CN111621437A - Otter escherichia coli LM-DK separated from oxidation pond of pig farm and application thereof - Google Patents
Otter escherichia coli LM-DK separated from oxidation pond of pig farm and application thereof Download PDFInfo
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Abstract
The invention discloses woodchuck Escherichia (Escherichia marmotae) LM-DK separated from bottom mud of a biogas slurry oxidation pond, which is the woodchuck Escherichia marmotae with the preservation number: CGMCC No. 18769. The invention also discloses the application of the woodchuck Escherichia (Escherichia marmotae) LM-DK: removing CODcr, BOD5, total phosphorus and ammonia nitrogen in the pig breeding wastewater.
Description
Technical Field
The invention relates to an escherichia woodchuck (Escherichia coli) LM-DK separated from bottom mud of a biogas slurry oxidation pond in a pig farm and application thereof in culture wastewater treatment.
Background
Pig farm culture water is one of important agricultural pollution sources influencing the quality of water environment in China, the existing wastewater treatment technology mainly comprises planting and breeding combination, standard discharge, an ectopic fermentation bed and the like, but the treatment technologies have certain defects, so that the wastewater treatment is not in place, the unqualified culture wastewater enters a water body frequently, the surface water and underground water quality of an area are influenced, and the bottleneck is formed for the development of the pig culture industry in order to protect the water environment, shut down the pig farm in a large range, establish a restricted culture area and the like in all parts of the country.
The livestock and poultry breeding wastewater has high pollutant concentration and simultaneously contains a large amount of pathogenic microorganisms, parasitic ova and residual antibiotics and heavy metal elements. The indexes of main pollutants in the livestock and poultry breeding wastewater such as suspended matter concentration, CODcr value and BOD5The value, the organic phosphorus content, the ammonia nitrogen content and the like are high, so that the sewage degradation treatment difficulty is high, in addition, the livestock and poultry breeding areas are dispersed in suburbs and vast rural areas, so that the livestock and poultry breeding wastewater is difficult to transport to sewage plants for treatment through transportation or pipelines, if the livestock and poultry breeding wastewater cannot be properly treated in the breeding areas and is discharged to the natural environment, the livestock and poultry breeding wastewater can cause serious harm to the environment such as water, soil and the like, the natural ecological balance is damaged, and the health of people and livestock is greatly threatened. Therefore, finding a method and a technology for efficiently treating livestock and poultry breeding wastewater becomes a current research hotspot, which is an urgent technical demand for breeding enterprises.
At present, the main methods for treating the livestock and poultry breeding wastewater pollution comprise a returning method, a natural treatment method and a biological treatment method. The biological treatment method is divided into an anaerobic treatment method, an aerobic treatment method and an anaerobic-aerobic combined method, the biological treatment method is an effective treatment method of wastewater with low cost and no secondary pollution, which is widely concerned by researchers, and pollutants in the wastewater are decomposed and converted by using microorganisms to eliminate potential harm of the pollutants in the wastewater to the environment.
At present, no report is found at home and abroad on the research of the application of the woodchuck Escherichia bacteria to sewage treatment, only a small amount of other Escherichia bacteria (and close genera) are reported to be used for sewage treatment, and are mainly used for denitrification, and the research shows that the other Escherichia bacteria (and close genera) mainly show stronger denitrification (Geraniin, Chensi, Liuna, Sichuan institute of technology and institute of technology (Nature science edition), 2013, 26 (4): 10-13; Dongliang, Pengying, environmental science bulletin, 2017, 37(9): 3349-3355; Ferguson SJ,1994, Antonie Van Leeuwenhoek,66(1/3): 89-110; HarnMF, HuS, ShiY, et al,2013, Nature,500(7464): 56570) which can perform denitrification on nitrate nitrogen in water bodies, remove nitrate nitrogen in water bodies, and is one of important biological micro-biological nitrogen for sewage denitrification, however, the other escherichia bacteria have single metabolism type on pollutants in sewage, focus on the denitrification of nitrogen, and have few research reports on the metabolism of other pollutants, and particularly, the research on the treatment of livestock and poultry breeding wastewater with high pollutant concentration and multiple varieties is not reported at home and abroad.
