CN111606533A - Plant-carried microbial inoculum for in-situ remediation of polluted bottom mud and preparation method thereof - Google Patents

Plant-carried microbial inoculum for in-situ remediation of polluted bottom mud and preparation method thereof Download PDF

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CN111606533A
CN111606533A CN202010381398.3A CN202010381398A CN111606533A CN 111606533 A CN111606533 A CN 111606533A CN 202010381398 A CN202010381398 A CN 202010381398A CN 111606533 A CN111606533 A CN 111606533A
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bacteria
plant
microbial inoculum
organic matter
microbial
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张宏耀
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China complete equipment import and Export Group Co.,Ltd.
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China Complete Equipment Import And Export Group Co ltd
China National Complete Plant Import & Export Corp ltd
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    • CCHEMISTRY; METALLURGY
    • C02TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02FTREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F11/00Treatment of sludge; Devices therefor
    • C02F11/02Biological treatment
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20Bacteria; Culture media therefor

Abstract

The invention discloses a plant-carried microbial agent for in-situ remediation of polluted bottom mud. The microbial inoculum comprises heterotrophic nitrifying bacteria, aerobic denitrifying bacteria, phosphorus accumulating bacteria and organic matter degrading bacteria, and is separated from root parts of the self-submerged plants. The plant-carried microbial inoculum can effectively improve polluted bottom sediment, eliminates ammonia nitrogen in the bottom sediment by more than 90 percent, provides favorable conditions for survival of submerged plants, improves the microbial remediation efficiency by 3-5 times by taking plant roots as adsorption carriers, and simultaneously enhances the water scouring resistance of the microbial inoculum. The microbial inoculum achieves the purpose of in-situ 'ecological dredging' on polluted bottom mud, avoids secondary pollution of ex-situ engineering dredging, and simultaneously realizes the long-term restoration of water ecology.

Description

Plant-carried microbial inoculum for in-situ remediation of polluted bottom mud and preparation method thereof
Technical Field
The invention relates to the technical field of sludge treatment, in particular to a plant-carried microbial agent for reducing polluted bottom sludge and a preparation method thereof.
Background
Formation of contaminated bottom sludge
The blackening and smelling of the water body sludge are mainly the result of series physical, chemical and biological effects of organic pollutants in the water body under the anoxic or anaerobic state. Organic pollutants entering the river are one of the main reasons for causing black and smelly sludge in the water body. The organic pollutants mainly comprise organic carbon source pollutants, organic nitrogen source pollutants and phosphorus-containing compounds, the pollutants are mainly from the decomposition of a large amount of organic matters settled in a water body, and a large amount of dissolved oxygen is consumed in the decomposition process, so that the bottom mud is anoxic, anaerobic microorganisms propagate in a large amount and decompose the organic matters to generate a large amount of black and odor causing substances. According to investigation, the thickness of the bottom mud polluted by the black and odorous water in rivers and lakes can reach 0.4-1 m at present, so that the upward flooding of the bottom mud is one of the main sources of water pollution.
Characteristics of polluting the bottom mud
The polluted substrate sludge is usually in a colloidal state, the permeability coefficient is very low, and due to the fact that the thickness of a biomembrane and an inorganic substance precipitate generated by metabolism of internal anaerobic microorganisms and the thickness of clay particle double-electric-layer layers is increased, effective pores in a soil body are reduced, connectivity of pores in a sample is reduced, and the permeability coefficient of a sludge sample is further reduced. Therefore, a single microbial remediation form stays on the surface layer of the bottom sediment for a long time, so that the microbial agent is overlong in the period of remedying the bottom sediment and is not thorough in remediation.
