CN116286511A - Flora with soil remediation function and application thereof - Google Patents

Abstract

The invention provides a flora with petroleum degradation function, which comprises Gordonia sp.Strain C1, cellulomicrobacteria C3 (Cellulomicrobacteria sp.Strain C3) or both, wherein the preservation numbers of the Gordonia C1 and the Cellulomicrobacteria C3 are CGMCC No.25708 and CGMCC No.25709 respectively. The flora and the culture formula provided by the invention have the advantages that the degradation rate of petroleum hydrocarbon in a petroleum degradation system with petroleum content of 60000-100000mg/L is more than 60%, the maximum degradation rate can reach more than 85%, the flora and the culture formula can be applied to a petroleum pollution treatment process with microorganisms as cores, the ecological management of petroleum pollution environment is facilitated, and the flora and the culture formula have wide application prospects in the field of ecological restoration of petroleum pollution soil or water bodies.

Description

Flora with soil remediation function and application thereof

Technical Field

The invention relates to the technical field of environmental biology, in particular to a flora with petroleum degradation function and application thereof.

Background

Petroleum is an important energy source for social development, and the petroleum industry has irreplaceable value in the aspects of promotion of world economic development, improvement of social life and the like. However, oil pollution accidents during exploitation, transportation, use, storage and the like are accompanied, and the frequency of oil leakage pollution is obviously increased. Because the compound in petroleum is very complex in type and contains a large amount of high-toxicity components which are difficult to degrade and easy to accumulate, the pollution caused by petroleum leakage seriously threatens the environmental safety, and further threatens the human health. With the gradual rise of social level and the gradual enhancement of human environmental awareness, how to cope with petroleum leakage and environmental pollution brought by petroleum leakage is one of the important problems of world concern.

Petroleum is a mixture of various organic matters, has large chemical component difference, and has the characteristics of uneven dispersion, high heterogeneity and the like in petroleum pollution environments, thereby providing higher requirements for effectively treating the petroleum pollution of the environment. Over the past few decades, petroleum pollution treatment processes have been developed with physical, chemical and biological cores, respectively. Among them, in the bioremediation technology, a biological treatment process for biodegrading petroleum-contaminated environments using microorganisms has been attracting attention because of its environmental friendliness, high efficiency and low cost, as compared with other strategic methods for treating petroleum contamination.

At present, researches on petroleum degradation microorganisms are actively carried out in China on the microbial remediation of petroleum polluted environments. The prior researches focus on single petroleum degrading microorganisms, and degradation culture conditions are not optimized according to strain characteristics, such as patent CN111718867B, CN105505812B, CN109777747A and the like. In addition, since the degradation effect of a single kind of degradation bacteria is limited, and the mixed bacteria degradation system can realize efficient and rapid degradation of petroleum pollutants, the construction of the mixed bacteria degradation system has attracted various attention, such as patents CN110511890a and CN111534462 a. However, most of the mixed systems have problems of low petroleum hydrocarbon degradation efficiency, low degradation concentration and the like. Therefore, there is an urgent need to develop a complex bacterial colony having petroleum degradation function, which can achieve efficient degradation of petroleum pollutants at high concentration.

Disclosure of Invention

In view of the above, the present invention provides a complex bacterial colony with petroleum degradation function and application thereof, so as to solve the above-mentioned technical problems.

In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:

the invention provides two strains of petroleum degrading functional bacteria, wherein a single strain thereof screens oil sludge separated from a sludge pit of a Qinghai oilfield oil extraction plant, and colony morphological characteristics and molecular biology identification are carried out on the two obtained strains of petroleum degrading functional bacteria. The identification shows that the obtained C1 colony is orange, opaque and convex; c3 colony, the color is changed from white to green, opaque and convex. Phylogenetic tree constructed using MEGA7.0 software based on Maximum likelihood method suggests that the strain is named Gordonia sp (Gordonia sp), gordonia sp.strain C1 and cellobiobacterium (Cellulosimicrobium sp) named cellulimium sp.strain C3 in order.

