Comamonas testosteroni G314 and application thereof

Abstract

The invention belongs to the field of environmental microbiology, and particularly relates to comamonas testosteroni (Comamonas testosteroni) G314 and application of the comamonas testosteroni G314. The technical key points are as follows: the comamonas testosteroni provided by the invention is preserved in China General Microbiological Culture Collection Center of Institute of Microbiology, Chinese Academy of Sciences, No. 3, No.1 yard, West Beichen Road, Chaoyang District, Beijing, China; the preservation number is CGMCC (China General Microbiological Culture Collection Center) No.32129, and the preservation time is September 30, 2024. According to the comamonas testosteroni provided by the invention, the capabilities of synthesizing the PHA by utilizing different organic acid carbon sources are different, and the type of the PHA can be regulated and controlled by regulating and controlling the composition of the volatile fatty acid, so that the performance of the PHA is improved, a new strain can be provided for efficiently producing the PHA, and the production cost of the PHA can also be remarkably reduced.

Description

Technical Field

[0001] The invention belongs to the field of environmental microbiology, and in particular relates to Comamonas testosteroni G314 and application thereof. Background Art

[0002] Polyhydroxyalkanoates (PHA) have physical properties comparable to those of traditional plastics, and also have biodegradability and biocompatibility that traditional plastics do not have, making them ideal alternative materials. However, the current production of PHA is limited and the production cost is high, which greatly restricts the commercial application of PHA.

[0003] At present, more than 300 kinds of microorganisms have been found in nature that can synthesize PHA. The PHA production capacity and required conditions of different species of microorganisms are different. Most of the known PHA synthesizing bacteria are aerobic microorganisms. When producing PHA under aerobic conditions, most of the carbon source enters the tricarboxylic acid cycle and is eventually converted into CO2. The remaining carbon source is used for strain growth and PHA synthesis. This process not only leads to a significant reduction in the carbon source converted into PHA, but also increases the cost of aeration.

[0004] Under hypoxic or anaerobic conditions, there is little or no carbon source for the CO2 produced by respiratory metabolism, which can significantly improve the utilization of carbon sources and obtain a higher amount of PHA synthesis. However, current research can only transform strains through gene editing technology to enable them to synthesize PHA under hypoxic conditions to increase PHA production. However, strains modified by gene editing technology can only synthesize PHA of a specific configuration, and the production efficiency is low. Summary of the invention

[0005] The present invention aims to provide Comamonas testosteroni G314, which can be cultured by using different types of carbon sources to regulate the configuration of polyhydroxyalkanoates, and the carbon source for synthesizing polyhydroxyalkanoates can be obtained by anaerobic fermentation of organic wastes such as kitchen waste or residual sludge, thereby achieving resource utilization of organic wastes and reduction of production costs of polyhydroxyalkanoates at the same time.

[0006] The above technical objectives of the present invention are achieved through the following technical solutions:

[0007] The present invention provides a Comamonas testosteroni G314, a strain screened from industrial sludge, which can anaerobically synthesize polyhydroxyalkanoates without using gene editing technology. The strain is deposited in the General Biological Center of the China Microbiological Culture Collection Committee of the Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, China; the deposit number is CGMCC No.32129, and the deposit time is: September 30, 2024.

[0008] The present invention provides an application of Comamonas testosteroni G314, which is to use Comamonas testosteroni G314 to synthesize polyhydroxyalkanoates under anoxic or anaerobic conditions. No oxygenation equipment is required during the fermentation process, which greatly simplifies the requirements for fermentation equipment and reduces the cost of equipment investment and maintenance. At the same time, the anaerobic environment also helps to inhibit the growth of other aerobic contaminating bacteria, thereby improving the purity and reliability of production.

