Lactobacillus paracasei, microbial inoculum and application of lactobacillus paracasei

Abstract

The invention belongs to the field of microorganisms for degrading polycyclic aromatic hydrocarbons, and relates to lactobacillus paracasei, a microbial agent and application of the lactobacillus paracasei. The preservation number of the lactobacillus paracasei is CGMCC (China General Microbiological Culture Collection Center) No.24463. The lactobacillus paracasei is stable in property, can rapidly grow in soil or underground water with polycyclic aromatic hydrocarbon as a unique carbon source, is excellent in polycyclic aromatic hydrocarbon degradation performance, and can be widely applied to remediation of soil or underground water containing polycyclic aromatic hydrocarbon.

Description

Technical Field

[0001] The invention belongs to the field of microorganisms for degrading polycyclic aromatic hydrocarbons, and specifically relates to a lactobacillus paracasei, a bacterial agent containing the lactobacillus paracasei, and applications of the lactobacillus paracasei and the bacterial agent in degrading polycyclic aromatic hydrocarbons. Background Art

[0002] Polycyclic aromatic hydrocarbons refer to aromatic hydrocarbons containing two or more benzene rings, referred to as PAHs. According to the different ways of connecting the benzene rings, they can be mainly divided into two categories. One category is the non-condensed ring type, which includes biphenyl, biphenyls and polyphenylated aliphatic hydrocarbons. The compound formed by the σ bond between benzene rings is called biphenyl. Biphenyl derivatives and biphenyl compounds are named after biphenyl as the parent. Polyphenylated aliphatic hydrocarbons are compounds formed by several benzene rings replacing hydrogen atoms in aliphatic hydrocarbons. This type of compound is named after phenyl as a substituent and aliphatic hydrocarbon as a parent. The structure and properties of non-condensed ring polycyclic aromatic hydrocarbons such as biphenyl, biphenyls and polyphenylated aliphatic hydrocarbons are similar to those of single-ring aromatic hydrocarbons. The other category is the condensed ring type, that is, two carbon atoms are shared by two benzene rings. For example, naphthalene, the molecular formula of naphthalene is C 10 H8, it is formed by two benzene rings sharing two adjacent carbon atoms. Naphthalene is a white flaky crystal with a melting point of 80°C and a boiling point of 218°C. It is insoluble in water but easily soluble in hot alcohol, ether and other organic solvents. It is volatile, easy to sublimate, and has a special smell. Naphthalene is the compound with the highest content in coal tar, accounting for about 10% in high-temperature coal tar. The molecular formula of anthracene is C 14 H 10 , which is composed of three benzene rings fused together. Anthracene exists in coal tar, with a content of about 0.25%. Phenanthrene also exists in coal tar and is an isomer of anthracene.

[0003] The sources of PAHs are divided into natural sources and anthropogenic sources. Natural sources mainly come from the biosynthesis process of terrestrial and aquatic plants and microorganisms. In addition, PAHs also exist in natural fires in forests and grasslands, volcanic eruptions, fossil fuels, lignin and sediments; anthropogenic sources are mainly formed by incomplete combustion of various mineral fuels (such as coal, oil and natural gas, etc.), wood, paper and other hydrocarbons, or pyrolysis under reducing conditions.

[0004] Unlike other persistent organic pollutants that are globally banned from production and use, the main sources of release of most PAHs are usually closely related to human activities. Therefore, the presence of PAHs in the environment is currently of great concern. PAHs are ubiquitous in various environments. They mainly enter surface water directly through atmospheric deposition, river runoff, urban and industrial wastewater, and leakage during transportation. Water is an important medium for the migration of PAHs in the environment. Surface runoff, atmospheric deposition, soil-sediment and air-water interface input exchange are all effective ways for their diffusion. This pollutant has extremely low solubility in water and has strong affinity and biochemical persistence. It has a strong adsorption effect on marine sediments and can often exist stably in different media in nature. However, when sediments are subjected to external forces such as waves, tides and ocean currents, trawling, ship traffic and water conservancy project operations, the pollutants in the sediments are released, exacerbating the pollution.

