CN113215025A - Oil-containing sludge degrading strain Bacillus velezensis D3 and application thereof - Google Patents
Oil-containing sludge degrading strain Bacillus velezensis D3 and application thereof Download PDFInfo
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Abstract
The invention provides an oil-containing sludge degrading strain Bacillus velezensis D3 and application thereof, wherein the strain is preserved in China general microbiological culture collection center in 2021 year, 1 month and 25 days, and the preservation number is CGMCC NO. 21728; the strain is separated from the oily sludge of an oil production plant, can be applied to microbial remediation of the oily sludge, can quickly become a dominant flora after entering the oily sludge, effectively promotes the degradation of petroleum hydrocarbons, optimizes a soil microbial system, improves the physical properties of soil, enhances the biological activity of the soil, and has a degradation rate of Total Petroleum Hydrocarbons (TPH) in the oily sludge of more than 92.21%.
Description
Technical Field
The invention relates to the technical field of biochemical treatment of oily sludge; in particular to an oil-containing sludge degrading strain Bacillus velezensis D3 and application thereof.
Background
The oily sludge is a solid waste which is generated in the processes of petroleum exploitation, oil and gas gathering and transportation, refining and processing and oily sewage treatment and consists of crude oil/water/heavy metal and sludge. The content of petroleum hydrocarbon (TPH) in oil sludge in China is generally 5-50%, the content of solid particles is 5-46%, the water content is generally 35-90%, even higher, the pH value is generally 6.5-7.5, and the oil sludge also contains a large amount of colloid, asphaltene, benzene series, anthracene, phenol, pyrene and other toxic and harmful substances, even radioactive substances and heavy metals, wherein a plurality of substances have a 'three-cause' effect (such as Polycyclic Aromatic Hydrocarbons (PAHs)). Oily sludge is listed in national hazardous waste list (2018) (HW08 waste mineral oil and mineral oil-containing waste) due to toxicity and inflammability, and can cause serious threats to ecological environment and human health if not properly treated.
The principle of the biological method for treating the oily sludge is as follows: petroleum hydrocarbon is used as carbon source for microbial metabolism, harmful substances in the oily sludge are assimilated and degraded by utilizing the metabolic activity of the microbes, and finally the harmful substances are completely converted into harmless CO2And H2O and the like, and realizes the harmless treatment of the oily sludge.
Bacillus velezensis was first isolated from soil samples from the river mouth of Belgium, Macla, province, Spain. The bacillus is a spore-forming gram-positive bacterium, and most of the bacillus has the advantages of wide antibacterial spectrum, rapid growth, easy separation and culture, strong stress resistance, high biological safety and the like, so that the bacillus is widely researched as a probiotic in the aspects of agriculture, food, industry, medicine, metallurgy, forestry, environmental protection, military and the like, but the application research of the bacillus in the aspect of oil-containing sludge remediation is not reported.
Disclosure of Invention
The invention aims to provide an oily sludge degrading strain Bacillus velezensis D3 and application thereof.
In a first aspect, the invention is realized by the following technical scheme:
an oil-containing sludge degrading strain Bacillus velezensis D3 (Bacillus belgii), which is preserved in China general microbiological culture collection center in 1 month and 25 days 2021 year, with the preservation number of CGMCC NO. 21728.
Further, the strain is isolated from oil-containing sludge of an oil production plant.
In one embodiment of the present application, the strain is isolated from the oily sludge of a guard edge oil extraction plant in an extended oil field.
In a second aspect, the present invention also relates to a method for separating an oily sludge degrading strain from an oily sludge of an oil production plant, comprising the steps of:
s1, collecting an oil-containing sludge sample from an oil production plant;
s2, placing the oily sludge sample into a conical flask filled with a broth culture medium, shaking the conical flask on a constant-temperature shaking table with the temperature of 30 ℃ and the rpm of 120, and carrying out enrichment culture for 48 hours; preparation of 10 by gradient dilution-2、10-3、10-4、10-5、10-6Serial dilutions, respectively coating 150 μ L of dilutions on nutrient agar culture medium; culturing the inoculated plate for 48h at 30 ℃ under the aerobic condition of 180 r/min; selecting bacterial colonies with rapid growth and large diameter;
s3, adding 2 wt% of crude oil into the inorganic salt culture medium, and carrying out autoclaving at 121 ℃ for 30 min. Activating the strain selected in S2, inoculating into sterilized oil-containing inorganic salt culture medium, controlling the temperature at 30 ℃ under aerobic condition, and continuously culturing for 2 weeks at the rotating speed of a shaking table of 180 r/min; measuring and recording the growth, oil content and emulsification condition of bacteria in the culture process; and finally, selecting bacteria with good emulsification and degradation effects on crude oil, inoculating the bacteria on an LB solid culture medium by adopting a plate streaking separation method, placing the bacteria in a constant-temperature incubator for inverted culture at 30 ℃ for 24h, picking out single colonies after the colonies grow out, repeating the streaking separation process until purified colonies with single morphology are formed, and inoculating the strains on a storage culture medium inclined plane to be stored in a refrigerator at 4 ℃ for later use.
