Method for modifying nylon filler with iron-doped graphene to mediate synchronous fragmentation of microplastics and reduction of antibiotic resistance genes by chironomus tentans larvae

Abstract

This invention belongs to the field of environmental pollutant remediation technology and discloses a method for simultaneously breaking down microplastics and reducing resistance genes in midge larvae mediated by iron-doped graphene-modified nylon packing. This invention employs a composite process combining iron-doped graphene with nylon blending to prepare the modified packing, improving its mechanical properties, reusability, and environmental safety. It enhances the activity of microplastic-degrading enzymes, selectively screens and enriches functional bacterial communities, and increases the density of midge larvae. Through the synergistic interaction between the larval intestinal mechanical action and the intestinal flora, the simultaneous treatment effect of microplastics and resistance genes is strengthened, preventing their accumulation and spread in wastewater and residual sludge. This invention features significant synergistic treatment effects with unique targets, environmental friendliness and controllable cost, simple operation and easy scaling, and high resource recycling rate.

Description

Technical Field

[0001] This invention belongs to the field of environmental pollutant remediation technology, and relates to a method for mediating the simultaneous fragmentation of microplastics and reduction of resistance genes in chironomid larvae mediated by iron-doped graphene-modified nylon fillers. Background Technology

[0002] Microplastic pollution is a global concern. Microplastic pollution in the ocean has been extensively studied and reported. Microplastics can be transferred to higher trophic levels through food webs, posing a serious threat to human health. Wastewater treatment plants, as point sources of microplastic pollution and transit points for domestic wastewater discharge into the environment, have become a hot topic in microplastic research. In wastewater treatment plants, microplastics can combine with other pollutants to amplify their impact. The mobility of microplastics can provide a carrier for the spread of other pollutants, posing a dual threat to ecosystems and human health. Resistance genes, as a novel environmental pollutant, can spread among microorganisms through horizontal gene transfer, leading to enhanced bacterial resistance. The presence of microplastics significantly promotes the accumulation and spread of resistance genes, exacerbating environmental risks.

[0003] Microplastics, as recalcitrant organic polymers, are difficult to degrade using conventional wastewater treatment processes. Physical interception is typically used to reduce microplastic levels in discharged water, but intercepted microplastics usually accumulate in sludge, and the reuse and landfilling of this sludge can pose a risk of secondary pollution. Microplastic removal has become a major challenge in the field of biological wastewater treatment, making the development of green and efficient biological microplastic removal technologies imperative. Existing research has found that some aquatic organisms (such as cladocerans and fish) can ingest microplastics, but most organisms can only excrete the microplastics intact, failing to effectively break them down and degrade them, and having no effect on reducing resistance genes attached to the microplastic surface. Resistance gene reduction technologies mainly focus on disinfection (such as chlorination and ultraviolet disinfection) and bioinhibition (such as probiotic competition and bacteriophage lysis), but disinfection technologies easily generate disinfection byproducts, while bioinhibition technologies suffer from poor targeting and susceptibility to environmental interference. The synergistic treatment technology for microplastic and resistance gene co-pollution is still in the exploratory stage, and there is a lack of practical technologies that can simultaneously achieve microplastic fragmentation and degradation and resistance gene reduction while maintaining good environmental compatibility.

