Will wheat be damaged by heavy metals on exposure to coal fly ash?

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Will wheat be damaged by heavy metals on exposure to coal fly ash? Xin Xiao, Kai Qin, Xiaofei Sun, Wang Hui∗, Limei Yuan, Lixin Wu College of Environment Science and Spatial Informatics, Chinese University of Mining Technology, 221116 Jiangsu, China

A R T I C L E I N F O

A B S T R A C T

Keywords: Coal-fired power plants Heavy metals Atmospheric particles Wheat seedling Ultrastructural alteration

The mining industry and coal-fired power plants are among the major sources of atmospheric particles that cause air pollution. Can crops be damaged as they are exposed to foliar deposits of particulate matter enriched with various metal(loid)s from coal-mining regions? This study investigated the effects of atmospheric particles on wheat seedlings grown in a region north of Xuzhou, East China, where five coal mines, three coal-fired power plants, two large fly ash yards, and several coal yards coexist. More than two-thirds of the land area of this region is cropland. Atmospheric deposition analysis showed that the daily bulk deposition fluxes of Cr were generally higher in this region than those in other regions worldwide, and the highest fluxes were found in the area between the two-major coal-fired power plants. Wheat was grown under different soil and atmospheric conditions in the coal-fired region (north of Xuzhou) and a non-coal-fired region (south of Xuzhou). Heavy metal analysis showed that the accumulation of Cu, Cd, Pb, Zn, Cr, and As in the cell wall, cell organelles, and soluble fraction of wheat seedling shoots, particularly the percentage in the cell wall, was higher in the coal-fired region than in the non-coal-fired region. Analysis of the changes in the ultrastructure of seedling root and leaf cells revealed that atmospheric particles released in the coal-fired region damages the cellular structure of various parts of the wheat seedling and affects photosynthetic processes by damaging the chloroplasts.

1. Introduction Coal mining and burning are major anthropogenic sources of atmospheric particles and heavy metals (Wang et al., 2011). Atmospheric particles can reduce the total irradiance and diffuse solar radiation reaching the Earth's surface (Chang et al., 2009; Kara et al., 2014; Kulmala et al., 2004), the main drivers of photosynthesis in terrestrial plants. The proportion of diffused radiation reaching the plant canopy affects the light-use efficiency for canopy photosynthesis (Cheng et al., 2015; Mercado et al., 2009). The plant response to diffused radiation is also dependent on the characteristics of the plant, such as functional type, leaf physiology, leaf area, leaf inclination, canopy structure, and shape (Kanniah et al., 2012). The deposited particles are inert, so airborne particles are important heavy-metal carriers, especially in coal-mining regions. Heavy metals are commonly taken up by the surface of dust particles and are normally present in loosely bound forms, which are highly mobile and potentially bioavailable (Marx et al., 2008). Atmospheric particles exist in various sizes; the residence time of the particles in the atmosphere and their association with heavy metals depend on the particle size. Coarse particles settle near the particle source, whereas fine particles settle slowly (Candeias et al., 2014). Thus, deposition is a significant

pathway for the transfer of heavy metals from the atmosphere to terrestrial surfaces and soils. Several studies have reported that the plant canopy can efficiently adsorb particulate matter (PM) and reduce the PM ratio by capturing airborne PM on their foliar parts (Shahid et al., 2017), strongly affecting the elemental composition of wheat grain in mining and industry polluted area (Bermudez et al., 2012). Additionally, the growth and yield of rice and wheat crops grown near to the thermal power plant in Uttar Pradesh were reduced compared with fields located farther away (Chakrabarti et al., 2014). The characteristics of the particles, such as physical, chemical, and nutrient properties, may affect vegetation health (Burkhardt, 2010; Pariyar et al., 2013; Yan et al., 2014). Heavy metals emitted from mining and industrial areas may attach to atmospheric particles in highly mobile and potentially bioavailable forms (Eqani et al., 2016). Foliar absorption of heavy metals due to PM deposition is therefore of great concern (Bermudez et al., 2011). However, unlike root absorption (Boussen et al., 2013; Jamali et al., 2009), fewer studies have focused on the uptake of heavy metals from plant shoots. In the present study, we investigated the plant response to phytotoxicity at the cellular level of wheat seedlings growing in a region where atmospheric particles are released from coal-fired electricity-

Peer review under responsibility of Turkish National Committee for Air Pollution Research and Control. ∗ Corresponding author. E-mail address: [email protected] (W. Hui). https://doi.org/10.1016/j.apr.2018.01.019 Received 9 August 2017; Received in revised form 26 January 2018; Accepted 31 January 2018 1309-1042/ © 2018 Turkish National Committee for Air Pollution Research and Control. Production and hosting by Elsevier B.V.

