Patrick Ollivier
email: p.ollivier@brgm.fr

Jérémy Engevin

Sébastien Bristeau

Christophe Mouvet

Laboratory study on the mobility of chlordecone and seven of its transformation products formed by chemical reduction in nitisol lysimeters of a banana plantation in Martinique (French Caribbean)

Keywords: Chlordecone, transformation products, mobility, nitisol, French Caribbean, In Situ Chemical Reduction

niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

Water and soil contamination by pesticides is an issue of major concern notably because of the persistence and toxicity of these contaminants. Moreover, pesticides can be transformed, by natural processes or as a result of decontamination processes, into degradation products that can be more persistent, mobile, and sometimes more toxic than the parent compound [START_REF] Silva | Pesticide residues in European agricultural soils -A hidden reality unfolded[END_REF]. In the French Caribbean (FC), soils and waters are contaminated by chlordecone (CLD; C10Cl10O; CAS number 143-50-0 -decachlorooctahydro-1,3,2-metheno-2Hcyclobuta [c,d]pentalen-2-one, also known as kepone), a highly persistent organochlorine insecticide. Forty-five % of agriculture area in Martinique are contaminated by CLD, 8% with concentrations above 1 mg CLD/kg (Le [START_REF] Déaut | Report on the Impacts of the Use of Chlordecone and Pesticides in the West Indies: Assessment and Future Prospects[END_REF]. CLD was used in banana plantations (1972-1978 and 1981-1993; 3 kg ha -1 year -1 ), to control the black banana Weevil (Cosmopolites sordidus). It is a highly hydrophobic molecule (Kow between 4.9 and 5.4), strongly sorbed to soil (log Koc = 4.1-4.2) and poorly soluble in water (0.020 mg/L at 25°C) (UNEP, 2007).

The CLD pollution of soils in the FC could last up to centuries if no remediation action is taken [START_REF] Cabidoche | Long-term pollution by chlordecone of tropical volcanic soils in the French West Indies: a simple leaching model accounts for current residue[END_REF][START_REF] Ollivier | Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France)[END_REF]. [START_REF] Chevallier | Natural Chlordecone Degradation Revealed by Numerous Transformation Products Characterized in Key French West Indies Environmental Compartments[END_REF] detected 17 transformation products (TPs) of CLD in soils and water samples from Martinique. This proof of the formation of CLD TPs under field conditions does not however prove that the degradation of CLD in soils under natural conditions is quantitatively significant. A significant decrease over time of CLD concentration in soils would more likely be generated by its transfer to another compartment of the environment than by its degradation.

CLD from contaminated soils can be taken up by crops leading to concentrations over the Maximum Residue Limit (MRL; 20 μg/kg) [START_REF] Cabidoche | Contamination of harvested organs in root crops grown on chlordecone-polluted soils[END_REF]. CLD accumulates also in the fatty tissues of living organisms with bioconcentration in the food chains. Involuntary soil ingestion by animals reared outdoors leads to CLD transfer in animal products that make a significant part of the diet of the local population [START_REF] Jondreville | Contamination of free-range ducks by chlordecone in Martinique (FrenchWest Indies): A field study[END_REF][START_REF] Fournier | Toxicokinetics of chlordecone in goats: implications for risk management in French West Indies[END_REF]. CLD is also found in aquatic coastal fauna [START_REF] Dromard | Different transfer pathways of an organochlorine pesticide across marine tropical food webs assessed with stable isotope analysis[END_REF] and even pelagic cetacean [START_REF] Méndez-Fernandez | From banana fields to the deep blue: Assessment of chlordecone contamination of oceanic cetaceans in the eastern Caribbean[END_REF].

FC human population is exposed to CLD through the consumption of contaminated foodstuffs and drinking water. Chronic exposure to CLD can have severe impacts on human health: increased prevalence of prostate cancers [START_REF] Multigner | Chlordecone exposure and risk of prostate cancer[END_REF] and suspected increase risk of preterm birth [START_REF] Multigner | Chlordecone exposure and adverse effects in French West Indies populations[END_REF]. In 2009, CLD was included on the list of priority pollutants by the Stockholm Convention (Stockholm Convention on Persistent Organic Pollutants, 2001).

To date, no remediation process of FC soils contaminated by CLD has been validated on a large scale in real conditions. Phytoremediation by plants adapted to tropical climates appears to be limited. Only 0.03% of the CLD spiked to a clay-loam soil to reach a CLD concentration of 1 mg/kg was extracted over 1 full growth cycle by two species of Miscanthus [START_REF] Liber | Growth parameters influencing uptake of chlordecone by Miscanthus species[END_REF]. The intensification of land use increases the amount of CLD absorbed by plants, but the soil-to-plant transfer rates remain very low [START_REF] Liber | Bayesian network approach for the identification of relationships between drivers of chlordecone bioaccumulation in plants[END_REF]. As for root vegetables, the soil to plant transfer of CLD is too low for phytoremediation [START_REF] Clostre | Organochlorine (chlordecone) uptake by root vegetables[END_REF].

Microbial biodegradation of CLD was observed in laboratory experiments using FC soils in anaerobic conditions [START_REF] Chaussonnerie | Microbial Degradation of a Recalcitrant Pesticide: Chlordecone[END_REF]Chevallier et al., 2017;[START_REF] Lomheim | Evidence for extensive anaerobic dechlorination and[END_REF] and the same TPs (hydrochlordecones and polychloroindenes) were measured in natural soils [START_REF] Chevallier | Natural Chlordecone Degradation Revealed by Numerous Transformation Products Characterized in Key French West Indies Environmental Compartments[END_REF]. Microorganisms capable of degrading CLD are therefore present in FC soils. However, the CLD biodegradation in contaminated soils in a microcosm under aerobic conditions lead to only 3.0 to 4.4% of CLD mineralization after 215 days (Fernández-Bayo et al., 2013). Ways to increase the efficiency of CLD biodegradation are needed for the development of bioremediation approaches.