[0005] Most PAHs have strong toxic side effects on organisms, including immunotoxicity and carcinogenicity. Among them, benzo[a]pyrene has been classified as Group 1 by the International Atomic Energy Agency and is carcinogenic to humans. Other PAHs are divided into Group 2A and Group 2B. At the same time, PAHs have high bioaccumulation, which makes the adverse effects more serious. Even trace amounts can cause various diseases and cause serious harm to the human body. There is increasing evidence that long-term exposure to PAHs can not only cause certain specific cancers, but also certain other diseases, such as obesity and type 2 diabetes (T2D).

[0006] PAHs pollution has traditionally been treated using physical and chemical methods. Conventional methods using temperature, electrokinetics and redox have the advantages of short cycles and quick results, but their high energy consumption and obvious damage to the original site cannot be ignored. Microbial degradation has gradually become the main research direction for PAHs degradation because it has the advantages of lower cost and more convenient operation than other degradation methods.

[0007] In the natural environment, the decomposition and conversion mechanism of organic matter by microorganisms is the main method for removing organic pollutants without human interference. This degradation and conversion mechanism is the core idea of ​​using microbial methods to treat organic pollutants. When the pollutants on the site are chain petroleum hydrocarbons or organic pesticides, the microbial method often has a better treatment effect, but when encountering PAHs with high hydrophobicity, the effect of the microbial method is often unsatisfactory. In order to achieve better remediation effects, it is necessary not only to improve the remediation methods, but also to clarify the characteristics of microbial degradation, screen and develop microorganisms that are consistent with the structural characteristics and physical and chemical properties of the target pollutants for remediation, so as to achieve better degradation effects. Summary of the invention

[0008] The invention aims to provide a Lactobacillus paracasei with excellent performance of degrading polycyclic aromatic hydrocarbons, a bacterial agent and an application thereof.

[0009] The first aspect of the present invention provides a Lactobacillus paracasei, wherein the deposit number of the Lactobacillus paracasei is CGMCC No.24463.

[0010] The second aspect of the present invention provides a bacterial agent, which contains the Lactobacillus paracasei.

[0011] The third aspect of the present invention provides the use of the Lactobacillus paracasei or the bacterial agent in degrading polycyclic aromatic hydrocarbons.

[0012] Furthermore, the application method comprises: contacting the Lactobacillus paracasei or the bacterial agent with pollutants containing polycyclic aromatic hydrocarbons.

[0013] Furthermore, the polycyclic aromatic hydrocarbons are condensed-ring polycyclic aromatic hydrocarbons or non-condensed-ring polycyclic aromatic hydrocarbons.

[0014] Furthermore, the condensed-ring polycyclic aromatic hydrocarbons are selected from naphthalene, anthracene, phenanthrene, pyrene, At least one of benzopyrene, benzanthracene, benzofluoranthene and indenopyrene.

[0015] Furthermore, the non-condensed-ring polycyclic aromatic hydrocarbon is selected from at least one of biphenyl, bipolyphenyl and polyphenylated aliphatic hydrocarbon.

[0016] Furthermore, the biphenyl is terphenyl; and the polyphenylated aliphatic hydrocarbon is at least one of diphenylmethane and diphenylethane.

[0017] Furthermore, the pollutant containing polycyclic aromatic hydrocarbons is soil containing polycyclic aromatic hydrocarbons or groundwater containing polycyclic aromatic hydrocarbons.

[0018] Furthermore, in the soil containing polycyclic aromatic hydrocarbons, the content of polycyclic aromatic hydrocarbons is 2-1000 mg / kg of soil, calculated as polycyclic aromatic hydrocarbons; in the groundwater containing polycyclic aromatic hydrocarbons, the content of polycyclic aromatic hydrocarbons is 2-1000 mg / L of groundwater, calculated as polycyclic aromatic hydrocarbons.

[0019] Furthermore, when the polycyclic aromatic hydrocarbon-containing pollutant is soil containing polycyclic aromatic hydrocarbons, the amount of the Lactobacillus paracasei or the bacterial agent is such that the number of viable bacteria in the soil containing polycyclic aromatic hydrocarbons is 10 7 -10 9 cfu / g; when the PAH-containing pollutant is groundwater containing PAHs, the amount of Lactobacillus paracasei or the bacterial agent is such that the number of viable bacteria in the groundwater containing PAHs is 10 7 -10 9 cfu / mL.

[0020] Furthermore, the contact conditions include: temperature of 15-40° C. and pH of 5-10.