The current commonly used sludge treatment technology and defects
At present, the pollution bottom mud control technology mainly comprises an ex-situ treatment technology and an in-situ treatment technology. The ectopic treatment technology is to excavate polluted bottom mud out and transport the polluted bottom mud to other places for treatment, namely, to transfer an internal pollution source of a water body away so as to prevent the pollution of the water body. The ectopic treatment technology mainly comprises dredging, ectopic leaching, vitrification and the like, but a large amount of dredged sediment is necessarily generated in the most widely applied dredging process, and because the sediment has the problems of high water content, difficult transportation, undefined pollution components, no place accumulation and the like, if reasonable treatment cannot be obtained, a large amount of land resources are wasted, and pollutants in the sediment can flow into soil and the surrounding environment along with surface runoff to cause secondary pollution to the environment around an accumulation area. The in-situ treatment technology is to seal the polluted bottom mud in situ, and measures are taken to prevent the bottom mud pollutants from entering the water body, namely the pollution path of an internal pollution source is cut off. The in-situ treatment technology mainly comprises covering, curing, oxidizing, water diversion, physical leaching, air injection, electrokinetic remediation, microbial remediation, phytoremediation and the like. Wherein, the pollution of the chemical drugs to the water body and the bottom mud often exceeds the restoration effect of the chemical drugs; the physical method can cause the damage of an ecological system, can only improve for a while, and can not maintain long-term ecological balance, and the biological remediation is one of important means for water body and sediment remediation due to the advantages of small investment, good effect, no secondary pollution and the like.
The microbial remediation technology in China is late, but the rapid development starts from the last 90 th century, and the figure of the microbial agent can be seen in various environmental protection fields. But the microbial inoculum used in the current market for repairing polluted bottom mud has a plurality of defects: firstly, the permeability of the bottom sediment repairing microbial inoculum to polluted bottom sediment is extremely low, so that the repairing effect of the microbial inoculum to the bottom sediment can only stay on the surface; secondly, the pertinence of the bottom sediment remediation microbial inoculum to different pollutions is poor, and if the removal efficiency of high ammonia nitrogen concentration in the bottom sediment is low, the bottom sediment recovery is obviously seriously influenced; finally, the anti-scouring capability of the bottom sediment repairing microbial inoculum is poor, and the simple microbial inoculum can not be reserved and play a role for a long time under a flowing water area. The concept of microbial-plant combined remediation has been proposed in recent years and is mostly in the laboratory research phase. At present, no connection exists between the applied plants and microorganisms, only by the random collocation of the single microbial inoculum and the single plants, the effective interaction research of the combination of the single microbial inoculum and the single plants is lacked, so that the single microbial inoculum can only perform single functions under most conditions, the real combination is not carried out, and the repairing effect does not achieve the long-term expected target. Therefore, providing and developing a plant-carried microbial agent with pertinence, stability and high efficiency and an in-situ ecological restoration technology of river sludge are particularly important for treating the river polluted bottom sludge.
Disclosure of Invention
Based on the problems in the prior art, the invention provides the following technical scheme.
1. The plant-carried microbial agent is characterized by comprising heterotrophic nitrifying bacteria, aerobic denitrifying bacteria, phosphorus accumulating bacteria and organic matter degrading bacteria;
the heterotrophic nitrifying bacteria, the aerobic denitrifying bacteria, the phosphorus accumulating bacteria and the organic matter degrading bacteria are obtained by screening submerged plant roots;
wherein the heterotrophic nitrifier is Pseudomonas putida (Pseudomonas allopurina) H15 strain, and the preservation number is CGMCC No. 19258;
the aerobic denitrifying bacteria are alicyclic-philic bacteria (a)Alicycliphilus denitrificans) D60 strain with preservation number of CGMCC No. 19260;
the polyphosphate accumulating bacteria is a Pseudomonas otitidis (Pseudomonas otitidis) P23 strain, and the preservation number is CGMCC No. 19257;
the organic matter degrading bacteria is Pseudomonas kunmingensis (Pseudomonas kunmingensis) M60 strain, and the preservation number is CGMCC No. 19259.
2. The plant-carried microbial agent according to 1, wherein the weight ratio of each component strain is as follows: 30-45% of heterotrophic nitrifying bacteria, 30-35% of aerobic denitrifying bacteria, 10-20% of phosphorus-accumulating bacteria and 15-20% of organic matter degrading bacteria.
3. The plant-carried microbial agent according to 1, wherein the total viable bacteria number in the plant-carried microbial agent is not less than 1.0 × 109cfg/g。
4.1 to 3, the method for preparing the plant-carried microbial agent is characterized by comprising the following steps:
collecting submerged plant roots for culture and screening to obtain heterotrophic nitrifying bacteria, aerobic denitrifying bacteria, phosphorus accumulating bacteria and organic matter degrading bacteria;
respectively activating heterotrophic nitrifying bacteria, aerobic denitrifying bacteria, phosphorus accumulating bacteria and organic matter degrading bacteria, fermenting and culturing, and then mixing fermentation liquor in proportion to obtain mixed fermentation liquor;
preferably, the method also comprises the steps of adsorbing the mixed fermentation liquor on a substrate, and then drying and granulating at low temperature to obtain the product.
