Abstract
Microplastics pollution in agricultural soil is increasingly recognized, but the specific situation varies with geography, climate conditions, and farming practices. The karst landscape, a typical geomorphology in China, demands a deeper understanding of microplastics pollution in such areas. This research zeroes in on Guizhou, a province known for its karst formations, by collecting soil samples from the mulched cultivation layer in ten counties and cities. The study employed metallographic microscopy, scanning electron microscopy (SEM-EDS), and Fourier-transform infrared spectroscopy (FT-IR) to analyze the presence and distribution of microplastics. Results show that polyethylene is the predominant component of microplastics in the mulched agricultural soils of Guizhou, primarily existing as irregular fragments in black, transparent, and translucent forms, with diameters of 40 ~ 120 μm and rough surfaces marked by significant erosion. The concentration of microplastics varies from 143.28 to 3,283.46 items/kg, averaging 1,150.60 ± 647.86 items/kg. The majority of particles accounting for 64.79% are sized between 10 ~ 100 μm. A highly significant positive correlation (p < 0.001) is found between mulching duration and microplastics concentration, indicating that prolonged mulching increases microplastics accumulation in farmlands. Additionally, crop type, irrigation method, and soil type also influence microplastics concentration. This study highlights the escalating issue of microplastics pollution in China’s karst regions, underscoring the need for attention.
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Introduction
Plastic plays a vital role in our daily lives, offering convenience and ubiquity. Global plastic production surged from 1.7 million tonnes in 1950 to 348 million tonnes in 20171. In agriculture, plastics are used to control weeds, conserve water, and increase soil temperature. The widespread adoption of mulching technology has led to a sharp increase in the global use of agricultural films. In 2019, China produced 2.4695 million tonnes of agricultural film, covering an area of over 25 million hectares, accounting for 61% of global usage2. Post-use, these films are prone to significant aging and are difficult to recycle, leaving large residues in the agricultural environment ranging from 50 to 260 kg/hectare. Of particular concern is the fragmentation of these films due to mechanical tilling and natural degradation, resulting in microplastics, which become a significant source of microplastics in soil, water, and air environments3,4.
Microplastics are tiny plastic fragments with a size of less than 5 mm, a term coined by Thompson in 20045. Microplastics, characterized by their relatively stable physicochemical properties, are resistant to degradation in natural environments and possess strong migration capabilities, leading to easily dispersed pollution. This has drawn widespread attention from scholars both domestically and internationally in recent years6. Microplastics are characterized by a large specific surface area and a propensity to adsorb other contaminants. Compared to pristine microplastics, those carrying pollutants are more prone to desorption during transport in acidic media or within organisms7,8,9. Once microplastics enter the soil, they directly alter soil properties, disrupting microbial habitats and altering the distribution of anaerobic and aerobic microorganisms10. Indirectly, they also cause shifts in plant rhizosphere microbial communities, affecting plant growth11,12,13. Microplastics items can infiltrate plant tissues, accumulate in leaves, stems, and roots, and can migrate to edible parts, posing threats to food safety and human health14,15.
Farmland soils in karst regions are particularly susceptible to microplastics pollution due to their unique geological characteristics and agricultural activities. The high permeability and complex terrain of karst landscapes facilitate the migration of microplastics through rock fractures, hydraulic transport, and other mechanisms16,17. Additionally, the recovery of agricultural films in these mountainous areas is more challenging, exacerbating the issue of microplastics contamination in farmland soils18. Studies have shown that the complex groundwater networks in karst regions lead to an uneven distribution of microplastics in soils, with potential hotspots of accumulation in certain areas19. Therefore, the problem of microplastics pollution in karst regions warrants significant attention.
The negative impacts of microplastics on soil and terrestrial ecosystems underscore the need for proactive prevention and mitigation measures. Current research primarily focuses on marine and aquatic pollution, with comparatively less attention on soil, particularly agricultural soil. This study, set in Guizhou Province, established 20 sampling sites across 10 counties, collecting topsoil samples from 0 to 30 cm depth. The abundance, shape, and size of soil microplastics were analyzed using metallographic microscopy and FT-IR, while their micro-morphology and elemental composition were observed with SEM-EDS. The study aims to: (1) ascertain the abundance and distribution of microplastics in 10 counties of Guizhou Province; (2) determine the morphological characteristics of microplastics in cultivated soils; and (3) investigate the factors influencing the abundance of microplastics in cultivated soils. The results will provide a theoretical basis for the study of microplastics pollution in arable soils.
