Arbuscular mycorrhizal fungi enhance maize cadmium resistance and reduce translocation: Dependence on microplastics concentration

Arbuscular mycorrhizal fungi enhance maize cadmium resistance and reduce translocation: Dependence on microplastics concentration | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Arbuscular mycorrhizal fungi enhance maize cadmium resistance and reduce translocation: Dependence on microplastics concentration Yi Lin, Xiaoli Sun, Jiping Chen This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8569352/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 12 You are reading this latest preprint version Abstract Background The coexistence of microplastics (MPs) and heavy metals (HMs) such as cadmium (Cd) in agricultural soils represents a growing threat to crop production and food security. While arbuscular mycorrhizal fungi (AMF) are recognized for their ability to enhance plant metal tolerance, their role in mediating crop responses under combined contamination with MPs and Cd, especially across different MPs concentrations, remains largely unexplored. This study was conducted to elucidate how AMF modulate maize growth, Cd accumulation, and soil biogeochemical processes under co-contamination with polyethylene (PE) (0, 0.5% and 5% w/w) and Cd (0, 20 mg kg − 1 ) with or without AMF. Results The addition of 5% PE-MPs significantly aggravated Cd toxicity in maize, elevating Cd translocation to shoots by 79.6% and causing severe growth suppression. PE-MPs also modified key soil characteristics, increasing organic matter content and pH, which promoted the transformation of Cd into less bioavailable fractions yet failed to counteract its direct phytotoxic effects. Inoculation with AMF markedly alleviated these stresses. Under Cd and 5% PE-MPs co‑contamination, mycorrhizal plants showed 87.5% higher shoot biomass, 39.6% greater phosphorus uptake, and 38.5% enhanced net photosynthesis compared to non‑inoculated plants. AMF further reduced oxidative damage, promoted Cd sequestration in cell walls, decreased the biologically active Cd pool in shoots, and lowered Cd bioavailability through shifts in soil bacterial community composition, particularly by restoring the abundance of Pseudomonadota . The beneficial effects of AMF were more evident at 0.5% PE-MPs than at the 5% concentration. Conclusions This study demonstrates that AMF confer dual protection in a PE-MPs concentration-dependent manner. AMF enhance plant physiological resilience by regulating antioxidant systems and Cd subcellular distribution, while reducing Cd bioavailability by modifying soil properties, soil bacterial diversity and Cd speciation. These phytoremediation combined PE-MPs + Cd pollution synergistic toxicity soil bacterial community heavy metals bioavailability Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 1. Background Soil contamination by heavy metals (HMs), particularly cadmium (Cd), has evolved into a critical global environmental crisis, severely endangering food security and ecosystem integrity 1 . Hou et al. 2 reported that approximately 14–17% of global croplands are contaminated by HMs, among which Cd exhibits the highest exceedance rate at 9%. Concurrently, microplastics (MPs), polymer-based particles with a diameter < 5 mm, have become ubiquitous pollutants posing a persistent anthropogenic challenge across terrestrial and aquatic ecosystems 3 . Global plastic production now exceeds 430 million tons annually with 10 to 40 million metric tons of MPs emitted into the environment each year 4 . A global soil survey 5 documented MPs concentrations ranging from 0 to 10 4 mg kg − 1 , with polyethylene (PE) identified as the most predominant type. In China, approximately 14.7% of agricultural soils are at risk of MPs pollution, with abundances varying from 7 to 3.61 × 10 5 items kg − 1 6 . MPs can enter plants through both foliar deposition and root uptake, and their presence has been confirmed in various food crops 7 . Alarmingly, MPs have also been detected in multiple human tissues and organs, with accumulating evidence indicating potential adverse impacts on human health 4 . Notably, the spatial distribution of MPs pollution shows remarkable overlap with HMs-contaminated areas 8 , suggesting their combined effects warrant thorough investigation. Given their high toxicity and strong bioaccumulation potential in the food chain, the co-occurrence of Cd and MPs in agricultural soils is of particular environmental concern, underscoring the urgent need for effective mitigation and remediation strategies 9 , 10 . MPs possess high surface reactivity and a large specific surface area, enabling them to act as carriers for HMs and potentially alter their environmental behavior 11 . Additionally, under HMs stress, MPs have been shown to induce the activation of the plant antioxidant defense system 12 . Zhao et al. 13 highlighted the potential of MPs to modulate the phytoremediation efficiency of Cd-contaminated soils. Despite the growing body of research investigating the combined effects of MPs and HMs co-pollution on plants, inconsistencies persist regarding whether their interactions exhibit synergistic, antagonistic, or neutral effects 14 – 16 . Specifically, Li et al. 15 reported that PE-MPs reduced the bioavailability of Cd in soil, thereby decreasing Cd uptake by pakchoi, without exerting significant impacts on plant biomass. In contrast, Zhao et al. 16 demonstrated that MPs increased the concentration of bioavailable Cd in soil and promoted Cd accumulation in maize. Similarly, Gan et al. 17 found that PE-MPs inhibited plant growth while enhancing Cd uptake, whereas another study documented a positive effect of MPs on tomato growth under Cd stress 18 . A recent meta-analysis further clarified that MPs tend to lower soil pH, which in turn increases the concentration of bioavailable Cd in soil and enhances Cd accumulation in plant shoots by 11.0% 19 . The complex interactions between MPs and HMs are further compounded by multiple influencing factors, including contaminant types, concentrations, MPs particle size, and inherent soil physicochemical properties 20 . Notably, most existing studies focusing on the toxicological effects of MPs and Cd on plants have primarily centered on plant physiological responses to external stressors, while frequently neglecting the pivotal regulatory role of soil microorganisms in mediating these biotic-abiotic interactions. Arbuscular mycorrhizal fungi (AMF) have emerged as a powerful biological tool for phytoremediation of contaminated soils. Extensive research, including our prior investigations 21 , 22 , has demonstrated that AMF inoculation significantly enhance phytoremediation efficiency in HMs-polluted soils through multiple mechanisms 23 . While the interactions between AMF and MPs have begun to attract scientific attention 24 – 26 , current understanding remains limited. AMF are widely recognized for mitigating pollutant toxicity in plants. However, Giambalvo et al. 24 observed that AMF reduced maize shoot biomass by an average of 7% in soils contaminated with MPs. Notably, the impact of AMF on plant tolerance to MPs is influenced by the size of the MPs 27 . Although existing studies have predominantly examined the individual effects of MPs or HMs in AMF-plant systems, emerging evidence suggests complex interactions among these factors. MPs may exacerbate the risk of HMs in the AMF-plant system 25 . AMF can alleviate the combined phytotoxicity of MPs and Cd to plants 28 , potentially by reducing the translocation of MPs to shoots 29 . Both MPs and AMF can significantly alter soil microbial communities and physicochemical properties, creating a dynamic rhizosphere environment that modulates contaminant bioavailability 30 , 31 . Nevertheless, the tripartite interactions among AMF, MPs, and HMs in co-contaminated soils remain poorly characterized. This is particularly true regarding their combined effects on plant growth, metal accumulation dynamics, and stress tolerance mechanisms. This critical knowledge gap underscores the urgency of comprehensive studies to decipher how AMF modulate plant adaptive responses within this MPs-HMs co-contaminated rhizosphere environment. Maize ( Zea mays L.), a globally critical staple crop that feeds nearly one-third of the world’s population, serves as an ideal model for this investigation owing to its rapid growth, high biomass, and documented tolerance to both HMs and MPs 32 , 33 . While the interactive effects between Cd and MPs in soil-maize systems have gained increasing attention, the regulatory mechanisms underlying AMF-mediated plant adaptation to co-exposure stresses remain largely elusive. Within the soil-AMF-plant system, it is therefore imperative to clarify how MPs modulate Cd translocation. Furthermore, soil MPs concentrations may alter the capacity of AMF to immobilize Cd. Consequently, elucidating the transport and immobilization processes of Cd at the soil-AMF-plant interface under varying MPs contamination levels is critical for developing effective remediation strategies. Notably, a significant knowledge gap persists regarding how different MPs concentrations regulate Cd dynamics in soil-AMF-crop systems. The present study investigates the effects of AMF inoculation on maize cultivated in sterilized soil under three distinct PE-MPs concentration gradients, with a specific focus on Cd speciation, translocation, and phytotoxicity. We hypothesize that PE-MPs concentration gradients will exert divergent impacts on Cd bioavailability and plant physiological responses in AMF-colonized systems. The specific objectives are to: (1) unravel the mechanisms governing Cd bioaccumulation and phytotoxicity in soil-maize systems under AMF mediation; (2) evaluate the regulatory role of AMF in maize growth parameters and Cd distribution patterns within plant tissues; and (3) quantify the concentration-dependent effects of PE-MPs on Cd uptake by maize and the associated phytotoxicological consequences. 2. Materials and methods 2.1. Soil, seeds, MPs and AMF inoculum Soil samples (0–20 cm depth) were collected from an agricultural field in Lishui City, Zhejiang Province, China (119°45’E, 28°02’N). After air-drying (14-day) and sieving (2 mm) for use. Soil properties were analyzed: pH (6.3), total nitrogen (2.90 g kg − 1 ), organic matter (52.63 g kg − 1 ), total phosphorus (665.0 mg kg − 1 ), available phosphorus (13.82 mg kg − 1 ), and background Cd (0.23 mg kg − 1 ). PE-MPs (100 µm) were procured from Telang Plastic Chemical Co., Ltd. (Dongguan, China) and pre-cleaned with 0.1 M HCl followed by deionized water rinsing. Maize seeds (Zhenuoyu 16, Cd-tolerant cultivar) were obtained from Zhejiang Kecheng Seed Industry Co., Ltd. The AMF Rhizophagus intraradices (BGC BJ09) was provided by the Institute of Plant Nutrition and Resources, Beijing Academy of Agriculture and Forestry. The inoculum contained spores (80 spores g − 1 soil), hyphae, root fragments, and sandy soil. 2.2. Experimental design and greenhouse setup Soil was spiked with CdCl₂ (0 and 20 mg Cd kg − 1 ) and equilibrated for two weeks. Three PE-MPs treatments were applied: 0% (PE0), 0.5% (PE0.5), and 5% (PE5) by dry soil weight. The 0.5% dose is commonly used in experiments involving MPs 10 , 34 . The 5% MPs concentration used in this study simulates extreme long-term accumulation scenarios. This level is particularly relevant to severely impacted areas like industrial mulch sites, where concentrations can reach up to 6.7% 35 . Employing such an elevated exposure aligns with established ecotoxicological approaches 17 , 33 , 36 and aids in identifying critical risk thresholds and elucidating underlying toxicological mechanisms. Although this concentration may exceed typical near-term environmental exposures, it is methodologically justified within ecotoxicology for revealing mechanistic tipping points and projecting long-term cumulative effects. Our experimental design thus establishes a continuous dose-response relationship, spanning from environmentally realistic levels to extreme concentrations. This range is essential for comprehensively assessing the ecological impacts of microplastics, particularly under conditions of combined pollution. Each pot (16 cm height × 17 cm diameter) contained 2.0 kg dry soil. The AMF treatment received 30 g of R. intraradices inoculum, while the non-mycorrhizal control was amended with autoclaved inoculum and 5 mL of filtered (15-µm) microbial washings to maintain comparable microbiota. Maize seeds were sequentially washed with deionized water and surface-sterilized in 2% (v/v) H₂O₂ for 12 h, followed by thorough rinsing with deionized water. Sterilized seeds were placed on moist absorbent cotton in Petri dishes and germinated at 25°C. Upon reaching 3–5 cm root length, seedlings were transplanted into pots. After six days of acclimation, two uniform seedlings were retained per pot. Soil moisture was maintained at 15–18% through daily irrigation with deionized water. Environmental conditions were controlled at 23 ± 3°C air temperature and 45 ± 5% relative humidity throughout the 40-day experimental period (January 21 to March 1, 2025). The experiment employed a fully factorial design with three replicates per treatment, resulting in 12 treatments across 36 pots. Detailed treatment combinations are listed in Supplementary Table S1 . 2.3. Plant growth and physiological measurements 2.3.1. Biomass and photosynthesis Shoots and roots were washed thoroughly with deionized water, and oven-dried (65°C, 48 h) for dry weight (DW) determination. Fresh leaves were homogenized with 95% ethanol at 4°C and a spectrophotometer (P4, Mapada, China) was used to measure the chlorophyll (Chl) content. The Chl levels (mg g − 1 FW) were calculated as follows 37 : Chl a = 12.7 × A663 − 2.69 × A645; Chl b = 22.9 × A645 − 4.68 × A663. Net photosynthetic rates (Pn) was measured between 9:00 and 11:00 am using a photosynthetic instrument (CIRAS-4, PP-Systems, Hitchin, UK) at 1200 µmol m − 2 s − 1 light intensity and 400 µmol mol − 1 CO 2 . 2.3.2. Elemental analysis Dried tissues were microwave-digested (HNO 3 :HClO 4 , 3:1 v/v), and Cd/P concentrations were determined via ICP-MS (iCAP RQ, Thermo Fisher Scientific, USA). Quality control included certified reference materials (National Center of Analysis and Testing, Beijing). The detection limits of Cd and P were 0.05 µg L − 1 . 2.4. Oxidative stress and cellular damage assays Fresh leaves were homogenized in PBS (0.05 M, pH 7.0) and to prepare a supernatant. The MDA, SOD, POD, and CAT levels were determined using assay kits (Nanjing Jiancheng Bioengineering Institute, China) according to the manufacturer’s protocols 38 . To determine cell death, leaves were immersed in a 0.25% Evans blue solution for 24 h and then boiled in 95% ethanol for 30 min to remove chlorophyll 39 . 2.5. Subcellular distribution, chemical forms of Cd Subcellular distribution analysis of Cd was conducted following the method described by previous study 40 . The samples were separated into different cellular fractions, including cell wall fraction, cell organelle fraction, and soluble fractions, using differential centrifugation at 4°C. Detailed extraction procedures can be found in Supplementary Text S1. The Cd content in the subcellular fractions was determined using ICP-MS. The chemical forms of Cd in the maize were determined using stepwise extraction 41 (Supplementary Text S2). 2.6. Soil chemical and enzymatic analyses 2.6.1. Cd fractions Root-attached soil particles were carefully separated by gentle shaking to obtain rhizosphere soil samples. The collected soil was homogenized by thorough mixing and divided into two aliquots. One portion was immediately stored at -80°C for subsequent soil microbial community analysis. The other portion was air-dried for two weeks at room temperature (20–25°C) and sieved (2 mm mesh) for physicochemical characterization. Soils (0.500 g ± 0.003) were digested with HCl-HNO 3 (3:1 v/v). The procedure of microwave digestion was as follows: 120°C for 10 mins, 150°C for 10 mins and 190°C for 40 mins. The three-step BCR sequential extraction procedure was employed to extract the different Cd fractions from the soil, including acid-soluble Cd, reducible Cd, oxidizable Cd, and residual Cd (Supplementary Table S2) 42 . The residual fraction was obtained by digesting the final residues from step 3 with concentrated HCl-HNO 3 (3:1, v/v). Bioavailable Cd was extracted using 0.005 M DTPA (pH 7.3) 43 . All Cd concentrations were quantified by ICP-MS (iCAP RQ, Thermo Fisher Scientific, USA). 