CLD sequestration in soils can be an alternative solution to soil decontamination by limiting the release of CLD into other compartments of the environment until efficient remediation techniques become available. Indeed, adding activated carbons (e.g., coconut shell) to contaminated soils (10 and 100 mg CLD/kg) decreased CLD bioavailability to liver and adipose tissue of piglets and goats by 65 to 99% [START_REF] Ranguin | Reduction of chlordecone environmental availability by soil amendment of biochars and activated carbons from lignocellulosic biomass[END_REF]. Crop contamination and leaching can also be reduced by adding compost in soils, compost acting through its strong capacity to sorb CLD (Clostre et al., 2014a) and its effect on soil porosity (Woignier et al., 2016).

By contrast, the In Situ Chemical Reduction (ISCR) treatment reduces significantly the CLD contamination in soils and water. ISCR consists in adding a mixture of a strongly reducing agent and an organic amendment to the soil brought close to its field capacity (Seech et al., 2010). After 6 months of treatment with the Daramend® amendment containing zero valent iron (ZVI), CLD concentrations in laboratory 80-kg mesocosms dropped by 74% in nitisol, 71% in ferralsol and 22% in andosol of FC [START_REF] Mouvet | Remediation by chemical reduction in laboratory mesocosms of three chlordecone-contaminated tropical soils[END_REF]. In the actual field conditions of a banana plantation, a 12-week soil treatment by ISCR lowered the CLD concentrations not only in the soil, but also in radishes, cucumbers and sweet potatoes that returned to values below the MRL [START_REF] Mouvet | Physico-chemical and agronomic results of soil remediation by In Situ Chemical Reduction applied to a chlordeconecontaminated nitisol at plot scale in a French Caribbean banana plantation[END_REF]. However, TPs (hydrochlordecones from mono-to pentadechlorinated) appeared in the soil and soil pore water. The formation of hydrochlordecones has also been reported in bacterial liquid cultures and microcosms inoculated with FC soils in the presence of ZVI (Chevallier et al., 2017). The ISCR results raise the question about the risk of hydrochlordecones contaminating the non-saturated zone beneath the treated topsoil layer and possibly the groundwater, as well as the evolution over time of this contamination. Batch experiments with a nitisol indicated that the TPs mono-and tri-hydroCLD are less sorbed by soil than the parent compound [START_REF] Mouvet | Sorption and desorption of chlordecone and two of its degradation products formed by reductive dechlorination[END_REF]. [START_REF] Ollivier | Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France)[END_REF] showed, using a 20-cm long nitisol column experiment under saturated conditions that dechlorination increases the mobility of the derivatives compared to the parent molecule (trihydroCLD > monohydroCLD > CLD). Such an increased mobility is likely to facilitate the transport of the TPs to soil layers deeper than those treated by ISCR. However, the experimental conditions of the two aforementioned studies were not fully representative of environmental conditions (e.g., sieved and unstructured soil, static batch systems or small columns, saturated conditions) making the results difficult to transpose to the natural environment. Columns filled with repacked soils do not maintain the original soil structure, horizons and macropore connections that influence the transport of contaminants [START_REF] Saporito | A Protocol for Collecting and Constructing Soil Core Lysimeters[END_REF].

Here, laboratory experiments are carried out using lysimeters (65-70-cm long and 20-cm in diameter) recovered with a minimum of disturbance from a CLD contaminated nitisol in FC.

A dedicated rain simulator generated the equivalent of 9.8 years of effective rainfall in Martinique, while maintaining an unsaturated flow regime in the soils. The objectives are to study i) the leaching of CLD from a contaminated nitisol, ii) the degradation of CLD and the production of its TPs resulting from a treatment simulating ISCR (Sim-ISCR) in the topsoil (15 cm), iii) the transport of these TPs deeper than the Sim-ISCR treated zone and notably iv) the evolution over time of their concentrations in the soil pore water and in the outflow water of the lysimeters.

Materials and methods

Soil sampling

The soil used in this study is a nitisol from a banana plantation in the plain of Lamentin (Martinique, French Caribbean). Three stainless steel lysimeters, 90 cm-long and 20 cm in diameter (named L1, L2 and L3), were gently driven into the soil by the bucket of a backhoe while trying to preserve the natural structure of the soil. Once the desired depth had been reached, the lysimeters were gently lifted out of the ground thanks to a rod inserted at the top and connected to the backhoe arm. The height of the soil inside the lysimeters was 69 cm (L1), 65 cm (L2) or 68 cm (L3). The top and bottom of the lysimeters were tightly sealed and flown to the laboratory in Orléans (mainland France) where the experiments were conducted.

Initial CLD soil concentrations had to be determined without disturbing the content of the lysimeters before the onstart of the experiments. To do so, triplicate soil samples were taken with a hand bulb planter from the walls of the holes resulting from the excavation of the lysimeters. The layers sampled were 0-30 cm, 30-60 cm and 60-90 cm for L2 and L3, and 0-30 cm, 30-55 cm and 55-80 cm for L6. In addition, the soil removed at the beginning of the experiment from inside the lysimeters to install porous cups was taken for the analysis of CLD and its TPs.

Design of the experiments

The experiment was designed to study i) the leaching of CLD from the contaminated nitisol under an unsaturated water flow regime, ii) the degradation of CLD and the production of its TPs resulting from a treatment in the topsoil (15 cm) simulating ISCR (Sim-ISCR), and iii) the fate of these TPs deeper than the treated zone, notably their concentrations in the outflow water of the lysimeters.

The very strong hydrophobicity of CLD makes it prone to sorption on organic components of experimental devices. To minimize such interferences, the entire system designed here consists of stainless steel, Teflon and glass. The experiment was carried out in a thermostatically controlled room (25 ± 2°C, covering the range of soil temperature at 10-30 cm depth in Martinique; [START_REF] Baran | Transfer process of phytosanitary products from soil to groundwater in Martinique[END_REF] and in the dark.

Each lysimeter was equipped with two sintered glass porous suction cups, at depths 15 and 49 cm for L1, 11 and 45 cm for L2, and 14 and 48 cm for L3 (Figure 1). The porous cup installed in the topsoil (15 cm) where Sim-ISCR was applied aimed at gaining information on the compounds released into the water in this area. The soil moisture content was measured at depth 35-cm every 3 minutes using time domain reflectometry (TDR) probes that had been previously calibrated in buckets of the nitisol over a range of water contents.

