Method for preparing methylselenoic acid using Delftia acidovorans R25

Abstract

The invention relates to the technical field of microbiology and selenium-contaminated environmental restoration, and provides a method for preparing methylselenic acid using Delftia acidovorans R25. The strain R25 can efficiently reduce selenite to elemental selenium, and efficiently convert selenite and elemental selenium into volatile selenium, and a method for converting selenite and elemental selenium in water into volatile selenium and removing them using the strain R25 is established, and the volatile selenium can be converted into methylselenic acid at the same time. The strain R25 provided by the present invention and the method for preparing methylselenic acid using the strain R25 can be used for bioremediation of selenium-contaminated environments and recovery and resource utilization of selenium.

Description

Technical Field

[0001] The invention relates to the technical field of microbiology and selenium-polluted environment restoration, and in particular to a method for preparing methylselenoic acid by utilizing Delftia acidovorans R25. Background Art

[0002] Selenium is an essential trace element for humans and animals, and its safe intake range is narrow. Selenium deficiency can reduce human immunity, and severe deficiency can even cause Keshan disease, Kashin-Beck disease and other diseases. Excessive intake of selenium also poses a risk of poisoning. Excessive selenium content in water and soil environments can cause toxic effects on organisms, and as it accumulates and amplifies in the food chain, it poses risks to the entire ecosystem and human health.

[0003] Industrial activities such as mining, metal smelting, and selenium product processing will discharge high-concentration selenium-containing wastewater, which needs to be treated before discharge to reduce environmental pollution. At present, the treatment of selenium-contaminated wastewater includes physical, chemical and biological methods. Among them, the physical and chemical methods have complex procedures, large engineering workload, high cost and are prone to secondary pollution. The biological treatment method is low-cost, environmentally friendly, and can treat multiple pollutants at the same time. It has been reported that a variety of microorganisms can convert selenate [Se(VI)] and selenite [Se(IV)] into volatile selenium compounds such as dimethyl selenium (DMSe), dimethyl diselenide (DMDSe), dimethyl selenyl sulfide (DMSeS), hydrogen selenide (H2Se) and methyl selenol (MeSeH) through the reduction and methylation of selenium (Kagami, Narita et al. 2013). The use of selenium volatilization microorganisms can remove selenium contamination in the environment and recycle selenium by collecting volatile selenium compounds. Studies have found that selenium compounds such as MeSeCys (selenomethylselenocysteine) and MSA (methylselenic acid) have anti-cancer activity. Among them, methylselenic acid (MSA) is very excellent in anti-tumor and has selective cytotoxicity. Appropriate concentrations of MSA can induce endoplasmic reticulum stress and even death of tumor cells without affecting the survival of normal cells (Varlamova and Turovsky 2021).

[0004] Delftia belongs to the family Comamonadaceae and can be used for bioremediation of pollutants such as microcystins (Zhou Jie, Yan Hai et al. 2006), polyethylene terephthalate (PET) (Liu, Xu et al. 2018), sodium dodecyl sulfate (SDS) (Yilmaz and Icgen 2014), and per- and polyfluoroalkyl compounds (Harris, Coon et al. 2022). At present, there are no reports on the production of volatile selenium by Delftia for selenium bioremediation and recycling of selenium resources. Delftia is used to convert selenium compounds in wastewater into volatile selenium, and then the volatile selenium is recovered by tail gas capture device and reprocessed into selenium products, which not only achieves the treatment of selenium-contaminated wastewater, but also facilitates the recycling and reuse of selenium resources. Summary of the invention

[0005] The purpose of the present invention is to provide a strain of Delftia sp. R25 capable of synthesizing volatile selenium and application thereof.

[0006] Another object of the present invention is to provide a method for preparing methylselenoic acid using Delftia acidovorans R25.

[0007] In order to achieve the purpose of the present invention, in the first aspect, the present invention provides a strain of Delftia acidovorans isolated and purified from high-selenium environment soil, classified and named Delftia as Delftiasp.R25, which has been deposited in the General Microbiology Center of China Culture Collection Administration, No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, Postal Code 100101, Deposit Number CGMCC No.25434, Deposit Date July 29, 2022.