2.6.2. Soil physicochemical properties Soil AP was measured using 0.5 M NaHCO 3 (pH 8.5) with the Mo–Sb anti-spectrophotometric method 44 . Soil OM was measured according to the potassium dichromate oxidation method 44 . Soil pH was measured with a pH meter (LC-PH-3S, Shanghai, China) at a water to soil ratio of 2.5:1 (w/v). Soil enzyme activities were quantified using standardized colorimetric assays following Wang et al. 45 with commercial test kits (Nanjing Jiancheng Bioengineering Institute, China). Three key enzymes were analyzed. Urease activity was determined by measuring the changes in absorbance at 578 nm using a spectrophotometer (T6-New Century; Beijing Purkinje General Instrument Co., Ltd., China) and reported in µg NH 3 –N g − 1 24h − 1 at 37°C. Acid phosphatase activity was determined by measuring the changes in absorbance at 400 nm and reported in µmol PNP g − 1 24h − 1 at 37°C. The CAT activity was determined by measuring changes in the absorbance at 240 nm and reported in µmol H 2 O 2 g − 1 h − 1 . 2.7. Soil Bacterial community analysis Rhizosphere soil DNA was extracted using the E.Z.N.A.® soil DNA Kit (OMEGA, USA), with three independent biological replicates of soil samples processed per treatment. The quality of the extracted DNA was evaluated via 1% agarose gel electrophoresis. Microbial community analysis was conducted by Shanghai Majorbio Bio-pharm Technology Co., Ltd. The V3–V4 hypervariable region of the bacterial 16S rRNA gene was amplified using the primers 338F (5’-ACTCCTACGGGAGGCAGCAG-3’) and 806R (5’-GGACTACHVGGGTW TCTAAT-3’). PCR amplification was performed following the protocol described by Xu et al. 30 . 2.8. Statistical analysis Data (mean ± standard deviation, n = 3) were analyzed using SPSS 26.0 (IBM, Armonk, NY, USA), and figures were generated with Origin 2024 (Northampton, MA, USA). A one-way analysis of variance (ANOVA) was conducted to compare significant differences among treatments at the same Cd level, with a significance threshold set at p < 0.05 using Duncan’s test. For assessing the effects of Cd, PE, AMF inoculation, and their interactions on plant growth parameters and soil-related experimental indices, two-way or three-way ANOVA was employed. Prior to ANOVA analysis, the normality and homogeneity of variance of the data were verified using the Shapiro-Wilk test. 3. Results and discussion 3.1 MPs and Cd effects on maize growth traits are modified by AMF Although the individual toxic effects of MPs and Cd on plants are well-documented, their combined impacts remain poorly understood. Cd, PE, and AMF significantly influenced the growth indices of maize (Table S3). As shown in Fig. 1 a, maize growth status at 30-day intervals demonstrates significant improvement with AMF inoculation. Quantitative analysis reveals that Cd exposure alone reduced maize dry weight by 46.0% in shoots and 37.1% in roots (Fig. 1 b). In Cd-free soil, PE exerted concentration-dependent physiological effects. Although shoot biomass remained unaffected, application of 5% PE significantly enhanced root biomass, achieving a 41.5% increase relative to the control. The three-way ANOVA revealed that root biomass was influenced by soil Cd and PE concentrations (Table S3). Notably, AMF inoculation significantly alleviated the combined toxicity of MPs and Cd, increasing shoot biomass by 87.5% and root biomass by 32.5% in the Cd+PE5 treatment ( p < 0.05; Fig. 1 b). The 5% PE addition exacerbated Cd toxicity, resulting in a 29.6% reduction in shoot biomass compared to Cd treatment alone. Across all treatments, AMF application enhanced maize growth, increasing shoot and root dry weight by 63.8% and 32.5%, respectively. The most pronounced growth promotion occurred in low dose PE (0.5%) with AMF, showing 17.9% and 41.3% increases in shoot biomass relative to PE5-Cd and PE0-Cd treatments. PE addition significantly altered biomass allocation, with 5% PE increasing the root-to-shoot ratio by 67.4% under Cd-free conditions and by 59.5% under Cd20 stress, whereas 0.5% PE reduced it by 18.9% under Cd exposure (Fig. 1 c). AMF-treated plants consistently exhibited lower root-to-shoot ratio, indicating preferential shoot growth promotion. Figure 1 d showed that more root at PE5 and AMF mostly. The findings demonstrate a concentration-dependent response to PE, inhibitory at high level and stimulatory at low level, which supports the potential of maize for remediating moderately contaminated soils. Photosynthetic analysis revealed that Cd stress dramatically reduced Pn, while PE alone showed negligible effects (Fig. 1 e). AMF inoculation alleviated photosynthetic inhibition under stress conditions, increasing Pn by 20.9% under Cd and 38.5% under Cd+PE5, whereas the Cd+PE5 treatment itself decreased chlorophyll content and Pn by 41.0% and 51.6%, respectively, relative to the control (Fig. 1 f). A study indicated MPs can affect photosynthetic ability and decrease the photosynthetic pigment content 46 . Under Cd stress, 10% PE-MPs inhibited Pn of maize 32 . The impact of MPs on plant growth is dual, potentially promoting or inhibiting growth, or having no noticeable effect. The effects vary across different plant organs and are influenced by the concentration and type of MPs, as well as the plant species and its growth environment 47 . Xiang et al. 48 found that PE had no effect on shoot biomass, but increased root biomass through a metal-analysis. Liu et al. 49 found that adding 0.2% polyester to Cd-polluted soils increased rice biomass, whereas 0.2% polyethylene terephthalate had no effect on biomass. The combined toxicity of Pb and MPs to plants is more evident than when they are present individually 50 . Although the mitigating or exacerbating effects of PE on plants in metal-polluted conditions have been documented, no studies have investigated the responses of the AMF-crop system to combined MPs and Cd stress. The toxicity of HMs may be amplified when combined with PE. Liu et al. 51 reported that addition of PE increased Cd toxicity in sorghum, as evidenced by a reduction in length and biomass. MPs can affect the biomass allocation and tend to increase root to shoot biomass ration 52 . A meta-analysis pointed out that low levels MPs promotion shoot and high levels promotion root in order to resist stress 53 . AMF enhanced phosphorus uptake, particularly at high PE concentration (Fig. 1 g), corresponding with observed growth improvements. PE and Cd damaged the photosynthetic system to different degrees, causing leaf cell more damaged (Fig. 1 h). These physiological changes were paired with distinct morphological alterations in both Cd-free and Cd-polluted soils, underscoring the intricate interactions between PE-MPs, AMF, and Cd stress. 3.2 Coexisting AMF and MPs influenced the uptake and transfer of Cd in plants, and exhibited a “dose effect” MPs concentration was one of the main factors affecting toxic elements accumulation in plants 54 . Shoot and root Cd concentrations were significantly influenced by Cd, PE, and their interaction, while the AMF × Cd interaction also significantly affected shoot Cd concentration (Table S3). Without Cd treatments, the concentration of Cd in shoots and roots remained stable and was unaffected by the presence of PE or AMF. Our results demonstrated that Cd accumulation in roots was significantly higher than in shoots, as evidenced by TF < 1 (Fig. 2 ). Maize, as an excluder plant (TF < 1), has been shown to primarily accumulate Cd in its roots, effectively immobilizing it and limiting its translocation to the aerial parts 55 . PE treatment significantly reduced Cd uptake in maize roots by 21.3–35.7% in non-inoculated plants and by 12.6–21.2% in AMF-treated plants. Interestingly, shoot Cd concentrations in AMF-treated maize remained stable regardless of PE addition. However, in non-AMF treatments, shoot Cd levels in the PE5 group were 20.4% higher than in the PE0.5 group. For most crops, AMF has been shown to inhibit the uptake of metals to prevent damage. Kuang et al. 56 reported that AMF decreased TF of Cd in maize. The effect of AMF on Cd translocation under MPs co-exposure was Cd-level dependent, significantly reducing the TF at 40 mg kg − 1 Cd but showing no effect at 80 mg kg − 1 Cd 29 . AMF inoculation alone decreased root Cd uptake in PE0 treatment by 14.4%, while PE addition attenuated this beneficial effect. In both AMF-inoculated and non-inoculated plants, the TF of Cd in maize exhibited a dose-dependent decrease with increasing PE concentration. Although PE generally enhanced Cd translocation to shoots by reducing root absorption, AMF inoculation effectively counteracted this phenomenon. The reduced Cd uptake in AMF-treated plants may be attributed to a dilution effect resulting from enhanced plant growth. Our previous research demonstrated a strong correlation between plant Cd accumulation and soil bioavailable Cd content 21 . The current study found that MPs reduced Cd bioavailability, a finding consistent with prior reports that 0.25 mg kg − 1 PE decreased Cd bioaccessibility by 13.3% 51 . These results suggested that MPs influenced plant Cd concentrations primarily by modifying soil physicochemical and microbial properties. Previous research indicated that MPs affected Cd absorption in a dose-dependent manner, with higher MPs concentrations promoting Cd accumulation in both shoots and roots 57 . 3.3. Antioxidant response of maize Both PE alone and the interaction between Cd and AMF significantly influenced all enzyme activities (Table S3). As shown in Fig. 3 a-d, combined contamination with Cd and PE elicits complex responses in the antioxidant enzyme system and promotes membrane lipid peroxidation in maize. In contrast, AMF inoculation alleviates oxidative damage under co-contamination conditions by modulating the antioxidant defense response. Under Cd-free conditions, SOD activity was enhanced by 5% PE but remained unaffected by AMF except in the absence of PE. In Cd-stressed soil, 5% PE increased SOD activity, while AMF inoculation suppressed this PE-induced enhancement. With respect to POD activity, the addition of PE (both 0.5% and 5%) tended to reduce it in Cd-free soil, whereas no significant effect was observed in Cd20 soil. Notably, AMF consistently up-regulated POD activity across all Cd and PE treatments. The CAT activity exhibited concentration- and condition-dependent behavior. In the absence of Cd, 0.5% PE significantly increased CAT activity regardless of AMF presence, whereas high-concentration PE showed no stimulatory effect. Under Cd20 conditions without AMF, both low and high PE concentrations tended to enhance CAT activity. In contrast, under Cd stress, AMF inoculation in combination with low PE reduced CAT activity. Unlike previous findings in Solanum nigrum 37 , where Cd stress elevated the activities of SOD, POD, and CAT, the present results demonstrate that Cd specifically induced POD and CAT activities, while SOD remained largely unaltered. The generation of reactive oxygen species (ROS), during oxidative bursts is a key mechanism underlying the toxic effects of Cd and MPs on plants 41 . The antioxidant system counteracts such damage primarily via a sequential enzymatic process involving SOD, which converts superoxide into H₂O₂, followed by CAT and POD, which catalyze the decomposition of H₂O₂ 37, 38 . By scavenging ROS, these enzymes help alleviate Cd phytotoxicity and limit its uptake and accumulation. Previous evidence shows that AMF colonization can significantly enhance enzyme activity under Cd stress 58 . The previous study demonstrated that increased Cd content in plants can boost antioxidant enzyme activity 37 . This study’s polynomial regression analysis identified a significant positive correlation between SOD activity and Cd accumulation in the aboveground plant parts, suggesting a nonlinear relationship (Fig. 3 e). However, CAT activity and POD levels showed no correlation with Cd content. The degree of membrane lipid peroxidation was assessed by measuring MDA. The levels of MDA in maize showed a positive correlation with shoot Cd concentration (Fig. 3 f). Importantly, AMF effectively reduced MDA levels under all treatment conditions, indicating alleviated oxidative membrane damage. The elevated MDA content induced by Cd was further exacerbated by high-concentration PE, highlighting the synergistic toxicity of combined pollutants. The modulation of antioxidant enzymes suggests that AMF enhances ROS scavenging capacity, particularly through sustained up-regulation of POD activity. The distinct responses of SOD and CAT indicate a shift in redox management strategy under mycorrhizal symbiosis. The reduction in MDA underscores the protective role of AMF at the cellular level, likely mediated through enhanced Cd immobilization and diminished ROS production 58 . Overall, AMF inoculation improves maize tolerance to combined Cd-PE stress by fine-tuning the antioxidant system and maintaining membrane integrity. 3.4. MPs and AMF effects on subcellular distribution and chemical forms of Cd are varying The subcellular distribution and chemical forms of HMs play a pivotal role in plant metals tolerance and detoxification through compartmentalization and chelation mechanisms 22 , 58 . The partitioning of metals within plant subcellular compartments serves as a critical determinant of their intracellular toxicity. Plants have evolved multiple strategies to mitigate metal toxicity, including chelation, vacuolar sequestration, and root immobilization to restrict translocation to aerial tissues 59 . Our findings demonstrate that AMF inoculation significantly reduced Cd content across all subcellular fractions in shoots (Fig. 4 a). The observed distribution pattern in maize shoots followed: cell wall fraction (F CW , 47.7–61.5%) > soluble fraction (F S , 25.0–31.4%) > organelle fraction (F O , 10.4–27.3%) (Fig. 4 b). This result suggests that cell wall deposition and vacuolar compartmentalization constitute primary detoxification pathways in maize, which is consistent with the response observed in Populus yunnanensis under Cd stress 40 . These results corroborate previous studies identifying F CW as the predominant Cd form in maize shoots where cell wall binding serves as a key mechanism for Cd toxicity alleviation 38 , 55 . In this study, the AMF-mediated subcellular redistribution exhibited PE concentration dependence. AMF increased F S (+ 21.7%) at PE0 while AMF increased F CW (+ 26.0%) at PE5. At low dose of PE, there was an increase in F CW and a decrease in F O , regardless of the presence of AMF. High dose PE increased F O (+ 35.3%), while AMF can offset that. At PE0.5 treatment, more Cd accumulated in F CW , thereby reducing its harmful effects. We observed a distinctive AMF-induced modification in leaf Cd partitioning, characterized by preferential accumulation in cell wall-integrated forms. While our previous research confirmed AMF’s capacity to enhance F CW 22 , elevated PE concentrations attenuated this protective effect. Notably, the F O fraction demonstrated an inverse relationship with F CW , decreasing with AMF colonization but increasing with PE addition. These findings align with Zhang et al.’s 55 report of AMF-enhanced F CW in maize roots. The increase in F O proportion induced by high-dose PE contributed to the exacerbation of Cd phytotoxicity. Chemical speciation analysis revealed substantial AMF-induced modifications under combined PE-Cd stress (Fig. 4 c and d). F E and F W exhibit strong migration abilities, while F NaCl has a lower migration ability and toxicity, and F HAc , F HCl , and F R show the weakest toxicity to plants. The distribution of Cd forms followed: F NaCl (31.9–51.0%) > F E (16.9–29.7%) > F W (18.8–26.8%) > F HAc (4.1–16.1%) > F HCl (1.7–3.5%), with F R below 0.5%. This pattern is consistent with established literature documenting F E , F W , and F NaCl as predominant Cd forms in maize shoots 38 . The phytotoxicity of Cd is primarily governed by its biological activity and chemical speciation within plant tissues. Our findings demonstrate that high dose PE (5%) significantly increased the proportions of F E (+ 20.0%) and F W (+ 28.2%), consequently exacerbating Cd toxicity. In contrast, AMF inoculation effectively mitigated Cd toxicity by reducing the fractions of biologically active Cd, with average reductions of 26.5% in F E and 12.1% in F W . Notably, AMF consistently promoted the transformation of Cd into F HAc across all PE treatment levels, while only enhancing F NaCl (+ 13.6%) at 0.5% PE. These findings are consistent with prior research, which indicates that AMF increases the distribution of Cd in inactive forms while reducing its toxic proportions 58 , 60 . The differential effects of AMF on Cd chemical speciation provide mechanistic insights into its protective role against Cd toxicity in plants. 