Brucella WYQ-2, bacterial agent and use thereof

Abstract

The present invention relates to a Brucella WYQ-2, a bacterial agent and a use thereof. The Brucella WYQ-2 is classified as Brucella ( Brucella sp. ), deposited in China Center for Type Culture Collection, with a deposit number of CCTCC No: M 20242175, and a deposit date of October 14, 2024; the thalli of Brucella WYQ‑2 are small dots, white and opaque, full in shape, and without flagella; the bacterial lawn grows along the streak. Brucella WYQ‑2 and the bacterial agent of the present invention, under the condition of having an electron donor, especially with reduced iron powder as an electron donor, can break through the chemical barrier of extremely high chemical energy required for the conversion of carbon dioxide into other high value-added products, and realize effective fixation of carbon dioxide. In addition, reduced iron powder can be used as a trace element to regulate the secretion of protein during the growth of microorganisms in the fixation reaction.

Description

Technical Field

[0001] The invention belongs to the technical field of biological carbon fixation, and in particular relates to a method for fixing carbon dioxide with Brucella. Background Art

[0002] At present, common CO2 utilization technologies include physical, chemical, and biological methods. Bioconversion technology has the advantages of mild reaction conditions, permanent carbon fixation, environmental friendliness, diverse products, and high product added value. It is currently one of the most important and effective ways to utilize CO2 resources in the world. The process of plant photosynthesis is relatively complex and easily affected by biochemical and diffusion factors, while the microbial fixation pathway is relatively simple. Therefore, using autotrophic microorganisms to fix CO2 and convert it into high-value-added products is an effective strategy to achieve CO2 resource utilization. In this process, microorganisms can capture and convert CO2 under relatively mild pressure and temperature conditions without secondary pollution. At the same time, they can also produce value-added compounds such as ethanol and acetic acid, which has attracted widespread attention.

[0003] Although hydrogen-oxidizing bacteria are highly efficient in converting carbon dioxide into organic carbon, the preparation, storage, transportation, and use of hydrogen require special hydrogen safety technologies, and therefore place high demands on the conditions of the entire laboratory, which hinders the large-scale application of this type of microorganism. Therefore, it is urgent to screen for a non-hydrogen-oxidizing bacterium that can convert CO2 into organic carbon. Summary of the invention

[0004] In view of the problems existing in the above-mentioned prior art, the present invention provides a Brucella WYQ-2, a bacterial agent and its use. The Brucella WYQ-2 and the bacterial agent of the present invention, under the condition of having an electron donor, especially when using reduced iron powder as an electron donor, can break through the chemical barrier that extremely high chemical energy is required for the conversion of carbon dioxide into other high value-added products, and realize the effective fixation of carbon dioxide. In addition, the reduced iron powder (after being converted from zero valent to divalent) in the fixation reaction can be used as a trace element to regulate the secretion of protein during the growth of microorganisms.

[0005] On the one hand, the present invention provides a Brucella WYQ-2, the microbial classification of the Brucella WYQ-2 is named Brucella ( Brucella sp. ), deposited in China Center for Type Culture Collection, with the deposit number being CCTCC No: M20242175, and the deposit date being October 14, 2024.

[0006] Furthermore, the Brucella WYQ-2 bacteria described in the present invention are in the form of small dots, are white and opaque, have a plump shape, and have no flagella.

[0007] Furthermore, the Brucella WYQ-2 bacterial lawn described in the present invention grows along the streak.

[0008] On the other hand, the present invention provides a bacterial agent, which includes the Brucella WYQ-2 described in any one of the above items.

[0009] Furthermore, the preparation method of the bacterial agent of the present invention comprises:

[0010] Step 1) Slant culture:

[0011] The Brucella WYQ-2 was inoculated into a solid culture medium and cultured for 48 to 72 hours at a culture temperature of 30° C. to obtain a slant bacterial cell;

[0012] The solid culture medium is prepared by adding 16 g of Na2HPO4·12H2O, 6.8 g of KH2PO4, 4 g of CH3COONa, 1 g of yeast powder, 0.002 g of FeSO4, 0.0002 g of (NH4)2MoO4, and 1 mL of trace element solution into 800 ml of distilled water to obtain solution A, dissolving 0.58 of MgSO4 and 0.067 of CaCl2 into 200 ml of distilled water to obtain solution B, slowly stirring and mixing solutions A and B to obtain a culture medium, then taking 100 ml of the liquid volume and adding it into a 250 ml serum bottle, adding 2 g of agar to prepare a solid culture medium;

[0013] Step 2) Expand the culture:

[0014] The slant bacteria obtained in step 1) are inoculated on a plate for 48 to 72 hours, and then inoculated into a liquid culture medium for shaking culture for 24 to 48 hours, and then centrifuged to remove the supernatant, and then washed with an inorganic salt solution, and then added with an inorganic salt solution and shaken thoroughly to obtain the bacterial agent;

[0015] The rotation speed of the shaking culture is 140-180 r / min, the temperature of the shaking culture is 25-30° C., and the pH of the liquid culture medium is 5-9.

