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Acosta2018
1. Accepted Manuscript
Phytoremediation of mine tailings with Atriplex halimus and organic/inorganic
amendments: a five-year field case study
J.A. Acosta, A. Abbaspour, G.R. Martínez, S. Martínez-Martínez, R. Zornoza, M.
Gabarrón, A. Faz
PII: S0045-6535(18)30669-6
DOI: 10.1016/j.chemosphere.2018.04.027
Reference: CHEM 21175
To appear in: Chemosphere
Received Date: 05 February 2018
Revised Date: 22 March 2018
Accepted Date: 04 April 2018
Please cite this article as: J.A. Acosta, A. Abbaspour, G.R. Martínez, S. Martínez-Martínez, R.
Zornoza, M. Gabarrón, A. Faz, Phytoremediation of mine tailings with Atriplex halimus and organic
/inorganic amendments: a five-year field case study, (2018), doi: 10.1016/j.Chemosphere
chemosphere.2018.04.027
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2. ACCEPTED MANUSCRIPT
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1 Phytoremediation of mine tailings with Atriplex halimus and organic/inorganic
2 amendments: a five-year field case study
3 Acosta1* J.A., Abbaspour2 A., Martínez1 G.R., Martínez-Martínez1 S., Zornoza1 R., Gabarrón1 M., Faz1 A.
4 1Sustainable use, management and reclamation of soil and water research group, Universidad Politécnica de
5 Cartagena. Paseo Alfonso XIII 48, 30203. Cartagena (Spain).*E-mail: ja.acosta@upct.es
6 2Department of Soil and Water, Faculty of Agriculture, Shahrood University of Technology, Semnan
7 province, Iran.
8
9 Abstract
10 Mine tailings have adverse chemical and physical conditions, including high concentrations of
11 metals and salts, low organic matter content, and unbalanced rates of nutrients which limit the
12 development of vegetation. A large scale field experiment was conducted to reclaim a tailing pond
13 by triggering the growth of native species by spontaneous colonization by tilling (TL) the tailing
14 pond surface and using marble waste (CaCO3; MW), pig slurry (PS) and their combination
15 (MW+PS) as soil amendments. Soil physicochemical properties and water and DTPA extractable
16 metal concentrations of bulk and rhizosphere soils were analyzed after five year from the
17 application of the treatments. In addition, plants of Atriplex halimus from each treatment were
18 collected and metals in roots, leaves and stems analyzed. Before amendments application, the
19 studied pond showed a neutral pH, high salinity and a moderate organic carbon content. After five
20 years, the pH value was significantly increased only in MW plot. The results showed significant
21 increases of DTPA-extractable Zn in MW and MW+PS plots, Pb in all treatments except MW plot,
22 Cd only in PS plot, and Cu only in MW+PS plot. A. halimus was the most dominant species,
23 growing spontaneously in all plots, with lower vegetation cover in CT and MW plots, 6% and 2%
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24 respectively. Application of MW increased leaf Pb accumulation by 2.5-fold and Cd by 55 %,
25 when compared to the CT. The high initial salinity and probable substitution of metals by Ca2+ on
26 exchangeable surfaces of soil particles may be the reasons for higher uptake of metals in MW plot
27 when compared to the other plots. Although this plant is widely utilized in contaminated sites for
28 phytostabilization purposes, it may absorb and translocate high concentrations of metals to the
29 aboveground tissues in saline contaminated sites.
30 Keywords: plant establishment, mining waste, phytomanagement, amendment
31
32 1. Introduction
33 Mine activity is considered as one of the most dangerous anthropogenic activities in the world,
34 which results in changes in landscapes, destruction of habitats, contamination of soil and water,
35 and degradation of land resources (European Environmental Bureau, 2000). Deforestation,
36 formation of acid mine drainage, erosion and sedimentation processes, release of metal(loid)s,
37 modification of habitats, and contamination of surface and groundwater are considered as main
38 environmental impacts generated by mining during the exploitation period (Doumett et al., 2008;
39 Ji et al., 2011; Kabas et al., 2012; Zornoza et al., 2012b). Sierra Minera, located between Cartagena
40 and La Unión cities in southeast of Spain, constituted one of the most important and the oldest
41 mining districts in Europe, which was one of the world's largest mines producing Pb/Zn in the late
42 nineteenth century (Faz et al., 2001; Kabas et al., 2012). With the development of the mining
43 activity, millions of tons of waste were accumulated on the surface of the land (Acosta et al., 2011).
44 These wastes led to changes in the visual characteristics of the area and produced physiographic
45 modifications and alterations in the landscape..
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46 There are many techniques for remediating metal contaminated soils, but most of these techniques
47 are expensive and environmentally invasive (Simon, 2005). The establishment of a stable
48 vegetative cover is considered an adequate option to obtain the long-term remediation of these
49 contaminated soils (Simon, 2005; Pardo et al., 2017), which is known as phytoremediation. Among
50 phytoremediation techniques, the most promising option for tailing ponds reclamation is the use
51 of phytostabilization. It is defined, according to Wong (2003), as the use of metal-tolerant plants
52 to immobilize metal(loid)s by accumulation in the roots or precipitation in the rhizosphere.
53 Phytostabilization of contaminated soils reduces the bioavailability of pollutants, increases the
54 organic matter content, improves its physical properties, reduces wind and water erosion, increases
55 biodiversity (plant and microbial) of the ecosystem and favors the development of natural
56 processes of soil (Zornoza et al., 2012a; Pardo et al., 2014) , ultimately creating a long-term, self-
57 sustaining ecosystem. Compared to other existing techniques, it is more economical, effective and
58 environmentally sustainable (Yang et al., 2010).
