Arbuscular mycorrhizal fungus symbiotic plant remediation method

Abstract

The invention discloses an arbuscular mycorrhizal fungus symbiotic plant remediation method. The method comprises the following steps: (1) removing heavy metal-antibiotic polluted sediment dry sample impurities; (2) transplanting an arbuscular mycorrhizal fungus-aquatic plant symbiotic system, and constructing a heavy metal-antibiotic combined pollution sediment remediation system; (3) repairing and harvesting. Aiming at the characteristics of limitation, long consumed time and the like of the existing single aquatic plant remediation, the polluted bottom mud is subjected to in-situ remediation by utilizing the arbuscular mycorrhizal fungi-aquatic plant symbiotic system. The method can be widely applied to heavy metal, antibiotic and heavy metal-antibiotic combined pollution tail water sediment treatment, and has a remarkable remediation effect and sustainable ecological benefits. According to the method, an arbuscular mycorrhiza-aquatic plant symbiotic system which is successfully pre-cultured is transplanted, so that the purpose of efficiently repairing the polluted bottom mud is achieved.

Description

Technical Field

[0001] The present invention belongs to the field of phytoremediation methods, and particularly relates to a method for phytoremediation by symbiotic plants with arbuscular mycorrhizal fungi. Background Art

[0002] The combined pollution of antibiotics and heavy metals widely exists in water systems and has received extensive attention. Both of these pollutants can have serious harmful effects on humans and ecosystems. Aquaculture, livestock and poultry farming, the land application of animal manure, and the centralized treatment of industrial wastewater have led to their combined pollution of surface or groundwater. Controlling and managing the combined pollution of antibiotics and heavy metals is a huge challenge. On the one hand, these two types of pollutants have different environmental behaviors in water; on the other hand, certain combinations of antibiotics and heavy metals are prone to form complexes, while others are not, which makes the situation of removing them more complex.

[0003] Phytoremediation is a remediation strategy based on the ability of plants to extract, promote rhizodegradation, or stabilize pollutants in the medium. For a long time, phytoremediation has been considered an environmentally friendly, in-situ treatment, low-cost, and secondary pollution controllable technical means. However, due to low biomass production, long time consumption, and the stress caused by the combined toxicity of the polluted medium on the plant body, these effects may weaken the remediation potential of the plant.

[0004] Arbuscular mycorrhizal fungi (AMF) are a common sediment microbial community that can symbiose with the roots of about 90% of the classified higher plants. Although the lifestyle of arbuscular mycorrhizal fungi is aerobic, in the observation of the roots of various aquatic and wetland plant species around the world, arbuscular mycorrhizal fungi are the most common type of mycorrhizal fungi, and arbuscular mycorrhizal fungi are considered to be equally important for wetland plants and terrestrial plants. Arbuscular mycorrhizal fungi have been widely present in wetland habitats such as mangroves, swamps, constructed wetlands, and natural wetlands in the past few decades. The addition of arbuscular mycorrhizal fungi can promote plant nutrient absorption, improve the habitat for sustainable remediation, and regulate plant resistance and tolerance mechanisms. These characteristics also address the limitations of single-plant phytoremediation. Using the arbuscular mycorrhizal fungi-plant symbiotic system is an effective way to improve plant growth and development and enhance the remediation efficiency of the plant for the polluted medium. Summary of the Invention

[0005] Object of the Invention: To overcome the deficiencies of the prior art, the object of the present invention is to provide a method for phytoremediation by symbiotic plants with arbuscular mycorrhizal fungi, which repairs the heavy metal-antibiotic contaminated sediment, so as to achieve the purpose of restoring the natural ecological environment of local water bodies and sediment.

[0006] Technical solution: The arbuscular mycorrhizal fungal symbiotic plant restoration method comprises the following steps:

[0007] (1) Remove impurities from dry samples of heavy metal-antibiotic contaminated sludge;

[0008] (2) Transplanting arbuscular mycorrhizal fungi-aquatic plant symbiotic system to construct a sediment remediation system for heavy metal-antibiotic combined pollution;

[0009] (3) Repair.