[0016] Furthermore, in the preparation method of the bacterial agent of the present invention, the composition of the liquid culture medium is: 5 g / L yeast powder, 10 g / L peptone, and 10 g / L NaCl;

[0017] Alternatively, the composition of the liquid culture medium is: 4.5 g / L Na2HPO4·12H2O, 1.0 g / L KH2PO4, 2.5 g / L (NH4)2SO4, 0.2 g / L MgSO4·7H2O, 0.023 g / L CaCl2, 1 mL / L trace element mother solution, and 1 mL / L vitamin mother solution.

[0018] Furthermore, in the preparation method of the microbial agent described in the present invention, the trace element mother solution includes: 1 g / L FeSO4, 0.05 g / L CuSO4, 0.014 g / L H3BO3, 0.1 g / L MnSO4·4H2O, 0.02 g / L Na2MoO4·2H2O, and 0.02 g / L CoCl2·6H2O.

[0019] The vitamin mother solution also includes: 0.2g / L biotin VH, 0.2g / L folic acid VB, 1g / L pyridoxine hydrochloride VB6, 0.5g / L riboflavin, 0.45-0.55g / L vitamin B2, 0.5g / L ammonium sulfate VB1, 0.5g / L nicotinic acid, 0.5g / L pantothenic acid VB5, 0.01g / L B-12, 0.5g / L p-aminobenzoic acid and 0.5g / L lipoic acid.

[0020] In another aspect, the present invention provides a use of the Brucella WYQ-2 described in any one of the above items, wherein the Brucella WYQ-2 is used to fix carbon dioxide and perform a fixation reaction under the condition of providing an electron donor;

[0021] or,

[0022] The Brucella WYQ-2 is used to fix carbon dioxide under the condition that reduced iron powder is used as an electron donor to perform a fixation reaction.

[0023] In another aspect, the present invention provides a use of any of the above-mentioned bacterial agents, wherein the bacterial agent is used to fix carbon dioxide under the condition that reduced iron powder is used as an electron donor to carry out a fixation reaction.

[0024] Furthermore, in the use of Brucella WYQ-2 or the bacterial agent described in the present invention, the volume fraction of the initial carbon dioxide concentration of the fixation reaction is 30 vol%~66 vol%, the concentration of the initial reduced iron powder is 0.1~2 g / 100mL, and the pH of the fixation reaction is 5~9.

[0025] Furthermore, the Brucella WYQ-2 of the present invention exhibits different fixation effects on CO2 at different electron donor concentrations. 600 When the concentration of electron donor (iron powder concentration) is 0.5~2 g / 100mL, it shows a relatively excellent degradation effect on CO2.

[0026] Furthermore, the effect of Brucella WYQ-2 on CO2 fixation provided by the present invention is affected by pH. 600When the pH value was 0.05, the volume fraction of CO2 introduced was 35 vol%, and the pH was in the range of 7-9, the bacteria were able to exhibit excellent fixation performance for CO2, which proved that the pH range suitable for the growth of the bacteria was alkaline conditions, which was consistent with the pH changes at the end of the previous sludge acclimation.

[0027] Compared with the prior art, the present invention has the following beneficial technical effects:

[0028] (1) The bacteria of the present invention, Brucella WYQ-2, is a non-oxidizing bacteria that can grow under conditions with CO2 as an inorganic carbon source and an electron donor, especially reduced iron powder as an electron donor, and can convert CO2 into high value-added products without hydrogen conditions. At the same time, compared with oxidizing bacteria that use hydrogen as an electron donor, using reduced iron powder as an electron donor is inexpensive, has higher experimental safety, mild reaction conditions, and is environmentally friendly.