[0021] The Lactobacillus paracasei of the present invention has stable properties, can grow rapidly in soil or groundwater containing polycyclic aromatic hydrocarbons as the sole carbon source, has excellent performance in degrading polycyclic aromatic hydrocarbons, and can be widely used in the restoration of soil or groundwater containing polycyclic aromatic hydrocarbons.

[0022] Other features and advantages of the present invention will be described in detail in the following detailed description.

[0023] Biological Deposit Description

[0024] The biological material Lactobacillus paracasei F1 of the present invention is classified and named Lactobacillus paracasei, and was deposited in the China General Microbiological Culture Collection Center (CGMCC) of the China Microbiological Culture Collection Committee on March 2, 2022, with the address being: Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number being CGMCC NO.24463. DETAILED DESCRIPTION

[0025] 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.

[0026] The invention provides a Lactobacillus paracasei, and the preservation number of the Lactobacillus paracasei is CGMCC No. 24463. The Lactobacillus paracasei (preservation number CGMCC No. 24463) provided by the invention is a facultative anaerobic bacterium, Gram-positive, and is obtained by screening and separation from polycyclic aromatic hydrocarbons-contaminated soil in Tianjin.

[0027] The specific screening and separation steps are:

[0028] (1) Domestication

[0029] Take 3-5g of soil and add it to the sterilized inorganic salt culture medium. The volume of the culture medium is 150mL / 250mL Erlenmeyer flask. Then add phenanthrene as the only carbon source, shake well, and culture in a shaking cabinet at 30℃ and 150r / min. After 5 days, take 1mL of the bacterial solution in the culture bottle for transfer, and repeat at least 3 times.

[0030] (2) Separation

[0031] After the acclimated bacterial solution is diluted to an appropriate dilution, the bacterial solution is streaked using the plate streak method. Three parallel samples are made for each concentration gradient. After streaking, the culture dish is placed in an incubator at 30°C and cultured for 3 days.

[0032] (3) Purification

[0033] Observe the various colonies growing on the surface of the agar plate, pay attention to the differences in size, shape, edge, surface structure, transparency, color, properties, etc., take photos and save them. Repeatedly purify the colonies that have not formed single bacteria until the size and shape of the colonies on the same plate are uniform.

[0034] The purified Lactobacillus paracasei (named F1) was sent to Beijing Meiji Sanger Biopharmaceutical Technology Co., Ltd. for 16S rDNA strain identification. The results of the spliced ​​gene sequence of the strain and the NCBI comparison showed that the strain had 100% homology with the gene sequence of Lactobacillus paracasei strain. The isolated strain F1 was identified as Lactobacillus paracasei.

[0035] The lactobacillus paracasei F1 colonies provided by the invention are milky white, round, mostly with a diameter of more than 1.0 mm, a smooth and raised surface, and neat edges.

[0036] The inventors of the present invention have found through research that the Lactobacillus paracasei (preservation number CGMCC No. 24463) has good degradation activity on various types of polycyclic aromatic hydrocarbons; adding a bacterial agent containing the Lactobacillus paracasei (preservation number CGMCC No. 24463) into soil or groundwater contaminated by polycyclic aromatic hydrocarbons can repair the polycyclic aromatic hydrocarbon pollution in the soil or groundwater.

[0037] The bacterial agent may further contain auxiliary materials, which may be auxiliary materials commonly used in the art and will not affect the activity of Lactobacillus paracasei.

[0038] The preparation method of the bacterial agent can be a conventional preparation method in the art. Preferably, the preparation method comprises: activating the above-mentioned Lactobacillus paracasei and culturing it in a culture medium to obtain a bacterial liquid stock solution; preparing the bacterial bodies obtained after centrifugation of the bacterial liquid stock solution into a bacterial suspension with sterile water (the number of viable bacteria in the bacterial suspension is usually 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 Lactobacillus paracasei.

[0039] The polycyclic aromatic hydrocarbons refer to aromatic hydrocarbons containing two or more benzene rings, including both condensed-ring polycyclic aromatic hydrocarbons and non-condensed-ring polycyclic aromatic hydrocarbons.

[0040] The condensed-ring polycyclic aromatic hydrocarbons include but are not limited to naphthalene, anthracene, phenanthrene, pyrene, At least one of benzopyrene, benzanthracene, benzofluoranthene and indenopyrene. 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.