Please cite this article as: Xiao, X., Atmospheric Pollution Research (2018), https://doi.org/10.1016/j.apr.2018.01.019

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Fig. 1. Experimental area and sampling sites located north of Xuzhou, northwest Jiangsu Province, east China. The major coal-fired power plants are Huarun, Huamei, Pengcheng, and Chacheng. The major coal mine is Pangzhuang

cropland, and almost all coal mines and electricity plants are surrounded by cropland. Wheat, paddy rice, maize, and soybean are the main crops produced in this area.

producing power plants. The results were compared with those for plants growing in a non-coal-fired region with relatively clean air. The aims of the study were to: (a) evaluate the atmospheric pollution in a coal-fired electricity-producing region via atmospheric deposition analysis; (b) determine the subcellular distribution of heavy metals in wheat seedling shoots exposed to aerosols using tissue fractionation; and (c) investigate the ultrastructural alterations of plant cells induced by heavy metals through autometallography.

2.2. Bulk deposition sampling Bulk deposition samples were collected using polyethylene containers with a 0.15-m-diameter mouth, which were covered with a polyethylene web to prevent large parts of materials entering (Bermudez et al., 2012). Six containers were located at six sites surround two primary sources of pollution from mining in the north and south of Xuzhou, respectively (Fig. 1) and fitted on artificial roofs at approximately 4 m above the ground. Both rainwater and the settling particles were collected over a period of 180 days from 1 November 2014 to 1 May 2015.

2. Materials and methods 2.1. Study site The study was conducted in a coal-fired electricity-producing region around 8 km northwest of Xuzhou, Jiangsu Province, China (34°32′–34°36′ N and 117°07′–117°19′ E, Fig. 1). The climate in this area is characterized by a typical warm humid monsoon with an average annual temperature of 14 °C and rainfall of 900 mm. The bedrock consists mainly of carboniferous grey limestone, and the microlandform is mainly composed of extensive diluvial plains and sporadic uplands. The main soil is typical fluvo-aquic soil formed on alluvium (Wang et al., 2005). Xuzhou has exploited coal mines for over 124 years. In 2006, the city produced 259.7 million tons of coal and generated 7.2 × 109 kWh of electric power (Huang et al., 2009). Most of the coal mines and coalfired power plants are located in the northern part of the city and bitumite is popular in the section. Our study site is more than 50 km2 contains five coal mines, three coal-fired electric plants, two large fly ash yards, and several coal yards. More than two-thirds of the area is

2.3. Plant growth and treatment Wheat (Triticum aestivum Anti-50) seeds were supplied by Xuzhou Seeds Company (Xuzhou, Jiangsu, China). All seeds were sterilized with 10% H2O2 for 20 min and then washed several times with ultrapure water before use. Soil was obtained from a non-coal-fired region located south of Xuzhou and the coal-fired electricity-producing region located north of Xuzhou. Table 1 shows the heavy-metal concentrations in each soil from the different regions of Xuzhou and Table 2 shows the daily bulk deposition fluxes of heavy metals in each region. Each sample was assayed in triplicate. The experiments were performed in plastic pots with a diameter of 35 cm and a depth of 30 cm during November 5-25, 2015. Each pot 2

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Table 1 Heavy-metal concentrations of the soil samples (mg kg−1). Sample

Zn

Cu

Pb

Cd

Cr

As

Soil from non-coal-fired region south of Xuzhou Soil from coal-fired region north of Xuzhou

65.30 ± 2.45 119.82 ± 3.74

16.96 ± 1.27 31.53 ± 2.79

15.68 ± 1.43 20.00 ± 1.94

0.5784 ± 0.12 1.18 ± 0.19

56.04 ± 3.87 80.34 ± 4.28

11.83 ± 1.04 32.69 ± 3.42

2.6. Tissue fractionation

contained 20 seedlings. The experiment consisted of three treatments: 1) no exposure (T1); 2) soil exposure (pollutants present in the soil but not in the PM) (T2); 3) soil and atmospheric exposure (T3). T3 was located north of Xuzhou, avoiding roads and factories. Each treatment was performed in triplicate in three independent experiments. All analyses were performed on plant shoots and roots of 15-day-old seedlings.

