Application Progress of Deinococcus radiodurans in Biological Treatment of Radioactive Uranium-Containing Wastewater

Abstract

Radioactive uranium wastewater contains a large amount of radionuclide uranium and other heavy metal ions. The radioactive uranium wastewater discharged into the environment will not only pollute the natural environment, but also threat human health. Therefore, the treatment of radioactive uranium wastewater is a current research focus for many researchers. The treatment in radioactive uranium wastewater mainly includes physical, chemical and biological methods. At present, the using of biological treatment to treat uranium in radioactive uranium wastewater has been gradually shown its superiority and advantages. Deinococcus radiodurans is a famous microorganism with the most radiation resistant to ionizing radiation in the world, and can also resist various other extreme pressures. D. radiodurans can be directly used for the adsorption of uranium in radioactive waste water, and it can also transform other functional genes into D. radiodurans to construct genetically engineered bacteria, and then applied to the treatment of radioactive uranium containing wastewater. Radionuclides uranium in radioactive uranium-containing wastewater treated by D. radiodurans involves a lot of mechanisms. This article reviews currently the application of D. radiodurans that directly or construct genetically engineered bacteria in the treatment of radioactive uranium wastewater and discusses the mechanism of D. radiodurans in bioremediation of uranium. The application of constructing an engineered bacteria of D. radiodurans with powerful functions in uranium-containing wastewater is prospected.

Introduction

There is a constantly growing demand for nuclear fuel globally. Nuclear energy is an important energy material, and it has been more and more widely used in the world as a kind of clean energy. However, a great number of radioactive wastes are often produced in the product process of nuclear industrialization. Radioactive uranium-containing wastewater is the most common and harmful one. The rapid development of the uranium mining industry has caused radioactive uranium wastewater in large numbers to be discharged into natural environment. Radioactive uranium wastewater mainly comes from the mining, smelting and processing of uranium mines, as well as the emission of uranium-containing wastewater from nuclear power plants, laboratories and factories [1]. The direct discharge of uranium-containing wastewater into the natural environment will not only cause serious damage to human health, but also cause serious pollution to the natural environment. Other heavy metal ions in the wastewater will affect the survival and reproduction of organisms in the water environment. These will eventually endanger the survival of mankind [2]. Therefore, the restoration of radioactive uranium wastewater is the most important aspect of uranium mining and metallurgical waste treatment.

Uranium is a naturally occurring radionuclide, and one of the most common radioactive pollutants in soil, sediment and groundwater [3, 4]. There are three main natural isotopes of uranium in nature: ²³⁸U, ²³⁵U and ²³⁴U, all of which are radioactive [5]. Uranium has a long half-life and will cause serious pollution to the environment after entering the environment. Food and water are the primary ways for humans to intake uranium. Uranium that enters the human body can cause serious damage to the liver and kidney, nervous system, immune system and so on, it can also cause cancer and even death, thereby endangering human health [6, 7]. The toxicity of uranium is usually determined by its chemical properties and radioactivity. The uranium in radioactive wastewater mainly exists in two valence forms: tetravalent uranium U(IV) and hexavalent uranium U(VI). The poorly soluble U(IV) sedimentary compounds usually exist as minerals and have low toxicity, while soluble U(VI) mainly exists in wastewater in the form of uranyl ionsU₂²⁺, and has high toxicity [8, 9]. Generally speaking, the bioremediation of radioactivity uranium wastewater is mainly to remove hexavalent uranium in wastewater [10].