[0041] The non-condensed-ring polycyclic aromatic hydrocarbons include, but are not limited to, at least one of biphenyl (i.e., biphenyl), bipolyphenyl, and polyphenylated aliphatic hydrocarbons. The bipolyphenyl may be biphenyl terphenyl, etc., and the polyphenylated aliphatic hydrocarbons may be diphenylmethane, diphenylethane, etc.

[0042] The Lactobacillus paracasei and bacterial agent of the present invention are suitable for various pollutants containing polycyclic aromatic hydrocarbons, including but not limited to soil containing polycyclic aromatic hydrocarbons or groundwater containing polycyclic aromatic hydrocarbons.

[0043] 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.

[0044] In the present invention, the amount of the Lactobacillus paracasei F1 or the bacterial agent 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 Lactobacillus paracasei F1 or the bacterial agent is such that the number of viable bacteria in the soil containing polycyclic aromatic hydrocarbons is 10 7 -10 9 cfu / g. Wherein, the soil weight is calculated on a dry basis.

[0045] 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.

[0046] In some embodiments of the present invention, when the PAH-containing pollutant is groundwater containing PAHs, the amount of Lactobacillus paracasei F1 or the bacterial agent is such that the number of viable bacteria in the groundwater containing PAHs is 10 7 -10 9 cfu / mL.

[0047] The conditions for contacting Lactobacillus paracasei F1 or the bacterial agent with pollutants containing polycyclic aromatic hydrocarbons are usually natural environmental conditions, such as a temperature of 15-40° C. The bacterial strain of the present invention can play a role in degrading polycyclic aromatic hydrocarbons in a wide pH range of 5-10.

[0048] The present invention will be further described below in conjunction with embodiments, but the scope of the present invention is not limited to these embodiments.

[0049] Soil model evaluation method for bioremediation of petroleum hydrocarbon contaminated soil

[0050] The quality of bioremediation methods for petroleum hydrocarbon contamination in soil is ultimately evaluated by the degradation effect of petroleum hydrocarbons in the soil. Therefore, a soil model evaluation method for bioremediation methods for petroleum hydrocarbon contaminated soil was established. The model evaluation method is as follows: ① Enrich the bacterial agent in an inorganic salt culture medium with the target pollutant as the only carbon source; ② Centrifugal separation to obtain the enriched bacterial agent; ③ Put the bacterial agent into the soil together with a water retaining agent, a soil conditioner, and nutrients; ④ Add the target pollutant to the soil; ⑤ Regularly test the content of petroleum hydrocarbons in the soil and the total number of soil bacteria, and calculate the degradation effect of petroleum hydrocarbons.

[0051] The specific steps are:

[0052] 1. Take 1.5mL of bacterial solution and put it into a conical flask containing 150mL of inorganic salt culture medium, then add the target pollutant, shake at 150r / min for 5-10 days (depending on the amount of bacteria) to obtain enriched bacterial solution.

[0053] 2. Centrifuge the obtained enriched bacterial liquid at 8000r / min for solid-liquid separation, discard the upper culture medium solution, add 100mL of inorganic salt culture medium, shake thoroughly, centrifuge again at 8000r / min for solid-liquid separation, discard the upper solution, and keep the bottom solid for later use.

[0054] 3. Add water retaining agent, soil conditioner, nutrients and bottom solids to the soil, stir thoroughly, and add deionized water to maintain the soil moisture content within a certain range.

[0055] 4. Add the target pollutant to the soil and stir well.

[0056] 5. Test the content of target pollutants in soil and the total number of soil bacteria every week.

[0057] 6. Add water every two days to keep the soil moisture content within a certain range, and turn the soil 1-3 times a week.

[0058] Inorganic salt culture medium: NH4Cl 0.67g, NaNO3 1.06g, MgSO4·7H2O 0.1g, CaCl20.1g, K2HPO4·3H2O 1.5g, KH2PO4 0.5g, FeCl3·6H2O 0.1g, deionized water 1L, pH 7.0-7.5.

[0059] Analytical methods

[0060] The method for determining heterotrophic bacteria refers to the standard plate counting method of "Industrial Circulating Cooling Water Quality Analysis Method" GB / T14643.1-1993.

[0061] The determination method of PAHs in soil is based on the "Gas Chromatography-Mass Spectrometry for the Determination of Semivolatile Organic Compounds in Soil and Sediment" HJ 834-2017. A dichloromethane-acetone mixed solution is used for pressurized fluid extraction of PAHs. When the constant volume is 1.0 mL and the injection volume is 1.0 μL, the detection limit of the PAHs determination method is 0.06-0.3 mg / kg, and the lower limit of determination is 0.24-1.20 mg / kg.