The subcellular distribution of heavy metals in the shoots was determined as the ratio of heavy metals found in each subcellular fraction according to the method of Xiong et al. (2016). The shoots were homogenized in chilled extraction buffer containing 50 mM HEPES (C8H18N2O4S, pH 7.5), 500 mM sucrose, 1 mM dithiothreitol, 5 mM ascorbate, and 1% polyvinylpolypyrrolidone. To isolate the cell wall fraction, the homogenate was centrifuged at 500 × g for 5 min. The supernatant was subsequently centrifuged at 20,000 × g for 45 min to isolate the cell organelles from the supernatant solution, which was the soluble fraction. All procedures were performed at 4 °C. After digestion (cell wall and organelle-containing fractions) with concentrated HNO3/ HClO4 (4:1, v/v), the metal concentrations in various fractions were quantified by inductively coupled plasma atomic emission spectroscopy.

2.4. Metal determination Heavy-metal assays were conducted for deposition, soil, and shoots of the wheat seedlings. The deposition and soil samples in each pot were each digested in a 16 mL tetra-acid mixture (made up of four 99.9% highly pure guarantee regents, HCl, HNO3, HF, and HClO4, in a 3:1:3:1 ratio) (GB17134-17141-1997). 20 plants in each pot was mixed as a composite sample and the seedlings in each pot were pre-washed in tap water, and then washed with distilled water or 1 M HCl followed by 1 M Na2EDTA to remove the metals bound to cell wall components. The processed shoots and roots were oven dried at 80 °C until their mass no longer changed, and then ground to a fine powder. Each sample was digested in a 7 mL mixture of HNO3 and HClO4 (6:1) and diluted in pure water up to 50 mL (Rascio et al., 2008). The concentrations of Zn, Cu, Pb, Cd, Cr, and As were measured by inductively coupled plasma atomic emission spectroscopy (PerkinElmer Optimal 8000, USA). The accuracy of the analytical procedure was checked using the standard materials GBW07427 and GBW07403 for deposition and soil samples, respectively, and GBW10046 for the seedling samples. All samples were assayed in triplicate.

2.7. Statistical analysis Data are means ± standard deviation of three replicates. One-way ANOVA was used to compare the differences (p < 0.05) in heavy-metal concentrations between the shoots of variously treated plants. Statistical analysis was performed using SPSS 12 software (IBM Crop. 2003). 3. Results and discussion 3.1. Heavy metals in atmospheric fallout The range of daily element bulk deposition (μg m−2 d−1) at all sampling sites occurred in the following descending order: Zn (82.30–137.18), Cr (28.65–39.07), Cu (11.31–28.20), Pb (9.36–20.02), and Cd (0.62–1.36). The daily bulk deposition fluxes of the analyzed metals at all sampling sites were compared with those of other areas (urban, suburban, and industrial areas) (Table 3). The Cd, Pb, Cu, and Zn fluxes obtained in this study were lower than those for the industrial area in Aliaga in Turkey, but similar to those for other mining and urban areas worldwide. Cr fluxes in the coal-fired area were higher than those of the other areas, except the industrial area in Aliaga. The deposition fluxes of Pb, Cd, Cu, and Zn were originated from PM emitted from metal working and road traffic (Mijic et al., 2010). Cu, Zn, Cd, Cr and Pb derived from resuspended road dust and road traffic (through emissions for fuel combustion, worn tires, brake linings, and road construction materials), as well as crystal particles (Amato et al., 2009; Sternbeck et al., 2002). The anthropogenic activity that produces the largest contribution to the total amount of atmospheric Cr is the metallurgical industry, followed by coal combustion (Pacyna et al., 2007). Metal deposition was highest in area between the Huarun and Huamei coal-fired power plants, followed by that near the plants and in areas downwind of the Huamei coal-fired power plant (Fig. 2). These

2.5. Light and electron microscopy Three leaves samples (second leaf in the subapical region) and root tips of plants in each pot of three treatment were fixed overnight at 4 °C in 3% glutaraldehyde in 0.1 M sodium cacodylate buffer (pH 6.9), postfixed for 2 h in 1% osmium tetroxide in the same buffer, and processed as described previously by Rascio et al. (1991). For light microscopy, leaves at no exposition and soil and atmospheric exposition were fixed thin sections (1 μm) cut by an ultramicrotome (Ultracut, Leica EM UC7, Italy) were stained with equal volumes of 1% toluidine blue and 1% sodium tetraborate and then examined (Ortholux, Leitz, Wetzlar, Germany). For electron microscopy, ultrathin sections (600 Å) of all three leaves samples were cut using the same ultramicrotome, stained with lead citrate, and observed under a transmission electron microscope (Tecnai G2 F20*, FEI, USA) operated at 75 kV. Three leaves were chosen in each pot and more than 20 ultrathin section for each sample in order to reduce the randomness of the TEM observation. More than ten photos were taking for each sample and the most typical visions were choose in the paper.