Disclosure of Invention
The invention aims to solve the technical problem of providing a strain-Escherichia coli (Escherichia marmotae) LM-DK capable of being used for livestock and poultry breeding wastewater treatment and application thereof.
In order to solve the technical problem, the invention provides an Escherichia coli (Escherichia marmotae) LM-DK separated from bottom mud of a biogas slurry oxidation pond in a pig farm, which is the Escherichia coli, and the preservation number is as follows: CGMCC No. 18769.
As an improvement of Escherichia marmotae (Escherichia marmotae) of the present invention: the 16S rDNA gene sequence of the strain is shown in SEQ ID NO. 1.
The invention also provides the application of the above-mentioned otter Escherichia coli (Escherichia marmotae): treating livestock and poultry breeding wastewater.
As an improvement of the use of the Escherichia woodchuck (Escherichia marmotae) of the present invention: removing CODcr and BOD in livestock and poultry breeding wastewater5Total phosphorus and ammonia nitrogen.
The preservation information of the strains of the invention is as follows:
the preservation name is: woodchuck Escherichia marmotae, collection unit: china general microbiological culture Collection center, preservation Address: the microbial research institute of the national academy of sciences No. 3, Xilu No.1, Beijing, Chaoyang, Beijing, with the preservation number: CGMCC No.18769, preservation date: 2019, 10 and 30.
The bacterial colony characteristics of the strain are as follows: the bacillus brevis is a bacillus brevis, has rough surface, pilus and no cytopenia, has small capsule and gram-positive bacteria, has the size of (0.9-1.2) Mumx (0.4-0.7) Mum, and has circular bacterial colony, convex and smooth surface, complete edge, beige color and opacity on a solid culture medium.
The 16S rDNA gene sequence of the strain is shown in SEQ ID NO. 1:
the strain is obtained by screening biogas slurry oxidation pond bottom sludge collected from a northwest live pig farm in Yushu district of Xinyu province, and according to a fatty acid map generated by qualitative and quantitative analysis of a Sherlock MIS software system on strain LM-DK, a Library database is compared, the strain LM-DK is preliminarily identified to be woodchuck Escherichia coli (Escherichia marmotae), and the similarity index SI (similarity index) is 0.987; the strain was also identified as Escherichia coli (Escherichia marmotae) by 16S rDNA sequencing, and the similarity index SI (similarity index) was 0.99.
Inoculating the strain of the invention into live pig breeding wastewater, and culturing at 15-35 ℃ for 150-200 r.min-1The culture is carried out for 2-3 d under the aerobic condition, and the CODcr and BOD in the wastewater can be effectively removed5Total phosphorus and ammonia nitrogen.
The livestock and poultry breeding wastewater is mainly live pig breeding wastewater, and the CODcr concentration of the livestock and poultry breeding wastewater is 8000-14000 mg.L-1、BOD5The concentration is 4000-8000 mg/L-1The total phosphorus concentration is 80-300 mg.L-1The concentration of ammonia nitrogen is 800-1500 mg.L-1。
The strain can be used for treating pig breeding wastewater, and the pH value is 6.8, the temperature is 25-30 ℃, and the OD is OD415nm0.2, the CODcr concentration in the pig breeding wastewater is 11000 mg.L-1、BOD5The concentration is 5500 mg.L-1The total phosphorus concentration was 246 mg.L-1The ammonia nitrogen concentration is 1300 mg.L-1At a rotation speed of 180 r.min-1The wastewater is cultured for 3 days and filtered, the removal rates of CODcr, BOD5, total phosphorus and ammonia nitrogen in the wastewater are respectively 98.13%, 98.54%, 98.17% and 97.24%, and the wastewater can reach the limit standard (the CODcr concentration is 400 mg.L) of the livestock and poultry breeding pollutant discharge standard (GB18596-2001)-1、BOD5The concentration is 150 mg.L-1The total phosphorus concentration was 8.0 mg.L-1The ammonia nitrogen concentration is 80 mg.L-1) And (5) discharging.