[0004] Biofilms in wastewater biological treatment systems have the ability to capture and intercept microplastics. However, the accumulation of microplastics in the biofilm induces oxidative stress in bacteria, increases cell membrane permeability, and may actually exacerbate the horizontal transfer of resistance genes. Therefore, in traditional wastewater treatment plants where bacteria are the primary functional organisms, it is difficult to achieve the simultaneous removal of microplastics and resistance genes through biological methods. Recently, the patent "A method for simultaneously removing antibiotics and resistance genes from wastewater by chironomid larvae using copper-doped graphene-modified fiber packing" describes a cascade wastewater biological treatment system constructed with copper-doped graphene-modified fiber packing. This system uses copper single atoms to increase the activity of superoxide dismutase in bacteria and chironomid larvae, reducing the destructive effect of reactive oxygen species induced by antibiotics and resistance genes on cell membrane permeability, reducing the horizontal transfer of antibiotic resistance genes, alleviating the toxic effects of antibiotics and resistance genes on chironomid larvae, and enabling the chironomid larvae to secrete DNase to degrade antibiotic resistance genes. Ultimately, this achieves the goal of simultaneously removing antibiotics and resistance genes from wastewater, without addressing microplastics, a recalcitrant pollutant. The literature "Biofragmentation of Polystyrene Microplastics: A Silent Process Performed by Chironomus sancticaroli Larvae" indicates that midge larvae can ingest and mechanically break down polystyrene microplastics, but it does not involve resistance genes. Previous literature reports that microplastics can significantly enrich resistance genes in wastewater (literature "Study on the Enrichment Characteristics of Intracellular and Extracellular Resistance Genes in Urban Wastewater by Microplastics"), exacerbating the co-occurrence and horizontal transfer of antibiotic resistance genes (literature "Microplastics exacerbate co-occurrence and horizontal transfer of antibiotic resistance genes"). Microplastic diversity increases the abundance of antibiotic resistance genes (literature "Microplastic diversity increases the abundance of antibiotic resistance genes in soil"), making the removal of antibiotic resistance genes in the presence of microplastics a challenge. Therefore, although the aforementioned reports indicate that metazoans, represented by midge larvae, have the potential to break down microplastics and remove resistance genes, the presence of microplastics exacerbates the horizontal transfer of resistance genes and increases their abundance. Whether existing technologies can effectively remove resistance genes in the presence of microplastics remains unknown. Furthermore, both pollutants cause oxidative stress in midge larvae, accelerating the production of reactive oxygen species, leading to cell membrane lipid peroxidation and accelerating larval death. This makes the goal of "simultaneously breaking down microplastics and reducing resistance genes in midge larvae" difficult to achieve.

[0005] The gut microbiota provides its host with a variety of beneficial services, such as providing sufficient nutrients, defending against pathogens, regulating the immune system, detoxifying invading insecticides, and promoting development. However, the structure and function of the gut microbiota in midge larvae exposed to the dual stresses of microplastics and resistance genes are still poorly understood. The ability and mechanism by which gut microbiota assists midge larvae in coping with microplastic and resistance gene stresses remain unclear. Whether targeted regulation of the gut microbiota can help midge larvae resist the dual stresses of microplastics and resistance genes, thereby achieving simultaneous microplastic fragmentation and resistance gene reduction, has not yet been reported.

[0006] This invention proposes a method for simultaneously breaking down microplastics and reducing resistance genes in chironomid larvae mediated by iron-doped graphene-modified nylon packing. This method overcomes the limitations of existing technologies that only focus on the removal efficiency of either microplastics or resistance genes by chironomid larvae. By using iron atoms, which have better environmental compatibility, as dopant, iron atoms act as enzyme activators, promoting the secretion of microplastic-degrading enzymes (such as polyesterase and lipase) by functional bacteria such as *Pseudomonas* and *Bacillus*, enhancing the activity of intestinal digestive enzymes and nucleases, inhibiting the growth of host bacteria of resistance genes, and decomposing resistance genes. Combining the mechanical advantages of nylon, graphene-modified nylon composite packing is prepared, and a modified packing-enhanced wastewater treatment reactor is constructed. Through the targeted enrichment of surface bacteria and habitat regulation by the packing, the growth and reproduction of chironomid larvae are promoted, and the intestinal flora of the larvae is regulated. This fosters a mutually beneficial cooperation between the larvae and the intestinal flora, constructing a three-in-one synergistic system of "iron-doped graphene-nylon composite packing - chironomid larva - functional bacteria." This provides a solution to the more complex environmental problem of combined pollution by microplastics and resistance genes, filling a gap in existing technologies. Summary of the Invention

[0007] This invention provides a method for simultaneously breaking down microplastics and reducing resistance genes in chironomid larvae using iron-doped graphene-modified nylon packing. The modified packing is prepared using a composite process combining iron-doped graphene with nylon blending, improving its mechanical properties, reusability, and environmental safety. This enhances the activity of microplastic-degrading enzymes, selectively enriches functional bacterial communities, and increases chironomid larval density. Through the synergistic interaction between the larval intestinal mechanical action and the intestinal flora, the simultaneous treatment of microplastics and resistance genes is strengthened, preventing their accumulation and spread in wastewater and residual sludge.

