation rate of naphthalene reaches more than 80 percent, the degradation rate of anthracene, benzene and toluene reaches more than 70 percent, and the material has good degradation capability for m-xylene, p-xylene, benzo (a) anthracene, benzo (b) fluoranthene and benzo (a) pyrene.
Example 6
The application of the organic pollution remediation microbial inoculum containing pseudomonas putida AM21 in the remediation of petroleum and coking composite polluted soil comprises the following steps:
(1) Mixing petroleum-polluted soil (oil content 4.3%) and coking-polluted soil according to a mass ratio of 1:1, and uniformly stirring;
(2) Adding the organic pollution remediation microbial inoculum prepared in the embodiment 3 into the soil uniformly stirred in the step (1) according to the mass ratio of 1:50;
(3) Adjusting the water content to 25% by using sterilized distilled water, and stacking and degrading for 60d at 30+ -5deg.C, wherein the water content is kept to 25% in the degradation process.
After the degradation is completed, the content of main pollutants in the soil is detected, and compared with the soil before restoration, the degradation rate of the organic pollutants is calculated, and the results are shown in the following table:
TABLE 5 degradation of organic pollutants in Petroleum and coking composite contaminated soil
As shown in Table 5, after the soil is restored for 60 days, the degradation rate of petroleum hydrocarbon reaches 64.1%, and the highest degradation rate of the petroleum hydrocarbon reaches 97.03% because the petroleum hydrocarbon can be effectively degraded for various polycyclic aromatic hydrocarbons. The degradation rate of the parabenzene series reaches 74.3% -83.4%. The result shows that the organic pollution remediation microbial inoculum has good remediation effect on petroleum and coking composite polluted soil.
Analysis of results:
according to the degradation data of the embodiment, the pseudomonas putida AM21 disclosed by the invention can be used for rapidly repairing petroleum-polluted soil of an oil field, has a good repairing effect on Polycyclic Aromatic Hydrocarbons (PAHs) polluted soil of a coking plant, can rapidly and effectively repair petroleum-coking composite polluted soil, widens the application range of the organic pollution repairing microbial inoculum, and has a relatively wide application prospect.
Claims (10)
1. The multifunctional microorganism strain is characterized by being pseudomonas putidaPseudomonas putida) AM21, deposited at the China general microbiological culture Collection center, accession number: the institute of microbiological science, the institute of China, national academy of sciences, no. 3, north Chen West Lu, no.1, chaoyang, beijing, city, accession number: CGMCC No. 27033.
2. The method for culturing a multifunctional microorganism strain according to claim 1, comprising the steps of:
(1) Taking pseudomonas putida @Pseudomonas putida) AM21 is inoculated in a solid activating culture medium, and activated strain is prepared after activating culture;
(2) Inoculating the activated strain prepared in the step (1) into a liquid culture medium for culture to prepare seed liquid;
(3) Transferring the seed liquid prepared in the step (2) into an expansion culture medium according to the volume percentage of 2-10%, and carrying out expansion culture to obtain pseudomonas putida @Pseudomonas putida) AM21 bacterial liquid.
3. The method of claim 2, wherein one or more of the following conditions are satisfied:
i. the pseudomonas putida in the step (1)Pseudomonas putida) AM21 streaks were inoculated onto solid activation media;
the components of the solid activation medium in step (1) are as follows:
10g/L peptone, 5g/L yeast extract powder, 10g/L sodium chloride, 20g/L agar, the balance of water and natural pH;
the conditions of the activation culture in step (1) are: culturing for 1-2 days at 33-37 ℃;
the liquid medium in step (2) and the expansion medium in step (3) have the following composition:
10g/L peptone, 5g/L yeast extract powder, 10g/L sodium chloride and the balance of water, and the pH is natural;
the culture conditions in step (2) are: culturing for 1-2 days at the temperature of 33-37 ℃ and the speed of 100-200 rpm;
the conditions of the expansion culture in the step (3) are as follows: and (3) performing expansion culture for 1-2 days under the conditions of 33-37 ℃ and dissolved oxygen of 20-30%.
4. Use of a multifunctional microbial strain according to claim 1 for remediation of petroleum and/or coked contaminated soil.
5. An organic pollution repair microbial agent, which is characterized by comprising pseudomonas putida according to claim 1Pseudomonas putida)AM21。
6. An organic pollution remediation agent according to claim 5 wherein the organic pollution remediation agent further comprises an organic carrier.
7. An organic pollution remediation microbial agent according to claim 6 wherein the organic carrier is peatmoss.
8. An organic contamination repair fungus agent according to claim 5, wherein the viable bacteria concentration of the organic contamination repair fungus agent is (2-5) x 10 9 cfu/g。
9. The use of an organic pollution remediation microbial agent of claim 5 for remediating petroleum and/or coked contaminated soil.
10. The use of an organic pollution remediation agent as claimed in claim 9, wherein the use comprises the steps of:
inoculating the organic pollution remediation microbial inoculum of claim 5 into petroleum and/or coked polluted soil with oil content of 3-8%, regulating the water content to 20-30% according to the mass ratio of the organic pollution remediation microbial inoculum to the polluted soil of (1-5):100, mixing uniformly, and stacking at 30+/-5 ℃ for degradation.
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