Materials and methods
Study area and sample collection
Guizhou is located on the Yunnan-Guizhou Plateau, spanning from 103°36′ E to 109°35′ E, and from 24°37′ N to 29°13′ N. It features a subtropical humid monsoon climate with an average annual temperature of 16 °C, sunlight duration of 1,200 to 1,600 h, average annual precipitation of 1,000 to 1,400 mm, and an average relative humidity of 79.4%. The study focused on fields where major crops such as vegetables, chili peppers, tobacco, and potatoes are cultivated under plastic mulch, identifying these as key monitoring sites for residual microplastics. Twenty monitoring sites for plastic mulch residues were established in ten counties across nine municipalities in Guizhou Province. Locations were determined using a GPS device (eTrex 221, Garmin) and marked on a map with ArcGIS software (Fig. 1). At each site, five quadrats measuring 100 × 100 cm were laid out using a ruler from the soil surface, and soil samples were collected to a depth of 30 cm, yielding approximately 1 kg of soil from the 0–30 cm layer for analysis.
Sample processing and microplastics extraction
The extraction process of sample microplastics was shown in Fig. 2. Air-dried samples were oven-dried at 60 °C for 24 h, and from these, 100 g of soil, excluding items over 5 mm such as plastic debris, stones, and weeds, were placed into a 250 mL wide-mouth conical flask. The flask was filled with 150 mL of saturated NaCl solution (density 1.2 g/mL) and subjected to 5 min of ultrasonication to break up the soil aggregates, followed by 30 min of shaking to release microplastics. The suspension was allowed to settle for 24 h before the supernatant, which now contained microplastics, was collected. This ultrasonication, shaking, and supernatant collection process was repeated three times to fully extract microplastics from the soil. The supernatant was then vacuum-filtered through a 0.45 μm glass fiber filter, and 20 mL of 30% hydrogen peroxide was added to digest organic matter. The mixture was agitated on a shaker at 50 °C for 72 h for complete digestion, followed by another filtration through a 0.45 μm glass fiber filter. The filtered samples were collected for microscopic analysis.
Observation and identification of microplastics
All substances were identified and observed for concentration, size, and type using an upright metallurgical microscope (L302-HK830, Shenzhen Aokes Optical Instrument Co.). The size, type, and shape of all microplastics were recorded. Microplastics items selected from the extracted samples were analyzed using a Fourier Transform Infrared Spectrometer (LUMOS II, Bruker), with spectral data collected in the range of 400 ~ 4000 cm− 1, scanning the sample 64 times at a resolution of 2 cm− 1. Spectra obtained were processed using OPUS software. The spectra were compared with standard spectra to characterize their chemical structures and morphological features were observed and recorded. The provenance of polythene powder and fresh plastic mulch from Dongguan Jinheng Plastic Co. All collected microplastics were coated with gold and analyzed for surface morphology and elemental composition using a Scanning Electron Microscope coupled with an Energy Dispersive Spectrometer (Sigma 300, ZEISS).
Experimental quality control
To avoid potential microplastics contamination during the experiment, all containers were rigorously cleaned with ultrapure water, ultrasonicated, dried, and wrapped in aluminum foil before use. Gloves and cotton lab coats were worn during sampling and laboratory analysis. Laboratory doors and windows were kept closed and unnecessary movement was minimized to reduce potential airborne contamination. Three blank control groups were set up to monitor environmental microplastics contamination during the experimental process. No microplastics were detected in these blank controls throughout the subsequent detection processes.
Data analysis
Data were processed using Microsoft Excel 2016 and SPSS 27.0, employing one-way Analysis of Variance (ANOVA) and non-parametric tests for single samples to examine the impact of environmental factors on the distribution characteristics of microplastics. Graphs were created using GraphPad Prism 9.0.0. Principal Component Analysis (PCA) was conducted with Canoco5 to comprehensively analyze the influence of environmental factors on microplastics and scatter plots were generated.
