CN114410509A - Bacterium agent and composition of enterobacter aerogenes combined klebsiella aerogenes and application of bacterium agent and composition - Google Patents
Bacterium agent and composition of enterobacter aerogenes combined klebsiella aerogenes and application of bacterium agent and composition Download PDFInfo
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Abstract
The invention provides a microbial inoculum of Enterobacter aerogenes combined Klebsiella aerogenes, which is used for treating lead-cadmium polluted soil and comprises the Enterobacter aerogenes and the Klebsiella aerogenes; the Enterobacter aerogenes is Enterobacter aerogenes (Enterobacter aerogenes) W6, is classified and named as Enterobacter aerogenes by Latin, is stored in China general microbiological culture Collection center (CGMCC), has the storage date of 2021, 7 and 13 days and has the storage number of CGMCC No. 22888; the Klebsiella aerogenes is Klebsiella aerogenes Wn, the Latin classification name is Klebsiella aerogenes, the Klebsiella aerogenes is stored in the common microorganism center (CGMCC) of the China Committee for culture Collection of microorganisms, the storage date is 2021, 7 and 13 days, and the storage number is CGMCC No. 22889.
Description
Technical Field
The invention relates to a microorganism and application thereof, in particular to a microbial inoculum, a composition and application of enterobacter aerogenes combined klebsiella aerogenes.
Background
After decades of industrial development, the heavy metal content of soil exceeds the standard due to a large amount of mining and smelting, and the threat to human survival is brought. According to statistics, about 500 million tons of lead (Pb) and about 3 million tons of cadmium (Cd) are discharged per year in the world on average. The lead and cadmium pollution in the soil has the migration capacity, and lead and cadmium contents of surface water and underground water can exceed the standard after the lead and cadmium pollution is subjected to rainwater leaching, vertical infiltration, surface runoff and the like. Health diseases such as carcinogenesis, diseases and mutation can be caused after drinking underground water polluted by heavy metal lead and cadmium or eating food chain products polluted by lead and cadmium. Therefore, the heavy metal pollution technology which has low cost, no pollution, high efficiency and can repair the soil for a long time becomes the current first consideration.
At present, the soil heavy metal remediation technology mainly comprises a physical remediation method, a chemical remediation method, a biological remediation method and the like. Compared with other restoration technologies, the biological-chemical composite restoration technology is a method for treating heavy metals in soil widely by using the heavy metal in-situ restoration technology, which has the advantages of wide application range, low cost and short restoration period. The essence of the biological-chemical composite remediation technology is that biological and chemical substrates are put into the heavy metal contaminated soil, and a series of complex reactions are generated between chemical reagents and heavy metals under the specific action of organisms, so that the aim of solidifying and stabilizing the heavy metals in the soil is fulfilled, for example: the method not only solves the difficulty of secondary pollution of a chemical remediation method but also solves the problem of long remediation period of a biological remediation method by adopting phosphorus-solubilizing microorganisms matched with an insoluble phosphorus source to treat the soil polluted by heavy metals.
Although the existing biological-chemical composite remediation technology can be used for treating lead and cadmium polluted soil to a certain extent, the microorganisms and the treatment method adopted in the prior art are not particularly outstanding in practical applicability and are easy to generate a reverse dissolution phenomenon. Up to now, the effectiveness and long-term stability of soil pollution remediation are still difficult technical problems to solve.
In view of the above, there is a need to provide a bacterium agent, a composition and an application of Enterobacter aerogenes combined with Klebsiella aerogenes, so as to solve or at least alleviate the technical defects of poor effectiveness and long-term stability of soil treatment.
Disclosure of Invention
The invention mainly aims to provide a bacterium agent of enterobacter aerogenes combined with klebsiella aerogenes, a composition and application, and aims to solve the technical problems of poor effectiveness and long-term stability of soil treatment in the prior art.
In order to achieve the purpose, the invention provides a bacterium agent of Enterobacter aerogenes combined Klebsiella aerogenes, which is used for treating lead-cadmium polluted soil and comprises the Enterobacter aerogenes and the Klebsiella aerogenes;
wherein the Enterobacter aerogenes is Enterobacter aerogenes (Enterobacter aerogenes) W6, the Latin article is classified and named as Enterobacter aerogenes, and is stored in China general microbiological culture Collection center (CGMCC), the storage date is 2021, 7 and 13 days, and the storage number is CGMCC No. 22888;
the Klebsiella aerogenes is Klebsiella aerogenes Wn, the Latin classification name is Klebsiella aerogenes, the Klebsiella aerogenes is stored in the common microorganism center (CGMCC) of the China Committee for culture Collection of microorganisms, the storage date is 2021, 7 and 13 days, and the storage number is CGMCC No. 22889.
The invention also provides application of the microbial agent in treating lead and cadmium polluted soil.
The invention also provides a composition for treating lead-cadmium polluted soil, which is characterized by comprising a component A, a component B, a component C and a component D;
wherein component a comprises a source of phosphorus;
the component B comprises the microbial agent as described in any one of the above;
the component C comprises an aqueous solution containing glucose, magnesium chloride, magnesium sulfate, ammonium sulfate and potassium chloride;
the component D comprises an aqueous solution containing tryptone, yeast extract and sodium chloride.
Further, the phosphorus source is tricalcium phosphate.
The invention also provides a method for treating lead and cadmium polluted soil, which adopts the composition to treat the lead and cadmium polluted soil.
Further, comprising: applying the component A and a first mixed liquor containing the component B and the component C to soil to be treated for a first period of time;
applying a second mixed liquor containing the component B and the component D to the soil to be remediated for a second time period;
wherein the second time period is later than the first time period.
Further, still include: and applying the first mixed liquor to the soil to be treated between the first time period and the second time period.
Further, applying the first mixed liquor to the soil to be treated for multiple times between the first time period and the second time period, wherein the single application amount is consistent with the total application amount of the first mixed liquor in the first time period;
and applying the second mixed liquor to the soil to be treated for multiple times in the second time period, wherein the single application amount is consistent with the total application amount of the first mixed liquor in the first time period.
Further, the interval duration of the first time period and the second time period is 14-16 days.
The Latin classification of the Enterobacter aerogenes W6 used in the invention is named as Enterobacter aerogenes, and is preserved in China general microbiological culture Collection center (CGMCC), address: no. 3 Xilu No. 1 Hospital, Chaoyang, China, the preservation date is 2021 years, 7 months and 13 days, and the preservation number is CGMCC No. 22888.