[0010] Furthermore, the method for constructing the heavy metal-antibiotic composite contaminated sediment remediation system is as follows:

[0011] (1) Remove impurities from the surface of the sediment;

[0012] (2) 2 kg of heavy metal-antibiotic contaminated sludge was placed in a small plastic bucket, and the arbuscular mycorrhizal fungi-aquatic plant symbiotic system was transplanted into the bucket;

[0013] (3) After the symbiotic system of arbuscular mycorrhizal fungi and aquatic plants was cultivated for 4 weeks, the small bucket was moved into a glass tank and immersed in water to construct a sediment remediation system for heavy metal-antibiotic combined pollution;

[0014] (4) After 20 weeks of restoration, the intact plants were harvested and the bottom mud samples were collected.

[0015] Furthermore, in the above method, the preparation method of the heavy metal-antibiotic complex polluted sludge comprises the following steps:

[0016] (1) Collect surface sediment (10-20 cm) without pollution history and air dry it for 6 weeks. After air drying, pass it through a 10-mesh sieve (2 mm) for later use, and then sterilize it in a high-pressure steam sterilizer at 121°C for 2 hours to kill indigenous fungi;

[0017] (2) Add the required amount of heavy metal salt solution and antibiotic liquid standard to the sterilized sludge. Use a spray bottle to evenly spray the heavy metal salt solution and antibiotic standard on the sludge several times, stir them thoroughly manually, and leave them in the dark for 2 weeks to balance and stabilize the heavy metals and antibiotics. Prepare heavy metal-antibiotic composite contaminated sludge;

[0018] (3) Before the test, the prepared sludge was sterilized again in a high-pressure steam sterilizer at 121°C for 2 h.

[0019] Furthermore, the arbuscular mycorrhizal fungi-aquatic plant symbiotic system comprises a host aquatic plant plant with an arbuscular mycorrhizal fungi infection rate greater than 70% after pre-culture, extra-root hyphae extending beyond the root system of the host aquatic plant, and a small amount of pre-cultured soil with residual roots.

[0020] Furthermore, the aquatic plant is one or more of Iris pseudacorus, Canna indica, Phragmites australis, Juncus effusus, Cyperus rotundus or Glyceria maxima.

[0021] Furthermore, the arbuscular mycorrhizal fungi are one or more of arbuscular mycorrhizal fungi in the Glomeromycota.

[0022] The arbuscular mycorrhizal fungi-aquatic plant symbiotic system includes host aquatic plants infected by arbuscular mycorrhizal fungi, extraradical hyphae extending outside the roots of the host plants, and a small amount of pre-cultured soil remaining in the roots.

[0023] Compared with the prior art, the present invention has the following beneficial effects:

[0024] (1) The present invention uses potted plants to construct an arbuscular mycorrhizal fungi-aquatic plant symbiotic system and transplants it into heavy metal-antibiotic composite contaminated sediment, which can significantly reduce the concentrations of heavy metals and antibiotics in the sediment. The arbuscular mycorrhizal fungi-aquatic plant symbiotic system can extend a wide hyphal network in the sediment. While expanding the contact area between the root system and the sediment, this hyphal network can accumulate heavy metals that are dozens of times higher than the root systems of the host plants. In addition, the increased exudation of root exudates mediated by arbuscular mycorrhizal fungi stimulates the growth and reproduction of microbial groups, and is beneficial to the activation and binding of organic pollutants to the roots, enhancing the bioavailability and biodegradation of organic pollutants. The above-mentioned effects have a significant promoting effect on the plant absorption and degradation of heavy metal-antibiotic composite pollution in the sediment, and can effectively reduce the concentrations of heavy metals and antibiotics in the sediment. And it has the characteristics of high efficiency, ecological safety, green sustainability and simple operation, providing an effective method for repairing heavy metal-antibiotic composite pollution in sediment.