Results and discussion
Morphology and composition of microplastics
Metallographic microscope analysis of microplastics isolated from soil samples, exemplified by plot QXG, is presented in Fig. 3. The results demonstrate that soil microplastics predominantly appear as irregular black fragments (Fig. 3a,c,e,g,i), transparent (Fig. 3b), or semi-transparent (Fig. 3d,f,h), with diameters ranging from 40 to 120 μm. The varied colors of microplastics might be attributed to the pigments of their precursor plastic films and age-related degradation in the environment20. It has been reported that pigments in microplastics degrade gradually under the influence of sunlight, microbes, and humidity, leading to a fading of color21,22. Similar distributions of colored microplastics have been noted by Ren in their studies on the contribution of agricultural mulching films to pollution in Chinese agricultural soils and surface waters23. Moreover, mechanical actions such as tilling and weeding can stretch and fragment the plastic films in the soil into various irregular shapes24. Studies have indicated that primary microplastics from industrial production and personal care products are typically spherical, while microplastics in water bodies are commonly fibrous25,26. Fibrous and fragmentary microplastics are the main forms found in the Yangtze River estuary and its sediments, with spherical microplastics also detected27,28. The diameter of microplastics is linked to the physicochemical properties of their precursors (such as tensile properties, and mechanical strength) as well as their exposure environment and duration. Generally, the poorer the mechanical properties of the microplastics precursors, the harsher the exposure environment, and the longer the exposure duration, the smaller the diameter of the resulting microplastics29.
To identify the primary constituents of microplastics, representative samples (black, transparent, and semi-transparent) were compared with polyethylene powder and fresh plastic mulching films using FTIR. The results (Fig. 4) showed that polyethylene powder primarily exhibited four characteristic absorption bands, representing C = C-H (2916, 2843, and 719 cm− 1) and C = C (1472 cm− 1)30. The absorption peaks of fresh plastic mulching films aligned perfectly with those of polyethylene powder, indicating polyethylene as their main component. It is noteworthy that the three types of microplastics exhibit characteristic peaks of polyethylene, as well as vibrational absorption peaks indicative of C-O-C/-OH groups (1183, 1116, 1083, 1021, and 1017 cm− 1) and halogen-associated absorption peaks (608, 531, and 465 cm− 1). This suggests that polyethylene constitutes the major component of these microplastics. The presence of halogen functional groups may stem from the adsorption and fixation of halogenated contaminants (e.g., pesticides) from the environment, while the oxygen-containing groups could be related to the aging/oxidation of plastics in environmental conditions31.
Microplastics morphology and composition
To observe the microstructure of microplastics more clearly, their surface morphology and corresponding composition were analyzed using SEM-EDS. The results show that the microplastics depicted in Fig. 5a–c and e–g have relatively rough surfaces with evident signs of erosion, forming grooves, pits, and holes. This suggests prolonged environmental exposure and stronger aging effects32. EDS analysis indicates that these microplastics primarily consist of chemical elements such as C, N, O, Na, Al, Si, Ca, Fe, K, and Cl (Fig. 5d,h,l,p). The elements O, Na, Al, Si, Ca, Fe, K, and Cl mainly originate from the interaction of microplastics with minerals during the aging process and their adsorption of contaminants including Cl33. Conversely, the microplastics in Fig. 5i–k have comparatively smooth and less eroded surfaces, indicating a shorter formation period. Accordingly, lesser hetero-elements (mainly C, N, O, Na, Al, Si, Ca, and Fe) are found on their surfaces. Figure 5m–o shows microplastics with numerous fine items and a rich porous structure on their surfaces, suggesting a different aging effect. Such porous and rough surfaces increase the surface area of the microplastics, potentially enhancing their adsorption capacity for other pollutants in the soil34,35. O-type functional groups can activate mineral surfaces, enhancing the hydrogen bonding, ligand exchange, and hydrophobicity of microplastics36. The varied apparent morphologies of microplastics indicate that due to the differences in the physicochemical properties of their precursors, the microplastics undergo different aging processes in the environment. The main hetero-elements increased in the microplastics are O, Na, Al, Si, Fe, K, and Cl, with slight variations observed.
