CN114854633B - Aerococcus and application thereof - Google Patents
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- CN114854633B CN114854633B CN202210516550.3A CN202210516550A CN114854633B CN 114854633 B CN114854633 B CN 114854633B CN 202210516550 A CN202210516550 A CN 202210516550A CN 114854633 B CN114854633 B CN 114854633B
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Abstract
The invention discloses a pneumococcus and application thereof, and belongs to the technical field of microorganisms. The aerococcus is named as YRD202201CF, and the classification is named as aerococcusAerococcus sp11.11 days in 2022, the strain is preserved in the China general microbiological culture Collection center, the preservation address is No. 3 of Xilu No.1 of Beijing university of Chaoyang, the postal code is 100101, and the preservation number is CGMCC No. 2458. The invention discloses that the aerococcus separated from the saline-alkali soil has the capability of utilizing various substances to generate carbonate minerals for the first time, improves the carbon reserve potential of the soil and fixes atmospheric CO for deeply understanding the soil carbon conversion process 2 The method has important scientific significance for slowing down the greenhouse effect of the atmosphere and relieving global warming.
Description
Technical Field
The invention belongs to the technical field of microorganisms, and particularly relates to aerococcus and application thereof.
Background
Saline land occupies one fourth of the global land area and is available at 1-15 × 10 annually 6 The rate of hectare is expanding. Because saline-alkali soil is uncoordinated in multi-water-heat resources and limited by soil saline-alkali stress, suitable plants are few, mainly halophytes are used as main plants, vegetation is rare, the contribution of the plants in the area to soil carbon reserves is limited, and the soil inorganic carbon reserves are higher than the soil organic carbon reserves.
At present, the capability of microorganisms for forming carbon-containing minerals is widely researched, and whether the microorganisms capable of forming the carbon-containing minerals exist in the soil stressed by saline alkali or not has an important effect on understanding the formation mechanism of the carbon reservoir in the saline-alkali soil and exploring the carbon accumulation process in the saline-alkali soil. Therefore, it is necessary to explore the ability of microorganisms in saline-alkali soil to synthesize carbonaceous minerals.
Disclosure of Invention
Aiming at the defects in the prior art, the invention provides a gas coccus and application thereof, and discloses the capability of the gas coccus separated from saline-alkali soil to generate carbonate minerals by utilizing various substances for the first time.
In order to achieve the purpose of the invention, the invention adopts the technical scheme that:
provides a kind of Aerococcus, named YRD202201CF, classified named Aerococcus sp, which is preserved in China general microbiological culture Collection center at 11.3.2022.s.with the preservation address of No. 3 Hospital No.1 Xilu of Chaoyang district, beijing, the postal code is 100101 and the preservation number is CGMCC No. 2458.
Further, the method comprises the following steps: the 16S rDNA gene sequence of the aerococcus is shown in SEQ ID No. 1.
Further, the method comprises the following steps: the aerococcus comes from saline-alkali soil.
Further, the method comprises the following steps: the screening method of the aerococcus comprises the following steps: diluting saline-alkali soil, taking soil diluent, inoculating the soil diluent into an inorganic salt MSM culture medium containing calcium oxalate, and culturing and separating to obtain aerococcus.
The invention also provides the application of the aerococcus in the metabolism and utilization of calcium oxalate to form carbon-containing minerals.
The invention also provides application of the aerococcus in inducing the generation of carbonate minerals.
Further: carbonaceous minerals include, but are not limited to, carbonaceous inorganic compounds, such as carbonates.
Further: carbonates include, but are not limited to, calcium carbonate and the allotropes of calcium carbonate, such as vaterite, calcite.
The beneficial effects of the invention are as follows:
the invention discloses that the aerococcus separated from the saline-alkali soil has the capability of utilizing various substances to generate carbonate minerals for the first time, and has important application potential and scientific significance for deeply understanding the soil carbon conversion process and improving the soil carbon reserve potential.