3.5. Interactive effects of MPs and AMF on Cd speciation and bioavailability in rhizosphere soil The chemical speciation of HMs fundamentally governs their bioavailability, mobility, and ecotoxicity. Both AMF and MPs can modify soil physicochemical properties and microbial communities, thereby mediating the transformation of trace metal speciation in soil systems 61 . However, the mechanisms underlying MPs-induced alterations in HMs distribution remain poorly characterized 62 . Figure 5 a shows soil Cd forms after 40-day of treatment with or without MPs and AMF. Consistent Cd fractionation patterns across treatments: acid soluble (56.1–61.3%) > reducible (30.5–39.8%) > oxidizable (3.5–11.0%) > residual (0.5–1.6%) (Fig. 5 b). Both with and without AMF inoculation, high-dose PE consistently reduced acid-soluble Cd while increasing the oxidizable fraction content. In contrast, low-dose PE enhanced the reducible Cd content. AMF inoculation preferentially decreased reducible Cd (12.0–18.0%) and increased oxidizable forms content (43.0–89.8%), with minimal effects on acid-soluble Cd. The highest residual Cd was observed in the PE5 + AMF treatment. As there was no significant difference in the final residual Cd content among all treatments (19.57 ± 0.62 mg kg − 1 ), the percentage changes in Cd forms in the soils mirrored the changes in Cd concentration, except for the high dose of PE, which had no significant effect on the acid-soluble Cd percentage. It was reported that MPs dose affected Cd bioavailability and phytotoxicity 17 . The results demonstrated that high-dose PE significantly increased the proportions of oxidizable Cd forms and decreased acid soluble Cd, indicating PE facilitate the transformation from active to organically-bound species. This transformation may be attributed to PE-induced increases in soil pH and OM content, coupled with the inherent adsorptive capacity of insoluble MPs 10 , 63 . However, Chen et al. 64 reported that MPs increased Pb availability with reducible Pb increased by 6–12%. MPs enhanced Cd availability in wheat rhizosphere by reduced soil pH and 5% PE more effectively increased bioavailable Cd than 1% PE 17 . The addition of MPs facilitates the conversion of inactive Cd into active Cd 19 . PE at low dose can decrease the concentration of available Cd, thus reducing environmental risk. Research shows that although MPs usually promote metal fixation through organic complexation, their impact on metal migration depends on environmental conditions and the properties of the MPs. AMF inoculation further reduced bioavailable Cd in the maize rhizosphere by facilitating the formation of insoluble Cd compounds via three key mechanisms 29 , 40 , 65 : (1) secretion of metal-chelating molecules (e.g., metallothioneins, glutathione, and phytochelatins); (2) complexation of Cd on the surface of AMF hyphae; (3) enhancement of microbial-mediated Cd immobilization. Notably, AMF exhibited superior efficacy in reducing Cd bioavailability compared to PE treatment alone, as evidenced by significantly lower DTPA-extractable Cd levels. However, a meta-analysis showed that AMF does not impact soil metal availability 62 . The bioavailable Cd content in rhizosphere soil ranged from 9.8 to 15.0 mg kg − 1 , with maximum values observed in PE0 treatment. Importantly, we identified a significant positive correlation (R 2 = 0.73, p < 0.01, Fig. 6 d) between DTPA-extractable Cd and the total content of acid soluble and reducible fractions, supporting the functional relationship between Cd speciation and bioavailability 21 . HMs phytoavailability indicates the proportion that plants can absorb. In this study, a positive relation was found between shoot Cd concentration and DTPA-Cd (R 2 = 0.33, p < 0.05, Fig. 6 e). The effects of AMF and PE on DTPA-Cd were significant, though no significant interaction between them was observed (Table S3). Our findings show that both AM inoculation and the MPs-AMF interaction significantly lowered the active Cd level, leading to reduced Cd absorption in maize tissues. Therefore, the outcomes support the hypothesis that AMF and MPs decrease Cd absorption and transfer through influencing Cd forms and mobility. 3.6. AMF inoculation regulates rhizosphere soil physicochemical features under Cd and PE stress Both AMF and MPs can change the cycling process of carbon and phosphorus in the soil, thus affecting properties of the soil. The impact of MPs and AMF on soil pH remains controversial. Our study identified Cd concentration and MPs dosage as the primary factors influencing pH dynamics. In Cd-free soil, PE showed no significant effect on pH, whereas AMF increased pH by 1.9% at the PE5 level. In Cd-contaminated soil, pH exhibited an upward trend with increasing PE levels, and AMF further elevated pH, though not significantly. Yu et al. 66 observed that PE addition led to minimal changes in soil pH, while Liu et al. 67 documented more substantial pH increases under similar conditions. In MPs-amended soil, the impact of AMF on soil pH varies with the size of MPs, as MPs of different sizes affect root exudates, including amino acids, differently 27 . Notably, Cd-polluted soils exhibited significantly higher pH than their Cd-free counterparts, potentially due to Cd-stimulated root exudation activity. Given that soil pH critically governs HMs speciation and bioavailability, our results demonstrate that the observed pH shifts correlate with reduced DTPA-Cd. Specifically, AMF-induced pH elevation enhanced Cd adsorption, decreasing bioavailable Cd. Soil OM increased with rising PE levels regardless of Cd content, and AMF further amplified OM accumulation. This aligns with Liu et al. 67 showed 10% PE enhance soil OM, possibly by altering microbial activity and enzyme-mediated OM mineralization. However, PE addition significantly reduced soil AP. In Cd-free soil, 5% PE decreased AP by 21.5% compared to PE0, while in Cd-contaminated soil, the reduction was 17.7%. AMF further lowered AP by 22.3–27.7% in Cd-PE polluted soils, likely due to enhanced plant P uptake. The combination of PE and AMF led to the highest pH and OM levels, but the lowest AP. The increased pH and OM under PE treatment promoted Cd immobilization, shifting it toward less bioavailable forms and reducing root uptake. Soil enzymes play crucial roles in nutrient cycling and serve as sensitive indicators of soil health, reflecting both soil physicochemical properties and microbial functionality 68 . Their activity levels directly influence plant growth performance and respond rapidly to environmental stressors, including heavy metal contamination. However, the response patterns of soil enzymes to combined Cd and MPs pollution remain controversial 69 . Our 40-day incubation study systematically evaluated the activities of urease, catalase, and acid phosphatase under various treatment conditions (Fig. 6 d-f). Key findings revealed that Cd exposure significantly ( p < 0.05) suppressed urease (-9.2%) and acid phosphatase (-28.0%) activities while enhancing catalase activity (+ 44.4%). High-dose PE (5%) amplified Cd’s inhibitory effect on urease (-14.9%) and acid phosphatase (-15.6%) compared to PE0, but further stimulated catalase activity (+ 22.2%). AMF generally increased urease activity by 14.6% and 18.3% in Cd-polluted soils at PE0.5 and PE5 levels, respectively. For catalase activity, PE increased it in Cd-polluted soils, but AMF reduced this effect. In Cd-free soils, AMF raised catalase activity by 27.1–52.6%, while PE had minimal impact ( p > 0.05). Regarding acid phosphatase, MPs generally suppressed its activity, but AMF counteracted this inhibition. In soils co-contaminated with Cd and PE, AMF promoted the synthesis of urease and acid phosphatase but decreased catalase activity. The observed increase of pH in Cd-PE contaminated soils likely contributed to acid phosphatase inhibition, as this pH-sensitive enzyme prefers acidic conditions. AMF’s positive effects on acid phosphatase may stem from: direct secretion of the enzyme by extraradical hyphae and enhanced rhizosphere microbial activity through hyphal exudates. This enzymatic regulation ultimately promoted phosphorus mineralization and plant uptake, explaining the corresponding decrease in AP content. 3.7. Regulation of rhizosphere soil microbial community by AMF inoculation The soil microbial community is pivotal for sustaining soil quality and plant health, as it mediates a wide range of essential biochemical processes 51 , 70 . This study evaluated soil bacterial community characteristics using α-diversity indices: ACE and Chao1 (for microbial richness) and Shannon and Simpson (for microbial diversity). Higher ACE/Chao1 values indicate greater richness, while higher Shannon values combined with lower Simpson values reflect enhanced diversity 31 .​ In the absence of AMF inoculation, Cd contamination significantly reduced ACE, Chao1, and Shannon indices; similarly, high-concentration PE exerted a negative effect, significantly decreasing ACE and Chao1 (Table 1 ). In contrast to the findings of Zhao et al. 16 , our results indicate a significant effect of MPs on rhizosphere bacterial diversity under Cd stress. In contrast, combined AMF inoculation and low-concentration PE application in Cd-contaminated soil yielded the highest ACE, Chao1, and Shannon values, indicating this treatment optimally enhanced bacterial richness and diversity. Our results uncover a novel role of low-concentration MPs in facilitating AMF to improve soil ecosystem resilience and health under Cd stress, revealing a previously unrecognized synergistic interaction. Previous studies have addressed the effects of Cd and MPs on soil bacterial community composition 51 , 71 . Given that plant adaptation to contaminated soils is closely linked to rhizosphere microbial characteristics, investigating shifts in bacterial communities is critical for elucidating the microbiological mechanisms underlying enhanced phytoremediation. As shown in Fig. 7 , the dominant bacterial phyla in maize rhizosphere soil were Pseudomonadota , Bacillota , Actinomycetota , and Bacteroidota , collectively accounting for over 95% of all identified dominant phyla. This study found that combined PE + Cd pollution decreased Pseudomonadota abundance while increasing Actinomycetota abundance, compared to PE-only treatment. This shift aligns with the findings of Zhang et al. 70 , who reported the dominance of Pseudomonadota and Actinomycetota in MPs-Cd co-contaminated soils due to their high adaptability and resistance. Further analysis revealed consistent trends in microbial responses to stress conditions. Cd stress reduced Pseudomonadota abundance regardless of PE presence, while increasing Actinomycetota abundance. Under Cd stress, high-dose PE shifted the soil microbial composition, driving a shift from Pseudomonadota to a dominance of stress-tolerant Bacillota , with negligible effects on other major phyla. This aligns with Xu et al. 30 , who reported that 5% PE reduced Pseudomonadota abundance. Notably, under combined Cd and high-concentration PE stress, AMF inoculation increased the relative abundance of Pseudomonadota from 32.76% (uninoculated group) to 37.59%, while reducing that of Bdellovibrionota from 1.21% to 0.17%. The rise in Pseudomonadota is ecologically significant, as members of this phylum are known for their tolerance to HMs stress and their ability to inhibit HMs transfer in plants 71 . Thus, the recovery of Pseudomonadota under AMF + low-dose PE implies restoration of its functional potential, thereby improving soil nutrient cycling and plant stress resistance. Table 1 Alpha diversity indicators of rhizosphere soil bacterial diversity. Soil Cd (mg kg − 1 ) AMF PE treatments ACE Chao1 Shannon Simpson 0 - PE0 468.65 ± 18.28a 463.20 ± 17.65a 4.64 ± 0.07a 0.025 ± 0.002b - PE0.5 431.84 ± 39.61ab 427.98 ± 39.40a 4.61 ± 0.02a 0.023 ± 0.002b - PE5 378.54 ± 16.93c 374.78 ± 15.29b 4.31 ± 0.07b 0.038 ± 0.005a + PE0 413.18 ± 29.98bc 411.10 ± 28.23ab 4.25 ± 0.14b 0.048 ± 0.011a + PE0.5 442.96 ± 17.29ab 438.82 ± 18.18a 4.54 ± 0.07a 0.021 ± 0.002b + PE5 440.87 ± 36.90ab 438.04 ± 35.98a 4.30 ± 0.09b 0.044 ± 0.007a 20 - PE0 373.88 ± 45.32cd 370.10 ± 45.58cd 4.22 ± 0.19c 0.035 ± 0.009a - PE0.5 395.40 ± 10.80bcd 391.24 ± 11.41bcd 4.31 ± 0.02bc 0.032 ± 0.003a - PE5 351.40 ± 54.04d 346.70 ± 51.52d 4.20 ± 0.14c 0.034 ± 0.006a + PE0 417.82 ± 31.88bc 414.32 ± 31.01bc 4.31 ± 0.08bc 0.034 ± 0.005a + PE0.5 496.93 ± 24.28a 492.92 ± 23.63a 4.62 ± 0.11a 0.028 ± 0.006a + PE5 441.72 ± 16.75ab 437.95 ± 17.06ab 4.45 ± 0.06ab 0.029 ± 0.004a Different lowercase letters denote significant differences among treatments with different PE and AMF inoculation under the same Cd stress level (Duncan test; p < 0.05). –AMF: non-inoculated control; +AMF: inoculated with AMF. 4. Conclusion This study elucidates how AMF alleviate the combined toxicity of Cd and PE-MPs in maize by reducing Cd bioavailability, modulating its subcellular distribution, and improving plant physiology. Notably, a low dose of PE-MPs (0.5%) enhanced the Cd-mitigating effect of AMF, revealing a potential synergistic interaction that warrants further mechanistic investigation. The capacity of AMF to simultaneously enhance plant metal tolerance and potentially influence contaminant bioavailability positions it as a promising bioremediation agent for co-contaminated soils. While this study reveals key mechanisms under controlled conditions, future work should focus on validating these findings under field conditions with environmentally realistic pollutant gradients. Further research should also investigate the long-term dynamics of both Cd and MPs fate in AMF-assisted systems, particularly focusing on the role of AMF in regulating the uptake and transport of MPs within plants. Additionally, exploring the tripartite interactions in diverse soil-plant systems will help develop more robust and widely applicable remediation strategies. Declarations Competing interests The authors declare that there are no conflicts of interest. Author Contribution Yi Lin: Conceptualization, Methodology, Formal analysis, Writing–original draft, Funding acquisition. Xiaoli Sun: Conceptualization, Writing–review & editing, Supervision. Jiping Chen: Supervision, Writing–review & editing. Acknowledgements This work was supported by the Joint Fund of the Zhejiang Provincial Natural Science Foundation of China (grant No. LLSQN25E030003); and the Public Welfare Technology Application Research Project of Lishui (grant No. 2023GYX10). Data Availability All data generated or analyzed during this study are included in this article. References Khan MA, Khan S, Khan A, Alam M. Soil contamination with cadmium, consequences and remediation using organic amendments. 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Kuang QQ, Wu YJ, Gao YM, An TT, Liu S, Liang LY, Xu BC, Zhang SQ, Yu M, Shabala S, Chen YL. Arbuscular mycorrhizal fungi mitigate cadmium stress in maize. Ecotoxicol Environ Saf. 2025;289:117600. https://doi.org/10.1016/j.ecoenv.2024.117600 . Zhao M, Li Y, Li C, Wang X, Cao B, Zhang J, Wang J, Zou G, Chen Y. Effects of polyurethane microplastics combined with cadmium on maize growth and cadmium accumulation under different long-term fertilisation histories. J Hazard Mater. 2024;473:134726. https://doi.org/10.1016/j.jhazmat.2024.134726 . Yang J, Ma YZ, Hao SY, Qin YX, Zhu HD, Wu FY. Arbuscular mycorrhizal fungi regulate cadmium uptake and detoxification in winter wheat via Cd dose-dependent molecular and cellular mechanisms. J Environ Sci. 2025. https://doi.org/10.1016/j.jes.2025.08.051 . Sun P, Chen Y, Li X, Liu L, Guo J, Zheng X, Liu X. Detoxification mechanisms of biochar on plants in chromium contaminated soil: Chromium chemical forms and subcellular distribution. Chemosphere. 2023;327:138505. https://doi.org/10.1016/j.chemosphere.2023.138505 . Li H, Luo N, Zhang LJ, Zhao HM, Li YW, Cai QY, Wong MH, Mo CH. Do arbuscular mycorrhizal fungi affect cadmium uptake kinetics, subcellular distribution and chemical forms in rice? Sci Total Environ. 