Pseudomonas geniculata, fungicide, application and method for degrading polycyclic aromatic hydrocarbon

Abstract

The invention belongs to the technical field of soil and groundwater pollution remediation and bioengineering, and relates to Pseudomonas geniculata, a microbial agent, application and a method for degrading polycyclic aromatic hydrocarbon. The preservation number of the pseudomonas geniculata is CGMCC (China General Microbiological Culture Collection Center) No. 22735. The Pseudomonas aeruginosa or microbial inoculum has efficient polycyclic aromatic hydrocarbon degradation capability, can be used for repairing polycyclic aromatic hydrocarbon polluted soil or underground water, and has a good application prospect in green and sustainable repairing of polycyclic aromatic hydrocarbon polluted sites.

Description

Technical Field

[0001] The present invention belongs to the fields of soil and groundwater pollution remediation technology and bioengineering technology, and specifically, relates to a Pseudomonas genu, a bacterial agent containing the Pseudomonas genu, the use of the Pseudomonas genu and the bacterial agent in degrading polycyclic aromatic hydrocarbons and remediating polycyclic aromatic hydrocarbon-contaminated soil or polycyclic aromatic hydrocarbon-contaminated groundwater, and a method for degrading polycyclic aromatic hydrocarbons. Background Art

[0002] PAHs are organic pollutants widely present in the environment. They are carcinogenic, teratogenic and mutagenic. They are easily accumulated in the environment and have long-term impacts on the ecological environment and human health. At present, the commonly used PAH pollution remediation methods are mainly thermal desorption, chemical oxidation and biodegradation. Compared with physical and chemical remediation, bioremediation has the advantages of low cost, simple operation and low secondary pollution. It is a green and sustainable environmental governance technology.

[0003] CN113755338B discloses a polycyclic aromatic hydrocarbon degrading bacterium Pyronema domesticum LJD-1, its bacterial agent and application. The strain was domesticated and isolated from a soil sample of a petroleum contaminated site and can use phenanthrene as a carbon source. After culturing for 7 days in an inorganic salt culture solution with an initial phenanthrene concentration of 50 mg / L, the phenanthrene degradation rate can reach 79.1%; when it is made into a bacterial agent, the phenanthrene degradation rate is 61.0%.

[0004] CN104004684B discloses a polycyclic aromatic hydrocarbons-degrading bacterium Bacillus sp.s8-t8-L9 and its application. The strain can use polycyclic aromatic hydrocarbons such as naphthalene, phenanthrene, pyrene and diesel as the only energy and carbon source for growth and reproduction, and can degrade naphthalene, phenanthrene, pyrene and diesel in an inorganic salt culture medium by 58.46%, 52.14%, 23.14% and 47.54% respectively in 20 days.

[0005] CN106434470B discloses a polycyclic aromatic hydrocarbon degrading bacterium Rhodococcus sp.YE-WL-2 and its application. The strain can grow in a solution with a fluoranthene concentration of up to 300 mg / L. When the initial fluoranthene concentration is 50 mg / L, the strain can degrade more than 40% of the fluoranthene in the culture medium within 7 days, the degradation rate of fluoranthene after 14 days of cultivation reaches more than 50%, and the degradation rate of fluoranthene after 21 days of cultivation reaches more than 60%.

[0006] In the prior art, the degradation rate of PAHs by strains is mostly less than 80%, and the low remediation efficiency has become a bottleneck in the application of microbial remediation technology. Therefore, the development of efficient PAH-degrading bacteria is of great significance for promoting the application of microbial remediation. Summary of the invention

[0007] The purpose of the present invention is to overcome the problems in the prior art that the degradation rate of PAHs by PAH-degrading bacteria is low and the efficiency of remediation of PAH-contaminated soil or groundwater is low, and to provide a Pseudomonas genu, a bacterial agent, a method for degrading PAHs and an application thereof. The Pseudomonas genu or bacterial agent of the present invention is used to degrade PAHs with high degradation efficiency.

[0008] To achieve the above object, the present invention provides a Pseudomonas geniculata in a first aspect, wherein the deposit number of the Pseudomonas geniculata is CGMCC No.22735.

[0009] A second aspect of the present invention provides a bacterial agent, which contains the Pseudomonas genu as described above.

[0010] A third aspect of the present invention provides use of the Pseudomonas geniculate or the bacterial agent described above in degrading polycyclic aromatic hydrocarbons.

[0011] A fourth aspect of the present invention provides the use of the Pseudomonas geniculate or the bacterial agent described above in remediating soil contaminated by polycyclic aromatic hydrocarbons or groundwater contaminated by polycyclic aromatic hydrocarbons.

[0012] A fifth aspect of the present invention provides a method for degrading polycyclic aromatic hydrocarbons, which comprises: contacting the Pseudomonas geniculate or the bacterial agent described above with pollutants containing polycyclic aromatic hydrocarbons.

[0013] The Pseudomonas geniculate or bacterial agent described in the present invention has a high degradation rate of polycyclic aromatic hydrocarbons. Specifically, the Pseudomonas geniculate in an inorganic salt liquid culture medium with a phenanthrene content of 100 mg / L, a temperature of 30.0±2.0°C, and a pH of 7.0±0.2, the phenanthrene degradation rate reaches more than 80% in 3 days, and the phenanthrene degradation rate reaches more than 95% in 5 days. In contaminated soil with a phenanthrene content of 100 mg / kg, the phenanthrene degradation rate reaches more than 90% in 14 days under the conditions of a temperature of 30.0±2.0°C and a pH of 7.0±0.2.

[0014] The Pseudomonas geniculate or bacterial agent of the present invention has a highly efficient ability to degrade polycyclic aromatic hydrocarbons, can be used for the restoration of soil or groundwater contaminated by polycyclic aromatic hydrocarbons, and has a good application prospect in the green and sustainable restoration of polycyclic aromatic hydrocarbon contaminated sites.

[0015] Other features and advantages of the present invention will be described in detail in the following detailed description. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Exemplary embodiments of the present invention will be described in more detail with reference to the accompanying drawings.

[0017] Figure 1The colony morphology of the Pseudomonas genu NAP-1 of the present invention on an agar solid medium plate is shown.

