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Article

Endogeic Earthworms Avoid Soil Mimicking Metal Pollution Levels in Urban Parks

by
Marion Chatelain
1,2
1
Department of Zoology, University of Innsbruck, Technikerstraβe 25, 6020 Innsbruck, Austria
2
Institute of Ecology and Environmental Sciences—Paris, Faculté des Sciences & Ingénierie, Sorbonne Université, Campus Pierre-et-Marie-Curie, 75005 Paris, France
Sustainability 2023, 15(15), 11513; https://doi.org/10.3390/su151511513
Submission received: 12 May 2023 / Revised: 5 July 2023 / Accepted: 13 July 2023 / Published: 25 July 2023
(This article belongs to the Section Sustainable Urban and Rural Development)
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Abstract

:
In response to long-lasting high levels of metallic trace elements (MTEs) in urban soils, we expect soil invertebrates inhabiting urban environments to have evolved detection and avoidance and/or tolerance mechanisms to MTE pollution. In this study, I used artificial soils with concentrations of lead, zinc, copper, chromium and nickel that reflect pollution levels in the soils of Parisian parks. Using choice experiments, I compared habitat preference (i.e., the occurrence of individuals in the polluted vs. unpolluted soil) and health status (i.e., body mass maintenance, mobility, mortality) between three species of endogeic earthworms—Aporrectodea caliginosa, Aporrectodea icterica and Allolobophora chlorotica—originating either from urban or rural grasslands. This study highlights a clear avoidance of MTE-polluted soils in all three species, as well as MTE-induced health impairments, especially in A. chlorotica. Interestingly, earthworm response to MTE exposure only slightly differed between earthworms of urban and rural origin, suggesting the absence of widespread acclimatization or adaptation mechanisms to MTE pollution in cities. As a consequence, MTE pollution is expected to significantly shape earthworm spatial distribution in both urban and rural environments and, as a consequence, affect ecosystem functioning.

Graphical Abstract

1. Introduction

Metallic trace elements (MTEs) are particularly abundant in urban soils; lead (Pb), copper (Cu), zinc (Zn), chromium (Cr), nickel (Ni), cadmium (Cd), mercury (Hg) and arsenic (As) are the main MTE pollutants in urban soils [1,2]. For instance, Pb concentrations can be fourteen times higher in urban than in rural soils [3]. For these reasons, soil organisms are likely to be affected by MTE pollution in urban soils. Yet, soil invertebrates fulfil numerous soil-based ecosystem services such as organic matter decomposition, regulation of microbial activity, soil structure, etc. [4,5]. Amongst others, earthworms are of primary importance for ecosystem functioning by modifying the availability of resources for other organisms through physical and chemical changes in their surrounding soil environment [6]. Therefore, understanding how soil invertebrate populations respond to MTE pollution in urban soils is key to comprehending variations in soil ecosystem functioning in an increasingly urbanized world.
Both experimental and correlative studies at industrially and geologically MTE-contaminated sites demonstrated profound effects of soil MTE pollution on soil invertebrates. For instance, MTE soil pollution is associated with reduced density, biomass, diversity index and richness of soil invertebrates, including earthworms, close to smelters or mining sites [7,8,9,10]. Earthworms were virtually absent in a 2 km radius around the smelter in Avonmouth (England), likely as a result of soil Zn pollution [11,12]. Moreover, experimental studies on Eisenia fetida and Eisenia andrei demonstrated negative effects of Cu, Cd, Cr, Pb, Ni and Zn exposure on survival, body mass maintenance, cocoon production and viability, growth or sexual development [11,12,13,14,15,16]. However, MTE concentrations at industrial sites are usually extremely high (e.g., 28,000 ppm Pb [17], 2500 ppm Cu [18], 19,000 ppm Zn [9]. Similarly, MTE concentrations used in experimental studies often exceed the mean concentrations measured in this study in Parisian gardens [11]. Maybe more importantly, toxicity assessments are based on exposure to a single MTE and do not account for synergetic, antagonistic or additive effects between MTEs [19]. Therefore, while MTE pollution may contribute to the lower species richness and the altered community composition of invertebrates in cities relative to less disturbed habitats [20,21], the effects of soil MTE pollution in urban environments on soil invertebrates have not been directly tested so far.
Soil MTE pollution may affect soil invertebrate spatial distribution by influencing habitat preference/avoidance [22,23]. Avoidance behavior toward MTE-polluted habitats varies between species of invertebrates [24]. For instance, several species of earthworms were shown to avoid Cu- and Zn-enriched soils, although the concentration threshold determining departure was species-specific [25,26]; interestingly, the threshold concentration for the avoidance of As contaminated soil was lower in individuals originating from uncontaminated soil, suggesting either acclimatization or adaptation mechanisms [27]. The evolution of avoidance behavior, which implies the capacity to detect MTEs and to move away, depends on the cost of exploration and/or dispersal [28]. In fragmented landscapes, such as urban environments [29], the energy expenditure and mortality risk associated with dispersal can be particularly high [28,30]. Populations may then evolve alternative strategies to overcome the adverse influence of otherwise toxic MTE concentrations, which strategies involve the absorption, immobilization and/or excretion of MTEs. The evolution of MTE-tolerant genotypes is well-known in vascular plants [31,32]. In invertebrates, MTE exposure, although not in the context of urbanization, is associated with genetic and phenotypic variations [33,34,35]. For instance, wild populations of common fruit fly Drosophila melanogaster that have been exposed to MTE pollution are endowed with a duplicated gene coding for metallothionein (Mtn), a protein involved in the binding, transport and detoxification of MTEs; this confers individuals a better tolerance to Cu and Cd exposure [36]. In the earthworm Dendrobaena octaedra, offspring had a higher survival rate when exposed to elevated cadmium concentrations if their parents originated from MTE-polluted than from unpolluted forests [37]. Experimentally comparing populations exposed, in their natural environment, to different levels of MTE pollution is an interesting avenue to gain insight into the effects of urbanization on soil invertebrates and to identify adaptation and/or acclimatization mechanisms shaping spatial distribution in cities [38].
Using soils with MTE concentrations mimicking MTE pollution levels in urban parks, I compared the effect of MTE soil pollution on habitat preference between three species of endogeic earthworms, namely Aporrectodea caliginosa, Aporrectodea icterica and Allolobophora chlorotica, originating either from urban or rural grasslands. The aim of this study was two-fold: to (1) understand whether soil MTE pollution explains earthworm movements and habitat selection and (2) test whether habitat preference depends on earthworm previous exposure to MTE-polluted soil. Earthworm response to soil MTE pollution was also characterized by measuring mortality, mobility, body mass maintenance and MTE accumulation.

2. Methods

2.1. Subjects and Housing

Free-living adult earthworms from three endogeic species—Aporrectodea caliginosa, Aporrectodea icterica and Allolobophora chlorotica (green morph)—were collected from February to May 2016 in gardens, squares or grasslands at 16 urban sites in Paris and 8 rural sites across France (urban and rural sites categorized based on land use; see Table S1). At each urban site, they were collected from three different locations less than 200 m apart. The rural and urban soils (0–30 cm deep) were similar in terms of physico-chemical characteristics other than MTEs (i.e., pH, total carbonates, total organic carbon, organic matter, organic nitrogen, electrical resistivity and assimilable phosphorus; see Table S2), except for the proportion of bigger particles (>6.3 mm) being more abundant in rural than in urban sites (see Table S3).
Earthworms were identified in the lab using morphological characteristics [39,40]. To reduce potential stress effects of capture on earthworm behavior, earthworms were kept in acclimatizing terrariums (12 cm × 10 cm × 8 cm) filled with suitable soil (i.e., grassland soil sampled from a brown earth at the Research Institute for Development (Bondy, France; 48°54′ N, 2°29′ E) hosting large earthworm populations); they were kept at a maximum density of 10 earthworms per terrarium, at 20 °C during the day and night. Acclimatization lasted at least 3 weeks but varied between individuals depending on their capture and trial dates. Twice a month, acclimatizing terrariums were humidified and enriched with a cupped hand of homogenized horse manure spread on the surface of the soil. At least a week before the start of the trial, earthworms were transferred in MTE-less control soil (see Section 2.2) enriched with homogenized horse manure. Each individual earthworm was used only once during the experiment. The species names used herein are conformed to the Fauna Europaea web site (https://fauna-eu.org/index.php, accessed on 8 May 2023).