[0062] Method for determination of total soil bacteria count

[0063] The total number of bacteria in the soil was determined by the plate culture colony counting method, and the culture medium for soil bacteria was meat extract peptone medium.

[0064] The specific measurement steps are:

[0065] Weigh 10g of soil sample, put it into a triangular flask containing 90mL of sterile deionized water, and place it on a shaker at room temperature for 4 hours at a speed of 160 times / min to fully mix the soil sample with water, fully disperse the microbial cells in the soil, and separate them from the soil. -1 Soil suspension: pipette 1 mL of the soil suspension into 9 mL of sterile water containing 8.5 g / L NaCl, pipette and pump 3 times with a sterile pipette to mix well, and make 10 -2 Soil suspension. And so on to make 10 -3 , 10 -4 , 10 -5 , 10 -6 , 10 -7 , 10 -8 Soil suspensions with different dilutions. After selecting the appropriate dilution, the total bacterial count of the soil suspension was determined by the mixed plate count method.

[0066] The total number of bacteria in the soil, x (individuals / g), is calculated according to the following formula.

[0067]

[0068] Where:

[0069] x——total number of soil bacteria, pieces / g;

[0070] e——Total number of bacteria in soil suspension, counts / mL;

[0071] V——volume of soil suspension, mL;

[0072] m——sampling volume, g;

[0073] w——soil moisture content, %.

[0074] Meat extract peptone culture medium composition: NaCl 10g, peptone 10g, beef extract 5g, agar 18g, deionized water 1L, pH value 7.2-7.4.

[0075] The following examples and comparative examples all adopt the following test methods:

[0076] The soil used was dry soil sterilized at 250℃ for 24h. Sawdust was added to improve soil air permeability; polyacrylic acid was added to improve soil water retention; urea was added to supplement the soil N source. Mix well. The solid part of the bacterial solution was fully mixed with deionized water and added to the soil. Finally, the target pollutant polycyclic aromatic hydrocarbons were dissolved in 3mL of acetone and added to the soil. The soil environment temperature range was 18-23℃.

[0077] The soil was turned over once a week and deionized water was added until the soil moisture content reached the initial value. No other intervention measures were taken.

[0078] The method for obtaining the solid part of the bacterial liquid comprises: (1) taking 1.5 mL of the bacterial liquid into a triangular flask containing 150 mL of inorganic salt culture medium, adding the target pollutant as the only carbon source, and placing it in a constant temperature shaking cabinet at 30° C. and 150 r / min for 5-10 days to obtain the OD 600 (2) The obtained enriched bacterial solution was centrifuged at 8000 r / min for solid-liquid separation, the upper culture medium solution was discarded, 100 mL of inorganic salt culture medium was added, and after sufficient shaking, the solid-liquid separation was centrifuged again at 8000 r / min, the upper solution was discarded, and the solid part of the bacterial solution was obtained.

[0079] Inorganic salt culture medium composition: NH4Cl 0.67g, NaNO3 1.06g, MgSO4·7H2O 0.1g, CaCl2 0.1g, K2HPO4·3H2O 1.5g, KH2PO4 0.5g, FeCl3·6H2O 0.1g, deionized water 1L, pH value 7.0~7.5.

[0080] In the following examples and comparative examples:

[0081] Biphenyl was purchased from Beijing Inokai Technology Co., Ltd.

[0082] 1,4-Terphenyl was purchased from Beijing Inokai Technology Co., Ltd.

[0083] Diphenylmethane was purchased from Beijing Inokai Technology Co., Ltd.

[0084] Naphthalene was purchased from Beijing Inokai Technology Co., Ltd.

[0085] Anthracene was purchased from Beijing Inokai Technology Co., Ltd.

[0086] Phenanthrene was purchased from Beijing Inokai Technology Co., Ltd.

[0087] Pyrene was purchased from Beijing Inokai Technology Co., Ltd.

[0088] Benzo[a]pyrene was purchased from Beijing Inokai Technology Co., Ltd.