Table 2 Daily bulk deposition fluxes of heavy metals in the study areas (μg m−2 d−1). Region

Zn

Cu

Pb

Cd

Cr

Non-coal-fired region south of Xuzhou Coal-fired region north of Xuzhou

127.16 ± 5.21 109.74 ± 27.44

13.14 ± 1.79 19.76 ± 8.45

13.35 ± 2.47 14.71 ± 5.35

0.87 ± 0.14 0.99 ± 0.37

15.48 ± 2.86 33.86 ± 5.21

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Table 3 Daily bulk deposition fluxes of heavy metals (μg m−2 d−1) in various regions worldwide. Location

Period

Cd

Cr

Pb

Cu

Zn

Reference

North of Xuzhou, China Aliaga, Turkey Industry area Residential area Córdoba, Argentina Southern Scandinavia Mining Area of San Jorge River Basin, Colombia

2014–2015 2009–2010

28.65–39.07 122 ± 97 16 ± 11 22.2 0.611

9.36–20.02 1033 ± 881 110 ± 86 17.794 2.583 15.79–36.02

11.31–28.20 238 ± 143 34 ± 22 29.956 3.611 8.27–158.17

82.30–137.18 4709 ± 3901 575 ± 437 297 22.222 59.84–338.98

This study (Kara et al., 2014)

2005–2007 2002–2011 2010–2011

0.62–1.36 22 ± 21 2.3 ± 1.9 0.369 0.111 ND-12.22

Pearl River Delta, China oakforests in Montseny, Spain Varanasi, India Athabasca Oil Sand Region, Canada Belgrade, Serbia

2001–2002 1995–1996 2003–2004 2012 2002–2006

0.192 ND 0.67–10.67 0.004–0.02 0.603

17.616

34.795 1.205–7.014 ND-11.67 0.24–2.77 59.452

50.959 3.562–10.411 9.67–35.16 0.56–3.9 94.521

284.931 69.590–115.890 30.00–300.00 1.5–9.2 112.603

0.12–2.01 4.493

(Bermudez et al., 2012) (Hovmand and Kystol, 2013) (Luis Marrugo-Negrete et al., 2015) (Wong et al., 2003) (Avila and Rodrigo, 2004) (Sharma et al., 2008) (Bari et al., 2014) (Mijic et al., 2010)

ND: not detectable

area (27.6, 51.3, 54.9, 35.9, 51.1, and 49.2%, respectively; Fig. 3). Similar heavy metal concentrations were observed in the protoplast of the shoot of plants grown in T1 and T2 except Cr. Cu concentrations in protoplast of the shoots in T3 were higher significantly than that of plants in T1 and T2 (Fig. 3). Given the distribution of 70–90% of the heavy metals in PM10 and the small particle size (Mohanraj et al., 2004), the atmospheric metals accumulate in the openings of the cutin layer of the epidermis and in the stomatal apparatus, resulting in an increased heavy-metal concentrations in the shoots (Uzu et al., 2010). In the present study, 52.6% Cd, 55.0% Pb, 51.1% Cr, 49.1% As, 27.6% Cu, and 35.2% Zn were accumulated in the apoplast of the shoot of plants in soil and atmospheric exposition; these amounts were significantly higher than those in the control (Fig. 3). Thus, the deposition of atmospheric particles evidently enhanced the heavy-metal absorption by the shoots of wheat seedlings.