2.2. Artificial Soil Preparation

To ensure that MTE-enriched and MTE-less soils differed only in MTE concentrations, artificial soils were created; they consisted of 70% sand, 20% clay and 10% sphagnum. Forty-eight hours before the start of the trial, dry soil was transferred into soil preparation buckets where soils were humidified up to 25% of the dry mass with distilled water (M−) or MTE-enriched solutions (chromium sulphate, copper sulphate, lead acetate, nickel acetate and zinc chloride; Sigma-Aldrich, St-Louis, MO, USA) to obtain an MTE-rich soil (M++: Cr 10 ppm, Cu 100 ppm, Pb 300 ppm, Ni 15 ppm and Zn 245 ppm) and an MTE-poor soil (M+: half concentrated compared to the MTE-rich soil) (Figure 1). MTE concentrations were chosen to mimic average MTE levels in soils of public green spaces in Paris (personal data from Florence Dubs collected in 2015), which concentrations are in the range of concentrations measured in urban soils worldwide [1,2,41]. MTE salts were chosen for their solubility in water. Although not significantly so, MTE-enriched soils (both M++ and M+) had systematically lower pH, carbon-to-nitrate ratio and assimilable phosphorus (see Table S2), while the pH (from 5.42 to 6.55) always remained very close to the 6.0 ± 0.5 pH advised in the OECD guideline for the testing of chemicals, the availability of MTEs in this experiment was likely higher than it would have been in a neutral to slightly alkaline soil [42,43].

2.3. Terrarium Set-Up for Behavioral Trials

Behavioral trials were carried out in terrariums (25 cm × 25 cm × 1 cm) with transparent walls to allow easy location of the earthworms and observation of the galleries (see Section 2.4). The soils were slightly compacted whenever needed to obtain a visually homogeneous and similar soil density among and between terrariums.
Set-up for choice trials: To test whether earthworm habitat preference depends on MTE exposure, the two sides of the terrariums were filled separately with the MTE-less soil and one of the two MTE-enriched soils (M−/M++ or M−/M+); the two soils were held apart by a divider which was subsequently removed; they were alternatively on the right or on the left of the terrarium. A total of 96 choice trials were carried out: eight trials per species (A. caliginosa, A. icterica and A. chlorotica) per origin (urban and rural) per MTE exposure (M−/M+ and M−/M++) (Figure 1).
Set-up for control trials: To ensure that earthworm location within the terrarium results from a choice rather than from impaired mobility caused by MTE exposure, both sections were filled with one of the MTE-enriched (M+/M+ or M++/M++) or the MTE-less soil (M−/M−; Figure 1). A total of 36 control trials were carried out: two per species per origin per MTE exposure.

2.4. Trial Procedure: Movement and Body Mass Maintenance Measurements

In order not to influence earthworm choice for one of the two sides of the terrariums, a hole of 2 cm of depth was perpendicularly dug from the top of the soil, such as it was in the middle of the terrarium. For each trial, a single worm was introduced to the hole, the anterior end first (Figure 1). The terrarium was immediately closed and left undisturbed in a dark room at 20 °C. During five consecutive days, earthworm location was marked (i.e., a cross was drawn on the terrarium at the middle of the earthworm’s body) every two hours from 9 a.m. to 5 p.m., resulting in 23 to 25 location marks per trial. At the end of both control and choice trials, the galleries were traced on both sides of the terrarium; a picture of the empty terrarium—allowing us to simultaneously see the galleries on both sides—was taken. Total gallery length in the MTE-enriched and MTE-less soils (i.e., the total length of all galleries formed on each side of the terrarium) was measured using the SmartRoot package in ImageJ [44]. Each earthworm was weighed before and at the end of a trial to measure mass change, calculated as the difference between post and pre-trial mass divided by pre-trial mass. At the end of the choice trials, earthworms were moved back into an acclimatizing terrarium; they were released into their environment at the end of the study. Earthworms from the control trials were individually kept fasting for 24 h in a Petri dish containing a wet sponge, and then froze at −20 °C until MTE analysis (see Section 2.6).

2.5. Movement Analyses

Earthworm movements were measured using five variables: space occupation and activity (in control trials) and specificity and soil choice (in choice trials). For each control trial (M−/M−, M+/M+ and M++/M++), space occupation was measured by counting the number of locations on the left and the right section of the terrarium to verify that earthworm movements were homogeneous, hence not biased, within the terrarium. Activity was measured by (i) counting the number of non-overlapping location marks (hereinafter referred to as “mobility”) and (ii) measuring the total gallery length. For each choice trial (M+/M− and M++/M−), specificity for the MTE-less soil was defined as 0 when the individual visited the MTE-enriched soil at least once and as 1 when all the locations were in the MTE-less soil. Moreover, soil choice was measured by (i) counting the number of locations and (ii) measuring the size of the galleries in the MTE-enriched soil (M+ or M++) and in the MTE-less soil (M−); the locations in the middle of the terrarium were excluded.

2.6. MTE Analyses

In earthworm native soils: To ensure that wild earthworms collected in rural and urban environments were exposed to different levels of MTE pollution, the soil of each site was collected, and MTE concentrations (i.e., Cr, Cu, Pb, Ni and Zn) were measured. For urban sites, soil was collected at three different locations not more than 200 m apart. Briefly, a 30 × 30 × 30 cm block of soil was extracted and from which at least 1 L of soil was sampled from the topsoil (from 0 to 5 cm below the surface) and from the subsoil (from 10 to 30 cm below the surface), separately. MTE analyses were carried out by the Laboratoire d’Agronomie de la ville de Paris (France): soil samples were dried at 40 °C, 2 mm sieved and 150 µm ground with a ball mill (MM400, Retsch, Haan, Germany). A total of 0.5 g per sample was then digested simultaneously in 1.2 mL HNO3 (65%) and 3.5 mL HCl (37%) for 16 h at room temperature, and then 2 h at 100 °C. Ultrapure water was added to reach a final volume of 50 mL and left to decant for 8 h. The total content of Cr (284.325 nm), Cu (324.754 nm), Pb (220.353 nm), Ni (231.604 nm) and Zn (213.856 nm) was determined using an inductively coupled plasma optical emission spectrometer (iCAP 7000 ICP-OES, Thermo Electron, Waltham, MA, USA). Details on the measurements are in the Supplementary Material. To estimate earthworm exposure to MTEs at each site, MTE concentrations in the topsoil and the subsoil were averaged. For urban sites, MTE concentrations at the three locations, less than 200 m apart within the site, were averaged.
In artificial soils: To test the validity of the MTE supplementation protocol, the MTE-less (M−), -poor (M+) and -rich soils (M++) were sampled twice during the study. Forty-eight hours after their preparation, ca. 1 L of soil was taken from the soil preparation buckets. MTE concentrations in artificial soil samples were measured using the same protocol as for native soil samples.
In earthworms: Earthworms that were used in the control trials were defrosted and dried for 48 h at 50 °C. They were weighed to the nearest 0.1 mg and then digested simultaneously in 1 mL HNO3 (68%) and 1 mL HF (40%) during 24 h at 80 °C. The product of digestion was transferred into plastic tubes, and ultrapure water was added to reach a final 1% acid concentration. The total content of Cr (52Cr and 53Cr), Cu (63Cu and 65Cu), Pb (206Pb, 207Pb and 208Pb), Ni (60Ni and 61Ni) and Zn (66Zn and 68Zn) was determined using an inductively coupled plasma mass spectrometer (NexION 300D ICP Mass Spectrometer, Perkin Elmer, Waltham, MA, USA) in the Biological and Chemical Research Centre (Faculty of Chemistry, University of Warsaw, Poland). A conventional Mainhardt nebulizer and a quartz cyclonic spray chamber were used for sample introduction. Details on the measurements are in the Supplementary Materials.