[0018] Biological Deposit Description

[0019] The Pseudomonas geniculata NAP-1 of the present invention was deposited on June 21, 2021 at the General Microbiology Center of China Culture Collection Administration (address: No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, Postal Code 100101) (the abbreviation of the depository unit is CGMCC), with the deposit number CGMCC No. 22735. DETAILED DESCRIPTION

[0020] The specific embodiments of the present invention are described in detail below. It should be understood that the specific embodiments described herein are only used to illustrate and explain the present invention, and are not used to limit the present invention.

[0021] The first aspect of the present invention provides a Pseudomonas geniculata, wherein the deposit number of the Pseudomonas geniculata is CGMCC No.22735.

[0022] The Pseudomonas geniculate of the present invention is screened from soil contaminated by polycyclic aromatic hydrocarbons in Tianjin.

[0023] The screening method includes: collecting contaminated soil, adding it to sterilized deionized water to prepare an inoculum; inoculating the inoculum into an inorganic salt culture medium with phenanthrene as the sole carbon source, and subculturing 3-5 times; diluting the above culture solution in a concentration gradient, and continuously streaking and culturing until a single colony is isolated; using LB culture medium to expand the above single colony, centrifuging to obtain a bacterial cell, i.e., the strain to be identified. The obtained bacterial cell is resuspended with sterilized physiological saline to obtain a bacterial suspension, and the bacterial suspension is mixed with 70% glycerol at a volume ratio of 1:1, and stored in a -80°C refrigerator.

[0024] The strain to be identified has a regular colony morphology on the agar solid culture medium plate, with yellow bead-like protrusions, a large single colony, a smooth and moist surface, a glossy feel, and is opaque. Figure 1 The strain was identified as Pseudomonas genus by 16S rDNA sequencing and named NAP-1.

[0025] Pseudomonas galactiformis NAP-1 is deposited in the General Microbiology Center of China Microorganism Culture Collection Administration (address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, Postal Code 100101) (the abbreviation of the depository is CGMCC), with the deposit number CGMCC No. 22735 and the deposit date of June 21, 2021.

[0026] A second aspect of the present invention provides a bacterial agent, which contains the Pseudomonas genu as described above.

[0027] In some embodiments of the present invention, the bacterial agent is a liquid bacterial agent and / or a solid bacterial agent. In the bacterial agent of the present invention, the concentration of the Pseudomonas geniculate is not particularly limited and can be selected according to specific circumstances. In some embodiments of the present invention, the number of viable bacteria in the liquid bacterial agent is 10 9 cfu / mL or more. In some embodiments of the present invention, the number of viable bacteria in the solid bacterial agent is 10 9 cfu / g or above.

[0028] In some embodiments of the present invention, the bacterial agent may further contain an auxiliary material. The auxiliary material may be an auxiliary material commonly used in the art that does not affect the activity of Pseudomonas genu.

[0029] In addition, the bacterial agent provided by the present invention can be prepared into different dosage forms according to different intended uses, such as powder, paste, liquid, etc.

[0030] The preparation method of the bacterial agent can be a conventional preparation method in the art. Preferably, the preparation method comprises: activating the Pseudomonas genu as described above and culturing it in a culture medium to obtain a bacterial liquid stock solution; preparing the bacterial bodies obtained by centrifuging the bacterial liquid stock solution into a bacterial suspension with sterile water and making the number of live bacteria in the bacterial suspension reach 10 9 cfu / mL or more to obtain a bacterial agent. Wherein, the culture medium can be a culture medium conventionally used in the art for culturing Pseudomonas genu, for example, it can be LB culture medium. The inventors have found that the Pseudomonas genu described in the present invention can be cultured in LB culture medium. Preferably, for the consideration of production cost, ease of operation, etc., the culture medium is LB culture medium.

[0031] In the present invention, the culture conditions may be conventional conditions for culturing Pseudomonas geniculate known in the art, and the present invention is not limited thereto.

[0032] A third aspect of the present invention provides use of the Pseudomonas geniculate or the bacterial agent described above in degrading polycyclic aromatic hydrocarbons.

[0033] A fourth aspect of the present invention provides the use of the Pseudomonas geniculate or the bacterial agent described above in remediating soil contaminated by polycyclic aromatic hydrocarbons or groundwater contaminated by polycyclic aromatic hydrocarbons.

[0034] A fifth aspect of the present invention provides a method for degrading polycyclic aromatic hydrocarbons, which comprises: contacting the Pseudomonas geniculate or the bacterial agent described above with pollutants containing polycyclic aromatic hydrocarbons.

[0035] In the present invention, the polycyclic aromatic hydrocarbons refer to at least one of the aromatic hydrocarbons containing at least two benzene rings, including both condensed-ring polycyclic aromatic hydrocarbons and non-condensed-ring polycyclic aromatic hydrocarbons.

[0036] Specifically, the polycyclic aromatic hydrocarbons include but are not limited to naphthalene, anthracene, phenanthrene, pyrene, At least one of benzopyrene, benzanthracene, benzofluoranthene, indenopyrene, biphenyl (i.e., biphenyl), biphenyl and polyphenyl aliphatic hydrocarbons. The benzopyrene includes benzo[a]pyrene and benzo[e]pyrene. The benzanthracene includes benzo[a]anthracene and dibenzo[a,h]anthracene. The indenopyrene includes indeno[1,2,3-cd]pyrene. The biphenyl may be terphenyl, etc., and the polyphenyl aliphatic hydrocarbon may be diphenylmethane, diphenylethane, etc.

[0037] In some embodiments of the present invention, the pollutant containing polycyclic aromatic hydrocarbons is soil containing polycyclic aromatic hydrocarbons or groundwater containing polycyclic aromatic hydrocarbons.

[0038] In some embodiments of the present invention, when the polycyclic aromatic hydrocarbon-containing pollutant is soil containing polycyclic aromatic hydrocarbons, the content of the polycyclic aromatic hydrocarbons is 2-1000 mg / kg soil, calculated as polycyclic aromatic hydrocarbons, for example, it can be any one of 2 mg / kg soil, 5 mg / kg soil, 10 mg / kg soil, 50 mg / kg soil, 100 mg / kg soil, 200 mg / kg soil, 300 mg / kg soil, 500 mg / kg soil, 700 mg / kg soil, 1000 mg / kg soil, or a value within the range of any two of the above values. Wherein, the weight of the soil is calculated on a dry basis.