Drawings
FIG. 1 is an electron microscope scanning image of the surface of a soil thin layer of an experimental group in example 2 of the present invention;
FIG. 2 is an electron microscope scanning image of the surface of a control group soil thin layer in example 2 of the present invention;
FIG. 3 is a graph of the energy dispersive diffraction analysis of the mineral energy on the surface of the soil thin layer in the experimental group of example 2;
FIG. 4 is a graph of the energy dispersive diffraction analysis of the mineral energy on the surface of the soil thin layer of the control group in example 2 of the present invention;
FIG. 5 is an electron micrograph of a precipitate obtained in example 3 of the present invention;
FIG. 6 is an energy dispersive diffraction analysis chart of the precipitate in example 3 of the present invention.
Detailed Description
The following description of the embodiments of the present invention is provided to facilitate the understanding of the present invention by those skilled in the art, but it should be understood that the present invention is not limited to the scope of the embodiments, and it will be apparent to those skilled in the art that various changes may be made without departing from the spirit and scope of the invention as defined and defined in the appended claims, and all matters produced by the invention using the inventive concept are protected.
Unless otherwise indicated, the examples follow conventional experimental conditions, such as the Molecular Cloning Laboratory Manual, sambrook, et al (Sambrook J & Russell DW, molecular Cloning: a Laboratory Manual, 2001), or following the conditions recommended by the manufacturer's instructions.
Example 1
Isolation and identification of strain YRD202201 CF:
taking soil of typical saline-alkali soil of Shandong yellow river delta as target soil, the specific screening steps are as follows:
1. collecting 0-10cm deep soil, collecting 1.0g soil, adding into 9mL sterilized water, and shaking for 10min to obtain soil solution. 1mL of the soil suspension was taken and placed in 9mL of sterilized water to dilute the soil solution. This operation was repeated to dilute the soil solution to 10 deg.f -3 And (4) doubling. 100. Mu.L of the diluted soil solution was inoculated into MSM medium containing calcium oxalate, and cultured at 25 ℃ for 7 days.
Specifically, the medium comprises the following components: na (Na) 2 HPO 4 ·2H 2 O，3.5g/L；KH 2 PO 4 ，1.0g/L；(NH 4 ) 2 SO 4 ，0.5g/L；MgCl 2 ·6H 2 O，0.1g/L；Ca(NO 3 ) 2 ·4H 2 O,0.05g/L; 1mL/L of trace element solution; caC 2 O 4 2g/L;1.5% agar, pH 7.25.
The formula of the trace element solution is as follows: feSO 4 ·7H 2 O，0.4g/L；ZnSO 4 ·7H 2 O，0.1g/L；MnCl 2 ·4H 2 O，0.3g/L；H 3 BO 3 ，0.3g/L；CuCl 2 ·2H 2 O，0.1g/L；NiCl 2 ·6H 2 O，0.2g/L；NaMoO 4 ，0.3g/L；CoCl 2 ·6H 2 O，0.1g/L；Na 2 SeO 4 ·2H 2 O，0.05g/L。
2. And (4) picking single colonies growing on the culture medium, coating the single colonies on an LB culture medium, and further separating and purifying. Strain YRD202201CF was obtained.
3. 16S rDNA sequencing of strain YRD202201 CF: the universal primer 27F and the 1492R primer are adopted to carry out PCR amplification on the genome DNA of the strain YRD202201CF, the amplified product is sequenced, and the sequencing result is shown as SEQ ID No. 1.
