The interaction effects of degradable microplastics and Cd to Folsomia candida soil collembolan

doi:10.21203/rs.3.rs-1656120/v1

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The interaction effects of degradable microplastics and Cd to Folsomia candida soil collembolan

https://doi.org/10.21203/rs.3.rs-1656120/v1

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published 17 Mar, 2023

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Microplastics have become a major safety concern for soil ecosystem, but in real field soil conditions, multiple chemicals exposure may be the real scenario for soil biota. The co-occurrence of microplastics and Cadmium (Cd) is common in soils, which may pose a potential risks to soil ecosystems. Meanwhile, although degradable microplastics are suggested to use, the potential effects on soil ecosystems are unknown. Therefore, a standard soil animal collembolan Folsomia candida (F. candida) was used to evaluation the single and interaction effects of degradable microplastics and Cd. The results showed that single and co-microplastics and Cd all had negative influences on the survival, reproduction and growth of F. candida, and the effects intensified with microplastics concentrations. But combined microplastic and Cd alleviated the toxicity of single chemical on F. candida at lower microplastics concentrations. Biochemical assays on antioxidant enzymes had the same results. Toxicity endpoints reproduction was not more sensitive than survival and growth. Antioxidant enzymes CAT and POD was more sensitive than SOD, thus may be promising biomarkers on soil collembolan for soil microplastics exposure.

Cadmium

Degradable microplastics

Collembolan

Antioxidant enzymes

Microplastics (MPs), as an emerging contaminant, have become a global environmental problem (Derraik, 2002). Plastics have been widely used in daily life for their convenience, cheapness, and water and chemical resistance (Barnes et al., 2009; Geyer et al., 2017). Thus the widely usage makes a large amount of plastic accumulate in the environment and gradually change to MPs (< 5 mm) by physical, chemical and other external forces decomposition (Serranti et al., 2018; Hu et al., 2019). 300–67,500 mg kg^− 1 and 8–540 mg kg^− 1 microplastics have been detected in industrial soils and farmlands, respectively (Fuller and Gautam 2016; Liu et al. 2018). Studies have found that microplastics had significant effects on soil environments. It can affect soil properties and soil structure, and cause adverse effects on soil biota, and finally induce dangers to the soil ecosystem (Blasing and Amelung, 2018; Machado et al., 2018a, b; Zhu et al., 2018). Although degradable microplastics are suggested to use, the intermediate or final metabolites produced by degradable microplastics may affect soil ecosystems (Qi et al., 2018; Rillig et al., 2019). Some studies found they have negative effects on plant growth and soil bacteria (Rillig et al., 2019; Qi et al., 2020).

In fields, the pollutants do not always exist in a single form, multiple pollutants always exist in soil environments, which may promote or inhibit the toxic effects. Heavily metal is a typical pollutant in soil worldly, especially Cadmium (Cd). According to the 2014 Chinese National Soil Contamination Survey, Cd was the most common metal pollutant of soils in China (Zhao et al., 2015). Cd had high solubility, transferability, phytoavailability and toxicity, which is the main danger to agroecosystems and human health (Wang et al. 2001; Li et al. 2014). The co-occurrence of microplastics and Cd is common in soils (Martin and Turner, 2019; Wan et al., 2021). The interactions between microplastics and the associated contaminants may alter their environmental behaviors, bioavailability and toxicity, leading to potential risks to soil ecosystems. Studies on mixture pollution seem more practical and ecological related to field soils. Microplastics have a large specific surface area, thus may serve as an adsorbent for metals, reducing the availability of metals (Koelmans et al., 2016; Hodson et al., 2017). Meanwhile, studies also found that microplastics in soils may act as a medium to increase the exposure of metals to soil biota, thus increasing the mobility and availability of pollutants (Ramos et al., 2015). To understand the possible effects of degradable microplastics on soil biota, the combined effects of degradable microplastics and metals should be considered (Wang et al., 2020a,b).