[0008] In a second aspect, the present invention provides a bacterial agent containing the Delftia sp.

[0009] In a third aspect, the present invention provides a method for biosynthesizing volatile selenium, the method comprising: inoculating the Delftia sp. into a selenium-containing culture medium for culturing;

[0010] Wherein, the selenium in the culture medium is inorganic selenium and / or elemental selenium.

[0011] The inorganic selenium includes selenites, such as sodium selenite.

[0012] When the selenium in the culture medium is sodium selenite,

[0013] The concentration of sodium selenite in the liquid culture medium is 1-100 mM (in terms of selenium);

[0014] The concentration of sodium selenite in the solid culture medium is 1-100 mM (in terms of selenium).

[0015] When the selenium in the culture medium is elemental selenium,

[0016] The concentration of elemental selenium in the liquid culture medium is 1-300 mM (in terms of selenium);

[0017] The concentration of elemental selenium in the solid culture medium is 1-300 mM (in terms of selenium).

[0018] In the present invention, the volatile selenium includes but is not limited to DMSeO2 (dimethyl selenone), DMDSe (dimethyl diselenide), DMDSSe (dimethylselenodisulfide) and DMDSeS (dimethyl diselenylsulfide).

[0019] In a fourth aspect, the present invention provides the use of the Delftia bacteria in the bioremediation of selenium-contaminated water and soil.

[0020] Specifically, the Delftia bacteria or its bacterial agent is brought into contact with selenium-contaminated water and soil to convert inorganic selenium and / or elemental selenium therein into volatile selenium, so as to achieve the purpose of biological removal of selenium.

[0021] Furthermore, the application also includes a step of recovering the volatile selenium.

[0022] In a fifth aspect, the present invention provides a method for preparing methylselenic acid using Delftia acidovorans R25, wherein the volatile selenium obtained by the method is contacted with concentrated nitric acid to cause an oxidation reaction, and the methylselenic acid is collected from the reaction products.

[0023] By means of the above technical solution, the present invention has at least the following advantages and beneficial effects:

[0024] The Delftia acidovorans R25 provided by the present invention can efficiently reduce selenite to elemental selenium, and efficiently convert selenite and elemental selenium into volatile selenium, and a method for converting selenite and elemental selenium in water into volatile selenium and removing them by using the strain R25 is established, and the volatile selenium can be converted into methylselenic acid at the same time. The strain R25 provided by the present invention and the method for preparing methylselenic acid by using the strain R25 can be used for bioremediation of selenium-contaminated environments and recovery and resource utilization of selenium. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 The figure is the 16S rRNA gene evolution tree of strain R25 of the present invention.

[0026] Figure 2 This is the whole genome phylogenetic tree of strain R25 of the present invention.

[0027] Figure 3 The figure shows the growth of strain R25 on a solid medium containing selenite in a preferred embodiment of the present invention.

[0028] Figure 4 The figure shows the growth of strain R25 in a preferred embodiment of the present invention in a liquid culture medium containing selenite.

[0029] Figure 5 This is a TEM photo of the strain R25 reducing selenite to synthesize nano-selenium in a preferred embodiment of the present invention.

[0030] Figure 6 The figure shows the growth of strain R25 in a preferred embodiment of the present invention on a solid culture medium containing elemental selenium.

[0031] Figure 7 The figure shows the growth of strain R25 in a preferred embodiment of the present invention in a liquid culture medium containing elemental selenium.

[0032] Figure 8 The volatilization efficiency of selenite and nano-selenium by strain R25 in the preferred embodiment of the present invention is shown.

[0033] Fig. 9 This is the identification of the type of volatile selenium synthesized by strain R25 in the preferred embodiment of the present invention.

[0034] Fig.10 This is the time kinetics of the synthesis of volatile selenium by strain R25 in a preferred embodiment of the present invention.

[0035] Fig.11 The figure shows the efficiency of removing selenite and nano-selenium by strain R25 in a simulated bioreactor in a preferred embodiment of the present invention.