[0008] The technical solution of this invention:

[0009] A method for simultaneously breaking down microplastics and reducing resistance genes in midge larvae mediated by iron-doped graphene-modified nylon fillers, comprising the following steps:

[0010] Step 1: Mix 3-10 layer graphene powder, ferric chloride, and nylon 6 chips at a mass ratio of 1:0.2:8-1:0.5:12, add to a twin-screw extruder, and melt-blend at 230-250℃ for 10-15 min to obtain a composite spinning masterbatch; feed the composite spinning masterbatch into a melt spinning machine with an aperture of 0.1-0.2 mm, and spin at 240-260℃ and a spinning speed of 800-1200 m / min to obtain nascent fibers; calcine at a heating rate of 5℃ / min, and place the nascent fibers in a 5wt.%-10wt.% borax solution at 30-50℃ for 1-2 h for crosslinking, then calcine at 300-400℃ for 2-3 h under a nitrogen atmosphere to remove residual impurities and enhance the bonding force between iron and graphene, obtaining a diameter of 50-100 mm. The iron-doped graphene-nylon composite packing material has a diameter of 1-2 mm. This material is then spun into multiple strands to form fine fiber ropes with a diameter of 1-2 mm, which are then braided into fiber ropes with a diameter of 0.5-1.5 cm for use in the subsequent construction of wastewater treatment reactors. The iron-doped graphene-nylon composite packing material has a specific surface area of ​​20-120 m² / g, a tensile strength ≥3.5 cN / dtex, a porous surface structure, and exhibits both hydrophobic and hydrophilic properties, as well as good biocompatibility.

[0011] Step 2: Collect and screen chironomid larvae from natural water body sediment or sewage treatment plants. Place the chironomid larvae in simulated polluted water for acclimatization and cultivation for 7-10 days. During the acclimatization process, maintain a dissolved oxygen content of 2-5 mg / L, a temperature of 20-25℃, a pH of 6.5-8.0, and a photoperiod of 12-16 h light + 12-8 h darkness. Iron-doped graphene-nylon composite packing material is pre-added to the simulated polluted water at a dosage of 5-10 g / L to provide attachment sites for the chironomid larvae. Simultaneously, a nutrient regulator with a mass concentration of 0.1-0.5 g / L is added to the simulated polluted water. The nutrient regulator is a mixture of yeast extract, peptone, and glucose in a mass ratio of 2:1:1 to maintain the physiological activity of the chironomid larvae and functional bacterial flora.

[0012] Step 3: Iron-doped graphene-nylon composite packing fiber ropes are laid in a curtain-like arrangement in the wastewater treatment reactor, with a filling rate of 30%-50%, forming a three-dimensional attachment space. Domesticated chironomid larvae are introduced into the reactor at a biomass density of 0.5-2 g / L. At this point, the iron-doped graphene-nylon composite packing increases the chironomid larva density to 1.5-3 g / L. The surface of the iron-doped graphene-nylon composite packing will selectively enrich functional bacteria such as *Pseudomonas* and *Bacillus*, increasing bacterial abundance by 2-3 orders of magnitude. Iron promotes the secretion of microplastic-degrading enzymes by these bacteria. Strong sunlight should be avoided directly in the wastewater treatment reactor. The bacterial structure and chironomid larvae attachment on the surface of the iron-doped graphene-nylon composite packing should be monitored regularly to ensure system stability.

[0013] Step 4: Treat wastewater containing microplastics and resistance genes using a synergistic system of "iron-doped graphene nylon composite packing material - chironomid larvae - functional bacteria". The reaction temperature is 18-30℃, dissolved oxygen content is 1.5-4 mg / L, water retention time is 10-48 h, and the photoperiod is 12-16 h light + 12-8 h darkness. The influent COD concentration is 80-250 mg / L, ammonia nitrogen concentration is 15-50 mg / L, microplastic concentration is 0.15-0.35 g / L, particle size is 10-100 cm, and resistance gene concentration is 0.5-10 ng / μL. 8 -10 9 Copy number / μL;

[0014] Step 5: Regularly sample and test the particle size distribution, concentration, and abundance of resistance genes of microplastics in the water to evaluate the treatment effect of the system; after the chironomid larvae emerge, they can be collected by trap nets and used as fish bait or made into bio-organic fertilizer. The iron-doped graphene-nylon composite filler can be reused for more than 10 years after cleaning.