5. The preparation method according to 4, wherein the substrate is diatomite or zeolite powder.
6. The preparation method according to 4, wherein in the mixed fermentation liquor, the weight proportions of the component strains are respectively as follows: 30-45% of heterotrophic nitrifying bacteria, 30-35% of aerobic denitrifying bacteria, 10-20% of phosphorus-accumulating bacteria and 15-20% of organic matter degrading bacteria.
7. Use of the plant-borne microbial agent of any one of claims 1-3 for the treatment of contaminated substrate sludge.
8. The method for treating the polluted bottom mud comprises the following steps:
1) adding the plant-carried microbial agent as described in any one of 1-3 into the polluted bottom sludge;
2) and (5) planting submerged plants.
9. The treatment method according to 8, wherein the submerged plant is tape grass, watermifoil and/or hornworts.
10. The treatment method according to 8, wherein the submerged plant is planted after the plant-carried microbial agent is added for 3-10 days.
[ biological Material Collection ]
The heterotrophic nitrifier strains of the invention are deposited in:
china general microbiological culture Collection center (CGMCC), the preservation date is 1, 2 and 2020, and the preservation number is CGMCC No. 19258; the address of the depository is: xilu No.1, Beijing, Chaoyang, Beijing, and institute for microbiology, China academy of sciences.
The aerobic denitrifying strains of the present invention are deposited in:
china general microbiological culture Collection center (CGMCC), the preservation date is 1, 2 and 2020, and the preservation number is CGMCC No. 19260; the address of the depository is: xilu No.1, Beijing, Chaoyang, Beijing, and institute for microbiology, China academy of sciences.
The phosphorus accumulating bacteria strain of the invention is preserved in:
china general microbiological culture Collection center (CGMCC), the preservation date is 1 month and 2 days in 2020, and the preservation number is CGMCC No. 19257; the address of the depository is: xilu No.1, Beijing, Chaoyang, Beijing, and institute for microbiology, China academy of sciences.
The organic matter degrading bacterial strain of the invention is preserved in:
china general microbiological culture Collection center (CGMCC), the preservation date is 1, 2 and 2020, and the preservation number is CGMCC No. 19259; the address of the depository is: xilu No.1, Beijing, Chaoyang, Beijing, and institute for microbiology, China academy of sciences.
Technical effects of the invention
The plant-carried microbial agent provided by the invention is formed by mixing multiple microorganisms, has high viable count, and has higher removal rate on total nitrogen, ammonia nitrogen, phosphorus and COD (chemical oxygen demand) than that of a single strain. The used strain has strong environmental adaptability and no secondary pollution to the environment, and is an environment-friendly microbial inoculum; the end products of the microbial reproductive metabolism are water, nitrogen and carbon dioxide, thereby fundamentally reducing ammonia nitrogen in sludge and sewage, ensuring the survival of plants, and being beneficial to the survival and growth of plants due to the transformation of phosphorus element and organic matters. Meanwhile, the root of the plant can be used as a carrier of microorganisms in the microbial inoculum due to the synergistic effect of the microbial inoculum and submerged plants, and the polluted bottom mud is deeply repaired along with the process that the roots of the plants are deeply rooted into the bottom mud, so that the problems of low permeability and poor anti-scouring capability of the conventional bottom mud microbial inoculum on the bottom mud are solved.
The plant-carried microbial agent can obviously improve the sludge, wherein the reduction rate of ammonia nitrogen can reach more than 90 percent, and the plant-carried microbial agent does not contain chemical substances, has no secondary pollution to the environment and is an environment-friendly microbial agent. Meanwhile, the compound microbial agent has simple preparation process and low production cost, and has wide prospects in the aspects of eutrophication black and odorous water body pollution sediment and water body remediation.