The Enterobacter aerogenes W6 related in the invention is a gram-negative bacterium separated from the bottom sediment of a domestic sewage sedimentation fermentation tank of the leading ocean lake reclaimed water Limited company in Hunan of Changsha, Hunan, and forms a round, convex and grey-white colony during culture, and the Enterobacter aerogenes W6 has the advantages of appropriate growth temperature of 30-35 ℃, appropriate growth PH of 6.9-7.2, and aerobic or facultative anaerobic property.
The latin classification of Klebsiella aerogenes Wn used in the present invention is named Klebsiella aerogenes, and has been deposited in the common microorganism center of china committee for culture collection and management (CGMCC), address: no. 3 Xilu No. 1 Bichen of Chaoyang district, Beijing, China, the preservation date is 7 months and 13 days in 2021, and the preservation number is CGMCC No. 22889.
The Klebsiella aerogenes Wn involved in the invention is gram-negative bacteria separated from the bottom sludge of the domestic sewage sedimentation fermentation tank of the lake-south leading ocean lake reclaimed water Limited company in the lake-south province of Changsha; the bacterial colony is in a circular shape with obvious bulges, and forms a larger white mucus bacterial colony on an agar culture medium, and the middle of the bacterial colony is provided with less yellow mucus. The Klebsiella aerogenes Wn has a suitable growth temperature of 30-35 deg.C, a suitable growth pH of 6.9-7.2, and is aerobic or facultative anaerobic.
Compared with the prior art, the invention has the following advantages:
the invention provides a microbial agent for treating lead-cadmium polluted soil, which is beneficial to the phosphorus-containing characteristics of Enterobacter aerogenes (Enterobacter aeogens) W6 and Klebsiella aerogenes (Klebsiella aeogens) Wn, and can be used together with two strains with the same original source, so that a synergistic effect can be generated between the strains, and the diversity of phosphate solubilizing bacteria in the soil is improved; in addition, the invention also utilizes the characteristics of acid and alkali production of microorganisms in different culture media, thereby not only promoting the phosphorus dissolution of the microorganisms, but also avoiding the reverse dissolution of passivation products.
Specifically, the Enterobacter aerogenes (Enterobacter aeogens) W6 can passivate 99.7% of lead and cadmium in 15 days in combination with tricalcium phosphate; in a soil test, on the 7 th day, the treatment of the Enterobacter aerogenes W6 composite tricalcium phosphate can lead the maximum removal rate of the effective state contents of Pb and Cd in soil to be 45.5 percent and 40.9 percent respectively; at day 15, the highest removal rate of the effective state content is respectively reduced to 30.1 percent and 23.5 percent, and at day 30, the removal rates of the effective state content of Pb and Cd respectively reach 45 percent and 42 percent again, so that the phenomenon of reverse dissolution is effectively inhibited.
The Klebsiella aerogenes Wn combined with tricalcium phosphate can passivate lead and cadmium by 100 percent in 1 day; in a soil test, at the 7 th day, the treatment of the Klebsiella aerogenes Wn composite tricalcium phosphate can lead the highest removal rate of the effective state contents of Pb and Cd in the soil to be 49 percent and 49.3 percent respectively; on day 15, the maximum removal rates of the effective state contents are respectively reduced to 45.1% and 46.4%, and on day 30, the removal rates of the effective state contents of Pb and Cd respectively reach 51% and 51.3%, so that the phenomenon of reverse dissolution is effectively inhibited.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, it is obvious that the drawings in the following description are only some embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to the structures shown in the drawings without creative efforts.
FIG. 1 shows the phosphorus solubilizing characteristics of Enterobacter aerogenes W6 and Klebsiella aerogenes Wn cultured on solid medium 2 for 7 days in example 1;
FIG. 2 is a graph showing the effect of phosphorus content in Enterobacter aerogenes W6 in example 1;
FIG. 3 is a graph showing the effect of phosphorus content in Klebsiella aerogenes Wn in example 1;
FIG. 4 is a construction diagram of a phylogenetic tree of Enterobacter aerogenes (Enterobacter aeogens) W6 in example 1;
FIG. 5 is a drawing showing the construction of a phylogenetic tree of Klebsiella aerogenes Wn in example 1;
FIG. 6 is an XRD pattern of the reaction products of Enterobacter aerogenes W6 and Klebsiella aerogenes Wn for phosphorus-solubilizing and lead-cadmium-inactivating in example 2;
FIG. 7 shows the phosphorus solubilizing effect and pH dependence of Enterobacter aerogenes W6 in example 3 in various media;
FIG. 8 shows the phosphorus solubilizing effect and pH dependence of Klebsiella aerogenes Wn in different media in example 3.
The implementation, functional features and advantages of the present invention will be further explained with reference to the accompanying drawings.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention, and it is apparent that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be obtained by a person skilled in the art without any inventive step based on the embodiments of the present invention, are within the scope of the present invention.
It should be noted that all the directional indicators (such as the upper and lower … …) in the embodiment of the present invention are only used to explain the relative position relationship, movement, etc. of the components in a specific posture (as shown in the drawings), and if the specific posture is changed, the directional indicator is changed accordingly.
In addition, the descriptions related to "first", "second", etc. in the present invention are only for descriptive purposes and are not to be construed as indicating or implying relative importance or implicitly indicating the number of technical features indicated. Thus, a feature defined as "first" or "second" may explicitly or implicitly include at least one such feature.
Moreover, the technical solutions in the embodiments of the present invention may be combined with each other, but it is necessary to be able to be realized by a person skilled in the art, and when the technical solutions are contradictory or cannot be realized, the combination of the technical solutions should be considered to be absent, and is not within the protection scope of the present invention.
In order to complete the treatment of lead and cadmium polluted soil, the invention provides a bacterium agent of Enterobacter aerogenes combined Klebsiella aerogenes, which is used for treating lead and cadmium polluted soil and comprises the Enterobacter aerogenes and the Klebsiella aerogenes;
wherein the Enterobacter aerogenes is Enterobacter aerogenes (Enterobacter aerogenes) W6, the Latin article is classified and named as Enterobacter aerogenes, and is stored in China general microbiological culture Collection center (CGMCC), the storage date is 2021, 7 and 13 days, and the storage number is CGMCC No. 22888;
the Klebsiella aerogenes is Klebsiella aerogenes Wn, the Latin classification name is Klebsiella aerogenes, the Klebsiella aerogenes is stored in the common microorganism center (CGMCC) of the China Committee for culture Collection of microorganisms, the storage date is 2021, 7 and 13 days, and the storage number is CGMCC No. 22889.