[0029] (2) The Brucella WYQ-2 of the present invention has excellent CO2 fixation effect, and the initial bacterial cell concentration OD 600 =0.05, it can still fix high concentration CO2, and can still fix CO2 with a volume fraction greater than or equal to 66%. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 It is a colony photo of Brucella WYQ-2 of the present invention;

[0031] Figure 2 It is a negative staining diagram of Brucella WYQ-2 of the present invention;

[0032] Figure 3 is a phylogenetic tree of Brucella WYQ-2 of the present invention;

[0033] Figure 4-1 It is a comparison diagram of the change of carbon dioxide gas phase concentration in the Brucella WYQ-2 carbon dioxide fixation reaction in Examples 4 to 7 and Comparative Examples 1 to 3;

[0034] Figure 4-2 It is a comparison chart of the change of TOC content of Brucella WYQ-2 in the carbon dioxide fixation reaction in Examples 4 to 7 and Comparative Examples 1 to 3;

[0035] Figure 4-3 It is a comparison diagram of the proteoglycan content change of Brucella WYQ-2 in the carbon dioxide fixation reaction in Examples 4 to 7 and Comparative Examples 1 to 3;

[0036] Figure 5-1 It is a comparison diagram of the change of carbon dioxide gas phase concentration in the Brucella WYQ-2 carbon dioxide fixation reaction in Examples 9 to 11 and Comparative Examples 4 to 6;

[0037] Figure 5-2 It is a comparison chart of the change in TOC content of Brucella WYQ-2 in the carbon dioxide fixation reaction in Examples 9 to 11 and Comparative Examples 4 to 6;

[0038] Figure 5-3 It is a comparison chart of the changes in proteoglycan content of Brucella WYQ-2 in the carbon dioxide fixation reaction in Examples 9 to 11 and Comparative Examples 4 to 6;

[0039] Figure 6-1 This is a comparison diagram of the change in carbon dioxide gas phase concentration of Brucella WYQ-2 in the carbon dioxide fixation reaction in Example 12;

[0040] Figure 6-2 This is a comparison chart of the changes in TOC content of Brucella WYQ-2 in the carbon dioxide fixation reaction in Example 12;

[0041] Figure 6-3 This is a comparison chart of the changes in proteoglycan content in the Brucella WYQ-2 carbon dioxide fixation reaction in Example 12. DETAILED DESCRIPTION

[0042] In order to have a clearer understanding of the technical features, purposes and effects of the present invention, the specific implementation methods of the present invention are now described in detail with reference to the accompanying drawings. The described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention. If the specific conditions are not specified in the specific implementation method, they shall be carried out according to the conventional conditions or the conditions provided by the manufacturer.

[0043] When the following description refers to the drawings, unless otherwise indicated, the same numbers in different drawings represent the same or similar elements. The embodiments described in the following exemplary embodiments do not represent all embodiments consistent with the present disclosure. Instead, they are merely examples of devices and methods consistent with some aspects of the present disclosure as detailed in the appended claims. The terms used in the present disclosure are only for the purpose of describing specific embodiments and are not intended to limit the present disclosure.

[0044] Implementation method 1:

[0045] A Brucella WYQ-2 of the present invention, the microbial classification of the Brucella WYQ-2 is named Brucella ( Brucella sp. ), deposited in China Center for Type Culture Collection, with the deposit number being CCTCC No: M 20242175, and the deposit date being October 14, 2024.

[0046] The above-mentioned Brucella WYQ-2 bacteria are in the shape of small dots, white and opaque, plump in shape, and without flagella.

[0047] The bacterial lawn of Brucella WYQ-2 mentioned above grows along the streaking line.

[0048] Implementation method 2:

[0049] A bacterial agent of the present invention comprises any one of the above-mentioned Brucella WYQ-2.

[0050] The preparation method of the above-mentioned bacterial agent comprises:

[0051] Step 1) Slant culture:

[0052] The Brucella WYQ-2 was inoculated into a solid culture medium and cultured for 48 to 72 hours at a culture temperature of 30° C. to obtain slant cells;

[0053] The solid culture medium is prepared by adding 16 g of Na2HPO4·12H2O, 6.8 g of KH2PO4, 4 g of CH3COONa, 1 g of yeast powder, 0.002 g of FeSO4, 0.0002 g of (NH4)2MoO4, and 1 mL of trace element solution into 800 ml of distilled water to obtain solution A, dissolving 0.58 of MgSO4 and 0.067 of CaCl2 into 200 ml of distilled water to obtain solution B, slowly stirring and mixing solutions A and B to obtain a culture medium, then taking 100 ml of the liquid volume and adding it into a 250 ml serum bottle, adding 2 g of agar to prepare a solid culture medium;