The invention has two main points of innovation:
firstly, separating and screening black and odorous water polluted substrate sludge repairing strains from submerged plant roots, and preferably obtaining plant-carried microbial inoculum to solve the problems of low repairing efficiency, poor permeability and easy scouring of the existing microbial inoculum;
and secondly, screening to obtain high-ammonia nitrogen-resistant heterotrophic nitrifiers and aerobic denitrifying bactericides, adding the heterotrophic nitrifiers and aerobic denitrifying bactericides into the bottom sludge remediation bactericides, pertinently solving the problem that the existing bactericides have low removal rate of high ammonia nitrogen in the bottom sludge, and providing favorable conditions for recovering an ecological system.
Drawings
FIG. 1 shows the quantitative results of heterotrophic nitrifying strains attached to the roots of submerged plants.
Fig. 2 shows the change in water body of the control group and the experimental group after the anti-scour simulation experiment was performed. Wherein A is the water state of the control group, and B is the water state of the experimental group.
FIG. 3 shows the remediation of ammonia nitrogen concentration in river polluted bottom mud by the synergistic remediation of the plant-carried microbial inoculum and plants of the invention.
Fig. 4 shows the surface layer change before and after the remediation of the river polluted bottom mud, wherein a is the surface layer condition of the river before the remediation and B is the surface layer condition of the river after the remediation.
Detailed Description
In order that the objects, technical solutions and advantages of the present invention will become more apparent, the present invention will be further described in detail with reference to the accompanying drawings in conjunction with the following specific embodiments.
The methods used in the following examples are conventional methods unless otherwise specified, and the reagents used are commercially available reagents unless otherwise specified.
Example 1 Strain screening
In the plant-carried microbial agent of the present invention, heterotrophic nitrifying bacteria for degrading contaminated substrate sludge are obtained by:
collecting complete roots of the river and lake submerged plants in a sterile collection bag, soaking the roots in normal saline, vortexing the roots at the maximum rotation speed for 2min, then oscillating the roots at 220rpm for 2h, and carrying out enrichment culture in an enrichment culture medium A with ammonium salt as a unique nitrogen source for 30 days. After gradient dilution, appropriate dilution concentrations are selected to coat solid culture media such as LB, 1/10LB and R2A, colonies with single morphology are selected and further purified and cultured. The obtained bacterial strains are subjected to functional screening, and the bacterial strain with the strongest nitrification capability is selected to be the plant-carried heterotrophic nitrifying bacteria, and the preservation number of the plant-carried heterotrophic nitrifying bacteria is CGMCC No. 19258.
Wherein the enrichment medium A is (1L): 0.5g of ammonium sulfate, 5g of sodium succinate, 2g of sodium acetate, 0.3g of sodium chloride and 50ml of trace elements. Wherein each 1L of the trace elements comprises: 5g of dipotassium phosphate, 2.5g of magnesium sulfate, 2.5g of sodium chloride, 0.05g of ferrous sulfate and 0.05g of manganese sulfate.
In the plant-carried microbial agent of the present invention, the aerobic denitrifying bacteria for degrading the contaminated substrate sludge are obtained by:
collecting complete roots of river and lake submerged plants in an aseptic collection bag, soaking the roots in normal saline, vortexing at the maximum rotation speed for 2min, oscillating the roots at 220rpm for 2h, carrying out enrichment culture in an enrichment culture medium B with nitrate as a unique nitrogen source for 30 days, then carrying out gradient dilution, selecting appropriate dilution concentration to coat solid culture media such as LB, 1/10LB and R2A, selecting bacterial colonies with single form, further carrying out purification culture, and carrying out functional verification to select a bacterial strain with the strongest denitrification capability, namely the plant-carried aerobic denitrifying bacteria with the preservation number of CGMCC No. 19260.
Wherein the enrichment medium B is (1L): potassium nitrate 0.7g, sodium succinate 5g, sodium acetate 2g, trace elements 50 ml. Wherein each 1L of the trace elements comprises: 5g of dipotassium phosphate, 2.5g of magnesium sulfate, 2.5g of sodium chloride, 0.05g of ferrous sulfate and 0.05g of manganese sulfate.