[0009] Furthermore, the polyhydroxyalkanoate is poly(3-hydroxybutyrate) or poly(3-hydroxybutyrate-co-3-hydroxyvalerate). Poly(3-hydroxybutyrate) or poly(3-hydroxybutyrate-co-3-hydroxyvalerate) are both biodegradable materials, which can be completely degraded by microorganisms in natural environments (such as soil, oceans, etc.) to produce harmless products such as carbon dioxide and water. Therefore, they will not cause white pollution and are an environmentally friendly green material that replaces traditional petroleum-based plastics. In addition, the raw materials of PHA in the present invention can be derived from organic waste, which makes its production process more sustainable, reduces dependence on petrochemical raw materials, and has the dual function of treating waste.

[0010] Poly(3-hydroxybutyrate) itself has high crystallinity, so it shows high rigidity and strength in mechanical properties. It is suitable as a material to replace certain hard plastics and is used in products that require wear resistance or high rigidity. By adding 3-hydroxyvaleric acid (3HV) units to poly(3-hydroxybutyrate) to form a copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate), the flexibility and ductility of the material can be significantly improved. The introduction of 3HV reduces the crystallinity of the polymer, making the material softer and more elastic, suitable for applications that require a certain degree of flexibility, such as packaging materials and medical products.

[0011] Furthermore, the Comamonas testosteroni is used to synthesize polyhydroxyalkanoates under anoxic or anaerobic conditions, comprising the following steps:

[0012] The Comamonas testosteroni was inoculated into LB medium for cultivation to obtain seed liquid;

[0013] After inoculating 1% to 10% (v / v) of seed liquid in the fermentation medium, the fermentation culture is carried out on a constant temperature shaker to obtain a fermentation liquid; the present invention can adjust the inoculation amount according to the fermentation scale and process requirements, thereby optimizing the production efficiency; the fermentation culture on a constant temperature shaker can ensure the precise control of the temperature and the shaker speed, thereby providing a stable environment for bacterial growth and PHA synthesis. This helps to improve the reproducibility and efficiency of the fermentation process and reduce the impact caused by environmental fluctuations.

[0014] The fermentation liquid is centrifuged to obtain the cells of Comamonas testosteroni, and the dried cells of Comamonas testosteroni are obtained after freeze drying; the fermented cells can be effectively collected after centrifugation. The centrifugation step is fast and efficient, and is suitable for large-scale production. The dried cells obtained by freeze drying have good stability and can maximize the structure and purity of PHA in the cells. This drying method will not destroy PHA, ensuring that high-quality PHA is obtained in the subsequent extraction process.

[0015] The dried cells of Comamonas testosteroni were mixed with chloroform and acidified methanol containing benzoic acid and reacted at 105°C for 6 hours; chloroform as an organic solvent can effectively dissolve PHA, while acidified methanol can help destroy the cell wall and release PHA. The extraction process has been optimized in terms of reaction time and temperature, and can efficiently extract PHA while avoiding PHA degradation caused by excessive high temperature, thereby obtaining high-purity PHA.

[0016] Distilled water is added, and the chloroform layer is transferred to anhydrous sodium sulfate for drying to obtain polyhydroxyalkanoate. After adding distilled water, the chloroform layer is transferred to anhydrous sodium sulfate for drying, which can effectively remove moisture in the organic solvent and improve the purity of PHA.

[0017] Furthermore, the initial pH value of the fermentation culture is 5 to 9, the fermentation culture temperature is 25 to 40°C, the rotation speed of the constant temperature shaker is 100 to 200 rpm, and the seed liquid culture time is 10 to 24 hours. This flexibility allows the fermentation conditions to be adjusted to suit different environments and culture objectives, thereby improving the fermentation efficiency.

[0018] Further, the initial pH value of the fermentation culture is 8, the temperature of the fermentation culture is 258°C, the speed of the constant temperature shaker is 180rpm, the seed liquid culture time is 22h, and the seed liquid inoculation amount is 5-10% (v / v), preferably 10% (v / v). pH 8 as an initial value can provide a suitable alkaline environment for the metabolism of Comamonas testosteroni and promote the utilization of carbon sources. Selecting a higher seed liquid inoculation amount (5-10%, preferably 10%) can accelerate the increase in the number of bacteria in the early stage of fermentation, shorten the production cycle, reduce the risk of contamination, and ensure the robustness and high yield of the fermentation.