Long-term exposure to uranium will lead to potential risks to human health. People are mostly exposed to uranium due to ingestion of drinking water, so it is crucial to monitoring and understanding the uranium concentration in local groundwater [11]. Radioactive uranium wastewater discharged from human activities enters groundwater through water and soil, causing groundwater pollution. Radioactive elements in geology can enter the groundwater environment through interaction with groundwater [12]. The form, migration and distribution of uranium in groundwater mainly depend on the pH, redox conditions, chemical composition and so on [13]. Uranium exists in groundwater in a variety of chemical forms, the oxidation state of groundwater is dominant. Uranium (VI) mainly exists in the form of free uranyl ions under acidic conditions (pH < 5), and it can combine with phosphate to form uranyl phosphate at pH < 6.5, but combines with hydroxyl ions and Carbonate at higher pH to form various complexes, and the uranium in groundwater can also combine with other substances, such as humus, colloids, etc. [14]. Uranium combines with inorganic or organic ligands to form complexes, which are not easily be adsorbed and decomposed, and further improves the fluidity of uranium [15]. The remediation of uranium in groundwater is mainly in-situ remediation. The adsorption of uranium by soil, sediments, minerals and microorganisms can reduce the migration of uranium. In addition, when repairing groundwater uranium pollution, it is an effective method to reduce soluble U(VI) to insoluble U(IV) by reducing microorganisms (iron-reducing bacteria and sulfate-reducing bacteria) under strong reducing conditions, it can also effectively reduce the migration of uranium [16]. Through a series of single-well and push–pull tests, Istok et al. [17] investigated the in-situ biological reduction of U(VI) and Tc(VII) in the presence of NO₃-co-contamination in a shallow unconfined aquifer in the United States, which NO₃- (up to 170 mM), U(VI) (up to 20 uM) and Tc(VII) (up to 30,000 pM). Through adding electron donors such as ethanol, acetate, or glucose, it was found that the growth and activity of local denitrification and metal reduction organisms could be stimulated, thus proving that biological immobilization is an feasible repair strategy in sites contaminated by mixture of U(VI) and Tc(VII). However, once some conditions of the groundwater change, the deposited mineral uranium is likely to enter the water again and cause pollution again. For example, oxygen and nitrate can reduce the insoluble U(IV) to U(VI) again [18]. The composition of groundwater environment is complex, and it was disturbed by many factors in the process of repairing uranium pollution. In general, uranium pollution in groundwater is a serious environmental pollution problem.

With the frequent occurrence of environmental pollution by radioactive uranium wastewater, how to treat these uranium-containing wastewater has attracted an increasing number of attention from the government and researchers. Radioactive uranium wastewater must be treated before it can be discharged into the natural environment. The treatment of uranium wastewater includes physical, chemical and biological methods [19]. Remediation utilizing microorganism is an emerging environmental pollutant purification technology. Physical and chemical treatment method of uranium has the advantages of rapid, can continuous operation, can automatic control and can also simultaneous treatment of multiple pollutants. However, their disadvantages are high economic cost, many influencing factors, complicated operations, and some processes are prone to produce secondary pollutants [20]. Although the service life of the microbial treatment method is limited, it offers advantages in some terms, such as high-efficiency, low cost, low energy consumption, simple operation, no secondary pollutants and a wide range of source of raw materials. In addition, many current studies have immobilized microorganisms and then used them to treat uranium wastewater, which can increase the repeatability of microorganisms [6, 21]. Therefore, the microbial treatment method has potential advantages in the treatment field of uranium wastewater. Microbial repair of uranium has attracted a lot of attention and studies of many scholars and researchers in the field of radioactive uranium containing wastewater treatment and achieved many gratifying research results [22, 23]. Wang et al.[24] have studied that the adsorption of uranium by Saccharomyces cerevisiae under different cell activity states, and the study has found that the heat-killing process destroys the cell membrane and enhances its permeability, resulting in the free diffusion of small molecules and decomposing substances from the inside of the cell. Since the released phosphate can combine with uranium, the rough surface and nanopores of the bacteria caused by autoclaving also provide more adsorption surface for the bacteria, which enhances the adsorption capacity of heat-killed cells to uranium. Compared with living cells that reduce uranium toxicity via metabolism-dependent, the heat-killed cells have better adsorption capacity for uranium, so the heat-killed yeast may be a suitable biological adsorbent for treating uranium wastewater.