The single petroleum degrading bacteria Gordonia C1 (Gordonia sp.Strain C1) and Cellulomicrobacteria C3 (Cellulomicrobium sp.Strain C3) obtained were subjected to strain-unique carbon source utilization tests.

The invention provides a culture formula and culture conditions of two petroleum degradation functional bacteria. The culture formula comprises the following steps: BO medium; culture conditions: 26-37 deg.c and 150-200rpm.

As another aspect of the present invention, there is provided application conditions of a complex bacterial liquid having petroleum degradation function, wherein the culture medium uses a modified BO culture formulation, and the culture conditions are 26-37 ℃ and 150-200rpm.

The invention also provides a culture and compounding method for the combined application of two petroleum degradation functional bacteria, and a single strainCulturing Gordonia C1 and Cellulomicrobacteria C3 separately; adjusting the concentration OD of two strains of bacteria ₆₀₀ And the weight ratio is 0.6-1.0, and synchronous mixing and compounding are carried out under the conditions of 50-70% and 30-50% respectively.

The invention also provides application of the flora in degrading petroleum hydrocarbon in petroleum pollution environment.

The invention also provides a formula for the flora in a petroleum degradation system.

The invention also provides a method for degrading petroleum hydrocarbon. Inoculating the mixed flora into a degradation system with petroleum mass concentration of 60000-100000mg/L at the inoculum size of 10% -20%, and culturing at 26-37deg.C and 150-200rpm for 8-24 days to degrade petroleum.

More specifically, the present invention includes the following:

1. a flora, characterized in that the flora comprises Gordonia (Gordonia sp.) C1 with a collection number of CGMCC No.25708, cellomicrozyme (cellomium sp.) C3 with a collection number of CGMCC No.25709, or both.

2. The flora according to claim 1, characterized in that it consists of 50-70% by volume of Gordonia sp C1 and 30-50% by volume of cellomicrozyme sp C3.

3. The method of culturing the flora according to item 1 or 2, wherein the culture medium used for the culture has a composition comprising: CH (CH) ₃ COONa 1g/L，C ₃ H ₃ NaO ₃ 0.5g/L，K ₂ HPO ₄ 0.5g/L，KNO ₃ 0.1g/L，NH ₄ Cl 0.05g/L，NaCl 25g/L，FeC ₆ H ₅ O ₇ 0.002g/L, natural pH; the culture conditions are as follows: 26-37 deg.c and 150-200rpm.

4. The application of the flora in the aspect 1 or 2 in the field of petroleum pollution environmental treatment.

5. The use according to item 4, wherein the petroleum-contaminated environment comprises petroleum-contaminated soil and petroleum-contaminated water.

6. A method for the microecological remediation of petroleum pollution, wherein the method is to degrade petroleum hydrocarbons in petroleum contaminated soil or water using the flora of item 1 or 2.

7. A method for degrading petroleum hydrocarbon, characterized in that the method is to inoculate the flora in claim 1 or 2 into a degradation system containing petroleum with mass concentration of 60000-100000mg/L, and culture at 26-37 ℃ and 150-200rpm.

8. The method of claim 7, wherein the nutrient component in the degradation system is CH ₃ COONa 0.3g/L，C ₃ H ₃ NaO ₃ 0.1g/L，K ₂ HPO ₄ 1g/L，KNO ₃ 2g/L，NH ₄ Cl0.5g/L，NaCl 10g/L，FeC ₆ H ₅ O ₇ 0.005g/L。

9. An article of manufacture for use in the remediation of petroleum contaminated environments, said article of manufacture comprising the population of claim 1 or 2.

10. The article of claim 9, wherein the article is a liquid microbial inoculant article.

Compared with the prior art, the invention has the following beneficial technical effects:

the flora with petroleum degradation function provided by the invention can realize high-efficiency degradation of petroleum pollutants with high concentration (60000-100000 mg/L), and the degradation efficiency of the two strains is obviously improved compared with that of a single strain when the two strains are used together.