The Enterobacter aerogenes (Enterobacter aerogenes) W6 and the Klebsiella aerogenes (Klebsiella aerogenes) Wn both have excellent phosphate solubilizing effect and belong to the category of phosphate solubilizing bacteria. Wherein, the phosphorus dissolving amount of the Enterobacter aerogenes (Enterobacter aerogenes) W6 can reach 622.2mg/L and the phosphorus dissolving amount of the Klebsiella aerogenes (Klebsiella aerogenes) Wn can reach 683.1mg/L within 7 days. The two strains can convert indissolvable compound phosphorus in the soil into soluble phosphorus, so that the two strains are used for passivating lead and cadmium, and the problems of water eutrophication, soil acidification, plant nutrition unbalance and the like caused by directly adding soluble phosphate compounds are avoided. Therefore, the lead-cadmium polluted soil can be effectively treated by using the Enterobacter aerogenes (Enterobacter aerogenes) W6 and the Klebsiella aerogenes (Klebsiella aerogenes) Wn.
Specific passivation processes may include: microorganisms, tricalcium phosphate and other insoluble phosphorus sources in the microbial inoculum enter polluted soil, heavy metals (such as Pb, Cd, Cu and Zn) can be fixed, and highly insoluble metal phosphate precipitates are formed, so that the migration rate and the availability of Pb and Cd are obviously reduced, wherein the passivating product of cadmium is Ca7.7Cd0.8(PO4)8(H2O)2.4(ii) a The passivating product of lead is Pb5(PO4)3Cl。
It is to be understood that the invention also carries out comparative research on some strains with phosphate solubilizing capability in the existing phosphate solubilizing bacteria. For example: in the existing research, according to the content disclosed in screening and verifying phosphate solubilizing effect of a strain of efficient phosphate solubilizing bacteria, Tang chen (2020) and the like screen out a strain of efficient phosphate solubilizing bacteria from a farmland: the bacillus beilesensis is identified by 16 sr DNA analysis, the phosphorus dissolution amount reaches 495.4mg/L, and the phosphorus dissolution amount reaches 582.4mg/L after condition optimization. However, the phosphorus-dissolving effect is still lower than that of W6 and Wn listed in the present embodiment of the invention.
On the basis, the Enterobacter aerogenes (Enterobacter aerogenes) W6 and the Klebsiella aerogenes (Klebsiella aegerenes) Wn are simultaneously used as microbial inoculum for treating the lead-isolated polluted soil, so that the diversity of phosphate solubilizing bacteria can be improved; moreover, since the original sources of the two strains of the invention are the same and the efficacies are similar, a synergistic effect can be produced, for example: both can produce acid, and the PH dropping speed in the environment can be increased under the combined action, so that the phosphorus dissolving efficiency can be further improved; in addition, it is to be noted that the Enterobacter aerogenes (Enterobacter aeogenes) W6 and the Klebsiella aerogenes (Klebsiella aerogenes) Wn can produce organic acids and carbon dioxide for promoting the stabilization of heavy metal minerals.
In view of the above, the invention further provides an application of the microbial inoculum according to any embodiment in treating lead-cadmium polluted soil.
In order to facilitate development of treatment work and further improve treatment efficiency, the invention also provides a composition for treating lead-cadmium polluted soil, which comprises a component A, a component B, a component C and a component D;
wherein, the component A comprises a phosphorus source, the phosphorus source refers to a difficultly soluble phosphorus source, and preferably can be tricalcium phosphate.
The component B comprises the microbial inoculum according to any embodiment above.
The component C comprises an aqueous solution containing glucose, magnesium chloride, magnesium sulfate, ammonium sulfate and potassium chloride.
The component C is liquid, and the preparation process of the component C can be as follows: weighing 48.7% of glucose, 24.3% of magnesium chloride, 1.2% of magnesium sulfate, 0.97% of ammonium sulfate, 0.49% of potassium chloride and 24.34% of tricalcium phosphate which account for the first preset mass according to mass percent, then weighing the glucose, the magnesium chloride, the magnesium sulfate, the ammonium sulfate and the potassium chloride (tricalcium phosphate can be scattered in soil, so tricalcium phosphate can not be added), and then adding water into a container containing the weighed components to fix the volume; the first preset mass is the sum of the masses of the glucose, the magnesium chloride, the magnesium sulfate, the ammonium sulfate, the potassium chloride and the tricalcium phosphate, and the mass-to-volume ratio of the first preset mass to the component C can be 2.055 g: 100 ml.
For example: when 100ml of component C is prepared, 1g of glucose, 0.5g of magnesium chloride, 0.025g of magnesium sulfate, 0.02g of ammonium sulfate, 0.01g of potassium chloride and 0.5g of tricalcium phosphate are added into 100ml of water according to calculation requirements; however, since tricalcium phosphate is applied to the soil in advance during actual treatment, tricalcium phosphate may not be added to the component C, and only 1g of glucose, 0.5g of magnesium chloride, 0.025g of magnesium sulfate, 0.02g of ammonium sulfate, and 0.01g of potassium chloride may be added to 100ml of water; of course, the solid may be weighed and then the volume may be increased to 100 ml.
The component D comprises an aqueous solution containing tryptone, yeast extract and sodium chloride.
The component D is liquid, and the preparation process of the component D can be as follows: weighing tryptone accounting for 40% of the second preset mass, yeast powder accounting for 20% of the second preset mass and sodium chloride accounting for 40% of the second preset mass according to mass percentage, and then adding water into a container containing the components to fix the volume; wherein the second predetermined mass is the total mass of tryptone, yeast extract and sodium chloride expected to be applied to the soil, and the mass to volume ratio of the second predetermined mass to the component D may be 2.5 g: 100 ml.
For example: when 100ml of component D is prepared, 1g of tryptone, 0.5g of yeast extract and 1g of sodium chloride may be added to 100ml of water; of course, the solid may be weighed and then the volume may be increased to 100 ml.
It is noted that although lead, cadmium and the like can be inactivated by the dual action of microorganisms and poorly soluble phosphorus sources, although, as the reaction time increases, desolvation phenomena occur, such as: the highly insoluble metal phosphate precipitate forms CdCO3、CdPO4And the like.
It is to be understood that, through experimental research, the microorganisms in the microbial inoculum can generate acid production phenomenon and phosphorus dissolution phenomenon when being cultured in the component C; while the occurrence of an alkali-producing phenomenon and the suppression of the occurrence of a reverse-dissolving phenomenon occur when cultured in the component D.