The experiment was designed to simulate the equivalent of several years of effective rainfall in Lamentin plain (Martinique), while maintaining an unsaturated flow regime in the soils.

Every 6 hours, Milli-Q H2O was sprayed for 4 minutes at 75 mL/min by a garden sprinkler at the top of the lysimiters (Figure 1). The spraying rates were regularly checked to correct a potential drift in the water supply. This regime of rain simulation led to an unsaturated water flow regime as evidenced by the fluctuations of the soils moisture content (Figure S1). Using an average value of 1,670 mm/yr for the effective rainfall in Lamentin plain [START_REF] Vittecoq | Assessment of water resources in Martinique: Spatial calculation of effective rainfall and validation at the watershed-scale[END_REF], the volume of water sprayed on the soil surface can be converted into "an equivalent number of years of effective rainfall". The daily water supply, 1.2 liters, corresponds to 38.2 mm of water, i.e., 8.35 days of effective rainfall.

Two months after the beginning of the experiment, a conservative tracer (Br, 100 mL at 600 mg Br/L) was sprayed at the top of the lysimeters to obtain information on the flow of water. After adding Br, the outflow water was collected every 12 hours for the first 3 days, less frequently for the next 17 days, to analyze Br by ion chromatography (DIONEX IC3000-SP-EG-DC system; Figure 2a).

The water leaving the lysimeters (i.e., outflow water) was continuously collected in 25L glass bottles through a tightly adapted funnel preventing evaporation. Every one to three weeks, after thorough mixing of the whole volume, an aliquot (500 mL) was sampled for the analysis of CLD and its TPs. One sample in triplicate and two samples in duplicate collected from each lysimeter and analyzed separately confirmed the good representativeness of the data from a single sample from the 25L bottle (± 0.3 to 5.9%, 1σ). After the 500 mL sampling, the remaining water collected was discarded and the bottles returned to the outlet of the lysimeters to collect the next samples.

CLD can be transported by suspended particulate matter (SPM) through the lysimeters.

However, CLD concentrations measured in 6 outflow waters filtered through Whatman GF/C glass filter (0.7 µm) were not different from those measured in unfiltered waters. If differences in CLD concentrations in outflow waters appear between the three lysimeters or over time, they will not be attributable to differences in SPM concentrations.

Soil pore water was collected every 4 to 6 weeks by applying a vacuum (500 mbar) for 24 h to the 2 porous suction cups installed in each lysimeter. The 30 to 800 mL of water collected were analyzed for CLD and its TPs.

After 140 days of experiment (corresponding to 3.2 years of effective rainfall), the water supply was stopped and the topsoil (15 cm, ~6 kg) of each column was removed from the column, dried (30°C) and transferred to sealed stainless steel containers to be treated by an in vitro simulation of In Situ Chemical Reduction (Sim-ISCR). The treatment consisted of a thorough mixing of ZVI (coarse grain size, 30% with a size < 50 µm) added to the soil at a rate of 4% (dry weight ZVI/dry weight soil) as described by [START_REF] Mouvet | Decontamination by In Situ Chemical Reduction of a nitisol and an alluvial soil polluted by chlordecone. Physico-chemical and agronomic results[END_REF]. The soil was then compacted and CaCl2 10 -2 M solution was added to obtain a soil moisture content close to its water holding capacity, favoring reducing conditions. Covers were placed on the containers to best guarantee the tightness of the systems. The redox potential was > 500 mV in untreated soils and dropped sharply to values < -500 mV after the addition of ZVI. It was stable during the treatment phase.

After 21 days of Sim-ISCR, the soil was retrieved from the containers and spread on aluminum trays to be gently stirred daily by hand, promoting the contact with air and enabling the soil to return to the redox conditions similar to those before Sim-ISCR. At the end of the treatment, triplicate samples were taken in treated soils and analyzed for CLD and its TPs. A conservative tracer (Br; 100 mL at 600 mg Br/L) was mixed with the treated soils in order to obtain, by monitoring Br concentrations in the outflow waters (Figure 2b), information on potential modifications of the water flow due to the treatment and the possible perturbations resulting from the removal of the top layer and subsequent refilling of the lysimeter. The treated soils were then put back in place at the top of the lysimeters. Water supply was started again as before the soil treatment. Water samples were taken once every 1 to 4 weeks for the outflow and 5 to 9 weeks for the suction cups, with a higher frequency just after the treated soil was put back in the lysimeter.

The whole experiment lasted 451 days. A total of 512.7 liters of water was sprayed at the surface of each lysimeter, corresponding to ~16,320 mm of water, i.e., an equivalent of ~9.8 years of effective rainfall in the plain of Lamentin.

Analytical method

The extraction and analytical methods for CLD and its TPs are fully described in [START_REF] Bristeau | Validation of chlordecone analysis for native and remediated French West Indies soils with high organic matter content[END_REF] and [START_REF] Mouvet | Remediation by chemical reduction in laboratory mesocosms of three chlordecone-contaminated tropical soils[END_REF].

The soil samples were dried (at 38°C for 72 h) before crushing and sieving to < 2 mm, followed by grinding to a particle size of <80 µm. Extraction was done using Pressurized Liquid Extraction with a mixture acetone/hexane 50:50 v/v at 100°C and 110 bars pressure.

Trans-nonachlor was used before extraction as surrogate to correct the results for extraction efficiency. CB53 and 13 C-CLD were added before analysis as two internal standards for the trans-nonachlor and CLD calibration, respectively. GC/MS/MS analyses were performed with a Bruker GC450 gas chromatograph, 1177 injector, automated sampler Combi Pal (CTC) and a 300MS triple quadrupole mass spectrometer.

The water samples had a volume of 500 mL for the outflow waters and 30 to 500 mL for the pore waters, depending on the volume collected by the suction cup. Analyses were carried out by liquid/liquid extraction dichloromethane. The final extract was done with 1ml of cyclohexane followed by GC/MS/MS. 13 C-CLD was added as an internal standard before analysis.