Collembolans are widespread soil micro-arthropods, and play an important role in nutrient cycling, energy transfer and maintenance of biodiversity in soils, they are real soil dwellers and thus are sensitive to soil pollution (Fountain and Hopkin 2005; OECD 2009). Several species have been adopted as standard animals for the assessment of soil pollution, especially Folsomia candida (F. candida). The cultivability, short generation time and high reproduction rate make F. candida feasible as a standard species in ecotoxicological tests (Fountain and Hopkin 2005; OECD 2009). Several reliable studies proved that F. candida could ingest microplastic < 66.0 +/- 10.9 mu m plastic, and the velocity and distance of F. candida movement were significantly reduced (Huerta Lwanga et al. 2016, 2017; Rodriguez-Seijo et al. 2017; Maass et al., 2017; Kim and An, 2019, 2020). Luo et al (2021) found that under the food exposure of microplastics, the molting frequency, reproduction and biomass of F. candida decreased significantly compared with control. Therefore, microplastics may have significant effects on the survival, reproduction, growth and locomotion behavior of F. candida. But the tested microplastics mostly belong to non-degradable microplastics such as Poly Ethylen (PE) and polystyrene (PS), the possible effects of degradable microplastics were few studied.

Except for traditional toxicity endpoints (survival, reproduction and growth et al) at individual and population levels, antioxidant enzymes at the cellular level have the advantages of high sensitivity and efficiency, and have been used to give early warning of soil pollution (Howcroft et al., 2009). They play an important role in maintaining the dynamic stability of oxidation-antioxidant action in organisms and ensuring the normal life activities of the organism (Modesto and Martinez 2010). The bioindicators of antioxidant enzymes include catalase (CAT), peroxidase (POD), superoxide dismutase (SOD), glutathione reductase (GR), glutathione-s-transferase (GST) et al on collembola had been investigated under exposure to many pollutants (Yuan et al., 2011; Maria, et al., 2014; Dai et al. 2018; Zheng et al. 2022). Maria et al (2014) and Li et al (2020) found under the exposure of Cd, the activity of CAT changed severely. With the increase of Cd concentrations, CAT activity firstly increased and decreased at higher Cd concentrations, which indicated the occurrence of oxidative damage. Microplastics also cause serious oxidative damage to Eisenia fetida, Cantareus aspersus and Enchytraeus crypticus (Pflugmacher et al., 2020; Sun et al., 2021; Colpaert et al., 2021). However, the potential effects of combined degradable microplastics and Cd on collembolans’ antioxidant systems in soil systems remain poorly studied.

Therefore, the aim of this study was to assess the signal and co-degradable microplastics and Cd effects on standard soil animal F. candida through standard species test and biochemical assays, which may benefit us in the understanding of combined microplastics and Cd environmental risks in soils.

2. 1 Test organism

Folsomia candida was reared in Petri dishes (height 10 mm, diameter 90 mm) with a layer of moist plaster of Paris mixed with activated charcoal (9: 1 w/w) at 20 ± 1 °C and 75% relative humidity with a photoperiod of 16/8 h (light/dark). The animals were fed with dried baker’s yeast (Angel Yeast Co., Ltd., Yichang, Hubei, China) twice a week. Distilled water was added to maintain the moisture content when necessary. Synchronized individuals of F. candida were used according to standardized methods of the Organization for Economic Cooperation and Development (OECD, 2009).

2. 2 Soil preparation

The commercial grade of biodegradable microplastics PLA 4032D (100-154 μm) was used in the test (Nature Works LLC, Minnetonka, MN, USA), with a density of 1.24 g cm^-3, a crystallinity of about 37%, and a melting point of 160 °C. It was cleaned by 0.1 M HCl and distilled water to remove potential metals on their surface and thoroughly mixed into the soil to achieve the target doses. Cd solutions were prepared by dissolving Cd(NO₃)₂·4H₂O in deionized water and adding to MPs mixture soils at target concentrations. The tested soil was taken from the surface of 20 cm in an uncontaminated natural forest with pH H₂O 6.2, soil water holding capacity (WHC) 40%, organic matter content (OM) 18.3 g kg^-1, cation exchange capacity (CEC) 17.2 cmol (+) kg^-1, and clay content of 19.3%. The soils were allowed to equilibrate for one week at 50% of the maximum water holding capacity (WHC) before use.