2016;571:1183–90. https://doi.org/10.1016/j.scitotenv.2016.07.124 . Chen W, Chen Z, Li F, Zhang J, Wu Y, Wang Y. Alfalfa growth, Pb accumulation and bacterial communities in response to co-contamination with microplastics and Pb. Appl Soil Ecol. 2025;213:106278. https://doi.org/10.1016/j.apsoil.2025.106278 . Tang YJ, Xing Y, Wang X, Ya HB, Zhang T, Lv MJ, Wang JC, Zhang H, Dai W, Zhang D, Zheng R, Jiang B. PET microplastics influenced microbial community and heavy metal speciation in heavy-metal contaminated soils. Appl Soil Ecol. 2024;201:105488. https://doi.org/10.1016/j.apsoil.2024.105488 . Yu H, Hou J, Dang Q, Cui D, Xi B, Tan W. Decrease in bioavailability of soil heavy metals caused by the presence of microplastics varies across aggregate levels. J Hazard Mater. 2020;395:122690. https://doi.org/10.1016/j.jhazmat.2020.122690 . Chen G, Huang X, Chen P, Gong X, Wang X, Liu S, Huang Z, Fang Q, Pan Q, Tan X. Polystyrene influence on Pb bioavailability and rhizosphere toxicity: Challenges for ramie ( Boehmeria nivea L.) in soil phytoremediation. Sci Total Environ. 2024;954:176322. https://doi.org/10.1016/j.scitotenv.2024.176322 . Hu S, Hu B, You L, Vogel-Mikuš K, Pongrac P, Vavpetič P, Chen Z, Zhao F. Pb uptake, translocation and allocation in Iris pseudacorus from arbuscular mycorrhizal fungi-assisted constructed wetlands. Chem Eng J. 2025;518:164888. https://doi.org/10.1016/j.cej.2025.164888 . Yu Q, Gao B, Wu P, Chen M, He C, Zhang X. Effects of microplastics on the phytoremediation of Cd, Pb, and Zn contaminated soils by Solanum photeinocarpum and Lantana camara . Environ Res. 2023;231:116312. https://doi.org/10.1016/j.envres.2023.116312 . Liu D, Iqbal S, Gui H, Xu J, An S, Xing B. Nano-iron oxide (Fe 3 O 4 ) mitigates the effects of microplastics on a ryegrass soil-microbe-plant system. ACS Nano. 2023;17:24867–82. https://doi.org/10.1021/acsnano.3c05809 . Seo Y, Lai Y, Chen G, Dearnaley J, Li L, Song P. Size and concentration-dependent effects of polyethylene microplastics on soil chemistry in a microcosm study. J Hazard Mater. 2025;497:139668. 10.1016/j.jhazmat.2025.139668 . https://doi-org.autorpa.ntunhs.edu.tw:8443/ . Wang Q, Wang Q, Wang T, Zhang S, Yu H. Impacts of polypropylene microplastics on the distribution of cadmium, enzyme activities, and bacterial community in black soil at the aggregate level. Sci Total Environ. 2024;917:170541. https://doi.org/10.1016/j.scitotenv.2024.170541 . Zhang Y, Chen Y, Jiao RQ, Gao SS, Li BL, Li YY, Han H, Chen ZJ. Beneficial microbial consortia effectively alleviated plant stress caused by the synergistic toxicity of microplastics and cadmium. Ind Crops Prod. 2025;225:120479. https://doi.org/10.1016/j.indcrop.2025.120479 . Ji JH, Zhong YY, Xiao ML, Wang XT, Hu ZE, Zhan MJ, Ding JN, Zhu ZK, Ge TD. Synergistic effect of microplastics and cadmium on microbial community and functional taxa in wheat rhizosphere soil. Soil Ecol Lett. 2025;7:240260. https://doi.org/10.1007/s42832-024-0260-4 . Additional Declarations No competing interests reported. Supplementary Files SupplementaryMaterial.doc Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Revision requested 01 Mar, 2026 Reviews received at journal 28 Feb, 2026 Reviews received at journal 09 Feb, 2026 Reviewers agreed at journal 02 Feb, 2026 Reviews received at journal 02 Feb, 2026 Reviewers agreed at journal 29 Jan, 2026 Reviewers agreed at journal 27 Jan, 2026 Reviewers agreed at journal 27 Jan, 2026 Reviewers invited by journal 27 Jan, 2026 Editor assigned by journal 25 Jan, 2026 Submission checks completed at journal 23 Jan, 2026 First submitted to journal 22 Jan, 2026 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-8569352","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":582683233,"identity":"7b87c196-4ec9-493c-826f-6c5eccbebe10","order_by":0,"name":"Yi Lin","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAsUlEQVRIiWNgGAWjYBACPmYgkVBhAeZIEKWFDazljAQpWkAEYxtJWth5j0k8nCdhb3CA+eBtHga7PCIcxpcmkbhNInHDAbZkax6G5GIitPCYgbQkGBzgMZPmYTiQ2ECcljkgh/F/I0VLgwTjhgM8bERrMbZIOCaROPMwm7HlHINkwlr4+c8Y3vxRY2PPd7z54Y03FXaEtQABCyQ6QHHKYECEepDaD8SpGwWjYBSMghELAL0kLklEhlIYAAAAAElFTkSuQmCC","orcid":"","institution":"Lishui University","correspondingAuthor":true,"prefix":"","firstName":"Yi","middleName":"","lastName":"Lin","suffix":""},{"id":582683236,"identity":"86dfd30b-d7b1-40f6-86f1-c553df8b91d9","order_by":1,"name":"Xiaoli Sun","email":"","orcid":"","institution":"Lishui University","correspondingAuthor":false,"prefix":"","firstName":"Xiaoli","middleName":"","lastName":"Sun","suffix":""},{"id":582683238,"identity":"bc2bc060-5ea3-4c03-9431-4df372af3bfe","order_by":2,"name":"Jiping Chen","email":"","orcid":"","institution":"Dalian Institute of Chemical Physics","correspondingAuthor":false,"prefix":"","firstName":"Jiping","middleName":"","lastName":"Chen","suffix":""}],"badges":[],"createdAt":"2026-01-10 15:53:24","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-8569352/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8569352/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":101752383,"identity":"052819e8-d34e-4709-a2d0-20ff38665dc0","added_by":"auto","created_at":"2026-02-03 10:27:09","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":1416687,"visible":true,"origin":"","legend":"\u003cp\u003eGrowth indices of maize under various treatments. (a) 30-day actual growth, (b) dry weights, (c) shoot height, (d) root system morphology, (e) net photosynthetic rate, (f) chlorophyll content, (g) phosphorus concentration in shoots and roots, (h) Evan’s blue staining of maize leaves exposed to Cd and PE with AMF inoculation. Different lowercase letters denote significant differences among treatments with varying PE and AMF inoculation under the same Cd stress level (Duncan test; p \u0026lt; 0.05). An asterisk indicates a significant difference between 20 mg kg\u003csup\u003e−1\u003c/sup\u003e Cd stress (Cd20) and Cd-free (Cd0) treatments (t-test; p \u0026lt; 0.05). ns denotes no significant difference at p \u0026gt; 0.05. –AMF: Treatment without AMF; +AMF: Treatment with AMF.\u003c/p\u003e","description":"","filename":"Picture1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8569352/v1/1827e10eb8b977b15e612869.jpg"},{"id":101752382,"identity":"c441fbf0-3e93-4b1d-83c2-d7271fdec499","added_by":"auto","created_at":"2026-02-03 10:27:09","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":1655480,"visible":true,"origin":"","legend":"\u003cp\u003eCd concentration in the shoot and root of maize grown under Cd-free (Cd0) and 20 mg kg\u003csup\u003e−1\u003c/sup\u003e Cd stress (Cd20) for 40 days and the Cd transfer factor (TF) in soil polluted with 20 mg kg\u003csup\u003e−1\u003c/sup\u003e Cd. Different lowercase letters denote significant differences among treatments with different PE and AMF inoculation under the same Cd stress level (Duncan test; p \u0026lt; 0.05). –AMF: non-inoculated control; +AMF: inoculated with AMF.\u003c/p\u003e","description":"","filename":"Picture2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8569352/v1/1ed2a49f98e1fd81ad02ad78.jpg"},{"id":101528275,"identity":"8d292fdf-cbfe-41d7-b9a7-a4cf876fdbe1","added_by":"auto","created_at":"2026-01-30 19:02:42","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":1126864,"visible":true,"origin":"","legend":"\u003cp\u003eEffects of different treatments on antioxidant enzyme activities and MDA content in maize shoots: (a) SOD activity, (b) POD activity, (c) CAT activity, (d) MDA content, (e) antioxidant enzyme activity in maize leaves in response to shoot Cd concentration, and (f) MDA in maize leaves in response to shoot Cd content. Different lowercase letters indicate significant differences among treatments with different PE and AMF inoculation under the same Cd stress level (Duncan test; \u003cem\u003ep\u003c/em\u003e \u0026lt; 0.05). An asterisk indicates a significant difference between 20 mg kg\u003csup\u003e−1 \u003c/sup\u003eCd stress (Cd20) and Cd-free (Cd0) treatments (t-test; \u003cem\u003ep\u003c/em\u003e \u0026lt; 0.05). –AMF: non-inoculated control; +AMF: inoculated with AMF.\u003c/p\u003e","description":"","filename":"Picture3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8569352/v1/aefca9c93b89a8b6740c217d.jpg"},{"id":101752469,"identity":"3b244676-cd0c-40c7-a8c7-c5ba37156ee9","added_by":"auto","created_at":"2026-02-03 10:27:41","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":1212221,"visible":true,"origin":"","legend":"\u003cp\u003eSubcellular distribution and chemical forms of Cd in maize: (a) Cd concentration in subcellular components; (b) distribution ratio of Cd in subcellular components; (c) Cd concentration in each chemical form; (d) distribution ratio of chemical forms of Cd. Different lowercase letters indicate significant differences among treatments with different PE and AMF inoculation (Duncan test; \u003cem\u003ep\u003c/em\u003e \u0026lt; 0.05). –AMF: non-inoculated control; +AMF: inoculated with AMF.\u003c/p\u003e","description":"","filename":"Picture4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8569352/v1/c871d44088478d67a910f106.jpg"},{"id":101528274,"identity":"6d614f86-0269-46a6-93f8-612fe787c8ca","added_by":"auto","created_at":"2026-01-30 19:02:42","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":1066167,"visible":true,"origin":"","legend":"\u003cp\u003eSoil Cd chemical fractions and bioavailability: (a) concentrations of different Cd fractions, (b) percentage distribution of Cd fractions, (c) bioavailable Cd (DTPA-extractable) concentration in soil, (d) relationship between DTPA-Cd and the sum of acid-soluble and reducible Cd fractions, (e) relationship between Cd concentration in maize shoots and soil DTPA-Cd content. Different lowercase letters indicate significant differences among treatments with different PE and AMF inoculation (Duncan test; \u003cem\u003ep\u003c/em\u003e\u0026lt; 0.05). –AMF: non-inoculated control; +AMF: inoculated with AMF.\u003c/p\u003e","description":"","filename":"Picture5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8569352/v1/2dcab6dcfe0bdb7dba9ac330.jpg"},{"id":101528276,"identity":"a75f3509-a51e-4ace-811d-cb5e7afb1e86","added_by":"auto","created_at":"2026-01-30 19:02:42","extension":"jpg","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":1186881,"visible":true,"origin":"","legend":"\u003cp\u003eEffects of AMF on soil properties under combined Cd and PE: (a) soil pH, (b) soil organic matter, (c) soil available phosphorus, (d) soil urease activity, (e) soil acid phosphatase activity, (f) soil catalase activity. Different lowercase letters indicate significant differences among treatments with different PE addition and AMF inoculation (Duncan test; \u003cem\u003ep\u003c/em\u003e \u0026lt; 0.05). An asterisk indicates a significant difference between 20 mg kg\u003csup\u003e−1 \u003c/sup\u003eCd stress (Cd20) and Cd-free (Cd0) treatments (t-test; \u003cem\u003ep\u003c/em\u003e \u0026lt; 0.05). –AMF: non-inoculated control; +AMF: inoculated with AMF.\u003c/p\u003e","description":"","filename":"Picture6.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8569352/v1/04b3a62dec8a404099b19e16.jpg"},{"id":101528277,"identity":"aabaa4d8-a184-4e6e-8892-b624ecdb065c","added_by":"auto","created_at":"2026-01-30 19:02:42","extension":"jpg","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":1273875,"visible":true,"origin":"","legend":"\u003cp\u003eShifts in bacterial community composition at the phylum level, driven by AMF inoculation under different Cd (0 or 20 mg kg\u003csup\u003e−1\u003c/sup\u003e) and PE-MPs (0, 0.5%, or 5%) concentrations.\u003c/p\u003e","description":"","filename":"Picture7.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8569352/v1/26bc7bd365679b1b41e1acbb.jpg"},{"id":101880907,"identity":"8202008b-354f-42e4-b08f-ff02a59ba40a","added_by":"auto","created_at":"2026-02-04 15:07:42","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":10153458,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8569352/v1/820e8200-52e3-4066-85de-1e3a2dacc301.pdf"},{"id":101528271,"identity":"4841e171-599f-433f-97ae-d50cf79d8476","added_by":"auto","created_at":"2026-01-30 19:02:42","extension":"doc","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":102912,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryMaterial.doc","url":"https://assets-eu.researchsquare.com/files/rs-8569352/v1/201129bb472014549d6bd7ad.doc"}],"financialInterests":"No competing interests reported.","formattedTitle":"Arbuscular mycorrhizal fungi enhance maize cadmium resistance and reduce translocation: Dependence on microplastics concentration","fulltext":[{"header":"1. Background","content":"\u003cp\u003eSoil contamination by heavy metals (HMs), particularly cadmium (Cd), has evolved into a critical global environmental crisis, severely endangering food security and ecosystem integrity \u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e. Hou et al. \u003csup\u003e2\u003c/sup\u003e reported that approximately 14\u0026ndash;17% of global croplands are contaminated by HMs, among which Cd exhibits the highest exceedance rate at 9%. Concurrently, microplastics (MPs), polymer-based particles with a diameter\u0026thinsp;\u0026lt;\u0026thinsp;5 mm, have become ubiquitous pollutants posing a persistent anthropogenic challenge across terrestrial and aquatic ecosystems \u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e. Global plastic production now exceeds 430\u0026nbsp;million tons annually with 10 to 40\u0026nbsp;million metric tons of MPs emitted into the environment each year \u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e. A global soil survey \u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e documented MPs concentrations ranging from 0 to 10\u003csup\u003e4\u003c/sup\u003e mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, with polyethylene (PE) identified as the most predominant type. In China, approximately 14.7% of agricultural soils are at risk of MPs pollution, with abundances varying from 7 to 3.61 \u0026times; 10\u003csup\u003e5\u003c/sup\u003e items kg\u003csup\u003e\u0026minus;\u0026thinsp;1 6\u003c/sup\u003e. MPs can enter plants through both foliar deposition and root uptake, and their presence has been confirmed in various food crops \u003csup\u003e\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e. Alarmingly, MPs have also been detected in multiple human tissues and organs, with accumulating evidence indicating potential adverse impacts on human health \u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e. Notably, the spatial distribution of MPs pollution shows remarkable overlap with HMs-contaminated areas \u003csup\u003e\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e, suggesting their combined effects warrant thorough investigation. Given their high toxicity and strong bioaccumulation potential in the food chain, the co-occurrence of Cd and MPs in agricultural soils is of particular environmental concern, underscoring the urgent need for effective mitigation and remediation strategies \u003csup\u003e\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eMPs possess high surface reactivity and a large specific surface area, enabling them to act as carriers for HMs and potentially alter their environmental behavior \u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e. Additionally, under HMs stress, MPs have been shown to induce the activation of the plant antioxidant defense system \u003csup\u003e\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u003c/sup\u003e. Zhao et al. \u003csup\u003e13\u003c/sup\u003e highlighted the potential of MPs to modulate the phytoremediation efficiency of Cd-contaminated soils. Despite the growing body of research investigating the combined effects of MPs and HMs co-pollution on plants, inconsistencies persist regarding whether their interactions exhibit synergistic, antagonistic, or neutral effects \u003csup\u003e\u003cspan additionalcitationids=\"CR15\" citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e. Specifically, Li et al. \u003csup\u003e15\u003c/sup\u003e reported that PE-MPs reduced the bioavailability of Cd in soil, thereby decreasing Cd uptake by pakchoi, without exerting significant impacts on plant biomass. In contrast, Zhao et al. \u003csup\u003e16\u003c/sup\u003e demonstrated that MPs increased the concentration of bioavailable Cd in soil and promoted Cd accumulation in maize. Similarly, Gan et al. \u003csup\u003e17\u003c/sup\u003e found that PE-MPs inhibited plant growth while enhancing Cd uptake, whereas another study documented a positive effect of MPs on tomato growth under Cd stress \u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e. A recent meta-analysis further clarified that MPs tend to lower soil pH, which in turn increases the concentration of bioavailable Cd in soil and enhances Cd accumulation in plant shoots by 11.0% \u003csup\u003e19\u003c/sup\u003e. The complex interactions between MPs and HMs are further compounded by multiple influencing factors, including contaminant types, concentrations, MPs particle size, and inherent soil physicochemical properties \u003csup\u003e\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u003c/sup\u003e. Notably, most existing studies focusing on the toxicological effects of MPs and Cd on plants have primarily centered on plant physiological responses to external stressors, while frequently neglecting the pivotal regulatory role of soil microorganisms in mediating these biotic-abiotic interactions.