2.7. Statistical Analyses

Statistical analyses were performed using R software (version 3.5.1).
MTE concentrations in natural soils: To ensure that the earthworms collected in urban and rural soils were in their natural environment and exposed to different levels of MTE pollution, the Cr, Cu, Pb, Ni and Zn concentrations in urban and rural soils where earthworms have been collected were compared using linear models (R built-in ‘lm’ function) with the log-transformed MTE concentration as the dependent variable and the origin (urban vs. rural) as the explanatory variable.
MTE-induced health impairments (control trials): To investigate MTE exposure health effects on earthworms, I performed linear mixed-effects models (‘lmer’ function from the ‘lme4′ package [45]) with either (i) mobility, (ii) log-transformed gallery length or (iii) mass loss as the dependent variable, and MTE exposure (M−, M++ or M++), earthworm origin (rural vs. urban) and species (A. caliginosa, A. chlorotica and A. icterica), and their two-way interactions as explanatory variables; the low sample size (i.e., two individuals per species per MTE exposure) prevented testing for the three-way interaction. The capture site was added as a random intercept. Moreover, to ensure that earthworm movements were homogeneous within the terrarium, I performed a generalized mixed-effects model (‘glmer’ function from the ‘lme4’ package) with the number of locations in the left section over the total number of locations (binomial distributions) as the dependent variable, and MTE exposure, earthworm origin and species, and their two-way interactions as the explanatory variables. The capture site was added as a random intercept. The trial identity was also added as a random intercept to correct for overdispersion.
MTE accumulation in earthworms (control trials): To test whether MTE accumulation differed in response to MTE exposure and depending on the species and the origin of the earthworm, I performed similar models as previously described to test MTE-induced health impairments, with log-transformed MTE concentrations (Cr, Cu, Pb, Ni or Zn) as explanatory variables.
MTE-linked habitat preference (choice trials): To explain the variability in earthworms’ movements in response to MTE exposure, I performed generalized mixed-effects models (‘glmer’ function) with the proportion of locations or galleries in the MTE-enriched soil (M+ or M++) as the dependent variable (binomial distributions), and MTE exposure (M+/M− vs. M++/M−), earthworm origin and species, and their two- and three-way interactions as the explanatory variables. The capture site was added as a random intercept. The trial identity was also added as a random intercept to correct for overdispersion. More than half of the earthworms did not visit the MTE-enriched soil. Visiting or not the MTE-enriched soil may reflect different habitat selection strategies (e.g., strict avoidance of MTE-polluted soils associated with high sensory capacities vs. tolerance to MTE pollution associated with a benefit for exploratory behavior). Therefore, in a second step, I analyzed the behaviour of earthworms that visited the MTE-less soil only and the ones that visited both soils, separately. Variation in specificity for the MTE-less soil was investigated using a similar generalized mixed-effects model as described above. However, the low sample size prevented testing for the three-way interaction. In individuals that visited both MTE-less and MTE-enriched soils, I investigated both soil choices (i.e., the proportion of locations or galleries in the MTE-enriched soil) using a similar generalized mixed-effects model as described above and learning—defined as the variation of earthworm location between the MTE-less and -enriched soils over the course of the trial—using a generalized mixed-effects model with earthworm location (in MTE-less or MTE-enriched soil) as the dependent variable (binomial distribution) and time (the number of location marks; from 1 to 23), MTE exposure, earthworm origin and species, and the two-way interactions with time as the explanatory variables. The capture site and the trial identity were added as random intercepts. The low sample size prevented testing for other interactions between the explanatory variables.
Linear mixed-effects models and generalized mixed-effects models were fitted using the restricted maximum likelihood (REML) and the Laplace approximation of the maximum likelihood methods, respectively. For each model, I performed a backward stepwise selection using the AIC. A Type III Wald chi-square test ANOVA was used to determine the significance of the retained variables in the final models. Contrasts among groups were tested using least-square mean pairwise comparisons (contrast function of the ‘lsmeans’ package [46]).

3. Results

3.1. MTE Concentrations in Natural and Artificial Soils

In natural soils: Cu and Pb concentrations were higher in urban than in rural soils (F = 5.27, p = 0.032 and F = 8.61, p = 0.008, respectively). On the contrary, Cr concentrations were lower in urban than in rural soils (F = 8.02, p = 0.010). No significant difference in Zn and Ni concentrations was measured (F = 3.33, p = 0.082 and F = 0.55, p = 0.466, respectively). However, Zn concentrations in 12 out of 16 urban sites were higher than the median Zn concentration in rural sites (82.5 ppm). Moreover, one of the rural sites had Zn concentrations 2.33 times higher than the mean Zn concentration in the other rural sites. When removing this point, Zn concentrations were higher in urban than in rural soils (F = 5.32, p = 0.031; Figure 2).
In artificial soils: As intended, MTE concentrations were higher in M++ than in M+, which in turn were higher than in M−. Except for Cr and Pb concentrations in M+, the enrichment was lower than intended (Table 1).

3.2. Space Occupation Homogeneity (Control Trials)

The proportion of locations on the left section of the terrarium varied from 0 to 1 with a mean ± se of 0.55 ± 0.06. This means that, on average, 55% of the locations were in the left section and 45% in the right section. Such a proportion differed between species (χ2 = 8.94, p = 0.011): it was lower in A. caliginosa (mean ± se = 0.38 ± 0.10) than in A. chlorotica (mean ± se = 0.79 ± 0.11; z = −2.99, p = 0.008). A. icterica space occupation was the most homogeneous (mean ± se = 0.53 ± 0.33).

3.3. MTE-Induced Health Impairment (Control Trials)

Three earthworms from A. chlorotica (two of rural and one from urban origin) died when exposed to M++; they were removed from the analysis. None of the explanatory variables were retained in the model investigating earthworm mobility variability. Gallery length tended to depend on the interaction between MTE exposure and species (χ2 = 8.80, p = 0.066): in A. icterica, gallery length was shorter in M++ than in M+ (t = 3.48, p = 0.046) and M− (t = 3.60, p = 0.036). The trend was similar in A. caliginosa and A. chlorotica, although non-significantly so (Figure 3a). Mass loss was significantly different between the three MTE exposures (χ2 = 15.56, p < 0.001): it was higher in M++ than in M+ (t = 3.01, p = 0.014) and M− (t = 3.51, p = 0.004; Figure 3b).

3.4. MTE Accumulation in Earthworms (Control Trials)

Cu, Pb and Zn concentrations in earthworms varied between MTE exposure (χ2 = 69.29, p < 0.001; χ2 = 101.79, p < 0.001 and χ2 = 15.97, p < 0.001, respectively): Cu and Pb were higher in earthworms exposed to M++ than to M+ (t = 3.71, p = 0.023 and t = 3.51, p = 0.005, respectively), which in turn were higher than in M− (t = 4.81, p < 0.001 and t = 6.88, p < 0.001, respectively). Zn was higher in earthworms exposed to M++ than M+ and M− (t = 3.90, p = 0.003 and t = 2.92, p = 0.022, respectively); Zn concentrations were not significantly different between earthworms exposed to M+ and M− (t = 0.89, p = 0.655). Moreover, Zn concentrations differed between species (χ2 = 9.82, p = 0.007): they were higher in A. caliginosa than in A. icterica (t = 3.11, p = 0.011). None of the explanatory variables were retained in the models investigating Cr and Ni concentration variability. Earthworm origin (rural vs. rural) was retained in none of the models (Figure 4).