59 Atriplex halimus L. is a shrub belonging to Amaranthaceae family, tolerates high concentration of
60 salts and metals in soils (Clemente et al., 2012; Pardo et al., 2017) and therefore could be used for
61 the stabilization of tailings affected by mining activity. Nonetheless, despite the high metal
62 tolerance and growth in tailings, some studies have reported that A. halimus shows a great capacity
63 to translocate metals from roots to shoot (Kabas et al., 2012; Pardo et al., 2017). Hence, a thorough
64 assessment of the potential risks of metal entry to the food chain by using A. halimus for the
65 phytomanagement of tailings and mine affected soils is essential to minimized hazards to
66 ecosystems.
67 Several types of amendments have been used for more effective phytostabilization of metals
68 depending on their characteristics and availability (Kabas et al., 2012; Lahori et al., 2017). The
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69 application of organic residues as soil improvers can decrease the bioavailability of metals and
70 improve soil fertility conditions, allowing the survival of plants and their growth. In this case the
71 reduction of metal bioavailability is due to the adsorption on the solid surfaces and the
72 complexation with humic substances (Perez-Esteban et al., 2014). This process of adsorption
73 depends on the metal and the type of soil involved, the degree of humification of the organic
74 matter, the content of metals and salts and the effects of the organic matter on the redox potential
75 and pH of the soil (Simon, 2005; Clemente et al., 2012; Kabas et al., 2012). In addition, alkaline
76 materials, such as cement, mussel shell, lime or marble waste have been used to improve soil
77 physical and chemical properties (de Mora et al., 2005; Pardo et al., 2011; Kabas et al., 2012;
78 Ahmad et al., 2014; Rajapaksha et al., 2015).
79 According to the latter approaches, a long-term field experiment was started in 2011, applying
80 marble waste (MW) and pig slurry (PS) to a tailing pond of Cartagena-La Unión mining district in
81 order to trigger the establishment of a spontaneous native vegetation cover which could contribute
82 to the stabilization of metals and improvement of soil quality and fertility (Kabas et al., 2012).
83 Five years from the amendments application, A. halimus was the most dominant species, growing
84 spontaneously in all plots. Therefore, the main objectives of this research were: i) to evaluate the
85 spontaneous colonization of A. halimus after five years of addition of MW, PS and MW+PS to
86 reclaim a tailings pond; ii) to assess the effect of the different amendments and their combined
87 application on the bioavailiability of metals (Cd, Cu, Pb and Zn) in the soil and their accumulation
88 in different tissues of A. halimus. This research will permit to identify if A. halimus is a suitable
89 candidate for phytostabilization of tailings and how the addition of different amendments can favor
90 the immobilization of metals or their uptake and translocation to the aerial parts.
91
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92 2. Materials and methods
93 2.1. Description of the studied area
94 Cartagena-La Unión mining district is located in SE Spain, between Cartagena and La Unión cities,
95 at the Mediterranean seaside. The climate of the area is semiarid Mediterranean, with an annual
96 average temperature of 18° C and average annual precipitation of 275 mm. The mining activity
97 developed in this mining district (over 50 km2) for more than 2500 years caused an enormous
98 landscape impact in the area, especially due to the accumulation of mine wastes in ponds. These
99 wastes pose the main problems and environmental health risks because of high pollution and
100 salinity of the wastes (Kabas et al., 2012). In September 2011, a remediation strategy was carried
101 out in one tailing pond (37º 35’ N, 0º 52’ W) through the application of organic and alkaline
102 amendments (European Commission Project FP7 IRIS). This tailing pond presents an area of 7400
103 m2, a depth of 14 m and a volume of 150 000 m3.
104 2.2. Experimental design
105 The tailing pond was divided into five different plots where the following treatments were applied
106 in September 2011 as described by Kabas et al. (2012) and shown in Fig.1a:
107 1- Untilled and unamended control (CT).
108 2- Surface tillage (0-50 cm) to improve tailings physical properties (TL).
109 3- Surface tillage and application of marble waste (MW) at a rate of 4 kg m-2. This dose was
110 calculated using the method proposed by Sobek et al. (1978), which provides an indication for the
111 quantity of lime required to neutralize all the potential acid according to the percentage of
112 sulphides present in the mine soil.
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113 4- Surface tillage and application of pig slurry (PS) at a rate of 3 L m-2. Dose for pig slurry was
114 established by thresholds imposed by legislation regarding the addition of total nitrogen to soil to
115 avoid contamination by nitrates (Council Directive 91/676/EEC).
116 5- Surface tillage and combined application of MW and PS (4 kg m-2 and 3 L m-2 respectively).
117 Some chemical and physical characteristics of the tailings in the different plots and properties of
118 MW and PS applied in this study have been described by Kabas et al. (2012) in detail. Briefly,
119 MW, consisting of calcium carbonate and free of toxic elements, contributes to neutralize the
120 acidity, immobilize metals and improve soil aggregation. Pig slurry was used as a source of organic
121 matter and nutrients to improve the soil quality and fertility. Once the amendments were applied,
122 different plant species spontaneously colonized the different plots, depending on the treatment
123 applied. After 5 years from the amendments application, A. halimus was the predominant species
124 in all plots.
125 Three samples of bulk soil were collected from the surface of the tailings pond in each plot (0-25
126 cm) and three A. halimus plants, with same size and without symptoms of diseases and pests, were
127 selected per plot and uprooted to collect rhizospheric soil and plant material for analyses. Soil
128 samples were dried at 40ºC for 48 hours and passed through a 2 mm sieve. A subsample of each
129 sample was ground using an agate mortar (RetchRM 100). For plants analyses, each plant was
130 separated in roots, stems and leaves. In order to eliminate all dust from the surface of the plants,
131 and soil adhered to the roots, each part was carefully washed with tap water and deionized water,
132 finally was dried at 50 ºC for 72h. Plant material was ground and preserved in polyethylene bags.