\u003c/p\u003e \u003cp\u003eArbuscular mycorrhizal fungi (AMF) have emerged as a powerful biological tool for phytoremediation of contaminated soils. Extensive research, including our prior investigations \u003csup\u003e\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e, \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u003c/sup\u003e, has demonstrated that AMF inoculation significantly enhance phytoremediation efficiency in HMs-polluted soils through multiple mechanisms \u003csup\u003e\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u003c/sup\u003e. While the interactions between AMF and MPs have begun to attract scientific attention \u003csup\u003e\u003cspan additionalcitationids=\"CR25\" citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u003c/sup\u003e, current understanding remains limited. AMF are widely recognized for mitigating pollutant toxicity in plants. However, Giambalvo et al. \u003csup\u003e24\u003c/sup\u003e observed that AMF reduced maize shoot biomass by an average of 7% in soils contaminated with MPs. Notably, the impact of AMF on plant tolerance to MPs is influenced by the size of the MPs \u003csup\u003e\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e. Although existing studies have predominantly examined the individual effects of MPs or HMs in AMF-plant systems, emerging evidence suggests complex interactions among these factors. MPs may exacerbate the risk of HMs in the AMF-plant system \u003csup\u003e\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e\u003c/sup\u003e. AMF can alleviate the combined phytotoxicity of MPs and Cd to plants \u003csup\u003e\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e, potentially by reducing the translocation of MPs to shoots \u003csup\u003e\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e. Both MPs and AMF can significantly alter soil microbial communities and physicochemical properties, creating a dynamic rhizosphere environment that modulates contaminant bioavailability \u003csup\u003e\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e, \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e\u003c/sup\u003e. Nevertheless, the tripartite interactions among AMF, MPs, and HMs in co-contaminated soils remain poorly characterized. This is particularly true regarding their combined effects on plant growth, metal accumulation dynamics, and stress tolerance mechanisms. This critical knowledge gap underscores the urgency of comprehensive studies to decipher how AMF modulate plant adaptive responses within this MPs-HMs co-contaminated rhizosphere environment.\u003c/p\u003e \u003cp\u003eMaize (\u003cem\u003eZea mays\u003c/em\u003e L.), a globally critical staple crop that feeds nearly one-third of the world\u0026rsquo;s population, serves as an ideal model for this investigation owing to its rapid growth, high biomass, and documented tolerance to both HMs and MPs \u003csup\u003e\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e, \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e\u003c/sup\u003e. While the interactive effects between Cd and MPs in soil-maize systems have gained increasing attention, the regulatory mechanisms underlying AMF-mediated plant adaptation to co-exposure stresses remain largely elusive. Within the soil-AMF-plant system, it is therefore imperative to clarify how MPs modulate Cd translocation. Furthermore, soil MPs concentrations may alter the capacity of AMF to immobilize Cd. Consequently, elucidating the transport and immobilization processes of Cd at the soil-AMF-plant interface under varying MPs contamination levels is critical for developing effective remediation strategies. Notably, a significant knowledge gap persists regarding how different MPs concentrations regulate Cd dynamics in soil-AMF-crop systems. The present study investigates the effects of AMF inoculation on maize cultivated in sterilized soil under three distinct PE-MPs concentration gradients, with a specific focus on Cd speciation, translocation, and phytotoxicity. We hypothesize that PE-MPs concentration gradients will exert divergent impacts on Cd bioavailability and plant physiological responses in AMF-colonized systems. The specific objectives are to: (1) unravel the mechanisms governing Cd bioaccumulation and phytotoxicity in soil-maize systems under AMF mediation; (2) evaluate the regulatory role of AMF in maize growth parameters and Cd distribution patterns within plant tissues; and (3) quantify the concentration-dependent effects of PE-MPs on Cd uptake by maize and the associated phytotoxicological consequences.\u003c/p\u003e"},{"header":"2. Materials and methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1. Soil, seeds, MPs and AMF inoculum\u003c/h2\u003e \u003cp\u003eSoil samples (0\u0026ndash;20 cm depth) were collected from an agricultural field in Lishui City, Zhejiang Province, China (119\u0026deg;45\u0026rsquo;E, 28\u0026deg;02\u0026rsquo;N). After air-drying (14-day) and sieving (2 mm) for use. Soil properties were analyzed: pH (6.3), total nitrogen (2.90 g kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e), organic matter (52.63 g kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e), total phosphorus (665.0 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e), available phosphorus (13.82 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e), and background Cd (0.23 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e). PE-MPs (100 \u0026micro;m) were procured from Telang Plastic Chemical Co., Ltd. (Dongguan, China) and pre-cleaned with 0.1 M HCl followed by deionized water rinsing. Maize seeds (Zhenuoyu 16, Cd-tolerant cultivar) were obtained from Zhejiang Kecheng Seed Industry Co., Ltd. The AMF \u003cem\u003eRhizophagus intraradices\u003c/em\u003e (BGC BJ09) was provided by the Institute of Plant Nutrition and Resources, Beijing Academy of Agriculture and Forestry. The inoculum contained spores (80 spores g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e soil), hyphae, root fragments, and sandy soil.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2. Experimental design and greenhouse setup\u003c/h2\u003e \u003cp\u003eSoil was spiked with CdCl₂ (0 and 20 mg Cd kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e) and equilibrated for two weeks. Three PE-MPs treatments were applied: 0% (PE0), 0.5% (PE0.5), and 5% (PE5) by dry soil weight. The 0.5% dose is commonly used in experiments involving MPs \u003csup\u003e\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e\u003c/sup\u003e. The 5% MPs concentration used in this study simulates extreme long-term accumulation scenarios. This level is particularly relevant to severely impacted areas like industrial mulch sites, where concentrations can reach up to 6.7% \u003csup\u003e35\u003c/sup\u003e. Employing such an elevated exposure aligns with established ecotoxicological approaches \u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e, \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e, \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e\u003c/sup\u003e and aids in identifying critical risk thresholds and elucidating underlying toxicological mechanisms. Although this concentration may exceed typical near-term environmental exposures, it is methodologically justified within ecotoxicology for revealing mechanistic tipping points and projecting long-term cumulative effects. Our experimental design thus establishes a continuous dose-response relationship, spanning from environmentally realistic levels to extreme concentrations. This range is essential for comprehensively assessing the ecological impacts of microplastics, particularly under conditions of combined pollution. Each pot (16 cm height \u0026times; 17 cm diameter) contained 2.0 kg dry soil. The AMF treatment received 30 g of \u003cem\u003eR. intraradices\u003c/em\u003e inoculum, while the non-mycorrhizal control was amended with autoclaved inoculum and 5 mL of filtered (15-\u0026micro;m) microbial washings to maintain comparable microbiota. Maize seeds were sequentially washed with deionized water and surface-sterilized in 2% (v/v) H₂O₂ for 12 h, followed by thorough rinsing with deionized water. Sterilized seeds were placed on moist absorbent cotton in Petri dishes and germinated at 25\u0026deg;C. Upon reaching 3\u0026ndash;5 cm root length, seedlings were transplanted into pots. After six days of acclimation, two uniform seedlings were retained per pot. Soil moisture was maintained at 15\u0026ndash;18% through daily irrigation with deionized water. Environmental conditions were controlled at 23\u0026thinsp;\u0026plusmn;\u0026thinsp;3\u0026deg;C air temperature and 45\u0026thinsp;\u0026plusmn;\u0026thinsp;5% relative humidity throughout the 40-day experimental period (January 21 to March 1, 2025). The experiment employed a fully factorial design with three replicates per treatment, resulting in 12 treatments across 36 pots. Detailed treatment combinations are listed in Supplementary Table \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3. Plant growth and physiological measurements\u003c/h2\u003e \u003cdiv id=\"Sec6\" class=\"Section3\"\u003e \u003ch2\u003e2.3.1. Biomass and photosynthesis\u003c/h2\u003e \u003cp\u003eShoots and roots were washed thoroughly with deionized water, and oven-dried (65\u0026deg;C, 48 h) for dry weight (DW) determination. Fresh leaves were homogenized with 95% ethanol at 4\u0026deg;C and a spectrophotometer (P4, Mapada, China) was used to measure the chlorophyll (Chl) content. The Chl levels (mg g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e FW) were calculated as follows \u003csup\u003e\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e\u003c/sup\u003e: Chl a\u0026thinsp;=\u0026thinsp;12.7 \u0026times; A663\u0026thinsp;\u0026minus;\u0026thinsp;2.69 \u0026times; A645; Chl b\u0026thinsp;=\u0026thinsp;22.9 \u0026times; A645\u0026thinsp;\u0026minus;\u0026thinsp;4.68 \u0026times; A663. Net photosynthetic rates (Pn) was measured between 9:00 and 11:00 am using a photosynthetic instrument (CIRAS-4, PP-Systems, Hitchin, UK) at 1200 \u0026micro;mol m\u003csup\u003e\u0026minus;\u0026thinsp;2\u003c/sup\u003e s\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e light intensity and 400 \u0026micro;mol mol\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e CO\u003csub\u003e2\u003c/sub\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section3\"\u003e \u003ch2\u003e2.3.2. Elemental analysis\u003c/h2\u003e \u003cp\u003eDried tissues were microwave-digested (HNO\u003csub\u003e3\u003c/sub\u003e:HClO\u003csub\u003e4\u003c/sub\u003e, 3:1 v/v), and Cd/P concentrations were determined via ICP-MS (iCAP RQ, Thermo Fisher Scientific, USA). Quality control included certified reference materials (National Center of Analysis and Testing, Beijing). The detection limits of Cd and P were 0.05 \u0026micro;g L\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003e2.4. Oxidative stress and cellular damage assays\u003c/h2\u003e \u003cp\u003eFresh leaves were homogenized in PBS (0.05 M, pH 7.0) and to prepare a supernatant. The MDA, SOD, POD, and CAT levels were determined using assay kits (Nanjing Jiancheng Bioengineering Institute, China) according to the manufacturer\u0026rsquo;s protocols \u003csup\u003e\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e\u003c/sup\u003e. To determine cell death, leaves were immersed in a 0.25% Evans blue solution for 24 h and then boiled in 95% ethanol for 30 min to remove chlorophyll \u003csup\u003e\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003e2.5. Subcellular distribution, chemical forms of Cd\u003c/h2\u003e \u003cp\u003eSubcellular distribution analysis of Cd was conducted following the method described by previous study \u003csup\u003e\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e\u003c/sup\u003e. The samples were separated into different cellular fractions, including cell wall fraction, cell organelle fraction, and soluble fractions, using differential centrifugation at 4\u0026deg;C. Detailed extraction procedures can be found in Supplementary Text S1. The Cd content in the subcellular fractions was determined using ICP-MS. The chemical forms of Cd in the maize were determined using stepwise extraction \u003csup\u003e\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e\u003c/sup\u003e (Supplementary Text S2).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec10\" class=\"Section2\"\u003e \u003ch2\u003e2.6. Soil chemical and enzymatic analyses\u003c/h2\u003e \u003cdiv id=\"Sec11\" class=\"Section3\"\u003e \u003ch2\u003e2.6.1. Cd fractions\u003c/h2\u003e \u003cp\u003eRoot-attached soil particles were carefully separated by gentle shaking to obtain rhizosphere soil samples. The collected soil was homogenized by thorough mixing and divided into two aliquots. One portion was immediately stored at -80\u0026deg;C for subsequent soil microbial community analysis. The other portion was air-dried for two weeks at room temperature (20\u0026ndash;25\u0026deg;C) and sieved (2 mm mesh) for physicochemical characterization. Soils (0.500 g\u0026thinsp;\u0026plusmn;\u0026thinsp;0.003) were digested with HCl-HNO\u003csub\u003e3\u003c/sub\u003e (3:1 v/v). The procedure of microwave digestion was as follows: 120\u0026deg;C for 10 mins, 150\u0026deg;C for 10 mins and 190\u0026deg;C for 40 mins. The three-step BCR sequential extraction procedure was employed to extract the different Cd fractions from the soil, including acid-soluble Cd, reducible Cd, oxidizable Cd, and residual Cd (Supplementary Table S2) \u003csup\u003e\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e\u003c/sup\u003e. The residual fraction was obtained by digesting the final residues from step 3 with concentrated HCl-HNO\u003csub\u003e3\u003c/sub\u003e (3:1, v/v). Bioavailable Cd was extracted using 0.005 M DTPA (pH 7.3) \u003csup\u003e43\u003c/sup\u003e. All Cd concentrations were quantified by ICP-MS (iCAP RQ, Thermo Fisher Scientific, USA).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section3\"\u003e \u003ch2\u003e2.6.2. Soil physicochemical properties\u003c/h2\u003e \u003cp\u003eSoil AP was measured using 0.5 M NaHCO\u003csub\u003e3\u003c/sub\u003e (pH 8.5) with the Mo\u0026ndash;Sb anti-spectrophotometric method \u003csup\u003e\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e\u003c/sup\u003e. Soil OM was measured according to the potassium dichromate oxidation method \u003csup\u003e\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e\u003c/sup\u003e. Soil pH was measured with a pH meter (LC-PH-3S, Shanghai, China) at a water to soil ratio of 2.5:1 (w/v). Soil enzyme activities were quantified using standardized colorimetric assays following Wang et al. \u003csup\u003e45\u003c/sup\u003e with commercial test kits (Nanjing Jiancheng Bioengineering Institute, China). Three key enzymes were analyzed. Urease activity was determined by measuring the changes in absorbance at 578 nm using a spectrophotometer (T6-New Century; Beijing Purkinje General Instrument Co., Ltd., China) and reported in \u0026micro;g NH\u003csub\u003e3\u003c/sub\u003e\u0026ndash;N g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e 24h\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e at 37\u0026deg;C. Acid phosphatase activity was determined by measuring the changes in absorbance at 400 nm and reported in \u0026micro;mol PNP g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e 24h\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e at 37\u0026deg;C. The CAT activity was determined by measuring changes in the absorbance at 240 nm and reported in \u0026micro;mol H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e h\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003e2.7. Soil Bacterial community analysis\u003c/h2\u003e \u003cp\u003eRhizosphere soil DNA was extracted using the E.Z.N.A.