[0089] Oligotrophomonas was purchased from China General Microbiological Culture Collection Center, with the accession number CGMCC1.6393;

[0090] Acinetobacter was purchased from China General Microbiological Culture Collection Center, with the collection number CGMCC1.10395 T ;

[0091] Dietrichia coli was purchased from China General Microbiological Culture Collection Center, with the collection number CGMCC1.6332 T .

[0092] Example 1

[0093] 6 g of polyacrylic acid was weighed and dissolved in 200 g of deionized water to form aqueous solution 1, 0.96 g of urea was weighed and dissolved in 100 g of deionized water to form aqueous solution 2, Lactobacillus paracasei was mixed in 300 g of deionized water to form aqueous solution 3, 45 g of sawdust was weighed and mixed in 2400 g of soil, and then aqueous solutions 1, 2, and 3 were added and mixed evenly. 150 mg of biphenyl was added to the evenly mixed soil.

[0094] Example 2

[0095] Weigh 6 g of polyacrylic acid and dissolve it in 200 g of deionized water to form aqueous solution 1, weigh 0.96 g of urea and dissolve it in 100 g of deionized water to form aqueous solution 2, take Lactobacillus paracasei and mix it in 300 g of deionized water to form aqueous solution 3, weigh 45 g of sawdust and mix it in 2400 g of soil, then add aqueous solutions 1, 2, and 3 and mix them evenly. Add 150 mg of 1,4-terphenyl to the evenly mixed soil.

[0096] Example 3

[0097] 6 g of polyacrylic acid was weighed and dissolved in 200 g of deionized water to form aqueous solution 1, 0.96 g of urea was weighed and dissolved in 100 g of deionized water to form aqueous solution 2, Lactobacillus paracasei was mixed in 300 g of deionized water to form aqueous solution 3, 45 g of sawdust was weighed and mixed in 2400 g of soil, and then aqueous solutions 1, 2, and 3 were added and mixed evenly. 150 mg of diphenylmethane was added to the evenly mixed soil.

[0098] Example 4

[0099] 6 g of polyacrylic acid was weighed and dissolved in 200 g of deionized water to form aqueous solution 1, 0.96 g of urea was weighed and dissolved in 100 g of deionized water to form aqueous solution 2, Lactobacillus paracasei was mixed in 300 g of deionized water to form aqueous solution 3, 45 g of sawdust was weighed and mixed in 2400 g of soil, and then aqueous solutions 1, 2, and 3 were added and mixed evenly. 150 mg of naphthalene was added to the evenly mixed soil.

[0100] Example 5

[0101] 6 g of polyacrylic acid was weighed and dissolved in 200 g of deionized water to form aqueous solution 1, 0.96 g of urea was weighed and dissolved in 100 g of deionized water to form aqueous solution 2, Lactobacillus paracasei was mixed in 300 g of deionized water to form aqueous solution 3, 45 g of sawdust was weighed and mixed in 2400 g of soil, and then aqueous solutions 1, 2, and 3 were added and mixed evenly. 150 mg of anthracene was added to the evenly mixed soil.

[0102] Example 6

[0103] 6 g of polyacrylic acid was weighed and dissolved in 200 g of deionized water to form aqueous solution 1, 0.96 g of urea was weighed and dissolved in 100 g of deionized water to form aqueous solution 2, Lactobacillus paracasei was mixed in 300 g of deionized water to form aqueous solution 3, 45 g of sawdust was weighed and mixed in 2400 g of soil, and then aqueous solutions 1, 2, and 3 were added and mixed evenly. 150 mg of phenanthrene was added to the evenly mixed soil.

[0104] Example 7

[0105] 6 g of polyacrylic acid was weighed and dissolved in 200 g of deionized water to form aqueous solution 1, 0.96 g of urea was weighed and dissolved in 100 g of deionized water to form aqueous solution 2, Lactobacillus paracasei was mixed in 300 g of deionized water to form aqueous solution 3, 45 g of sawdust was weighed and mixed in 2400 g of soil, and then aqueous solutions 1, 2, and 3 were added and mixed evenly. 150 mg of pyrene was added to the evenly mixed soil.

[0106] Example 8

[0107] 6 g of polyacrylic acid was weighed and dissolved in 200 g of deionized water to form aqueous solution 1, 0.96 g of urea was weighed and dissolved in 100 g of deionized water to form aqueous solution 2, Lactobacillus paracasei was mixed in 300 g of deionized water to form aqueous solution 3, 45 g of sawdust was weighed and mixed in 2400 g of soil, and then aqueous solutions 1, 2, and 3 were added and mixed evenly. 150 mg of benzo[a]pyrene was added to the evenly mixed soil.