[0036] Fig.12 It is the volatile selenium product produced by strain R25 in a simulated bioreactor in a preferred embodiment of the present invention. DETAILED DESCRIPTION

[0037] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art, and the raw materials used are all commercially available products.

[0038] Example 1 Isolation and Identification of Delftia acidovorans R25 Strain

[0039] 5 g of soil was collected from the selenium fish pond in Enshi, Hubei Province, and 95 mL of sterile saline was added. The soil was placed in a shaker at 25 °C and 150 rpm for 30 min. The soil suspension was diluted 10 times by the 10-fold gradient dilution method. 1 -10 6Times. Take 100 μL of each dilution multiple bacterial solution, spread evenly on NA plate with 100 mM selenite added (NA medium: peptone 10.0 g / L, beef extract 3.0 g / L, sodium chloride 5.0 g / L, agar 15.0 g / L, pH 7.3 ± 0.1, sterilized at 121 ° C for 15 min), and culture in a 28 ° C constant temperature incubator for 2 days to obtain selenite tolerant bacteria. Among them, in the process of culturing in liquid containing selenite, the bacterial solution of strain R25 first turns red and then fades, the generated elemental selenium decreases and produces a garlic-like smell, indicating that strain R25 can produce volatile selenium.

[0040] The genomic DNA of strain R25 was extracted using a bacterial genome extraction kit (TIANGEN), and its 16S rRNA gene was amplified using universal primers 27F (5'-AGAGTTTGATCCTGGCTCAG-3') and 1492R (5'-GGTTACCTTGTTACGACTT-3'). The PCR product was purified and sequenced, and the sequencing results were spliced ​​using DNAMAN software to obtain the 16S rRNA sequence of strain R25 (SEQID NO: 1). The BLAST program of the NCBI database (https: / / blast.ncbi.nlm.nih.gov / Blast.cgi) was used for nucleotide similarity comparison, and the results showed that strain R25 had a similarity to Delftia acidovorans IAM 12409. T The consistency of the strains reached 100%. The 16S rRNA gene phylogenetic tree constructed using the neighbor-joining method (Kimura 2-parameter model) using MEGA software is shown in Figure 1 As shown, strain R25 was classified into the same clade as Delftia.

[0041] The whole genome draft of R25 strain was sequenced using Illumina HiSeq PE150 platform, and the genome size of R25 strain was 7.09Mb, the GC content was 66.54mol%, and there were 6401 coding genes. The genome of R25 was identified by TYGS analysis, and the whole genome phylogenetic tree of strain R25 was obtained ( Figure 2 ), strain R25 and Delftiaacidovorans NBRC 14950 T The strains were classified into a monophyletic group, further confirming that R25 belonged to Delftia acidovorans.

[0042] The average nucleotide similarity (ANI) between strain R25 and other Delftia species was calculated using JSpeciesWS (https: / / jspecies.ribohost.com / jspeciesws / #analyse) (Table 1), and the digital DNA-DNA hybridization (Dddh) between strain R25 and other Delftia species was calculated using GGDC Calculator 3.0 (http: / / ggdc.dsmz.de) (Table 1). The results showed that strain R25 had a similar genome to Delftia acidovorans IAM 12409. T The ANI and dDDH values ​​were 96.9% and 79.6%, respectively, both higher than the species threshold (ANI>95%, dDDH>70%), indicating that the R25 strain belonged to Delftia acidovorans.

[0043] Table 1 ANI and dDDH values ​​of R25 and other Delftia species

[0044] Reference model strains ANIb(%) <![CDATA[dDDH(%),d4]]> <![CDATA[Delftia acidovorans IAM 12409 T ]]> 96.9 79.6 <![CDATA[Delftia lacustris LMG 24775 T ]]> 93.8 58.5 <![CDATA[Delftia tsuruhatensis NBRC 16741 T ]]> 93.7 58.9 <![CDATA[Comamonas phosphati CGMCC 1.12294 T ]]> 79.5 25.4 <![CDATA[Comamonas terrae NBRC 106524 T ]]> 79.3 25.2 <![CDATA[Comamonas granuli NBRC 101663 T ]]> 76.3 22.5