In the plant-carried microbial agent, the phosphorus-accumulating bacteria for degrading the polluted substrate sludge are obtained by the following steps:
collecting complete roots of river and lake submerged plants in an aseptic collection bag, soaking the roots in normal saline, vortexing at the maximum rotation speed for 2min, oscillating the roots at 220rpm for 2h, carrying out enrichment culture in a high-phosphate enrichment culture medium C for 30 days, carrying out gradient dilution, selecting appropriate dilution concentration, coating LB, 1/10LB, R2A and other solid culture media, selecting colonies with single form, further carrying out purification culture, carrying out functional verification, and selecting a strain with the strongest phosphorus accumulation capacity, namely the plant-carried phosphorus accumulation bacteria with the preservation number of CGMCC No. 19257.
Wherein the enrichment medium C is (1L): 0.06g of monopotassium phosphate, 2g of ammonium sulfate, 5g of sodium acetate, 0.2g of calcium chloride and 50ml of trace elements. Wherein each 1L of the trace elements comprises: 5g of dipotassium phosphate, 2.5g of magnesium sulfate, 2.5g of sodium chloride, 0.05g of ferrous sulfate and 0.05g of manganese sulfate.
In the plant-carried microbial agent, the organic matter degrading bacteria for degrading the polluted bottom mud are obtained by the following steps:
collecting complete roots of river and lake submerged plants in a sterile collection bag, soaking the roots in normal saline, vortexing at the maximum rotation speed for 2min, oscillating the roots at 220rpm for 2h, carrying out enrichment culture in an enrichment culture medium D with high COD for 30 days, then carrying out gradient dilution, selecting appropriate dilution concentration to coat solid culture media such as LB, 1/10LB and R2A, selecting colonies with single form, further carrying out purification culture, carrying out functional verification, and selecting a strain with the strongest COD removal capacity as the plant carried organic matter degrading bacteria with the preservation number of CGMCC No. 19259.
Wherein the enrichment medium D is (1L): 2g of sodium succinate, 1g of glucose, 5g of sodium acetate, 2g of ammonium sulfate, 0.5g of sodium nitrate and 50ml of trace elements. Wherein each 1L of the trace elements comprises: 5g of dipotassium phosphate, 2.5g of magnesium sulfate, 2.5g of sodium chloride, 0.05g of ferrous sulfate and 0.05g of manganese sulfate.
Example 2 preparation of microbial Agents
The preparation method of the compound microbial agent comprises the following steps:
firstly, screening a submerged plant root system by adopting the method of embodiment 1 to obtain heterotrophic nitrifying bacteria with the preservation number of CGMCC No.19258, aerobic denitrifying bacteria with the preservation number of CGMCC No.19260, phosphorus accumulating bacteria with the preservation number of CGMCC No.19257 and degraded organic matter bacteria with the preservation number of CGMCC No. 19259; the following operations are then carried out:
1) preparation of fermentation seeds: heterotrophic nitrifying bacteria with the preservation number of CGMCC No.19258, aerobic denitrifying bacteria with the preservation number of CGMCC No.19260, phosphorus accumulating bacteria with the preservation number of CGMCC No.19257 and organic matter degrading bacteria with the preservation number of CGMCC No.19259 are respectively inoculated on a seed culture medium (LB culture medium) according to the inoculation amount of 2-3% for activation, the culture temperature is 30 ℃, the rotation speed is 150rpm, the culture is carried out for 18-24 hours, and the OD600 reaches 0.8-1.
2) Fermentation culture of functional microbial inoculum: inoculating each strain into a fermentation culture medium according to the inoculum size of 5%, wherein the fermentation conditions are that the temperature is 30-35 ℃, the pH is about 6.5-7.5, the rotation speed is 150-300rpm, the dissolved oxygen is more than 20%, the tank pressure is 0.02-0.05MPa, the fermentation culture is 10-20 h (preferably 12-20 h), and the OD of the fermentation liquid is600And (3) mixing the strains according to a proportion to obtain a mixed fermentation liquor according to the actual pollution condition of the bottom mud, wherein the proportion is more than 10.
Wherein, the fermentation medium of each strain is shown in table 1:
TABLE 1
3) Directly adsorbing the mixed fermentation liquor on a substrate, preferably diatomite and zeolite powder, and then drying and granulating at low temperature.