Therefore, the technology uses the characteristics of microorganisms cultured in different components from the perspective of reaction products to regulate and control the pH intensity from acid (cultured in the component C) to alkali (cultured in the component D) so as to naturally fix and stabilize the lead and the cadmium, avoid the occurrence of reverse dissolution and creatively find a long-acting mechanism for producing phosphorus by regulating and controlling the pH and applying the phosphorus to the heavy metal pollution control.
Based on the composition, the invention also provides a method for treating lead and cadmium polluted soil, which adopts the composition in any embodiment to treat the lead and cadmium polluted soil.
Specifically, the method for treating the lead-cadmium polluted soil comprises the following steps: applying the component A and a first mixed liquor comprising the component B and the component C to the soil to be remediated for a first period of time.
It should be noted that the first mixed solution may be similar to a culture solution obtained by breeding the microorganisms in the microbial inoculum in the component C, and the two microorganisms in the microbial inoculum may be applied to the soil after being bred in the component C separately or after being co-cultured. In the specific test process, the liquid is usually taken directly from the culture solution of the microorganism, and during the previous culture of the strain, the culture solution contains part of tricalcium phosphate, so that tricalcium phosphate can be contained in the first mixed solution, and the actual effect is not influenced. In a specific application, a person skilled in the art can selectively determine whether to add part of the tricalcium phosphate in the component C according to actual conditions.
Applying a second mixed liquor containing the component B and the component D to the soil to be remediated for a second time period; the second mixed solution can be similar to a culture solution obtained after the microorganisms propagate in the component D, and two microorganisms in the microbial inoculum can be applied to soil after the components D are cultured separately or after the components D are cultured together.
Wherein the second time period is later than the first time period.
In addition, between the first time period and the second time period, the first mixed solution can be applied to the soil to be treated so as to maintain the soil water content and the activity of bacteria.
As an explanation of the above embodiment: the first time period is the initial time, namely the component A (including the indissolvable phosphorus source) and the first mixed solution (namely the component C with the microorganisms in the microbial inoculum) are applied to the soil to be treated from the beginning, so that the lead and cadmium in the soil are passivated.
The interval between the first time period and the second time period may be 14 to 16 days, specifically, the interval between the second time period and the first time period may be 15 days, that is, after 15 days of the first time period, the second mixed solution may be applied to the soil to be treated, so as to inhibit the occurrence of a reverse dissolution phenomenon, thereby enhancing a passivation effect.
In addition, the first mixed liquid is applied between the first time period and the second time period, so that the phosphorus dissolving efficiency is enhanced, the growth of microorganisms is promoted, and meanwhile, the phosphorus dissolving effect is ensured to be generated in the early stage, and the passivation of lead and cadmium in the early stage is ensured.
Further, the first mixed liquor is applied to the soil to be treated for multiple times between the first time period and the second time period, and the single application amount is consistent with the total application amount of the first mixed liquor in the first time period. And applying the second mixed liquor to the soil to be treated for multiple times in the second time period, wherein the single application amount is consistent with the total application amount of the first mixed liquor in the first time period.
To facilitate a further understanding of the invention, reference will now be made to the following examples:
example 1
Screening and identification of Enterobacter aerogenes W6
1. The culture conditions are as follows: aerobically, at 30 ℃, at an initial PH of 7 (i.e. the initial PH of the medium is 7); wherein, the shaking table culture at 150rpm/min is adopted when the culture is carried out under the liquid condition.
2. Preparing a culture medium: liquid culture medium 1, liquid culture medium 2, solid culture medium 2.
Wherein, the preparation process of the liquid culture medium 1 comprises the following steps: calculating the mass of the solid component according to the capacity and the solid-liquid concentration (25g/L) which are prepared as required, then weighing 40% of tryptone, 20% of yeast powder and 40% of sodium chloride which account for the mass percent of the solid component, and fixing the volume after weighing;
the preparation process of the liquid culture medium 2 comprises the following steps: calculating the mass of a solid component according to the capacity and the solid-liquid concentration (20.55g/L) which are prepared as required, then weighing 48.7 percent of glucose, 24.3 percent of magnesium chloride, 1.2 percent of magnesium sulfate, 0.97 percent of ammonium sulfate, 0.49 percent of potassium chloride and 24.34 percent of tricalcium phosphate which account for the mass percent of the solid component, and fixing the volume after weighing;
3. Weighing 1g of bottom mud obtained from domestic sewage precipitation fermentation tank of the pilot ocean lake reclaimed water of the limited company of Hunan, Changsha, Hunan, putting into a sterilized centrifuge tube, adding 9ml of 0.9% normal saline to prepare 10-1The bacterial suspension is put into a shaking incubator with the speed of 150rpm/min to shake for 20min, and then is taken out and stands for 2 h.
4. Standing the suspension, taking the supernatant, inoculating 2% of the supernatant to a liquid culture medium 2, and carrying out specific culture; placing on a shaking bed, keeping the temperature at 30 ℃, performing shake culture at 150rpm/min for 2-3 days. After culturing for a period of time, taking another 0.4ml of bacterial suspension and 4.5ml of ultrapure water for gradient dilution with the dilution multiple of 10-1、10-2、10-3、10-4、10-5、10-6Sample bacterial suspension of (4), 10-2、10-4、10-6And (3) taking 200 mu l of bacterial suspension with the concentration gradient, sequentially and uniformly coating the bacterial suspension on the solid culture medium 2, inverting the coated culture dish, putting the culture dish into a constant-temperature incubator, performing constant-temperature culture at the temperature of 30 ℃ for 24 hours, observing and recording the sizes of bacterial colonies and phosphorus dissolving rings, marking strains, and screening 5 strains of phosphorus dissolving bacteria.
5. Respectively carrying out plate streaking purification on the screened 5 phosphorus-solubilizing bacteria on a solid culture medium 2 (selecting standard plates of the phosphorus-solubilizing bacteria with obvious phosphorus-solubilizing rings from the 5 phosphorus-solubilizing bacteria in an aseptic operation table, streaking the standard plates to different solid culture mediums 2, placing the plates in an incubator at 30 ℃ for culture for a week), and then further selecting two phosphorus-solubilizing bacteria with obvious phosphorus-solubilizing effects, and marking the bacteria as W6 and Wn; wherein the phosphorus melting ring characteristics of W6 and Wn are shown in FIG. 1.
6. After dissolving phosphorus by the phosphorus dissolving bacteria, qualitatively analyzing the effective phosphorus content in the fermentation liquor: molybdenum antimony colorimetric resistance.