SEQ ID No.1 is shown below:
CGGGGGGGATGCCTATAGATGCAGTCGAGCGAACAGATGAAGTGCTTGCACTTCTGACGTTAGCGGCGAACGGGTGAGTAACACGTAAGGAATCTACCTATAAGCGGGGGATAACATTCGGAAACGGGTGCTAATACCGCATAATATCTTCTTCCGCATGGAAGAAGATTGAAAGACGGCTCTGCTGTCACTTATAGATGACCTTGCGGTGCATTAGTTAGTTGGTGGGGTAACGGCCTACCAAGACGATGATGCATAGCCGACCTGAGAGGGTGATCGGCCACATTGGGACTGAGACACGGCCCAAACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCGCAATGGGCGAAAGCCTGACGGAGCAATGCCGCGTGAGTGAAGAAGGCCTTCGGGTCGTAAAACTCTGTTATAAGAGAAGAACAAATTGTAGAGTAACTGCTACAGTCTTGACGGTATCTTATCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGATTTATTGGGCGTAAAGGGAGCGCAGGTGGTTTCTTAAGTCTGATGTGAAAGCCCACGGCTTAACCGTGGAGGGTCATTGGAAACTGGGAAACTTGAGTACAGAAGAGGAATGTGGAACTCCATGTGTAGCGGTGGAATGCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGACATTCTGGTCTGTTACTGACACTGAGGCTCGAAAGCGTGGGGAGCAAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAGGTGTTGGAGGGTTTCCGCCCTTCAGTGCCGCAGTTAACGCATTAAGCACTCCGCCTGGGGAGTACGACCGCAAGGTTGAAACTCAAAGGAATTGACGGGGACCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAAGTCTTGACATCCTTTGACCACCCTAGAGATAGGGCTTTCCCTTCGGGGACAAAGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCCTATTATTAGTTGCCAGCATTCAGTTGGGCACTCTAATGAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCAGCATGCCCCTTATGACTTGGGCTACACACGTGCTACAATGGATGGTACAACGAGTCGCAAACCCGCGAGGGCAAGCAAATCTCTTAAAGCCATTCTCAGTTCGGATTGCAGGCTGCAACTCGCCTGCATGAAGCCGGAATCGCTAGTAATCGTGGATCAGCACGCCACGGTGAATACGTTCCCGGGTCTTGTACACACCGCCCGTCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTTATGGAGCCAGCCGCCGAAGGGGGGATAC
BLAST analysis was performed based on the sequence of the 16S rDNA gene using NCBI database, and the results showed that the 16S rDNA gene sequence was homologous to Aerococcus (99.52% homology to Aerococcus sp.train 201707CJKOP-Y33, genBank: CP 080013.1), and the strain was finally determined to be Aerococcus by combining the morphological, structural and physiological and biochemical characteristics of the strain YRD202201CF and was named as YRD202201CF.
Example 2
Strain YRD202201CF forms carbon-containing minerals with hardly available organic matter (calcium oxalate):
1. and (3) acidifying, neutralizing, leaching and drying the saline-alkali soil to obtain the soil without inorganic carbon. And taking 10g of the treated soil sample, adding 200mg of calcium oxalate powder serving as a carbon source substance and 20mg of polyethylene glycol serving as a soil binder, uniformly mixing, putting the mixed sample into 50mL of deionized water, and oscillating for 5min on a vortex oscillator to prepare a suspension. A200. Mu.L suspension was placed in a 35mm diameter petri dish and dried in a dust-free environment, at which time a thin layer of soil was formed in the petri dish. The thin layer is an observation interface for observing the conversion of the calcium oxalate into the carbon-containing minerals by the microorganisms.
2. Sterilizing a culture dish containing a soil thin layer for 30min at 121 ℃ to remove the pollution of mixed bacteria, adding 2mL of sterilized deionized water to infiltrate the soil thin layer into water, and inoculating a strain YRD202201CF into the culture dish, wherein a control group is a strain subjected to inoculation and inactivation treatment, and other conditions are the same. The petri dish was capped and incubated at room temperature for 3 months.
3. And after the culture is finished, observing the change conditions of the carbon-containing minerals on the soil thin layers of the experimental group and the control group by adopting a scanning electron microscope and an energy dispersion diffraction analysis method. As shown in FIGS. 1 and 2, the surface of the soil thin layer of the experimental group is found to have a lot of minerals and complex structures through the analysis of a scanning electron microscope, while the surface of the control group is smooth and only a few mineral structures exist. The energy dispersive diffraction analysis (fig. 3 and 4) is performed on the typical points on fig. 1 and fig. 2, the compositions of the minerals on the surfaces of the experimental group and the control group (table 1) both contain carbon, oxygen, calcium, magnesium, aluminum, silicon and other elements, and the carbon content of the experimental group is obviously higher than that of the control group, namely the strain YRD202201CF can indeed form the carbon-containing minerals.