[0054] Step 2) Expand the culture:

[0055] The slant bacteria obtained in the above step 1) are inoculated on a plate for 48 to 72 hours, and then inoculated into a liquid culture medium for shaking culture for 24 to 48 hours, and then centrifuged to remove the supernatant, and then washed with an inorganic salt solution, and then added with an inorganic salt solution and shaken thoroughly to obtain the above bacterial agent;

[0056] The rotation speed of the shaking culture is 140-180 r / min, the temperature of the shaking culture is 25-30° C., and the pH of the liquid culture medium is 5-9.

[0057] In some embodiments, the composition of the liquid culture medium is: 5 g / L yeast powder, 10 g / L peptone, and 10 g / L NaCl;

[0058] Alternatively, in some embodiments, the composition of the above-mentioned liquid culture medium is: 4.5 g / L Na2HPO4·12H2O, 1.0 g / L KH2PO4, 2.5 g / L (NH4)2SO4, 0.2 g / L MgSO4·7H2O, 0.023 g / L CaCl2, 1 mL / L trace element mother solution, and 1 mL / L vitamin mother solution.

[0059] In some embodiments, the above-mentioned trace element mother solution includes: 1 g / L FeSO4, 0.05 g / L CuSO4, 0.014 g / L H3BO3, 0.1 g / L MnSO4·4H2O, 0.02 g / L Na2MoO4·2H2O, and 0.02 g / L CoCl2·6H2O.

[0060] In some embodiments, the vitamin mother solution further includes: 0.2 g / L biotin VH, 0.2 g / L folic acid VB, 1 g / L pyridoxine hydrochloride VB6, 0.5 g / L riboflavin, 0.45-0.55 g / L vitamin B2, 0.5 g / L ammonium sulfate VB1, 0.5 g / L niacin, 0.5 g / L pantothenic acid VB5, 0.01 g / L B-12, 0.5 g / L p-aminobenzoic acid and 0.5 g / L lipoic acid.

[0061] Implementation method three:

[0062] A use of any one of the above-mentioned Brucella WYQ-2 of the present invention, wherein the Brucella WYQ-2 is used to fix carbon dioxide and perform a fixation reaction under the condition of providing an electron donor;

[0063] or,

[0064] The Brucella WYQ-2 is used to fix carbon dioxide under the condition that reduced iron powder is used as an electron donor to carry out a fixation reaction.

[0065] In some embodiments, the volume fraction of the initial carbon dioxide concentration of the above-mentioned fixation reaction is 30 vol%~66 vol%, the concentration of the initial reduced iron powder is 0.1~2 g / 100 mL, and the pH of the above-mentioned fixation reaction is 5~9.

[0066] Implementation method 4:

[0067] A use of any one of the above-mentioned bacterial agents of the present invention is to fix carbon dioxide under the condition that reduced iron powder is used as an electron donor to carry out a fixation reaction.

[0068] In some embodiments, the volume fraction of the initial carbon dioxide concentration of the above-mentioned fixation reaction is 30 vol%~66 vol%, the concentration of the initial reduced iron powder is 0.1~2 g / 100 mL, and the pH of the above-mentioned fixation reaction is 5~9.

[0069] The present invention is further described in detail below with reference to specific embodiments.

[0070] Embodiment 1:

[0071] A Brucella WYQ-2 of the present invention, the microbial classification of Brucella WYQ-2 is named Brucella ( Brucella sp. ), deposited in China Center for Type Culture Collection, with the deposit number being CCTCC No: M 20242175, and the deposit date being October 14, 2024.

[0072] The bacteria of Brucella WYQ-2 are small dots, white and opaque, plump in shape, and without flagella.

[0073] Brucella WYQ-2 bacterial lawn grows along the streak.

[0074] The deposit address of Brucella WYQ-2 is Wuhan University, Wuhan, China, zip code 430072.

[0075] Embodiment 2:

[0076] Isolation, purification and identification of Brucella WYQ-2 of the present invention

[0077] (1) Isolation and purification of Brucella WYQ-2

[0078] Anaerobic sludge was collected on-site from a sewage treatment plant in Zhoushan, Zhejiang. After standing and settling for 3 hours, the supernatant and suspended impurities were removed, leaving the sludge with fine particles. 10 ml of the sludge was mixed with 100 ml of inorganic salt culture medium (1 g of reduced iron powder was added), and CO2 gas with a purity of 99.999% was used as the inorganic carbon source for microbial growth. The culture medium was aerated with nitrogen for 30 minutes before each use to remove the dissolved oxygen in the culture medium.