Bioremediation has important research significance in the field of treatment of radioactive uranium wastewater. Many microorganisms are used for the treatment of uranium wastewater, but common microorganisms are sensitive to radiation. The radiotoxicity of the radionuclide uranium will affect the survival, and certain specific functions of ordinary microorganisms, as well as high radiation levels and its chemical hazards are extremely destructive to organisms and often leading to cell death, thereby limiting their ability to repair uranium, they may only be suitable for lower radiation levels environment [25–28]. Deepti Appukuttan et al. constructed recombinant strains E. coli-phoN and recombinant strains Deino-phoN, it was found that after 6 kGy irradiation, the recombinant strains Deino-phoN still maintained their uranium precipitation ability. However, E. coli-phoN clones could not perform optimally founction even at low doses (1 kGy), and exhibited severe inhibition of PhoN activity at high doses (3–6 kGy) [29]. The development of microbial methods to treat relatively high-level radioactive uranium wastewater depends to a large extent on the ability of microorganisms to perform the required functions to survive and work under radiation stress. Therefore, it is necessary to find a microorganism with anti-radiation ability and can repair toxic metals [30]. D. radiodurans is the most radiation resistant microorganism on earth. It is a gram-positive bacterium, non-pathogenic, and has significant resistance to ionizing radiation, ultraviolet rays (UV), desiccation, mitomycin C and oxidative stress, and has a strong ability to make reparation of DNA damage [31]. It is a microorganism that suitable for DNA repair and antioxidant research. D. radiodurans is a kind of microorganism originally isolated in 1956 by Anderson team from a canned of meat that was still spoiled after being sterilized by γ-ray, which shows a strong resistance to radiation [32]. The cell wall of D. radiodurans basically has two layered structures, one is a peptidoglycan layer, and the other has an unknown layered structure, which is still under study. Observing the cell through electron microscope, its cell wall can be divided into six layers, and its ultra-thick cell wall characteristics also enhance its ability to resist extreme environments [33, 34]. After being exposed to 6000 Gy of ionizing radiation, genome of the D. radiodurans was broken down into hundreds of gene fragments. The radiation activated the PprI protein (pprI is a general switch gene for DNA damage repair), and then the DdrO protein (the total regulator of the RDR regulon in D. radiodurans) was cleaved of DdrO by activated PprI protein, so that RDR regulon genes (e.g., recA, pprA, gyrB and ddrO) were expressed. These proteins started to take radiation response after expression, and the DNA damage was repaired. after 6–9 h, the broken genome fragments can be recombined and restored, so that the D. radiodurans could return to normal without being killed [35, 36]. D. radiodurans has a strong radiation resistance, no pathogenicity, and high tolerance to oxidative stress, making it an ideal microorganism in the application of treatment of uranium-containing wastewater.

D. radiodurans, which strong radiation resistance, have great significance for the study of water environmental restoration and treatment in low-concentration uranium mining and metallurgical areas. This article focuses on reviewing the application of D. radiodurans in radioactive uranium wastewater and the mechanism of its enrichment uranium, and prospects the application of D. radiodurans to radioactive uranium-containing waste water.