[0019] Furthermore, the carbon source in the fermentation medium is any one or more of glucose, sucrose, glycerol or volatile fatty acids. The diversity of carbon sources increases the flexibility and adaptability of fermentation, and the most suitable carbon source can be selected according to the availability and cost of raw materials. For example, glucose and sucrose can provide rapidly available energy, while glycerol and volatile fatty acids are suitable for promoting PHA synthesis under specific conditions. In addition, the use of a combination of multiple carbon sources can optimize bacterial growth and PHA accumulation and increase the final yield.

[0020] Furthermore, the volatile fatty acid is any one or more of sodium acetate, sodium propionate, butyric acid or valeric acid. The metabolic pathways of these fatty acids directly promote the accumulation of PHA synthetic monomers. At the same time, using different types of volatile fatty acids can also regulate the composition of PHA, so that the final product has better properties, such as flexibility and ductility.

[0021] Furthermore, the composition of the fermentation medium includes: carbon source 3.3g / L, KH2PO4 0.08g / L, NH4Cl 0.15g / L, MgSO4 0.1g / L, CaCl2·2H2O 0.01g / L, FeCl3·6H2O1.5mg / L and trace elements 1.0mL / L. Among them, the concentration of the carbon source is moderate, which can promote growth and achieve the optimal ratio of PHA accumulation. KH2PO4 provides phosphorus to stabilize the pH of the culture medium; NH4Cl provides nitrogen source to promote protein and cell proliferation; MgSO4 and CaCl2 provide important ions to maintain the ion balance inside and outside the cell; FeCl3 acts as a cofactor to promote the activity of multiple enzymes. The composition of the culture medium has been optimized to ensure the required nutritional balance for the growth of Comamonas testosteroni and PHA synthesis, maximize cell proliferation and PHA synthesis efficiency, and reduce the waste of nutrients.

[0022] Furthermore, trace elements include: CuSO4·5H2O 0.01g / L, ZnSO4·H2O 0.1g / L, MnCl2·4H2O0.03g / L, H3BO3 0.3g / L, CoCl2·6H2O 0.2g / L, NiCl2·6H2O 0.02g / L and NaMoO4·2H2O 0.03g / L. The supplementation of trace elements can effectively improve the growth and metabolic activity of bacteria. Among them, Cu and Zn play an important role in redox reactions and enzyme activity; Mn contributes to protein synthesis and enzyme stability; Co is a component of vitamin B12 and is essential for cell metabolism; Ni and Mo are related to specific enzyme actions, especially for the synthesis pathway of PHA. The precise addition of these trace elements ensures the metabolic efficiency of Comamonas testosteroni and the synthesis rate of PHA, greatly improving the stability of fermentation production and the yield of PHA.

[0023] In summary, the present invention has the following beneficial effects:

[0024] A kind of Comamonas testosteroni G314 provided by the present invention is cultivated with sodium acetate under anoxic conditions and anaerobic conditions, and the PHB accumulation amount obtained is 69.0% and 63.7% of the cell dry weight respectively, which is higher than the PHB accumulation amount under aerobic conditions, meaning that the carbon utilization efficiency of the thalline in the metabolic process is higher, indicating that under anoxic or anaerobic environment, the carbon source is more preferentially used for PHA synthesis rather than other metabolic pathways. Since anoxic and anaerobic conditions do not require forced oxygen supply, energy consumption can be greatly reduced, especially in large-scale industrial fermentation, this reduction can not only save energy costs, but also reduce the investment and maintenance of process equipment (such as oxygenation system), and significantly improve economic benefits. At the same time, lower oxygen demand also reduces the impact on the environment, which is in line with the production concept of green environmental protection.