[0079] During the experiment of acclimating and screening carbon dioxide-fixing bacteria in anaerobic sludge, the culture medium was firstly injected with a concentration of 230 g / m 3 The CO2 gas was removed and the CO2 concentration in the gas phase, the TOC content in the liquid phase and the OD in the whole culture system were detected after 12 h. 600 (Characterizing microbial growth).

[0080] TOC (Total Organic Carbon) content refers to the total amount of carbon in dissolved and suspended organic matter in water. The higher the TOC content, the higher the organic matter content in the water, so it can be used as an indicator to evaluate organic pollution in water quality. 600 It refers to the optical density value measured when the wavelength of the spectrophotometer is set to 600nm. It is one of the commonly used biomass determination methods in the process of microbial fermentation and can indirectly reflect the concentration of the fermentation liquid. 600 Proportional to the bacterial concentration, OD 600 The higher the value, the higher the bacterial concentration. 600 The lower the value, the lower the bacterial concentration, which is a proportional relationship rather than a linear relationship.

[0081] The study found that the CO2 concentration in the gas phase showed a decreasing trend, and the liquid phase TOC content and OD 600 The values ​​increased, proving that CO2-fixing bacteria existed in the culture system. Iron powder could be reduced as an electron donor and CO2 gas could be used as an inorganic carbon source for microbial growth. The sludge in the bottle was then acclimated for one month, with each cycle lasting 7 days for a total of 4 cycles. The CO2-fixing strains were isolated and purified, and 350 g / m 3 After each cycle, 5 ml of the acclimated sample in the shake flask was transferred to other shake flasks for further performance testing. The culture was carried out at a culture speed of 160 r / min and a culture temperature of 30°C. The CO2 concentration in the gas phase, the TOC content in the liquid phase, and the OD 600 and pH, and found that the transferred acclimated sludge could stably fix CO2 in a shake flask and convert it into organic matter. -1 ~10 -6 After multiple dilution, the plate was coated and cultured in a 30°C constant temperature incubator for 3-5 days, and a single colony on the plate was picked for streaking separation to obtain a pure colony. After functional verification, a strain of Brucella WYQ-2 of the present invention was obtained, and the Brucella WYQ-2 can fix CO2.

[0082] (2) Identification of Brucella WYQ-2

[0083] like Figure 1 and Figure 2 As shown, the morphological characteristics of the Brucella WYQ-2 of the present invention are: the colonies are small dot-shaped, white, opaque, full in shape, without flagella, easy to pick up, and the bacterial lawn grows along the streaks.

[0084] The Brucella WYQ-2 CO2-fixed strain obtained by screening, separation and purification was entrusted to Shanghai Sangon Biotechnology Co., Ltd. for PCR amplification and sequencing, and the 16SrRNA sequencing results were as follows (Genbank accession number is PQ628109):

[0085]

[0086] The sequencing results were uploaded to Ezbiocloud.net and compared with the standard strains on the website. The bacterial phylogenetic tree was constructed using the Neighbor-Joining method using MEGA-X software and evaluated using the Bootstrap method (repeated 1000 times). The constructed phylogenetic tree is shown in Figure 3 As shown, the strain WYQ-2 was determined to be Brucella, named Brucella WYQ-2, and deposited in China Center for Type Culture Collection on October 14, 2024, with a deposit number of CCTCC No: M 20242175, address: Wuhan University, Wuhan, China, Postal Code 430072.

[0087] Embodiment 3:

[0088] Expansion culture of Brucella WYQ-2 of the present invention

[0089] After the Brucella WYQ-2 is expanded and cultured, a bacterial solution containing Brucella WYQ-2 is obtained. The specific process is as follows:

[0090] Step 1) Slant culture:

[0091] The isolated and purified Brucella WYQ-2 was inoculated into the solid culture medium UPM and cultured at 30°C for 48-72 h to obtain slant bacteria.