[0015] Microplastics include one or more of the following: polyvinyl chloride (PVC), polyethylene (PE), polypropylene (PP), and polystyrene (PS).

[0016] The host bacteria for resistance genes include plasmids or bacterial groups carrying one or more of the following: sulfonamides, tetracyclines, and chloramphenicol.

[0017] The beneficial effects of this invention are as follows: The few-layer graphene used in this invention has higher conductivity and electron migration efficiency than multilayer or larger graphene systems, thus promoting stronger microbial metabolic performance and requiring less dosage. The defective structure of graphene itself coordinates with iron ions, complexing them into the graphene structure. This stabilizes the iron ions and enhances the conductivity of the graphene itself, simultaneously fixing them, simplifying the preparation process, increasing efficiency, extending service life, and saving costs. The iron-doped graphene-nylon composite filler, with its high specific surface area, porous structure, and excellent mechanical properties, provides a stable attachment carrier for midge larvae, increasing their density in the reactor by 30%-60% compared to the unfilled system, preventing larvae from being lost with the water flow. The iron atoms in the filler are not only environmentally safe but also act as enzyme activators, promoting the secretion of microplastics by functional bacteria such as Pseudomonas and Bacillus. The larvae of the midge (Centella asiatica) ingest enzymes (such as polyesterase and lipase) and inhibit the growth of host bacteria carrying resistance genes. They exhibit high feeding activity, efficiently consuming microplastic particles and functional bacteria from the water and packing material surfaces via their mouthparts. Microplastics are mechanically broken down within their digestive tract, significantly reducing their particle size. After ingestion, some of the bacteria enriched in the packing material form synergistic interactions with the larvae's existing gut flora. With the synergistic effect of iron, this enhances the activity of intestinal digestive enzymes, accelerating the erosion and chemical destruction of microplastic surfaces, and increasing the secretion of nucleases in the gut. For resistance genes and host bacteria attached to the microplastic surface, the midge larvae can directly ingest the host bacteria during feeding. The digestive enzymes, symbiotic microorganisms, and functional bacteria in the gut collectively disrupt the host bacterial cell structure, leading to the release of resistance genes. These released resistance genes are then degraded into nucleotide fragments by highly active nucleases, achieving resistance gene reduction. Furthermore, the metabolic activities of the midge larvae and the metabolic products of the bacteria enriched in the packing material mutually promote each other, further optimizing the aquatic microbial community structure, inhibiting horizontal transfer of resistance genes, and reducing the risk of transmission.

[0018] (1) The synergistic treatment effect is significant and the target is unique: Compared with previous patents or literature that focused on the single removal of microplastics or resistance genes, this invention innovatively achieves the simultaneous breaking of microplastics and reduction of resistance genes. Under the mediation of iron-doped graphene modified nylon fiber filler, the mechanical breaking of chironomid larvae and the biodegradation of microbial community work synergistically. The amount of microplastics ingested by chironomid larvae can reach 10-50 MPs / item, and the abundance of resistance genes is reduced by 30-100%.

[0019] (2) Environmentally friendly and cost-controllable: Chironomid larvae are widely distributed in natural water bodies and have strong reproductive capacity, making them easy to obtain, domesticate and cultivate on a large scale; the iron-doped graphene-nylon composite filler used in this invention uses iron atoms, which have good biocompatibility and low leaching risk, and the combination with nylon blending improves the mechanical properties and reusability of the filler (it can be reused for more than 10 years); the water quality index range is wide, and it can achieve high stability under water quality fluctuation conditions.

[0020] (3) Simple operation and easy to scale up: The treatment system of the present invention is simple to construct, the environmental parameters are easy to control, and it is suitable for different types of water bodies containing microplastics and resistance genes (such as municipal sewage, aquaculture wastewater, pharmaceutical wastewater, plastic processing wastewater, decentralized domestic sewage, etc.), which facilitates large-scale engineering applications.

[0021] (4) High resource recycling rate: The chironomid larvae after treatment can be used as fish feed or made into bio-organic fertilizer, realizing the recycling of biological resources and improving the overall benefits of the technology; at the same time, after the chironomid larvae lay eggs, they can be cultivated in the sewage treatment reactor for generations, with a survival period of about 6-8 months, a long treatment cycle, and a high resource recycling rate. Attached Figure Description

[0022] Figure 1 This is an image of the curtain-like fiber ropes in the constructed wastewater treatment reactor.