133 2.3. Analytical methods
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134 Electrical conductivity (EC) and pH were measured in deionized water (1:5 and 1:1 w/v,
135 respectively) (Soil Survey Staff, 2004).The soil samples were digested using US-EPA 3051
136 protocol, 0.5 g of the ground soil sample was weighed into a Teflon digestion tube (MARSXpress),
137 followed by addition of 10 mL of 69% nitric acid (HNO3). Availability of metals was determined
138 by chelation method with DTPA (Lindsay and Norvell, 1978). The concentration of water-soluble
139 metals was extracted with deionized water (1:5 w/v) by continuous stirring for 6 h (Ernst, 1996).
140 Metal concentration in the different plant tissues (root, stem and leaves) was determined by the
141 US-EPA 3052 protocol in a microwave digestion system (Cem Corporation, Matthews, USA): 0.5
142 g of the plant sample was weighed into a Teflon digestion tube (MARS Xpress), followed by the
143 addition of 0.5 ml of 37% hydrochloric acid (Suprapur, Merk), 9 ml nitric acid, (Suprapur, Merk),
144 and 1 ml of 30% hydrogen peroxide (Sigma - Aldrich). The concentration of Cd, Cu, Pb and Zn in
145 the soil and plant samples was measured using atomic absorption spectrophotometer in triplicate
146 (AA240FS series from Varian Australia Pty Ltd). A certified reference material (BAM-U110,
147 purchased from Federal Institute of Germany for Materials Research and Testing) were also used
148 to verify the quality assurance of the analyses. We obtained recoveries of 95-104% for Cd, 93-
149 99% for Cu, 91-103% for Pb, 97-102% for Zn.
150
151 2.4. Metal bioaccumulation and translocation indices
152 Metal bioaccumulation index (BI) and translocation index (TI) were calculated to assess the metals
153 accumulation in different tissues of the plant. The BI was calculated as the ratio of a metal
154 concentration in different tissues of a plant to its concentration in soil. The BI higher than the unit
155 reveals that a certain metal is effectively taken up by leaves, stems, or roots from soil (Kandziora-
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156 Ciupa et al., 2017). The TI was calculated as the ratio of a metal concentration in stems or leaves
157 to its concentration in roots. TF higher than the unit reveals that plant translocate a certain metal
158 efficiently from root to shoot (Kandziora-Ciupa et al., 2017).
159 2.5. Data analysis
160 To ensure the fitting of the data to a normal distribution the Kolmogorov-Smirnov test was applied.
161 Some data do not followed normal distribution and log transformation was done. ANOVA test
162 was used to identify differences among treatments using a Tukey’s post hoc with a significance
163 p<0.05. Data that failed on normality, even log-transforming, were submitted to non-parametric
164 test such as Kruskal-Wallis test and Mann-Whitney U test at p<0.05 to assess the differences
165 between variables. The relationship among soil properties and metals uptake by Atriplex halimus
166 were studied by Spearman correlations. All statistical analysis was performed using the statistics
167 software SPSS 23 (IBM).
168 3. Results and discussion
169 3.1. Effect of the amendments five years after application in chemical properties and
170 metals
171 The initial pH of the tailings was above 7 in all plots, being generally neutral (Table 1). There was
172 no significant difference (p ≤ 0.05) for pH in the plots except in MW plot, where a value of 8.4
173 was reached after 5 years. The initial electrical conductivity (EC) of the tailings ranged from 2.6
174 to 7.6 dS m-1, being the highest in MW plot, probably due to the accumulation of salts in this area
175 of the pond favored by its topography (Kabas et al., 2012). Nevertheless, the species of A. halimus
176 could grow, being very resistant to the salinity. Before application of the amendment, the organic
177 carbon (OC) content was moderate in all plots, with a maximum value of 13.2 g kg-1 in MW plot
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178 (Table 1). After 5 years of the amendments application, the levels of organic carbon decreased in
179 TL, and MW plots, due to the mineralization of the organic matter.
180 The total metal content in the different plots was variable, ranging 7500 -12741 mg kg-1 for Zn,
181 3254-5041 mg kg-1 for Pb, 24-33 mg kg-1 for Cd, and 176-223 mg kg-1 for Cu (data not showed),
182 indicating the high heterogeneity of this mining waste.
183 The initial concentrations of DTPA extractable (D-Ext) Zn (Table 2) were high (up to 402 mg kg-1)
184 in all plots except in the control (CT), in which the concentration reached 135 mg kg-1. After 5
185 years of amendments application, Zn availability significantly decreased (p ≤ 0.05) in TL and PS,
186 while significantly increased in MW and MW+PS (Table 2). Contrary to the fact that the addition
187 of alkaline compounds to soil reduces the availability of metals through pH increase (Lahori et al.,
188 2017), the application of MW did not reduce the availability of this element in the tailings after
189 five years.
190 The initial concentration of available Pb was very high in CT (345 mg kg-1), but ranged from 35
191 to 62 mg kg-1 in the other plots. After 5 years of amendments application, a significant increase in
192 the availability of Pb was observed in all plots except for CT and MW. These increases may be
193 due to the association of this element with the organic compounds in those plots where pig slurry
194 was applied (PS and MW+PS) and the oxidation of the sulfides in TL plot. Tailings typically
195 contain minerals rich in metal sulfides (Acosta et al., 2011) and gradual oxidation of these minerals
196 generates metal sulfates, being in a more available form (Castillo et al., 2013).
197 The concentration of available Cd was high and very variable among the plots before the
198 application of amendments, with values ranging from 1.1 to 6.1 mg kg-1. After 5 years of
199 amendments application, available Cd significantly (p ≤ 0.05) decreased in TL and MW, but
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200 increase in PS. In the plots treated with MW, the calcium carbonate applied may form insoluble
201 compounds like Cd carbonate, thereby reducing Cd bioavailability. In contrast, organic materials
202 application has been able to form soluble organic complexes with Cd during the decomposition
203 that may increase the availability of this element (Abbaspour et al., 2008; Khan et al., 2017).