Application of D. radiodurans in Uranium Wastewater

D. radiodurans Directly Applied to the Treatment of Uranium Containing Wastewater

D. radiodurans is used in the treatment of radioactive uranium containing wastewater due to its extreme radiation resistance. D. radiodurans not only has a much thicker cell wall than that of ordinary bacteria, but also has many radiation resistant genes, such as pprI, pprM, ddrA and RecA genes, etc. [37, 38].These properties endow D. radiodurans with strong radiation resistance. The RecA repair enzyme encoded by the recA gene of D. radiodurans is easily combined with DSBs (double-stranded DNA fragments), so it can repair broken DNA quickly and efficiently [39]. Therefore, many researchers use the extremely strong radiation resistance of D. radiodurans to directly apply it to the treatment of uranium-containing wastewater, and have achieved many good results. The inherent radiation resistance of D. radiodurans enables it to absorb uranyl ions in uranium-containing wastewater under uranium stress and form flake-like sediment on the surface of the bacteria [40]. Yang et al. [41] have used D. radiodurans to perform uranium adsorption experiments targeted to remove uranyl ions in uranium-containing wastewater and researched its effect factors on adsorption of uranium. Study showed that D. radiodurans adsorbed uranium mainly by ion exchange or surface complexation mechanism.

Application of D. radiodurans Engineering Bacteria in the Treatment of Uranium Wastewater

With the increasing development of science and technology and the broading demand for practical applications, the directly application of D. radiodurans to wastewater treatment has gradually shown its limitations (Fig. 1). D. radiodurans is relatively easy to carry out genetically transformation and genetically manipulation, and is considered to be a good model organism that can be used to construct genetically engineered bacteria in radioactive environments [42, 43]. Brim et al. [44] have studied that the tod and xyl genes from Pseudomonas putida were cloned to D. radiodurans to make the recombinant Tod/Xyl strain, which could naturally reduce Cr(VI) to Cr(III)(less mobile and less toxic), was engineered for complete toluene degradation. This engineered Tod/Xyl strain can mineralize toluene and can utilize the energy of toluene catabolism combined with its natural ability to reduces Cr(VI) to repair heavy metals in radioactive waste. Since radioactive uranium-containing wastewater discharged from uranium mining not only contains radioactive uranium, but also has multiple stresses such as fluoride ion stress and other heavy metal interference, D. radiodurans will also be interfered by many factors in the process of treating uranium-containing wastewater [45]. Therefore, researchers began to try to transform other functional genes into D. radiodurans for conferring the powerful functions on the D. radiodurans and expanding its application range in the treatment of uranium-containing wastewater (Fig. 1). After transferring other functional genes into D. radiodurans, the bacteria can have some functions that themselves don't possess, and make it more effective to adsorb uranium in wastewater. Misra et al. [46] have transformed the plasmid pSN4 or pPN1 carrying Pssb-phoN or PgroESL-phoN into D. radiodurans, and used radiation-induced promoter Pssb to enhance the bioprecipitation of uranium by recombinant bacteria (pSN4). Deinococcus (pSN4) cells could precipitated 1.8 g uranium/g dry biomass in 24 h and 3.6 g uranium/g dry biomass in 48 h from 5 and 10 mM uranyl nitrate. Deinococcus (pSN4) cells could precipitate 70% of uranium in 4 days when the uranium concentration is 20 mM, while Deinococcus (pPN1) cells could only precipitate 20% of uranium. The inherent radiation tolerance and high level of PhoN activity of the recombinant bacteria (pSN4), which make it able to withstand long-term radiation and rapidly precipitate uranium.

Fig. 1.

Uranium treatment model of wild-type D. radiodurans and genetically engineered bacteria D. radiodurans

Mechanism of D. radiodurans in the Bioremediation of Radioactivity Uranium Containing Wastewater

Microbes can remove the radionuclide uranium in uranium wastewater through their own inherent adsorption or precipitation capacity (Fig. 2). The mechanisms of remediation of uranium wastewater by methods of microorganism mainly include biological reduction, biological adsorption, and biological mineralization, they have their own advantages and disadvantages (Table 1) [47].

Fig. 2.

The mechanism of D. radiodurans repairing U(VI); (1) Biosorption; (2) Bioreduction; (3) Biomineralization

Table 1.