3.5. MTE-Linked Habitat Preference (Choice Trials)

Two individuals from A. chlorotica died during the choice trials. They died after spending 3.5 days in M− then 1 day in M++, and 5 days in M++, respectively.
The proportion of locations in the MTE-enriched soil (M+ or M++) varied from 0 to 1; on average, 20% of the locations were in the MTE-enriched soil (M++ or M+; mean ± se = 0.20 ± 0.03). This proportion depended on MTE exposure (χ2 = 17.71, p < 0.001): it was lower when earthworms were exposed to M++/M− than to M+/M− (z = −4.21, p < 0.001; Figure 5a). Similarly, the proportion of galleries in the MTE-enriched soil varied from 0 to 1; on average, 24% of the galleries were in the MTE-enriched soil (M++ or M+; mean ± se: 0.24 ± 0.03). This proportion depended on MTE exposure (χ2 = 20.20, p < 0.001): it was lower when earthworms were exposed to M++/M− than to M+/M− (z = −4.49, p < 0.001; Figure 5b).
In 54 out of 96 choice trials, earthworms were observed in M− only, meaning that in those trials, none of the locations were in the MTE-enriched soil. Specificity for M− depended on MTE exposure (χ2 = 10.82, p = 0.001): it was higher in trials with M++ than in trials with M+ as an alternative soil (z = 3.29, p < 0.001). Specificity for M− also differed between species (χ2 = 6.35, p = 0.015): it tended to be lower in A. icterica than in A. chlorotica (z = −2.30, p = 0.056) and A. caliginosa (z = −2.13, p = 0.084; Figure 6). In 6 out of 95 trials, earthworms were observed in the MTE-enriched soil only: five and one in M+ and M++, respectively. Four were A. chlorotica (two from each origin), and two were A. icterica (one from each origin).
Thirty-five earthworms visited both M− and MTE-enriched soils (M+ or M++). In these individuals, the proportion of locations in the MTE-enriched soil varied from 0.05 to 0.91; on average, 37% of the locations were in the MTE-enriched soil (mean ± se = 0.37 ± 0.04). This proportion depended on the interaction between MTE exposure and earthworm origin (χ2 = 5.36, p = 0.021): in earthworms of urban origin, it was lower in individuals exposed to M++/M− than to M+/M− (z = −2.70; Figure 7a). Similarly, the proportion of galleries in the MTE-enriched soil varied from 0.04 to 0.99; on average, 36% of the galleries were in the MTE-enriched soil (mean ± se: 0.36 ± 0.04). This proportion depended on MTE exposure (χ2 = 6.51, p = 0.011): it was lower in earthworms exposed to M++/M− than to M+/M− (z = −2.55, p = 0.011; Figure 7b). The proportion of galleries in the MTE-enriched soil also varied between species (χ2 = 8.80, p = 0.012): it was lower in A. icterica than in A. chlorotica (z = −2.70, p = 0.011) and A. caliginosa, although not significantly so (z = −2.16, p = 0.077). Finally, earthworm location depended on the interaction between time (i.e., the number of location marks) and species (χ2 = 12.86, p = 0.002): the slope between earthworm location in the MTE-enriched soil and time tended to be slightly negative in A. icterica (r = −0.10, t = −1.89, p = 0.06), slightly positive in A. chlorotica (r = 0.10, t = 1.81, p = 0.072) but not significantly different from 0 in A. caliginosa (r = 0.09, t = 1.45, p = 0.15). However, the post hoc model failed to detect significant differences between the three slopes (P > 0.368; Figure 7c). Earthworm location also depended on the interaction between time and site (χ2 = 6.16, p = 0.013): the slope between earthworm location in the MTE-enriched soil and time tended to be slightly positive in earthworms of urban origin (r = 0.09, t = 1.76, p = 0.079), while it was not significantly different from 0 in earthworms of rural origin (r = −0.07, t = −1.26, p = 0.209). However, again, the post hoc model failed to detect significant differences between the two slopes (p = 0.176; Figure 7d).

4. Discussion

4.1. MTE Exposure Decreases Body Mass Maintenance

MTE pollution at levels similar to the ones measured in Parisian parks had significant effects on some of the health proxies measured in this study. First, earthworms exposed to M++ lost more weight over the course of the trial than earthworms exposed to M−; however, mass loss was not significantly different between individuals exposed to M+ and M−. Second, three individuals from A. chlorotica died while exposed to M++, and two others died during the choice trial after visiting M++. Third, earthworms built shorter galleries when exposed to MTEs; however, this pattern was significant for A. icterica only. Earthworm mobility was unaffected by MTE concentrations in the soil. However, the death of three out of the four individuals from A. chlorotica exposed to M++ prevented the assessment of the effect of M++ exposure on mobility in this species. Still, the fact that four individuals from A. chlorotica were found in the MTE-enriched soil only during the choice trials strongly suggests that MTE exposure hindered earthworm movements.
All in all, this study showed that earthworm health was impaired in M++, meaning that earthworms were sensitive to a cocktail of around 70 ppm Cu, 240 ppm Pb, 8 ppm Ni and 190 ppm Zn. Cu, Pb and Zn concentrations in M++ were largely similar to the levels measured in Square des Batignolles and were lower than concentrations measured in four of the 16 Parisian parks (see Table S1). Moreover, these concentrations were lower than the average concentrations in urban areas [1,2]. For instance, out of 86 cities where Pb concentrations in the soil have been measured, 14 and 12 have concentrations higher than 200 and 500 ppm, respectively [2]. While MTE concentrations in urban soils can vary between and within cities [2], these results suggest that urban green spaces can be poor-quality habitats for earthworms, especially for some species such as A. chlorotica. This study underlines the importance of conducting more studies investigating the effects of cocktails of “low” concentrations of MTEs on soil invertebrates to better estimate acceptable MTE levels in soils [47]. Moreover, comparing earthworm sensitivity to MTE exposure in cities more or less polluted with MTEs is an interesting avenue to investigate whether earthworm susceptibility can vary as a response to MTE exposure. Measuring differences in earthworm susceptibility may reveal the evolution of tolerance mechanisms; alternatively, the absence of variation in earthworm susceptibility would suggest a strong impact of MTE pollution on population dynamics and viability in urban environments.

4.2. MTE Exposure Strongly Affects Habitat Preference

On average, earthworms were detected only 1 time out of 5 in the MTE-enriched soil. Importantly, the percentage of occurrence in the MTE-enriched soil dropped from around 35% in the M+/M− trials to only 10% in the M++/M− trials. This result underlines the fact that, even though MTE concentrations in M+, which reproduce MTE pollution in Paris squares and parks fairly well (see Figure 2), did not hinder earthworm survival, mobility and body mass maintenance (see Section 4.1), they did significantly influence earthworm habitat preference. Previous studies also showed that earthworms avoided MTE-contaminated soils even though MTE concentrations were below the mortality and reproduction endpoints [48,49,50]. The evolution of such MTE exposure avoidance probably results from adverse effects on health proxies that were not measured in this study (e.g., immunity, oxidative balance, etc. [51]).
Interestingly, after inoculation, a large proportion of earthworms exclusively visited M−; this proportion increased with increasing MTE concentrations in the alternative soil. This result suggests that habitat preference in earthworms relies on high sensory capacities. Indeed, the presence of chemoreceptors and sensory tubercles makes earthworms highly sensitive to chemicals in the environment [52,53,54]. While the mechanisms by which earthworms discriminate soils with different MTE pollution levels are poorly understood, earthworm avoidance of Zn contamination would result from a direct effect of Zn2+ ions on epidermal chemosensitive receptors [55]. Surprisingly, in individuals that did visit the MTE-enriched soil, the visiting frequency of this soil did not decrease over time. This suggests that earthworm avoidance of MTE-polluted soil does not result from a learning process but rather from an innate mechanism. Overall, MTE pollution-linked habitat preference strongly suggests that MTE pollution at levels measured in urban parks is likely to affect earthworm distribution and, consequently, urban soil services.