In water and soil water, the CLD molecule presents a gem diol function instead of the ketone function but the temperature of the GC-MS injector dehydrates chlordecone gem-diol [START_REF] Wilson | Structures of some Kepone photoproducts and related chlorinated pentacyclodecanes by carbon-13 and proton nuclear magnetic resonance[END_REF]. Consequently, it is impossible in this paper to distinguish the hydrated from the non-hydrated form of CLD. Besides CLD, analytical standards enabling quantification were available for only 4 TPs, 2 monohydroCLD, 5-and 5b-hydroCLD (CAS nomenclature), 1 trihydroCLD and 1 tetrahydroCLD. For the 2 latter compounds, the position of dechlorination could not be determined. For 3 additional TPs, the degree of dechlorination (penta-to heptadechlorination) was identified for sure but the position of dechlorination could not be determined either and no standard was available to enable quantification. The relative peak areas will be used as a proxy for semi-quantification of these 3 compounds [START_REF] Mouvet | Decontamination by In Situ Chemical Reduction of a nitisol and an alluvial soil polluted by chlordecone. Physico-chemical and agronomic results[END_REF].

Laboratory studies on CLD biodegradation with bacterial consortia [START_REF] Chaussonnerie | Microbial Degradation of a Recalcitrant Pesticide: Chlordecone[END_REF]Chevallier et al, 2017;[START_REF] Lomheim | Evidence for extensive anaerobic dechlorination and[END_REF] and field data from FC soils and waters [START_REF] Chevallier | Natural Chlordecone Degradation Revealed by Numerous Transformation Products Characterized in Key French West Indies Environmental Compartments[END_REF][START_REF] Lomheim | Evidence for extensive anaerobic dechlorination and[END_REF] identified chloroindenes, most notably the polychloroindene C9Cl5H3 (B1 compound in Chevallier et al, 2017) as important CLD TPs. P.L. Saaïdi from the Genoscope team provided an aliquot of B1 that enabled us to conduct a qualitative search for the compound in our samples. No quantification could however be achieved because the standard was described as « non stable » and because the means available in our study did not enable a validation of the full analytical procedure for B1 in natural soils and soil waters (in particular the extraction step) according to standard procedures (NF T90-210, 2018;ISO 11352, 2013). As a result, only qualitative data on the detection of the compound will be provided here.

In soils, the limit of quantification (LOQ) was 0.03 mg/kg for CLD, 5-hydroCLD, the trihydroCLD and the tetrahydroCLD, and 0.05 mg/kg for 5b-hydroCLD. In water samples, the LOQ for CLD, 5-hydroCLD, 5b-hydroCLD, the trihydroCLD and the tetrahydroCLD were 0.02 µg/L.

Results and discussion

Untreated soil

CLD concentrations in the top 30-cm of soils collected on the walls of the holes resulting from the excavation of the lysimeters were (n = 3) 0.51 ± 0.06 mg/kg for L1, 0.65 ± 0.04 mg/kg for L2 and 0.47 ± 0.04 mg/kg for L3, respectively. Within the lysimeters, they were 0.34 mg/kg at 15 cm-depth in L1, 0.53 mg/kg at 11 cm-depth in L2, and 0.40 mg/kg at 14 cm-depth in L3. These values are slightly lower than those measured in plots of the same field by [START_REF] Mouvet | Physico-chemical and agronomic results of soil remediation by In Situ Chemical Reduction applied to a chlordeconecontaminated nitisol at plot scale in a French Caribbean banana plantation[END_REF] at a depth of 0 to 40 cm (0.71 ± 0.26 mg/kg) and significantly lower than those found by [START_REF] Ollivier | Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France)[END_REF] in the top 30-cm of soils (1.52 ± 0.12 mg/kg) collected at other locations in the same banana plantation. Such a spatial variability of CLD concentrations in soils at plot scale was reported elsewhere [START_REF] Clostre | Soil type and growing conditions influence uptake and translocation of organochlorine (chlordecone) by cucurbitaceae species. Water, Air[END_REF] and our values are consistent with the literature data (between 0.31 and 2.9 mg CLD/kg in nitisol; [START_REF] Cabidoche | Long-term pollution by chlordecone of tropical volcanic soils in the French West Indies: a simple leaching model accounts for current residue[END_REF][START_REF] Cattan | Crisis management of chronic pollution: contaminated soil and human health[END_REF][START_REF] Woignier | Physical limitation of pesticides (chlordecone) decontamination in volcanic soils: fractal approach and numerical simulation[END_REF].

CLD concentrations in soils varied also with depth, with the highest values measured in the topsoil. They were 0.29 ± 0.05 mg/kg and 0.19 ± 0.02 mg/kg at depths 30-55 cm and 55-80 cm respectively near L1, 0.62 ± 0.15 mg/kg and 0.39 ± 0.03 mg/kg at depths 30-60 cm and 60-90 cm respectively near L2, while they were 0.51 ± 0.09 mg/kg and 0.27 ± 0.05 mg/kg at depths 30-60 cm and 60-90 cm respectively near L3. The soil samples recovered from inside the lysimeters (when setting up the suction cups) showed CLD concentrations of 0.17 mg/kg at 49 cm-depth in L1, 0.32 mg/kg at 45 cm-depth in L2, and 0.34 mg/kg at 48 cm-depth in L3.

The fact that the contamination by CLD is not limited to the tilling depth (0-60 cm soil horizon) indicates that a part of CLD was transported deeper than the topsoil where it was applied decades ago.

None of the hydroCLD identified neither the polychloroindene C9Cl5H3 were detected in the soils before the Sim-ISCR treatment.