[0025] (2) The arbuscular mycorrhizal fungi-aquatic plant symbiotic system phytoremediation technology significantly improves the removal rate of sediment heavy metal-antibiotic pollution by phytoremediation, effectively overcoming the limitations of single-plant phytoremediation such as long time consumption, low efficiency and being stressed by pollutant toxicity. Description of the Drawings

[0026] Figure 1 It is a schematic diagram of a potted plant immersed in an arbuscular mycorrhizal fungi symbiotic system;

[0027] Figure 2 It is the residual amounts of Cu and Zn in the sediment under different treatment conditions;

[0028] Figure 3 It is the removal rates of Cu and Zn in the sediment under different treatment conditions;

[0029] Figure 4 It is the residual amounts of TC and OTC in the sediment under different treatment conditions;

[0030] Figure 5 is the removal rate of TC and OTC in sediment under different treatment conditions. DETAILED DESCRIPTION

[0031] In order to make the purpose, technical solution and advantages of the present invention clearer, the technical solution of the present invention will be further described below.

[0032] The arbuscular mycorrhizal fungi symbiotic system plant restoration technology of this embodiment includes the following steps:

[0033] (1) Collect surface sediment (10-20 cm) without pollution history and air dry it for 6 weeks. After air drying, pass it through a 10-mesh sieve (2 mm) for later use, and then sterilize it in a high-pressure steam sterilizer at 121°C for 2 hours to kill indigenous fungi;

[0034] (2) Add the required amount of heavy metal salt solution and antibiotic liquid standard to the sterilized sludge. Use a spray bottle to evenly spray the heavy metal salt solution and antibiotic standard on the sludge several times, stir them manually, and leave them in the dark for 2 weeks to balance and stabilize the heavy metals and antibiotics. Before the test, the prepared heavy metal-antibiotic composite contaminated sludge was sterilized again in a high-pressure steam sterilizer at 121°C for 2 hours;

[0035] (3) Place the prepared heavy metal-antibiotic complex contaminated sludge in a small plastic bucket at a rate of 2 kg per bucket;

[0036] (4) transplanting the pre-cultured arbuscular mycorrhizal fungi-aquatic plant symbiotic system (arbuscular mycorrhizal fungi infection rate greater than 70% of the host aquatic plant, extra-root hyphae extending beyond the host aquatic plant root system, and a small amount of pre-cultured soil remaining at the root) into a small bucket;

[0037] (5) After the transplanted plants were cultured for 4 weeks, the small bucket was moved into a glass tank and immersed in water. Water was added to the glass tank manually to keep 3 cm of shallow water covering the bottom mud, thus constructing a heavy metal-antibiotic composite contaminated mud remediation system;

[0038] (6) The waterlogged pots were restored for 20 weeks, the intact plants were harvested, and bottom mud samples were collected.

[0039] The exemplary embodiments of the present invention are described in more detail below in conjunction with the accompanying drawings. In order to make the purpose, technical solutions and advantages of the present invention clearer and more understandable, the present invention is further described in detail below in conjunction with embodiments. The embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0040] Example 1: Experiment on the remediation effect of the root cyst system of Rhizospora endothelii-Iris calamus on heavy metal-antibiotic contaminated sediment

[0041] 1. Using the method of the present invention

[0042] In this example, laboratory-simulated composite polluted sediment was used. The sediment was collected from a lotus pond in Xuzhou City, Jiangsu Province. The background values of the physical and chemical properties of the sediment are shown in Table 1:

[0043]

[0044] The experiment conducted in this example includes the following steps:

[0045] 1. Remove the impurities on the surface of the sediment;

[0046] 2. Put the heavy metal-antibiotic composite polluted sediment into small buckets at a rate of 2 kg per bucket;

[0047] The preparation method of the heavy metal-antibiotic polluted sediment includes the following steps:

[0048] (1) Air-dry the sediment naturally for 6 weeks. After air-drying, sieve it through a 10-mesh sieve (2 mm) and reserve it. Then sterilize it in a high-pressure steam sterilizer at 121 °C for 2 h to kill the indigenous fungi;

[0049] (2) For Cu and Zn, according to the dosing amounts of 40 mg / kg and 270 mg / kg of the sediment dry weight respectively, select CuSO4 and ZnCl2 to prepare solutions. For tetracycline (TC) and oxytetracycline (OTC), the dosing amounts are 20 mg / kg and 12 mg / kg of the sediment dry weight. Spray the required amounts of heavy metal salt solutions and liquid antibiotic standards evenly on the sediment with a sprayer, stir well and let it stand in the dark for 2 weeks to reach equilibrium and stability. Before the experiment, sterilize the prepared composite polluted sediment again in a high-pressure steam sterilizer at 121 °C for 2 h;

[0050] (3) Put the dry samples of the heavy metal-antibiotic composite polluted sediment into multiple small buckets (height 17 cm, upper diameter 16 cm, lower diameter 13 cm), and put 2.0 kg of dry sediment samples in each small bucket.

[0051] 3. Transplant the Glomus intraradices-Iris pseudacorus symbiotic system into the small buckets in step 2;

[0052] 4. After transplantation, cultivate for 4 weeks, then move the small buckets into a glass tank (length 30 cm, width 30 cm, height 25 cm) and immerse them in water. During daily management, add water manually to the large bucket to keep the overlying water on the sediment in the small bucket at 3 cm. After cultivating for 20 weeks, collect the sediment and host plant samples for analysis and determination.

[0053] II. Adopt the method of the control example

[0054] This experiment was set with two treatments, including an Iris pseudacorus control treatment (I) and a Glomus intraradices-Iris pseudacorus symbiotic system (HEMI) treatment, with three parallels for each treatment. The experimental methods were the same and the experimental time was 20 weeks.

[0055] III. Analytical and Determination Methods Adopted

[0056] Remove foreign matters such as twigs, leaves, and stones from the collected sediment samples, and then air-dry and grind them until they pass through a sieve with a pore size of 100 mesh. Weigh 0.1 g of the sample to be tested (accurate to 0.0001 g), place it in a 100 mL conical flask fumigated with aqua regia, add 6 mL of aqua regia solution, put on a glass funnel, and keep the aqua regia in a slightly boiling state on an electric hot plate for digestion for 2 h. After digestion, let it stand and cool to room temperature, filter the extract with a slow quantitative filter paper and collect it in a 50 mL volumetric flask. After the extract is completely filtered, wash the glass funnel, conical flask, and filter residue with a small amount of 0.5 mol / L nitric acid solution at least 3 times, filter the washing liquid and collect it in the volumetric flask, and make up the volume to the scale with ultrapure water. Then, measure the heavy metal concentration with an Agilent ICP-MS 7700.

[0057] Remove foreign matters such as branches, leaves, and stones from the sediment samples, take an appropriate amount and put it into a freeze dryer for dehydration. Grind the freeze-dried sample with a ball mill and pass it through a 60-mesh (pore size 0.25 mm) sieve for extraction. Weigh 2 g (accurate to 0.01 g) of the pre-sample into a 50 mL polypropylene centrifuge tube, extract it twice with acetonitrile-McIlvaine buffer solution, and combine the two extraction liquids. Dilute the extract with 1000 mL of pure water and adjust the pH value to about 3 with 4 mol / L sulfuric acid. Connect a SAX anion exchange column and an HLB solid-phase extraction column in series head-to-tail for solid-phase extraction. Elute it twice with 5 mL of methanol at a flow rate of about 0.5 mL / min, and collect all the eluates. Slowly blow the eluate to near dryness with high-purity nitrogen at 40 °C, make up the volume to 1.0 mL with a mixed solution of formic acid-ammonium acetate-acetonitrile, filter it with a 0.22 μm nylon filter membrane, and then measure the antibiotic concentration with a Thermo Fisher LC-MS / MS.