Drawings

FIG. 1 is a photograph showing a culture of Gordonia C1 (Gordonia sp. Strain C1);

FIG. 2 is a photograph showing a culture of Cellulomicrobium sp.Strain C3;

FIG. 3 shows growth curves of strains at 37℃under different medium conditions;

FIG. 4 shows growth curves of strains under different temperature conditions in BO medium;

FIG. 5 is a graph showing the degradation rate of petroleum hydrocarbon after the strain is degraded in a degradation system with a petroleum mass concentration of 80000mg/L for 24 days;

FIG. 6 shows the strain after 24 days of degradation in a degradation system with a petroleum mass concentration of 80000mg/L ₁₀ -C ₄₀ GC-MS analysis peak patterns of aliphatic hydrocarbons;

FIG. 7 is a graph showing the degradation rate of petroleum hydrocarbon after 8 days, 16 days and 24 days of biodegradation of the composite bacterial liquid in a degradation system with the petroleum mass concentration of 80000 mg/L.

Biological material sample preservation information:

gordonia C1 (Gordonia sp. Strain C1), class designation: gordonia sp.15 days 2022 was preserved in China general microbiological culture Collection center (CGMCC) with the address of Beijing Kogyo-Chao North West Lu No. 1, 3, china academy of sciences microbiological study with the preservation number of CGMCC No.25708.

Cellulomicrobium sp.Strain C3), class designation: cellulomictibium sp.9.15 of 2022 is preserved in China general microbiological culture Collection center (CGMCC) with the preservation address of Beijing, chaoyang area, north Chenxi Lu No. 1, 3 and the preservation number of CGMCC No.25709.

Detailed Description

The present invention will be further described in detail below with reference to specific embodiments and with reference to the accompanying drawings, in order to make the objects, technical solutions and advantages of the present invention more apparent. This example is intended to be illustrative of the invention and is not intended to be limiting. The experimental methods in the following examples are conventional experimental procedures unless otherwise specified. All of the instruments, consumables, reagents, etc. in the following examples are commercially available unless otherwise specified. Quantitative experiments in the following examples were performed with three biological replicates per experiment.

The medium formulation used in the following examples was:

microorganism enrichment medium: 20mM NH ₄ Cl，5mM K ₂ HPO ₄ Natural pH.

LB1 medium: 15g/L NaCl, 3g/L yeast, 7g/L tryptone and natural pH.

Biodegradable (BO) medium: CH (CH) ₃ COONa 1g/L，C ₃ H ₃ NaO ₃ 0.5g/L，K ₂ HPO ₄ 0.5g/L，KNO ₃ 0.1g/L，NH ₄ Cl 0.05g/L，NaCl 25g/L，FeC ₆ H ₅ O ₇ 0.002g/L, natural pH. Sterilizing at 115 deg.C for 30min.

Improved BO medium: CH (CH) ₃ COONa 0.3g/L，C ₃ H ₃ NaO ₃ 0.1g/L，K ₂ HPO ₄ 1g/L，KNO ₃ 2g/L，NH ₄ Cl 0.5g/L，NaCl 10g/L，FeC ₆ H ₅ O ₇ 0.005g/L, natural pH. Sterilizing at 115 deg.C for 30min.

Petroleum degradation culture medium: 80000mg/L petroleum was added to BO medium and sterilized at 115℃for 45min.

Petroleum screening plate medium: sequentially adding Tween 80 2.5mL/L, sodium hexametaphosphate 0.2g/L, polyvinylpyrrolidone K30.05 g/L,10% SDS 0.5mL/L, pH=8.0EDTA 1mL/L, petroleum 100000mg/L and agar 15g/L on the basis of microorganism enrichment culture medium, thoroughly mixing, and sterilizing at 115 ℃ for 45min.

Gas chromatograph-mass spectrometer (GC-MS-QP 2010 Ultra, japan), multifunctional enzyme-labeled instrument (molecular devices SpectraMax i X, USA).