Release of CLD and its

TPs from the soil before the ISCR treatment

In outflow waters

During the first 140 days of the experiments, corresponding to 3.2 years of effective rainfall, CLD concentrations varied from 7.3 to 11.1 µg/L, from 10.9 to 15.5 µg/L, and from 8.1 to 12.1 µg/L in outflow waters of L1, L2 and L3, respectively (Figure 3). The highest concentration was found in the first two water samples collected, but no clear decreasing trend was observed over time. [START_REF] Ollivier | Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France)[END_REF] reported also the absence of significant variation in CLD concentrations in waters leaving a 20cm-long nitisol column over the equivalent of 3.5 years of effective rainfall. Using all measured CLD concentrations and the volumes of water that passed through the lysimeters, the mean CLD concentration in outflow waters was calculated to be 8.41 µg/L (L1), 12.4 µg/L (L2) and 9.25 µg/L (L3). The highest concentrations in waters are thus found at the outlet of L3, the lysimeter showing the highest CLD concentrations in soil. However, the CLD concentration in water leaching a lysimeter does not depend solely on the CLD concentration in the soil. It may also be influenced by the contact time between the water and the soil [START_REF] Ollivier | Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France)[END_REF]. Actually, the conservative tracer (Br) shows differences in water flow between the three lysimeters (Figure 2a). The residence time of water in L3 is longer compared to the other two lysimeters, with the peak of Br concentrations reached after ~6.5 days in L2 against 3.5 days for both L1 and L3. Note that Br is measured in the first outflow water sample (less than 1 day after Br adding) from L1 indicating a strong connection between the deepest soil layer and the soil surface of the lysimeter where Br was added. In the outflow waters of L3 and L2, Br is detected after 1 day and 1.5 days, respectively. Though 5b-hydroCLD was not detected in the soils, its concentrations in the outflow waters of the three lysimeters ranged from 0.03 to 0.22, 0.05 to 0.34 and 0.06 to 0.22 µg/L in L1, L2 and L3, respectively (Figure 4). The detection of 5b-hydroCLD in the outflow waters of the lysimeters before any remediation treatment reflects the fact that 5b-hydroCLD has been produced as an impurity during CLD manufacturing [START_REF] Devault | Natural transformation of chlordecone into 5b-hydrochlordecone in French West Indies soils: statistical evidence for investigating long-term persistence of organic pollutants[END_REF]. However, 5b-hydroCLD concentrations in soils from Martinique banana plantations cannot be justified by its sole input as an impurity present in the commercial formulations [START_REF] Devault | Natural transformation of chlordecone into 5b-hydrochlordecone in French West Indies soils: statistical evidence for investigating long-term persistence of organic pollutants[END_REF]. It can also be generated in soils by photolysis and microbial degradation [START_REF] Orndorff | Microbial transformation of Kepone[END_REF]. The 5b-hydroCLD concentrations were, however, 36 to 238 times lower than the CLD concentrations in the same sample. Previous studies showed also that 5b-hydroCLD is present at much lower levels than CLD in surface and ground-waters in Martinique [START_REF] Clostre | Organochlorine (chlordecone) uptake by root vegetables[END_REF][START_REF] Cattan | A conceptual model of organochlorine fate from a combined analysis of spatial and mid-to long-term trends of surface and groundwater contamination in tropical areas (FWI)[END_REF].

Because the outflow waters were continuously collected and analyzed for CLD and its TPs, the quantity of CLD and 5b-hydroCLD leached from the soil can be calculated. During the first 140 days of experiments (i.e., 3.2 years of effective rainfall), 1.4, 2.08 and 1.54 mg of CLD was leached from L1, L2 and L3, respectively, and 18.5, 26.6 and 25.7 µg for 5b-hydroCLD from L1, L2 and L3, respectively. From the mass of soil in the lysimeters (27.1, 25.6 and 26.7 kg dw, in L1, L2 and L3, respectively) and assuming that the CLD concentration measured in the sample taken from each soil horizon (0-30 cm, 30-55 cm or 30-60 cm, and 55-80 cm or 60-90 cm) corresponds to the mean concentration of the whole horizon, the quantity of CLD in the soil of each lysimeter was estimated to be 9.3 mg (L1), 14.1 mg (L2) and 11.1 mg (L3). On these bases, the % of CLD that was leached from the soils after 3.2 years of effective rainfall was calculated to be 15.1, 14.7 and 13.9%. These values are consistent with previous studies that indicated that, in the absence of any attenuation mechanisms other than leaching, CLD in a typical nitisol in the FC would persist for several decades [START_REF] Cabidoche | Long-term pollution by chlordecone of tropical volcanic soils in the French West Indies: a simple leaching model accounts for current residue[END_REF][START_REF] Ollivier | Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France)[END_REF]. Neither 5-hydroCLD, nor any of the identified tri-to heptahydroCLD, nor the polychloroindene C9Cl5H3 were detected in the outflow waters.

In pore waters

CLD was found in the pore waters, with the lowest values from the suction cup installed in the topsoil (Figure 5). They varied between 4.9 and 6.8 µg/L at 15 cm-depth and between 7.5 and 8.8 µg/L at 49 cm-depth in L1, between 4.4 and 7.6 µg/L at 11 cm-depth and between 10.7 and 11.7 µg/L at 45 cm-depth in L2, and between 2.9 and 3.5 µg/L at 14 cm-depth and between 4.5 and 8.9 µg/L at 48 cm-depth in L3. The concentrations greater at the 45 -49 cm depths than at the 11 -15 cm depths in all 3 lysimeters indicate that, from the 11 -15 cm deep layer down to the 45 -49 cm depth, the soil acts more as a source of CLD than as a sink. 5b-hydroCLD was also detected in the pore waters but, contrary to CLD, the highest values were measured in the topsoil area (Figure 6). 5b-hydroCLD/CLD ratios in the topsoil waters reached 0.14, 0.09 and 0.23 in L1, L2 and L3, respectively. These values are much higher than those in the commercial products Kepone® and Curlone® used in the FC (5b-hydroCLD/CLD mass ratio = 0.00055-0.00109, with a geometrical mean of 0.00077; [START_REF] Devault | Natural transformation of chlordecone into 5b-hydrochlordecone in French West Indies soils: statistical evidence for investigating long-term persistence of organic pollutants[END_REF] suggesting that 5b-hydroCLD was actually produced in the soil after CLD was applied decades ago. Microbial degradation and photoreaction with UV light can indeed produce 5b-hydroCLD at the soil surface, which could explain why the highest concentrations are found in the topsoil. Neither 5-hydroCLD, nor any of the identified tri-to heptahydroCLD, nor the polychloroindene C9Cl5H3 were detected in the pore waters.