[0092] The preparation method of the solid culture medium is as follows: 16g of Na2HPO4·12H2O, 6.8g of KH2PO4, 4g of CH3COONa, 1g of yeast powder, 0.002g of FeSO4, 0.0002g of (NH4)2MoO4, and 1mL of trace element solution are added into 800ml of distilled water to obtain solution A, 0.58 of MgSO4 and 0.067 of CaCl2 are dissolved into 200ml of distilled water to obtain solution B, solutions A and B are slowly stirred and mixed to obtain a culture medium, and then 100ml of liquid volume is added into a 250ml serum bottle, and 2g of agar is added to prepare a solid culture medium.

[0093] Step 2) Expand the culture:

[0094] From the slant bacteria on the slant in step 1), pick an inoculation loop of colonies and inoculate them on a plate for 48-72 h. Pick a single colony from the plate and inoculate it into the sterilized liquid culture medium. Incubate it in a constant temperature shaker (temperature: 30°C, speed: 160 r / min) for 24-48 h until the logarithmic growth phase. Centrifuge it at 4°C, 8000 r / min for 10 min, discard the supernatant, and then wash it three times with a sterilized inorganic salt solution. Finally, add 5-15 mL of the inorganic salt solution to the centrifuged bacteria and shake it completely to obtain the OD 600 =1 bacterial solution.

[0095] The inorganic salt solution in this embodiment 3 is a mixture of 4.5g / L Na2HPO4·12H2O, 1.0g / L KH2PO4, 2.5g / L (NH4)2SO4, 0.2g / L MgSO4·7H2O, 0.023g / L CaCl2, 1mL / L trace element mother solution, and 1mL / L vitamin mother solution.

[0096] Embodiment 4:

[0097] The bacterial agent prepared in Example 3 was inoculated into 100 mL of a sterilized inorganic salt medium containing 1 g of reduced iron powder. Before use, the inorganic salt medium was purged with 99.999% N2 for 30 min to remove dissolved oxygen in the medium. The initial OD value of Brucella WYQ-2 in the inorganic salt medium was adjusted. 600 =0.05, the initial pH of the inorganic salt medium was 6.8, and the volume fraction of CO2 introduced into each sample was 30 vol%. The samples were shaken and cultured in a constant temperature shaker at 30℃ and 160 r / min. Samples were taken at regular intervals to measure the gas phase CO2 concentration, liquid phase TOC content, and proteoglycan content in the reaction system. The results are as follows: Figure 4-1 to Figure 4-3 shown.

[0098] Embodiment 5:

[0099] The only difference between Example 4 and Example 5 is that the volume fraction of CO2 introduced into the sample is 35 vol%. The gas phase CO2 concentration, liquid phase TOC content, and proteoglycan content in the reaction system were measured. The results are as follows: Figure 4-1 to Figure 4-3 shown.

[0100] Embodiment 6:

[0101] The only difference between Example 6 and Example 5 is that the volume fraction of CO2 introduced into the sample is 38 vol%. The gas phase CO2 concentration, liquid phase TOC content, and proteoglycan content in the reaction system were measured. The results are as follows: Figure 4-1 to Figure 4-3 shown.

[0102] Embodiment 7:

[0103] The only difference between Example 4 and Example 5 is that the volume fraction of CO2 introduced into the sample is 66 vol%. The gas phase CO2 concentration, liquid phase TOC content, and proteoglycan content in the reaction system were measured. The results are as follows: Figure 4-1 to Figure 4-3 shown.

[0104] Comparative Example 1:

[0105] The difference between this comparative example 1 and example 5 is that the volume fraction of CO2 introduced into the sample is 13 vol%. The gas phase CO2 concentration, liquid phase TOC content, and proteoglycan content in the reaction system were measured. The results are as follows: Figure 4-1 to Figure 4-3 shown.

[0106] Comparative Example 2:

[0107] The difference between this comparative example 2 and example 5 is that the volume fraction of CO2 introduced into the sample is 23 vol%. The gas phase CO2 concentration, liquid phase TOC content, and proteoglycan content in the reaction system were measured. The results are as follows: Figure 4-1 to Figure 4-3 shown.

[0108] Comparative Example 3:

[0109] The only difference between Comparative Example 3 and Example 5 is that the bacterial agent prepared in Example 3 is not used in Comparative Example 3. Six identical samples are prepared, and the volume fractions of CO2 introduced into each sample are 13 vol%, 23 vol%, 30 vol%, 35 vol%, 38 vol% and 66 vol%, respectively. The gas phase CO2 concentration in the reaction system is measured, and the results are as follows: Figure 4-1 shown.