The invention discovers for the first time that the escherichia woodchuck can effectively treat the livestock and poultry breeding wastewater with high pollutant concentration and various types, and can remove CODcr, BOD5, total phosphorus and ammonia nitrogen in the livestock and poultry breeding wastewater in a metabolizing way, which is different from other reported escherichia bacteria, and fills up the blank of related research.
In conclusion, the otter Escherichia coli (Escherichia marmotae) LM-DK for treating the livestock and poultry breeding wastewater is obtained through screening, and has the potential of efficiently treating the livestock and poultry breeding wastewater; provides a technical basis for controlling the influence of the livestock and poultry breeding wastewater discharge on the environment, and has wide application prospect.
Drawings
The following describes embodiments of the present invention in further detail with reference to the accompanying drawings.
FIG. 1 is an Escherichia marmotae (Escherichia marmotae) LM-DK (magnification 20000 times);
FIG. 2 shows the result of 16S rDNA PCR amplification (DK: strain LM-DK; Marker: DNA standard molecular weight);
FIG. 3 is a strain LM-DK phylogenetic tree based on the 16S rDNA sequence.
Detailed Description
The invention will be further described with reference to specific examples, but the scope of the invention is not limited thereto:
example 1 screening and identification of degrading Strain
1 materials and methods
1.1 media and reagents
Enrichment culture medium: potassium nitrate (2, 5, 10, 20 or 30g, gradually increased and added for acclimatization and enrichment), CaCl2·6H2O0.1g，MgCl20.25g，K2HPO41.5g，NH4Cl 1g, peptone 9g, H2O 1000mL，pH＝6.5～7.0；
Basic culture medium: NH (NH)4NO31.00g，MgSO4·7H2O 0.5g，(NH4)2SO40.5g，KH2PO40.5g，NaCl0.5g，K2HPO41.5g，H2O 1000mL，pH4.0；
Adding 30g potassium nitrate into each 1000ml basal medium to obtain a medium containing 30 g.L-1A liquid basal medium of potassium nitrate;
adding 30g potassium nitrate and 15g agar into 1000ml basal medium to obtain culture medium containing 30 g.L-1A solid basal medium of potassium nitrate;
all media need to be sterilized at high temperature before use (conventional high temperature sterilization, generally sterilization at 1.1 atm and 121 ℃ for 20min), which is common knowledge.
The pH of the pig breeding wastewater is 6.8, and the CODcr concentration is 11000 mg.L-1、BOD5The concentration is 5500 mg.L-1The total phosphorus concentration was 246 mg.L-1The ammonia nitrogen concentration is 1300 mg.L-1The pig breeding wastewater.
1.2 determination of treatment efficiency of strain on pig-containing breeding wastewater
The single strain after purification is OD415nmInoculating to 100mL of pig-containing breeding wastewater (pH6.8, CODcr concentration of 11000 mg.L) at a concentration of 0.2-1、BOD5The concentration is 5500 mg.L-1The total phosphorus concentration was 246 mg.L-1The ammonia nitrogen concentration is 1300 mg.L-1) In a 250mL triangular flask, the mixed wastewater without inoculation is used as a reference, and the temperature is 30 ℃, and the time is 180 r.min-1The aerobic culture is carried out for 3 days on a constant temperature shaking table. After the culture is finished, filtering with 0.20 μm membrane to remove mycelium, and using filtrate for CODcr and BOD5And determination of total phosphorus and ammonia nitrogen (see: State environmental protection administration Water and wastewater monitoring analysis method committeeWater and wastewater monitoring and analysis method (fourth edition), 2002, environmental science press).
Removal rate (%) × 100 (control sample residual amount-treated sample residual amount) × 100/control sample residual amount.
1.3 dominant degradation Strain Breeding
1.3.1 sources of strains
Collecting biogas slurry oxidation pond sediment of a water north town live pig farm in Yu water areas of Xinyu province in Jiangxi province.