CLD and its TPs after the Sim-ISCR treatment

In the treated topsoil

Assuming that 15.1% (L1), 14.7% (L2) and 13.9% (L3) of CLD left the lysimeters by leaching before the topsoil treatment (see above), the CLD concentrations before the soil treatment was estimated as 0.29 mg/kg at 15 cm-depth in L1, 0.46 mg/kg at 11 cm-depth in L2 and 0.34 mg/kg at 14 cm-depth in L3. After the Sim-ISCR treatment, CLD concentrations were 0.15 ± 0.02 mg/kg, 0.27 ± 0.08 mg/kg and 0.20 ± 0.02 mg/kg in the topsoil (0-15 cm) of L1, L2 and L3, respectively. Sim-ISCR thus led to a reduction of 47 ± 8%, 41 ± 18% and 42 ± 4% in the CLD content in the topsoil of L1, L2 and L3, respectively. These values agree with those previously reported in the literature. [START_REF] Mouvet | Physico-chemical and agronomic results of soil remediation by In Situ Chemical Reduction applied to a chlordeconecontaminated nitisol at plot scale in a French Caribbean banana plantation[END_REF] indicated that ISCR with addition of 4.0% dw/dw of ZVI in the 0-40-cm soil layer of a nitisol in a FC banana plantation lowered the initial CLD concentration by up to 55% after 21 days of treatment. [START_REF] Ollivier | Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France)[END_REF] showed a reduction of 52% of the CLD concentration in the top 30-cm of a nitisol after 15 days of ISCR treatment (ZVI; 4.3% dw/dw). Calculated from the difference in CLD concentrations before and after Sim-ISCR treatment, the total mass of CLD degraded in the topsoil by Sim-ISCR was 0.8 mg in L1, 1.12 mg in L2 and 0.84 mg in L3.

Sim-ISCR generated 5-hydroCLD, with concentrations of 0.062 ± 0.005 mg/kg, 0.073 ± 0.018 mg/kg and 0.051 ± 0.005 mg/kg in the topsoil of L1, L2 and L3, respectively, corresponding to a total mass of 5-hydroCLD of 0.37 mg, 0.44 mg and 0.30 mg. A mass balance (calculated on the basis of µmol) shows that 50, 42 and 39% of CLD were transformed into 5-hydroCLD in the topsoil of L1, L2 and L3, respectively. Neither 5b-hydroCLD nor any of the identified tri-to heptahydroCLD were detected in the soils. At least 50% of the CLD was therefore transformed into molecules other than those that were analyzed. The polychloroindene C9Cl5H3 was not detected either (Figure S2), in line with the observation that chlorinated indene TPs are not formed as a result of the action of ZVI only while they are found in reactions using sodium sulfide or titanium(III) citrate and vitamin B12 (Chevallier et al, 2017). The use of complementary extraction and analytical techniques, notably liquid chromatography [START_REF] Chevallier | Natural Chlordecone Degradation Revealed by Numerous Transformation Products Characterized in Key French West Indies Environmental Compartments[END_REF] would have proven interesting for a more complete mass balance.

CLD and its TPs in the soil pore and outflow waters

CLD concentrations in pore waters decreased by 48 to 73% in the zone treated by Sim-ISCR, and remained rather stable until the end of the experiment (Figure 5). They varied between 1.3 and 2 µg/L (vs 4.9 and 6.8 µg/L before treatment) in L1, between 2.1 and 2.9 µg/L (vs 4.4 and 7.6 µg/L before treatment) in L2, and between 1.4 and 2.0 µg/L (vs 2.9 and 3.5 µg/L before treatment) in L3. As previously reported [START_REF] Ollivier | Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France)[END_REF], the decrease in the CLD concentration is more pronounced in water (48 to 73%) than in soil (~42 to 47%). This may be due to Sim-ISCR being more effective in degrading the CLD present on the surface of the soil grains than the CLD in the microporosity, the former being probably more easily released into the water [START_REF] Woignier | Physical limitation of pesticides (chlordecone) decontamination in volcanic soils: fractal approach and numerical simulation[END_REF]. 5b-hydroCLD concentrations in pore waters in the treated zone decreased also greatly after treatment, dropping below 0.1 µg/L vs up to 0.8 µg/L before the treatment (Figure 6).

While 5-hydroCLD was not found in the pore water before the treatment, it was quantified in the pore water of the treated zone, with the highest values in the first sample collected after the Sim-ISCR treatment: 1.38 (L1), 2.38 (L2), and 1.43 µg/L (L3) (Figure 5). These values are similar to the 2.2 µg/L measured by [START_REF] Ollivier | Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France)[END_REF] in waters that passed through a 20-cm long column containing nitisol treated by ISCR. Afterwards, 5-hydroCLD concentration decreased but remained above 0.1 µg/L (corresponding to the precautionary limit value for each pesticide in drinking water; European Union, 2000). After 6 years of equivalent effective rainfall following the Sim-ISCR treatment, 5-hydroCLD concentrations were 0.2, 0.1 and 0.23 µg/L in pore waters in the treated zone of L1, L2 and L3, respectively.

None of the identified tri-to heptahydroCLD were detected in the pore waters.

In outflow waters, CLD concentrations decreased after the Sim-ISCR treatment of the topsoil (Figure 3). Over the 6.5 years of effective rainfall after the treatment, CLD concentrations varied between 5.4 and 7.7 µg/L (with a mean value of 6.3 µg/L vs 8.4 µg/L before treatment) in L1, between 9.5 and 12.2 µg/L (mean of 10.8 µg/L vs 12.4 µg/L before treatment) in L2, and between 6.3 and 9.0 µg/L (mean of 7.1 µg/L vs 9.2 µg/L before treatment) in L3. The reduction in CLD concentrations was thus only 25, 23 and 13% in outflow waters of the L1, L2 and L3, respectively. This somewhat limited effect results from the treatment addressing only the top 15-cm of the soil profile. The flux of CLD from the remaining untreated 50-55 cm of the profile had no reason to decrease and "buffered" the effect generated in the topsoil.

5b-hydroCLD concentrations in outflow waters after treatment dropped below 0.1 µg/L (between 0.02 and 0.08 µg/L against 0.05 to 0.34 µg/L before the treatment), i.e., similar values to those measured in topsoil pore waters indicating that the flux of 5b-hydroCLD from the remaining untreated 50-55 cm of the profile was low (Figure 6). That is consistent with the hypothesis that 5b-hydroCLD is mainly produced in the untreated soil by microbial degradation and photoreaction with UV light at the soil surface. Sim-ISCR treatment of the topsoil can thus be effective to reduce 5b-hydroCLD in waters, even in the underlying groundwater.