Example 1: separation and identification of petroleum degradation functional bacteria and carbon utilization test

About 30g of crude oil contaminated soil was added to 200mL of the microorganism enrichment medium, and sterilized petroleum (5-10 g) was supplemented for an average of 30 days, 5-20mL of the microorganism enrichment medium, to construct a 200mL petroleum degradation functional microorganism enrichment reactor. After 180 days from the construction of the petroleum degradation functional microorganism enrichment reactor, the microorganism is transferred into an enrichment culture medium containing 15% crude oil (m/v) with 20% of inoculum size, and enrichment selection culture is carried out for 6 cycles again with 15-20 days as a cycle. 30-37 deg.c and 150-180rpm.

Sequentially diluting the enriched bacterial solution to 10 by using 1 XPBS (phosphate buffer solution) and adopting a gradient dilution method ^-6 、10 ^-7 And 10 ^-8 And single colony streaking is carried out on a petroleum screening flat-plate culture medium which takes petroleum as the only carbon source, and the culture is carried out for 2-4 days at 37 ℃. Selecting colonies with different sizes, colors and morphologies, and streaking and culturing the same colony for 3-5 timesFinally, two potential petroleum degradation functional bacteria are obtained. And (3) carrying out colony morphological characteristics and molecular biological identification on the obtained two petroleum degrading bacteria. The identification shows that the obtained C1 colony is orange, opaque and convex (shown in figure 1); c3 colonies, which changed from white to green in color, were opaque and convex (as shown in fig. 2). Phylogenetic tree was constructed using MEGA7.0 software based on Maximum likelihood method, and identified as Gordonia sp.strain C1 and cellobiobacillus C3 (cellobiomicrobium sp.strain C3), respectively.

The Gordonia sp.Strain C1 has been preserved in China general microbiological culture Collection center (CGMCC) with the address of Beijing, chaoyang, no. 1, west North, and China center for China with the preservation number of CGMCC No.25708 at 9 months and 15 days of 2022. The corresponding Genbank accession No. OP389054.1.

The Cellulomicrobium sp.Strain C3 has been preserved in China general microbiological culture Collection center (CGMCC) with the address of Hospital No. 3 in the North Chen West Lu 1 of the Korean region of Beijing city and the preservation number of CGMCC No.25709 in the year 2022, 9 and 15. The corresponding Genbank accession No. OP389052.1.

1 percent (according to the characteristics of the selected carbon source, the liquid culture medium adopts volume fraction, the solid culture medium adopts mass fraction) of different carbon sources are added into the microorganism enrichment culture medium, the carbon source utilization condition of two strains is tested, and detailed results are shown in table 1, wherein "+" indicates that the strains grow, and "—" indicates that the strains do not grow.

TABLE 1 carbon source utilization Table

Carbon source	Gordonia C1	Cellulomicrobacteria C3
			Glucose	﹣	﹣
Fructose	﹣	﹣
			Sucrose	﹢	﹢
Trehalose	﹢	﹢
			Cellobiose	﹣	﹣
Acetic acid sodium salt	﹢	﹢
			Pyruvic acid sodium salt	﹢	﹢
Succinic acid sodium salt	﹢	﹢
			Sodium fumarate	﹢	﹣
Sodium citrate	﹣	﹣
			Gluconic acid sodium salt	﹢	﹢
Glycerol	﹢	﹢
			N-butanol	﹢	﹣
Tween 80	﹢	﹢
			N-hexadecane	﹢	﹢
Paraffin oil	﹢	﹢

Analysis of the utilization conditions of different carbon sources by the strain shows that Gordonia C1 and Cellularomyces C3 have great application potential in alkane degradation, and simultaneously lay a foundation for designing specific nutrition culture formulas (such as an improved BO culture medium). In addition, because the two carbon sources are utilized differently, the combined application has wider carbon source utilization.

Example 2: characterization of the functional Strain traits of Gordonia C1 and Cellulomicrobacteria C3 cultivated using a Biodegradable (BO) Medium

The functional properties of the strain mainly comprise growth potential, emulsifying property, cell surface hydrophobicity and petroleum degradation potential.