2. 3 The single species tests with soil

In China, 5 mg kg^-1 of Cd represents most of Cd pollution level (Wang et al., 2015). Thus, the test included two levels of Cd (0 and 5 kg^-1 ) and four levels of microplastics (0, 1, 10 and 100 mg kg^-1). A total of 8 treatments were included and each treatment had three replicates. Ten cleaned and synchronized (10-12 d) F. candida individuals were introduced to a test vessel containing 30 g spiked soil at 20 ± 1 °C, 75% relative humidity and a 16: 8 h photoperiod. Distilled water was supplied twice a week to maintain moist conditions. After exposure for 28 days, the numbers of adults and juveniles in the two tests were determined under a stereomicroscope (Leica S8 APO, Wetzlar, Germany) and the body lengths of adults and juveniles from the leading edge of the head to the end of the abdomen were measured under a microscope camera (MS60, Mingmei Optoelectronic Technology Co. Ltd., Guangzhou, China).

2. 4 Biochemical assays

The soil spiking method and the treatments were as described above. The activities of the antioxidant enzymes superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) were selected in the test and were determined using kits from Nanjing Jiancheng Bioengineering Institute, Nanjing, east China.

Fifty cleaned adult animals were transferred to the test vessels containing 30 g prepared spiked field soil and were exposed for 7 days at 20 ± 1 °C, 75% relative humidity and a 16: 8 h photoperiod. After the test, the animals were carefully collected from the soil. After being washed and homogenized using a tissue grinder (Coyote Co., Ltd, Beijing, China) (1: 9) in 0.1 M K-phosphate buffer (pH 7.4), the tissue homogenate was centrifuged for 10 min at 2500 × g (4 °C) (5417R, Eppendorf, Hamburg, Germany) to obtain the post-mitochondrial supernatant (PMS) and the resulting supernatants were used for the analysis of antioxidant enzymes. CAT activity was determined by measuring the decomposition of the substrate H₂O₂ at 240 nm. POD activity was determined by catalyzing into H₂O₂ at 420 nm. SOD activity determination by superoxide radicals produced by the xanthine–xanthine oxidase system at 550 nm. The analysis was conducted by calibrating the optical density (OD) values with a range of standard concentrations provided by the kits. SOD and POD activities were expressed as U mg per of protein, CAT activity was expressed as U g per protein.

2. 5 Statistical analysis

All data are expressed as mean ± standard error (SE). Significant differences were determined by one-way analysis of variance (ANOVA) and were compared by the Duncan’s multiple range test at P < 0.05. In the analysis of the enzymes, a two-way ANOVA was conducted for differences in the treatments and exposure periods. Statistical analysis was conducted using Microsoft Excel 2016 and the SPSS 21.0 software package (SPSS Inc., Chicago, IL).

3. 1 The effects on collembolan survival and reproduction

The 28-d reproduction test results were shown in Fig. 1 and Fig. 2. Specifically, microplastics (0-10mg kg^-1) with or without the addition of Cd in soils had no adverse effects on the number of surviving adults, but it decreased at the highest microplastics concentrations (100 mg kg^-1) without or with Cd addition, the decreasing rates were 20 and 13.3%, respectively compared with the control (F_{7, 23} = 4.82, P < 0.005).

The number of juveniles had significant differences among the treatments (F_{7, 23} = 120, P < 0.001). Compared with the control group, the number of juveniles decreased by 11.6, 8.37 and 24.2 % at MPs-1, MPs-10 and MPs-100 treatments, but the decreasing rates were lower after the addition of Cd, which were 96.33, 91.05 and 80.4% of MPs-0 + Cd treatment at MPs-1 + Cd, MPs-10 + Cd and MPs-100 + Cd treatments. The single addition of Cd in soils had the most toxic effects on the juvenile numbers, but the combination of microplastics in soils increased the number of juveniles. The number of juveniles increased by 34.3, 25.7 and 9.63 % at MPs-1 + Cd, MPs-10 + Cd and MPs-100 + Cd treatments compared with MPs-1, MPs-10 and MPs-100 treatments, respectively.