204 Nonetheless, some organic amendments with high rate of humified organic materials can reduce
205 Cd availability by adsorption reactions (Kabas et al., 2012; Pardo et al., 2014). It should be pointed
206 out that the highest concentration of available Cd was observed with application of MW followed
207 by MW+PS. In addition, these plots had the highest EC values (Table 1). The increases in the
208 availability of Cd (and to some extent that of Zn) with the increase in the soil salinity could be
209 mainly attributed to the ion pairs and complexes formed with the inorganic anion ligands such as
210 chlorides and sulfates (Ghallab and Usman, 2007; Abbaspour et al., 2008).
211 Similar to available Cd, the concentration of available Cu was highly variable, with values ranging
212 between 2.3 and 4.6 mg kg-1. After 5 years of amendments application, the availability of Cu
213 significantly decreased in TL, whereas it significantly increased in MW+PS plot. This increase
214 may be due to the formation of organic complexes between Cu and dissolved organic ligands. The
215 complexation of Cu by dissolved organic materials present in pig slurry may reduce the adsorption
216 reactions of Cu on soil colloids and subsequently increase its availability (Ashworth and Alloway,
217 2007; Abbaspour et al., 2008).
218 The concentration of soluble Zn in all plots before application of amendments was < 2 mg kg-1
219 except for CT (7.5 mg kg-1) (Table 2). After 5 years of amendments application, there was an
220 increase in soluble Zn in all plots except for CT, so that both tillage and application of the
221 amendments triggered a higher release of this element to the soil solution, possibly due to the
222 activation of oxidation processes (Castillo et al., 2013).
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223 At the beginning of the experiment, the concentration of soluble Pb ranged from 0.2 mg kg-1 in CT
224 to 0.6 mg kg-1 in MW+PS, indicating the lower mobility of this metal, assuming no risk of
225 dispersion by runoff or leachate waters. Contrary to available Pb, the soluble fraction of Pb
226 significantly decreased in all plots, except CT. This was in agreement with the results of some
227 researchers who applied hydrated lime and compost to slightly acid tailings (Pardo et al., 2017) as
228 well as pig slurry and compost to acid tailings (Clemente et al., 2012), reporting significant
229 decreases in the Pb solubility.
230 The soluble Cd concentration was below 0.11 mg kg-1 in all plots before to the application of
231 amendments. After 5 years of application, an increase in the soluble fraction of Cd was observed
232 in plots amended with PS and PS+MW, owing to the formation of Cd organic complexes. Several
233 studies have indicated that soluble organic wastes are effective in raising the solubility of heavy
234 metals ( Ashworth and Alloway, 2007; Abbaspour et al., 2008; Antoniadis et al., 2017). Usman et
235 al. (2004) also found the significant increases of Cd, Zn, and Cu concentrations, extracted by
236 NH4NO3, during 90 days incubation of a calcareous soil amended with sewage sludge, composted
237 turf and plant residues. Possible mechanisms explained by the authors consisted of the decreased
238 soil pH and the formed metals-organic complexes by adding the organic wastes.
239 Initially, the soluble fraction of Cu was low in all plots (< 0.03 mg kg-1), but after 5 years it
240 significantly increased to 0.08 mg kg-1 and 0.06 mg kg-1 in PS and MW+PS, respectively. The
241 increases may be attributed to the organic material provided by the pig slurry. In general, the
242 mobility of metals in soil depends highly on pH, ionic strength, and type and amount of organic
243 matter (Ashworth and Alloway, 2007). The PS is substantially consisted of dissolved organic
244 matter which may forms Cu complexation, thereby increasing mobility of Cu in soil solution.
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245 3.3. Effect of the A. halimus rhizosphere in chemical properties and metals mobility
246 The pH was slightly alkaline in the rhizosphere soil (Table 3), observing a higher value than that
247 of in the bulk soil for CT and PS plots, which indicates that the root zone produced an increase in
248 pH in these treatments. Nye (1981) indicated that rhizosphere increases soil pH when the plant
249 absorb nitrates and other anions in higher quantities than cations, and therefore roots release
250 bicarbonates to maintain electrical neutrality, increasing soil pH. The EC was significantly
251 (p≤0.05) higher in the rhizosphere than in the bulk soil, except for CT and PS plots, implying that
252 water and nutrients taken up by the plant root produces an accumulation of salts or dissolve some
253 easily weathered minerals in this area by the root exudates (Séguin et al., 2004). The OC content
254 in all plots was significantly (p≤0.05) higher in the rhizospheric soil than in the bulk soil. In
255 general, the plant roots excrete into the rhizosphere some organic and inorganic compounds,
256 consisting of a mixture of sugars, organic acids, vitamins, and ions (e.g. OH−, H+ and HCO3
−),
257 thereby increasing microbial activities, which may alter the pH, EC and OC content in the
258 rhizosphere (Dakora and Phillips, 2002; Séguin et al., 2004; Abbaspour et al., 2012).
259 Compared to the bulk, the rhizospheric soil had the higher concentration of available Zn only in
260 CT plot, and that of soluble Zn in all plots. The available and soluble Pb increases were observed
261 for the rhizospheric soil in MW plot and in all plots, respectively. The slightly same trends were
262 found for those of Cd and Cu, indicating that the exudates produced by the plant roots solubilize
263 the metals, highlighting the higher concentrations found in MW+PS plot. It should be pointed out
264 that the pH and OC are two fundamental factors affecting on the metal solubility in soil (Séguin et
265 al., 2004). Although the pH value in the rhizosphere did not decrease by the root exudates, the
266 increased OC content could enhance the soluble metal concentrations. Organic acids released from
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267 the roots can solubilize insoluble metal minerals to the more available forms, thereby increasing
268 the metals in soil solution (Séguin et al., 2004; Abbaspour et al., 2012).