Gordonia C1 (Gordonia sp.Strain C1) and Cellulomicrobacteria C3 (Cellulomicrobium sp.Strain C3) obtained in example 1 were activated with LB1 medium, and cultured at 37℃and 180rpm for 60 hours. Cells were washed 2 times with 1 XPBS and strain suspension OD was adjusted ₆₀₀ =1. Inoculating 1%o of inoculum size into LB1 culture medium and BO culture medium respectively, culturing at 37deg.C and 200rpm for 144 hr, sampling at regular time to determine bacterial liquid OD ₆₀₀ . The results showed that in LB1 medium the growth of the strain was relatively large, but it took longer to reach stationary phase and had a longer delay period than when BO medium was used (FIG. 3). Furthermore, the Cellularomyces C3 has a relatively high growth.

On the other hand, the cell suspension after the activation treatment is respectively inoculated into BO culture medium with an inoculum size of 1 per mill, and is respectively cultured at 26 ℃,30 ℃ and 37 ℃ and 200rpm for 120 hours, and the bacterial liquid OD is sampled and measured at regular time ₆₀₀ . The results show that the strain grows consistently at 3 different temperatures (fig. 4), and that the growth of the cellomicrozyme C3 is relatively higher than that of gordonia C1, indicating that the strain has potential for room temperature applications.

By MATH test ^[1] The surface hydrophobicity of the cells was evaluated and the results are shown in table 2. As can be seen from table 2, the cell surface hydrophobicity of gordonia C1 is significantly higher than that of cellobioc 3, especially under the conditions of culture using BO medium, indicating that the use of BO medium is most conducive to cell adhesion to the petroleum hydrocarbon surface and degradation of petroleum hydrocarbons.

TABLE 2 cell surface hydrophobicity (CSH%) of strains after cultivation in different media

The surfactant has the ability to enhance the dissolution of petroleum hydrocarbon contaminants, promote entry of hydrophobic materials into cells and accelerate their biodegradation. The bacterial emulsifying property is evaluated by E24 value by taking paraffin oil as crude oil substitute, thereby reflecting the capacity of the bacterial strain to produce the surfactant. As can be seen from Table 3, the E24 value of Cellularomyces C3 was significantly higher than that of Yu Gedeng, C1, indicating higher emulsifying properties.

TABLE 3E 24 of supernatants of strains after cultivation in different media

	LB1-E24	BO-E24
			Gordonia C1	12.66±2.51 b	6.67±2.89 b
Cellulomicrobacteria C3	75±5 a	34.33±4.04 a

Gordonia C1 and Cellulomicrobacteria C3 of example 1 were inoculated into LB1 medium (petroleum content 80000 mg/L) to which petroleum was added and petroleum degradation medium, respectively, at 37℃and 200rpm, and cultured for 24 days at 15% inoculum size.

Extracting petroleum residues in the fermentation broth after biodegradation by an ultrasonic method, measuring the total petroleum hydrocarbon degradation rate by a gravimetric method, and evaluating the degradation performance of single bacteria ^[2] . As a result, as shown in FIG. 5, when LB1 medium was used, the maximum degradation rate of total petroleum hydrocarbon was 43.36%, whereas when petroleum degradation medium was used, the totalThe degradation rate of petroleum hydrocarbon is more than 60 percent.

Extraction of Petroleum residues in Petroleum degradation Medium Using n-hexane, by GC-MS ^[3],[4] For C remaining in the experimental group (group with added microbial agent) and the blank group (group without added microbial agent) ₁₀ -C ₄₀ Is analyzed for aliphatic hydrocarbons. Measured according to national standard "gas chromatography for determination of petroleum hydrocarbons (C10-C40) in HJ 1021-2019 soil and sediments". The results are shown in FIG. 6, and GC-MS analysis peak shows the strain pair C ₁₀ -C ₄₀ Has good degradation effect on aliphatic hydrocarbon, C ₁₀ -C ₄₀ The degradation rates of (a) were 79.37% and 72.94%, respectively.