Further, the 16S rRNA gene sequences of strains isolated from oil-containing sludge from oil production plants were determined by analytical biology methods such as Seq ID No: 1. the inventors used MEGA 4.1 software to compare Seq ID No: 1 gene sequence, finding that the strain belongs to Bacillus belgii, and is named as Bacillus velezensis D3.
The 16S rRNA gene sequence of the strain Bacillus velezensis D3 is shown as Seq ID No: 1.
preferably, the physicochemical properties of the oily sludge sample are analyzed before the experiment in S1.
Preferably, prior to starting the experiment in S1, a sample of the oily sludge is crushed, passed through a 2mm sieve and refrigerated at 4 ℃.
Preferably, the components of the broth culture in S2 include peptone 10.0g/L, beef extract 5.0g/L and sodium chloride 5.0g/L, pH 7.2-7.4.
Preferably, the components of the mineral salts medium in S3 include crude oil 20g/L, K2HPO4 5.0g/L、MgSO40.25 g/L、NH4NO32.0 g/L、NaCl 5.0g/L、KH2PO41.0 g/L、MgSO4·7H2O2.0 g/L and CaCl2·2H2O 1.0g/L，pH 7.0-7.2。
The optimal growth conditions of the strain Bacillus velezensis D3 are as follows: the temperature is 30-35 ℃, the salt tolerance is lower than 6 percent, and the pH value is 6-9.
In a third aspect, the invention also relates to an application of the strain Bacillus velezensis D3 in degrading oily sludge, and the technical scheme is as follows:
the bacterial strain Bacillus velezensis D3 is applied to degradation of oily sludge, bacterial liquid is inoculated into the oily sludge in an inoculation amount of 5-15 wt%, in order to increase air permeability of the oily sludge in the fermentation process, 5-10 wt% of sawdust is added into the oily sludge, in addition, a nitrogen source and a phosphorus source are supplemented according to the petroleum hydrocarbon content in the oily sludge, so that C, N, P is 90:5:1, the water content is controlled to be 20-40 wt%, the temperature is 15-45 ℃, and the pH is 5.0-10.0; and after uniformly stirring, stacking and fermenting, wherein the stacking thickness is 50-80 cm, stirring and supplementing water every 3 days, and the treatment period is 60-90 days.
Preferably, the water content of the oily sludge is controlled to be 30%, the temperature is controlled to be 35 ℃, and the pH value is controlled to be 8.0.
Further, the nitrogen source is any one or a combination of several of ammonium chloride, urea and ammonium nitrate, and the phosphorus source is any one or a combination of several of potassium hydrogen phosphate, potassium dihydrogen phosphate, sodium dihydrogen phosphate, dipotassium hydrogen phosphate and sodium hydrogen phosphate.
Further, in the application process of the strain Bacillus velezensis D3 in degrading the oily sludge, sampling is carried out once every 10 days, and the content of Total Petroleum Hydrocarbon (TPH) in the oily sludge is determined by adopting an infrared spectrophotometer.
The invention has the following advantages:
(1) the invention provides a high-efficiency petroleum hydrocarbon degrading bacterium Bacillus velezensis D3 separated from oily sludge, which can be applied to the microbial remediation of the oily sludge;
(2) the Bacillus velezensis D3 separated by the method is derived from oil-containing sludge, so that the strain becomes a dominant flora soon after entering the oil-containing sludge, the degradation of petroleum hydrocarbons is effectively promoted, a soil microbial system is optimized, the physical properties of soil are improved, and the biological activity of the soil is enhanced;
(3) the Bacillus velezensis separated by the method is an advantageous strain screened by taking crude oil as a unique carbon source, and the strain can metabolize rhamnolipid in the growth and propagation process, so that the property of petroleum hydrocarbon substances in the oily sludge is effectively changed, and the degradation rate of microorganisms on the petroleum hydrocarbon substances in the oily sludge is increased;
(4) the Bacillus velezensis D3 obtained by separation has high degradation efficiency on petroleum hydrocarbon, can degrade petroleum pollutants in the oily sludge in a short time, and has the degradation rate of Total Petroleum Hydrocarbon (TPH) in the oily sludge of more than 92.21 percent within 2 weeks.
