Carbonate mineralizing bacterium and application thereof

Abstract

According to the carbonate mineralizing bacteria and the application thereof, firstly, the bacteria C7-12 with the best cadmium removal effect are screened out from the carbonate mineralizing bacteria separated from lead-zinc mining area polluted soil and are preserved in Guangdong Microbial Culture Collection Center (GDMCC) at present, and the preservation number is GDMCC 65568. The carbonate mineralizing bacteria provided by the invention have strong Cd resistance, and the Cd removal rate of 1mg / L Cd solution can reach 85% after the carbonate mineralizing bacteria are cultured for 48 hours under the conditions that the pH is 6, the temperature is 30 DEG C and the urea concentration is 20g / L; the bacterial strain can simultaneously reduce the concentrations of Cu < 2 + >, Pb < 2 + >, Zn < 2 + > and Cd < 2 + > in a soil solution, the removal rates are 33-65% (Cd), 28-32% (Pb), 22-49% (Zn) and 38-44% (Cu) respectively, the heavy metal contaminated soil solution is repaired, and the carbonate mineralized bacteria provided by the invention have the advantages of no secondary pollution, obvious repair effect and easiness in operation, and have wide application prospects in the aspect of soil heavy metal pollution repair.

Description

Technical Field

[0001] The present invention relates to the technical field of soil heavy metal remediation, and specifically to a carbonate mineralizing bacterium and its application. Background Art

[0002] Frequent human activities, such as industrial emissions and agricultural activities, have increased the activation and migration of heavy metals. Heavy metals eventually accumulate in the soil, leading to soil heavy metal pollution. Due to the characteristics of heavy metal pollution such as concealment, persistence, environmental mobility, and bioaccumulation, they are difficult to directly detect with the naked eye and can accumulate in the soil for a long time, posing a potential threat to the environment. Among common heavy metals, Cd has the greatest biological toxicity and has been harming the entire ecosystem. Therefore, remediating heavy metal pollution, especially Cd pollution, is particularly important for maintaining the stability and health of the ecosystem.

[0003] Traditional soil heavy metal remediation methods include soil replacement, chemical passivation, etc. However, these methods are costly and may damage the soil structure, which is not conducive to the sustainable use of the soil. Microbial remediation refers to using microbial metabolic activities to reduce the bioavailability and environmental mobility of heavy metals, thereby alleviating heavy metal pollution. Carbonate mineralizing bacteria are widely present in heavy metal polluted soils and have high urease production ability. Their characteristic is to use biomineralization, that is, by secreting urease to hydrolyze urea to produce CO3 2- 2−, inducing heavy metal ions to form stable mineral compounds, and adsorbing the mineral compounds on the cell surface to effectively reduce the availability and migration ability of heavy metal ions. Another product NH4 + produced by urea hydrolysis can increase the pH value of the environment, avoiding the competition between surrounding H + + and heavy metal ions for adsorption sites and reducing the possibility of environmental acidification damaging the mineral stability. Summary of the Invention

[0004] Aiming at the above problems, the present invention provides a carbonate mineralizing bacterium and its application, which can efficiently remove Cd 2+ from the soil solution, and its removal ability for various heavy metals in the soil solution proves the feasibility of using carbonate mineralizing bacteria to remediate actual soil heavy metal pollution.

[0005] One technical object of the present invention is to provide a carbonate mineralizing bacterium, which is Serratia marcescens C7-12. The preservation unit is Guangdong Provincial Microbial Culture Collection Center, the preservation date is December 02, 2024, and the preservation number is GDMCC NO.65568.

[0006] Further preferably, the application of the carbonate mineralizing bacterium in the removal of heavy metal ions.

[0007] Further preferably, the carbonate mineralizing bacteria are used in the preparation of a heavy metal ion removal bacterial suspension.

[0008] Further preferably, the OD600 value of the bacterial suspension is 0.4 - 0.5.