[0039] In the present invention, the amount of the Pseudomonas geniculate or the bacterial agent used can be selected within a wide range. In some embodiments of the present invention, when the polycyclic aromatic hydrocarbon-containing pollutant is soil containing polycyclic aromatic hydrocarbons, the amount of the Pseudomonas geniculate or the bacterial agent used is such that the number of viable bacteria in the soil containing polycyclic aromatic hydrocarbons is 10 6 -10 8 cfu / g. Wherein, the soil weight is calculated on a dry basis.

[0040] In some embodiments of the present invention, the contact conditions include: a temperature of 20-40°C and a pH of 5-10. The temperature may be any one of 20°C, 23°C, 26°C, 28°C, 30°C, 33°C, 35°C, 38°C, and 40°C, or a value within the range of any two of the above values. The pH may be any one of 5.0, 5.3, 5.5, 5.7, 6.0, 6.3, 6.6, 6.8, 7.0, 7.2, 7.5, 7.8, 8.0, 8.3, 8.6, 8.8, 9.0, 9.2, 9.4, 9.8, and 10.0, or a value within the range of any two of the above values. Preferably, in order to further improve the degradation rate of polycyclic aromatic hydrocarbons, the contact conditions are: a temperature of 25-38°C and a pH of 5.8-9.2. The inventors have found that when the contact conditions are within the above preferred range, the degradation rate of polycyclic aromatic hydrocarbons can be further improved.

[0041] In some embodiments of the present invention, when the pollutant containing PAHs is groundwater containing PAHs, the concentration of PAHs, calculated as PAHs, is 2-1000 mg / L groundwater, for example, it can be 2 mg / L groundwater, 5 mg / L groundwater, 10 mg / L groundwater, 50 mg / L groundwater, 100 mg / L groundwater, 200 mg / L groundwater, 300 mg / L groundwater, 500 mg / L groundwater, 700 mg / L groundwater, 1000 mg / L groundwater, any one of the values ​​or a value within the range consisting of any two of the above values.

[0042] In some embodiments of the present invention, when the polycyclic aromatic hydrocarbon-containing pollutant is groundwater containing polycyclic aromatic hydrocarbons, the amount of the Pseudomonas geniculate or the bacterial agent is such that the number of viable bacteria in the groundwater containing polycyclic aromatic hydrocarbons is 10 6 -10 8 cfu / mL.

[0043] The present invention will be described in detail below by way of examples. In the following examples, unless otherwise specified, all reagents and raw materials used can be obtained commercially, and all methods used are conventional methods in the art.

[0044] In the following embodiments:

[0045] LB medium: yeast extract 5 g / L, peptone 10 g / L, NaCl 10 g / L, deionized water 1 L.

[0046] Inorganic salt medium: NH 4 Cl 0.67g / L, NaNO 3 1.06g / L, MgSO 4 7H 2 O 0.1g / L, CaCl2 0.1g / L, K 2 HPO 4 ·3H 2 O 1.5g / L, KH 2 PO 4 0.5g / L, FeCl 3 6H 2 O 0.1g / L, deionized water 1L.

[0047] Phenanthrene was purchased from Beijing Inokai Technology Co., Ltd.

[0048] Naphthalene was purchased from Beijing Inokai Technology Co., Ltd.

[0049] Purchased from Beijing Inokai Technology Co., Ltd.

[0050] Benz[a]anthracene was purchased from Beijing Inokai Technology Co., Ltd.

[0051] Benzo[b]fluoranthene was purchased from Beijing Inokai Technology Co., Ltd.

[0052] Benzo[a]pyrene was purchased from Beijing Inokai Technology Co., Ltd.

[0053] Dibenzo[a,h]anthracene was purchased from Beijing Inokai Technology Co., Ltd.

[0054] Indeno[1,2,3-cd]pyrene was purchased from Beijing Inokai Technology Co., Ltd.

[0055] The strain Pyronema domesticum LJD-1 was purchased from Guangdong Microbiological Culture Collection Center.

[0056] The strain Bacillus sp.s8-t8-L9 was purchased from the General Microbiology Center of China Culture Collection Administration.

[0057] Pseudomonas geniculata NAP-1 was deposited on June 21, 2021 at the General Microbiology Center of China Culture Collection Administration (address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, Postal Code 100101) (the abbreviation of the depository is CGMCC), with the deposit number CGMCC No.22735.

[0058] The detection method of the PAH content in contaminated soil is carried out in accordance with "Gas chromatography-mass spectrometry for the determination of semi-volatile organic matter in soil and sediment" HJ 834-2017. Among them, a dichloromethane-acetone mixed solution is used to extract PAHs under pressure. When the constant volume is 1.0mL and the injection volume is 1.0μL, the detection limit of the method for determining PAHs is 0.06-0.3mg / kg, and the lower limit of determination is 0.24-1.20mg / kg.

[0059] The detection method of the concentration of polycyclic aromatic hydrocarbons in the inorganic salt culture medium is based on the "Determination of polycyclic aromatic hydrocarbons in water by liquid-liquid extraction and solid-phase extraction high performance liquid chromatography" HJ 478-2009.

[0060] PAH degradation rate in inorganic salt medium (%) = (initial PAH concentration - residual PAH concentration) / initial PAH concentration. The initial PAH concentration refers to the PAH concentration in the inorganic salt medium without inoculation of bacterial suspension.

[0061] PAH degradation rate in contaminated soil (%) = (initial PAH content - residual PAH content) / initial PAH content. The initial PAH content refers to the PAH content in contaminated soil that has not been inoculated with bacterial suspension.

[0062] Example 1

[0063] This example is used to illustrate the degradation effect of Pseudomonas genuformis NAP-1 on phenanthrene.

[0064] Preparation of simulated PAH-contaminated groundwater: prepare a phenanthrene solution with n-hexane, filter it with a sterile filter membrane, add it to a sterile conical flask, and add it to a sterile inorganic salt culture medium after the n-hexane is completely evaporated so that the phenanthrene concentration is 100 mg / L, to obtain an inorganic salt culture medium with a phenanthrene concentration of 100 mg / L.

[0065] The preserved Pseudomonas genu NAP-1 cells were activated and cultured in LB medium to obtain a stock solution. The cells obtained after centrifugation of the stock solution were prepared into a bacterial suspension with sterile water, and the live bacterial content was about 10 9 cfu / mL.