[0108] Comparative Example 1

[0109] Weigh 6g of polyacrylic acid and dissolve it in 200g of deionized water to form aqueous solution 1, weigh 0.96g of urea and dissolve it in 100g of deionized water to form aqueous solution 2, take oligotrophomonas and mix it in 300g of deionized water to form aqueous solution 3, weigh 45g of sawdust and mix it in 2400g of soil, then add aqueous solutions 1, 2, and 3 and mix them evenly. Add 150mg of biphenyl to the evenly mixed soil.

[0110] Comparative Example 2

[0111] Weigh 6 g of polyacrylic acid and dissolve it in 200 g of deionized water to form aqueous solution 1, weigh 0.96 g of urea and dissolve it in 100 g of deionized water to form aqueous solution 2, take Acinetobacter and mix it in 300 g of deionized water to form aqueous solution 3, weigh 45 g of sawdust and mix it in 2400 g of soil, then add aqueous solutions 1, 2, and 3 and mix them evenly. Add 150 mg of diphenylmethane to the evenly mixed soil.

[0112] Comparative Example 3

[0113] 6 g of polyacrylic acid was weighed and dissolved in 200 g of deionized water to form aqueous solution 1, 0.96 g of urea was weighed and dissolved in 100 g of deionized water to form aqueous solution 2, Dietrichia was mixed in 300 g of deionized water to form aqueous solution 3, 45 g of sawdust was weighed and mixed in 2400 g of soil, and then aqueous solutions 1, 2, and 3 were added and mixed evenly. 150 mg of phenanthrene was added to the evenly mixed soil.

[0114] Comparative Example 4

[0115] Weigh 6 g of polyacrylic acid and dissolve it in 200 g of deionized water to form aqueous solution 1, weigh 0.96 g of urea and dissolve it in 100 g of deionized water to form aqueous solution 2, take Dietrichia and mix it in 300 g of deionized water to form aqueous solution 3, weigh 45 g of sawdust and mix it in 2400 g of soil, then add aqueous solutions 1, 2, and 3 and mix them evenly. Add 150 mg of benzo[a]pyrene to the evenly mixed soil.

[0116] Comparative Example 5

[0117] 6 g of polyacrylic acid was weighed and dissolved in 200 g of deionized water to form aqueous solution 1, 0.96 g of urea was weighed and dissolved in 100 g of deionized water to form aqueous solution 2, 45 g of sawdust was weighed and mixed in 2400 g of soil, and then aqueous solutions 1, 2 and 300 g of deionized water were added and mixed well. 150 mg of benzo[a]pyrene was added to the mixed soil.

[0118] Test Example 1

[0119] Soil samples of the above embodiments and comparative examples were taken regularly to measure the residual PAH content in the soil samples and investigate the degradation of PAHs. The results are shown in Table 1.

[0120] Table 1 Polycyclic aromatic hydrocarbons content of soil samples in Examples and Comparative Examples

[0121] initial 7d 14d 21d 28d 35d 42d 49d Example 1 50 48.1 43.8 29.4 17.6 8 2.3 0.3 Example 2 50 48.5 44.1 32.4 17.8 9.1 4.1 1.7 Example 3 50 48.5 44.5 31.8 18.6 10.8 5.4 1.9 Example 4 50 48.7 45.8 34.2 20.3 12.6 7.4 2.3 Example 5 50 48.8 46.1 33.8 21.4 13.2 9.8 2.5 Example 6 50 48.4 46.2 35.6 21.5 13.5 9.8 2.5 Example 7 50 49.6 49.2 44.8 38.1 29.8 22.4 15.0 Example 8 50 49.8 49.4 46.2 41.0 33.4 28.4 22.4 Comparative Example 1 50 49.8 48.1 43.9 39.6 38.1 36.4 34.3 Comparative Example 2 50 49.9 48.6 44.0 39.2 38.2 37.4 35.9 Comparative Example 3 50 49.9 48.9 46.7 42.5 41.8 40.7 39.9 Comparative Example 4 50 49.9 49.8 48.7 47.6 47.1 47.0 47.1 Comparative Example 5 50 49.9 49.9 49.8 49.6 49.2 49.1 49.1

[0122] Note: The unit of PAH content is mg / kg.