[0009] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0010] The present invention screens out a native carbonate mineralizing bacterium C7-12, optimizes the culture medium conditions of strain C7-12, and obtains the culture conditions under the best cadmium removal rate. In addition, strain C7-12 can also remove multiple heavy metal ions in the soil solution simultaneously, and has the ability to repair actual heavy metal pollution. Using carbonate mineralizing bacteria has no secondary pollution, obvious repair effect, and is easy to operate, and has broad application prospects in the repair of soil heavy metal pollution. Description of the Drawings

[0011] Figure 1 For the Cd removal effects of 5 Cd-tolerant urease-producing bacteria at a Cd concentration of 5 mg / L 2+ concentration of 5 mg / L 2+ removal effects;

[0012] Figure 2 For the effects of environmental factors (temperature, pH, urea concentration, bacterial suspension dosage, culture time, and initial Cd concentration) on the Cd removal effect of strain C7-12;

[0013] Figure 3 For the removal of Cu2+, Pb2+, Zn2+, and Cd2+ by strain C7-12. Detailed Embodiments

[0014] In order to enable those skilled in the art to better understand the technical solutions of the present invention, the present invention will be further described in detail below in conjunction with embodiments. Those skilled in the art will understand that the following embodiments are only used to illustrate the present invention and should not be construed as limiting the scope of the present invention.

[0015] Example 1

[0016] Five urease-producing bacteria screened from the farmland soil around a lead-zinc mine in Huize County, Yunnan Province were used as the test strains. One loop of bacteria was picked and cultured on a solid plate for 24 h for amplification. The amplified bacteria were transferred to an LB liquid medium and cultured at 30 °C and 160 rpm for 24 h, centrifuged at 4 °C and 8000 rpm for 10 min, and the bacterial cell precipitate was resuspended in sterile water to prepare a bacterial suspension (OD 600 = 0.4). The bacterial suspension was added at 2% (v / v) to the Cd concentration (5, 10, 20, 40 mg·L -1In the LB medium, three replicates were set for each treatment and cultured at 30 °C and 160 rpm for 48 h. Centrifuge at 4 °C and 8000 rpm for 10 min, and use a flame atomic spectrophotometer to measure the remaining Cd concentration in the supernatant. Through differential analysis, the urease-producing bacteria with the strongest Cd removal ability were finally screened out. The Cd removal rate calculation formula is as follows:

[0017]

[0018] As Figure 1 shown, under 5-40 mg / L Cd stress, the Cd 2+ removal effect of 5 Cd-tolerant urease-producing bacteria was the best when the Cd 2 + concentration was 5 mg / L. Differential analysis found that when the Cd 2+ concentration was 5 mg / L, the removal rate of strain C7-12 was significantly higher than that of other strains (P<0.05), which was 72%. With the increase of concentration, C7-12 still showed good Cd removal effect. When the Cd 2+ concentration was 40 mg / L, its Cd removal rate was significantly lower than that of other urease-producing bacteria. In summary, strain C7-12 will be used as the test strain for subsequent research and will be preserved on December 02, 2024. It is currently preserved in the Guangdong Microbial Culture Collection Center (GDMCC), and the preservation number is GDMCC NO.65568.

[0019] Example 2

[0020] (1) The strain C7-12 bacterial suspension was inoculated into the urea (30 g / L) LB liquid medium with Cd concentrations of 0, 1, and 5 mg / L at a volume ratio of 2%, and cultured at 30 °C and 160 rpm for 96 h. Samples were continuously taken at 0, 4, 8, 12, 24, 36, 48, 60, 72, and 96 h. Centrifuge at 4 °C and 8000 rpm for 10 min, measure the Cd concentration in the supernatant, calculate the Cd removal rate, and determine the initial Cd concentration and culture time when the removal rate is the highest.

[0021] (2) The strain C7-12 bacterial suspension was inoculated into the urea (30 g / L) LB liquid medium with a Cd concentration of 1 mg / L at volume ratios of 1%, 2%, 4%, and 8%, and cultured at 30 °C and 160 rpm for 48 h. Centrifuge at 4 °C and 8000 rpm for 10 min, measure the Cd concentration in the supernatant, calculate the Cd removal rate, and determine the dosage of the bacterial suspension when the removal rate is the highest.