[0023] Figure 2 These are images of chironomid larvae in a wastewater treatment reactor constructed with modified packing material.

[0024] Figure 3 This refers to the uptake of 10μm and 100μm polyethylene microplastics by midge larvae.

[0025] Figure 4 It is the Raman spectrum of 10 μm polyethylene microplastics after being ingested by midge larvae.

[0026] Figure 5 It is the Raman spectrum of 100 μm polyethylene microplastics after being ingested by midge larvae.

[0027] Figure 6 It represents the abundance of resistance genes in midge larvae after they ingest polyethylene microplastics. Detailed Implementation

[0028] The specific embodiments of the present invention are described in detail below with reference to the technical solutions and accompanying drawings.

[0029] Example 1: Treatment of simulated water containing polyethylene microplastics and the sul1 resistance gene

[0030] (1) Construction of modified packing enhanced wastewater treatment reactor: Iron-doped graphene-nylon composite packing was prepared according to step 1 (graphene:ferric chloride:nylon 6 chip mass ratio 1:0.3:10, melt spinning temperature 250℃, spinning speed 1000 m / min, borax solution concentration 8%, calcination at 350℃ for 2.5 h), and laid in a curtain-like arrangement in a 30 L wastewater treatment reactor ( Figure 1The packing material filling rate was 40%. Activated sludge mixture was taken from a well-functioning wastewater treatment plant as inoculum sludge. After acclimatization, it was inoculated into the wastewater treatment process enhanced by modified packing material. The reaction device was started and aerated for 15 days to allow the modified packing material to grow a biofilm naturally. Subsequently, the reactor was fed with normal water, with an influent COD concentration of 100-200 mg / L, an ammonia nitrogen concentration of 25-45 mg / L, an influent flow rate maintained at 18 L / h, an internal recirculation rate of 100%, an aeration rate of 0.5 L / min, and a hydraulic retention time of 24 h.

[0031] (2) Screening and domestication of red chironomid larvae: Chironomid larvae were collected from the sewage treatment plant and screened by microscopic observation (body length 3-4 mm, head reddish-brown, abdomen with bare beard-like appendages); the red chironomid larvae were placed in the above-mentioned sewage treatment reactor for domestication, and the density of red chironomid larvae increased after domestication. Figure 2 Furthermore, it can continuously reproduce and grow. With a density of 20-40 cells / L, dissolved oxygen controlled at 3 mg / L, temperature at 20.0±2.0℃, pH at 7.0±0.5, and a low-light environment maintained with a light cycle of 16 h light / 8 h darkness, the abundance of Pseudomonas spp. on the packing surface increased by 2.5 orders of magnitude compared to the initial level.

[0032] (3) Construction and regulation of the treatment system: Polyethylene microplastics and resistance gene plasmids of different particle sizes were added to the influent. The polyethylene microplastics had particle sizes of 10 μm and 100 μm, and a concentration of 250 mg / L. The plasmids carrying the sul1, cml_e3, and tetx resistance genes had a concentration of 1 ng / μL (10 8 The microplastic ingestion, microplastic fragmentation, and resistance gene abundance of chironomid larvae were examined after 16 days of treatment (copy number / μL).

[0033] (4) Monitoring of treatment effect: The test results showed that the average number of 10 and 100 μm polyethylene microplastics ingested by midge larvae in the water was 28 and 20 per larva, respectively. The intake of small-particle (10 μm) polyethylene microplastics was higher than that of large-particle microplastics. Figure 3 Raman spectroscopy showed that the typical peaks of polyethylene microplastics significantly decreased after being ingested by larvae, indicating that significant physical fragmentation had occurred. Figure 4 , Figure 5 Under both particle size microplastics, the abundance of sul1, cml_e3, and tetx resistance genes was significantly reduced, with the largest decrease observed during the uptake of smaller-sized microplastics. The abundance of sul1, cml_e3, and tetx resistance genes decreased by 55%, 55%, and 100% and 32%, 44%, and 67%, respectively, during the breakage of 10 and 100 μm polyethylene microplastics (see appendix). Figure 6Increased abundance of Acinetobacter and Glutamicibacter in the gut microbiota enhanced the degradation capacity of pollutants. Simultaneously, both chironomid larvae and the gut microbiota upregulated the ribosome and cysteine / methionine metabolism pathways to promote the synthesis of antioxidant enzymes and reduce oxidative damage. At the same time, larvae downregulated the ABC transporter and alanine, aspartate, and glutamate metabolism pathways to conserve energy during detoxification, while these pathways were upregulated in the gut microbiota, thus strategically providing nutrition to the larval host and enhancing its tolerance to pollution stress. Iron leaching was less than 0.02 mg / L, meeting national wastewater discharge standards.