Example 3: efficient treatment of high ammonia nitrogen polluted bottom mud by composite microbial inoculum
The compound microbial inoculum used in the embodiment comprises the following strains in mass fraction based on the weight of the total strains contained in the microbial inoculum:
wherein the total viable bacteria number in the microbial inoculum is 5.0 × 109cfg/g。
Placing typical high ammonia nitrogen river pollution bottom mud (ammonia nitrogen concentration is 625mg/kg) into a glass cylinder with length, width and height of 60 x 50 x 40cm, adding black mud with thickness of 15cm, adding black smelly sewage with depth of 20cm, and mixing according to the ratio of 50mg/cm2Adding a compound microbial inoculum, and adding a micro aeration device to ensure that the dissolved oxygen is more than 3 mg/L. Meanwhile, a control test without adding the microbial inoculum is set, and other conditions are completely the same.
After 7 days of fungus feeding, the thickness of the sediment of the experimental group is reduced to a certain extent, wherein the yellowing thickness of the surface layer of the polluted sediment is obviously increased, the ammonia nitrogen is reduced to 42mg/kg, the removal rate reaches 93.3 percent, and the contrast group does not have obvious change in the same period. After 30 days of fungus feeding, the thickness of black mud in an experimental group is obviously reduced, the ammonia nitrogen is further reduced to 11mg/kg, the removal rate is up to 98%, and the experimental group and a control group have obvious difference.
The result proves that the compound microbial agent has high tolerance to ammonia nitrogen, has effective improvement capability on seriously polluted bottom mud, and can effectively reduce the thickness of the polluted bottom mud and remove high-load nutrient substances in the polluted bottom mud. Indexes of ammonia nitrogen and the like of the sediment of the experimental group and the contrast group are shown in the following table:
TABLE 2 improvement of contaminated bottom sediment by 7 and 30 days of inoculum addition in the experimental groups
TABLE 3 sediment indices of control group versus experimental group at the same time period
Example 4: synergistic remediation of polluted bottom sediment by using composite microbial inoculum and adsorption carrier (plant)
The compound microbial inoculum used in the embodiment comprises the following strains in mass fraction based on the weight of the total strains contained in the microbial inoculum:
wherein the total viable bacteria number in the microbial inoculum is 1.0 × 109cfg/g。
Placing typical river pollution bottom mud with high ammonia nitrogen into a glass cylinder with the length, width and height of 60 x 50 x 40cm, adding black mud with the thickness of 15cm, adding black and odorous sewage with the depth of 20cm, and mixing according to the ratio of 50mg/cm2Adding a microbial inoculum, and adding a micro aeration device to ensure that the dissolved oxygen is more than 3 mg/L. And (3) planting 20 tape grass 7 days after the compound microbial inoculum is added, and setting a control test without adding the microbial inoculum.
After 7 days of fungus feeding, the thickness of the experimental sediment is reduced to a certain extent, wherein the thickness of the polluted sediment which turns yellow on the surface layer is obviously increased, and the thickness of the control group is unchanged. After 5 days of tape grass planting, total DNA of bacteria loaded on the root system is extracted, and quantitative determination is carried out on heterotrophic nitrifying bacteria of the microbial inoculum by utilizing qPCR (see figure 1), and a large amount of heterotrophic nitrifying bacteria are found to be adsorbed on the root system of the experimental group plant. After 30 days, the thickness of the black mud is obviously reduced, and the experimental group and the control group have obvious difference.
The results prove that the composite microbial inoculum has the effects of efficiently reducing the polluted bottom mud and improving the black and odorous water body, and can effectively improve the survival rate of submerged plants (by 60%).
According to the experiment of the embodiment 3, the compound microbial agent disclosed by the invention has effective improvement capacity on the polluted bottom mud, but the result of multiple experiments shows that when the thickness of the polluted bottom mud is reduced by about 1.5cm, the polluted bottom mud is difficult to further reduce, the ammonia nitrogen of the surface bottom mud is obviously reduced due to the function of the microbial agent, but the ammonia nitrogen of the deeper bottom mud is not effectively removed, which is supposed to be mainly because microorganisms cannot enter the deep black mud to play a role.
According to the invention, through planting and rooting of submerged vegetation, the effect of reducing the thickness of the sediment is 1 time higher than that of the treatment effect of a simple microbial agent. And the control group experiment proves that the effect is not from the repair of plants.