\u0026reg; soil DNA Kit (OMEGA, USA), with three independent biological replicates of soil samples processed per treatment. The quality of the extracted DNA was evaluated via 1% agarose gel electrophoresis. Microbial community analysis was conducted by Shanghai Majorbio Bio-pharm Technology Co., Ltd. The V3\u0026ndash;V4 hypervariable region of the bacterial 16S rRNA gene was amplified using the primers 338F (5\u0026rsquo;-ACTCCTACGGGAGGCAGCAG-3\u0026rsquo;) and 806R (5\u0026rsquo;-GGACTACHVGGGTW TCTAAT-3\u0026rsquo;). PCR amplification was performed following the protocol described by Xu et al. \u003csup\u003e30\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003e2.8. Statistical analysis\u003c/h2\u003e \u003cp\u003eData (mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation, n\u0026thinsp;=\u0026thinsp;3) were analyzed using SPSS 26.0 (IBM, Armonk, NY, USA), and figures were generated with Origin 2024 (Northampton, MA, USA). A one-way analysis of variance (ANOVA) was conducted to compare significant differences among treatments at the same Cd level, with a significance threshold set at \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05 using Duncan\u0026rsquo;s test. For assessing the effects of Cd, PE, AMF inoculation, and their interactions on plant growth parameters and soil-related experimental indices, two-way or three-way ANOVA was employed. Prior to ANOVA analysis, the normality and homogeneity of variance of the data were verified using the Shapiro-Wilk test.\u003c/p\u003e \u003c/div\u003e"},{"header":"3. Results and discussion","content":"\u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003e3.1 MPs and Cd effects on maize growth traits are modified by AMF\u003c/h2\u003e \u003cp\u003eAlthough the individual toxic effects of MPs and Cd on plants are well-documented, their combined impacts remain poorly understood. Cd, PE, and AMF significantly influenced the growth indices of maize (Table S3). As shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea, maize growth status at 30-day intervals demonstrates significant improvement with AMF inoculation. Quantitative analysis reveals that Cd exposure alone reduced maize dry weight by 46.0% in shoots and 37.1% in roots (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb). In Cd-free soil, PE exerted concentration-dependent physiological effects. Although shoot biomass remained unaffected, application of 5% PE significantly enhanced root biomass, achieving a 41.5% increase relative to the control. The three-way ANOVA revealed that root biomass was influenced by soil Cd and PE concentrations (Table S3). Notably, AMF inoculation significantly alleviated the combined toxicity of MPs and Cd, increasing shoot biomass by 87.5% and root biomass by 32.5% in the Cd+PE5 treatment (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05; Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb). The 5% PE addition exacerbated Cd toxicity, resulting in a 29.6% reduction in shoot biomass compared to Cd treatment alone. Across all treatments, AMF application enhanced maize growth, increasing shoot and root dry weight by 63.8% and 32.5%, respectively. The most pronounced growth promotion occurred in low dose PE (0.5%) with AMF, showing 17.9% and 41.3% increases in shoot biomass relative to PE5-Cd and PE0-Cd treatments.\u003c/p\u003e \u003cp\u003ePE addition significantly altered biomass allocation, with 5% PE increasing the root-to-shoot ratio by 67.4% under Cd-free conditions and by 59.5% under Cd20 stress, whereas 0.5% PE reduced it by 18.9% under Cd exposure (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec). AMF-treated plants consistently exhibited lower root-to-shoot ratio, indicating preferential shoot growth promotion. Figure\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ed showed that more root at PE5 and AMF mostly. The findings demonstrate a concentration-dependent response to PE, inhibitory at high level and stimulatory at low level, which supports the potential of maize for remediating moderately contaminated soils. Photosynthetic analysis revealed that Cd stress dramatically reduced Pn, while PE alone showed negligible effects (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ee). AMF inoculation alleviated photosynthetic inhibition under stress conditions, increasing Pn by 20.9% under Cd and 38.5% under Cd+PE5, whereas the Cd+PE5 treatment itself decreased chlorophyll content and Pn by 41.0% and 51.6%, respectively, relative to the control (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ef). A study indicated MPs can affect photosynthetic ability and decrease the photosynthetic pigment content \u003csup\u003e\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e\u003c/sup\u003e. Under Cd stress, 10% PE-MPs inhibited Pn of maize \u003csup\u003e\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u003c/sup\u003e. The impact of MPs on plant growth is dual, potentially promoting or inhibiting growth, or having no noticeable effect. The effects vary across different plant organs and are influenced by the concentration and type of MPs, as well as the plant species and its growth environment \u003csup\u003e\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e\u003c/sup\u003e. Xiang et al. \u003csup\u003e48\u003c/sup\u003e found that PE had no effect on shoot biomass, but increased root biomass through a metal-analysis. Liu et al. \u003csup\u003e49\u003c/sup\u003e found that adding 0.2% polyester to Cd-polluted soils increased rice biomass, whereas 0.2% polyethylene terephthalate had no effect on biomass. The combined toxicity of Pb and MPs to plants is more evident than when they are present individually \u003csup\u003e\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eAlthough the mitigating or exacerbating effects of PE on plants in metal-polluted conditions have been documented, no studies have investigated the responses of the AMF-crop system to combined MPs and Cd stress. The toxicity of HMs may be amplified when combined with PE. Liu et al. \u003csup\u003e51\u003c/sup\u003e reported that addition of PE increased Cd toxicity in sorghum, as evidenced by a reduction in length and biomass. MPs can affect the biomass allocation and tend to increase root to shoot biomass ration \u003csup\u003e\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e\u003c/sup\u003e. A meta-analysis pointed out that low levels MPs promotion shoot and high levels promotion root in order to resist stress \u003csup\u003e\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e\u003c/sup\u003e. AMF enhanced phosphorus uptake, particularly at high PE concentration (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eg), corresponding with observed growth improvements. PE and Cd damaged the photosynthetic system to different degrees, causing leaf cell more damaged (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eh). These physiological changes were paired with distinct morphological alterations in both Cd-free and Cd-polluted soils, underscoring the intricate interactions between PE-MPs, AMF, and Cd stress.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003e3.2 Coexisting AMF and MPs influenced the uptake and transfer of Cd in plants, and exhibited a \u0026ldquo;dose effect\u0026rdquo;\u003c/b\u003e \u003c/p\u003e \u003cp\u003eMPs concentration was one of the main factors affecting toxic elements accumulation in plants \u003csup\u003e\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e\u003c/sup\u003e. Shoot and root Cd concentrations were significantly influenced by Cd, PE, and their interaction, while the AMF \u0026times; Cd interaction also significantly affected shoot Cd concentration (Table S3). Without Cd treatments, the concentration of Cd in shoots and roots remained stable and was unaffected by the presence of PE or AMF. Our results demonstrated that Cd accumulation in roots was significantly higher than in shoots, as evidenced by TF\u0026thinsp;\u0026lt;\u0026thinsp;1 (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Maize, as an excluder plant (TF\u0026thinsp;\u0026lt;\u0026thinsp;1), has been shown to primarily accumulate Cd in its roots, effectively immobilizing it and limiting its translocation to the aerial parts \u003csup\u003e\u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e\u003c/sup\u003e. PE treatment significantly reduced Cd uptake in maize roots by 21.3\u0026ndash;35.7% in non-inoculated plants and by 12.6\u0026ndash;21.2% in AMF-treated plants. Interestingly, shoot Cd concentrations in AMF-treated maize remained stable regardless of PE addition. However, in non-AMF treatments, shoot Cd levels in the PE5 group were 20.4% higher than in the PE0.5 group. For most crops, AMF has been shown to inhibit the uptake of metals to prevent damage. Kuang et al. \u003csup\u003e56\u003c/sup\u003e reported that AMF decreased TF of Cd in maize. The effect of AMF on Cd translocation under MPs co-exposure was Cd-level dependent, significantly reducing the TF at 40 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e Cd but showing no effect at 80 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e Cd \u003csup\u003e29\u003c/sup\u003e. AMF inoculation alone decreased root Cd uptake in PE0 treatment by 14.4%, while PE addition attenuated this beneficial effect. In both AMF-inoculated and non-inoculated plants, the TF of Cd in maize exhibited a dose-dependent decrease with increasing PE concentration. Although PE generally enhanced Cd translocation to shoots by reducing root absorption, AMF inoculation effectively counteracted this phenomenon.\u003c/p\u003e \u003cp\u003eThe reduced Cd uptake in AMF-treated plants may be attributed to a dilution effect resulting from enhanced plant growth. Our previous research demonstrated a strong correlation between plant Cd accumulation and soil bioavailable Cd content \u003csup\u003e\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e. The current study found that MPs reduced Cd bioavailability, a finding consistent with prior reports that 0.25 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e PE decreased Cd bioaccessibility by 13.3% \u003csup\u003e51\u003c/sup\u003e. These results suggested that MPs influenced plant Cd concentrations primarily by modifying soil physicochemical and microbial properties. Previous research indicated that MPs affected Cd absorption in a dose-dependent manner, with higher MPs concentrations promoting Cd accumulation in both shoots and roots \u003csup\u003e\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003e3.3. Antioxidant response of maize\u003c/h2\u003e \u003cp\u003eBoth PE alone and the interaction between Cd and AMF significantly influenced all enzyme activities (Table S3). As shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea-d, combined contamination with Cd and PE elicits complex responses in the antioxidant enzyme system and promotes membrane lipid peroxidation in maize. In contrast, AMF inoculation alleviates oxidative damage under co-contamination conditions by modulating the antioxidant defense response. Under Cd-free conditions, SOD activity was enhanced by 5% PE but remained unaffected by AMF except in the absence of PE. In Cd-stressed soil, 5% PE increased SOD activity, while AMF inoculation suppressed this PE-induced enhancement. With respect to POD activity, the addition of PE (both 0.5% and 5%) tended to reduce it in Cd-free soil, whereas no significant effect was observed in Cd20 soil. Notably, AMF consistently up-regulated POD activity across all Cd and PE treatments. The CAT activity exhibited concentration- and condition-dependent behavior. In the absence of Cd, 0.5% PE significantly increased CAT activity regardless of AMF presence, whereas high-concentration PE showed no stimulatory effect. Under Cd20 conditions without AMF, both low and high PE concentrations tended to enhance CAT activity. In contrast, under Cd stress, AMF inoculation in combination with low PE reduced CAT activity. Unlike previous findings in \u003cem\u003eSolanum nigrum\u003c/em\u003e \u003csup\u003e\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e\u003c/sup\u003e, where Cd stress elevated the activities of SOD, POD, and CAT, the present results demonstrate that Cd specifically induced POD and CAT activities, while SOD remained largely unaltered.\u003c/p\u003e \u003cp\u003eThe generation of reactive oxygen species (ROS), during oxidative bursts is a key mechanism underlying the toxic effects of Cd and MPs on plants \u003csup\u003e\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e\u003c/sup\u003e. The antioxidant system counteracts such damage primarily via a sequential enzymatic process involving SOD, which converts superoxide into H₂O₂, followed by CAT and POD, which catalyze the decomposition of H₂O₂ \u003csup\u003e37, 38\u003c/sup\u003e. By scavenging ROS, these enzymes help alleviate Cd phytotoxicity and limit its uptake and accumulation. Previous evidence shows that AMF colonization can significantly enhance enzyme activity under Cd stress \u003csup\u003e\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e\u003c/sup\u003e. The previous study demonstrated that increased Cd content in plants can boost antioxidant enzyme activity \u003csup\u003e\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e\u003c/sup\u003e. This study\u0026rsquo;s polynomial regression analysis identified a significant positive correlation between SOD activity and Cd accumulation in the aboveground plant parts, suggesting a nonlinear relationship (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ee). However, CAT activity and POD levels showed no correlation with Cd content. The degree of membrane lipid peroxidation was assessed by measuring MDA. The levels of MDA in maize showed a positive correlation with shoot Cd concentration (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ef). Importantly, AMF effectively reduced MDA levels under all treatment conditions, indicating alleviated oxidative membrane damage. The elevated MDA content induced by Cd was further exacerbated by high-concentration PE, highlighting the synergistic toxicity of combined pollutants. The modulation of antioxidant enzymes suggests that AMF enhances ROS scavenging capacity, particularly through sustained up-regulation of POD activity. The distinct responses of SOD and CAT indicate a shift in redox management strategy under mycorrhizal symbiosis. The reduction in MDA underscores the protective role of AMF at the cellular level, likely mediated through enhanced Cd immobilization and diminished ROS production \u003csup\u003e\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e\u003c/sup\u003e. Overall, AMF inoculation improves maize tolerance to combined Cd-PE stress by fine-tuning the antioxidant system and maintaining membrane integrity.