Drawings
FIG. 1 is a colony morphology diagram of the strain Bacillus velezensis D3;
FIGS. 2 and 3 are scanning electron micrographs at different magnifications of the strain Bacillus velezensis D3, respectively;
FIG. 4 is a 16S rRNA phylogenetic tree of strain Bacillus velezensis D3;
FIG. 5 is a graph of the emulsification effect of a strain Bacillus velezensis D3 on crude oil in a fermentation process with glucose and crude oil as the sole carbon sources (A-glucose as a carbon source; B-crude oil as a carbon source; C-a control experiment);
FIG. 6 gas chromatograms of crude oil before and after treatment with the strain Bacillus velezensis D3 (D-before microbial treatment; B-after microbial treatment);
FIG. 7 degradation kinetics curves of strain Bacillus velezensis D3 for crude oil;
FIG. 8 the degradation effect of Bacillus velezensis D3 on extended oil field oily sludge at different inoculum sizes;
FIG. 9 the degradation effect of Bacillus velezensis D3 on extended oil field oily sludge at different temperatures;
FIG. 10 the degradation effect of Bacillus velezensis D3 on extended oil field oily sludge under different water content.
Detailed Description
The present invention will be described in detail with reference to specific examples. It should be noted that the following examples are only illustrative of the present invention, but the scope of the present invention is not limited to the following examples.
Example 1
This example provides an oily sludge degrading strain Bacillus velezensis D3 (Bacillus belgii) and a method for isolating the strain from oily sludge.
An oil-containing sludge degrading strain Bacillus velezensis D3 (Bacillus belgii), which is preserved in China general microbiological culture collection center in 1 month and 25 days 2021 year, with the preservation number of CGMCC NO. 21728.
In one embodiment of the present application, the strain is isolated from the oily sludge of a guard edge oil extraction plant in an extended oil field.
The method for separating the oily sludge degrading strain from the oily sludge of the oil production plant comprises the following steps:
s1, collecting an oily sludge sample from a Jing-edge oil extraction plant of the extension oil field by adopting a plastic container with the capacity of 1L, and analyzing the physical and chemical properties of the oily sludge sample before an experiment. Before starting the experiment, a sample of oily sludge was crushed, passed through a 2mm sieve and refrigerated at 4 ℃.
S2, weighing 10g of oily sludge sample, putting the oily sludge sample into a conical flask filled with 100mL of broth culture medium, shaking the mixture on a constant temperature shaking table at 30 ℃ and 120rpm, and carrying out enrichment culture for 48 h. The components of the broth culture medium are as follows: 10.0g/L of peptone, 5.0g/L of beef extract, 5.0g/L of sodium chloride, pH7.2-7.4, and sterilizing at 121 ℃ for 20 minutes. Preparation of 10 by gradient dilution-2、 10-3、10-4、10-5、10-6Serial dilutions were made by plating 150. mu.L of each dilution on nutrient agar media. The inoculated plates were incubated aerobically at 30 ℃ and 180 r/min. Bacterial colonies with rapid growth and large diameter were selected.
S3, adding 2 wt% of crude oil into an inorganic salt culture medium, and carrying out high-pressure sterilization at 121 ℃ for 30 min; the selected strains are activated and inoculated into 200mL of inorganic salt culture medium (containing crude oil 20g/L, K)2HPO4 5.0g/L、MgSO4 0.25g/L、NH4NO3 2.0g/L、 NaCl 5.0g/L、KH2PO4 1.0g/L、MgSO4·7H2O 2.0g/L、CaCl2·2H2O1.0 g/L, pH 7.0.0-7.2, autoclaving at 121 ℃ for 30min) in a 500mL conical flask, and continuously culturing for 2 weeks at 30 ℃ under aerobic conditions and 180r/min of shaking table rotation speed. Bacterial growth, oil content and emulsification were measured and recorded during the culture. Finally, selecting bacteria with good emulsification and degradation effects on crude oil, inoculating the bacteria on an LB solid culture medium by adopting a plate streaking separation method, placing the bacteria in a constant-temperature incubator for inverted culture at 30 ℃ for 24h, picking out a single colony after the colony grows out, repeating the streaking separation process until a purified colony with a single shape is formed, and inoculating the purified colony on a storage culture medium inclined plane to be stored in a refrigerator at 4 ℃ for later use.