4.3. Response to MTE Exposure Only Slightly Differs between Species

Five individuals from A. chlorotica died when exposed to M++ or M++/M−, while no losses were recorded in the other two species. This suggests that A. chlorotica was more sensitive to MTE exposure than the two other species. This variability in species susceptibility to MTE pollution could be due to the size of the earthworms. Indeed, the three species used in this experiment vary greatly in mass; from the smallest to the largest: A. chlorotica (≈300 ± 20 g), A. caliginosa (≈420 ± 30 g) and A. icterica (≈740 ± 40 g, calculated from earthworm mass before the trials. Similarly, smaller earthworm species were more sensitive to cadmium exposure than larger species (i.e., they suffered from higher DNA damage; [56]). More generally, sensitivity to xenobiotics decreases with increasing body size in invertebrates [57,58]. This could be due to the fact that small organisms absorb greater quantities of toxicants through their epidermis, due to their greater body surface area relative to their volume, compared with larger organisms [59]. Surprisingly, the high sensitivity of A. chlorotica to MTE exposure did not translate into a stronger choice for M− compared to the other two species.
Overall, A. caliginosa, A. icterica and A. chlorotica exhibited similar behavioural responses to MTE exposure: the three endogeic species preferred the MTE-less soil to the MTE-enriched soils. While the behavioral response to MTE exposure differed between A. icterica and the other two species, the differences were not consistent between the variables that were measured. Previous studies have measured species differences in the avoidance behavior of earthworms toward MTE-contaminated soils. For instance, avoidance behavior toward MTE pollution was lower in epigeic than in anecic and endogeic species, suggesting that epigeic species would be less sensitive to MTEs [25,48], but see [60]. Comparing habitat preferences in response to urban soil pollution between species with different ecological characteristics—exposed differently to MTE pollution and with different mobility and/or dispersal capacities—is an interesting avenue for better understanding how MTE pollution can affect the composition of earthworm communities in cities.

4.4. Little Evidence of Evolutionary Divergence between Urban and Rural Populations

Species vary in their ability to adapt to the often-drastic physical changes along the urban–rural gradient [61]. Several invertebrate species evolved in response to urbanization, even though examples of local adaptation are still scarce [62]. Of interest, urban and rural populations of the mosquitos Anopheles coluzii and Anopheles gambiae show signs of evolutionary divergence in genes involved in xenobiotic tolerance [63]. As earthworms in urban environments are exposed to higher concentrations of potentially toxic MTEs—particularly Pb, Cu and Zn—than their counterparts in rural environments, I expected earthworms in urban habitats to have developed mechanisms for tolerating or avoiding MTE pollution [64]. In other words, I expected earthworms from urban sites to be both less sensitive to MTE exposure (i.e., to show fewer signs of altered health) and less selective when given a choice between an MTE-enriched and an MTE-less soil [27]. Alternatively, earthworms chronically exposed to MTEs could have evolved improved sensory mechanisms allowing a better detection of MTEs and an escape response when exposed to an MTE-enriched soil. Yet, this study highlighted only a few differences in earthworm response to MTE exposure between individuals from urban and rural origin: earthworms of rural and urban origin showed similar patterns of mobility, gallery length, body mass maintenance, mortality and MTE accumulation. On average, their percentage of occurrence in the MTE-enriched soil was 18% and 22%, respectively. Moreover, a similar proportion of rural and urban earthworms visited the MTE-enriched soil. However, in earthworms that visited both MTE-enriched and MTE-less soils, while individuals of rural origin preferred the MTE-less soil whatever the level of MTE pollution in the MTE-enriched soil, individuals from urban soils preferred the MTE-less soil only when given the choice with the MTE-rich soil; when given the choice with the MTE-poor soil, earthworms were detected almost one out of two times in the MTE-enriched soil. I expect the variation in earthworm selectivity to increase with decreasing MTE pollution levels. Conducting similar habitat preference experiments while using lower pollution levels than the ones tested in this study would allow us to identify the “No Observed Effect Concentration” (NOEC), meaning the maximum MTE pollution level at which earthworms from urban and rural populations express a choice between the MTE-enriched and -less soils. Ideally, future experiments should also measure fitness proxies to test whether earthworm habitat preference confers any fitness advantage and, therefore, could be selected.

5. Conclusions

Soil pollution with metallic trace elements (MTEs) poses a serious threat to biodiversity close to mining sites. This study confirmed that, similarly to mining sites, soils of urban green spaces contain high levels of lead, zinc and copper compared to rural grasslands. By submitting three species of endogeic earthworms to a choice experiment, this study highlighted a clear avoidance of soils spiked with MTEs at concentrations close to the ones measured in Parisian parks; such an avoidance behavior was observed even with MTE levels that did not affect earthworm survival, body mass maintenance, or mobility. Therefore, it is likely that MTE concentrations in urban soils shape earthworm distribution and densities, with significant consequences on soil services. To better understand the impact of urban soil pollution on soil ecosystems, future research should focus on comparing earthworm response to soil pollution between different functional groups and using different pollution levels and soil physico-chemical properties (e.g., pH and OM) that mimic as best natural conditions in urban areas. Experimental studies coupled with community analyses will allow us to reveal the causal link between soil pollution heterogeneity and variations in earthworm communities.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/su151511513/s1. A: Earthworm sampling sites & Soil characteristics; Table S1. Localisation of earthworm sampling sites. Table S2. Chemical parameters of artificial and natural soils. Table S3. Particle size of artificial and natural soils. B: Metallic trace element analyses.

Funding

This work was supported by the Austian Science Fund (FWF) [grant numbers M2628-B25].

Institutional Review Board Statement

Ethical review and approval were waived for this study as earthworms are not animals covered by the rules governing animal experimentation in France (Article R214-87 in section 6 of the “Code rural et de la pêche maritime”).

Data Availability Statement

The data presented in this study are openly available in OSF at 10.17605/OSF.IO/MEU3C.

Conflicts of Interest

The author declares no competing interest.