Respectively placing single strains of W6 and Wn in different liquid culture media 2 to culture for 7 days, and then detecting, wherein the specific detection process comprises the following steps: taking 1% of bacterial suspension into a sterilized centrifugal tube, centrifuging under the condition of 10000rpm, taking 100 mu L of supernatant, adding the supernatant into a 50ml colorimetric tube, adding ultrapure water to two thirds of a scale mark, then adding 1ml of the prepared solution A and 2ml of the prepared solution B, uniformly mixing, fixing the volume to the scale mark, standing for developing for 15min, measuring the absorbance value on an ultraviolet visible spectrophotometer, setting the wavelength to be 700nm, and calculating the corresponding effective phosphorus content according to a standard curve. In addition, in this example, a blank test of sterility was also conducted under the same conditions.
7. After the determination, it was concluded that: referring to FIGS. 2 and 3, the concentration of available phosphorus in the blank control was much lower than that in the culture solutions of W6 and Wn, and W6 and Wn were excellent in phosphate solubilizing effect when medium 2 was used as a nutrient source. Within 7 days, the phosphorus dissolving amount of W6 can reach 622.2mg/L, and the phosphorus dissolving amount of Wn can reach 683.1 mg/L.
8. Identification of phosphate solubilizing bacteria W6 and Wn: the strains were amplified and forward sequenced using primers and by comparing the sequences to those in the NCBI database.
(1) The similarity of W6 and Enterobacter aerogenes reaches 99%, and the Enterobacter aerogenes can be identified by constructing a developmental tree as shown in FIG. 4.
The 16s DNA gene sequence splicing determination result of the enterobacter aerogenes W6 strain is as follows:
Tgcaagtcgagcggtagcacagagagcttgctctcgggtgacgagcggcggacgggtgagtaatgtctgggaaactgcctgatggagggggataactactggaaacggtagctaataccgcataacgtcgcaagaccaaagtgggggaccttcgggcctcatgccatcagatgtgcccagatgggattagctagtaggtggggtaatggctcacctaggcgacgatccctagctggtctgagaggatgaccagccacactggaactgagacacggtccagactcctacgggaggcagcagtggggaatattgcacaatgggcgcaagcctgatgcagccatgccgcgtgtatgaagaaggccttcgggttgtaaagtactttcagcgaggaggaaggcgttaaggttaataaccttagcgattgacgttactcgcagaagaagcaccggctaactccgtgccagcagccgcggtaatacggagggtgcaagcgttaatcggaattactgggcgtaaagcgcacgcaggcggtctgtcaagtcggatgtgaaatccccgggctcaacctgggaactgcattcgaaactggcaggctagagtcttgtagaggggggtagaattccaggtgtagcggtgaaatgcgtagagatctggaggaataccggtggcgaaggcggccccctggacaaagactgacgctcaggtgcgaaagcgtggggagcaaacaggattagataccctggtagtccacgccgtaaacgatgtcgacttggaggttgtgcccttgaggcgtggcttccggagctaacgcgttaagtcgaccgcctggggagtacggccgcaaggttaaaactcaaatgaattgacgggggcccgcacaagcggtggagcatgtggtttaattcgatgcaacgcgaagaaccttacctactcttgacatccagagaacttagcagagatgctttggtgccttcgggaactctgagacaggtgctgcatggctgtcgtcagctcgtgttgtgaaatgttgggttaagtcccgcaacgagcgcaacccttatcctttgttgccagcgattcggtcgggaactcaaaggagactgccagtgataaactggaggaaggtggggatgacgtcaagtcatcatggcccttacgagtagggctacacacgtgctacaatggcatatacaaagagaagcgacctcgcgagagcaagcggacctcataaagtatgtcgtagtccggattggagtctgcaactcgactccatgaagtcggaatcgctagtaatcgtagatcagaatgctacggtgaatacgttcccgggccttgtacacaccgcccgtcacaccatgggagtgggttgcaaaagaagtaggtagcttaaccttcgggagggcgcttacc
(2) the similarity between Wn and the Klebsiella aerogenes reaches 99%, and the Klebsiella aerogenes can be identified by constructing a developmental tree as shown in figure 5.
The 16s DNA gene sequence splicing determination result of the Klebsiella aerogenes Wn strain is as follows:
Gccctcccgaaggttaagctacctacttcttttgcaacccactcccatggtgtgacgggcggtgtgtacaaggcccgggaacgtattcaccgtagcattctgatctacgattactagcgattccgacttcatggagtcgagttgcagactccaatccggactacgacatactttatgaggtccgcttgctctcgcgaggtcgcttctctttgtatatgccattgtagcacgtgtgtagccctactcgtaagggccatgatgacttgacgtcatccccaccttcctccagtttatcactggcagtctcctttgagttcccgaccgaatcgctggcaacaaaggataagggttgcgctcgttgcgggacttaacccaacatttcacaacacgagctgacgacagccatgcagcacctgtctcagagttcccgaaggcaccaaagcatctctgctaagttctctggatgtcaagagtaggtaaggttcttcgcgttgcatcgaattaaaccacatgctccaccgcttgtgcgggcccccgtcaattcatttgagttttaaccttgcggccgtactccccaggcggtcgacttaacgcgttagctccggaagccacgcctcaagggcacaacctccaagtcgacatcgtttacggcgtggactaccagggtatctaatcctgtttgctccccacgctttcgcacctgagcgtcagtctttgtccagggggccgccttcgccaccggtattcctccagatctctacgcatttcaccgctacacctggaattctacccccctctacaagactctagcctgccagtttcgaatgcagttcccaggttgagcccggggatttcacatccgacttgacagaccgcctgcgtgcgctttacgcccagtaattccgattaacgcttgcaccctccgtattaccgcggctgctggcacggagttagccggtgcttcttctgcgagtaacgtcaatcgctaaggttattaaccttaacgccttcctcctcgctgaaagtactttacaacccgaaggccttcttcatacacgcggcatggctgcatcaggcttgcgcccattgtgcaatattccccactgctgcctcccgtaggagtctggaccgtgtctcagttccagtgtggctggtcatcctctcagaccagctagggatcgtcgcctaggtgagccattaccccacctactagctaatcccatctgggcacatctgatggcatgaggcccgaaggtcccccactttggtcttgcgacattatgcggtattagctaccgtttccagtagttatccccctccatcaggcagtttcccagacattactcacccgtccgccgctcgtcacccgagagcaagctctctgtgttaccgctcgacttgca
example 2
Preparing lead and cadmium mother liquor: 1.5985g of lead nitrate and 20.317g of cadmium chloride are weighed and respectively dissolved in a beaker filled with 500ml of ultrapure water, simultaneously a glass rod is continuously stirred until the lead nitrate and the cadmium chloride are dissolved, then the lead nitrate and the cadmium chloride are respectively transferred into a 1L volumetric flask to be constant volume to scale marks, and labels are pasted for standby.