The abundance of microplastics
As shown in Fig. 6 (p < 0.05), the microplastics content in Guizhou’s mulched farmland ranges from 143.28 to 3,283.46 items/kg, with an average of 1,150.60 ± 647.86 items/kg. The maximum and minimum values are found to be the GD and QXG, respectively. The highest average concentration of microplastics is at site GD with 469.63 ± 437.43 items/kg, followed by site PB with an average of 1,622.82 ± 776.72 items/kg. The lowest average concentration is at site QXG with 1,835.75 ± 973.00 items/kg, and then site HP with an average of 797.98 ± 480.49 items/kg. These results indicate that microplastics are widely distributed in agricultural soils. The microplastics contents are comparable to those in Shaanxi’s agricultural soils (1430–3410 items/kg), significantly lower than in Yunnan’s planting areas (7100–42960 items/kg) and Wuhan’s vegetable farmlands (1.6 × 105 items/kg), but higher than in agricultural lands in Jiangsu (420–1290 items/kg) and Zhejiang (average 571items/kg)37,38,39,40. Additionally, this is higher compared to the average microplastics content in coastal sediments near the Yangtze River Estuary of 121 ± 9 items/kg and in Swiss lake and beach sediments of 593 items/kg26,28. Studies suggest that inland areas away from water bodies accumulate more microplastics than coastal buffer zones and marine sediments due to the hydrological effects that remove microplastics, thus soils on land are more prone to microplastics accumulatio41,42.In addition to the influence of different agricultural practices and climate factors on the distribution and abundance of microplastics in different regions, the regional characteristics of agricultural management practices (plastic mulching, irrigation, and fertilization) also contribute to the variability of microplastics pollution in farmland soils43,44,45,46.
The size distribution of microplastics
The size distribution of microplastics across different regions is depicted in Fig. 7. In Guizhou’s mulched farmlands, microplastics within the 10–100 μm range constitute the highest proportion, accounting for 64.79%, followed by the 1–10 μm range at 34.49%. Items smaller than 1 μm are the least common, making up only 0.19%, with those in the 100–1000 μm range slightly more common at 0.53%. Microplastics smaller than 1 μm are only detected at the HP and BZ sampling sites, with respective proportions of 1.83% and 0.48%. Microplastics in the 1–10 μm range have the highest proportion at the BZ site (65.14%) and the lowest at the WN site (9.90%). For microplastics in the 10–100 μm range, the highest proportion is at the WN site (90.10%), and the lowest is at the HP site (33.03%). Microplastics within the 100–1000 μm range are found at the GD, PZ, HP, BZ, and QXG sites, with the highest proportion at the GD site (65.13%) and the lowest at the BZ site (0.24%).
Microplastics of small size are prevalent across all sampling sites; however, larger items (> 1 μm) are not detected. Studies indicate that after significant weathering and degradation, the size distribution of microplastics is positively skewed, with the proportion of MPs decreasing as particle size increases47,48. This trend does not entirely align with our study. Larger microplastics tend to accumulate in surface soils, while smaller ones are more likely to be found in deeper layers49,50. This may be due to larger microplastics being more affected by external environmental factors such as wind and rainfall, leading to their migration to other environmental media48. Smaller microplastics migrate to deeper soil layers under the influence of soil biological activity and surface runoff51,52,53. Overall, larger microplastics are more susceptible to physical fragmentation, UV radiation, high temperatures, and mechanical abrasion, leading to their breakdown into smaller pieces. This, combined with the effects of climatic conditions and farming practices, results in a higher proportion of items in the 10–100 μm and 1–10 μm size ranges.
Impact of mulching duration and farming practices on microplastics distribution
The distribution of microplastics in mulched farmland in Guizhou is influenced by the duration of mulch use (Fig. 8a). Microplastics concentrations range from 143.28 to 1,325.32 items/kg for 1–6 years of mulching, 704.45 to 2,376.03 items/kg for 7–11 years, 895.49 to 2,706.37 items/kg for 12–16 years, 740.27 to 3,283.46 items/kg for 17–21 years, and 149.20 to 1,890.48 items/kg for 22–30 years, respectively. The highest average microplastics concentration is found in soils with 17–21 years of mulching, at 1,693.47 ± 1,107.45 items/kg, followed by 22–30 years with an average of 1,624.81 ± 257.26 items/kg. The lowest average concentrations are observed for 1–6 years, at 607.94 ± 356.40 items/kg, and for 7–11 years, at 1,167.78 ± 477.29 items/kg. A linear regression analysis shows a significant positive correlation between mulching duration and microplastics content (p < 0.001), with the equation y = 49.806x + 657.528 and R = 0.542 (Fig. 9). Overall, microplastics content increases with the duration of mulch use, indicating that plastic mulch contributes to significant microplastics pollution, consistent with prior studies24,46,48. Huang et al. found that microplastics content significantly increases with prolonged mulching duration, with concentrations rising from 80.3 particles/kg in soils mulched for five years to 308 particles/kg and 1,075.6 particles/kg in soils mulched for 15 and 24 years, respectively32. While plastic mulch technology is a common method to enhance agricultural yields, it also increases the concentration of microplastics in the soil, exacerbating the burden of plastic decomposition. Notably, the study observes a slight decrease in microplastics content for 22–30 years compared to 17–21 years, a phenomenon also observed in Yu’s research48.