1.3.2 isolation, purification and screening of degrading strains
Taking 10g of bottom sludge from the biogas slurry oxidation pond of each pig farm, respectively adding the bottom sludge into a 250mL triangular flask filled with 100mL of sterile liquid enrichment medium under the sterile condition, and carrying out 180 r-min at 30 DEG C-1After culturing for 7 days on a shaking table, the cells were transferred to the next enrichment medium according to the inoculum size of 10% (potassium nitrate gradient is 2, 5, 10, 20 or 30 g.L in sequence)-1) And domesticating and culturing for 7 days under the same condition. By analogy, the potassium nitrate concentration is 30 g.L-1And (5) enriching culture solution corresponding to the culture medium.
The culture solution is transferred to a medium containing 30 g.L according to the inoculation amount of 10 percent-1Continuously culturing in liquid basic culture medium containing potassium nitrate for 7 days, and continuously inoculating for 2 times; taking 0.1mL of the obtained fermentation broth to a fermentation broth content of 30 g.L-1Repeatedly carrying out plate streaking separation and purification on a solid basic culture medium of potassium nitrate until a single bacterial colony is obtained by screening, inoculating the pure bacterial colony on a slope, and storing in a refrigerator at 4 ℃.
Propagating the purified series of single purified strains through a basal culture medium, and specifically comprises the following steps: inoculating one loop of the purified strain into 100mL of basal medium, and culturing at 30 deg.C for 180r min-1Culturing on a shaking table for 7 days; the obtained culture solution is transferred into 1000mL of basal culture medium according to the inoculation amount of 10 percent, and the culture is continued for 3 days under the same culture conditions; and obtaining the propagation bacterium liquid. Measured as bacterial load OD415nm0.2 to 100mL of the waste water, that is, the above-mentioned expanded bacterial solution (about 10mL) was inoculated into a 250mL triangular flask containing 100mL of the waste water, at which the bacterial amount OD in the waste water415nm0.2; compared with the live pig breeding wastewater without inoculation,at 30 ℃ and 180 r.min-1The aerobic culture is carried out for 3 days on a constant temperature shaking table. After the culture is finished, the removal rate of four pollution indexes of CODcr, BOD5, total phosphorus and ammonia nitrogen in the pig breeding wastewater is measured according to the method 1.2. And (4) performing subsequent strain identification, strain morphological characteristic observation and physiological and biochemical characteristic determination on the strain with the highest removal rate.
1.3.3 Strain identification
The strain identification adopts a Sherlock Microbiological Identity System (MIS) software system of MIDI company in America, and the system extracts and analyzes fatty acid of a single colony purified on a basic culture medium according to the operation specification of the MIDI company, qualitatively (type) and quantitatively (content) analyzes the fatty acid component of a microorganism and generates a fatty acid map, the generated map is compared with a database (Library), unknown strains are identified according to a similarity index SI (similarity index), and a certain microorganism can be basically determined if the similarity index SI is more than 0.9. The system is a microorganism identification system which is relatively quick and has rich bacteria library, and has been widely applied (Wu Hamni, Xujian, Wang Haizhen, etc. SherlockMIS system is applied to the research of soil bacteria identification. soil bulletin, 2006, 43(4): 642-647).
In addition, 16S rDNA identification method was also used for verification and comparison with Sherlock Microbiological Identity System (MIS). Using LM-DK strain total DNA as template, using 16S rDNA gene general primer to make PCR amplification, recovering and sequencing the obtained amplified fragment to define its size, then using BLAST software to make homology comparison with sequence in GenBank.
1.3.4 Observation of morphological characteristics of bacterial strains and measurement of physiological and biochemical characteristics
Inoculating the strain to a culture medium containing 30 g.L-1In a solid basic culture medium of potassium nitrate, observing the morphological characteristics of the strains by an electron microscope after 48 hours; taking the logarithmic phase of the growth of the purified strain to perform gram and crystal violet simple capsule staining and the like; the physiological and biochemical characteristics are determined according to the handbook of identification of common bacterial systems (Dongxu pearl, Chuia Miao Yin).