Function of Enterobacter aerogenes W6 in lead and cadmium passivation
1. A single colony was picked from a plate of W6 strain and cultured overnight in 20mL of liquid medium 1 (same as in example 1), and then transferred to 100mL of liquid medium 1 for expansion culture.
2. The growth density of the W6 strain was observed, and its growth curve was measured at a wavelength of 600nm using an ultraviolet spectrophotometer to determine the logarithmic growth phase and stationary phase OD of the W6 strain600The time point 1 was used for subsequent experiments; at 5.5h of culture, W6 reached OD6001 stable period.
4. Setting up a lead-cadmium solution experiment, adding 1ml of lead mother liquor and 1ml of cadmium mother liquor into a sterilized liquid culture medium 2 (same as example 1) through a sterile filter head (1 ml of lead mother liquor and 1ml of cadmium mother liquor are added into 100ml of liquid culture medium 2), and simultaneously carrying out expansion culture to OD600The bacterial suspension was inoculated into about 1% of the culture medium, and then cultured in a water bath with shaking at 30 ℃ and an initial pH of 7 at 150rpm/min, and the Pb content in the solution was measured periodically by ICP2+、Cd2+The concentration of (c) is varied.
In addition, a blank test of sterility was also performed under the same conditions in this example.
The results show that: under the conditions of aerobic shaking culture at constant temperature of 150rpm and constant temperature of 7 pH at 30 ℃ under normal pressure, the experimental group can hardly detect Pb on day 152+、Cd2+Is present. The passivation rate of lead and cadmium is 99.7%.
Second, application of Klebsiella aerogenes Wn (Klebsiella aerogenes) in lead and cadmium passivation
1. A single colony was picked from the Wn strain plate and cultured overnight in 20mL of liquid medium 1 (same as example 1), and then transferred to 100mL of liquid medium 1 for scale-up culture.
2. Observing the growth density of Wn strain, and determining its growth curve at 600nm wavelength by ultraviolet spectrophotometer to determine logarithmic growth phase and stationary phase OD of Wn strain600The time point 1 was used for subsequent experiments; wn reached OD at 5h of culture6001 stable period.
4. Setting up a lead-cadmium solution experiment, adding 1ml of lead mother liquor and 1ml of cadmium mother liquor into a sterilized liquid culture medium 2 (same as example 1) through a sterile filter head (1 ml of lead mother liquor and 1ml of cadmium mother liquor are added into 100ml of liquid culture medium 2), and simultaneously carrying out expansion culture to OD600The bacterial suspension was inoculated into about 1% of the culture medium, and then cultured in a water bath with shaking at 30 ℃ and an initial pH of 7 at 150rpm/min, and the Pb content in the solution was measured periodically by ICP2+、Cd2+The concentration of (c) is varied.
In addition, a blank test of sterility was also performed under the same conditions in this example.
The results show that: under the conditions of aerobic shaking culture at constant temperature of 150rpm/min at 30 ℃ and initial pH 7 under normal pressure, the ICP can not measure Pb on the 1 st day2+、Cd2+In this case, the lead-cadmium passivation is 100%.
Thirdly, identification of passivation products
The passivation reaction products after W6 and Wn were centrifuged respectively (centrifugation condition: 10000rpm centrifugation for 10min), the reaction precipitate remained (washed by 3 times with ultrapure water and centrifuged) was put into a 30-degree oven to be dried for one week, and the reaction precipitate was ground into powder after being completely dried to carry out XRD test.
The results show that: as will be appreciated with reference to FIG. 6, in both sets of W6 and Wn experiments, the reaction product of cadmium included Ca7.7Cd0.8(PO4)8(H2O)2.4(ii) a The reaction products of lead each include Pb5(PO4)3Cl。
Example 3
The liquid medium 1 used in this example was the same as that used in example 1, and glucose and tricalcium phosphate were additionally added in amounts equivalent to those of the liquid medium 2; the liquid medium 2 used in this example was the same as that used in example 1; the initial pH of the medium used in this example was 7.
Effect of different media on Enterobacter aerogenes W6
Absorbing 1% of culture bacteria liquid of W6, respectively putting the culture bacteria liquid into a culture medium 1 and a culture medium 2 as different nutrient sources for culture, placing the culture medium on a shaking table at a constant temperature of 30 ℃ and 150rpm/min, shaking and culturing for one week, taking 3ml of sample on an aseptic operating platform at the same time point every day into a 5ml centrifuge tube, centrifuging for 5 minutes at 8000rpm, taking supernatant, measuring the content change of available phosphorus by a molybdenum-antimony colorimetric method, and recording the corresponding pH value.
The results after the measurement showed that: as can be understood from FIG. 7, when Medium 2 was used as the nutrient source, the phosphate solubilizing effect was remarkable, and the pH value in the medium solution system was drastically lowered from neutral to about 4.
The control experiment, medium 1, showed no phosphorus dissolution and a steady increase in pH to about 9, indicating that the phosphorus removal efficiency of W6 correlated well with the pH.
Note that the solid line in fig. 7 corresponds to the amount of dissolved phosphorus, and the dotted line corresponds to PH.
Second, the action of different culture media on Klebsiella aerogenes Wn
And (3) respectively putting 1% Wn of culture bacteria liquid into a culture medium 1 and a culture medium 2 as different nutrient sources for culture, placing the culture medium on a shaking table at a constant temperature of 30 ℃ and at a speed of 150rpm/min, carrying out shake culture for one week, taking 3ml of sample on an aseptic operating platform at the same time point every day into a 5ml centrifuge tube, centrifuging for 5 minutes at 8000rpm, taking supernatant, measuring the content change of available phosphorus by a molybdenum-antimony colorimetric method, and recording the corresponding pH value.
The results after the measurement showed that: as can be understood from FIG. 8, when Medium 2 was used as the nutrient source, the phosphate solubilizing effect was remarkable, and the pH value in the medium solution system was drastically lowered from neutral to about 4.
The control experiment, medium 1, showed no phosphorus dissolution and a steady increase in pH to about 9, indicating that all of the phosphorus dissolution efficiencies of Wn have a certain correlation with pH.
Note that the solid line in fig. 8 corresponds to the amount of dissolved phosphorus, and the dotted line corresponds to PH.
To sum up: medium 1 and Medium 2 acted equally on W6 and Wn, allowing both strains to produce acid in Medium 2 and base in Medium 1.