[0025] In addition, when the strain was cultured with valeric acid as the carbon source, the highest PHBV accumulation was obtained under anaerobic conditions, reaching 74.3% of the cell dry weight, of which HV accounted for as high as 91.1%. This high HV proportion means that the obtained PHA is mainly poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), and the flexibility is significantly improved. The increase in HV content can significantly improve the mechanical properties of the polymer, such as reducing brittleness, increasing ductility and elasticity, making this material more suitable for applications requiring high flexibility and plasticity, such as biomedical materials, packaging films, etc.

[0026] The present invention can regulate the composition and properties of PHA by changing the type of carbon source, for example, synthesizing more PHB or PHBV under different environments and adjusting the ratio of HV. This method of regulating the configuration of PHA by carbon source can provide diversified material options for industrial applications, allowing the physical and chemical properties of PHA to be flexibly changed according to different needs, thereby expanding its application range, for example, in multiple fields such as biodegradable plastics, food packaging, and biomedical devices to be better adapted.

[0027] Biomaterial Deposit

[0028] Comamonas testosteroni G314 was deposited at the General Biological Center of China Microbiological Culture Collection, Institute of Microbiology, Chinese Academy of Sciences on September 30, 2024, with the deposit number CGMCC No.32129, and the deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] The present invention is further described below in conjunction with the accompanying drawings and embodiments.

[0030] Figure 1 This is a phylogenetic tree constructed from the 16S rRNA sequence of Comamonas testosteroni G314.

[0031] Figure 2 The growth and PHA production of Comamonas testosteroni G314 under different oxygen conditions using sodium acetate as the carbon source.

[0032] Figure 3 The growth and PHA synthesis of Comamonas testosteroni G314 using sodium propionate as carbon source.

[0033] Figure 4 The growth and PHA synthesis of Comamonas testosteroni G314 using butyrate as carbon source.

[0034] Figure 5 The growth and PHA synthesis of Comamonas testosteroni G314 using valeric acid as the carbon source. DETAILED DESCRIPTION

[0035] In order to further explain the technical means and effects adopted by the present invention to achieve the predetermined purpose of the invention, a Comamonas testosteroni G314 and its application proposed by the present invention, its specific implementation method, characteristics and effects are described in detail as follows.

[0036] The experimental methods in the following examples, unless otherwise specified, are all conventional methods, and are performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials, reagents, etc. used in the following examples, unless otherwise specified, can all be obtained from commercial channels.

[0037] At present, PHA can be divided into homopolymers or copolymers according to the monomer configuration of the polymer. Common homopolymers include poly (3-hydroxybutyrate) (P (3HB), abbreviated as PHB in the text), poly (4-hydroxybutyrate) (P (4HB)), and copolymers include poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (P (3HB-co-3HV), abbreviated as PHBV in the text). The physical and chemical properties of PHA depend on the chemical and structural composition of the monomer units. Compared with homopolymers, the mechanical properties of copolymers are greatly improved. With the increase of monomer content in the copolymer, the material has a lower Young's modulus, higher elongation at break and better mechanical properties. Carbon source is the most important culture parameter in the PHA synthesis process, which greatly affects the yield and type of PHA synthesized by microorganisms.

[0038] Therefore, this specific embodiment screened a microorganism that can synthesize PHA under anoxic / anaerobic conditions. The ability to synthesize PHA using different organic acid carbon sources is different. The type of PHA can be regulated by regulating the composition of volatile fatty acids, thereby improving the performance of PHA. This can not only provide new strains for the efficient production of PHA, but also significantly reduce the production cost of PHA, especially the high cost caused by aeration during large-scale production.

[0039] Example 1: Screening and identification of Comamonas testosteroni G314

[0040] The Comamonas testosteroni G314 provided in this embodiment is deposited in the General Biological Center of the China Microbiological Culture Collection Administration Committee, Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, China; the deposit number is CGMCC No.32129, and the deposit time is: September 30, 2024.

[0041] The culture medium used included:

[0042] Luria-Bertani (LB) medium: peptone 10.0 g / L, yeast powder 5.0 g / L, NaCl 10.0 g / L, take 20 g and add ultrapure water to 1 L. Sterilize in an autoclave at 121°C for 20 min before use. You can also add 2% agar powder to prepare LB solid medium.