TABLE 1 surface mineral element composition of experimental and control groups
Example 3
The strain YRD202201CF uses easily available organic substances (organic substances rich in carbon and nitrogen) to generate carbonate:
100mL of liquid culture medium is prepared, placed in a 250mL triangular flask, and sterilized at 121 ℃ for 30min. Specifically, the liquid medium comprises the following components: 10g/L of tryptone, 5g/L of yeast extract, 10g/L of sodium chloride and 5g/L of calcium chloride.
Inoculating the strain YRD202201CF into the culture medium by using an inoculating needle, inoculating the inactivated strain into a control group, keeping the conditions consistent, and culturing at room temperature. Whether precipitates are generated or not is observed every other week, precipitates are generated in the experimental group is observed at week 4, and the process is cultured for 60 days in order to obtain enough precipitates for subsequent analysis. After the culture was completed, the precipitates formed in the experimental group were collected and treated with hydrogen peroxide solution to remove the residues of microorganisms, etc., to obtain 0.2799g of the precipitates in total, while no precipitate was formed in the control. And (4) performing electron microscope scanning and energy dispersion diffraction analysis on the precipitate.
The result is shown in fig. 5, and the scanning electron microscope analysis shows that the precipitates generated in the experiment are hemispherical, and the hemispherical structural layers are laminated together and have a porous structure, which is a direct evidence of the action of the microorganism, and shows that the microorganisms are continuously generated by the precipitates in the surrounding environment during the life process, and are laminated together and gathered around the microorganisms; as shown in fig. 6 and table 2, according to the energy dispersive diffraction analysis, the main components of the precipitate were carbon, oxygen, calcium, and calcium carbonate by atomic percentage and weight percentage analysis, indicating that the strain YRD202201CF has the ability to generate carbon-containing minerals. Furthermore, by X-ray diffraction analysis, these calcium carbonates were found to be mainly composed of 80% vaterite and 20% calcite, demonstrating that strain YRD202201CF indeed forms carbonate mineral crystals.
TABLE 2 carbon-containing mineral element composition analysis Table (number of weight and atom)
The invention discloses that the aerococcus separated from the saline-alkali soil has the capability of utilizing various substances to generate carbonate minerals for the first time, and has important scientific significance for deeply understanding the soil carbon conversion process and improving the soil carbon reserve potential.
It will be evident to those skilled in the art that the invention is not limited to the details of the foregoing illustrative embodiments, and that the present invention may be embodied in other specific forms without departing from the spirit or essential attributes thereof. The present embodiments are therefore to be considered in all respects as illustrative and not restrictive, the scope of the invention being indicated by the appended claims rather than by the foregoing description, and all changes which come within the meaning and range of equivalency of the claims are therefore intended to be embraced therein. Any reference sign in a claim should not be construed as limiting the claim concerned.
Furthermore, it should be understood that although the present specification describes embodiments, not every embodiment includes only a single embodiment, and such description is for clarity purposes only, and it is to be understood that all embodiments may be combined as appropriate by one of ordinary skill in the art to form other embodiments as will be apparent to those of skill in the art from the description herein.