Tracking and finding heterotrophic nitrifying bacteria: the microbial agent enters the bottom sediment along with the root system, and 10 percent of eluent is contained in per milliliter of eluent4The copy number is of order of magnitude, which indicates that the roots of the plants can be used as microorganism adsorption carriers to enter the deep part of the bottom sediment, thereby further efficiently reducing and repairing the polluted bottom sediment. The indexes of the sediment of the experimental group and the control group are shown in the following table:
TABLE 4
After the experimental group plants the tape grass for 23 days (namely 30 days after the fungus feeding), the survival rate of the plants is 100 percent and the plants grow vigorously, wherein the index change of the bottom mud of the experimental group is shown in the following table.
TABLE 5
The survival rate of the plants is 40% and the growth condition is poor after the control group is planted with the tape grass for 23 days (namely, 30 days after the fungus feeding). The bottom mud index change of the control group is shown in the following table.
TABLE 6
Example 5: the complex microbial inoculum has high-efficiency anti-scouring capability
The compound microbial inoculum used in the embodiment comprises the following strains in mass fraction based on the weight of the total strains contained in the microbial inoculum:
wherein the total viable bacteria number in the microbial inoculum is 3.0 × 109cfg/g。
Placing typical river pollution bottom mud into a glass cylinder with length, width and height of 60 x 50 x 40cm, adding black mud with thickness of 15cm, adding black smelly sewage with depth of 20cm, and mixing according to 50mg/cm2Adding a microbial inoculum, and adding a micro aeration device to ensure that the dissolved oxygen is more than 3 mg/L. And (4) planting 20 tape grass on the 7 th day after adding, and setting a control test without adding the microbial inoculum.
After 7 days of fungus feeding, the thickness of the bottom mud is reduced to a certain extent, wherein the thickness of the polluted bottom mud surface layer which turns yellow is obviously increased. After 10 days of planting the tape grass, introducing sewage with the flow of 0.2L/S into one end of the surface of the water body for 10min by using a water pipe with the diameter of 3cm, and performing a scouring simulation experiment on the water body and the sediment. After 3 days, the water in the experimental group is clarified and basically recovered to the original state; the control group had turbid water and the leaves of the aquatic weeds were stained with soil (see FIG. 2). After the tape grass is planted for 23 days, the thickness of the black mud is obviously reduced, and the experimental group and the control group have obvious difference.
The result proves that the plant-carried microorganism composite microbial inoculum has high-efficiency anti-scouring capability, and the microbial inoculum microorganism takes the root system and the leaves of the plant as a carrier, so that the fixing capability of the microbial inoculum is obviously improved to resist the scouring of the external water flow, and the function of improving the polluted bottom mud and the water quality is effectively exerted for a long time.
The water quality indexes before and after the washing of the experimental group and the control group are shown in the following table:
the water quality change of the experimental group is shown in the table 7.
TABLE 7
The water quality changes of the control group at the same time are shown in Table 8.
TABLE 8
Example 6: in-situ remediation experiment of plant-carried compound microbial agent
The plant-carried microbial inoculum used in the present example comprises the following strains in mass fraction, based on the total weight of strains contained in the inoculum:
wherein the total viable bacteria number in the microbial inoculum is 5.0 × 109cfg/g。
The test area used in the embodiment is a typical black and odorous water river channel, the width is 5 meters, the depth is 1.5 meters on average, the bottom mud is seriously polluted and is in a completely black and odorous colloid state, the ammonia nitrogen content is more than 400mg/kg, and the whole river channel is not grown.
A 10-meter-long river channel is enclosed by a purse net, then plant-carried compound microbial agents are uniformly scattered, carbon sources and aeration are timely added, and intermittent aeration is adopted during operation to maintain the dissolved oxygen at more than 3 mg/L.
Submerged plants (e.g., hornworts and watermifoil) were grown several times on day 7 of the inoculation, at a water temperature of 25 ℃.
After a week of microbial inoculum remediation, the ammonia nitrogen is reduced to about 100mg/kg, and submerged plants are planted at the moment, so that the survival of the plants is basically ensured. Then, after a month, the plant roots are used as carriers, and the microbial inoculum enters the deep part of the bottom sediment to play a role, so that the bottom sediment is further improved (see figure 4). The ammonia nitrogen index and the surface change of the sediment are respectively shown in figure 3 and figure 4.