The bacterial strain is gram-negative bacilli, and the bacterial colony on the TSA culture medium is light yellow, irregular in shape, high-convex in total section, smooth and opaque, and regular in edge, as shown in figure 1. The thallus is rod-shaped with different lengths, and the shape of the bacterium is shown in a scanning electron microscope picture 2 and a picture 3.
The 16S rRNA gene sequence of the strain separated from the oil-containing sludge of the oil production plant is determined by adopting an analytical biology method, such as Seq ID No: 1, the phylogenetic tree of 16S rRNA and its relatives constructed by the neighbor joining method was retrieved from GenBank database as shown in FIG. 4. The related sequences were aligned using MEGA 4.1 software and the strain was found to belong to Bacillus belgii, named Bacillus velezensis D3.
The strain Bacillus velezensis D3 can rapidly grow and propagate no matter glucose or crude oil is used as a unique carbon source, a large amount of biosurfactant rhamnolipid can be metabolized, fermentation liquor has a good emulsification effect on crude oil, and complete dispersion and emulsification can be realized on 5% of added crude oil within 8 hours, as shown in figure 5.
The strain Bacillus velezensis D3 has high degradation efficiency on petroleum hydrocarbon in the oily sludge, and the gas chromatographic analysis results of crude oil components before and after microbial degradation are shown in figure 6.
Example 2
Bacillus velezensis D3 is used for the degradation repair test of the oil-containing sludge of the prolonged oil field.
In a third aspect, the invention also relates to an application of the strain Bacillusvelezensiss D3 in degrading oily sludge, and the technical scheme is as follows:
taking the oil-containing sludge of the extended oil field, air-drying, grinding, and sieving by a 2mm sieve for later use. The strain Bacillus velezensiss D3 was inoculated into a broth culture medium and the Erlenmeyer flask was activated for 24h at 30 ℃ at 120rpm in a shaker. Weighing 20kg of the oil-containing sludge subjected to air drying, grinding and sieving, inoculating a bacterial solution into the oil-containing sludge in an inoculation amount of 5-15 wt%, adding 1kg of sawdust into the oil-containing sludge in order to increase air permeability in a fermentation process of the oil-containing sludge, adding 200g of ammonium chloride and 45g of potassium dihydrogen phosphate at the same time to ensure that C: N: P is 90:5:1, controlling the water content to be 20-40%, controlling the temperature to be 20-40 ℃, stacking and fermenting after uniformly stirring, wherein the stacking thickness is 50-80 cm, stirring and supplementing water every 3 days, and the treatment period is 60 days.
During the microbial treatmentSampling every 10 days, and measuring the content of Total Petroleum Hydrocarbon (TPH) in the oily sludge and the content of crude petroleum hydrocarbon C in the oily sludge by using an infrared spectrophotometer10-C40The determination of the component content is based on the environmental protection standard HJ 1021 and 2019 < petroleum hydrocarbon (C) of soil and sediment10-C40) Measurement of gas chromatography (CRC) & gt. The gas chromatographic analysis conditions were: a sample inlet: the temperature is 300 ℃; and (3) sample introduction mode: no shunt sampling; column temperature: maintaining the initial temperature at 50 deg.C for 2min, increasing to 230 deg.C at 40 deg.C per minute, and increasing to 320 deg.C at 20 deg.C per minute for 20 min; gas flow rate: 1.5mL/min of high-purity helium, 30mL/min of hydrogen and 300mL/min of air; detector temperature: 325 ℃; sample introduction amount: 1 μ L. In addition, the method requires the use of n-decanone and n-forty alkane to determine the retention window time, using C10-C40The standard solution establishes a standard curve.
The rate of decomposition of petroleum hydrocarbon (TPH) in the oily sludge by Bacillus velezensis D3 was analyzed by an infrared oil analyzer, and the results are shown in FIG. 7.
As can be seen from figure 7, the TPH degradation rate gradually increases with the extension of the degradation time, and reaches 78.01% -86.42% by 60 days, which indicates that the strain has good bioremediation effect on the actual oily sludge.