[0066] The bacterial suspension was inoculated at a volume of 1 vol.% (10 9cfu) were inoculated in the above-mentioned inorganic salt medium with a phenanthrene concentration of 100 mg / L. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas genu NAP-1) were set up, the residual phenanthrene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas genu NAP-1 on phenanthrene in the inorganic salt medium and the degradation effect of 5 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The phenanthrene degradation effects of Pseudomonas genu NAP-1 under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 3 days and 5 days are shown in Table 1.

[0067] Preparation of simulated PAH-contaminated soil: Take soil that is not contaminated by PAHs, sieve it to remove small stones, grass roots and other impurities in the soil after pretreatment and let it air-dry naturally, weigh a certain amount of phenanthrene, dissolve it in n-hexane solution, and spray it evenly on the above soil sample so that the phenanthrene content is 100 mg / kg (wherein the weight of the soil is based on dry basis), and wait until the n-hexane is completely volatilized to obtain contaminated soil with a phenanthrene content of 100 mg / kg.

[0068] The bacterial suspension was inoculated at a volume of 1 (v / w)% (10 9 cfu) were inoculated in the above-mentioned contaminated soil with a phenanthrene content of 100 mg / kg. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas genu NAP-1) were set up, the residual phenanthrene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas genu NAP-1 on phenanthrene in contaminated soil and the degradation effect of 20 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The phenanthrene degradation effects of Pseudomonas genu NAP-1 under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 14 days and 20 days are shown in Table 1.

[0069] Example 2

[0070] This example is used to illustrate the degradation effect of Pseudomonas genuformis NAP-1 on naphthalene.

[0071] Preparation of simulated PAH-contaminated groundwater: prepare naphthalene solution with n-hexane, filter with a sterile filter membrane, add to a sterile conical flask, and add to a sterile inorganic salt culture medium after the n-hexane is completely evaporated so that the naphthalene concentration is 100 mg / L, to obtain an inorganic salt culture medium with a naphthalene concentration of 100 mg / L.

[0072] The preserved Pseudomonas genu NAP-1 cells were activated and cultured in LB medium to obtain a stock solution. The cells obtained after centrifugation of the stock solution were prepared into a bacterial suspension with sterile water, and the live bacterial content was about 10 9 cfu / mL.

[0073] The bacterial suspension was inoculated at a volume of 1 vol.% (10 9 cfu) were inoculated in the above-mentioned inorganic salt medium with a naphthalene concentration of 100 mg / L. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas genu NAP-1) were set up, the residual naphthalene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas genu NAP-1 on naphthalene in the inorganic salt medium and the degradation effect of 5 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The naphthalene degradation effects of Pseudomonas genu NAP-1 under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 3 days and 5 days are shown in Table 1.

[0074] Preparation of simulated PAH-contaminated soil: Take soil that is not contaminated by PAHs, sieve it to remove small stones, grass roots and other impurities in the soil after pretreatment and let it air-dry naturally, weigh a certain amount of naphthalene, dissolve it in n-hexane solution, and spray it evenly on the above soil sample so that the naphthalene content is 100 mg / kg (wherein the weight of the soil is based on dry basis), and wait until the n-hexane is completely volatilized to obtain contaminated soil with a naphthalene content of 100 mg / kg.

[0075] The bacterial suspension was inoculated at a volume of 1 (v / w)% (10 9 cfu) were inoculated in the above-mentioned contaminated soil with a naphthalene content of 100 mg / kg. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas genu NAP-1) were set up, the residual naphthalene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas genu NAP-1 on naphthalene in contaminated soil and the degradation effect of 20 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The naphthalene degradation effects of Pseudomonas genu NAP-1 under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 14 days and 20 days are shown in Table 1.

[0076] Example 3

[0077] This example is used to illustrate the effect of Pseudomonas genu NAP-1 on degradation effect.

[0078] Preparation of simulated PAH-contaminated groundwater: Preparation with n-hexane The solution was filtered with a sterile filter membrane and added to a sterile conical flask. After the n-hexane was completely volatilized, it was added to the sterile inorganic salt medium to make The concentration is 100 mg / L, and the Inorganic salt culture medium with a concentration of 100 mg / L.

[0079] The preserved Pseudomonas genu NAP-1 cells were activated and cultured in LB medium to obtain a stock solution. The cells obtained after centrifugation of the stock solution were prepared into a bacterial suspension with sterile water, and the live bacterial content was about 10 9 cfu / mL.

[0080] The bacterial suspension was inoculated at a volume of 1 vol.% (10 9 cfu) were inoculated in the above In an inorganic salt medium with a concentration of 100 mg / L, with temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas geniculate NAP-1) were set up, and the remaining content, calculate the degradation rate of PAHs, and determine the degradation of Pseudomonas genu NAP-1 to inorganic salt medium The optimal degradation conditions for Pseudomonas genu NAP-1 and the degradation effect under the optimal degradation conditions for 5 days. The results showed that the optimal degradation conditions for Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effects are shown in Table 1.

[0081] Preparation of simulated PAH-contaminated soil: Take soil that is not contaminated by PAHs, sieve it to remove small stones, grass roots and other impurities in the soil after pretreatment and dry it naturally, and weigh a certain amount of Dissolve in n-hexane solution and spray evenly on the above soil samples to make The content is 100 mg / kg (wherein the weight of the soil is calculated on a dry basis), and after the n-hexane is completely volatilized, Contaminated soil with a content of 100 mg / kg.

[0082] The bacterial suspension was inoculated at a volume of 1 (v / w)% (10 9 cfu) were inoculated in the above In the contaminated soil with a content of 100 mg / kg, the experimental group and the control group (the control group was not inoculated with Pseudomonas geniculate NAP-1) were set up with temperature and pH as the influencing factors, and the residual content, calculate the degradation rate of PAHs, and determine the degradation of Pseudomonas galactiformis NAP-1 to the contaminated soil The optimal degradation conditions for Pseudomonas genu NAP-1 and the degradation effect under the optimal degradation conditions for 20 days. The results showed that the optimal degradation conditions for Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effects are shown in Table 1.

[0083] Example 4

[0084] This example is used to illustrate the degradation effect of Pseudomonas genu NAP-1 on benz[a]anthracene.

[0085] Preparation of simulated PAH-contaminated groundwater: prepare a benzo[a]anthracene solution with n-hexane, filter with a sterile filter membrane, add it to a sterile conical flask, and add it to a sterile inorganic salt culture medium after the n-hexane is completely evaporated so that the benzo[a]anthracene concentration is 100 mg / L, to obtain an inorganic salt culture medium with a benzo[a]anthracene concentration of 100 mg / L.