Example 4
The liquid medium 1 used in this example was the same as the liquid medium 1 used in example 1; the liquid medium 2 contained in the bacterial liquid sprayed for the first time in this example was the same as in example 1; in the following interval sprayed microorganism-containing medium 2, tricalcium phosphate was not added to the liquid medium 2 to avoid the influence of the continuous addition of tricalcium phosphate on the soil, and the amounts of the remaining components of the liquid medium 2 were kept in accordance with those in example 1.
Application of Enterobacter aerogenes W6 in lead and cadmium polluted soil
1. Weighing 10g (40 meshes) of air-dried soil, placing the air-dried soil into a 50ml centrifuge tube, adding a phosphorus source (an insoluble phosphorus source: tricalcium phosphate) in a direct putting mode, and stirring by using a clean plastic rod to uniformly distribute the tricalcium phosphate in a soil sample;
2. spraying a prepared bacterial solution of enterobacter aerogenes W6 (the bacterial agent grows to OD in the culture medium 2) into the conical flask6001 stationary phase or logarithmic growth phase) and uniformly stirring to finally make the water content of the soil (bacterial liquid) be 30%;
3. medium 2(OD ═ 1) containing W6 was added every 3 days for the first 15 days to maintain 30% water content in the soil, and medium 1(OD ═ 1) containing W6 was added every 3 days for the last 15 days to maintain 30% water content in the soil. The content of the added phosphate radical and the contents of two heavy metal elements, namely Pb and Cd, in the soil keep a certain proportional relationship, specifically, the adding amount of tricalcium phosphate is related to the molar mass ratio of Pb (P: HMs-1: 1) to Cd (P: HMs-5: 1) in the soil, and finally the adding amount is determined to be 4 mg/g.
In addition, the present example also performed a sterile blank test under the same conditions.
The experimental results show that: compared with a blank control test, in 7 days, the highest removal rate of the effective state contents of Pb and Cd in the soil is 45.5% and 40.9% respectively by the treatment of the microorganism W6 microbial inoculum composite tricalcium phosphate; at 15 days, the maximum removal rate of the content of the effective state is respectively reduced to 30.1 percent and 23.5 percent, which shows that the back dissolution can occur only by the phosphorus dissolving effect. At 30 days, the removal rates of the effective state contents of Pb and Cd reach 45% and 42% respectively, which shows that the phenomenon of reverse dissolution can be inhibited by the matched use of the culture medium.
Second, application of Klebsiella aerogenes Wn in lead and cadmium polluted soil
1. Weighing 10g (40 meshes) of air-dried soil, placing the air-dried soil into a 50ml centrifuge tube, adding a phosphorus source (an insoluble phosphorus source: tricalcium phosphate) in a direct putting mode, and stirring by using a clean plastic rod to uniformly distribute the tricalcium phosphate in a soil sample;
2. spraying a bacterial solution of the prepared klebsiella aerogenes Wn in advance (the bacterial agent grows to OD in the culture medium 2)6001 stationary phase or logarithmic growth phase) and uniformly stirring to finally make the water content of the soil (bacterial liquid) be 30%;
3. medium 2 containing Wn (OD ═ 1) was added every 3 days for the first 15 days to maintain a water content of 30% in the soil, and medium 1 containing Wn (OD ═ 1) was added every 3 days for the last 15 days to maintain a water content of 30% in the soil. The content of the added phosphate radical and the contents of two heavy metal elements, namely Pb and Cd, in the soil keep a certain proportional relationship, specifically, the adding amount of tricalcium phosphate is related to the molar mass ratio of Pb (P: HMs-1: 1) to Cd (P: HMs-5: 1) in the soil, and finally the adding amount is determined to be 4 mg/g.
In addition, the present example also performed a sterile blank test under the same conditions.
The experimental results show that: compared with a blank control test, in 7 days, the treatment of the microbial Wn microbial inoculum composite tricalcium phosphate ensures that the highest removal rates of the effective state contents of Pb and Cd in the soil are 49% and 49.3% respectively; at 15 days, the highest removal rate of the content of the effective state is respectively reduced to 45.1 percent and 46.4 percent, which shows that the back dissolution can occur only by the phosphorus dissolving effect. At 30 days, the removal rates of the effective state contents of Pb and Cd reach 51% and 51.3% respectively again, which shows that the phenomenon of reverse dissolution can be inhibited by the matching use of the culture medium.
In the above technical solutions, the above are only preferred embodiments of the present invention, and the technical scope of the present invention is not limited thereby, and all the technical concepts of the present invention include the claims of the present invention, which are directly or indirectly applied to other related technical fields by using the equivalent structural changes made in the content of the description and the drawings of the present invention.
Sequence listing
<110> university of south-middle school
<120> Enterobacter aerogenes combined Klebsiella aerogenes microbial inoculum, composition and application
<160> 2
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1410
<212> DNA
<213> Enterobacter aerogenes
<400> 1
tgcaagtcga gcggtagcac agagagcttg ctctcgggtg acgagcggcg gacgggtgag 60
taatgtctgg gaaactgcct gatggagggg gataactact ggaaacggta gctaataccg 120
cataacgtcg caagaccaaa gtgggggacc ttcgggcctc atgccatcag atgtgcccag 180
atgggattag ctagtaggtg gggtaatggc tcacctaggc gacgatccct agctggtctg 240
agaggatgac cagccacact ggaactgaga cacggtccag actcctacgg gaggcagcag 300
tggggaatat tgcacaatgg gcgcaagcct gatgcagcca tgccgcgtgt atgaagaagg 360
ccttcgggtt gtaaagtact ttcagcgagg aggaaggcgt taaggttaat aaccttagcg 420
attgacgtta ctcgcagaag aagcaccggc taactccgtg ccagcagccg cggtaatacg 480
gagggtgcaa gcgttaatcg gaattactgg gcgtaaagcg cacgcaggcg gtctgtcaag 540
tcggatgtga aatccccggg ctcaacctgg gaactgcatt cgaaactggc aggctagagt 600