Example 3
Degradation effect of Bacillus velezensis D3 on prolonging oil-containing sludge of oil field under different inoculation amounts
Taking the oil-containing sludge of the extended oil field, air-drying, grinding, and sieving by a 2mm sieve for later use. The strain Bacillus velezensis D3 was inoculated into a broth culture medium and the Erlenmeyer flask was activated for 24h at 30 ℃ at 120rpm in a shaker. Weighing 20kg of the oil-containing sludge subjected to air drying, grinding and sieving, respectively inoculating the oil-containing sludge into the oil-containing sludge according to the inoculation amounts of 5%, 7.5%, 10%, 12.5% and 15% by mass, simultaneously adding 1kg of sawdust, 200g of ammonium chloride and 45g of potassium dihydrogen phosphate, controlling the water content to be 30%, controlling the temperature to be 30 ℃, uniformly stirring, stacking and fermenting, wherein the stacking thickness is 50-80 cm, stirring and supplementing water every 3 days, and analyzing the TPH content in the oil-containing sludge after 60-day biodegradation treatment, wherein the result is shown in figure 8, and the result is shown in figure 8.
Example 4
Bacillus velezensis D3 for prolonging degradation effect of oil-containing sludge of oil field at different temperatures
Taking the oil-containing sludge of the extended oil field, air-drying, grinding, and sieving by a 2mm sieve for later use. The strain Bacillus velezensis D3 was inoculated into a broth culture medium and the Erlenmeyer flask was activated for 24h at 30 ℃ at 120rpm in a shaker. Weighing 20kg of the oil-containing sludge subjected to air drying, grinding and sieving, inoculating the oil-containing sludge into the oil-containing sludge according to the inoculation amount of 10 wt%, simultaneously adding 1kg of sawdust, 200g of ammonium chloride and 45g of monopotassium phosphate, controlling the water content to be 30 wt%, uniformly stirring, then stacking and fermenting, wherein the stacking thickness is 50-80 cm, the fermentation temperature is controlled to be 20 ℃, 25 ℃, 30 ℃, 35 ℃ and 40 ℃, stirring and supplementing water every 3 days, and carrying out biodegradation treatment for 60 days to analyze TPH in the oil-containing sludge, wherein the result is shown in figure 9, and the result is shown in figure 9.
Example 5
Degradation effect of Bacillus velezensis D3 on prolonging oil-containing sludge of oil field under different water content conditions
Taking the oil-containing sludge of the extended oil field, air-drying, grinding and sieving by a 2mm sieve for later use. The strain Bacillus velezensis D3 was inoculated into a broth culture medium and the Erlenmeyer flask was activated for 24h at 30 ℃ at 120rpm in a shaker. Weighing 20kg of the oil-containing sludge subjected to air drying, grinding and sieving, inoculating the oil-containing sludge into the oil-containing sludge according to the inoculation amount of 10 wt%, simultaneously adding 1kg of sawdust, 200g of ammonium chloride and 45g of monopotassium phosphate, respectively controlling the water content to be 20%, 25%, 30%, 35% and 40%, uniformly stirring, then stacking and fermenting, wherein the stacking thickness is 50-80 cm, controlling the fermentation temperature to be 35 ℃, stirring and replenishing water every 3 days, and carrying out biodegradation treatment for 60 days to analyze TPH in the oil-containing sludge, wherein the result is shown in figure 10, and the strain Bacillus velezensis D3 has a certain removal effect on TPH in the oil-containing sludge within the range of 20% -40% of the water content, wherein the effect is optimal when the water content is 30%.
The foregoing description of specific embodiments of the present invention has been presented. It is to be understood that the present invention is not limited to the specific embodiments described above, and that various changes or modifications may be made by one skilled in the art within the scope of the appended claims without departing from the spirit of the invention.
Sequence listing
<110> Xian Petroleum university
<120> oil-containing sludge degrading strain Proteus mirabilis SB and application thereof
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<170> SIPOSequenceListing 1.0
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Claims (10)
1. An oil-containing sludge degrading strain Bacillus velezensis D3 is characterized in that the strain is preserved in China general microbiological culture collection center in 2021 year, 1 month and 25 days, and the preservation number is CGMCC NO. 21728.
2. The oily sludge degrading strain Bacillus velezensis D3 as claimed in claim 1, wherein the strain is isolated from an oily sludge of a production plant.