Mechanism, advantages and disadvantages of bioremediation of uranium in wastewater containing uranium

Mechanism	Advantage	Disadvantage
Biosorption	There are many kinds of microorganisms available, fast adsorption rate and large capacity	The product is unstable and desorption occurs after adsorption
Bioreduction	Using the principle of oxidation and reduction, a relatively stable circulation system is formed and contributes to uranium enrichment, and is environmentally frienhment	The product is not stable and is prone to reoxidation under aerobic conditions
Biomineralization	Its product uranyl phosphate is extremely stable and can remove uranium more effectively	Requires the presence of organophosphate to react under the mediation of phosphatase

Biosorption

Microbial adsorption method is a novel treatment technology for radioactive uranium wastewater, it makes use of the chemical structure and composition characteristics of microorganism itself to adsorb uranium in wastewater, thereby removing uranium from the uranium wastewater [48]. Biosorption is considered to be a kind of high-efficiency, economic and easy-to-operate and environmental friendly biological purification technology with wide applicability, it has a good application prospect in the research of repairing radioactive uranium wastewater [49]. The biological adsorption process includes adsorption kinetics and adsorption thermodynamics, and also has different adsorption models, which mainly include the Freundlich adsorption isotherm model, the Langmuir adsorption isotherm model and the BET adsorption isotherm model [50, 51]. The surface of bacteria contains many adsorption sites (surface active groups), such as NH, CN, hydroxyl, carboxyl, carbonyl, etc., these are the main adsorption functional groups,which can combine with heavy metal ions in wastewater to achieve the purpose of removing heavy metal ions from wastewater [52, 53]. Thus, some researchers used wild type D. radiodurans as an adsorbent to adsorb uranium in uranium-containing wastewater. Liu et al. [54] have studied the biosorption of uranium by D. radiodurans under culture conditions. The biosorption results have shown that the D. radiodurans cells have higher adsorption capability for uranium under culture conditions, its maximum biosorption efficiency is 86% and the maximum biosorption quantity is 230 mg/g.

Immobilizing bacteria and then applying them to the treatment of uranium-containing wastewater is a popular research method that has emerged during recent years. Immobilization is a method that can improve the stability of bacteria so that they can be reused, but doesn’t reduce the biological activity of bacteria [55]. Use physical or chemical methods to immobilize the bacteria on the immobilized carrier, and then apply the immobilized carrier containing the bacteria to the treatment of uranium-containing wastewater, this immobilization makes the bacteria have better mechanical strength and stability, and the carrier can also be reused, which improves the practicability of bacteria [56]. According to the saturated boric acid-alginate calcium cross-linking method, Can Chen et al. [57] by embedding sodium sulfate into living yeast cells of Saccharomyces cerevisiae, have prepared a novel of biosorbent:sulfate-strengthened immobilized gel beads containing living yeast cells. The characteristics of the immobilized microorganisms were investigated and the factors affecting the adsorption of uranium by the new biosorbent were explored. Qin et al. [58] have immobilized live D.radiodurans with sodium alginate (SA) to studied the adsorption and elution of uranium by immobilized D.radiodurans, and the ashing volume reduction and the chemical form of uranium in ashing. The adsorption efficiency of immobilized D.radiodurans on uranium could reach more than 95%. In addition, the immobilized D.radiodurans can be reused to remove uranium through an attachment-desorption cycle.

In addition to applying D. radiodurans live cells directly to uranium-containing wastewater for uranium adsorption, some researchers use it to form biofilms as an adsorbent to treat uranium-containing wastewater. Biofilm is a community of microorganisms that relies on extracellular products to attach to the surface of organisms, and bacteria are wrapped in their self-produced matrix [59].While enhanceing the metabolism of bacteria and their resistance to the environment and polymetallic, the biofilm can also reduce the damage of heavy metal ions to bacteria [60]. Manobala et al.[61] have studied the ability of biofilm formed by recombinant D. radiodurans (DR1-bf⁺) to remove uranium. The research results have shown that the DR1-bf⁺ biofilm removes uranium by adsorption. When Ca²⁺ presence in the solution, the amount of uranyl ions removed by DR1-bf⁺ biofilm was obviously higher than that of wild-type DR1 planktonic cells, DR1-bf⁺ planktonic cells and DR1-bf⁺control biofilm (grown without Ca²⁺). In a few minutes, the recombinant Deinococcus radiodurans biofilm (DR1-bf⁺) grown for four days can obviously remove about 75% UO₂²⁺. Obviously, D. radiodurans, as a microorganism with extreme radiation resistance, has significant research significance as a bioadsorbent for the treatment of radionuclides in a radioactive environment.