The content and distribution of microplastics vary with different crops cultivated (Fig. 8b). Microplastics content ranges from 549.23 to 2,706.37 items/kg when vegetables are grown, 143.28 to 2,376.03 items/kg for hot peppers, 417.89 to 3,283.46 items/kg for tobacco, and 1,158.17 to 1,790.98 items/kg for potatoes, respectively. Potatoes have the highest average microplastics concentration at 1,462.63 ± 274.79 items/kg, followed by vegetables at 1,303.83 ± 585.19 items/kg. The lowest averages are found in tobacco at 1,117.57 ± 740.50 items/kg, and hot peppers at 1,053.64 ± 646.27 items/kg. Potato fields show the highest levels of microplastics, possibly due to the mechanical fragmentation of larger plastic mulch pieces during harvest and the subsequent movement of plastic obstructed by the root system or adhered to microplastics deeper into the soil with tillage40. This suggests that cultivation methods may influence the persistence and distribution of microplastics in the soil to some degree42.
In Guizhou’s plastic mulched farmlands, the irrigation method influences microplastics distribution (Fig. 8c). Contents range from 334.32 to 1,890.48 items/kg under sprinkler irrigation and 417.89 to 3,283.46 items/kg without irrigation. Furrow irrigation shows 143.28 to 1,468.60 items/kg, and drip irrigation has 441.77 to 2,706.37 items/kg. The highest average of 1,247.16 ± 609.06 items/kg is observed without irrigation, followed by drip irrigation at 1,104.93 ± 752.36 items/kg. The lowest averages are furrow irrigation at 811.91 ± 693.54 items/kg and sprinkler irrigation at 1,026.83 ± 666.03 items/kg. The microplastic content was slightly higher in the fields with non-irrigation methods and slightly lower in furrow irrigation methods. This could be due to Guizhou’s plentiful rainfall, which reduces irrigation and surface runoff, thereby exacerbating microplastics accumulation. The clean water source in furrow irrigation, with no detected microplastics pollution, facilitates hydraulic transport, thus reducing microplastics content in fields54. Microplastics distribution in cultivated soil is affected by the source, volume, and frequency of irrigation46.
Microplastics distribution varies with soil texture (Fig. 8d). In clay, loam, and sandy soils, microplastics contents are 740.27 ~ 2,706.37, 143.28 ~ 2,376.03, and 334.32 ~ 3,283.46 items/kg, with averages of 1,347.88 ± 1,054.39, 1,327.31 ± 618.55, and 1,050.71 ± 543.68 items/kg, respectively. Yu et al. found that sandy soils exhibit higher microplastics levels than silty loams and loams, aligning with the findings of the present study48. Sandy soils, with larger particles and pores, contrast with the smaller particles and pore spaces in clay and loam, affecting leaching processes and thus influencing microplastics transport and accumulation52,53. Contrarily, Scheurer and Bigalke’s findings suggest no significant correlation between MPs abundance and soil texture, possibly due to a variety of factors affecting MPs accumulation in soils, such as diverse sources like agricultural plastic waste, soil amendments, and irrigation practices in different regions26. Huang’s study shows no apparent pattern or correlation between soil chemical properties and microplastics content, indicating that soil properties are not a key factor in determining microplastics abundance35. The impact of soil texture and properties on MPs abundance, migration, and transformation requires further investigation.