[0110] According to the results of Examples 1 to 7 and Comparative Examples 1 to 3, Figure 4-1 to Figure 4-3 It can be seen that: within the volume fraction range of 13~66 vol%, Brucella WYQ-2 can grow and reproduce by fixing CO2 as a carbon source and energy source. The gas phase CO2 concentrations of all blank groups and experimental groups showed a downward trend. The reason for the decrease in the concentration in the blank group may be that, on the one hand, part of the CO2 dissolves in water as the culture time increases, and on the other hand, it is because the Fe in the system reacts with CO2 to form FeCO3. At low CO2 concentrations of 13~23 vol%, the CO2 concentration in the blank group decreased faster than that in the experimental group with added strain WYQ-2; but at medium and high CO2 concentrations of 30~38 vol%, the gas phase CO2 concentration in the experimental group with added strain WYQ-2 decreased significantly faster than the blank control group as the culture time increased. Figure 4-2 As shown in the figure, the TOC content in the liquid phase increases with the increase of CO2 concentration, proving that WYQ-2 can effectively fix inorganic carbon and convert it into organic matter. Figure 4-3 As shown in the figure, WYQ-2 has a higher proteoglycan content when the CO2 concentration is 35 vol%, which proves that WYQ-2 grows faster at this concentration, showing a higher biological growth rate. In the volume fraction range of 35~66 vol%, WYQ-2 can show a good fixation effect on CO2.

[0111] Embodiment 8:

[0112] The only difference between Example 8 and Example 5 is that the composition of the liquid culture medium of Example 8 is: 4.5 g / L Na2HPO4·12H2O, 1.0 g / L KH2PO4, 2.5 g / L (NH4)2SO4, 0.2 g / L MgSO4·7H2O, 0.023 g / L CaCl2, 1 mL / L trace element stock solution, and 1 mL / L vitamin stock solution.

[0113] The trace element mother solution includes: 1g / L FeSO4, 0.05g / L CuSO4, 0.014g / L H3BO3, 0.1g / L MnSO4·4H2O, 0.02g / L Na2MoO4·2H2O, and 0.02g / L CoCl2·6H2O.

[0114] The vitamin mother solution includes: 0.2g / L biotin VH, 0.2g / L folic acid VB, 1g / L pyridoxine hydrochloride VB6, 0.5g / L riboflavin, 0.45~0.55g / L vitamin B2, 0.5g / L ammonium sulfate VB1, 0.5g / L niacin, 0.5g / L pantothenic acid VB5, 0.01g / L B-12, 0.5g / L p-aminobenzoic acid and 0.5g / L lipoic acid.

[0115] Embodiment 9:

[0116] The bacterial agent prepared in Example 3 was inoculated into a sterilized 100 mL inorganic salt medium to obtain a sample. Before use, the inorganic salt medium was purged with 99.999% pure N2 for 20 min to remove dissolved oxygen in the inorganic salt medium. The initial OD of the experimental group was adjusted to 600 =0.05, the initial pH of the inorganic salt medium = 6.8, the content of reduced iron powder in the sample was 0.5g, and then 35 vol% CO2 was introduced, and the culture was shaken in a constant temperature shaker at 30℃ and 160r / min. Samples were taken after a certain period of time to determine the gas phase CO2 concentration, liquid phase TOC content and proteoglycan content in the reaction system. The results are shown in Figures 5-1 to 5-3 shown.

[0117] Embodiment 10:

[0118] The difference between Example 10 and Example 9 is that the content of reduced iron powder in the sample is 1g. The gas phase CO2 concentration, liquid phase TOC content, and proteoglycan content in the reaction system were measured. The results are as follows: Figure 5-1 to Figure 5-3 shown.

[0119] Embodiment 11:

[0120] The difference between Example 11 and Example 9 is that the content of reduced iron powder in the sample is 2 g. The gas phase CO2 concentration, liquid phase TOC content, and proteoglycan content in the reaction system were measured. The results are as follows: Figure 5-1 to Figure 5-3 shown.

[0121] Comparative Example 4:

[0122] The difference between this comparative example 4 and example 9 is that the content of reduced iron powder in the sample is 0.1 g. The gas phase CO2 concentration, liquid phase TOC content, and proteoglycan content in the reaction system were measured. The results are as follows: Figure 5-1 to Figure 5-3 shown.