[0086] The preserved Pseudomonas genu NAP-1 cells were activated and cultured in LB medium to obtain a stock solution. The cells obtained after centrifugation of the stock solution were prepared into a bacterial suspension with sterile water, and the live bacterial content was about 10 9 cfu / mL.

[0087] The bacterial suspension was inoculated at a volume of 1 vol.% (10 9 cfu) were inoculated in the above-mentioned inorganic salt medium with a benz[a]anthracene concentration of 100 mg / L. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas genu NAP-1) were set up, the remaining benz[a]anthracene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas genu NAP-1 on benz[a]anthracene in the inorganic salt medium and the degradation effect under the optimal degradation conditions for 5 days were determined. The results showed that the optimal degradation conditions for Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effect of Pseudomonas genu NAP-1 on benz[a]anthracene under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 5 days is shown in Table 1.

[0088] Preparation of simulated PAH-contaminated soil: Take soil that is not contaminated by PAHs, sieve it to remove small stones, grass roots and other impurities in the soil after pretreatment and let it air-dry naturally, weigh a certain amount of benz[a]anthracene, dissolve it in n-hexane solution, and spray it evenly on the above soil sample so that the benz[a]anthracene content is 100 mg / kg (wherein the weight of the soil is based on dry basis), and wait until the n-hexane is completely volatilized to obtain contaminated soil with a benz[a]anthracene content of 100 mg / kg.

[0089] The bacterial suspension was inoculated at a volume of 1 (v / w)% (10 9 cfu) were inoculated in the above-mentioned contaminated soil with a benz[a]anthracene content of 100 mg / kg. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas genu NAP-1) were set up. The remaining benz[a]anthracene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas genu NAP-1 on benz[a]anthracene in contaminated soil and the degradation effect of 20 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effect of Pseudomonas genu NAP-1 on benz[a]anthracene under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 20 days is shown in Table 1.

[0090] Example 5

[0091] This example is used to illustrate the degradation effect of Pseudomonas genu NAP-1 on benzo[b]fluoranthene.

[0092] Preparation of simulated PAH-contaminated groundwater: Prepare benzo[b]fluoranthene solution with n-hexane, filter with a sterile filter membrane, add to a sterile conical flask, and add to a sterile inorganic salt culture medium after the n-hexane is completely evaporated so that the concentration of benzo[b]fluoranthene is 70 mg / L, to obtain an inorganic salt culture medium with a benzo[b]fluoranthene concentration of 70 mg / L.

[0093] The preserved Pseudomonas genu NAP-1 cells were activated and cultured in LB medium to obtain a stock solution. The cells obtained after centrifugation of the stock solution were prepared into a bacterial suspension with sterile water, and the live bacterial content was about 10 9 cfu / mL.

[0094] The bacterial suspension was inoculated at a volume of 1 vol.% (10 9 cfu) were inoculated in the above-mentioned inorganic salt medium with a benzo[b]fluoranthene concentration of 70 mg / L. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas genu NAP-1) were set up, the remaining benzo[b]fluoranthene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas genu NAP-1 on benzo[b]fluoranthene in the inorganic salt medium and the degradation effect under the optimal degradation conditions for 5 days were determined. The results showed that the optimal degradation conditions for Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effect of Pseudomonas genu NAP-1 on benzo[b]fluoranthene under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 5 days is shown in Table 1.

[0095] Preparation of simulated PAH-contaminated soil: Take soil that is not contaminated by PAHs, sieve it to remove small stones, grass roots and other impurities in the soil after pretreatment and let it air-dry naturally, weigh a certain amount of benzo[b]fluoranthene, dissolve it in n-hexane solution, and spray it evenly on the above soil sample so that the benzo[b]fluoranthene content is 70 mg / kg (wherein the weight of the soil is based on dry basis), and wait until the n-hexane is completely volatilized to obtain contaminated soil with a benzo[b]fluoranthene content of 70 mg / kg.

[0096] The bacterial suspension was inoculated at a volume of 1 (v / w)% (10 9 cfu) were inoculated in the above-mentioned contaminated soil with a benzo[b]fluoranthene content of 70 mg / kg. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas genu NAP-1) were set up. The remaining benzo[b]fluoranthene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas genu NAP-1 on benzo[b]fluoranthene in contaminated soil and the degradation effect of 20 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effect of Pseudomonas genu NAP-1 on benzo[b]fluoranthene for 20 days under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) is shown in Table 1.

[0097] Example 6

[0098] This example is used to illustrate the degradation effect of Pseudomonas genu NAP-1 on benzo[a]pyrene.

[0099] Preparation of simulated PAH-contaminated groundwater: prepare benzo[a]pyrene solution with n-hexane, filter with a sterile filter membrane, add to a sterile conical flask, and add to a sterile inorganic salt culture medium after the n-hexane is completely evaporated so that the benzo[a]pyrene concentration is 30 mg / L, to obtain an inorganic salt culture medium with a benzo[a]pyrene concentration of 30 mg / L.

[0100] The preserved Pseudomonas genu NAP-1 cells were activated and cultured in LB medium to obtain a stock solution. The cells obtained after centrifugation of the stock solution were prepared into a bacterial suspension with sterile water, and the live bacterial content was about 10 9 cfu / mL.

[0101] The bacterial suspension was inoculated at a volume of 1 vol.% (10 9cfu) were inoculated in the above-mentioned inorganic salt medium with a benzo[a]pyrene concentration of 30 mg / L. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas genu NAP-1) were set up, the residual benzo[a]pyrene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas genu NAP-1 on benzo[a]pyrene in the inorganic salt medium and the degradation effect of 5 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effect of Pseudomonas genu NAP-1 on benzo[a]pyrene under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 5 days is shown in Table 1.

[0102] Preparation of simulated PAH-contaminated soil: Take soil that is not contaminated by PAHs, sieve it to remove small stones, grass roots and other impurities in the soil after pretreatment and let it air-dry naturally, weigh a certain amount of benzo[a]pyrene, dissolve it in n-hexane solution, and spray it evenly into the above soil sample so that the benzo[a]pyrene content is 30 mg / kg (wherein the weight of the soil is based on dry basis), and wait until the n-hexane is completely volatilized to obtain contaminated soil with a benzo[a]pyrene content of 30 mg / kg.