5-hydroCLD was present in outflow waters with clear differences between lysimeters (Figure 3). 5-hydroCLD was found for L1 in the first outflow sample (0.39 µg/L, after ~1 month of effective rainfall) after the treatment while it was detected later in the outflow water of L2 (0.02 µg/L) and L3 (0.02 µg/L), after an equivalent of ~10.3 and ~3 months of effective rainfall, respectively. A similar trend was found in the Br tracing, with a later breakthrough in L2 compared to L3 and L1, the latter with a value in the first sample near to the maximum reached, strongly suggesting some preferential flow (Figure 2b). However, significant delays in 5-hydroCLD transport were observed compared to the conservative tracer. The highest 5-hydroCLD concentrations were reached after an equivalent of 1.4 years (0.6 µg/L, L1), 3.5 years (0.54 µg/L, L2) and 4 years (0.41 µg/L, L3) of effective rainfall following the soil treatment, whilst the maximum concentrations of Br were measured after an equivalent of 9, 38 and 13 days of effective rainfall. Although 5-hydroCLD is more mobile than CLD [START_REF] Ollivier | Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France)[END_REF], its breakthrough at the outlet strongly retarded compared to that of Br indicates that it interacts strongly with the soil. Its detection in outflow water samples over the whole duration of the experiment indicates some persistence in pore water. Concentrations in outflow waters increased when those in pore water in the treated zone decreased (Figure 5). They were higher than those measured in pore waters of the treated zone after 1.4 years (in L1), 2.8 years (in L2) and 4 years (in L3) of effective rainfall following the treatment. The highest values measured in outflow water were, however, lower by a factor of 2.3 (L1), 4.4 (L2) and 3.5 (L3) than those in pore waters, showing a natural attenuation with depth. After 6.5 years of effective rainfall following the Sim-ISCR treatment, 5-hydroCLD was still present in outflow waters with a concentration of 0.27, 0.32 and 0.29 µg/L for L1, L2 and L3, respectively. A further attenuation by a factor of around 3 would bring the outflow water in line with the maximum concentration acceptable for drinking water, 0.1 µg/L. The identified trihydroCLD, tetrahydroCLD, pentahydroCLD and heptahydroCLD were not detected in outflow waters, except for L1 (Figure 6, SI-3 and SI-4). The highest concentrations (or relative peak intensities) were found just after the Sim-ISCR treatment.

They were no longer detected after 10.3 months (trihydroCLD), 4.5 months (tetrahydroCLD and pentahydroCLD) and 2.9 months (heptahydroCLD) of effective rainfall. This sequence of disappearance is consistent with previous works suggesting an increase in the mobility of derivatives with increasing degree of dechlorination [START_REF] Dolfing | Gibbs free energy of formation of chlordecone and potential degradation products: implications for remediation strategies and environmental fate[END_REF][START_REF] Mouvet | Sorption and desorption of chlordecone and two of its degradation products formed by reductive dechlorination[END_REF][START_REF] Ollivier | Transport of chlordecone and two of its derivatives through a saturated nitisol column (Martinique, France)[END_REF]. Just like in the treated soil, and for the same reason, the polychloroindene C9Cl5H3 was not detected in the soil pore and outflow waters after the Sim-ISCR treatment (Figure S2).

Conclusions and environmental relevance

The results presented here confirm the effectiveness of ISCR adapted to laboratory conditions, with a decrease in CLD concentration of 41 to 47% in the soil and 48 to 73% in the soil pore waters of the treated 15 top cm layer. However, the CLD contamination affects more than the topsoil, notably the water percolating through the soils to the groundwater.

Of greatest interest to this respect are two key results from our study. Firstly, 3.2 years of effective rainfall leached less than 15% of the CLD present in 65-70 cm soil lysimeters. In the absence of any attenuation mechanism other than leaching, the CLD in the soil will continue to act as a source of CLD for decades. Secondly, the Sim-ISCR remediation process applied to the top 15 cm layer resulted in a rather low, 13 to 25%, decrease in CLD concentrations in the water leached at the bottom of the 65-70 cm soil profile. This somewhat limited effect is mainly due to the fact that the CLD in the soil between the 15 and 65 -70 cm depths, 0.17 to 0.62 mg/kg, remains a major source term and conceal the benefit of remediating the top 15 cm. This effect illustrated by our experimental data will be all the more significant in natural systems where CLD is certainly present deeper than 70 cm. Indeed, CLD cannot have reached the aquifers without having passed through and contaminated the whole unsaturated zone, locally several tens of meters thick. Remediating by ISCR the 15 (this work) to 30-cm [START_REF] Mouvet | Physico-chemical and agronomic results of soil remediation by In Situ Chemical Reduction applied to a chlordeconecontaminated nitisol at plot scale in a French Caribbean banana plantation[END_REF] top of the soils will therefore not be sufficient for preventing further CLD contamination of the underlying groundwater. It must be stressed, however, that this limitation, resulting from the intrinsic characteristics of the media to be restored, is not specific to ISCR.

The potential benefits or drawbacks of ISCR must also be evaluated on the basis of the CLD TPs formed by the process. In the soil, CLD was mainly transformed into 5-hydroCLD, representing 39% to 50% of the total CLD degraded. In the outflow waters of the lysimeters, in addition to 5-hydroCLD, a trihydroCLD, a tetrahydroCLD, a pentahydroCLD and an heptahydroCLD were also detected. The 5-hydroCLD concentrations in outflow waters, 2.3 to 4.4 lower than those in soil pore water in the treated zone, indicate some natural attenuation with depth. The mobility of the CLD TPs was greater than that of CLD but they were not flushed out completely of the 60 cm soil profile over the duration of our study. Indeed, after 6.5 years of effective rainfall following the treatment, the 5-hydroCLD concentration in the outflow waters (~50-55 cm below the soil treated) was still in the 0.3 µg/L range (Figure 3). However, the presence of these molecules in water recharging aquifers possibly used as source for drinking water should not be considered a greater concern than the existing contamination by CLD. Indeed, Legeay et al. (2017) showed that a monohydroCLD and a trihydroCLD formed by ISCR are devoid of mutagenicity and genotoxicity and have lower proangiogenic properties than CLD. An in vivo test with a murine model of human prostate tumor confirmed that the proangiogenic effect of a mono-, a tri-and a tetrahydroCLD formed by ISCR was lower than that of the parent molecule [START_REF] Alibrahim | In vivo comparison of the proangiogenic properties of chlordecone and three of its dechlorinated derivatives formed by in situ chemical reduction[END_REF]. Lastly, [START_REF] Mouvet | Remediation by chemical reduction in laboratory mesocosms of three chlordecone-contaminated tropical soils[END_REF] did not observe any greater ecotoxicological response in the soils treated by ISCR than in the untreated soil.