[0045] According to the conventional microbial identification methods in the Manual of Common Bacteria Identification (Dong Xiuzhu and Cai Miaoying, 2001), strain R25 was physiologically and biochemically identified. Strain R25 is a Gram-negative bacterium, with rod-shaped cells, a diameter of 0.6-0.9 μm, and a length of 1.5-2.6 μm. In LB medium (LB medium: yeast extract 5g / L, tryptone 10g / L, sodium chloride 10g / L, pH7.0-7.2, sterilized at 121℃ for 20min) cultured at 28℃ for 2d, R25 colonies were 1-2mm in diameter, beige, translucent, moist on the surface, and rounded on the edges; it grows facultatively anaerobic, can reduce nitrate, can produce oxidase, catalase, and urease, but cannot produce H2S, DNA enzymes or arginine dihydrolase, and cannot hydrolyze starch. The growth stress resistance characteristics of strain R25 are shown in Table 2. R25 was cultured on LB solid medium in incubators at 4°C, 15°C, 20°C, 28°C, 32°C, 37°C, 40°C and 42°C for 7 days, indicating that it can grow at 4°C-37°C. TSB liquid medium with a NaCl concentration of 0-7% (w / v) was prepared, and fresh bacterial solution was inoculated with an inoculum of 1%, and placed in a shaker at 28°C and 150 rpm for 7 days. The absorbance value (OD 600) to determine that R25 can tolerate 2%-5% salt; prepare TSB liquid culture medium with different pH (3-11) values ​​using buffer, inoculate fresh bacterial liquid at 1% inoculation volume, place in a shaker at 28°C and 150rpm, shake and culture for 7 days, take the bacterial liquid to measure the absorbance value (OD 600 ), indicating that strain R25 can grow in the pH range of 5-11.

[0046] Table 2 Growth and stress resistance characteristics of strain R25 (Li, Yan et al. 2015)

[0047] index R25 <![CDATA[Delftia acidovorans NBRC 14950 T ]]> Growth temperature, °C 4-37 10-45 Salt tolerance (NaCl) 2％-5％ 0-3％ pH tolerance 5-11 6-10

[0048] Based on phylogenetic analysis, genome ANI and dDDH calculation results, and physiological and biochemical characteristics analysis, strain R25 can be identified as Delftia acidovorans, classified and named Delftia sp. R25, and the preservation number is CGMCC No. 25434.

[0049] Example 2 Tolerance and reducing ability of strain R25 to selenite

[0050] 1. Tolerance and reduction ability of strain R25 to selenite on solid medium

[0051] Pick a single colony of R25 and inoculate it into an LB tube. Cultivate it at 28°C and 150 rpm for 12 h to activate it. Adjust the OD 600 The concentration of LB was 0.8, which was used as the seed solution. The sterilized LB solid medium was cooled to about 60°C, and the filtered and sterilized sodium selenite solution was added. The mixture was gently shaken to prepare a plate containing 1-300 mM sodium selenite (in terms of selenium). The R25 seed solution was diluted step by step with sterile physiological saline in a 10-fold gradient, and 2.5 μL was added to the selenium-containing plate, dried with sterile air, and then cultured at 28°C for 48 hours.

[0052] The growth of strain R25 on solid medium containing sodium selenite is shown in Figure 3 The results showed that R25 could tolerate up to 100 mM (in terms of selenium) selenite. At a concentration of 1-100 mM, the R25 colony appeared bright red, indicating that selenite was reduced to synthesize elemental selenium.

[0053] 2. Tolerance and reduction ability of strain R25 to selenite in liquid culture medium

[0054] The R25 seed solution was inoculated at 1% inoculum into LB liquid medium to which sterile sodium selenite solution had been added to a final concentration of 1-300 mM (in terms of selenium), and cultured at 28° C. and 150 rpm for 48 hours, and then the bacterial growth was observed.