2. Results
2.1 isolation and screening of the strains
1 strain of bacteria which can effectively treat the pig breeding wastewater and is named as LM-DK and the others are 30 g.L is obtained through separation, purification and screening-1The degradation capability of strains (such as LM-DKq, LM-DKs and the like) which can grow on a solid basic culture medium of potassium nitrate is declined to different degrees, and after 3 days of culture, pollutants CODcr and BOD in LM-DK live pig breeding wastewater are found5The removal rates of total phosphorus and ammonia nitrogen are respectively 98.13%, 98.54%, 98.17% and 97.24%.
2.2 identification of the strains
2.2.1 basic morphology and physiological and biochemical characteristics of LM-DK strain
The bacillus is bacillus, has rough surface, pilus, no production of spores, small capsule and gram-positive bacteria, the size of the bacteria is about 0.9-1.2 μm long and 0.4-0.7 μm wide (as shown in figure 1), and on a solid culture medium, the bacterial colony is circular, has convex and smooth surface and complete edge, and is beige and opaque. The reaction of the formation test of the acetyl methyl methanol (v-p) is negative, the indole test is positive, the gelatin can not be liquefied, and other physiological and biochemical characteristics are shown in the table 1.
TABLE 1 physiological and biochemical Properties of Escherichia marmotae (Escherichia marmotae strain) LM-DK
| Test items | Results | Physiological and biochemical test | Results |
| Gram stain | + | V-P experiment | - |
| Methyl Red | - | Indole experiments | + |
| Lipase enzyme | - | Hydrogen sulfide | + |
| Lysine decarboxylation | + | Phenylalanine deaminase | - |
| Ornithine-ammonia decarboxylase | + | Oxidase enzyme | - |
| Citric acid salt | + | Urease hydrolysis | - |
| Xylose | + | Arginine double hydrolase | + |
| Glucose gas production | + | Acid production from sucrose | - |
| Malonic acid | - | Acid production from maltose | - |
| Lactose | + | Acid production by dulcitol | - |
| L-alanine | + | Denitrification | + |
| Salicylic acid | - | Liquefied gelatin | - |
Note: + positive reaction; negative reaction
2.2.2 identification of LM-DK Strain
(1) Identification of LM-DK by Sherlock MIS system
According to a fatty acid map generated by qualitative and quantitative analysis of bacterial LM-DK fatty acid by a Sherlock MIS software system, and by comparing with a Library database, the bacterial LM-DK is preliminarily identified to be Escherichia coli (Escherichia marmotae), and the similarity index SI (similarity index) is 0.987.
(2) Validation of LM-DK by 16S rDNA identification method
DNA extraction results
PCR amplification using total DNA of LM-DK strain as template and 16S rDNA gene universal gene resulted in 1 fragment of about 1kb (FIG. 2). The amplified fragment is recovered and sequenced to confirm that the size of the amplified fragment is 1328bp (SEQ ID NO. 1).
II.16S rDNA Gene PCR amplification and sequence analysis
The homology comparison of the sequencing result with the sequence in GenBank by using BLAST software shows that the LM-DK strain has higher homology with the Escherichia marmotae (Escherichia marmotae), the homology is close to 99 percent, and the genetic distance is closer.
From the combination of the physiological and biochemical properties of LM-DK, Sherlock MIS (MIDI identification System) and the phylogenetic analysis of the 16S rDNA system, it can be seen that the bacterium belongs to the species Escherichia marmota (Escherichia marmotae).
3 conclusion
1 high-efficiency strain LM-DK (CGMCC No.18769) capable of effectively treating the live pig breeding wastewater is obtained by separating biogas slurry oxidation pond bottom sludge from a live pig breeding farm in northwest urban areas of Yushui province, Xianan, and is identified as Escherichia coli (Escherichia coli) through a Sherlock MIS system and 16 SrDNA.
LM-DK was deposited with the following information:
the preservation name is: woodchuck Escherichia (Escherichia marmotae), collection unit: china general microbiological culture Collection center, preservation Address: the microbial research institute of the national academy of sciences No. 3, Xilu No.1, Beijing, Chaoyang, Beijing, with the preservation number: CGMCC No.18769, preservation date: 2019, 10 and 30.
Example 2 detection of treatment Performance of strains on pig-containing aquaculture wastewater
1 materials and methods
1.1 media and reagents
Basic culture medium: the same as in example 1.