3. 2 The effects on collembolan growth

The adult and juvenile body lengths had significant differences among the treatments (Fig. 3). With the increase of microplastics in soils, the body length of adults significantly decreased no matter the addition of Cd in soils (F_{7, 23} = 16.7, P < 0.001). Compared with the control group, the body length of adults decreased by 10.2, 14.1 and 22.9% at MPs-1, MPs-10 and MPs-100 treatments, but the decreasing trends were lower after the addition of Cd, it decreased by 5.50, 9.95 and 17.6% at MPs-1 + Cd, MPs-10 + Cd and MPs-100 + Cd treatments compared with MPs-0 + Cd treatment (Fig. 3A). The addition of Cd in microplastic contaminated soils increased the body length of adults compared with single microplastics treatments, it increased by 4.41, 3.77 and 4.49% at MPs-1 + Cd, MPs-10 + Cd and MPs-100 + Cd treatments, respectively compared with MPs-1, MPs-10 and MPs-100 treatments.

The same trend was also found in the body length of juveniles. With the increase of microplastics in soils, the body length of juveniles decreased dramatically no matter the addition of Cd in soils (F_{7, 23} = 11.7, P < 0.001). Compared with the control group, the body length of juveniles decreased by 15.8, 22.5 and 32.2% at MPs-1, MPs-10 and MPs-100 treatments, but the decreasing trends were lower after the addition of Cd with 3.37, 8.33 and 26.8% at MPs-1 + Cd, MPs-10 + Cd and MPs-100 + Cd treatments (Fig. 3 B). The addition of Cd increased the body length of juveniles compared with single microplastics treatments, they increased by 12.1, 6.92 and 4.23% at MPs-1 + Cd, MPs-10 + Cd and MPs-100 + Cd treatments.

3. 2 The effects on the antioxidant systems of collembolan

Enzyme activities of CAT, POD and SOD on F. candida were induced and had significant differences among the treatments and among the exposure times. The effects on CAT and POD activities were dependent between the treatments and exposure times (Table 1, Fig. 4).

As can be seen in Fig. 4 A, generally, the CAT activity of F. candida was sensitive to microplastics in soils. It increased significantly after 2 days, which was 1.02, 1.51 and 2.00 times higher at MPs-1, MPs-10 and MPs-100 treatment, respectively, than that at MPs-0 treatment. While it declined significantly with the increase of microplastics in soils and exposure times. On day 4, it started to decline at MPs-100 treatment, but the declining trend occurred early at MPs-10 treatment on day 7 and day 14. Cadmium had significant effects on CAT activities, it was induced quickly but declined with the increasing of exposure times. On day 7, it induced to its maximum, which was 1.72 times higher than that on day 2, but it declined by 71.5% on day 14 compared with that on day 7. In the combined microplastics and Cd treatments, the CAT activity of F. candida had no significant differences in lower microplastics treatments (1~10 mg kg-¹), but the combined 100 mg kg^-1 microplastics and Cd significantly increased the CAT activity of F. candida. It increased by 41.6% compared with that at MPs-0 + Cd treatment. Comparing the addition of Cd in microplastics-contaminated soils, the inducement of CAT activity was less sensitive with the increase of microplastics in soils, and the declined effects were less sensitive as the exposure time increased. For example, CAT activity increased 100% at MPs-100 treatment, but it only increased by 41.6% at MPs-100 + Cd treatment. It declined by 26.1, 54.4 and 64.5% at MPs-100 treatment on days 4, 7 and 14 compared with that on day 2, but the declining trend was lower at MPs-100 + Cd treatment, it declined by 16.7, 33.6 and 42.0% on days 4, 7 and 14 compared with the corresponding values on day 2.