References

  1. Biasioli, M.; Barberis, R.; Ajmonemarsan, F. The Influence of a Large City on Some Soil Properties and Metals Content. Sci. Total Environ. 2006, 356, 154–164. [Google Scholar] [CrossRef]
  2. Ajmone-Marsan, F.; Biasioli, M. Trace Elements in Soils of Urban Areas. Water Air Soil Pollut. 2010, 213, 121–143. [Google Scholar] [CrossRef]
  3. Roux, K.E.; Marra, P.P. The Presence and Impact of Environmental Lead in Passerine Birds along an Urban to Rural Land Use Gradient. Arch. Environ. Contam. Toxicol. 2007, 53, 261–268. [Google Scholar] [CrossRef] [PubMed]
  4. Jones, C.G.; Lawton, J.H.; Shachak, M. Organisms as Ecosystem Engineers. In Ecosystem Management; Springer: New York, NY, USA, 1994; pp. 130–147. ISBN 978-0-387-94667-2. [Google Scholar]
  5. Jones, C.G.; Gutiérrez, J.L.; Byers, J.E.; Crooks, J.A.; Lambrinos, J.G.; Talley, T.S. A Framework for Understanding Physical Ecosystem Engineering by Organisms. Oikos 2010, 119, 1862–1869. [Google Scholar] [CrossRef]
  6. Blouin, M.; Hodson, M.E.; Delgado, E.A.; Baker, G.; Brussaard, L.; Butt, K.R.; Dai, J.; Dendooven, L.; Peres, G.; Tondoh, J.E.; et al. A Review of Earthworm Impact on Soil Function and Ecosystem Services: Earthworm Impact on Ecosystem Services. Eur. J. Soil Sci. 2013, 64, 161–182. [Google Scholar] [CrossRef]
  7. Bengtsson, G.; Tranvik, L. Critical Metal Concentrations for Forest Soil Invertebrates: A Review of the Limitations. Water Air Soil Pollut. 1989, 47, 381–417. [Google Scholar] [CrossRef]
  8. Lévêque, T.; Capowiez, Y.; Schreck, E.; Mombo, S.; Mazzia, C.; Foucault, Y.; Dumat, C. Effects of Historic Metal(Loid) Pollution on Earthworm Communities. Sci. Total Environ. 2015, 511, 738–746. [Google Scholar] [CrossRef]
  9. Nahmani, J.; Lavelle, P. Effects of Heavy Metal Pollution on Soil Macrofauna in a Grassland of Northern France. Eur. J. Soil Biol. 2002, 38, 297–300. [Google Scholar] [CrossRef]
  10. Tyler, G.; Påhlsson, A.-M.B.; Bengtsson, G.; Bååth, E.; Tranvik, L. Heavy-Metal Ecology of Terrestrial Plants, Microorganisms and Invertebrates: A Review. Water Air Soil Pollut. 1989, 47, 189–215. [Google Scholar] [CrossRef]
  11. Spurgeon, D.J.; Hopkin, S.P. Effects of Cadmium, Copper, Lead and Zinc on Growth, Reproduction and Survival of the Earthworm Eisenia Fetida (Savigny): Assessing the Environmental Impact of Point-Source Metal Contamination in Terrestrial Ecosystems. Environ. Pollut. 1994, 84, 123–130. [Google Scholar] [CrossRef]
  12. Spurgeon, D.J.; Hopkin, S.P. Effects of Metal-Contaminated Soils on the Growth, Sexual Development, and Early Cocoon Production of the EarthwormEisenia Fetida, with Particular Reference to Zinc. Ecotoxicol. Environ. Saf. 1996, 35, 86–95. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  13. Neuhauser, E.F.; Loehr, R.C.; Milligan, D.L.; Malecki, M.R. Toxicity of Metals to the Earthworm Eisenia Fetida. Biol. Fertil. Soils 1985, 1, 149–152. [Google Scholar] [CrossRef]
  14. van Gestel, C.A.M.; van Dis, W.A.; Dirven-van Breemen, E.M.; Sparenburg, P.M.; Baerselman, R. Influence of Cadmium, Copper, and Pentachlorophenol on Growth and Sexual Development of Eisenia Andrei (Oligochaeta; Annelida). Biol. Fertil. Soils 1991, 12, 117–121. [Google Scholar] [CrossRef]
  15. Van Gestel, C.A.M.; Dirven-Van Breemen, E.M.; Baerselman, R.; Emans, H.J.B.; Janssen, J.A.M.; Postuma, R.; Van Vliet, P.J.M. Comparison of Sublethal and Lethal Criteria for Nine Different Chemicals in Standardized Toxicity Tests Using the Earthworm Eisenia Andrei. Ecotoxicol. Environ. Saf. 1992, 23, 206–220. [Google Scholar] [CrossRef] [PubMed]
  16. Lock, K.; Janssen, C.R. Effect of New Soil Metal Immobilizing Agents on Metal Toxicity to Terrestrial Invertebrates. Environ. Pollut. 2003, 121, 123–127. [Google Scholar] [CrossRef]
  17. Bisessar, S. Effect of Heavy Metals on Microorganisms in Soils near a Secondary Lead Smelter. Water Air. Soil Pollut. 1982, 17, 305–308. [Google Scholar] [CrossRef]
  18. Bengtsson, G.; Rundgren, S. Ground-Living Invertebrates in Metal-Polluted Forest Soils. Ambio 1984, 13, 29–33. [Google Scholar]
  19. Spurgeon, D.J.; Jones, O.A.H.; Dorne, J.-L.C.M.; Svendsen, C.; Swain, S.; Stürzenbaum, S.R. Systems Toxicology Approaches for Understanding the Joint Effects of Environmental Chemical Mixtures. Sci. Total Environ. 2010, 408, 3725–3734. [Google Scholar] [CrossRef]
  20. McKinney, M.L. Effects of Urbanization on Species Richness: A Review of Plants and Animals. Urban Ecosyst. 2008, 11, 161–176. [Google Scholar] [CrossRef]
  21. Raupp, M.J.; Shrewsbury, P.M.; Herms, D.A. Ecology of Herbivorous Arthropods in Urban Landscapes. Annu. Rev. Entomol. 2010, 55, 19–38. [Google Scholar] [CrossRef] [Green Version]
  22. Morris, D.W. Temporal Variation, Habitat Selection and Community Structure. Oikos 1990, 59, 303–312. [Google Scholar] [CrossRef]
  23. Dunning, J.B.; Danielson, B.J.; Pulliam, H.R. Ecological Processes That Affect Populations in Complex Landscapes. Oikos 1992, 65, 169–175. [Google Scholar] [CrossRef] [Green Version]
  24. Mogren, C.L.; Trumble, J.T. The Impacts of Metals and Metalloids on Insect Behavior. Entomol. Exp. Appl. 2010, 135, 1–17. [Google Scholar] [CrossRef]
  25. Lukkari, T.; Haimi, J. Avoidance of Cu- and Zn-Contaminated Soil by Three Ecologically Different Earthworm Species. Ecotoxicol. Environ. Saf. 2005, 62, 35–41. [Google Scholar] [CrossRef]
  26. Lukkari, T.; Aatsinki, M.; Väisänen, A.; Haimi, J. Toxicity of Copper and Zinc Assessed with Three Different Earthworm Tests. Appl. Soil Ecol. 2005, 30, 133–146. [Google Scholar] [CrossRef]
  27. Langdon, C.J.; Piearce, T.G.; Meharg, A.A.; Semple, K.T. Survival and Behaviour of the Earthworms Lumbricus Rubellus and Dendrodrilus Rubidus from Arsenate-Contaminated and Non-Contaminated Sites. Soil Biol. Biochem. 2001, 33, 1239–1244. [Google Scholar] [CrossRef]
  28. Bonte, D.; Van Dyck, H.; Bullock, J.M.; Coulon, A.; Delgado, M.; Gibbs, M.; Lehouck, V.; Matthysen, E.; Mustin, K.; Saastamoinen, M.; et al. Costs of Dispersal. Biol. Rev. 2012, 87, 290–312. [Google Scholar] [CrossRef]
  29. Grimm, N.B.; Faeth, S.H.; Golubiewski, N.E.; Redman, C.L.; Wu, J.; Bai, X.; Briggs, J.M. Global Change and the Ecology of Cities. Science 2008, 319, 756–760. [Google Scholar] [CrossRef] [Green Version]
  30. Cote, J.; Bestion, E.; Jacob, S.; Travis, J.; Legrand, D.; Baguette, M. Evolution of Dispersal Strategies and Dispersal Syndromes in Fragmented Landscapes. Ecography 2017, 40, 56–73. [Google Scholar] [CrossRef] [Green Version]
  31. Singh, M.; Kumar, J.; Singh, S.; Singh, V.P.; Prasad, S.M.; Singh, M. Adaptation Strategies of Plants against Heavy Metal Toxicity: A Short Review. Biochem. Pharmacol. Open Access 2015, 04, 161. [Google Scholar] [CrossRef] [Green Version]
  32. Singh, S.; Parihar, P.; Singh, R.; Singh, V.P.; Prasad, S.M. Heavy Metal Tolerance in Plants: Role of Transcriptomics, Proteomics, Metabolomics, and Ionomics. Front. Plant Sci. 2016, 6, 1143. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  33. Posthuma, L.; Van Straalen, N.M. Heavy-Metal Adaptation in Terrestrial Invertebrates: A Review of Occurrence, Genetics, Physiology and Ecological Consequences. Comp. Biochem. Physiol. C Pharmacol. Toxicol. Endocrinol. 1993, 106, 11–38. [Google Scholar] [CrossRef]
  34. Morgan, A.J.; Kille, P.; Stürzenbaum, S.R. Microevolution and Ecotoxicology of Metals in Invertebrates. Environ. Sci. Technol. 2007, 41, 1085–1096. [Google Scholar] [CrossRef]