Example 2 shows that the Gordonia C1 provided by the invention has strong petroleum degradation capability in a Biodegradation (BO) culture medium and has good application prospect in petroleum microorganism ecological restoration.

Examples 3 to 6: evaluation of Petroleum degradation Performance of two Petroleum degrading bacteria under different culture conditions

The results of the different culture conditions were set as described in example 2, inoculated into a petroleum degradation medium at 15% of the inoculum size, and the petroleum degradation performance of the single strain was evaluated by a gravimetric method after 24 days of culture, and the results are shown in Table 4.

TABLE 4 Total Petroleum Hydrocarbon degradation Rate under different culture conditions

Under different culture conditions, the petroleum hydrocarbon degradation rate of two strains of petroleum degrading bacteria is above 60%, which indicates that the single strain provided by the invention has flexible culture application conditions.

Example 7: evaluation of petroleum degradation performance of composite bacterial liquid

The Gordonia C1 (Gordonia sp. Strain C1) and the fiber obtained in example 1 were activated, respectivelyMicrobacterium C3 (Cellulomictiobium sp.Strain C3), washing and modulating the cell suspension to OD ₆₀₀ =1, the gordonia C1 and the cellomyces C3 were synchronously mixed at a volume ratio of 60% and 40%, respectively, to prepare a composite bacterial solution.

Inoculating the composite bacterial liquid into a modified BO culture medium with petroleum mass concentration of 80000mg/L at 15%, culturing at 37deg.C and 180-200rpm for 24 days with a blank of modified BO culture medium without bacterial liquid with petroleum mass concentration of 80000mg/L, and performing destructive sampling on day 0, day 8, day 16 and day 24 respectively ^[3] 。

The total petroleum hydrocarbon degradation rate of the composite bacterial liquid on the 8 th day, the 16 th day and the 24 th day is measured by a weight method. As a result, as shown in FIG. 7, the total petroleum hydrocarbon degradation rate was 48.43% on the 8 th day, 69.36% on the 16 th day and 85.35% on the 24 th day. The results show that the composite flora has higher petroleum degradation capability and higher degradation capability than that of a single strain. The cell surface hydrophobicity of the Gordonia C1 is obviously higher than that of the Cellularomyces C3, which is favorable for adhering cells to the surface of petroleum hydrocarbon and degrading the petroleum hydrocarbon, has stronger petroleum pollution restoration capability, and has more obvious degradation effect when the duty ratio is increased; the growth amount of the fiber microzyme C3 is relatively higher than that of the Gordonia C1, and the degradation of partial alkyl substances is promoted.

Examples 8 to 9: influence of single-bacterium proportion in composite bacterial liquid on petroleum degradation rate

The different volume fractions of the single bacteria in the composite bacterial liquid were set as described in example 7, and the capacity of degrading petroleum was evaluated by gravimetric methods on days 8, 16 and 24 of cultivation, and the results are shown in Table 5.

TABLE 5 Total Petroleum Hydrocarbon degradation Rate at different Mono-bacterial ratios

The different volume ratios of the single bacteria in the composite flora have little influence on the final petroleum degradation performance, the total petroleum hydrocarbon degradation rate of the composite flora is above 60 percent, and the highest total petroleum hydrocarbon degradation rate is 86.52 percent, which indicates that the composite flora provided by the invention has flexible application mode. The main functional degradation bacteria are Gordonia C1, and the growth amount of the fiber micro-bacteria C3 is faster, so that degradation is promoted.

Examples 10 to 11: capability of composite bacterial liquid for treating different petroleum concentrations

The degradation performance of the composite bacterial liquid on different petroleum concentrations was evaluated by a gravimetric method on the 8 th, 16 th and 24 th days of cultivation as described in example 7, and the results are shown in Table 6.