Bioreduction

Bioreduction of uranium is a reduction of soluble U(VI) in wastewater to insoluble U(IV) deposits by microorganisms through their own reduction functions (such as expression of reductase, etc.) or other means, thereby reducing the concentration and mobility of uranium in wastewater [62]. Bioreduction has the advantages of environmental friendliness, low cost, can operate in large quantity and green environmental protection, and has obvious advantages in the research of radioactive uranium-containing wastewater treatment [63]. There are many complicated mechanisms for the reduction of soluble U(VI) to sparingly U(IV). Some bacteria can reduce U(VI) with using various c-type cytochrome-mediated extracellular electron transfers, such as iron-reducing bacteria and sulfate-reducing bacteria [64]. Some enzymes of membrane-bound, periplasmic and intracellular can involve in the reduction of U(VI), some microbial pili and electron shuttle compounds also participate in the reduction of U(VI), and electron donors, such as acetic acid and lactic acid, etc., promote the reduction of U(VI) by providing electrons [60, 65]. Fredrickson et al. [66] have coupled the reduction of humic acid analog anthraquinone-2,6-disulfonate (AQDS) by D. radiodurans and the reduction of heavy metals and radionuclides to treat heavy metals and radionuclides in wastewater. Results have shown that D. radiodurans can reduce U(VI) to U(IV) in the presence of AQDS with lactic acid as the electron donor, D. radiodurans removed 95–100% of U(VI) with a concentration ranging from 5 to 100 mM within 21 days.

The process of bioreduction of uranium will also be affected by many other factors, such as electron donor concentration, (IV) forms, bioreduction rate, U(VI) bioavailability, pH value and Stability of U(IV) and so on [67]. After the soluble U(VI) in wastewater is reduced to insoluble U(IV) sediments, it is easily reoxidized to soluble U(VI) under aerobic conditions [68]. Therefore, the bioreduction of uranium has certain limitations, it is necessary to continue researches for expand the practical application of bioreduction of uranium.

Biomineralization

Since the uranyl ions in the solution are easily reoxidized under reduction conditions after bioreduction to insoluble U(IV), if the underground environmental conditions change to an oxidized state, the U(IV) sediments after bioreduction are likely to be re-oxidized to soluble uraniumU(VI), which will pollute the underground environment again. Studies have shown that it can promote the precipitation of poorly soluble uranium phosphates via the addition of an organophosphate to the uranium-containing waste liquid, making it more recalcitrant to reoxidize, this kind of mineral is more stable and harder to reoxidize than the product of microbial reduction of U(VI), so it is feasible to prevent U(IV) precipitation of microbial reduction from reoxidation by adding organic phosphate [69]. Therefore, in the presence of organic phosphate, the biomineralization of uranium may be a more attractive method for remediation of radioactive uranium-containing wastewater.