[0043] Minimal medium (MM) culture medium: KH2PO4 (1.5g / L); NH4Cl (0.2g / L); MgSO4 (0.1g / L); Na2HPO4 (3.5g / L); CaCl2·2H2O (0.01g / L); ammonium ferric citrate (0.06g / L); 1.0mL / L trace elements (MnCl2·4H2O (0.03g / L); ZnSO4·H2O (0.1g / L); H3BO3 (0.3g / L); CoCl2·6H2O (0.2g / L); CuSO4·5H2O (0.01g / L); NaMoO4·2H2O (0.03g / L); NiCl2·6H2O (0.02g / L)), and 4g / L sodium acetate.

[0044] The pH value of the culture medium was adjusted to 7.0 with 2 M NaOH, and calcium salts and trace elements were added after filtering with a syringe (0.22 μm) before inoculation. The remaining components were sterilized in an autoclave at 121° C. for 20 min before use.

[0045] The separation and screening specifically includes the following steps:

[0046] (1) The strain was screened from the sludge of a sewage treatment plant in Shenzhen. 10 g of the sludge sample from a sewage treatment plant in Shenzhen was placed in a 250 mL Erlenmeyer flask, 90 mL of sterile water was added, and the suspension was shaken at 30 °C and 150 rpm for 30 min to obtain a suspension. Then, a gradient dilution was performed and 10 -2 ~10 -6 The dilutions were spread on LB agar plates (with 50 μg / mL Nile blue added), 3 plates were spread for each gradient, 100 μL of the diluted sample suspension was spread on each plate, the plates were sealed and numbered, and then placed in a 30°C incubator for 48 h.

[0047] Single fluorescent colonies were picked from different plates for separation and purification, and pure bacteria were obtained after repeated zoning and streaking.

[0048] The obtained single colony was inoculated into MM medium for rescreening, and cultured in a shaker at 30°C and 150rpm for 72h. Finally, the strain with the highest PHA accumulation was screened out, and Comamonas testosteroni G314 was obtained. It was stored in 25% glycerol and placed in a -20°C refrigerator.

[0049] Identification of strain 16SrDNA: The isolated strain G314 was classified and identified using the 16s rRNA gene sequence method. The measured 16s rRNA gene sequence was compared with the sequence in GenBank. The results showed that the homology of strain G314 with Comamonas testosteroni GCF_014076415.1 reached 100%, and it had a similarity of 99.8% with Comamonas phosphati. Therefore, it was determined that the strain was a species of Comamonas, specifically Comamonas testosteroni. The phylogenetic tree of Comamonas testosteroni G314 can be found in Figure 1 .

[0050] Example 2: Determination of the ability of Comamonas testosteroni G314 to produce PHA under different oxygen conditions

[0051] Production of PHA: The strain Comamonas testosteroni G314 obtained in Example 1 was re-streaked onto LB solid medium and cultured in a constant temperature incubator at 30° C. for 24 h to obtain an activated strain;

[0052] A single colony was hooked from the LB plate into a high-temperature sterilized LB liquid culture medium, and cultured at 25°C and 180rpm for 18h to obtain a seed culture solution; according to an inoculation amount of 10%, the seed solution was inoculated into a 500mL conical flask containing 200mL of sterilized PHA-producing liquid culture medium, and cultured at a constant temperature of 25°C and 180rpm for 72h.

[0053] Extraction of PHA: The fermentation broth was transferred to a 50 mL centrifuge tube, centrifuged at 7800 rpm for 15 min, and the supernatant was removed; the cell pellet was pre-frozen at -80 ° C for 12 h, and then freeze-dried to obtain a freeze-dried sample of the strain; 20 mg of the freeze-dried sample of the strain was weighed and placed in a 10 mL esterification tube, and in a fume hood, 2 mL of chloroform and 2 mL of acidified methanol solution containing 0.2% benzoic acid were added to the esterification tube, mixed evenly, and then placed in an oven at 105 ° C for 6 h for digestion;

[0054] After the reaction was completed, the mixture was cooled to room temperature, and then 1 mL of deionized water was added to the esterification tube. After being fully mixed for 30 seconds, the mixture was allowed to stand for stratification. The lower organic phase was transferred to 0.75 g of anhydrous sodium sulfate for drying, and then transferred to a gas chromatography analysis bottle.