[0103] The bacterial suspension was inoculated at a volume of 1 (v / w)% (10 9 cfu) were inoculated in the above-mentioned contaminated soil with a benzo[a]pyrene content of 30 mg / kg. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas geniculate NAP-1) were set up. The remaining benzo[a]pyrene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas geniculate NAP-1 on benzo[a]pyrene in contaminated soil and the degradation effect of 20 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pseudomonas geniculate NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effect of Pseudomonas geniculate NAP-1 on benzo[a]pyrene under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 20 days is shown in Table 1.

[0104] Example 7

[0105] This example is used to illustrate the degradation effect of Pseudomonas genu NAP-1 on dibenzo[a,h]anthracene.

[0106] Preparation of simulated PAH-contaminated groundwater: prepare dibenzo[a,h]anthracene solution with n-hexane, filter with a sterile filter membrane, add to a sterile conical flask, and add to a sterile inorganic salt culture medium after the n-hexane is completely evaporated so that the dibenzo[a,h]anthracene concentration is 30 mg / L, to obtain an inorganic salt culture medium with a dibenzo[a,h]anthracene concentration of 30 mg / L.

[0107] The preserved Pseudomonas genu NAP-1 cells were activated and cultured in LB medium to obtain a stock solution. The cells obtained after centrifugation of the stock solution were prepared into a bacterial suspension with sterile water, and the live bacterial content was about 10 9 cfu / mL.

[0108] The bacterial suspension was inoculated at a volume of 1 vol.% (10 9 cfu) were inoculated in the above-mentioned inorganic salt medium with a dibenzo[a,h]anthracene concentration of 30 mg / L. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas genu NAP-1) were set up. The remaining dibenzo[a,h]anthracene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas genu NAP-1 on dibenzo[a,h]anthracene in inorganic salt medium and the degradation effect under the optimal degradation conditions for 5 days were determined. The results showed that the optimal degradation conditions for Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effect of Pseudomonas genu NAP-1 on dibenzo[a,h]anthracene under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 5 days is shown in Table 1.

[0109] Preparation of simulated PAH-contaminated soil: Take soil that is not contaminated by PAHs, sieve it to remove small stones, grass roots and other impurities in the soil after pretreatment and let it air-dry naturally, weigh a certain amount of dibenzo[a,h]anthracene, dissolve it in n-hexane solution, and spray it evenly into the above soil sample so that the dibenzo[a,h]anthracene content is 30 mg / kg (wherein the weight of the soil is based on dry basis), and wait until the n-hexane is completely volatilized to obtain contaminated soil with a dibenzo[a,h]anthracene content of 30 mg / kg.

[0110] The bacterial suspension was inoculated at a volume of 1 (v / w)% (10 9 cfu) were inoculated in the above-mentioned contaminated soil with a dibenzo[a,h]anthracene content of 30 mg / kg. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas genu NAP-1) were set up. The remaining dibenzo[a,h]anthracene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas genu NAP-1 on dibenzo[a,h]anthracene in contaminated soil and the degradation effect of 20 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pseudomonas genu NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effect of Pseudomonas genu NAP-1 on dibenzo[a,h]anthracene under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 20 days is shown in Table 1.

[0111] Example 8

[0112] This example is used to illustrate the degradation effect of Pseudomonas geniculate NAP-1 on indeno[1,2,3-cd]pyrene.

[0113] Preparation of simulated PAH-contaminated groundwater: prepare an indeno[1,2,3-cd]pyrene solution with n-hexane, filter with a sterile filter membrane, add the solution to a sterile conical flask, and add the solution to a sterile inorganic salt culture medium after the n-hexane is completely evaporated so that the indeno[1,2,3-cd]pyrene concentration is 30 mg / L, thereby obtaining an inorganic salt culture medium with an indeno[1,2,3-cd]pyrene concentration of 30 mg / L.

[0114] The preserved Pseudomonas genu NAP-1 cells were activated and cultured in LB medium to obtain a stock solution. The cells obtained after centrifugation of the stock solution were prepared into a bacterial suspension with sterile water, and the live bacterial content was about 10 9 cfu / mL.

[0115] The bacterial suspension was inoculated at a volume of 1 vol.% (10 9 cfu) were inoculated in the above-mentioned inorganic salt medium with an indeno[1,2,3-cd]pyrene concentration of 30 mg / L. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas geniculate NAP-1) were set up. The remaining indeno[1,2,3-cd]pyrene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas geniculate NAP-1 for indeno[1,2,3-cd]pyrene in inorganic salt medium and the degradation effect of 5 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pseudomonas geniculate NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effect of Pseudomonas geniculate NAP-1 on indeno[1,2,3-cd]pyrene for 5 days under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) is shown in Table 1.

[0116] Preparation of simulated PAH-contaminated soil: Take soil that is not contaminated by PAHs, sieve it to remove small stones, grass roots and other impurities in the soil after pretreatment and let it air-dry naturally, weigh a certain amount of indeno[1,2,3-cd]pyrene, dissolve it in n-hexane solution, and spray it evenly into the above soil sample so that the indeno[1,2,3-cd]pyrene content is 30 mg / kg (wherein the weight of the soil is based on dry basis), and wait until the n-hexane is completely volatilized to obtain contaminated soil with an indeno[1,2,3-cd]pyrene content of 30 mg / kg.

[0117] The bacterial suspension was inoculated at a volume of 1 (v / w)% (10 9cfu) were inoculated in the above-mentioned contaminated soil with an indeno[1,2,3-cd]pyrene content of 30 mg / kg. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pseudomonas geniculate NAP-1) were set up. The remaining indeno[1,2,3-cd]pyrene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pseudomonas geniculate NAP-1 on indeno[1,2,3-cd]pyrene in contaminated soil and the degradation effect of 20 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pseudomonas geniculate NAP-1 were 30.0±2.0℃ and pH 7.0±0.2. The degradation effect of Pseudomonas geniculate NAP-1 on indeno[1,2,3-cd]pyrene under the optimal conditions (temperature 30.0±2.0℃, pH 7.0±0.2) for 20 days is shown in Table 1.

[0118] Comparative Example 1

[0119] This comparative example is used to compare the degradation effect of Pyronema domesticum LJD-1 (GDMCC No. 61797) on phenanthrene.