Live pig breeding wastewater: pH6.8, CODcr concentration of 11000 mg.L-1、BOD5The concentration is 5500 mg.L-1The total phosphorus concentration was 246 mg.L-1The ammonia nitrogen concentration is 1300 mg.L-1。
1.2 Strain pair high concentration pig breeding waste water treatment performance
Propagating a single purified woodchuck Escherichia (Escherichia marmotae) LM-DK strain through a basal culture medium, and specifically comprising the following steps: inoculating Escherichia marmotae (Escherichia marmotae) LM-DK to 100mL of basal medium, and inoculating at 30 deg.C for 180r min-1Culturing on a shaking table for 7 days; the resulting culture broth was adjusted to 10% (by volume%) Inoculating the strain into 1000mL of a basic culture medium, and continuously culturing for 3 days under the same culture conditions; obtaining bacterial liquid.
Measured as bacterial load OD415nmInoculating 0.2 to 100mL of pig breeding wastewater, namely inoculating the strain of Escherichia marmotae (Escherichia marmotae) LM-DK (about 10mL) to a 250mL triangular flask containing 100mL of pig breeding wastewater, wherein the strain amount OD in the wastewater is415nm0.2; using non-inoculated live pig breeding wastewater as a control, and controlling the temperature at 30 ℃ and 180 r.min-1The aerobic culture is carried out for 3 days on a constant temperature shaking table. After the culture is finished, the strain LM-DK is used for measuring CODcr and BOD in the breeding wastewater of the live pigs according to the method of example 15And the removal rate of four pollution indexes of total phosphorus and ammonia nitrogen.
2. Results
The strain LM-DK has the pH value of 6.8 and the OD value415nm0.2, rotation speed 180 r.min-1And the biological pig breeding wastewater CODcr and BOD under the aerobic environment condition5The removal rates of total phosphorus and ammonia nitrogen are respectively 98.13%, 98.54%, 98.17% and 97.24%. CODcr, BOD5The total phosphorus and ammonia nitrogen residual concentrations are 205.7 mg.L respectively-1、80.3mg·L-1、4.6mg·L-1And 35.9 mg. L-1Can reach the discharge standard of pollutants for livestock and poultry breeding (GB18596-2001) limit standard (CODcr concentration is 400 mg.L)-1、BOD5The concentration is 150 mg.L-1The total phosphorus concentration was 8.0 mg.L-1The ammonia nitrogen concentration is 80 mg.L-1) And (4) discharging, which shows that the otter Escherichia coli (Escherichia marmotae strain) separated and screened by the research is a high-efficiency strain capable of effectively treating the live pig breeding wastewater, and has certain application potential for controlling the influence of the live pig breeding wastewater on the environment.
Comparative experiment:
the remaining strains obtained during the screening process of the present invention, as well as other currently existing Escherichia bacteria having a certain denitrification (as described in Table 2 below) and the domestic exclusively reported Escherichia marmot HT07316 were tested according to the method described in example 2 after replacing LM-DK, and the results obtained are compared with the results of LM-DK as described in Table 2 below. It can be seen that Escherichia marmot (Escherichia mar) is found in the present inventionmotae strain) LM-DK is an efficient strain capable of effectively treating pig breeding wastewater and simultaneously efficiently removing CODcr and BOD in the wastewater5Total phosphorus and ammonia nitrogen, and has certain application potential for controlling the influence of the live pig breeding wastewater on the environment; while other related genera do not have this capability or are less capable.
TABLE 2
Example 3: detection of treatment performance of strain on low-concentration cultivation wastewater generated by water-washed fence cultivation mode
1 materials and methods
1.1 media and reagents
Basic culture medium: the same as in example 1.
The water-washed fence breeding mode generates pig breeding wastewater: pH6.9, CODcr concentration of 5700 mg.L-1、BOD5The concentration is 2850 mg.L-1The total phosphorus concentration was 96 mg.L-1The ammonia nitrogen concentration is 414 mg.L-1。
1.2 treatment Performance of the strains on Low-concentration aquaculture wastewater generated by a water-flushed fence aquaculture mode
The water-washed fence breeding mode-generated live pig breeding wastewater replaces the live pig breeding wastewater in the embodiment 2, and the rest is the same as the embodiment 2.