269
270 3.4. Metal contents in different parts of A. halimus
271 A. halimus spontaneously grew in all plots, the percentages of cover in each plots for Atriplex
272 were: 2% MW, 6% CT, 19% MW+PS, 32% PS and 38% TL. The lowest vegetation cover was
273 reported in MW plot (Fig. 1b), likely owing to its high initial EC (EC 7.6 dS m-1; Table 1),
274 following by CT plot, owing to adverse physical properties as high compaction and weak aeration
275 (Kabas et al., 2012).
276 The Zn concentration was statistically higher in leaves than in roots and stems in CT, PS and MW
277 (Fig. 2a). Concentrations were > 350 mg kg-1, which could generate a risk of entry to the food
278 chain. However, the phytotoxic concentration of Zn in leaf tissue for various species reported to
279 be 100-400 mg kg-1 (Kabata-Pendias, 2010). In this sense, Clemente et al. (2012) found higher leaf
280 Zn content (up to 1200 mg kg-1) in this species, growing on a highly contaminated soil (total Zn
281 of 9686 mg kg-1 and pH of 6.2). Kabas et al. (2012) also found the concentration of higher than
282 700 mg kg-1 in the shoots of A. halimus, growing in the same tailings but at the first year of the PS
283 treatment. This revealed the fact that A. halimus was unaffected by high Zn concentration in the
284 tailings, probably through the formation of Zn oxalate in the leaves (Lutts et al., 2004). The Zn BI
285 was < 1 only in MW and MW+PS, indicating that the application of calcium carbonate limited the
286 absorption of Zn by the plant (Fig. 2b). In addition, the leaf Zn TI was significantly higher than
287 the unit in all plots, indicating the ineffective stabilization of Zn by the plant (Fig. 2c).
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288 Considering the phytotoxic range of Pb (30-300 mg kg-1) in leaf of various species (Kabata-
289 Pendias, 2010), the highest accumulation of Pb was detected in the plant leaves in MW and
290 MW+PS (> 200 mg kg-1; Fig. 3a). This was in agreement with the results of Kabas et al. (2012)
291 who reported that, among four native species growing on the same tailings, A. halimus was the
292 only species accumulating the higher content of Pb in shoots than in roots. However, contrary to
293 our findings, Pb is known as an immobile element in plant tissues, whose translocation from root
294 to shoot is limited (Abbaspour et al., 2012; Badrloo et al., 2016; Antoniadis et al., 2017). The Pb
295 BI indicated a remarkable stem and mostly leaf Pb accumulation in MW and MW+PS plots (Fig.
296 3b). In the other plots there was no Pb bioaccumulation, since the values were, as an average, < 1.
297 Besides, the highest Pb TI was found in leaves in both plots treated with MW (Fig. 3c). The higher
298 uptake and translocation of Pb with addition of MW may be likely due to the highest salinity (Table
299 1). As a consequence, Pb2+ is replaced in the exchange sites of the soil by some other cation species,
300 thereby increasing Pb mobility in the respective plots. Several authors have reported that the
301 increased salinity enhance the concentration of metals in the soil solution through the formation of
302 soluble inorganic complexes, depending on the particular metal and type and concentration of salts
303 (Ghallab and Usman, 2007; Abbaspour et al., 2008). Considering the fact that A. halimus was
304 reported as a excluder species to stabilize metals in soil (Clemente et al., 2012; Pardo et al., 2014;
305 Pardo et al., 2017;), Pb may precipitate as insoluble compounds like chloropyromorphite
306 (Pb5(PO4)3Cl) on the root surface (Cao et al., 2002) and in the soil (Abbaspour and Golchin, 2011;
307 Badrloo et al., 2016). Therefore, high EC and Ca2+ concentration obtained by MW addition may
308 inhibit the formation of such compounds, thereby increasing the Pb uptake. Unfortunately, the
309 concentration of soluble species of Ca2+ and PO4
3- was not determined in the current study, though
310 soluble Pb concentration was highest in MW after 5 years of reclamation (Table 2) despite the
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311 highest pH (Table 2). Compared to the bulk soil, highest EC and available Pb concentration in the
312 rhizosphere soil, without any significant change in the pH value (Table 1), may prove this
313 hypothesis.
314 The highest concentration of Cd was observed in the leaves of MW plot, followed by CT, with
315 significant differences among plant parts (Fig. 4a). No significant differences among root, stem
316 and leaf were observed in the rest of plots. A significant decrease in Cd BI was found in all
317 treatments when compared to CT (Fig. 4b), implying that the treatments hinder the Cd uptake. The
318 Cd TI showed the highest values in leaves of plants growing in MW plot (Fig. 4c). This is probably
319 because of the highest salinity in the MW plot (Table 2), related to higher concentration of
320 available Cd. Among all parameters measured in this study, the best correlation was distinguished
321 between the leaf Cd concentration and EC (p˂0.05, n=15, data not shown), indicating the effect of
322 salinity on more Cd bioavailability.
323 The highest Cu concentration was observed in leaves of CT, PS and MW (Fig. 5a), being lower
324 than the minimum concentration of phytotoxic, reported by Kabata-Pendias (2010) . Cu BI was >
325 1 in all plots and in all plant tissues, being the highest for leaves in CT and PS (Fig. 5b). The Cu
326 TI showed also the highest values in leaves of plants growing in CT and PS (Fig. 5c). It indicated
327 that MW amendment was more efficient than PS to stabilize Cu in the plant roots or in the tailings.