[0055] Results Figure 4 , strain R25 can grow in liquid medium containing 1-100mM sodium selenite (in terms of selenium) and can tolerate up to 100mM sodium selenite (in terms of selenium). The bacterial liquid appears bright red, and in the TEM photo ( Figure 5 ) observed a large number of elemental selenium particles, and the Na2S spectrophotometric method measured that the maximum elemental selenium yield was 2.05mM under the condition of 40mM sodium selenite (in terms of selenium). Figure 5 It can be seen that in the medium containing 10mM sodium selenite, the diameter and length of R25 bacteria were reduced, and single selenium particles were produced. The single selenium particles had a diameter between 200-400nm and were spherical.

[0056] Example 3 Tolerance of strain R25 to elemental selenium

[0057] 1. Tolerance of strain R25 to elemental selenium on solid culture medium

[0058] After the LB solid medium is completely melted, it is cooled to about 60°C, and the filtered and sterilized elemental selenium solution is added. The mixture is gently shaken to prepare a plate containing 1-300mM elemental selenium (in terms of selenium). The R25 seed solution is diluted step by step with sterile physiological saline in a 10-fold gradient, and 2.5μL is added to the selenium-containing plate. After being dried with sterile air, it is placed at 28°C for 48h. The growth of strain R25 on the solid medium containing elemental selenium is shown in Figure 6 The results showed that R25 could tolerate more than 300 mM (in terms of selenium) of elemental selenium.

[0059] 2. Tolerance of strain R25 to elemental selenium in liquid culture medium

[0060] The R25 seed solution was inoculated into LB liquid medium supplemented with sterile elemental selenium at a 1% inoculation rate (final concentration was 1-300 mM, in terms of selenium), and the culture was shaken in an incubator at 28°C and 150 rpm for 48 hours, and the growth of the bacteria was observed. Figure 7 , strain R25 can grow in liquid culture medium containing 1-300 mM elemental selenium (calculated as selenium) and can tolerate elemental selenium above 300 mM (calculated as selenium).

[0061] Example 4 Volatilization efficiency of strain R25 for selenite and elemental selenium

[0062] In a 150mL shake flask filled with 50mL LB medium, 0.5mL R25 seed solution and the corresponding volume of sodium selenite or elemental selenium mother solution (filter sterilized) were inoculated to make the initial selenium concentrations 0.0625mM, 0.125mM, 0.25mM, 0.5mM, 1.0mM, 1.5mM, 2.0mM (in terms of selenium), respectively, and cultured at 28°C and 150rpm for 96h. The volatilization rate of selenium was calculated by the reduction of total selenium in the bacterial solution after 96h of culture:

[0063] Selenium volatilization rate (%) = (total selenium in bacterial solution 0h -Total selenium in bacterial solution 96h ) / Total selenium in bacterial solution 0h ×100%

[0064] The total selenium in the bacterial solution was determined by atomic fluorescence spectrometry (HG-AFS) and the selenium volatilization rate was calculated. The results showed that after 96 hours of inoculation with R25, the selenium in the culture medium decreased significantly, and a strong garlic smell was smelled during the culture process, indicating that volatile selenium compounds were produced. At the addition amount of 0.0625-0.25mM, the volatilization rate of R25 for both inorganic selenium, selenite and elemental selenium, reached more than 85%. Among them, the volatilization rate reached the highest at the addition amount of 0.25mM, with the volatilization rates of selenite and elemental selenium being 90.7% and 90.3% respectively; at the addition amount of 0.5mM, the volatilization rates of selenite and elemental selenium were 49.6% and 43.1% ( Figure 8 ).

[0065] Example 5 Identification of volatile selenium synthesis by strain R25

[0066] The R25 seed solution was inoculated into a 3 mL LB headspace bottle containing 0.5 mM selenite or 0.5 mM elemental selenium (in terms of selenium) at a 1% inoculation rate and cultured at 28°C and 150 rpm. After 96 h of culture, the headspace gas was identified for selenium speciation using gas chromatography-mass spectrometry (GC-MS). The results showed that ( Fig. 9 ), R25 converted Se(IV) into DMSeO2 (dimethyl selenone), DMDSe (dimethyl diselenide), DMDSSe (dimethylselenodisulfide) and DMDSeS (dimethyl diselenylsulfide), and 1,2,4-triselenocyclopentane was also observed at 7.9-8.0min, without the production of DMSe (dimethylselenide).