For the POD activity of F. candida, it increased with the increase of microplastics in soils at shorter exposure periods but decreased with the increase of microplastics and longer exposure periods no matter the addition of Cd in soils. On day 2, it increased by 19.9, 14.4 and 20.2% at MPs-1, MPs-10 and MPs-100 treatment, and increased by 12.5, 18.8 and 42.9% at MPs-1 + Cd, MPs-10 + Cd and MPs-10 + Cd treatment compared with the corresponding control. But on day 14, the POD activity decreased by 75.0% at MPs-100 treatment compared with MPs-0 treatment, and it was 55.8 % of the values on day 7. The same trend also found in the combined MPs and Cd treatments. On day 14, the POD activity decreased by 35.5% at MPs-100 + Cd treatment compared with, and it was 77.4 % of the values on day 7. The single contamination of Cd in soils significantly decreased the POD activity of F. candida as the exposure periods increased, it decreased by 12.5, 21.9 and 37.5% on days 4, 7 and 14 compared with that on day 2. But the addition of different concentrations of microplastics in soils increased the POD activity of F. candida at all MPs + Cd treatments and exposure periods. Compared with the effects of Cd addition on microplastics contaminated soils, the induced maximum values of POD activities were higher in single microplastics treatments at lower concentrations or higher concentrations with shorter exposure periods. At higher microplastics concentrations, the declined trend was more severe with the exposure time at single microplastics treatments. For example, on day 14, the POD activity declined by 75.0% at MPs-100 treatment compared with that on day 2, but the declining trend was gentler at MPs-100 + Cd treatments.

The SOD activity of F. candida was less sensitive than that of CAT and POD activities at shorter exposure periods and at lower microplastics concentration no matter the addition of Cd in soils, and the significant differences were only found at the highest microplastics treatments (MPs-100 and MPs-100 + Cd treatments ). At MPs-100 treatment, SOD activity increased with the exposure period, it increased by 6.00, 11.5 and 31.0% on days 4, 7 and 14 compared with that on day 2. While, at MPs-100 + Cd treatment, SOD activity decreased with the increasing exposure period, it decreased by 4.68, 24.0 and 33.6% on days 4, 7 and 14 compared with that on day 2. Cadmium had significant effects on the SOD activity of F. candida, it was induced firstly to its highest values on day 7 but decreased by 71.5% on day 14. The SOD activity of F. candida at combined microplastics and Cd treatments had no significant differences with single Cd treatments in the lower exposure period, but it decreased dramatically at higher microplastic concentrations and longer exposure time. For example, on day 7, the SOD activity of F. candida decreased by 45.2% at MPs-100 + Cd treatments compared with that at MPs-0 + Cd treatment.

4. 1 Survival, reproduction and growth

Generally, no matter the addition of Cd in soils, the lower microplastics in soils (0-10 mg kg^-1) had no significant effects on the numbers of adults, but the number of juveniles and the body lengths of adults and juveniles decreased significantly. They may be due to the sensitivity of different toxicity endpoints on microplastics. Numerous studies had confirmed that although the survival data is easy to obtain, it is not sensitive compared with the data on reproduction and growth (Lin et al., 2019, Li et al., 2021). Zhu et al (2018) obtained the same results that under the 5 mg kg^-1 microplastics in soils, low mortality of F. candida was observed, but significant decreases in reproduction and body weight were found. Cao et al (2017) found that under lower microplastics ( < 5 mg kg^-1) exposure, the survival and reproduction rates of earthworms had no significant difference with the control, but with the increase of microplastics ( > 10 mg kg^-1) in soils, the survival and reproduction rates of earthworms were inhibited dramatically. In our study, the adverse effects of degradable microplastics on F. candida reproduction and growth indicated a risk of microplastics on soil biota. Collembolan can ingest microplastics, which may disturb the gut microbiota, metabolism and the absorption of nutrient elements to induce toxic effects (Zhu et al., 2018, Kim and An, 2019, 2020). In addition, studies had found that microplastics have negative effects on the hatching of eggs, some eggs may lay on the surfaces of microplastics but cannot be hatched successfully (Goldstein et al., 2012, Ja and Costa, 2014). This may be another reason for the decrease in reproduction exposed to microplastics.

When Cd and degradable microplastics were combined, the negative effect of Cd on F. candida was correlated with the concentrations of microplastics in soils. The combined Cd and lower concentrations of microplastics in soils seemed to decrease the toxic effects of single microplastic and single Cd on F. candida, but the decreasing effects were less significant at higher concentrations of microplastics. This may be because the higher concentration of microplastics in soil increase the exposure of Cd to soil animals, thus increasing the toxic effects of pollutants on F. candida (Hodson et al., 2017, Feng et al., 2022). Toxicity endpoints at cellular levels may provide some insights into the mechanism of combined microplastics and metals of organisms.