cttgtagagg ggggtagaat tccaggtgta gcggtgaaat gcgtagagat ctggaggaat 660
accggtggcg aaggcggccc cctggacaaa gactgacgct caggtgcgaa agcgtgggga 720
gcaaacagga ttagataccc tggtagtcca cgccgtaaac gatgtcgact tggaggttgt 780
gcccttgagg cgtggcttcc ggagctaacg cgttaagtcg accgcctggg gagtacggcc 840
gcaaggttaa aactcaaatg aattgacggg ggcccgcaca agcggtggag catgtggttt 900
aattcgatgc aacgcgaaga accttaccta ctcttgacat ccagagaact tagcagagat 960
gctttggtgc cttcgggaac tctgagacag gtgctgcatg gctgtcgtca gctcgtgttg 1020
tgaaatgttg ggttaagtcc cgcaacgagc gcaaccctta tcctttgttg ccagcgattc 1080
ggtcgggaac tcaaaggaga ctgccagtga taaactggag gaaggtgggg atgacgtcaa 1140
gtcatcatgg cccttacgag tagggctaca cacgtgctac aatggcatat acaaagagaa 1200
gcgacctcgc gagagcaagc ggacctcata aagtatgtcg tagtccggat tggagtctgc 1260
aactcgactc catgaagtcg gaatcgctag taatcgtaga tcagaatgct acggtgaata 1320
cgttcccggg ccttgtacac accgcccgtc acaccatggg agtgggttgc aaaagaagta 1380
ggtagcttaa ccttcgggag ggcgcttacc 1410
<210> 2
<211> 1403
<212> DNA
<213> Klebsiella aerogenes (Klebsiella aerogenes)
<400> 2
gccctcccga aggttaagct acctacttct tttgcaaccc actcccatgg tgtgacgggc 60
ggtgtgtaca aggcccggga acgtattcac cgtagcattc tgatctacga ttactagcga 120
ttccgacttc atggagtcga gttgcagact ccaatccgga ctacgacata ctttatgagg 180
tccgcttgct ctcgcgaggt cgcttctctt tgtatatgcc attgtagcac gtgtgtagcc 240
ctactcgtaa gggccatgat gacttgacgt catccccacc ttcctccagt ttatcactgg 300
cagtctcctt tgagttcccg accgaatcgc tggcaacaaa ggataagggt tgcgctcgtt 360
gcgggactta acccaacatt tcacaacacg agctgacgac agccatgcag cacctgtctc 420
agagttcccg aaggcaccaa agcatctctg ctaagttctc tggatgtcaa gagtaggtaa 480
ggttcttcgc gttgcatcga attaaaccac atgctccacc gcttgtgcgg gcccccgtca 540
attcatttga gttttaacct tgcggccgta ctccccaggc ggtcgactta acgcgttagc 600
tccggaagcc acgcctcaag ggcacaacct ccaagtcgac atcgtttacg gcgtggacta 660
ccagggtatc taatcctgtt tgctccccac gctttcgcac ctgagcgtca gtctttgtcc 720
agggggccgc cttcgccacc ggtattcctc cagatctcta cgcatttcac cgctacacct 780
ggaattctac ccccctctac aagactctag cctgccagtt tcgaatgcag ttcccaggtt 840
gagcccgggg atttcacatc cgacttgaca gaccgcctgc gtgcgcttta cgcccagtaa 900
ttccgattaa cgcttgcacc ctccgtatta ccgcggctgc tggcacggag ttagccggtg 960
cttcttctgc gagtaacgtc aatcgctaag gttattaacc ttaacgcctt cctcctcgct 1020
gaaagtactt tacaacccga aggccttctt catacacgcg gcatggctgc atcaggcttg 1080
cgcccattgt gcaatattcc ccactgctgc ctcccgtagg agtctggacc gtgtctcagt 1140
tccagtgtgg ctggtcatcc tctcagacca gctagggatc gtcgcctagg tgagccatta 1200
ccccacctac tagctaatcc catctgggca catctgatgg catgaggccc gaaggtcccc 1260
cactttggtc ttgcgacatt atgcggtatt agctaccgtt tccagtagtt atccccctcc 1320
atcaggcagt ttcccagaca ttactcaccc gtccgccgct cgtcacccga gagcaagctc 1380
tctgtgttac cgctcgactt gca 1403
Claims (9)
1. The bacterium agent of the combination of the enterobacter aerogenes and the klebsiella aerogenes is characterized in that the bacterium agent is used for treating lead-cadmium polluted soil and comprises the enterobacter aerogenes and the klebsiella aerogenes;
wherein the Enterobacter aerogenes is Enterobacter aerogenes (Enterobacter aerogenes) W6, the Latin article is classified and named as Enterobacter aerogenes, and is stored in China general microbiological culture Collection center (CGMCC), the storage date is 2021, 7 and 13 days, and the storage number is CGMCC No. 22888;
the Klebsiella aerogenes is Klebsiella aerogenes Wn, the Latin classification name is Klebsiella aerogenes, the Klebsiella aerogenes is stored in the common microorganism center (CGMCC) of the China Committee for culture Collection of microorganisms, the storage date is 2021, 7 and 13 days, and the storage number is CGMCC No. 22889.
2. The application of the microbial inoculum according to claim 1 in treating lead-cadmium polluted soil.
3. The composition for treating lead-cadmium polluted soil is characterized by comprising a component A, a component B, a component C and a component D;
wherein component a comprises a source of phosphorus;
the component B comprises the microbial agent of claim 1;
the component C comprises an aqueous solution containing glucose, magnesium chloride, magnesium sulfate, ammonium sulfate and potassium chloride;
the component D comprises an aqueous solution containing tryptone, yeast extract and sodium chloride.
4. The composition of claim 3, wherein the phosphorus source is tricalcium phosphate.
5. A method for treating lead-cadmium contaminated soil, which comprises treating the lead-cadmium contaminated soil with the composition according to claim 3 or 4.
6. The method for treating lead-cadmium contaminated soil according to claim 5, comprising: applying the component A and a first mixed liquor containing the component B and the component C to soil to be treated for a first period of time;
applying a second mixed liquor containing the component B and the component D to the soil to be remediated for a second time period;
wherein the second time period is later than the first time period.
7. The method for treating lead-cadmium contaminated soil according to claim 6, further comprising: and applying the first mixed liquor to the soil to be treated between the first time period and the second time period.
8. The method for treating lead-cadmium contaminated soil according to claim 7,
applying the first mixed liquor to the soil to be treated for multiple times between the first time period and the second time period, wherein the single application amount is consistent with the total application amount of the first mixed liquor in the first time period;
and applying the second mixed liquor to the soil to be treated for multiple times in the second time period, wherein the single application amount is consistent with the total application amount of the first mixed liquor in the first time period.
9. The method for treating lead-cadmium polluted soil according to any one of claims 6 to 8, wherein the interval between the first time period and the second time period is 14 to 16 days.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115232765A (en) * | 2022-06-16 | 2022-10-25 | 贵州民族大学 | Klebsiella WH-15 and application thereof |
| CN115232765B (en) * | 2022-06-16 | 2023-06-16 | 贵州民族大学 | Klebsiella WH-15 and application thereof |
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