[0034] Example 2: Treatment of simulated wastewater containing polystyrene microplastics and sul2 and tetg resistance genes.

[0035] The modified packing material was prepared according to the steps in Example 1, a wastewater treatment reactor was constructed, and chironomid larvae were domesticated. 10 μm polystyrene microplastics and plasmids carrying the sul2 and tetg resistance genes were added to the influent. The concentration of polyethylene microplastics was 200 mg / L, and the concentration of plasmids carrying the sul2 and tetg resistance genes was 0.5 ng / μL. After 3 days of treatment, the ingestion of microplastics by chironomid larvae, the degree of microplastic fragmentation, and the abundance of resistance genes were investigated. The number of polystyrene microplastics ingested by chironomid larvae in the water was approximately 15-45 per larva, and the abundance of the sul2 and tetg resistance genes decreased by 34% and 21%, respectively.

[0036] Example 3

[0037] Based on Example 1, the conditions for the domestication process are changed to:

[0038] The dissolved oxygen content was maintained at 5 mg / L, the temperature at 25℃, the pH at 8.0, and the photoperiod was 12 hours of light + 12 hours of darkness.

[0039] Example 4

[0040] Based on Example 1, the conditions for treating wastewater containing microplastics and resistance genes were changed to:

[0041] The reaction was conducted at 18℃, with a dissolved oxygen content of 1.5 mg / L, a water residence time of 10 h, a light cycle of 12 h light + 12 h dark, and influent COD concentration of 80 mg / L, ammonia nitrogen concentration of 15 mg / L, polyethylene microplastic concentration of 0.15 g / L, and plasmid concentration carrying sul1, cml_e3, and tetx resistance genes of 0.5 ng / μL. 9 Copy number / μL.

[0042] Comparative Example 1: Control group without modified filler reinforcement and microbial regulation, and fed with midge larvae.

[0043] When midge larvae were placed in an aquatic environment without fillers or biofilms, and the same influent and treatment conditions were maintained as in Example 1, the amount of polyethylene microplastics ingested was basically similar. However, after 48 hours of treatment, the mortality rate of midge larvae reached 40%, and they could not continue to ingest and decompose pollutants, nor could they maintain molting, reproduction, and high-density growth.

[0044] Comparative Example 2: Control group with modified filler and no chironomid larvae

[0045] A modified packing-enhanced wastewater treatment reactor was constructed according to the steps in Example 1, without the addition of chironomid larvae, to treat 10 μm, 250 mg / L polyethylene microplastics, 1 ng / μL (10 8 The plasmid containing the sul1 resistance gene (copy number / μL) showed no significant change in microplastics after 3 days of treatment, remaining in the biofilm, and the abundance of the sul1 resistance gene was not significantly different.

[0046] Comparative Example 3: A control group using commercially available nylon fiber filler and without chironomid larvae feeding.

[0047] A wastewater treatment reactor of the same size was constructed following the steps in Example 1. The packing material used was nylon fiber of similar size with no added midge larvae. It was used to treat 10 μm, 250 mg / L polyethylene microplastics at a concentration of 1 ng / μL (10 8 The plasmid containing the sul1 resistance gene (copy number / μL) showed no significant change in microplastics after 3 days of treatment, remaining in the biofilm, while the abundance of the sul1 resistance gene increased by 35%.

[0048] The above description, in conjunction with specific preferred embodiments, provides a further detailed explanation of the present invention, enabling those skilled in the art to understand and apply it. However, it should not be construed that the specific implementation of the present invention is limited to these descriptions. For those skilled in the art, several simple deductions or substitutions can be made without departing from the concept of the present invention, without requiring creative effort. Therefore, any simple improvements made to the present invention by those skilled in the art based on the disclosure of the present invention should be within the scope of protection of the present invention.