Principal component analysis (PCA) of microplastics content and environmental factors (Fig. 10) identified four main components: the first accounting for 20.64%, the second for 17.65%, the third for 14.96%, and the fourth for 11.20%, cumulatively contributing 64.36%. Factors such as mulching duration, crop type, irrigation method, and soil texture, along with climate and topography, are significant and influence microplastics content and spatial distribution to some extent. In Fig. 10a, the arrows for VE and DI point in similar directions, indicating that these factors exhibit similar variation trends across the samples. In Fig. 10b, the clustering of sample points such as PB1, PB2, PB3, PB4, and PZ1 suggests that these locations are influenced by similar environmental factors affecting microplastics distribution. Conversely, sample points like QXG1 and GD3 are more isolated and distant from other groups, indicating significant differences in their characteristics. Guizhou’s ___location on the Yunnan-Guizhou Plateau with high solar radiation, intense ultraviolet (UV) rays, and relatively high wind speeds are also influential. UV radiation and weathering erosion are major factors in microplastics aging55. Plastics exposed to the environment absorb high-energy wavelengths of UV radiation, inducing photochemical reactions, followed by oxidation, which increases brittleness and reduces elasticity, leading to smaller plastics upon thermal degradation55,56. Previous studies indicate that weathering can cause surface color changes, cracking, and disintegration of plastic items57,58,59. Considering the combined effects of sunlight, oxidants, and physical stress, Guizhou’s climate conditions may promote the decomposition and fragmentation of plastic polymers, making microplastics formation more likely in its agricultural production42,57.
Principal Component Analysis (PCA) of the influence of agricultural environmental factors on microplastic abundance (a and b). LP, MA, CL, VE, DI, NI, SI, FI, CH, SA, LO, TO, and PO represent mulching years, microplastic abundance, clay, vegetables, drip irrigation, non- irrigation, sprinkler irrigation, furrow irrigation, chili, sandy soil, loamy soil, tobacco, and potato, respectively.
Conclusion
In the mulched agricultural soils of Guizhou, polyethylene is the predominant microplastics component, mainly appearing as black, transparent, and translucent irregular fragments ranging from 40 to 120 μm in diameter. These microplastics have rough surfaces with evident signs of erosion and are rich in oxygen-containing (C-O-C/-OH) and halogen functional groups. Quantitative analysis reveals a microplastics concentration of 143.28 to 3,283.46 items/kg, averaging 1,150.60 ± 647.86 items/kg. Items sized 10 to 100 μm constitute the majority at 64.79%, followed by those 1 to 10 μm. There is a highly significant positive correlation between microplastics abundance and years of mulch use (p < 0.001). The highest average concentration of microplastics is found in potato crops, at 1,462.63 ± 274.79 items/kg, followed by vegetables, chili peppers, and tobacco. Soils without irrigation display the highest microplastics average (1,247.16 ± 609.06 items/kg), followed by drip and furrow irrigation methods. Clay soil has the highest microplastics content, followed by loam and sand. Principal component analysis indicates that in addition to mulching duration, crop type, irrigation method, and soil texture, climate, and topography are significant factors that affect microplastics content and spatial distribution to a certain extent.
Data availability
The study’s original contributions are detailed in the article/supplementary material; any further questions should be directed to the corresponding author.
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Funding
This work was supported through financial support from National Natural Science Foundation of China (Grant No. 42167067), Department of Education of Guizhou Province (No. QianJiaoJi[2023]034), the Key Program for Science and Technology of CNTC (No.110202202030), the Science and Technology Program of Guizhou Provincial Branch of the CNTC (Nos.2023-13,2022520500240192) and the Scientific Research Platform of Guizhou Minzu University (GZMUGCZX [2021]02).
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M.H.Y.: Conceptualization, Methodology, Validation, Formal Analysis, Investigation, and Writing–Original Draft. Z.S.Y.: Methodology, Validation, Formal Analysis, Investigation, and Writing–Original Draft. G.W.C.: Supervision, Writing – Review & Editing. Z.J.Y: Formal Analysis, and Investigation.C.H.Q: Formal Analysis, and Investigation.W.L.J: Investigation. L.J.C: Investigation. W.Z.H: Supervision, Writing – Review & Editing. L.T.Z.: Supervision, Writing – Review & Editing. All authors have read and agreed to the published version of the manuscript.
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Miao, H., Zhang, S., Gao, W. et al. Microplastics occurrence and distribution characteristics in mulched agricultural soils of Guizhou province. Sci Rep 14, 21505 (2024). https://doi.org/10.1038/s41598-024-72829-7
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DOI: https://doi.org/10.1038/s41598-024-72829-7
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