Biomineralization of uranium means that the combination of U(VI) and enzymes (such as phosphatase) produced by microorganisms forms a precipitate, thereby precipitating the uranium in the uranium wastewater [1, 70]. The phosphatase secreted by microorganisms can hydrolyze the organophosphorus compounds in the solution and then release inorganic phosphate, the latter reacts with uranium to form a stable uranyl phosphate precipitate, which is not easily oxidized under aerobic conditions [71]. Phosphatases can be divided into acidic and alkaline phosphatases, uranium can be treated by microorganisms with containing the corresponding phosphatase in acidic or alkaline environments [72]. Therefore, an increasing number of researchers have transformed acid or alkaline phosphatase genes into D. radiodurans for the treatment of uranium in radioactive uranium wastewater. Appukuttan et al. [29] have constructed a recombinant D. radiodurans strain containing the non-specific acid phosphatase gene (PhoN), and successfully expressed the active protein PhoN in D. radiodurans. After the recombinant strain Deino-PhoN was irradiated with 6 kGy ⁶⁰Co γ-rays, it still maintained PhoN activity and radiation resistance, and 90% uranium was precipitated from a 0.8 mM uranyl nitrate solution within 6 h. Kulkarni et al. [73] have introduced the alkaline phosphatase (PhoK) gene from Sphingomonas sp. into D. radiodurans, the recombinant strain with high phosphatase activity can effectively precipitate uranium in dilute alkaline solutions, so it may be applied to the bioremediation of radioactive uranium mine wastewater. This inherent radiation resistance ability of this microorganism can help researchers use engineered strains to perform in-situ bioremediation of radioactive waste, and can recover uranium from the low concentration and high radiation environment unique to dilute nuclear waste.

The composition of radioactive uranium-containing wastewater is complex, and there are many other heavy metal ions in the wastewater, such as chromium (VI), fluorine, copper and so on, which can interfere with the biological precipitation of uranium [30, 30]. In order to overcome the interference of other heavy ions in wastewater, researchers began to seek other methods to eliminate ion interference. Xu et al. [74] have transformed the phosphatase gene phoN and chromium reductase gene YieF into D. radiodurans to construct a recombinant D. radiodurans containing phoN and YieF, and studied the uranium precipitating ability of the recombinant Deino-phoN-YieF strain in the presence of chromium (VI) in wastewater. Studies have shown that Deino-phoN-YieF can reduce Cr(VI) to Cr(III) and effectively remove uranium from radioactive uranium-containing wastewater. In general, lyophilized microorganisms can reduce their volume and increase stability, but don’t reduce the activity and survival of bacteria. Appukuttana et al. [75] have lyophilized the cells of the recombinant D. radiodurans strain that containing non-specific acid phosphatase phoN gene. It has been found that lyophilized recombinant D. radiodurans cells can still maintain its viability and PhoN activity, and can also retain uranium precipitation ability after six months of storage at room temperature. The lyophilized of microorganisms and then treatment of radioactive uranium-containing wastewater may be a new research direction.

Summary and Outlook

When the concentration of radioactive uranium-containing wastewater is slightly higher, it is easy to pack tightly. Factors such as radiation and fluoride ion stress will cause the "active inactivation" of the bioconcentration agent and impair the uranium enrichment ability. In the complex components of uranium-containing wastewater, fluoride ions can inhibit the enzyme activity of indigenous bacteria, inhibit microbial metabolism, and have a strong inhibitory or killing effect on the growth of microorganisms [76].The indigenous reducing bacteria SRB to treat groundwater from in-situ leaching of uranium was studied and found that SRB lacks ideal resistance to some ions such as uranium and fluorine, so its application range is limited in practical applications [77]. It is precisely because of the complexity of the components of uranium-containing wastewater and the multiplicity of biological stress factors that the simple indigenous bacteria are not enable to fully play their role in reducing uranium to repairing the environment.

D. radiodurans has a strong tolerance to radiation. In recent years, many good results have been achieved by applying D. radiodurans directly to the treatment of uranium-containing wastewater or applying genetic engineering bacteria of D. radiodurans to the treatment of uranium-containing wastewater. However, due to the complexity and multiple stress factors of radioactive uranium wastewater, there will be many disturbances in the actual treatment process. Therefore, the construction of a D. radiodurans genetically engineered bacteria that can withstand multiple extreme stresses has great important practical significance for the treatment of radioactive uranium-containing wastewater and also has good development prospects in the research of uranium-containing wastewater biological treatment.