A limitation of the soil treatment by ISCR is its cost (~19 €/m 2 ; [START_REF] Mouvet | Decontamination by In Situ Chemical Reduction of a nitisol and an alluvial soil polluted by chlordecone. Physico-chemical and agronomic results[END_REF] when applied as done for the field validation conducted in a banana plantation [START_REF] Mouvet | Physico-chemical and agronomic results of soil remediation by In Situ Chemical Reduction applied to a chlordeconecontaminated nitisol at plot scale in a French Caribbean banana plantation[END_REF].

Most of the cost is linked to that of ZVI itself and to the cost of its delivering to the FC. A major reduction of cost would be achieved if the development of specific microbial populations capable of reducing the iron oxides naturally present in the soils could be stimulated without costly inputs. The Fe(II) produced could be an electron donor as effective as Fe0 in degrading CLD (Hervé Macarie, personal communication;[START_REF] Devault | Natural transformation of chlordecone into 5b-hydrochlordecone in French West Indies soils: statistical evidence for investigating long-term persistence of organic pollutants[END_REF].

Further investigations under way in this perspective may pave the way for a significant contribution of soil remediation processes to the management of the CLD issue in the FC.

Even though our study focused on the fate of CLD and some of its hydrochlordecone TPs, recent works show the presence of polychloroindene TPs in natural FC soils [START_REF] Chevallier | Natural Chlordecone Degradation Revealed by Numerous Transformation Products Characterized in Key French West Indies Environmental Compartments[END_REF][START_REF] Lomheim | Evidence for extensive anaerobic dechlorination and[END_REF]. Further quantitative investigations on the degradation of polychloroindene TPs by ISCR treatment in soil and water and the later fate of these new TPs are needed. The results would enable to gain insight on the need to consider such TPs when looking at possible cocktail effects influencing the overall eco-toxicity of CLD and its TPs in the FC.

FiguresFigure 1 .

 1 Figures Figure 1. Experimental set-up. The depth of the porous cups differs slightly between the lysimeters. Here is shown the schematic for L1 (see text for details). Sim-ISCR: Simulated In Situ Chemical Reduction. ZVI: Zero Valent Iron.

Figure 2 .

 2 Figure 2. Br tracing as a function of time (days of experiment or equivalent daily effective rainfall in Lamentin; see text for details) before (a) and after (b) the treatment of the topsoil by Sim-ISCR (15 cm).

Figure 3 .

 3 Figure 3. CLD (left) and 5-hydroCLD (right) concentrations in the outflow water samples of the lysimeters (L1, L2 and L3). Open symbols correspond to samples analyzed after the Sim-ISCR treatment of the topsoil (15 cm) (see text for details).

Figure 4 .

 4 Figure 4. 5b-hydroCLD (left) and trihydroCLD (right) concentrations in the outflow water samples of the lysimeters (L1, L2 and L3). Open and orange symbols correspond to samples analyzed after the Sim-ISCR treatment of the topsoil (15 cm) (see text for details).

Figure 5 .

 5 Figure 5. CLD (left) and 5-hydroCLD (right) concentrations in pore water samples collected, according to L1, L2 and L3, at depths 11, 14 or 15 cm (red symbols) and 45, 48 or 49 cm (green symbols), and in the outflow water as a function of water input expressed as "Equivalent yearly effective rainfall in Lamentin". Open symbols show the samples collected after the Sim-ISCR treatment of the topsoil (15 cm; see text for details).

Figure 6 .

 6 Figure 6. 5b-hydroCLD (left) and trihydroCLD (right) concentrations in pore water samples collected, according to L1, L2 and L3, at depths 11, 14 or 15 cm (red symbols) and 45, 48 or 49 cm (green symbols), and in the outflow water as a function of water input expressed as "Equivalent yearly effective rainfall in Lamentin". Open symbols show the samples collected after the Sim-ISCR treatment of the topsoil (15 cm; see text for details).

Table 2

 2 Composition of the substrates (sludge, olive pomace and macroalgal residues). Olive pomace), MAR (macroalgal residues), TWAS (thickened waste activated sludge).

	Substrates	OP	TWAS:OP	TWAS	TWAS:MAR	MAR
	C (% TS)	52±0.2	-	38.3±0.6	-	38.8±0.2
	H (% TS)	7.2±0.3	-	5.68±0.05	-	6.1±0.3
	N (% TS)	1.0±0.2	-	6.13±0.06	-	4.0±0.2
	S (% TS)	0.10±0.01	-	1.12±0.03	-	0.65±0.01
	C/N	52.0	12.0	6.7	7.6	9.7
	P (g /kg TS)	10.1±0.4	-	39.2±0.4	-	11.5±0.3
	K (g /kg TS)	65±4	-	37.3±0.1	-	3.6±0.4
	Lipids (%)	16.4±0.8	-	-	-	-
	OP (

Acknowledgments

This work was supported by the French Ministry for the Ecological and Inclusive Transition.

We gratefully acknowledge the financial support provided to the PIVOTS project by the Région Centre -Val de Loire (ARD 2020 program and CPER 2015 -2020) and the French Ministry of Higher Education and Research (CPER 2015 -2020 and public service subsidy to Brgm). This operation is co-funded by European Union. Europe is committed to the Center-Val de Loire region with the European Regional Development Fund.