TABLE 6 Total Petroleum Hydrocarbon degradation Rate at different Petroleum concentrations

The total petroleum hydrocarbon degradation rate of the compound bacteria in the improved BO culture medium added with petroleum (the petroleum concentration is 60000-100000 mg/L) is 77.02% -86% 15% after 24 days, which shows that the compound bacteria provided by the invention has higher degradation rate, high petroleum pollution treatment concentration and wider application range.

The foregoing description of the embodiments has been provided for the purpose of illustrating the general principles of the invention, and is not meant to limit the invention thereto, but to limit the invention thereto, and any modifications, equivalents, improvements and equivalents may be made thereto without departing from the spirit and principles of the invention.

Reference is made to:

[1] huang Xiangfeng, fangzheng, huang Wei, peng Kaiming, liu Lijun, liu Jia. MATH method characterizes the progress of investigation of the hydrophobicity of the cell surface of environmental microorganisms [ J ]. Microbiological bulletins, 2015,42 (01): 200-206;

[2] packing, huang Lixin, jianlong, ilina, development of oil and gas field oil-containing sludge environmental protection treatment technology [ J ]. Application chemical industry, 2021,50 (09): 2602-2608;

[3]Geng Pengxue,Ma Anzhou,Wei Xiaoxia,Chen Xianke,Yin Jun,Hu Futang,Zhuang Xuliang,Song Maoyong,Zhuang Guoqiang.Interaction and spatio-taxonomic patterns of the soil microbiome around oil production wells impacted by petroleum hydrocarbons.[J].Environmental pollution(Barking,Essex:1987),2022,307；

[4] guo Yan, ma Jian, yang Zongzheng, zhang Tian Yu, sun, wu Zhiguo. Screening and identification of Petroleum hydrocarbon degrading bacteria Mycolicibacterium fluoranthenivorans Y3 and degradation Property research [ J ]. Environmental science journal, 2022,41 (05): 1-7.

Claims (10)

1. A flora, characterized in that the flora comprises Gordonia (Gordonia sp.) C1 with a collection number of CGMCC No.25708, cellomicrozyme (cellomium sp.) C3 with a collection number of CGMCC No.25709, or both.

2. The flora according to claim 1, characterized in that it consists of 50-70% by volume of Gordonia sp C1 and 30-50% by volume of cellomicrozyme sp C3.

3. A method of culturing a bacterial population according to claim 1 or 2, characterized in that the culture medium used for the culture has the following composition formula: CH (CH) ₃ COONa 1g/L，C ₃ H ₃ NaO ₃ 0.5g/L，K ₂ HPO ₄ 0.5g/L，KNO ₃ 0.1g/L，NH ₄ Cl 0.05g/L，NaCl 25g/L，FeC ₆ H ₅ O ₇ 0.002g/L, natural pH; the culture conditions are as follows: 26-37 deg.c and 150-200rpm.

4. Use of the flora according to claim 1 or 2 in the field of petroleum pollution environmental remediation.

5. The use of claim 4, wherein said petroleum-contaminated environment comprises petroleum-contaminated soil and petroleum-contaminated water.

6. A method for the microecological remediation of petroleum pollution, characterized in that the flora of claim 1 or 2 is used to degrade petroleum hydrocarbons in petroleum contaminated soil or water.

7. A method for degrading petroleum hydrocarbon, characterized in that the method is to inoculate the flora in claim 1 or 2 into a degradation system containing petroleum with mass concentration of 60000-100000mg/L, and culture at 26-37 ℃ and 150-200rpm.

8. The method of claim 7, wherein the nutrient component in the degradation system is CH ₃ COONa 0.3g/L，C ₃ H ₃ NaO ₃ 0.1g/L，K ₂ HPO ₄ 1g/L，KNO ₃ 2g/L，NH ₄ Cl 0.5g/L，NaCl 10g/L，FeC ₆ H ₅ O ₇ 0.005g/L。

9. An article of manufacture for the remediation of petroleum-contaminated environments, said article comprising the flora of claim 1 or 2.

10. The article of claim 9, wherein the article is a liquid microbial inoculant article.

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