[0055] Determination of PHA: The above samples were analyzed by an Agilent 7890AGC-FID system (Agilent Technologies), which used a DB-5 column with a length of 30m, an inner diameter of 0.25mm, and a film thickness of 0.25μm. The injection port temperature was set to 200°C, the split ratio was 25:1, and the injection volume was 1μL. The temperature program was set to 40°C, maintained for 3min, then increased to 140°C at 10°C / min, maintained for 2min, and then increased to 200°C at a rate of 20°C / min. Hydrogen was used as the carrier gas with a flow rate of 40mL / min and an air flow rate of 450mL / min. Purchased PHB and PHBV (Sigma-Aldrich) were used as external standards.

[0056] PHA content (%) = PHA concentration (g / L) / CDW (g / L) × 100%,

[0057] CDW is the cell dry weight of the strain.

[0058] Results Figure 2 Although strain G314 grew slowly in the first 24 hours of culture under anaerobic conditions, it maintained a good growth trend in any oxygen environment after 24 hours. After 48 hours of culture, the PHB accumulation of the strain under anoxic and anaerobic conditions was always higher than that under aerobic conditions. This shows that anaerobic conditions not only do not inhibit the growth and reproduction of strain G314, but also promote the synthesis of PHB by the strain.

[0059] Example 3: Determination of the ability of Comamonas testosteroni G314 to produce PHA using volatile fatty acids

[0060] The strain Comamonas testosteroni G314 was inoculated into LB medium and cultured for 18 hours to obtain seed liquid, which was inoculated into a 500 mL conical flask containing 200 mL fermentation medium at a 10% inoculation amount, and cultured anaerobically on a constant temperature shaker at 25°C and 180 rpm for 72 hours. The fermentation medium components: volatile fatty acids (3.3 g / L), KH2PO4 (0.08 g / L), NH4Cl (0.15 g / L), MgSO4 (0.1 g / L), CaCl2·2H2O (0.01 g / L), FeCl3·6H2O (1.5 mg / L) and 1.0 mL / L of trace elements.

[0061] After fermentation started, the carbon source was supplemented every 24 hours. The extraction and determination methods of PHA were the same as those in Example 2. The results of PHA accumulation were shown in Figure 3 , 4 、5.

[0062] The results showed that when the strain G314 used sodium propionate as the carbon source, that is, when the volatile fatty acids selected sodium propionate, Figure 3 , cell dry weight and PHBV production gradually increased with time, reaching the maximum at 48h, which were 1.14g / L and 0.55g / L, respectively. At this time, PHBV accumulation reached 48.3% of cell dry weight, of which HV monomer accounted for 14.3%.

[0063] When strain G314 uses butyrate as the carbon source, i.e., when the volatile fatty acid selects butyrate, Figure 4 The cell dry weight and PHB production increased first and then gradually decreased with time, reaching the maximum at 24h, which were 1.23g / L and 0.81g / L, respectively. At this time, the PHB accumulation reached 58.2% of the cell dry weight.

[0064] When strain G314 uses valeric acid as the carbon source, that is, when volatile fatty acids select valeric acid, Figure 5 The cell dry weight and PHBV production increased first and then decreased slightly over time, reaching their maximum values ​​of 1.60 g / L and 1.20 g / L, respectively, at 48 h. At this time, the PHBV accumulation also reached its maximum value, which was 74.3% of the cell dry weight, of which HV monomer accounted for as high as 91.1%.