[0067] Example 6 Time dynamics of volatile selenium synthesis by strain R25

[0068] The R25 seed solution was inoculated into LB medium supplemented with sterile selenium at an inoculum volume of 1% (the final concentration was 0.5 mM sodium selenite or 0.5 mM elemental selenium (in terms of selenium)), and cultured at 28° C. and 150 rpm with shaking for 96 h.

[0069] Samples were taken at regular intervals and the total selenium content in the culture medium was determined using an atomic fluorescence spectrometer (HG-AFS). The volatilization rate was calculated based on the reduction in the total selenium in the bacterial culture. The results showed that ( Fig.10 ), 0-48h, the volatility of R25 to Se(IV) and SeNPs was less, with a volatility rate of less than 20%. At 48-96h, the volatility rates of R25 to Se(IV) and SeNPs doubled, reaching 40.6% and 41.58%, respectively.

[0070] Example 7 Removal of selenite and elemental selenium from water and recovery of methylselenoic acid using strain R25 in a simulated bioreactor

[0071] The R25 seed solution was inoculated into the simulated bioreactor device at an inoculum of 1%. The culture medium in the reactor was 6 L of sterile LB medium containing 0.25 mM Se (IV) and 0.25 mM Se NPs (in terms of selenium). The ventilation volume of the experimental device was 0.18 m 3 / h, tank pressure 0.05-0.06MPa, stirring 150rpm, fermentation at 28±0.5℃ for 48h. The tail gas enters the concentrated nitric acid (pure nitric acid) absorption liquid through the pipeline for absorption: tail gas→buffer bottle (1L)→absorption bottle ① (1L with 850mL concentrated nitric acid)→absorption bottle ② (1L with 500mL concentrated nitric acid)→water (1L with 500mL)→exhaust. The total selenium content in the culture medium is determined regularly using an atomic fluorescence spectrometer (HG-AFS), and the selenium volatilization rate is calculated using the reduction in total selenium in the bacterial solution. The results show that ( Fig.11 ), strain R25 was able to remove Se(IV) and SeNPs simultaneously, and the selenium volatilization rate reached 91% at 48 h.

[0072] The results of the identification of selenium forms in the 48h absorption bottle ① using high performance liquid chromatography-hydride generation-atomic fluorescence spectrometry (HPLC-HG-AFS) showed that ( Fig.12 ), the selenium compound in the capture solution is methylseleninic acid (MSA), which is the oxidation product of DMDSe (dimethyl diselenide) (Winkel, Feldmann et al. 2010), which can be further processed to produce anti-tumor drugs after collection. R25 does not generate DMSe (dimethyl selenium), and there is little interference from other selenium compounds such as dimethyl oxide (DMSeO, the oxidation product of DMSe) in the capture solution, making it a potential strain for producing anti-tumor drugs.

[0073] Although the present invention has been described in detail above with general descriptions and specific embodiments, it is obvious to those skilled in the art that some modifications or improvements can be made based on the present invention. Therefore, these modifications or improvements made without departing from the spirit of the present invention all fall within the scope of protection claimed by the present invention.

[0074] References:

[0075] [1] Burra, R., GAPradenas, RAMontes, CCVasquez and TGChasteen (2010). "Production of dimethyl triselenide and dimethyl diselenenyl sulfidein the headspace of metalloid-resistantBacilusspecies grown in the presence of selenium oxyanions." Analytical Biochemistry 396(2):217-222.

[0076] [2] Harris, JD, CMCoon, MEDoherty, EAMcHugh, MCWarner, CLWalters, OMOrahood, AELoesch, DCHatfield, JCSitko, EAAlmand andJ.J.Steel (2022). "Engineering and characterization of dehalogenase enzymes from Delftia acidovoransin bioremediation of perfluorinated compounds." Synthetic and Systems Biotechnology 7(2):671-676.