328 Conclusion
329 Tailings showed a high variability in both the concentration of metals and the different
330 physicochemical properties, indicating materials with high heterogeneity. Organic compounds
331 applied with pig slurry increased the concentration of available Pb, Cu and Cd, and soluble Cd and
332 Cu after five years. Contrarily, the soluble fraction of Pb significantly decreased after PS and MW
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333 applications. In addition, the application of MW did not reduce the availability of Zn in the tailings
334 after five years. Tillage led to a better growth of A. halimus, probably through improving physical
335 properties of the tailings, and a significant reduction of Cd uptake by the plant. After five years of
336 the application, pig slurry did not alter the pH and organic carbon content of the tailings, but
337 increased Cu concentration in leaves, likely due to the formation of dissolved organic complexes
338 with Cu2+ species. Marble waste increased Cd and Pb concentration in the leaves, likely attributed
339 to the high initial salinity, resulting in the formation of some soluble inorganic complexes. In
340 general, A. halimus showed the highest concentrations of metals preferentially in leaves.
341 Therefore, this plant was not able to stabilize metals, especially Pb and Cd in the tailings, despite
342 previous studies suggested the use of A. halimus to phytostabilize soil metals. These disagreements
343 would indicate that there are some other factors that could make the plant change its physiological
344 processes and accumulate metals in its different organs, aspect that should be further studied in
345 detail.
346 References
347 Abbaspour A., Arocena J., Kalbasi M. (2012) Uptake of phosphorus and lead by Brassica juncea
348 and Medicago sativa from chloropyromorphite. International journal of phytoremediation
349 14:531-542.
350 Abbaspour A., Golchin A. (2011) Immobilization of heavy metals in a contaminated soil in Iran
351 using di-ammonium phosphate, vermicompost and zeolite. Environmental Earth Sciences
352 63:935-943.
353 Abbaspour A., Kalbasi M., Hajrasuliha S., Fotovat A. (2008) Effect of Organic Matter and Salinity
354 on Ethylenediaminetetraacetic Acid–Extractable and Solution Species of Cadmium and
18. ACCEPTED MANUSCRIPT
17
355 Lead in Three Agricultural Soils. Communications in soil science and plant analysis
356 39:983-1005.
357 Ahmad M., Lee S.S., Lim J.E., Lee S.E., Cho J.S., Moon D.H., Hashimoto Y., Ok Y.S. (2014)
358 Speciation and phytoavailability of lead and antimony in a small arms range soil amended
359 with mussel shell, cow bone and biochar: EXAFS spectroscopy and chemical extractions.
360 Chemosphere 95: 433-441.
361 Acosta J., Faz A., Martínez-Martínez S., Zornoza R., Carmona D., Kabas S. (2011) Multivariate
362 statistical and GIS-based approach to evaluate heavy metals behavior in mine sites for
363 future reclamation. Journal of Geochemical Exploration 109:8-17.
364 Faz A., Raquel A.L., Miguel C.A.H., Fernandez G. (2001) Soils affected by mining and industrial
365 activities in Cartagena (SE Spain): classification problems, European Soil
366 Bureau,Research pp. 165–170.
367 Antoniadis V., Levizou E., Shaheen S.M., Ok Y.S., Sebastian A., Baum C., Prasad M.N., Wenzel
368 W.W., Rinklebe J. (2017) Trace elements in the soil-plant interface: Phytoavailability,
369 translocation, and phytoremediation–A review. Earth-Science Reviews 171:621-645.
370 Ashworth D., Alloway B. (2007) Complexation of copper by sewage sludge-derived dissolved
371 organic matter: effects on soil sorption behaviour and plant uptake. Water, air, and soil
372 pollution 182:187-196.
373 Badrloo T.A., Nobariyan M.R.S., Bostani A., Ghorbani H., Abbaspour A. (2016) Solubility and
374 Phytoavailability of Phosphorus and Lead in a Contaminated Soil Amended with Two
375 Phosphorus Fertilizers. Communications in Soil Science and Plant Analysis 47:1967-1974.
19. ACCEPTED MANUSCRIPT
18
376 Cao X., Ma L.Q., Chen M., Singh S.P., Harris W.G. (2002) Impacts of phosphate amendments on
377 lead biogeochemistry at a contaminated site. Environmental Science & Technology
378 36:5296-5304.
379 Castillo S., Jesús D., de la Campa A.M.S., González-Castanedo Y., Fernández-Caliani J.C.,
380 Gonzalez I., Romero A. (2013) Contribution of mine wastes to atmospheric metal
381 deposition in the surrounding area of an abandoned heavily polluted mining district (Rio
382 Tinto mines, Spain). Science of the Total Environment 449:363-372.
383 Clemente R., Walker D.J., Pardo T., Martínez-Fernández D., Bernal M.P. (2012) The use of a
384 halophytic plant species and organic amendments for the remediation of a trace elements-
385 contaminated soil under semi-arid conditions. Journal of hazardous materials 223:63-71.
386 Dakora F.D., Phillips D.A. (2002) Root exudates as mediators of mineral acquisition in low-
387 nutrient environments. Plant and soil 245:35-47. DOI: 10.1023/A:1020809400075.
388 de Mora A.P., Ortega-Calvo J.J., Cabrera F., Madejón E. (2005) Changes in enzyme activities and
389 microbial biomass after “in situ” remediation of a heavy metal-contaminated soil. Applied
390 Soil Ecology 28:125-137.
391 Doumett S., Lamperi L., Checchini L., Azzarello E., Mugnai S., Mancuso S., Petruzzelli G., Del
392 Bubba M. (2008) Heavy metal distribution between contaminated soil and Paulownia
393 tomentosa, in a pilot-scale assisted phytoremediation study: influence of different
394 complexing agents. Chemosphere 72:1481-1490.