  35. Janssens, T.K.S.; Roelofs, D.; van Straalen, N.M. Molecular Mechanisms of Heavy Metal Tolerance and Evolution in Invertebrates. Insect Sci. 2009, 16, 3–18. [Google Scholar] [CrossRef]
  36. Maroni, G.; Wise, J.; Young, J.E.; Otto, E. Metallothionein Gene Duplications and Metal Tolerance in Natural Populations of Drosophila Melanogaster. Genetics 1987, 117, 739–744. [Google Scholar] [CrossRef] [PubMed]
  37. Rożen, A. Effect of Cadmium on Life-History Parameters in Dendrobaena Octaedra (Lumbricidae: Oligochaeta) Populations Originating from Forests Differently Polluted with Heavy Metals. Soil Biol. Biochem. 2006, 38, 489–503. [Google Scholar] [CrossRef]
  38. Beeby, A.; Richmond, L. Adaptation by an Urban Population of the Snail Helix Aspersa to a Diet Contaminated with Lead. Environ. Pollut. 1987, 46, 73–82. [Google Scholar] [CrossRef]
  39. Bouché, M.B. Lombriciens de France: Écologie et Systématique; Institut National de la Recherche Agronomique: Paris, France, 1972. [Google Scholar]
  40. Sims, R.W.; Gerard, B.M. Earthworms: Keys and Notes for the Identification and Study of the Species; No. 31 of the Synopses of the British Fauna (New Series); Kermack, D.M., Barnes, R.S.K., Eds.; The Linnean Society of London and The Estuarine and Brackish-Water Biological Association: London, UK, 1985. [Google Scholar]
  41. Manta, D.S.; Angelone, M.; Bellanca, A.; Neri, R.; Sprovieri, M. Heavy Metals in Urban Soils: A Case Study from the City of Palermo (Sicily), Italy. Sci. Total Environ. 2002, 300, 229–243. [Google Scholar] [CrossRef]
  42. Hou, S.; Zheng, N.; Tang, L.; Ji, X.; Li, Y. Effect of soil pH and organic matter content on heavy metals availability in maize (Zea mays L.) rhizospheric soil of non-ferrous metals smelting area. Environ. Monit. Assess. 2019, 191, 634. [Google Scholar] [CrossRef]
  43. Zeng, F.; Ali, S.; Zhang, H.; Ouyang, Y.; Qiu, B.; Wu, F.; Zhang, G. The influence of pH and organic matter content in paddy soil on heavy metal availability and their uptake by rice plants. Environ. Pollut. 2011, 159, 84–91. [Google Scholar] [CrossRef]
  44. Lobet, G.; Pages, L.; Draye, X. A Novel Image-Analysis Toolbox Enabling Quantitative Analysis of Root System Architecture. Plant Physiol. 2011, 157, 29–39. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  45. Bates, D.; Mächler, M.; Bolker, B.; Walker, S. Fitting Linear Mixed-Effects Models Using Lme4. J. Stat. Softw. 2015, 67, 1–48. [Google Scholar] [CrossRef]
  46. Lenth, R.V. Least-Squares Means: The R Package Lsmeans. J. Stat. Softw. 2016, 69, 1–33. [Google Scholar] [CrossRef] [Green Version]
  47. Monchanin, C.; Devaud, J.-M.; Barron, A.B.; Lihoreau, M. Current Permissible Levels of Metal Pollutants Harm Terrestrial Invertebrates. Sci. Total Environ. 2021, 779, 146398. [Google Scholar] [CrossRef] [PubMed]
  48. Lowe, C.N.; Butt, K.R.; Cheynier, K.Y.-M. Assessment of Avoidance Behaviour by Earthworms (Lumbricus Rubellus and Octolasion Cyaneum) in Linear Pollution Gradients. Ecotoxicol. Environ. Saf. 2016, 124, 324–328. [Google Scholar] [CrossRef] [Green Version]
  49. Sivakumar, S. Effects of Metals on Earthworm Life Cycles: A Review. Environ. Monit. Assess. 2015, 187, 530. [Google Scholar] [CrossRef]
  50. Syed, Z.; Alexander, D.; Ali, J.; Unrine, J.; Shoults-Wilson, W.A. Chemosensory Cues Alter Earthworm (Eisenia Fetida) Avoidance of Lead-Contaminated Soil: Chemosensory and Earthworm Avoidance. Environ. Toxicol. Chem. 2017, 36, 999–1004. [Google Scholar] [CrossRef]
  51. Spurgeon, D.J.; Weeks, J.M.; Van Gestel, C.A.M. A Summary of Eleven Years Progress in Earthworm Ecotoxicology. Pedobiologia 2003, 47, 588–606. [Google Scholar] [CrossRef]
  52. Laverack, M.S. Tactile and Chemical Perception in Earthworms —I. Responses to Touch, Sodium Chloride, Quinine and Sugars. Comp. Biochem. Physiol. 1960, 1, 155–163. [Google Scholar] [CrossRef]
  53. Laverack, M.S. Tactile and Chemical Perception in Earthworms —II. Responses to Acid PH Solutions. Comp. Biochem. Physiol. 1961, 1, 22–34. [Google Scholar] [CrossRef]
  54. Stephenson, G.L.; Kaushik, A.; Kaushik, N.K.; Solomon, K.R.; Steele, T.; Scroggins, R.P. Use of an Avoidance-Response Test to Assess the Toxicity of Contaminated Soils to Earthworms. Adv. Earthworm Ecotoxicol. 1998, 67–81. [Google Scholar]
  55. Ma, W.-C.; Bonten, L.T.C. Bioavailability Pathways Underlying Zinc-Induced Avoidance Behavior and Reproduction Toxicity in Lumbricus Rubellus Earthworms. Ecotoxicol. Environ. Saf. 2011, 74, 1721–1726. [Google Scholar] [CrossRef] [PubMed]
  56. Fourie, F.; Reinecke, S.A.; Reinecke, A.J. The Determination of Earthworm Species Sensitivity Differences to Cadmium Genotoxicity Using the Comet Assay. Ecotoxicol. Environ. Saf. 2007, 67, 361–368. [Google Scholar] [CrossRef] [PubMed]
  57. Baird, D.J.; Van den Brink, P.J. Using Biological Traits to Predict Species Sensitivity to Toxic Substances. Ecotoxicol. Environ. Saf. 2007, 67, 296–301. [Google Scholar] [CrossRef]
  58. Ippolito, A.; Todeschini, R.; Vighi, M. Sensitivity Assessment of Freshwater Macroinvertebrates to Pesticides Using Biological Traits. Ecotoxicology 2012, 21, 336–352. [Google Scholar] [CrossRef]
  59. Klaassen, C.D. Absorption, Distribution, and Excretion of Toxicants. In Casarett & Doull’s Toxicology: The Basic Science of Poisons; McGraw Hill: New York, NY, USA, 1991; pp. 50–87. [Google Scholar]
  60. Langdon, C.J.; Hodson, M.E.; Arnold, R.E.; Black, S. Survival, Pb-Uptake and Behaviour of Three Species of Earthworm in Pb Treated Soils Determined Using an OECD-Style Toxicity Test and a Soil Avoidance Test. Environ. Pollut. 2005, 138, 368–375. [Google Scholar] [CrossRef]
  61. McKinney, M.L. Urbanization, Biodiversity, and Conservation. BioScience 2002, 52, 883. [Google Scholar] [CrossRef]
  62. Johnson, M.T.J.; Munshi-South, J. Evolution of Life in Urban Environments. Science 2017, 358, eaam8327. [Google Scholar] [CrossRef] [Green Version]
  63. Kamdem, C.; Fouet, C.; Gamez, S.; White, B.J. Pollutants and Insecticides Drive Local Adaptation in African Malaria Mosquitoes. Mol. Biol. Evol. 2017, 34, 1261–1275. [Google Scholar] [CrossRef] [Green Version]
  64. Sih, A.; Ferrari, M.C.O.; Harris, D.J. Evolution and Behavioural Responses to Human-Induced Rapid Environmental Change: Behaviour and Evolution. Evol. Appl. 2011, 4, 367–387. [Google Scholar] [CrossRef]
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Figure 1. Schematic representation of the experimental design. The study consisted of a total of (i) 96 choice trials—i.e., eight trials per species (Aporrectodea caliginosa, Aporrectodea icterica and Allolobophora chlorotica) per origin (urban and rural) and per MTE exposure (M−/M+ and M−/M++), and 36 control trials—i.e., two trials per species per origin and per MTE exposure (M−, M+ and M++).
Figure 1. Schematic representation of the experimental design. The study consisted of a total of (i) 96 choice trials—i.e., eight trials per species (Aporrectodea caliginosa, Aporrectodea icterica and Allolobophora chlorotica) per origin (urban and rural) and per MTE exposure (M−/M+ and M−/M++), and 36 control trials—i.e., two trials per species per origin and per MTE exposure (M−, M+ and M++).
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Figure 2. MTE concentrations in natural soils. Mean ± se Cr, Cu, Ni, Pb, and Zn concentrations in the soils of rural and urban origins where earthworms were samples. Significant differences in MTE concentrations between the two origins are highlighted by asterisks (* p ≤ 0.05, ** p ≤ 0.01, ns p > 0.05); they account for the capture site.