[0120] Preparation of simulated PAH-contaminated groundwater: prepare a phenanthrene solution with n-hexane, filter it with a sterile filter membrane, add it to a sterile conical flask, and add it to a sterile inorganic salt culture medium after the n-hexane is completely evaporated so that the phenanthrene concentration is 100 mg / L, to obtain an inorganic salt culture medium with a phenanthrene concentration of 100 mg / L.

[0121] After the Pyronema domesticum LJD-1 bacteria were activated, they were cultured in LB medium to obtain a stock solution. After the stock solution was centrifuged, the bacteria obtained were prepared into a bacterial suspension with sterile water, and the live bacterial content was about 10 9 cfu / mL.

[0122] The bacterial suspension was inoculated at a volume of 1 vol.% (10 9 cfu) were inoculated in the above-mentioned inorganic salt medium with a phenanthrene concentration of 100 mg / L. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pyronemadomesticum LJD-1) were set up, the residual phenanthrene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pyronemadomesticum LJD-1 on phenanthrene in the inorganic salt medium and the degradation effect of 5 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pyronema domesticum LJD-1 were 28.0±2.0℃ and pH 6.0±0.2. The phenanthrene degradation effects of Pyronema domesticum LJD-1 under the optimal conditions (temperature 28.0±2.0℃, pH 6.0±0.2) for 3 days and 5 days are shown in Table 1.

[0123] Preparation of simulated PAH-contaminated soil: Take soil that is not contaminated by PAHs, sieve it to remove small stones, grass roots and other impurities in the soil after pretreatment and let it air-dry naturally, weigh a certain amount of phenanthrene, dissolve it in n-hexane solution, and spray it evenly on the above soil sample so that the phenanthrene content is 100 mg / kg (wherein the weight of the soil is based on dry basis), and wait until the n-hexane is completely volatilized to obtain contaminated soil with a phenanthrene content of 100 mg / kg.

[0124] The bacterial suspension was inoculated at a volume of 1 (v / w)% (10 9 cfu) were inoculated in the above-mentioned contaminated soil with a phenanthrene content of 100 mg / kg. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pyronema domesticum LJD-1) were set up. The residual phenanthrene content was regularly tested, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pyronema domesticum LJD-1 on phenanthrene in contaminated soil and the degradation effect of 20 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pyronema domesticum LJD-1 were 28.0±2.0℃ and pH 6.0±0.2. The phenanthrene degradation effects of Pyronema domesticum LJD-1 under the optimal conditions (temperature 28.0±2.0℃, pH 6.0±0.2) for 14 days and 20 days are shown in Table 1.

[0125] Comparative Example 2

[0126] This comparative example is used to compare the degradation effect of Pyronema domesticum LJD-1 (GDMCC No. 61797) on benzo[a]pyrene.

[0127] Preparation of simulated PAH-contaminated groundwater: prepare benzo[a]pyrene solution with n-hexane, filter with a sterile filter membrane, add to a sterile conical flask, and add to a sterile inorganic salt culture medium after the n-hexane is completely evaporated so that the benzo[a]pyrene concentration is 30 mg / L, to obtain an inorganic salt culture medium with a benzo[a]pyrene concentration of 30 mg / L.

[0128] After the Pyronema domesticum LJD-1 bacteria were activated, they were cultured in LB medium to obtain a stock solution. After the stock solution was centrifuged, the bacteria obtained were prepared into a bacterial suspension with sterile water, and the live bacterial content was about 10 9 cfu / mL.

[0129] The bacterial suspension was inoculated at a volume of 1 vol.% (10 9cfu) were inoculated in the above-mentioned inorganic salt medium with a benzo[a]pyrene concentration of 30 mg / L. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pyronemadomesticum LJD-1) were set up. The residual benzo[a]pyrene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pyronemadomesticum LJD-1 for benzo[a]pyrene in the inorganic salt medium and the degradation effect of 5 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pyronema domesticum LJD-1 were 28.0±2.0℃ and pH6.0±0.2. The degradation effect of Pyronema domesticum LJD-1 on benzo[a]pyrene under the optimal conditions (temperature 28.0±2.0℃, pH6.0±0.2) for 5 days is shown in Table 1.

[0130] Preparation of simulated PAH-contaminated soil: Take soil that is not contaminated by PAHs, sieve it to remove small stones, grass roots and other impurities in the soil after pretreatment and let it air-dry naturally, weigh a certain amount of benzo[a]pyrene, dissolve it in n-hexane solution, and spray it evenly into the above soil sample so that the benzo[a]pyrene content is 30 mg / kg (wherein the weight of the soil is based on dry basis), and wait until the n-hexane is completely volatilized to obtain contaminated soil with a benzo[a]pyrene content of 30 mg / kg.

[0131] The bacterial suspension was inoculated at a volume of 1 (v / w)% (10 9 cfu) were inoculated in the above-mentioned contaminated soil with a benzo[a]pyrene content of 30 mg / kg. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Pyronemadomesticum LJD-1) were set up. The residual benzo[a]pyrene content was regularly tested, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Pyronemadomesticum LJD-1 on benzo[a]pyrene in contaminated soil and the degradation effect of 20 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Pyronema domesticum LJD-1 were 28.0±2.0℃ and pH 6.0±0.2. The degradation effect of Pyronema domesticum LJD-1 on benzo[a]pyrene under the optimal conditions (temperature 28.0±2.0℃, pH 6.0±0.2) for 20 days is shown in Table 1.

[0132] Comparative Example 3

[0133] This comparative example is used to compare the degradation effect of Bacillus sp.s8-t8-L9 (CGMCC No.9032) on phenanthrene.

[0134] Preparation of simulated PAH-contaminated groundwater: prepare a phenanthrene solution with n-hexane, filter it with a sterile filter membrane, add it to a sterile conical flask, and add it to a sterile inorganic salt culture medium after the n-hexane is completely evaporated so that the phenanthrene concentration is 100 mg / L, to obtain an inorganic salt culture medium with a phenanthrene concentration of 100 mg / L.

[0135] After the Bacillus sp.s8-t8-L9 bacteria were activated, they were cultured in LB medium to obtain a stock solution; the bacteria obtained after centrifugation of the stock solution were prepared into a bacterial suspension with sterile water, and the live bacterial content was about 10 9 cfu / mL.