The invention proves that the plant-carried microbial inoculum can effectively remove high ammonia nitrogen concentration polluting bottom mud, can carry out deep repair on the sludge and resist the scouring of rivers under the synergistic effect of the plant-carried microbial inoculum and submerged plants, and has practical application value.
The invention breaks the limitation of the in-situ ecological restoration of the existing microbial agent, achieves the aim of high-efficiency and long-term ecological restoration, and provides an important 'ecological dredging' idea.
The above-mentioned embodiments are intended to illustrate the objects, technical solutions and advantages of the present invention in further detail, and it should be understood that the above-mentioned embodiments are only exemplary embodiments of the present invention and are not intended to limit the present invention, and any modifications, equivalents, improvements and the like made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims (10)

1. The plant-carried microbial agent is characterized by comprising heterotrophic nitrifying bacteria, aerobic denitrifying bacteria, phosphorus accumulating bacteria and organic matter degrading bacteria;
the heterotrophic nitrifying bacteria, the aerobic denitrifying bacteria, the phosphorus accumulating bacteria and the organic matter degrading bacteria are obtained by screening submerged plant roots;
wherein the heterotrophic nitrifier is Pseudomonas putida (Pseudomonas allopurina) H15 strain, and the preservation number is CGMCC No. 19258;
the aerobic denitrifying bacteria are alicyclic denitrifying bacteria (Alicyclobacillus Denitrificas) D60 strains, and the preservation number is CGMCC No. 19260;
the polyphosphate accumulating bacteria is a Pseudomonas otitidis (Pseudomonas otitidis) P23 strain, and the preservation number is CGMCC No. 19257;
the organic matter degrading bacteria is Pseudomonas kunmingensis (Pseudomonas kunmingensis) M60 strain, and the preservation number is CGMCC No. 19259.
2. The plant-carried microbial agent according to claim 1, wherein the weight ratio of each component strain is respectively as follows: 30-45% of heterotrophic nitrifying bacteria, 30-35% of aerobic denitrifying bacteria, 10-20% of phosphorus-accumulating bacteria and 15-20% of organic matter degrading bacteria.
3. The plant-borne microbial inoculant according to claim 1, wherein the total viable bacteria count in the plant-borne microbial inoculant is not less than 1.0 × 109cfg/g。
4. A method of producing a plant-supported microbial inoculant according to any one of claims 1 to 3, comprising the steps of:
collecting submerged plant roots for culture and screening to obtain heterotrophic nitrifying bacteria, aerobic denitrifying bacteria, phosphorus accumulating bacteria and organic matter degrading bacteria;
respectively activating heterotrophic nitrifying bacteria, aerobic denitrifying bacteria, phosphorus accumulating bacteria and organic matter degrading bacteria, fermenting and culturing, and then mixing fermentation liquor in proportion to obtain mixed fermentation liquor;
adsorbing the mixed fermentation liquor on a substrate, and then drying and granulating at low temperature to obtain the product.
5. The method according to claim 4, wherein the substrate is diatomaceous earth or zeolite powder.
6. The preparation method according to claim 4, wherein the weight ratio of each component strain in the mixed fermentation broth is as follows: 30-45% of heterotrophic nitrifying bacteria, 30-35% of aerobic denitrifying bacteria, 10-20% of phosphorus-accumulating bacteria and 15-20% of organic matter degrading bacteria.
7. Use of the plant-borne microbial agent of any one of claims 1-3 for the treatment of contaminated substrate sludge.
8. The method for treating the polluted bottom mud comprises the following steps:
1) adding a plant-borne microbial agent as defined in any one of claims 1 to 3 to the contaminated substrate sludge;
2) and (5) planting submerged plants.
9. The treatment method according to claim 8, wherein the submerged plant is tape grass, watermifoil and/or hornworts.
10. The treatment method according to claim 8, wherein the submerged plant is planted 3 to 10 days after the addition of the plant-borne microbial agent.
CN202010381398.3A 2020-05-08 2020-05-08 Plant-carried microbial inoculum for in-situ remediation of polluted bottom mud and preparation method thereof Pending CN111606533A (en)

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