[0077] [3]Kagami,T.,T.Narita,M.Kuroda,E.Notaguchi,M.Yamashita,K.Sei,S.Sodaand M.Ike(2013)."Effective selenium volatilization under aerobic conditionsand recovery from the aqueous phase byPseudomonas stutzeriNT-I."WaterResearch 47(3):1361-1368.

[0078] [4]Liu,J.,G.Xu,W.Dong,N.Xu,F.Xin,J.Ma,Y.Fang,J.Zhou and M.Jiang(2018)."Biodegradation of diethyl terephthalate and polyethyleneterephthalate by a novel identified degraderDelftiasp WL-3and its proposedmetabolic pathway."Letters in Applied Microbiology67(3):254-261.

[0079] [5]Moreno-Martin,G.,J.Sanz-Landaluze,M.E.Leon-Gonzalez and Y.Madrid(2021)."In vivo quantification of volatile organoselenium compounds releasedby bacteria exposed to selenium with HS-SPME-GC-MS.Effect of selenite andselenium nanoparticles."Talanta 224.

[0080] [6]Varlamova,E.G.and E.A.Turovsky(2021)."The Main Cytotoxic Effectsof Methylseleninic Acid on Various Cancer Cells."International Journal ofMolecular Sciences 22(12).

[0081] [7]Winkel, L., J. Feldmann and AAMeharg (2010). "Quantitative and Qualitative Trapping of Volatile Methylated Selenium Species Entrainedthrough Nitric Acid." Environmental Science & Technology 44(1):382-387.

[0082] [8] Yilmaz, F. and B. Icgen (2014). "Characterization of SDS-degradingDelftia acidovorans and in situ monitoring of its temporal succession in SDS-contaminated surface waters." Environmental Science and Pollution Research 21(12):7413-7424.

[0083] [9] Dong Xiuzhu and Cai Miaoying (2001). Manual of systematic identification of common bacteria, Manual of systematic identification of common bacteria.

[0084]

[10] Zhou Jie, Yan Hai, He Hongsheng, Zhong Guangrong, Zhang Chao and Lin Hai (2006). "Study on the biodegradation activity of microcystin by Delftia acidovorans USTB04." Science Technology and Engineering 6(2):5.

[0085]

[11] Li, C.-T., Z.-F. Yan, X. Chu, F. Hussain, W.-D. 107(6):1445-1450.

Claims

1. Delftia sp. R25, deposited with CGMCC No. 25434.

2. A bacterial agent containing the Delftia bacteria according to claim 1.

3. A method for biosynthesizing volatile selenium, characterized in that: The method comprises: inoculating the Delftia bacteria described in claim 1 into a selenium-containing culture medium for culturing; Wherein, the selenium in the culture medium is inorganic selenium and / or elemental selenium.

4. The method according to claim 3, characterized in that The inorganic selenium is selenite.

5. The method according to claim 3, characterized in that: When the selenium in the culture medium is sodium selenite, The concentration of sodium selenite in the liquid culture medium is 1-100 mM, calculated as selenium; The concentration of sodium selenite in the solid medium is 1-100 mM, calculated as selenium; When the selenium in the culture medium is elemental selenium, The concentration of elemental selenium in the liquid culture medium is 1-300 mM, calculated as selenium; The concentration of elemental selenium in the solid culture medium is 1-300 mM, calculated as selenium.

6. The method according to any one of claims 3 to 5, characterized in that: The volatile selenium is selected from DMSeO2, DMDSe, DMDSSe and DMDSeS.

7. Use of the Delftia bacteria described in claim 1 in the bioremediation of selenium-contaminated water and soil.

8. The use according to claim 7, characterized in that: The Delftia bacteria or its bacterial agent is brought into contact with selenium-contaminated water and soil to convert inorganic selenium and / or elemental selenium therein into volatile selenium, thereby achieving the purpose of biological removal of selenium.

9. The use according to claim 7 or 8, characterized in that: The application further comprises a step of recovering the volatile selenium.

10. A method for preparing methylselenoic acid using Delftia acidovorans R25, characterized in that: The volatile selenium obtained by the method according to any one of claims 3 to 6 is contacted with concentrated nitric acid to cause an oxidation reaction, and methylselenic acid is collected from the reaction product.

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