[0136] The bacterial suspension was inoculated at a volume of 1 vol.% (10 9 cfu) were inoculated in the above-mentioned inorganic salt medium with a phenanthrene concentration of 100 mg / L. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Bacillus sp.s8-t8-L9) were set up, the residual phenanthrene content was regularly detected, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Bacillus sp.s8-t8-L9 for phenanthrene in the inorganic salt medium and the degradation effect of 5 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Bacillus sp.s8-t8-L9 were 25.0±2.0℃ and pH 7.5±0.2. The phenanthrene degradation effects of Bacillus sp.s8-t8-L9 for 3 days and 5 days under the optimal conditions (temperature 25.0±2.0℃, pH 7.5±0.2) are shown in Table 1.

[0137] Preparation of simulated PAH-contaminated soil: Take soil that is not contaminated by PAHs, sieve it to remove small stones, grass roots and other impurities in the soil after pretreatment and let it air-dry naturally, weigh a certain amount of phenanthrene, dissolve it in n-hexane solution, and spray it evenly on the above soil sample so that the phenanthrene content is 100 mg / kg (wherein the weight of the soil is based on dry basis), and wait until the n-hexane is completely volatilized to obtain contaminated soil with a phenanthrene content of 100 mg / kg.

[0138] The bacterial suspension was inoculated at a volume of 1 (v / w)% (10 9cfu) were inoculated in the above-mentioned contaminated soil with a phenanthrene content of 100 mg / kg. With temperature and pH as influencing factors, an experimental group and a control group (the control group was not inoculated with Bacillus sp.s8-t8-L9) were set up, the residual phenanthrene content was regularly tested, the degradation rate of polycyclic aromatic hydrocarbons was calculated, and the optimal degradation conditions of Bacillus sp.s8-t8-L9 on phenanthrene in contaminated soil and the degradation effect of 20 days under the optimal degradation conditions were determined. The results showed that the optimal degradation conditions of Bacillus sp.s8-t8-L9 were 25.0±2.0℃ and pH 7.5±0.2. The phenanthrene degradation effects of Bacillus sp.s8-t8-L9 under the optimal conditions (temperature 25.0±2.0℃, pH 7.5±0.2) for 14 days and 20 days are shown in Table 1.

[0139] Table 1 PAH degradation effect of Pseudomonas galactiformis NAP-1

[0140]

[0141] It can be seen from the data in Table 1 that the Pseudomonas genu NAP-1 of the present invention has a good degradation effect on various polycyclic aromatic hydrocarbons in soil or groundwater. In an inorganic salt liquid culture medium with a phenanthrene content of 100 mg / L, at a temperature of 30.0±2.0°C and a pH of 7.0±0.2, the phenanthrene degradation rate reaches more than 95% after 5 days. In contaminated soil with a phenanthrene content of 100 mg / kg, at a temperature of 30.0±2.0°C and a pH of 7.0±0.2, the phenanthrene degradation rate reaches more than 95% after 20 days; the degradation rate is significantly better than that of the control strain.

[0142] The embodiments of the present invention have been described above, and the above description is exemplary, not exhaustive, and is not limited to the disclosed embodiments. Many modifications and changes will be apparent to those skilled in the art without departing from the scope and spirit of the described embodiments.

[0143] The endpoints and any values ​​of the ranges disclosed in this article are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of each range, the endpoint values ​​of each range and the individual point values, and the individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed in this article.

Claims

1. A Pseudomonas geniculata, characterized in that: The deposit number of the Pseudomonas geniculate is CGMCC No.22735.

2. A bacterial agent, characterized in that The bacterial agent contains the Pseudomonas genu as described in claim 1.

3. The bacterial agent according to claim 2, wherein The bacterial agent is a liquid bacterial agent and / or a solid bacterial agent; The number of viable bacteria in the liquid bacterial agent is 10 9 cfu / mL or more; The number of viable bacteria in the solid bacterial agent is 10 9 cfu / g or above.

4. Use of the Pseudomonas geniculate described in claim 1 or the bacterial agent described in claim 2 or 3 in degrading polycyclic aromatic hydrocarbons.

5. Use of the Pseudomonas geniculate described in claim 1 or the bacterial agent described in claim 2 or 3 in remediating soil contaminated by polycyclic aromatic hydrocarbons or groundwater contaminated by polycyclic aromatic hydrocarbons.

6. A method for degrading polycyclic aromatic hydrocarbons, characterized in that: The method comprises: contacting the Pseudomonas genu described in claim 1 or the bacterial agent described in claim 2 or 3 with pollutants containing polycyclic aromatic hydrocarbons.

7. The method according to claim 6, wherein: The polycyclic aromatic hydrocarbon is at least one of aromatic hydrocarbons containing at least two benzene rings.

8. The method according to claim 7, wherein: The polycyclic aromatic hydrocarbons are selected from naphthalene, anthracene, phenanthrene, pyrene, At least one of benzopyrene, benzanthracene, benzofluoranthene, indenopyrene, biphenyl, bipolyphenyl and polyphenylated aliphatic hydrocarbons.

9. The method according to any one of claims 6 to 8, wherein: The pollutant containing polycyclic aromatic hydrocarbons is soil containing polycyclic aromatic hydrocarbons or groundwater containing polycyclic aromatic hydrocarbons.

10. The method according to claim 9, wherein: In the soil containing polycyclic aromatic hydrocarbons, the content of the polycyclic aromatic hydrocarbons is 2-1000 mg / kg soil in terms of polycyclic aromatic hydrocarbons; In the groundwater containing polycyclic aromatic hydrocarbons, the content of the polycyclic aromatic hydrocarbons is 2-1000 mg / L groundwater, calculated as polycyclic aromatic hydrocarbons.

11. The method according to claim 9, wherein: When the polycyclic aromatic hydrocarbon-containing pollutant is soil containing polycyclic aromatic hydrocarbons, the amount of the Pseudomonas geniculate or the bacterial agent is such that the number of viable bacteria in the soil containing polycyclic aromatic hydrocarbons is 10 6 -10 8 cfu / g; When the polycyclic aromatic hydrocarbon-containing pollutant is groundwater containing polycyclic aromatic hydrocarbons, the amount of the Pseudomonas genu or the bacterial agent is such that the number of viable bacteria in the groundwater containing polycyclic aromatic hydrocarbons is 10 6 -10 8 cfu / mL.

12. The method according to any one of claims 6 to 8, wherein: The contact conditions include: temperature of 20-40° C. and pH of 5-10.

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