Figure 2. MTE concentrations in natural soils. Mean ± se Cr, Cu, Ni, Pb, and Zn concentrations in the soils of rural and urban origins where earthworms were samples. Significant differences in MTE concentrations between the two origins are highlighted by asterisks (* p ≤ 0.05, ** p ≤ 0.01, ns p > 0.05); they account for the capture site.
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Figure 3. Health impairment. Mean ± se (a) mobility (calculated as the number of non-overlapping locations at the end of a trial) in earthworms of rural and urban origin exposed to MTE-less (M−), -poor (M+) and -rich soils (M++), (b) gallery length in A. caliginosa, A. chlorotica and A. icterica exposed to M−, M+ and M++, and (c) mass loss (calculated as the difference between post and pre-trial mass divided by pre-trial mass) in earthworms of rural and urban origin after exposure to M−, M+ and M++. Significant differences between species and MTE exposure (in (b)) and between MTE exposure (in (c)) are highlighted by different letters. Mobility did not significantly differ between MTE exposure, origin and species.
Figure 3. Health impairment. Mean ± se (a) mobility (calculated as the number of non-overlapping locations at the end of a trial) in earthworms of rural and urban origin exposed to MTE-less (M−), -poor (M+) and -rich soils (M++), (b) gallery length in A. caliginosa, A. chlorotica and A. icterica exposed to M−, M+ and M++, and (c) mass loss (calculated as the difference between post and pre-trial mass divided by pre-trial mass) in earthworms of rural and urban origin after exposure to M−, M+ and M++. Significant differences between species and MTE exposure (in (b)) and between MTE exposure (in (c)) are highlighted by different letters. Mobility did not significantly differ between MTE exposure, origin and species.
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Figure 4. MTE accumulation. Mean ± se Cr, Cu, Ni, Pb, and Zn concentrations in earthworms of rural and urban origin and exposed to MTE-less (M−), -poor (M+) and rich soils (M++). For graphical purposes, Cu and Pb concentrations have been divided by 10 and Zn concentrations by 100. For each MTE, significant differences in concentrations between MTE exposure (M−, M+ and M++) are highlighted by different letters. MTE concentrations did not vary according to earthworm origin.
Figure 4. MTE accumulation. Mean ± se Cr, Cu, Ni, Pb, and Zn concentrations in earthworms of rural and urban origin and exposed to MTE-less (M−), -poor (M+) and rich soils (M++). For graphical purposes, Cu and Pb concentrations have been divided by 10 and Zn concentrations by 100. For each MTE, significant differences in concentrations between MTE exposure (M−, M+ and M++) are highlighted by different letters. MTE concentrations did not vary according to earthworm origin.
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Figure 5. Habitat preference. Mean ± se proportion of (a) locations, (b) galleries in the MTE-enriched soil in earthworms of rural and urban origin and exposed to MTE-poor (M+/M−) and –rich soils (M++/M−). Significant differences between MTE exposure are highlighted by different letters. Earthworm movements did not significantly vary according to earthworm origin and species.
Figure 5. Habitat preference. Mean ± se proportion of (a) locations, (b) galleries in the MTE-enriched soil in earthworms of rural and urban origin and exposed to MTE-poor (M+/M−) and –rich soils (M++/M−). Significant differences between MTE exposure are highlighted by different letters. Earthworm movements did not significantly vary according to earthworm origin and species.
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Figure 6. Specificity. Mean ± se specificity for the MTE-less soil (M−) in earthworms of rural and urban origin and exposed to MTE-poor (M+/M−) and -rich soils (M++/M−). For graphical purposes, significant differences between MTE exposure and species are highlighted by different letters. However, only the simple effects of MTE exposure and species significantly explained specificity; the interaction between the two variables was not retained in the model.
Figure 6. Specificity. Mean ± se specificity for the MTE-less soil (M−) in earthworms of rural and urban origin and exposed to MTE-poor (M+/M−) and -rich soils (M++/M−). For graphical purposes, significant differences between MTE exposure and species are highlighted by different letters. However, only the simple effects of MTE exposure and species significantly explained specificity; the interaction between the two variables was not retained in the model.
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Figure 7. Choice mechanism. Mean ± se proportion of (a) locations and (b) galleries in the MTE-enriched, and of earthworms in the MTE-enriched soil according to (c) earthworm species and (d) origin in earthworms that visited both MTE-enriched and -less soils. For graphical purposes, significant differences between MTE exposure and earthworm origin and MTE exposure and species are highlighted by different letters, respectively, in (a,b). However, only the simple effects of MTE exposure and species significantly explained the proportion of galleries in the MTE-enriched soil; the interaction between the two variables was not retained in the model.
Figure 7. Choice mechanism. Mean ± se proportion of (a) locations and (b) galleries in the MTE-enriched, and of earthworms in the MTE-enriched soil according to (c) earthworm species and (d) origin in earthworms that visited both MTE-enriched and -less soils. For graphical purposes, significant differences between MTE exposure and earthworm origin and MTE exposure and species are highlighted by different letters, respectively, in (a,b). However, only the simple effects of MTE exposure and species significantly explained the proportion of galleries in the MTE-enriched soil; the interaction between the two variables was not retained in the model.
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Table
Table 1. MTE concentrations in artificial soils.
Table 1. MTE concentrations in artificial soils.
CrCuPbNiZn
M−5.5 ± 0.25.5 ± 5.538.8 ± 6.3BDL12.5 ± 12.5
M+6.2 ± 0.4 (5.0)42.0 ± 3.0 (50.0)154.3 ± 9.8 (150.0)2.9 ± 2.8 (7.5)106.0 ± 7.0 (122.5)
M++7.1 ± 0.1 (10.0)69.5 ± 7.5 (100.0)243.8 ± 28.8 (300.0)7.9 ± 0.0 (15.0)189.5 ± 12.5 (245.0)
Mean ± se measured (in black) and expected (in grey) MTE concentrations (in ppm) in artificial MTE-rich (M++), MTE-poor (M+) and MTE-less (M−) soils. Ni concentrations in M− were below detection limit (BDL).
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Chatelain, M. Endogeic Earthworms Avoid Soil Mimicking Metal Pollution Levels in Urban Parks. Sustainability 2023, 15, 11513. https://doi.org/10.3390/su151511513

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Chatelain M. Endogeic Earthworms Avoid Soil Mimicking Metal Pollution Levels in Urban Parks. Sustainability. 2023; 15(15):11513. https://doi.org/10.3390/su151511513

Chicago/Turabian Style

Chatelain, Marion. 2023. "Endogeic Earthworms Avoid Soil Mimicking Metal Pollution Levels in Urban Parks" Sustainability 15, no. 15: 11513. https://doi.org/10.3390/su151511513

APA Style

Chatelain, M. (2023). Endogeic Earthworms Avoid Soil Mimicking Metal Pollution Levels in Urban Parks. Sustainability, 15(15), 11513. https://doi.org/10.3390/su151511513

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