2. Results
The strain LM-DK has the pH value of 6.9 and the OD value415nm0.2, rotation speed 180 r.min-1And the pig breeding wastewater CODcr and BOD generated by the water-washed fence breeding mode under the aerobic environment condition5The removal rates of total phosphorus and ammonia nitrogen are 97.95%, 98.96%, 98.13% and 98.21% respectively. CODcr, BOD5The total phosphorus and ammonia nitrogen residual concentrations are respectively 116.8 mg.L-1、26.8mg·L-1、1.8mg·L-1And 7.4 mg. L-1Can reach the discharge standard of pollutants for livestock and poultry breeding (GB18596-2001) limit standard (CODcr concentration is 400 mg.L)-1、BOD5The concentration is 150 mg.L-1The total phosphorus concentration was 8.0 mg.L-1And the concentration of ammonia nitrogenIs 80 mg.L-1) And (4) discharging, which shows that the separated and screened Escherichia coli (Escherichia marmotae strain) is a high-efficiency strain capable of effectively treating the live pig breeding wastewater, and has certain application potential for controlling the influence of the low-concentration live pig breeding wastewater generated by the water-washed fence breeding mode on the environment.
Example 4 detection of treatment Performance of the strains on Low carbon-nitrogen ratio live pig Breeding wastewater
1 materials and methods
1.1 media and reagents
Basic culture medium: the same as in example 1.
The low carbon-nitrogen ratio live pig breeding wastewater: pH6.8, CODcr concentration of 2130 mg.L-1、BOD5The concentration is 1340 mg.L-1The total phosphorus concentration was 96 mg.L-1The ammonia nitrogen concentration is 423 mg.L-1。
1.2 treatment performance of the strain on the low carbon-nitrogen ratio live pig breeding wastewater
The 'live pig breeding wastewater' in the example 2 is replaced by 'live pig breeding wastewater with a low carbon-nitrogen ratio', and the aerobic culture time is changed from 3 days to 2 days, and the rest is equal to the example 2.
2. Results
The strain LM-DK has the pH value of 6.8 and the OD value415nm0.2, rotation speed 180 r.min-1And the pig breeding wastewater CODcr and BOD with low carbon-nitrogen ratio under the aerobic environment condition5The removal rates of total phosphorus and ammonia nitrogen are 96.83%, 97.69%, 94.27% and 95.31% respectively. CODcr, BOD5The total phosphorus and ammonia nitrogen residual concentrations are respectively 67.5 mg.L-1、30.9mg·L-1、5.5mg·L-1And 19.8 mg. L-1Can reach the discharge standard of pollutants for livestock and poultry breeding (GB18596-2001) limit standard (CODcr concentration is 400 mg.L)-1、BOD5The concentration is 150 mg.L-1The total phosphorus concentration was 8.0 mg.L-1The ammonia nitrogen concentration is 80 mg.L-1) And (4) discharging, which shows that the separated and screened otter Escherichia coli (Escherichia marmotae strain) is a high-efficiency strain capable of effectively treating live pig breeding wastewater, and has certain application potential for controlling the influence of the low-carbon-nitrogen-ratio live pig breeding wastewater on the environmentForce.
Finally, it is also noted that the above-mentioned lists merely illustrate a few specific embodiments of the invention. It is obvious that the invention is not limited to the above embodiments, but that many variations are possible. All modifications which can be derived or suggested by a person skilled in the art from the disclosure of the present invention are to be considered within the scope of the invention.