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec18\" class=\"Section2\"\u003e \u003ch2\u003e3.4. MPs and AMF effects on subcellular distribution and chemical forms of Cd are varying\u003c/h2\u003e \u003cp\u003eThe subcellular distribution and chemical forms of HMs play a pivotal role in plant metals tolerance and detoxification through compartmentalization and chelation mechanisms \u003csup\u003e\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e, \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e\u003c/sup\u003e. The partitioning of metals within plant subcellular compartments serves as a critical determinant of their intracellular toxicity. Plants have evolved multiple strategies to mitigate metal toxicity, including chelation, vacuolar sequestration, and root immobilization to restrict translocation to aerial tissues \u003csup\u003e\u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e\u003c/sup\u003e. Our findings demonstrate that AMF inoculation significantly reduced Cd content across all subcellular fractions in shoots (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea). The observed distribution pattern in maize shoots followed: cell wall fraction (F\u003csub\u003eCW\u003c/sub\u003e, 47.7\u0026ndash;61.5%) \u0026gt; soluble fraction (F\u003csub\u003eS\u003c/sub\u003e, 25.0\u0026ndash;31.4%) \u0026gt; organelle fraction (F\u003csub\u003eO\u003c/sub\u003e, 10.4\u0026ndash;27.3%) (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eb). This result suggests that cell wall deposition and vacuolar compartmentalization constitute primary detoxification pathways in maize, which is consistent with the response observed in \u003cem\u003ePopulus yunnanensis\u003c/em\u003e under Cd stress \u003csup\u003e\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e\u003c/sup\u003e. These results corroborate previous studies identifying F\u003csub\u003eCW\u003c/sub\u003e as the predominant Cd form in maize shoots where cell wall binding serves as a key mechanism for Cd toxicity alleviation \u003csup\u003e\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e, \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eIn this study, the AMF-mediated subcellular redistribution exhibited PE concentration dependence. AMF increased F\u003csub\u003eS\u003c/sub\u003e (+\u0026thinsp;21.7%) at PE0 while AMF increased F\u003csub\u003eCW\u003c/sub\u003e (+\u0026thinsp;26.0%) at PE5. At low dose of PE, there was an increase in F\u003csub\u003eCW\u003c/sub\u003e and a decrease in F\u003csub\u003eO\u003c/sub\u003e, regardless of the presence of AMF. High dose PE increased F\u003csub\u003eO\u003c/sub\u003e (+\u0026thinsp;35.3%), while AMF can offset that. At PE0.5 treatment, more Cd accumulated in F\u003csub\u003eCW\u003c/sub\u003e, thereby reducing its harmful effects. We observed a distinctive AMF-induced modification in leaf Cd partitioning, characterized by preferential accumulation in cell wall-integrated forms. While our previous research confirmed AMF\u0026rsquo;s capacity to enhance F\u003csub\u003eCW\u003c/sub\u003e \u003csup\u003e\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u003c/sup\u003e, elevated PE concentrations attenuated this protective effect. Notably, the F\u003csub\u003eO\u003c/sub\u003e fraction demonstrated an inverse relationship with F\u003csub\u003eCW\u003c/sub\u003e, decreasing with AMF colonization but increasing with PE addition. These findings align with Zhang et al.\u0026rsquo;s \u003csup\u003e55\u003c/sup\u003e report of AMF-enhanced F\u003csub\u003eCW\u003c/sub\u003e in maize roots. The increase in F\u003csub\u003eO\u003c/sub\u003e proportion induced by high-dose PE contributed to the exacerbation of Cd phytotoxicity.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eChemical speciation analysis revealed substantial AMF-induced modifications under combined PE-Cd stress (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ec and d). F\u003csub\u003eE\u003c/sub\u003e and F\u003csub\u003eW\u003c/sub\u003e exhibit strong migration abilities, while F\u003csub\u003eNaCl\u003c/sub\u003e has a lower migration ability and toxicity, and F\u003csub\u003eHAc\u003c/sub\u003e, F\u003csub\u003eHCl\u003c/sub\u003e, and F\u003csub\u003eR\u003c/sub\u003e show the weakest toxicity to plants. The distribution of Cd forms followed: F\u003csub\u003eNaCl\u003c/sub\u003e (31.9\u0026ndash;51.0%) \u0026gt; F\u003csub\u003eE\u003c/sub\u003e (16.9\u0026ndash;29.7%) \u0026gt; F\u003csub\u003eW\u003c/sub\u003e (18.8\u0026ndash;26.8%) \u0026gt; F\u003csub\u003eHAc\u003c/sub\u003e (4.1\u0026ndash;16.1%) \u0026gt; F\u003csub\u003eHCl\u003c/sub\u003e (1.7\u0026ndash;3.5%), with F\u003csub\u003eR\u003c/sub\u003e below 0.5%. This pattern is consistent with established literature documenting F\u003csub\u003eE\u003c/sub\u003e, F\u003csub\u003eW\u003c/sub\u003e, and F\u003csub\u003eNaCl\u003c/sub\u003e as predominant Cd forms in maize shoots \u003csup\u003e\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e\u003c/sup\u003e. The phytotoxicity of Cd is primarily governed by its biological activity and chemical speciation within plant tissues. Our findings demonstrate that high dose PE (5%) significantly increased the proportions of F\u003csub\u003eE\u003c/sub\u003e (+\u0026thinsp;20.0%) and F\u003csub\u003eW\u003c/sub\u003e (+\u0026thinsp;28.2%), consequently exacerbating Cd toxicity. In contrast, AMF inoculation effectively mitigated Cd toxicity by reducing the fractions of biologically active Cd, with average reductions of 26.5% in F\u003csub\u003eE\u003c/sub\u003e and 12.1% in F\u003csub\u003eW\u003c/sub\u003e. Notably, AMF consistently promoted the transformation of Cd into F\u003csub\u003eHAc\u003c/sub\u003e across all PE treatment levels, while only enhancing F\u003csub\u003eNaCl\u003c/sub\u003e (+\u0026thinsp;13.6%) at 0.5% PE. These findings are consistent with prior research, which indicates that AMF increases the distribution of Cd in inactive forms while reducing its toxic proportions \u003csup\u003e\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e, 60\u003c/sup\u003e. The differential effects of AMF on Cd chemical speciation provide mechanistic insights into its protective role against Cd toxicity in plants.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec19\" class=\"Section2\"\u003e \u003ch2\u003e3.5. Interactive effects of MPs and AMF on Cd speciation and bioavailability in rhizosphere soil\u003c/h2\u003e \u003cp\u003eThe chemical speciation of HMs fundamentally governs their bioavailability, mobility, and ecotoxicity. Both AMF and MPs can modify soil physicochemical properties and microbial communities, thereby mediating the transformation of trace metal speciation in soil systems \u003csup\u003e\u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e\u003c/sup\u003e. However, the mechanisms underlying MPs-induced alterations in HMs distribution remain poorly characterized \u003csup\u003e\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e\u003c/sup\u003e. Figure\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ea shows soil Cd forms after 40-day of treatment with or without MPs and AMF. Consistent Cd fractionation patterns across treatments: acid soluble (56.1\u0026ndash;61.3%) \u0026gt; reducible (30.5\u0026ndash;39.8%) \u0026gt; oxidizable (3.5\u0026ndash;11.0%) \u0026gt; residual (0.5\u0026ndash;1.6%) (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eb). Both with and without AMF inoculation, high-dose PE consistently reduced acid-soluble Cd while increasing the oxidizable fraction content. In contrast, low-dose PE enhanced the reducible Cd content. AMF inoculation preferentially decreased reducible Cd (12.0\u0026ndash;18.0%) and increased oxidizable forms content (43.0\u0026ndash;89.8%), with minimal effects on acid-soluble Cd. The highest residual Cd was observed in the PE5\u0026thinsp;+\u0026thinsp;AMF treatment. As there was no significant difference in the final residual Cd content among all treatments (19.57\u0026thinsp;\u0026plusmn;\u0026thinsp;0.62 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e), the percentage changes in Cd forms in the soils mirrored the changes in Cd concentration, except for the high dose of PE, which had no significant effect on the acid-soluble Cd percentage.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eIt was reported that MPs dose affected Cd bioavailability and phytotoxicity \u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e. The results demonstrated that high-dose PE significantly increased the proportions of oxidizable Cd forms and decreased acid soluble Cd, indicating PE facilitate the transformation from active to organically-bound species. This transformation may be attributed to PE-induced increases in soil pH and OM content, coupled with the inherent adsorptive capacity of insoluble MPs \u003csup\u003e\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e\u003c/sup\u003e. However, Chen et al. \u003csup\u003e64\u003c/sup\u003e reported that MPs increased Pb availability with reducible Pb increased by 6\u0026ndash;12%. MPs enhanced Cd availability in wheat rhizosphere by reduced soil pH and 5% PE more effectively increased bioavailable Cd than 1% PE \u003csup\u003e17\u003c/sup\u003e. The addition of MPs facilitates the conversion of inactive Cd into active Cd \u003csup\u003e19\u003c/sup\u003e. PE at low dose can decrease the concentration of available Cd, thus reducing environmental risk. Research shows that although MPs usually promote metal fixation through organic complexation, their impact on metal migration depends on environmental conditions and the properties of the MPs. AMF inoculation further reduced bioavailable Cd in the maize rhizosphere by facilitating the formation of insoluble Cd compounds via three key mechanisms \u003csup\u003e\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e, \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e, \u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e\u003c/sup\u003e: (1) secretion of metal-chelating molecules (e.g., metallothioneins, glutathione, and phytochelatins); (2) complexation of Cd on the surface of AMF hyphae; (3) enhancement of microbial-mediated Cd immobilization. Notably, AMF exhibited superior efficacy in reducing Cd bioavailability compared to PE treatment alone, as evidenced by significantly lower DTPA-extractable Cd levels. However, a meta-analysis showed that AMF does not impact soil metal availability \u003csup\u003e\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e\u003c/sup\u003e. The bioavailable Cd content in rhizosphere soil ranged from 9.8 to 15.0 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, with maximum values observed in PE0 treatment. Importantly, we identified a significant positive correlation (R\u003csup\u003e2\u003c/sup\u003e\u0026thinsp;=\u0026thinsp;0.73, \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.01, Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ed) between DTPA-extractable Cd and the total content of acid soluble and reducible fractions, supporting the functional relationship between Cd speciation and bioavailability \u003csup\u003e\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e. HMs phytoavailability indicates the proportion that plants can absorb. In this study, a positive relation was found between shoot Cd concentration and DTPA-Cd (R\u003csup\u003e2\u003c/sup\u003e\u0026thinsp;=\u0026thinsp;0.33, \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05, Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ee). The effects of AMF and PE on DTPA-Cd were significant, though no significant interaction between them was observed (Table S3). Our findings show that both AM inoculation and the MPs-AMF interaction significantly lowered the active Cd level, leading to reduced Cd absorption in maize tissues. Therefore, the outcomes support the hypothesis that AMF and MPs decrease Cd absorption and transfer through influencing Cd forms and mobility.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec20\" class=\"Section2\"\u003e \u003ch2\u003e3.6. AMF inoculation regulates rhizosphere soil physicochemical features under Cd and PE stress\u003c/h2\u003e \u003cp\u003eBoth AMF and MPs can change the cycling process of carbon and phosphorus in the soil, thus affecting properties of the soil. The impact of MPs and AMF on soil pH remains controversial. Our study identified Cd concentration and MPs dosage as the primary factors influencing pH dynamics. In Cd-free soil, PE showed no significant effect on pH, whereas AMF increased pH by 1.9% at the PE5 level. In Cd-contaminated soil, pH exhibited an upward trend with increasing PE levels, and AMF further elevated pH, though not significantly. Yu et al. \u003csup\u003e66\u003c/sup\u003e observed that PE addition led to minimal changes in soil pH, while Liu et al. \u003csup\u003e67\u003c/sup\u003e documented more substantial pH increases under similar conditions. In MPs-amended soil, the impact of AMF on soil pH varies with the size of MPs, as MPs of different sizes affect root exudates, including amino acids, differently \u003csup\u003e\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e. Notably, Cd-polluted soils exhibited significantly higher pH than their Cd-free counterparts, potentially due to Cd-stimulated root exudation activity. Given that soil pH critically governs HMs speciation and bioavailability, our results demonstrate that the observed pH shifts correlate with reduced DTPA-Cd. Specifically, AMF-induced pH elevation enhanced Cd adsorption, decreasing bioavailable Cd. Soil OM increased with rising PE levels regardless of Cd content, and AMF further amplified OM accumulation. This aligns with Liu et al.\u003csup\u003e67\u003c/sup\u003e showed 10% PE enhance soil OM, possibly by altering microbial activity and enzyme-mediated OM mineralization. However, PE addition significantly reduced soil AP. In Cd-free soil, 5% PE decreased AP by 21.5% compared to PE0, while in Cd-contaminated soil, the reduction was 17.7%. AMF further lowered AP by 22.3\u0026ndash;27.7% in Cd-PE polluted soils, likely due to enhanced plant P uptake. The combination of PE and AMF led to the highest pH and OM levels, but the lowest AP. The increased pH and OM under PE treatment promoted Cd immobilization, shifting it toward less bioavailable forms and reducing root uptake.\u003c/p\u003e \u003cp\u003eSoil enzymes play crucial roles in nutrient cycling and serve as sensitive indicators of soil health, reflecting both soil physicochemical properties and microbial functionality \u003csup\u003e\u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e68\u003c/span\u003e\u003c/sup\u003e. Their activity levels directly influence plant growth performance and respond rapidly to environmental stressors, including heavy metal contamination. However, the response patterns of soil enzymes to combined Cd and MPs pollution remain controversial \u003csup\u003e\u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e69\u003c/span\u003e\u003c/sup\u003e. Our 40-day incubation study systematically evaluated the activities of urease, catalase, and acid phosphatase under various treatment conditions (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ed-f). Key findings revealed that Cd exposure significantly (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) suppressed urease (-9.2%) and acid phosphatase (-28.0%) activities while enhancing catalase activity (+\u0026thinsp;44.4%). High-dose PE (5%) amplified Cd\u0026rsquo;s inhibitory effect on urease (-14.9%) and acid phosphatase (-15.6%) compared to PE0, but further stimulated catalase activity (+\u0026thinsp;22.2%). AMF generally increased urease activity by 14.6% and 18.3% in Cd-polluted soils at PE0.5 and PE5 levels, respectively. For catalase activity, PE increased it in Cd-polluted soils, but AMF reduced this effect. In Cd-free soils, AMF raised catalase activity by 27.1\u0026ndash;52.6%, while PE had minimal impact (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;0.05). Regarding acid phosphatase, MPs generally suppressed its activity, but AMF counteracted this inhibition. In soils co-contaminated with Cd and PE, AMF promoted the synthesis of urease and acid phosphatase but decreased catalase activity. The observed increase of pH in Cd-PE contaminated soils likely contributed to acid phosphatase inhibition, as this pH-sensitive enzyme prefers acidic conditions. AMF\u0026rsquo;s positive effects on acid phosphatase may stem from: direct secretion of the enzyme by extraradical hyphae and enhanced rhizosphere microbial activity through hyphal exudates. This enzymatic regulation ultimately promoted phosphorus mineralization and plant uptake, explaining the corresponding decrease in AP content.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec21\" class=\"Section2\"\u003e \u003ch2\u003e3.7. Regulation of rhizosphere soil microbial community by AMF inoculation\u003c/h2\u003e \u003cp\u003eThe soil microbial community is pivotal for sustaining soil quality and plant health, as it mediates a wide range of essential biochemical processes \u003csup\u003e\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e, \u003cspan citationid=\"CR70\" class=\"CitationRef\"\u003e70\u003c/span\u003e\u003c/sup\u003e. This study evaluated soil bacterial community characteristics using α-diversity indices: ACE and Chao1 (for microbial richness) and Shannon and Simpson (for microbial diversity). Higher ACE/Chao1 values indicate greater richness, while higher Shannon values combined with lower Simpson values reflect enhanced diversity \u003csup\u003e\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e\u003c/sup\u003e.