395 Ernst W. (1996) Bioavailability of heavy metals and decontamination of soils by plants. Applied
396 geochemistry 11:163-167.
20. ACCEPTED MANUSCRIPT
19
397 European Environment Bureau (EEB) (2000) The Environmental Performance of the Mining
398 Industry and the Action Necessary to Strengthen EU Legislation. In: Hontelez, J. (Ed.),
399 Document no. 2000/016, Brussels, p. 28
400 Ghallab A., Usman A.R.A. (2007) Effect of sodium chloride-induced salinity on phyto-availability
401 and speciation of Cd in soil solution. Water, air, and soil pollution 185:43-51.
402 Ji P., Sun T., Song Y., Ackland M.L., Liu Y. (2011) Strategies for enhancing the phytoremediation
403 of cadmium-contaminated agricultural soils by Solanum nigrum L. Environmental
404 pollution 159:762-768.
405 Kabas S., Faz A., Acosta J., Zornoza R., Martínez-Martínez S., Carmona D., Bech J. (2012) Effect
406 of marble waste and pig slurry on the growth of native vegetation and heavy metal mobility
407 in a mine tailing pond. Journal of Geochemical Exploration 123:69-76.
408 Kabata-Pendias A. (2010) Trace elements in soils and plants CRC press.
409 Kandziora-Ciupa M., Nadgórska-Socha A., Barczyk G., Ciepał R. (2017) Bioaccumulation of
410 heavy metals and ecophysiological responses to heavy metal stress in selected populations
411 of Vaccinium myrtillus L. and Vaccinium vitis-idaea L. Ecotoxicology:1-15.
412 Khan M.A., Khan S., Khan A., Alam M. (2017) Soil contamination with cadmium, consequences
413 and remediation using organic amendments. Science of the Total Environment 601:1591-
414 1605.
415 Lahori A.H., Zhang Z., Guo Z., Mahar A., Li R., Awasthi M.K., Sial T.A., Kumbhar F., Wang P.,
416 Shen F. (2017) Potential use of lime combined with additives on (im) mobilization and
417 phytoavailability of heavy metals from Pb/Zn smelter contaminated soils. Ecotoxicology
418 and Environmental Safety 145:313-323.
21. ACCEPTED MANUSCRIPT
20
419 Lindsay W.L., Norvell W.A. (1978) Development of a DTPA soil test for zinc, iron, manganese,
420 and copper. Soil science society of America journal 42:421-428.
421 Lutts S., Lefevre I., Delpérée C., Kivits S., Dechamps C., Robledo A., Correal E. (2004) Heavy
422 metal accumulation by the halophyte species Mediterranean saltbush. Journal of
423 Environmental Quality 33:1271-1279.
424 Nye (1981). Changes of pH across the rhizosphere induced by roots. Plant and Soil
425 Pardo T., Bernal M., Clemente R. (2017) Phytostabilisation of severely contaminated mine tailings
426 using halophytes and field addition of organic and inorganic amendments. Chemosphere
427 178:556-564.
428 Pardo T., Clemente R., Bernal M.P. (2011). Effects of compost, pig slurry and lime on trace
429 element solubility and toxicity in two soils differently affected by mining activities.
430 Chemosphere 84: 642-650.
431 Pardo T., Clemente R., Epelde L., Garbisu C., Bernal M. (2014) Evaluation of the
432 phytostabilisation efficiency in a trace elements contaminated soil using soil health
433 indicators. Journal of hazardous materials 268:68-76.
434 Pérez-Esteban J., Escolástico C., Masaguer A., Vargas C., Moliner A. (2014) Soluble organic
435 carbon and pH of organic amendments affect metal mobility and chemical speciation in
436 mine soils. Chemosphere 103: 164-171.
437 Rajapaksha A.U., Ahmad M., Vithanage M., Kim K.R., Chang J.Y., Lee S.S., Ok Y.S. (2015) The
438 role of biochar, natural iron oxides, and nanomaterials as soil amendments for
439 immobilizing metals in shooting range soil. Environmental Geochemistry and Health 37:
440 931-942.
22. ACCEPTED MANUSCRIPT
21
441 Séguin V., Gagnon C., Courchesne F. (2004) Changes in water extractable metals, pH and organic
442 carbon concentrations at the soil-root interface of forested soils. Plant and Soil 260:1-17.
443 Simon L. (2005) Stabilization of metals in acidic mine spoil with amendments and red fescue
444 (Festuca rubra L.) growth. Environmental Geochemistry and Health 27:289-300.
445 Soil Survey Staff, 2004. Soil Survey Laboratory Methods Manual. Version No. 4.0. USDA-NRCS.
446 Soil Survey Investigations Report No. 42. U.S. Govt. Print. Office, Washington (407 pp).
447 Usman A., Kuzyakov Y., Stahr K. (2004) Dynamics of organic C mineralization and the mobile
448 fraction of heavy metals in a calcareous soil incubated with organic wastes. Water, Air,
449 and Soil Pollution 158:401-418.
450 Wong M. (2003) Ecological restoration of mine degraded soils, with emphasis on metal
451 contaminated soils. Chemosphere 50:775-780.
452 Yang S.-X., Liao B., Li J.-t., Guo T., Shu W.-S. (2010) Acidification, heavy metal mobility and
453 nutrient accumulation in the soil–plant system of a revegetated acid mine wasteland.
454 Chemosphere 80:852-859.
455 Zornoza R., Faz A., Carmona D., Martínez-Martínez S., Acosta J. (2012a) Plant cover and soil
456 biochemical properties in a mine tailing pond five years after application of marble wastes
457 and organic amendments. Pedosphere 22:22-32.