[0065] It can be seen that when acetic acid and butyric acid are used as carbon sources, the strain mainly synthesizes HB monomers, and the content of synthesized HV monomers is very small and almost negligible. However, when propionic acid or valeric acid is used as the carbon source, the content of HV monomers synthesized by the strain increases significantly, especially when valeric acid is used as the carbon source, which can significantly promote the synthesis of HV monomers. This indicates that valeric acid is more suitable than propionic acid for regulating the proportion of HV monomers in PHBV.

[0066] The above description is only a preferred embodiment of the present invention and does not constitute any form of limitation to the present invention. Although the present invention has been presented as a preferred embodiment as above, it is not intended to limit the present invention. Any technician familiar with the profession can make some changes or modify the technical contents disclosed above into equivalent embodiments without departing from the scope of the technical solution of the present invention. However, any simple modification, equivalent change and modification made to the above embodiments based on the technical essence of the present invention without departing from the content of the technical solution of the present invention still falls within the scope of the technical solution of the present invention.

Claims

1. A Comamonas testosteroni G314, characterized in that It is deposited in the General Biological Center of China Microbiological Culture Collection Committee, Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, China; the deposit number is CGMCC No.32129, and the deposit date is September 30, 2024.

2. The use of Comamonas testosteroni G314 according to claim 1, characterized in that: The Comamonas testosteroni is used to synthesize polyhydroxyalkanoates under anoxic or anaerobic conditions.

3. The use of Comamonas testosteroni G314 according to claim 2, characterized in that: The polyhydroxyalkanoate is poly(3-hydroxybutyrate) or poly(3-hydroxybutyrate-co-3-hydroxyvalerate).

4. The use of Comamonas testosteroni G314 according to claim 2, characterized in that: The method of using the Comamonas testosteroni to synthesize polyhydroxyalkanoates under anoxic or anaerobic conditions comprises the following steps: The Comamonas testosteroni was inoculated into LB medium for cultivation to obtain seed liquid; After inoculating 1% to 10% (v / v) of seed liquid into a fermentation medium, fermentation and culturing are carried out on a constant temperature shaker to obtain a fermentation liquid; Centrifuging the fermentation liquid to obtain the bacterial cells of Comamonas testosteroni, and freeze-drying to obtain the dry bacterial cells of Comamonas testosteroni; The dried cells of Comamonas testosteroni were mixed with chloroform and acidified methanol containing 0.2% benzoic acid, and reacted at 100-105°C for 4-6 hours; Distilled water was added, and the chloroform layer was transferred to anhydrous sodium sulfate and dried to obtain polyhydroxyalkanoate.

5. The use of Comamonas testosteroni G314 according to claim 4, characterized in that: The initial pH value of the fermentation culture is 5 to 9, the fermentation culture temperature is 25 to 40° C., the rotation speed of the constant temperature shaker is 100 to 200 rpm, and the seed liquid culture time is 10 to 24 hours.

6. The use of Comamonas testosteroni G314 according to claim 4, characterized in that: The carbon source in the fermentation medium is any one or more of glucose, sucrose, glycerol or volatile fatty acids.

7. The use of Comamonas testosteroni G314 according to claim 6, characterized in that: The volatile fatty acid is any one or more of sodium acetate, sodium propionate, butyric acid or valeric acid.

8. The use of Comamonas testosteroni G314 according to claim 4, characterized in that: The fermentation medium comprises: 2-7 g / L of carbon source, 0.08-0.17 g / L of KH2PO4, 0.15-0.56 g / L of NH4Cl, 0.08-0.12 g / L of MgSO4, 0.008-0.012 g / L of CaCl2·2H2O, 1.2-1.8 mg / L of FeCl3·6H2O and 0.8-1.2 mL / L of trace elements.

9. The use of Comamonas testosteroni G314 according to claim 8, characterized in that: The trace elements include: CuSO4·5H2O 0.008-0.012 g / L, ZnSO4·H2O 0.08-0.12 g / L, MnCl2·4H2O 0.025-0.035 g / L, H3BO3 0.25-0.35 g / L, CoCl2·6H2O 0.16-0.24 g / L, NiCl2·6H2O 0.02-0.024 g / L and NaMoO4·2H2O 0.025-0.035 g / L.

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