4. 2 Antioxidant enzymes

Antioxidant capacity is commonly used in resistance tests at the cellular level, which is essential in maintaining the dynamic stability of oxidation-antioxidant action in organisms and ensuring the normal life activities of the organism. The antioxidant enzymes like SOD, CAT, and POD et al play an essential role in resisting excess reactive oxygen (ROS) and oxidative damage, which can be used as biomarkers to identify pollutants at early stages and at non-lethal concentrations (Howcroft et al., 2009). In addition, the changes in antioxidant systems at the cellular level are relevant to endpoints at individual and population levels like behavior, growth, survival and reproduction (Novais et al., 2011, Li et al., 2021).

In our studies, the significant difference in antioxidant enzyme activities under different exposure periods and treatments indicated that single and combined Cd or degradable microplastics caused oxidative stress to F. candida. Generally, the antioxidant enzyme activities induced at shorter exposure periods and lower pollution levels, but decreased as exposure periods and pollution levels increased. This may be because the produced excess ROS cannot be counterbalanced by antioxidant enzymes, the activities of antioxidant enzymes were inhibited and further, the oxidative damage occurred (Howcroft et al., 2009, Lushchak, 2011). Li et al (2020) had the same results that antioxidant enzyme activities were increased in Cd contaminated soils, but decreased as the Cd concentration increased. Numerous studies confirmed that microplastics can cause oxidative stress and trigger antioxidant upregulation on soil invertebrates, and the toxic effects are gradually intensified with the increase of exposure periods (Pflugmacher et al., 2020, Sun et al., 2021, Li et al., 2022). But the effects are depended on the microplastic type, shape, size and exposure time (Trestrail et al., 2020). The ingestion of microplastics may cause intestinal damage which leads to the increase of ROS in the body and triggers the imbalance of the antioxidant system (Dupré-Crochet et al., 2013). In addition, the associated chemicals like flame retardants, plasticizers, stabilizers and heavy metals may contribute to the possible oxidative stress (Jemec et al., 2012, Lambert et al., 2017). For example, tire particles as one of the largest sources of microplastics to the environment, contained a huge amount of heavy metals and organic pollutants (Selonen et al., 2021), which may have significant effects on the antioxidant system of collembolan. In our study, the selected PLA belongs to biodegradable microplastic, which is low persistence and easy to be decomposed by soil microorganisms, thus its toxic effects may be more severe than nondegradable microplastics, and the intermediate or final metabolites produced by degradation may affect soil animals (Lo Piparo et al., 2006, Qi et al., 2018, Rillig et al., 2019). Furthermore, Pflugmacher et al (2020) pointed out that the changes in soil properties by microplastics may also alter the activities of antioxidant enzymes and cause oxidative stress. The addition of microplastics could significantly influence soil pH, which has been considered as one of the most limited soil factors for collembolan (Fountain and Hopkin 2005, Dai et al., 2018), thus the potential oxidative damage may occur.

No matter the addition of Cd, the CAT and POD activities induced quickly at shorter exposure periods and microplastics treatments, which indicated that CAT and POD were sensitive to microplastics. While SOD activity had no significant effects at lower microplastics and short exposure periods. CAT and POD enzymes are mainly to cope with H₂O₂, but SOD is mainly in the removal of O^2-, and the activity rates increase with the production of ROS (Modesto and Martinez, 2010). Therefore, H₂O₂ may be the mainly ROS produced by F. candida under microplastics and Cd exposure over the short exposure time, or other antioxidant enzymes acted firstly than SOD (Howcroft et al., 2009, Krifka et al., 2013). Li et al (2022) had the same results that CAT activity was sensitive to microplastic exposure and thus can be used as oxidative stress biomarkers in marine bivalves. Therefore, antioxidant enzymes CAT and POD may be good biomarkers on soil collembolan for soil microplastic exposure.