Sequence listing
<110> institute of geoscience and resource of Chinese academy of sciences
<120> Aerococcus and application thereof
<141> 2022-05-09
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1450
<212> DNA
<213> Aerococcus YRD202201CF (Aerococcus sp. YRD202201 CF)
<400> 1
cgggggggat gcctatagat gcagtcgagc gaacagatga agtgcttgca cttctgacgt 60
tagcggcgaa cgggtgagta acacgtaagg aatctaccta taagcggggg ataacattcg 120
gaaacgggtg ctaataccgc ataatatctt cttccgcatg gaagaagatt gaaagacggc 180
tctgctgtca cttatagatg accttgcggt gcattagtta gttggtgggg taacggccta 240
ccaagacgat gatgcatagc cgacctgaga gggtgatcgg ccacattggg actgagacac 300
ggcccaaact cctacgggag gcagcagtag ggaatcttcc gcaatgggcg aaagcctgac 360
ggagcaatgc cgcgtgagtg aagaaggcct tcgggtcgta aaactctgtt ataagagaag 420
aacaaattgt agagtaactg ctacagtctt gacggtatct tatcagaaag ccacggctaa 480
ctacgtgcca gcagccgcgg taatacgtag gtggcaagcg ttgtccggat ttattgggcg 540
taaagggagc gcaggtggtt tcttaagtct gatgtgaaag cccacggctt aaccgtggag 600
ggtcattgga aactgggaaa cttgagtaca gaagaggaat gtggaactcc atgtgtagcg 660
gtggaatgcg tagatatatg gaagaacacc agtggcgaag gcgacattct ggtctgttac 720
tgacactgag gctcgaaagc gtggggagca aacaggatta gataccctgg tagtccacgc 780
cgtaaacgat gagtgctagg tgttggaggg tttccgccct tcagtgccgc agttaacgca 840
ttaagcactc cgcctgggga gtacgaccgc aaggttgaaa ctcaaaggaa ttgacgggga 900
cccgcacaag cggtggagca tgtggtttaa ttcgaagcaa cgcgaagaac cttaccaagt 960
cttgacatcc tttgaccacc ctagagatag ggctttccct tcggggacaa agtgacaggt 1020
ggtgcatggt tgtcgtcagc tcgtgtcgtg agatgttggg ttaagtcccg caacgagcgc 1080
aacccctatt attagttgcc agcattcagt tgggcactct aatgagactg ccggtgacaa 1140
accggaggaa ggtggggatg acgtcaaatc agcatgcccc ttatgacttg ggctacacac 1200
gtgctacaat ggatggtaca acgagtcgca aacccgcgag ggcaagcaaa tctcttaaag 1260
ccattctcag ttcggattgc aggctgcaac tcgcctgcat gaagccggaa tcgctagtaa 1320
tcgtggatca gcacgccacg gtgaatacgt tcccgggtct tgtacacacc gcccgtcaca 1380
ccacgagagt ttgtaacacc cgaagtcggt gaggtaacct ttatggagcc agccgccgaa 1440
ggggggatac 1450
Claims (5)
1. A pneumococcus (Aerococcus sp.) characterized by: the Aerococcus is named as YRD202201CF, is classified and named as Aerococcus sp, and is preserved in the common microorganism center of China general microbiological culture Collection management Committee in 2022, 3 months and 11 days, wherein the preservation address is No. 3 of Xilu No.1 of Chao Xinyang district, beijing, the postal code is 100101, and the preservation number is CGMCC No. 2458.
2. Use of a gasococcus according to claim 1 for metabolizing calcium oxalate to form carbonaceous minerals.
3. Use of a pneumococcus bacterium according to claim 2, in the metabolic utilization of calcium oxalate to form carbonaceous minerals, wherein: the carbonaceous minerals include, but are not limited to, carbonaceous inorganic compounds.
4. Use of an aerococcus bacterium according to claim 1 for inducing carbonate mineral production.
5. Use of a gasococcus according to claim 4 for inducing carbonate mineral formation, wherein: the carbonates include, but are not limited to, calcium carbonate and the allotrope of calcium carbonate.
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| CN102757925A (en) * | 2012-08-06 | 2012-10-31 | 山东轻工业学院 | Marine bacterium Aerococcus urinaeequi HZ and polysaccharide produced by bacterium |
| CN109182205A (en) * | 2018-10-09 | 2019-01-11 | 北京林业大学 | Rhodococcus sp and its application with carbon sequestration capacity |
| CN109266621A (en) * | 2018-09-25 | 2019-01-25 | 吉林大学 | One plant of new light green color aerococcus bacteriophage AVP and application thereof |
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| CN102757925A (en) * | 2012-08-06 | 2012-10-31 | 山东轻工业学院 | Marine bacterium Aerococcus urinaeequi HZ and polysaccharide produced by bacterium |
| CN109266621A (en) * | 2018-09-25 | 2019-01-25 | 吉林大学 | One plant of new light green color aerococcus bacteriophage AVP and application thereof |
| CN109182205A (en) * | 2018-10-09 | 2019-01-11 | 北京林业大学 | Rhodococcus sp and its application with carbon sequestration capacity |
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