[0123] It can be seen from the results in Table 1 that the Lactobacillus paracasei of the present invention has an obvious degradation effect on various types of polycyclic aromatic hydrocarbons, and is better than the control strains.

[0124] Test Example 2

[0125] Soil samples of the above embodiments and comparative examples were taken regularly to measure the total number of bacteria in the soil samples and to investigate the growth of microorganisms. The results are shown in Table 2.

[0126] Table 2 Growth of soil microorganisms in the examples and comparative examples

[0127] initial 7d 14d 21d 28d 35d 42d 49d Example 1 37 135 657 1358 971 758 297 124 Example 2 38 124 599 1247 912 701 288 136 Example 3 35 130 641 1178 863 640 325 150 Example 4 35 112 495 1046 579 376 298 145 Example 5 36 108 481 1033 531 344 284 123 Example 6 37 116 452 1002 458 325 278 117 Example 7 32 86 245 699 430 298 239 98 Example 8 38 74 213 644 398 245 216 69 Comparative Example 1 39 119 478 1095 795 567 221 89 Comparative Example 2 38 114 465 937 659 463 216 99 Comparative Example 3 35 102 298 682 203 186 145 63 Comparative Example 4 38 65 117 367 198 116 86 41 Comparative Example 5 0.2 0.3 0.9 8 12 7 4.1 0.4

[0128] Note: The total number of bacteria is 10 6 cfu / g soil.

[0129] It can be seen from the results in Table 2 that with the increase of treatment time, the growth of Lactobacillus paracasei in the soil reached a peak at 21 days and dropped to a level equivalent to 7 days at 49 days. During this period, the content of polycyclic aromatic hydrocarbons in the soil samples continued to decrease, indicating that the reduction in the content of polycyclic aromatic hydrocarbons in the soil was due to the degradation of Lactobacillus paracasei.

[0130] 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.

[0131] 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 Lactobacillus paracasei, characterized in that The preservation number of the Lactobacillus paracasei is CGMCC No.24463.

2. A bacterial agent, characterized in that The bacterial agent contains the Lactobacillus paracasei according to claim 1.

3. Application of the Lactobacillus paracasei described in claim 1 or the bacterial agent described in claim 2 in degrading polycyclic aromatic hydrocarbons.

4. The use according to claim 3, wherein: The application method comprises: contacting the Lactobacillus paracasei described in claim 1 or the bacterial agent described in claim 2 with pollutants containing polycyclic aromatic hydrocarbons.

5. The use according to claim 3, wherein: The polycyclic aromatic hydrocarbons are condensed-ring polycyclic aromatic hydrocarbons or non-condensed-ring polycyclic aromatic hydrocarbons.

6. The use according to claim 5, wherein: The condensed-ring polycyclic aromatic hydrocarbons are selected from naphthalene, anthracene, phenanthrene, pyrene, At least one of benzopyrene, benzanthracene, benzofluoranthene and indenopyrene.

7. The use according to claim 5, wherein: The non-condensed-ring polycyclic aromatic hydrocarbon is selected from at least one of biphenyl, bipolyphenyl and polyphenylated aliphatic hydrocarbon.

8. The use according to claim 7, wherein: The biphenyl is terphenyl; the polyphenylated aliphatic hydrocarbon is at least one of diphenylmethane and diphenylethane.

9. The use according to claim 4, wherein: The pollutant containing polycyclic aromatic hydrocarbons is soil containing polycyclic aromatic hydrocarbons or groundwater containing polycyclic aromatic hydrocarbons.

10. The use 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 use according to claim 9, wherein: When the polycyclic aromatic hydrocarbon-containing pollutant is soil containing polycyclic aromatic hydrocarbons, the amount of the Lactobacillus paracasei or the bacterial agent is such that the number of viable bacteria in the soil containing polycyclic aromatic hydrocarbons is 10 7 -10 9 cfu / g; When the polycyclic aromatic hydrocarbon-containing pollutant is groundwater containing polycyclic aromatic hydrocarbons, the amount of the Lactobacillus paracasei or the bacterial agent is such that the number of viable bacteria in the groundwater containing polycyclic aromatic hydrocarbons is 10 7 -10 9 cfu / mL.

12. The use according to any one of claims 4 to 9, wherein: The contact conditions include: temperature of 15-40° C. and pH of 5-10.

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