[0022] (3) The strain C7-12 bacterial suspension was inoculated into LB liquid medium containing different urea concentrations (0, 10, 20, 30, 40, 50 g / L) with a Cd concentration of 1 mg / L at a volume ratio of 2%, cultured at 30 °C and 160 rpm for 48 h, centrifuged at 4 °C and 8000 rpm for 10 min, the Cd concentration in the supernatant was measured, the Cd removal rate was calculated, and the dosage of the bacterial suspension when the removal rate was the highest was determined.

[0023] (4) The strain C7-12 bacterial suspension was inoculated into LB liquid medium containing a urea concentration of 20 g / L with a Cd concentration of 1 mg / L at a volume ratio of 2%, cultured at 160 rpm for 48 h at temperatures of 20, 25, 30, 35 °C respectively, centrifuged at 4 °C and 8000 rpm for 10 min, the Cd concentration in the supernatant was measured, the Cd removal rate was calculated, and the culture temperature when the removal rate was the highest was determined.

[0024] (5) The strain C7-12 bacterial suspension was inoculated into LB liquid medium containing urea (20 g / L) with a Cd concentration of 1 mg / L at a volume ratio of 2%, the pH of the medium was 4, 5, 6, 7, 8, 9, 10 respectively, cultured at 30 °C and 160 rpm for 48 h, centrifuged at 4 °C and 8000 rpm for 10 min, the Cd concentration in the supernatant was measured, the Cd removal rate was calculated, and the pH when the removal rate was the highest was determined.

[0025] As Figure 2 shown, for the effects of environmental factors (temperature, pH, urea concentration, dosage of bacterial suspension, culture time and initial Cd concentration) on the Cd removal effect of strain C7-12, it can be known from the experimental results that under the conditions of a culture time of 48 h, an initial Cd concentration of 1 mg / L, a temperature of 30 °C, a pH of 6.0, a urea concentration of 20 g / L, and a dosage of the bacterial suspension of 2%, the Cd removal rate of the solution reached 85%.

[0026] Example 3

[0027] (1) Soil solution medium: 10 g / L of tryptone, 5 g / L of sodium chloride, and 5 g / L of yeast powder were added to 1 L of slag, contaminated soil and farmland soil solution, the pH was 6.0, sterilized at 121 °C for 30 min, after cooling, the urea solution (20 g / L) was added to the medium using a syringe-type sterile filter (0.45 μm filter membrane).

[0028] (2) The strain C7-12 bacterial suspension was inoculated into the medium in step (1) at a volume ratio of 2%, cultured at 30 °C and 160 rpm for 48 h, centrifuged at 4 °C and 8000 rpm for 10 min, the heavy metal (Cd, Pb, Cu, Zn) concentrations in the supernatant were measured, and the heavy metal ion removal rate was calculated.

[0029] AsFigure 3 As shown, from the experimental results, Cd 2+ , Pb 2+ , Zn 2+ and Cu 2+ had removal rates of 33 - 65% (Cd), 28 - 32% (Pb), 22 - 49% (Zn), and 38 - 44% (Cu), respectively.

[0030] The above embodiments are merely explanations of the present invention and do not limit the present invention. Those skilled in the art can make modifications to these embodiments without creative contributions according to their needs after reading this specification, but as long as they are within the scope of the claims of the present invention, they are protected by the patent law.

Claims

1. A carbonate-mineralizing bacterium, characterized in that: The carbonate mineralizing bacterium is Serratia marcescens C7-12, preservation unit: Guangdong Microbial Culture Collection Center, preservation date: December 2, 2024, preservation number: GDMCC NO. 65568.

2. Use of a carbonate mineralizing bacterium according to claim 1, characterized in that: Application of the carbonate mineralizing bacterium in heavy metal ion removal.

3. Use of a carbonate mineralizing bacterium according to claim 2, characterized in that: Application of the carbonate mineralizing bacterium in the preparation of a heavy metal ion removal bacterial suspension.

4. Use of a carbonate mineralizing bacterium according to claim 3, characterized in that, The OD600 value of the bacterial suspension is 0.4 - 0.5.

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