In our study, the alleviation effects of the addition of Cd in microplastics contaminated soils to F. candida had been found on the population and individual levels. The same results were also found at the cellular level at lower microplastics concentrations and shorter exposure periods. Therefore, the conclusion can be made that the interaction of microplastic and Cd could alleviate the single toxic effects of microplastic and Cd to some extent. Lian et al (2020) and Feng et al (2022) had the same results that the combined microplastics and Cd relieved the toxic effects on the germination, root and bud length of wheat seeds and reduced the accumulation of Cd in shoots. And Duan et al (2021) found the lower size microplastics had better performance in inhibiting the Cd toxicity to the embryonic development of zebrafish. Microplastics could affect the bioavailability of heavy metals by adsorption and changing soil properties. Nicole et al (2017) through a column filtration test showed that the aged microplastics can not only increase the adsorption of Cu, Zn and Ca, but also weakened their desorption. And results found that microplastics could promote the transformation of metals from bioavailable to organic-bound species, thus decreasing the bioavailability of metals (Wang et al., 2020b, Yu et al., 2020, 2021). In addition, Lian et al (2020) found that microplastics could resist Cd toxicity by elevating the metabolism of carbohydrates and amino acid in wheat. However, some studies had different results. Hodson et al (2017) found that microplastics could increase Zn bioavailability in soils. Zhang et al (2019) also found that microplastics increased metal toxicity in water environments. And Liao and Yang (2022) established a digestive in-vitro method and noticed that the loaded Cd on microplastic could release in the animal body, which provides a new view on the potential environmental hazard of microplastics and heavy metals on soil animals. Therefore, the long-term effects of co-microplastics and Cd in soils are suggested to be further studied.

In our studies, the single and combined effects of degradable microplastics and Cd were investigated through a standard single species test and biochemical assays by F. candida. The results showed that single and co-microplastics and Cd all had negative influences on the survival, reproduction and growth of F. candida, and the effects intensified with microplastics concentrations. But combined microplastic and Cd alleviated the toxicity of single chemical on F. candida at lower microplastics concentrations. Biochemical assays on antioxidant enzymes had the same results. Toxicity endpoints reproduction was not more sensitive than survival and growth. Antioxidant enzymes CAT and POD was more sensitive than SOD, thus may be promising biomarkers on soil collembolan for soil microplastics exposure. The toxicity mechanism of co-microplastics and Cd on collembolan is complex, the long-term effects are suggested to be further studied.

Data availability

The data and materials presented in this study are available on request from the corresponding author.

Author contributions: Guoqiang Liu and Yonghua Liu designed the research, Guoqiang Liu and Xuanzhu Gu performed the research, Guoqiang Liu and Xuanzhu Gu analyzed and interpreted the data, Guoqiang Liu wrote original draft, Jing Wu, Haidong Li, Lianghu Su, Mei Chen, Sujuan Chen and reviewed and edited the draft.

Acknowledgement

This work was funded by supported National Natural Science Foundation of China (Grant No. 72174127).

Ethics approval and consent to participate

All procedures performed in studies involving animals were in accordance with the ethical standards. And all the listed authors have participated actively in the study.

Consent for publication

The manuscript is an original work and has not been submitted or is under consideration for publication in another journal. All the listed authors have read and agreed to the published version of the manuscript.

Competing interests

The authors declare no competing interests.

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Table 1 Two-way analysis of variance F-values of differences among treatments and exposure times (2, 4 , 7 and 14 days) of antioxidant enzyme activities (CAT, POD and SOD).

	df	CAT	POD	SOD
Treatment	7	29.0 ***	19.5***	14.4***
Time	3	36.6***	52.3***	3.57
Concentration × time	21	38.3***	167***	24.2***

*, P < 0.05, **, P < 0.01, ***, P < 0.001.

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The interaction effects of degradable microplastics and Cd to Folsomia candida soil collembolan

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Abstract

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1. Introduction

2. Materials And Methods

2. 1 Test organism

2. 2 Soil preparation

2. 3 The single species tests with soil

2. 4 Biochemical assays

2. 5 Statistical analysis

3. Results

3. 1 The effects on collembolan survival and reproduction

3. 2 The effects on collembolan growth

3. 2 The effects on the antioxidant systems of collembolan

4. Discussion

4. 1 Survival, reproduction and growth

4. 2 Antioxidant enzymes

5. Conclusions

Declarations

References

Tables

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