​ In the absence of AMF inoculation, Cd contamination significantly reduced ACE, Chao1, and Shannon indices; similarly, high-concentration PE exerted a negative effect, significantly decreasing ACE and Chao1 (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). In contrast to the findings of Zhao et al. \u003csup\u003e16\u003c/sup\u003e, our results indicate a significant effect of MPs on rhizosphere bacterial diversity under Cd stress. In contrast, combined AMF inoculation and low-concentration PE application in Cd-contaminated soil yielded the highest ACE, Chao1, and Shannon values, indicating this treatment optimally enhanced bacterial richness and diversity. Our results uncover a novel role of low-concentration MPs in facilitating AMF to improve soil ecosystem resilience and health under Cd stress, revealing a previously unrecognized synergistic interaction. Previous studies have addressed the effects of Cd and MPs on soil bacterial community composition \u003csup\u003e\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e, \u003cspan citationid=\"CR71\" class=\"CitationRef\"\u003e71\u003c/span\u003e\u003c/sup\u003e. Given that plant adaptation to contaminated soils is closely linked to rhizosphere microbial characteristics, investigating shifts in bacterial communities is critical for elucidating the microbiological mechanisms underlying enhanced phytoremediation. As shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003e, the dominant bacterial phyla in maize rhizosphere soil were \u003cem\u003ePseudomonadota\u003c/em\u003e, \u003cem\u003eBacillota\u003c/em\u003e, \u003cem\u003eActinomycetota\u003c/em\u003e, and \u003cem\u003eBacteroidota\u003c/em\u003e, collectively accounting for over 95% of all identified dominant phyla. This study found that combined PE\u0026thinsp;+\u0026thinsp;Cd pollution decreased \u003cem\u003ePseudomonadota\u003c/em\u003e abundance while increasing \u003cem\u003eActinomycetota\u003c/em\u003e abundance, compared to PE-only treatment. This shift aligns with the findings of Zhang et al. \u003csup\u003e70\u003c/sup\u003e, who reported the dominance of \u003cem\u003ePseudomonadota\u003c/em\u003e and \u003cem\u003eActinomycetota\u003c/em\u003e in MPs-Cd co-contaminated soils due to their high adaptability and resistance. Further analysis revealed consistent trends in microbial responses to stress conditions. Cd stress reduced \u003cem\u003ePseudomonadota\u003c/em\u003e abundance regardless of PE presence, while increasing \u003cem\u003eActinomycetota\u003c/em\u003e abundance. Under Cd stress, high-dose PE shifted the soil microbial composition, driving a shift from \u003cem\u003ePseudomonadota\u003c/em\u003e to a dominance of stress-tolerant \u003cem\u003eBacillota\u003c/em\u003e, with negligible effects on other major phyla. This aligns with Xu et al. \u003csup\u003e30\u003c/sup\u003e, who reported that 5% PE reduced \u003cem\u003ePseudomonadota\u003c/em\u003e abundance. Notably, under combined Cd and high-concentration PE stress, AMF inoculation increased the relative abundance of \u003cem\u003ePseudomonadota\u003c/em\u003e from 32.76% (uninoculated group) to 37.59%, while reducing that of \u003cem\u003eBdellovibrionota\u003c/em\u003e from 1.21% to 0.17%. The rise in \u003cem\u003ePseudomonadota\u003c/em\u003e is ecologically significant, as members of this phylum are known for their tolerance to HMs stress and their ability to inhibit HMs transfer in plants \u003csup\u003e\u003cspan citationid=\"CR71\" class=\"CitationRef\"\u003e71\u003c/span\u003e\u003c/sup\u003e. Thus, the recovery of \u003cem\u003ePseudomonadota\u003c/em\u003e under AMF\u0026thinsp;+\u0026thinsp;low-dose PE implies restoration of its functional potential, thereby improving soil nutrient cycling and plant stress resistance.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eAlpha diversity indicators of rhizosphere soil bacterial diversity.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"7\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSoil Cd (mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAMF\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePE treatments\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eACE\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eChao1\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eShannon\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eSimpson\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePE0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e468.65\u0026thinsp;\u0026plusmn;\u0026thinsp;18.28a\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e463.20\u0026thinsp;\u0026plusmn;\u0026thinsp;17.65a\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e4.64\u0026thinsp;\u0026plusmn;\u0026thinsp;0.07a\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.025\u0026thinsp;\u0026plusmn;\u0026thinsp;0.002b\u003c/p\u003e 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colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePE5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e378.54\u0026thinsp;\u0026plusmn;\u0026thinsp;16.93c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e374.78\u0026thinsp;\u0026plusmn;\u0026thinsp;15.29b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e4.31\u0026thinsp;\u0026plusmn;\u0026thinsp;0.07b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.038\u0026thinsp;\u0026plusmn;\u0026thinsp;0.005a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePE0\u003c/p\u003e \u003c/td\u003e 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align=\"left\" colname=\"c5\"\u003e \u003cp\u003e438.82\u0026thinsp;\u0026plusmn;\u0026thinsp;18.18a\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e4.54\u0026thinsp;\u0026plusmn;\u0026thinsp;0.07a\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.021\u0026thinsp;\u0026plusmn;\u0026thinsp;0.002b\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePE5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e440.87\u0026thinsp;\u0026plusmn;\u0026thinsp;36.90ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e438.04\u0026thinsp;\u0026plusmn;\u0026thinsp;35.98a\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e4.30\u0026thinsp;\u0026plusmn;\u0026thinsp;0.09b\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.044\u0026thinsp;\u0026plusmn;\u0026thinsp;0.007a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePE0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e373.88\u0026thinsp;\u0026plusmn;\u0026thinsp;45.32cd\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e370.10\u0026thinsp;\u0026plusmn;\u0026thinsp;45.58cd\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e4.22\u0026thinsp;\u0026plusmn;\u0026thinsp;0.19c\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.035\u0026thinsp;\u0026plusmn;\u0026thinsp;0.009a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePE0.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e395.40\u0026thinsp;\u0026plusmn;\u0026thinsp;10.80bcd\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e391.24\u0026thinsp;\u0026plusmn;\u0026thinsp;11.41bcd\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e4.31\u0026thinsp;\u0026plusmn;\u0026thinsp;0.02bc\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.032\u0026thinsp;\u0026plusmn;\u0026thinsp;0.003a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e 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\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePE0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e417.82\u0026thinsp;\u0026plusmn;\u0026thinsp;31.88bc\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e414.32\u0026thinsp;\u0026plusmn;\u0026thinsp;31.01bc\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e4.31\u0026thinsp;\u0026plusmn;\u0026thinsp;0.08bc\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.034\u0026thinsp;\u0026plusmn;\u0026thinsp;0.005a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePE0.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e496.93\u0026thinsp;\u0026plusmn;\u0026thinsp;24.28a\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e492.92\u0026thinsp;\u0026plusmn;\u0026thinsp;23.63a\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e4.62\u0026thinsp;\u0026plusmn;\u0026thinsp;0.11a\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.028\u0026thinsp;\u0026plusmn;\u0026thinsp;0.006a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e+\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePE5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e441.72\u0026thinsp;\u0026plusmn;\u0026thinsp;16.75ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e437.95\u0026thinsp;\u0026plusmn;\u0026thinsp;17.06ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e4.45\u0026thinsp;\u0026plusmn;\u0026thinsp;0.06ab\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.029\u0026thinsp;\u0026plusmn;\u0026thinsp;0.004a\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eDifferent lowercase letters denote significant differences among treatments with different PE and AMF inoculation under the same Cd stress level (Duncan test; \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05). \u0026ndash;AMF: non-inoculated control; +AMF: inoculated with AMF.\u003c/p\u003e \u003c/div\u003e"},{"header":"4. Conclusion","content":"\u003cp\u003eThis study elucidates how AMF alleviate the combined toxicity of Cd and PE-MPs in maize by reducing Cd bioavailability, modulating its subcellular distribution, and improving plant physiology. Notably, a low dose of PE-MPs (0.5%) enhanced the Cd-mitigating effect of AMF, revealing a potential synergistic interaction that warrants further mechanistic investigation. The capacity of AMF to simultaneously enhance plant metal tolerance and potentially influence contaminant bioavailability positions it as a promising bioremediation agent for co-contaminated soils. While this study reveals key mechanisms under controlled conditions, future work should focus on validating these findings under field conditions with environmentally realistic pollutant gradients. Further research should also investigate the long-term dynamics of both Cd and MPs fate in AMF-assisted systems, particularly focusing on the role of AMF in regulating the uptake and transport of MPs within plants. Additionally, exploring the tripartite interactions in diverse soil-plant systems will help develop more robust and widely applicable remediation strategies.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e \u003ch2\u003eCompeting interests\u003c/h2\u003e \u003cp\u003eThe authors declare that there are no conflicts of interest.\u003c/p\u003e \u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eYi Lin: Conceptualization, Methodology, Formal analysis, Writing\u0026ndash;original draft, Funding acquisition. Xiaoli Sun: Conceptualization, Writing\u0026ndash;review \u0026amp; editing, Supervision. Jiping Chen: Supervision, Writing\u0026ndash;review \u0026amp; editing.\u003c/p\u003e\u003ch2\u003eAcknowledgements\u003c/h2\u003e \u003cp\u003eThis work was supported by the Joint Fund of the Zhejiang Provincial Natural Science Foundation of China (grant No. LLSQN25E030003); and the Public Welfare Technology Application Research Project of Lishui (grant No. 2023GYX10).\u003c/p\u003e\u003ch2\u003eData Availability\u003c/h2\u003e\u003cp\u003eAll data generated or analyzed during this study are included in this article.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eKhan MA, Khan S, Khan A, Alam M. Soil contamination with cadmium, consequences and remediation using organic amendments. 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Soil Ecol Lett. 2025;7:240260. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1007/s42832-024-0260-4\u003c/span\u003e\u003cspan address=\"10.1007/s42832-024-0260-4\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"chemical-and-biological-technologies-in-agriculture","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"Learn more about [Chemical and Biological Technologies in Agriculture](https://chembioagro.springeropen.com/)","snPcode":"40538","submissionUrl":"https://submission.nature.com/new-submission/40538/3","title":"Chemical and Biological Technologies in Agriculture","twitterHandle":"@SpringerPlants","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"phytoremediation, combined PE-MPs + Cd pollution, synergistic toxicity, soil bacterial community, heavy metals bioavailability","lastPublishedDoi":"10.21203/rs.3.rs-8569352/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8569352/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e \u003cp\u003eThe coexistence of microplastics (MPs) and heavy metals (HMs) such as cadmium (Cd) in agricultural soils represents a growing threat to crop production and food security. While arbuscular mycorrhizal fungi (AMF) are recognized for their ability to enhance plant metal tolerance, their role in mediating crop responses under combined contamination with MPs and Cd, especially across different MPs concentrations, remains largely unexplored. This study was conducted to elucidate how AMF modulate maize growth, Cd accumulation, and soil biogeochemical processes under co-contamination with polyethylene (PE) (0, 0.5% and 5% w/w) and Cd (0, 20 mg kg\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e) with or without AMF.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eThe addition of 5% PE-MPs significantly aggravated Cd toxicity in maize, elevating Cd translocation to shoots by 79.6% and causing severe growth suppression. PE-MPs also modified key soil characteristics, increasing organic matter content and pH, which promoted the transformation of Cd into less bioavailable fractions yet failed to counteract its direct phytotoxic effects. Inoculation with AMF markedly alleviated these stresses. Under Cd and 5% PE-MPs co‑contamination, mycorrhizal plants showed 87.5% higher shoot biomass, 39.6% greater phosphorus uptake, and 38.5% enhanced net photosynthesis compared to non‑inoculated plants. AMF further reduced oxidative damage, promoted Cd sequestration in cell walls, decreased the biologically active Cd pool in shoots, and lowered Cd bioavailability through shifts in soil bacterial community composition, particularly by restoring the abundance of \u003cem\u003ePseudomonadota\u003c/em\u003e. The beneficial effects of AMF were more evident at 0.5% PE-MPs than at the 5% concentration.\u003c/p\u003e\u003ch2\u003eConclusions\u003c/h2\u003e \u003cp\u003eThis study demonstrates that AMF confer dual protection in a PE-MPs concentration-dependent manner. AMF enhance plant physiological resilience by regulating antioxidant systems and Cd subcellular distribution, while reducing Cd bioavailability by modifying soil properties, soil bacterial diversity and Cd speciation. These\u003c/p\u003e","manuscriptTitle":"Arbuscular mycorrhizal fungi enhance maize cadmium resistance and reduce translocation: Dependence on microplastics concentration","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-01-30 19:02:37","doi":"10.21203/rs.3.rs-8569352/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2026-03-01T15:18:37+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-01T03:10:07+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-02-09T10:46:59+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"179859040036169066799771792399104957256","date":"2026-02-02T14:01:02+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-02-02T07:57:42+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"57159993024726170090205783773312298901","date":"2026-01-29T21:20:21+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"90171523310354520032285867761535186189","date":"2026-01-28T03:10:26+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"42909388908855087102098305228775653396","date":"2026-01-28T01:57:33+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-01-28T00:34:48+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-01-25T11:49:19+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-01-23T12:08:04+00:00","index":"","fulltext":""},{"type":"submitted","content":"Chemical and Biological Technologies in Agriculture","date":"2026-01-22T16:25:04+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"chemical-and-biological-technologies-in-agriculture","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"Learn more about [Chemical and Biological Technologies in Agriculture](https://chembioagro.springeropen.com/)","snPcode":"40538","submissionUrl":"https://submission.nature.com/new-submission/40538/3","title":"Chemical and Biological Technologies in Agriculture","twitterHandle":"@SpringerPlants","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"aa42c60e-7bd1-4317-9fed-f0d8d79b390d","owner":[],"postedDate":"January 30th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-04-09T08:27:34+00:00","versionOfRecord":[],"versionCreatedAt":"2026-01-30 19:02:37","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8569352","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8569352","identity":"rs-8569352","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}