458 Zornoza R., Acosta J.A., Carmona D., Martínez-Martínez S., Weiss, N., Faz A. (2012b) The effect
459 of former mining activities on contamination dynamics in sediments, surface water and
460 vegetation in El Avenque stream, SE Spain. Water, Air and Soil Pollution 223:519-532.
461
462
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Figure 1. Scheme of the five field-scale plots setup in the tailings pond before amendments application (a) and
the spontaneous vegetation cover after five years of the treatment (b). MW: marble waste; PS: pig slurry; TL:
tilled; CT: control (untilled and unamended).
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Figure 2. Distribution of Zn in the different parts of A. halimus (a), bioaccumulation index (b) and translocation
index (c). MW: marble waste; PS: pig slurry; TL: tilled; CT: control (n=3).
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Figure 3. Distribution of Pb in the different parts of A. halimus (a), bioaccumulation index (b) and translocation
index (c). MW: marble waste; PS: pig slurry; TL: tilled; CT: control (n=3).
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Figure 4. Distribution of Cd in the different parts of A. halimus (a), bioaccumulation index (b) and translocation
index (c). MW: marble waste; PS: pig slurry; TL: tilled; CT: control (n=3).
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Figure 5. Distribution of Cu in the different parts of A. halimus (a), bioaccumulation index (b) and translocation
index (c). MW: marble waste; PS: pig slurry; TL: tilled; CT: control (n=3).
28. ACCEPTED MANUSCRIPT
Highlights
Tailings reclamation by plants was studied at presence of pig slurry and marble waste
After five years, A. halimus was the most dominant species growing on the tailing
A. halimus accumulated metals in leaf higher than in stem and root
The treatments decreased bioaccumulation index of Zn and Cd but increased that of Pb
Salinity increased leaf Pb and Cd rates in marble waste plot
29.
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Table 1. Soil properties in bare soil before and after 5-years of the amendments application (n=3)
pH EC† (dS m-1) OC (g kg-1)Treatment
Before application
CT* 7.4 ±0.2b1 2.6 ±0.5c 9.5 ±0.3b
TL 7.6 ±0.3b 3.3 ±0.6bc 11.0 ±0.4b
PS 7.5 ±0.4b 2.6 ±0.3c 11.4 ±0.3b
MW 7.8 ±0.2b 7.6 ±0.8a 13.2 ±0.2a
MW+PS 7.9 ±0.1ab 3.8 ±0.8b 11.3 ±0.3b
5-years after application
CT 7.2 ±0.1b 2.7 ±0.0c 10.8 ±1.0b
TL 7.6 ±0.0b 2.7 ±0.0c 7.6 ±0.4c
PS 7.3 ±0.0b 2.7 ±0.0c 9.4 ±0.9bc
MW 8.4 ±0.0a 7.1 ±0.2a 9.9 ±0.6b
MW+PS 7.6 ±0.1b 3.0 ±0.0bc 10.8 ±0.7b
*CT: Untilled and unamended tailings (Control), TL: Tilled tailings, PS: Pig slurry application, MW: Marble waste application. †EC:
electrical conductivity; OC: organic carbon. 1Different letters indicate significant differences (p<0.05) between means after an ANOVA
test.
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Table 2. Total, DTPA and water- extractable metals in bare soils sampled before and after five years of the
amendments application (mean ± standard deviation)(n=3)
DTPA-extractable, mg kg-1 Water-extractable, mg kg-1
Treatment
Zn Pb Cd Cu Zn Pb Cd Cu
Before amendments application
CT* 135 ±38d1 345 ±29a 1.1 ±0.5d 2.3 ±0.3cd 7.5 ±0.3c 0.2 ±0.1c 0.11 ±0.01bc 0.01 ±0.00c
TL 392 ±27b 52 ±19d 3.6±0.1b 4.6 ±0.9b 1.1 ±0.2d 0.4 ±0.0b 0.07 ±0.01bc 0.02 ±0.01b
PS 402 ±27b 62 ±23d 1.6 ±0.2d 3.1 ±1.3c 1.7 ±0.1d 0.3 ±0.0bc 0.04 ±0.01c 0.02 ±0.01b
MW 398 ±3b 38 ±27d 6.1 ±0.2a 4.4 ±0.6b 1.1 ±0.5d 0.4 ±0.0b 0.10 ±0.01b 0.03 ±0.01b
MW+PS 402 ±10b 35 ± 17d 4.1 ±0.1b 3.7 ±0.4bc 1.0 ±0.3d 0.6 ±0.1a <dl 0.03 ±0.01b
Five years after amendment application
CT 128 ±2d 350 ±5a 1.0 ±0.0d 2.1 ±0.2cd 6.4 ±0.4c 0.2 ±0.0c 0.08 ±0.00bc <dl
TL 251 ±4c 143 ±3b 1.2 ±0.1d 2.5 ±0.4cd 7.0 ±0.1c <dl 0.09 ±0.01bc 0.03 ±0.00b
PS 262 ±5c 106 ±4c 2.2 ±0.2c 2.3 ±0.2cd 9.7 ±0.1b <dl 0.15 ±0.01b 0.08 ±0.01a
MW 481 ±14a 42 ±1d 5.0 ±0.1b 4.6 ±0.3b 11.4 ±0.6b 0.2 ±0.1c 0.12 ±0.03b 0.02 ±0.00b
MW+PS 527 ±13a 103 ±5c 3.4 ±0.5b 6.4 ±0.5a 22.5 ±2.1a <dl 0.23 ±0.02a 0.07 ±0.02a
*CT: Untilled and unamended tailings (Control), TL: Tilled tailings, PS: Pig slurry application, MW: Marble waste application.
<dl: below detection limit. For Pb <0.05 mg kg-1, for Cu< 0.01 mg kg-1; for Cd < 0.01 mg kg-1. 1Different letters indicate significant
differences (p<0.05) between means after an ANOVA test.