Claims

1. A method for simultaneously breaking down microplastics and reducing resistance genes in chironomid larvae mediated by iron-doped graphene-modified nylon fillers, characterized in that... The steps are as follows: Step 1: Graphene powder, ferric chloride, and nylon 6 chips are mixed to prepare a composite spinning masterbatch; the composite spinning masterbatch is further spun to obtain nascent fibers; the nascent fibers are crosslinked in a 5wt.%-10wt.% borax solution at 30-50℃ for 1-2 hours, and then calcined at 300-400℃ for 2-3 hours under a nitrogen atmosphere to remove residual impurities and enhance the bonding force between iron and graphene, resulting in an iron-doped graphene-nylon composite filler with a diameter of 50-100 μm; the iron-doped graphene-nylon composite filler is multi-stranded and spun into fiber ropes with a diameter of 1-2 mm, and then mixed and woven into fiber ropes with a diameter of 0.5-1.5 cm; Step 2: Collect and screen midge larvae, and place them in simulated polluted water for acclimatization and cultivation for 7-10 days; pre-add iron-doped graphene-nylon composite filler to the simulated polluted water at a dosage of 5-10 g / L to provide attachment sites for the midge larvae; at the same time, add a nutrient regulator at a mass concentration of 0.1-0.5 g / L to the simulated polluted water. Step 3: Lay fiber ropes in a curtain-like arrangement in the wastewater treatment reactor, controlling the filling rate of iron-doped graphene-nylon composite packing to 30%-50%, forming a three-dimensional attachment space; add domesticated chironomid larvae into the wastewater treatment reactor at a biomass density of 0.5-2 g / L, and the iron-doped graphene-nylon composite packing increases the chironomid larvae density to 1.5-3 g / L; avoid direct sunlight on the wastewater treatment reactor, and regularly test the bacterial community structure on the fiber rope surface and the attachment of chironomid larvae to ensure system stability; Step 4: Treat wastewater containing microplastics and resistance genes using a synergistic system of iron-doped graphene nylon composite filler, chironomid larvae, and functional microbial communities; Step 5: Regularly sample and test the particle size distribution, concentration, and abundance of resistance genes of microplastics in the water to evaluate the treatment effect of the system; after the midge larvae emerge, they are collected by trap nets and used as fish bait or made into bio-organic fertilizer. The fiber ropes can be reused after washing.

2. The method for simultaneously breaking down microplastics and reducing resistance genes in chironomid larvae mediated by iron-doped graphene-modified nylon filler according to claim 1, characterized in that... In step 1: Graphene powder with 3-10 layers, ferric chloride, and nylon 6 chips are mixed at a mass ratio of 1:0.2:8-1:0.5:

12. The mixture is added to a twin-screw extruder and melt-blended at 230-250℃ for 10-15 min to obtain composite spinning masterbatch; The composite spinning masterbatch is fed into a melt spinning machine with an aperture of 0.1-0.2 mm, and nascent fibers are obtained by spinning at 240-260℃ and a spinning speed of 800-1200 m / min.

3. The method for simultaneously breaking down microplastics and reducing resistance genes in chironomid larvae mediated by iron-doped graphene-modified nylon filler according to claim 1, characterized in that... In step 2: During the acclimatization process, the dissolved oxygen content should be maintained at 2-5 mg / L, the temperature at 20-25℃, the pH at 6.5-8.0, and the photoperiod should be 12-16 h of light + 12-8 h of darkness. The nutritional regulator is a mixture of yeast extract, peptone, and glucose in a mass ratio of 2:1:

1.

4. The method for simultaneously breaking down microplastics and reducing resistance genes in chironomid larvae mediated by iron-doped graphene-modified nylon filler according to claim 1, characterized in that... The microplastics include one or more of polyvinyl chloride, polyethylene, polypropylene, and polystyrene.

5. The method for simultaneously breaking down microplastics and reducing resistance genes in chironomid larvae mediated by iron-doped graphene-modified nylon filler according to claim 1, characterized in that... The host bacteria of the resistance gene include one or more combinations of strains or bacterial groups carrying plasmids of sulfonamide resistance genes, tetracycline resistance genes, and chloramphenicol resistance genes.

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