[0058] IV. Detection Results

[0059] 1. After 20 weeks of remediation, the residual amounts of Cu and Zn (i.e., the concentrations of Cu and Zn) in the sediment of each sample of the present invention are significantly lower than those of the control example. Compared with the control group (CK), in the treatment group transplanted with the Glomus intraradices-Iris pseudacorus symbiotic system (HEMI), the Cu removal rate increased by 74%, and the Zn removal rate increased by 60%.

[0060] 2. After 20 weeks of remediation, the residual amounts of TC and OTC (i.e., the concentrations of TC and OTC) in the sediment of each sample of the present invention are significantly lower than those of the control example. Compared with the control group (CK), in the treatment group transplanted with the Glomus intraradices-Iris pseudacorus symbiotic system (HEMI), the TC removal rate increased by 13%, and the OTC removal rate increased by 87%.

[0061] The above are only the preferred embodiments of the present invention and do not impose any limitation on the present invention. Any person skilled in the art, within the scope of the technical solution of the present invention, makes any form of equivalent replacement or modification and other changes to the technical solution and technical content disclosed by the present invention, which are all within the content of the technical solution of the present invention and still fall within the protection scope of the present invention.

Claims

1. A method for symbiotic phytoremediation of arbuscular mycorrhizal fungi, characterized in that the steps Including: (1) Remove impurities from the dried sample of sediment contaminated with heavy metals and antibiotics; (2) Transplant the arbuscular mycorrhizal fungi-aquatic plant symbiotic system to construct a sediment remediation system for heavy metal-antibiotic combined pollution; (3) Conduct repair and harvesting.

2. The method for repairing plants symbiotic with arbuscular mycorrhizal fungi according to claim 1, wherein The preparation method of the sediment contaminated with heavy metals and antibiotics: (1) After collecting the surface sediment without pollution history and air-drying it naturally for 6 weeks, sieve it and reserve it, and sterilize it under high pressure to kill indigenous fungi; (2) Add the required amount of heavy metal salt solution and antibiotic liquid standard to the sterilized sediment, spray it evenly multiple times, stir well, and let it stand in the dark for 2 weeks to balance and stabilize heavy metals and antibiotics, and obtain the sediment contaminated with heavy metal-antibiotic combination. Sterilize the prepared contaminated sediment again before the experiment.

3. The arbuscular mycorrhizal fungus symbiotic plant remediation method according to claim 2, characterized in that, In step (1), sterilize it in a high-pressure steam sterilizer at 121 °C for 2 h.

4. The arbuscular mycorrhizal fungus symbiotic plant remediation method according to claim 2, characterized in that, Sterilize it again in a high-pressure steam sterilizer at 121 °C for 2 h before the experiment.

5. The arbuscular mycorrhizal fungus symbiotic plant remediation method according to claim 2, wherein The surface sediment is the sediment at a depth of 10-20 cm.

6. The arbuscular mycorrhizal fungus symbiotic plant remediation method according to claim 1, characterized in that, The aquatic plant is one or more of Iris pseudacorus, Canna indica, Phragmites australis, Juncus effusus, Cyperus rotundus or Glyceria maxima.

7. The arbuscular mycorrhizal fungus symbiotic plant remediation method according to claim 1, characterized in that, The arbuscular mycorrhizal fungi are one or more of arbuscular mycorrhizal fungi in the Glomeromycotina.

8. The arbuscular mycorrhizal fungus symbiotic plant remediation method according to claim 1, characterized in that, The arbuscular mycorrhizal fungi-aquatic plant symbiotic system includes the host aquatic plant infected by arbuscular mycorrhizal fungi, the extraradical mycelium extending outside the roots of the host plant, and a small amount of pre-cultured soil remaining in the roots.

9. The arbuscular mycorrhizal fungus symbiotic plant remediation method according to claim 1, wherein, The heavy metals are Cu and Zn.

Images