Sequence listing
<110> New Yuhe-Biotech Co., Ltd
<120> escherichia woodchuck LM-DK separated from oxidation pond of pig farm and application thereof
<160>1
<170>SIPOSequenceListing 1.0
<210>1
<211>1328
<212>DNA
<213> Escherichia marmotae (Escherichia marmotae)
<400>1
tgacgggcgg tgtgtacaag gcccgggaac gtattcaccg tggcattctg atccacgatt 60
actagcgatt ccgacttcat ggagtcgagt tgcagactcc aatccggact acgacgcact 120
ttatgaggtc cgcttgctct cgcgaggtcg cttctctttg tatgcgccat tgtagcacgt 180
gtgtagccct ggtcgtaagg gccatgatga cttgacgtca tccccacctt cctccagttt 240
atcactggca gtctcctttg agttcccggc cggaccgctg gcaacaaagg ataagggttg 300
cgctcgttgc gggacttaac ccaacatttc acaacacgag ctgacgacag ccatgcagca 360
cctgtctcac ggttcccgaa ggcaccttct catctctgaa aagttccgtg gatgtcaaga 420
ccaggtaagg ttcttcgcgt tgcatcgaat taaaccacat gctccaccgc ttgtgcgggc 480
ccccgtcaat tcatttgagt tttaaccttg cggccgtact ccccaggcgg tcgacttaac 540
gcgttagctc cggaagccac gcctcaaggg cacaacctcc aagtcgacat cgtttacggc 600
gtggactacc agggtatcta atcctgtttg ctccccacgc tttcgcacct gagcgtcagt 660
cttcgtccag ggggccgcct tcgccaccgg tattcctcca gatctctacg catttcaccg 720
ctacacctgg aattctaccc ccctctacga gactcaagct tgccagtatc agatgcagtt 780
cccaggttga gcccggggat ttcacatctg acttaacaaa ccgcctgcgt gcgctttacg 840
cccagtaatt ccgattaacg cttgcaccct ccgtattacc gcggctgctg gcacggagtt 900
agccggtgct tcttctgcgg gtaacgtcaa tgagcaaagg tattaacttt actcccttcc 960
tccccgctga aagtacttta caacccgaag gccttcttca tacacgcggc atggctgcat 1020
caggcttgcg cccattgtgc aatattcccc actgctgcct cccgtaggag tctggaccgt 1080
gtctcagttc cagtgtggct ggtcatcctc tcagaccagc tagggatcgt cgcctaggtg 1140
agccgttacc ccacctacta gctaatccca tctgggcaca tccgatggca agaggcccga 1200
aggtccccct ctttggtctt gcgacgttat gcggtattag ctaccgtttc cagtagttat 1260
ccccctccat caggcagttt cccagacatt actcacccgt ccgccactcg tcagcgaagc 1320
agcaagct 1328
Claims (4)
1. Woodchuck Escherichia (Escherichia marmotae) LM-DK separated from bottom mud of a biogas slurry oxidation pond is characterized in that: is woodchuck Escherichia marmotae, and has a preservation number: CGMCC No. 18769.
2. The woodchuck Escherichia (Escherichia marmotae) LM-DK according to claim 1, characterized in that: the 16S rDNA gene sequence of the strain is shown in SEQ ID NO. 1.
3. Use of the Escherichia woodchuck (LM-DK) according to claim 1 or 2, characterized in that: treating the pig breeding wastewater.
4. Use of the Escherichia woodchuck (Escherichia marmotae) LM-DK according to claim 3, characterized in that: removing CODcr, BOD5, total phosphorus and ammonia nitrogen in the pig breeding wastewater.
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| CN114874947A (en) * | 2022-05-31 | 2022-08-09 | 湖南农业大学 | Escherichia coli, and product and application thereof |
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| WO2018122225A1 (en) * | 2016-12-27 | 2018-07-05 | Inbiose N.V. | In vivo synthesis of sialylated compounds |
| CN110055191A (en) * | 2019-04-08 | 2019-07-26 | 浙江大学 | It is isolated from Bai Yelin kirschner bacterium LM-W and its application of painting wastewater bed mud |
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| LIU S等: "Genomic and molecular characterisation of Escherichia marmotae from wild rodents in Qinghai-Tibet plateau as a potential pathogen", 《SCI REP》 * |
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| CN114874947A (en) * | 2022-05-31 | 2022-08-09 | 湖南农业大学 | Escherichia coli, and product and application thereof |
| CN114874947B (en) * | 2022-05-31 | 2023-09-05 | 湖南农业大学 | Coli and product and application thereof |
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