[0123] Comparative Example 5:

[0124] The difference between this comparative example 5 and example 9 is that the content of reduced iron powder in the sample is 0.2 g. The gas phase CO2 concentration, liquid phase TOC content, and proteoglycan content in the reaction system were measured. The results are as follows: Figure 5-1 to Figure 5-3 shown.

[0125] Comparative Example 6:

[0126] The only difference between Comparative Example 6 and Example 5 is that the bacterial agent prepared in Example 9 is not used in Comparative Example 6. Six identical samples were prepared, and the contents of reduced iron powder in the samples were 0.1 g, 0.2 g, 0.5 g, 1 g, and 2 g, respectively. The gas phase CO2 concentration in the reaction system was measured, and the results were as follows: Figure 5-1 shown.

[0127] According to the results of Examples 9 to 11 and Comparative Examples 4 to 6, Figure 5-1 to Figure 5-3 It can be seen that when the amount of reduced iron powder added was 1 g, the gas phase CO2 concentration decreased fastest. When the amount of reduced iron powder added was 0.5~2 g, the proteoglycan content of strain WYQ-2 was higher, showing a higher biological growth rate. When the amount added was 1 g, the growth and reproduction rate was the fastest, corresponding to the fixation performance. The TOC content in the liquid phase increased with the increase in the amount of iron powder added. When the amount of iron powder added was 0.5~2 g, the TOC content was higher, corresponding to the fixation effect and growth of strain WYQ-2.

[0128] Embodiment 12:

[0129] The inorganic salt culture medium was adjusted to five samples with different pH values ​​of 5.0, 6.0, 7.0, 8.0 and 9.0 using 1 mol / L NaOH aqueous solution or 1 mol / L HCl aqueous solution; the bacterial agent prepared in Example 3 was inoculated into the sterilized culture medium containing 100 ml of inorganic salt solution containing 1 g of reduced iron powder, so that the initial bacterial cell concentration was OD 600 =0.05, the volume fraction of CO2 introduced was 35 vol%. The samples were shaken and cultured in a constant temperature shaker at 30℃ and 160 r / min. Samples were taken at regular intervals to measure the gas phase CO2 concentration, proteoglycan content and liquid phase TOC content in the reaction system. The results are as follows: Figures 6-1 to 6-3 shown.

[0130] from Figures 6-1 to 6-3 It can be seen that strain WYQ-2 can fix CO2 in the pH range of 5-9, and has a relatively excellent fixation effect at pH 7-9. This may be related to the environment of the early domestication and screening of CO2-fixing bacteria. At the same time, the growth amount and TOC value of Brucella WYQ-2 are also consistent with the CO2 fixation effect.

[0131] The raw materials and equipment used in the present invention, unless otherwise specified, are all commonly used raw materials and equipment in the art; the methods used in the present invention, unless otherwise specified, are all conventional methods in the art.

[0132] The present invention is described by the above specific embodiments, and those skilled in the art should understand that various changes and equivalent substitutions can be made to the present invention without departing from the scope of the present invention. The parts not described in detail in the present specification are well-known technologies to those skilled in the art. In addition, various modifications can be made to the present invention for specific situations or specific circumstances without departing from the scope of the present use. Therefore, the present invention is not limited to the specific embodiments disclosed, but should include all implementation methods falling within the scope of the claims of the present invention.

Claims

1. A Brucella species ( Brucella sp. )WYQ-2, characterized in that The microbial classification of the Brucella WYQ-2 is named Brucella ( Brucella sp. ), deposited in China Center for Type Culture Collection, with the deposit number CCTCCNo: M 20242175 and the deposit date being October 14, 2024.

2. A bacterial agent, characterized in that The bacterial agent is a bacterial suspension comprising the Brucella WYQ-2 described in claim 1.

3. The use of Brucella WYQ-2 as claimed in claim 1, characterized in that The Brucella WYQ-2 is used to fix carbon dioxide under the condition that reduced iron powder is used as an electron donor to perform a fixation reaction.

4. A use of the bacterial agent as claimed in claim 2, characterized in that: The bacterial agent is used to fix carbon dioxide under the condition that reduced iron powder serves as an electron donor to carry out a fixation reaction.

5. The use according to claim 3 or 4, characterized in that The volume fraction of the initial carbon dioxide concentration of the fixing reaction is 30 vol% to 66 vol%, the concentration of the initial reduced iron powder is 0.1 to 2 g / 100 mL, and the pH of the fixing reaction is 5 to 9.

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