3. A method for separating oily sludge degrading strains from oily sludge in an oil production plant is characterized by comprising the following steps:
s1, collecting an oil-containing sludge sample from an oil production plant;
s2, placing the oily sludge sample into a conical flask filled with a broth culture medium, shaking the conical flask on a constant-temperature shaking table with the temperature of 30 ℃ and the rpm of 120, and carrying out enrichment culture for 48 hours; preparation of 10 by gradient dilution-2、10-3、10-4、10-5、10-6Serial dilutions, respectively coating 150 μ L of dilutions on nutrient agar culture medium; culturing the inoculated plate at 30 ℃ under the aerobic condition of 180 r/min; selecting and purifying bacterial colonies with rapid growth and large diameter;
s3, adding 2 wt% of crude oil into an inorganic salt culture medium, and carrying out high-pressure sterilization at 121 ℃ for 30 min; activating the strain selected in S2, inoculating into sterilized oil-containing inorganic salt culture medium, controlling the temperature at 30 deg.C under aerobic condition, and culturing for 2 weeks at shaker rotation speed of 180 r/min; measuring and recording the growth, oil content and emulsification condition of bacteria in the culture process; and finally, selecting bacteria with good emulsification and degradation effects on crude oil, inoculating the bacteria on an LB solid culture medium by adopting a plate streaking separation method, placing the bacteria in a constant-temperature incubator for inverted culture at 30 ℃ for 24h, picking out single colonies after the colonies grow out, repeating the streaking separation process until purified colonies with single morphology are formed, and inoculating the strains on a storage culture medium inclined plane to be stored in a refrigerator at 4 ℃ for later use.
4. The strain Bacillus velezensis D3 according to any one of claims 1 to 3, wherein the strain has a 16S rRNA gene sequence as shown in Seq ID No: 1.
5. the method for separating the oily sludge degrading strain from the oily sludge of the oil production plant as claimed in claim 3, wherein the physicochemical properties of the oily sludge sample are analyzed before the experiment in S1; before starting the experiment in S1, a sample of oily sludge was crushed, passed through a 2mm sieve and refrigerated at 4 ℃.
6. The method of claim 3, wherein the components of the broth culture of S2 include peptone 10.0g/L, beef extract 5.0g/L and sodium chloride 5.0g/L, and pH 7.2-7.4.
7. The method of claim 3, wherein the mineral salts medium of S3 comprises crude oil 20g/L, K g2HPO4 5.0g/L、MgSO4 0.25g/L、NH4NO32.0g/L、NaCl 5.0g/L、KH2PO4 1.0g/L、MgSO4·7H2O2.0 g/L and CaCl2·2H2O 1.0g/L，pH 7.0-7.2。
8. The application of the strain Bacillus velezensis D3 in degrading oily sludge according to any one of claims 1-3, wherein the strain liquid is inoculated into the oily sludge in an inoculation amount of 5-15 wt%, 5-10 wt% of sawdust is added into the oily sludge at the same time, and then a nitrogen source and a phosphorus source are supplemented according to the content of petroleum hydrocarbon in the oily sludge, so that the C, N, P is 90:5:1, the water content is controlled to be 20-40 wt%, the temperature is 15-45 ℃, and the pH is 5.0-10.0; and after uniformly stirring, stacking and fermenting, wherein the stacking thickness is 50-80 cm, stirring and supplementing water every 3 days, and the treatment period is 60-90 days.
9. The application of the strain Bacillus velezensis D3 in degrading oily sludge according to claim 7, wherein the moisture content of the oily sludge is controlled to be 30%, the temperature is 35 ℃, and the pH is 8.0.
10. The application of the strain Bacillus velezensis D3 in degrading oily sludge according to claim 7, wherein the nitrogen source is any one or combination of ammonium chloride, urea and ammonium nitrate, and the phosphorus source is any one or combination of potassium hydrogen phosphate, potassium dihydrogen phosphate, sodium dihydrogen phosphate, dipotassium hydrogen phosphate and sodium hydrogen phosphate.
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| CN108034626A (en) * | 2018-02-06 | 2018-05-15 | 北京大学 | A kind of degradation bacteria strains JN1 of oily sludge petrochina hydro carbons and its application |
| CN112094775A (en) * | 2020-09-22 | 2020-12-18 | 中国石油天然气集团有限公司 | Bacillus belgii and screening culture method and application thereof |
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| CN108034626A (en) * | 2018-02-06 | 2018-05-15 | 北京大学 | A kind of degradation bacteria strains JN1 of oily sludge petrochina hydro carbons and its application |
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