3.3. Subcellular distribution of heavy metals in wheat shoots Most of the six metal concentrations in the cell wall, organelles, and soluble fraction of shoots exposed to atmospheric particles coal fire area were higher than those in no exposure and soil exposure. In general, the cell walls had the highest proportion of heavy metals (26.7–64.8%), followed by the soluble fractions (18.9–53.6%), with the lower proportion found in the organelles (3.9–26.5%). Almost all of the six metals in each subcellular fraction of shoots increased on exposure to the polluted soil and polluted atmosphere, and the increase rate in the organelles and soluble fractions was higher than that in the cell wall. The proportions of Cu, Zn, Pb, Cd, and Cr in shoot cell walls grown in the coal-fired area were higher than those in the control, whereas As decreased from 55.41 to 39.9%, compared with the control. The percentage of most metals in organelles of total concentration increased on exposure to polluted air compared with the control, except Cu and Cr, whereas the percentage in the soluble fraction decreased (Table 4 and Fig. 4). Compared with the control, the percentage of metal was lower in the cell wall but higher in organelles in the shoots of the atmospheric control. The percentage of metal in the cell wall increased on exposure to the polluted atmosphere, except As. In terms of heavy-metal subcellular distribution, plants usually bind most metals to the cell wall (Allan and Jarrell, 1989) or accumulate metals in the vacuoles instead of in the highly active parts of cells (Li et al., 2013). Cell walls contain proteins and polysaccharides, which have great potential for acting as ligands for binding metals (Zeng et al., 2011). The final step of metal detoxification in the cell is the sequestration into vacuoles, which mainly depends on glutathione and phytochelatins (Song et al., 2010). Our results showed that the airborne heavy metals increased the metal percentage in the cell wall, compared with the atmospheric control.

Fig. 2. Spatial variations in deposition fluxes of particulate matter at the sampling sites. 1. Heizhang is located between two major coal-fire power plant; 2. Zhangxialou is nearby a coal mine well; 3. Huamei is located in the place near the fly ash field; 4. Lizhuang is located the south of a main road; 5. shixi is located in the south of section; 6. Jiahe hospital is located in Jiahe coal mine.

results indicated that coal combustion in the study area is an important source of heavy metals in the atmosphere. 3.2. Heavy metals in plant shoots The wheat seedlings were exposed to various soil and atmospheric conditions. The shoots of the plants grown in T3 showed the highest heavy-metal concentrations (p < 0.05; Fig. 3), Cu, Zn, Pb and Cr in T3 were 8.1%, 10.0%, 33.1%, and 35.1% higher than T2, respectively and 33.4%, 18.1%, 44.4% and 47.6% higher than T1, respectively. These decreased after washing the shoots with EDTA because the metals bound to the cell wall were removed (Xiong et al., 2016), and it can be consider as metals accumulate at protoplast of the shoot. The concentrations of the six analyzed metals in the shoots washed with EDTA in T1 and T2 decreased compared with those of the shoots washed with H2O, the highest decline were shown as 30.3% (Pb in T1) and 35.6% (Cu in T2). The rate of reduction in the concentrations of Cu, Cd, Pb, Zn, Cr, and As was higher in the shoots of plants located in the coal-fired 4

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Fig. 3. Heavy-metal concentrations in shoots of wheat seedlings grown under the three different treatments after washing with H2O or EDTA (DW = dry weight). Treatment 1, soil obtained from the non-coal-fired region and located in the non-coal-fired region (no exposure); Treatment 2, soil obtained from the coal-fired electricity-producing region and located in the non-coal-fired region (soil exposure); and Treatment 3, soil obtained from the coal-fired electricity-producing region and located in the coal-fired electricity-producing region with polluted air (soil and atmospheric exposure). Data are the means ± standard deviations of three replicates. Bars represent standard deviation of three independent replicates. Bars that do not share the same letters within the same fraction are significantly different at P < 0.05 as determined by a least significant difference multiple range test.

Table 4 Heavy-metal distribution in cell wall, cell organelles, and soluble fraction of wheat shoots under different soil and atmospheric conditions. Subcellular fraction (mg kg−1)

Cu

Cell wall

2.57 3.29 3.71 1.21 1.83 1.53 3.30 3.26 3.87

Cell organelles

Soluble fraction

T1 T2 T3 T1 T2 T3 T1 T2 T3

± ± ± ± ± ± ± ± ±

0.009c 0.014b 0.011a 0.004c 0.002a 0.011b 0.009b 0.020b 0.005a

Zn

As

Pb

7.66 ± 0.009b 11.48 ± 0.006b 15.88 ± 0.015a 5.64 ± 0.004c 8.40 ± 0.019a 7.13 ± 0.007b 15.40 ± 0.029a 11.81 ± 0.009b 12.18 ± 0.007b

0.34 ± 0.010b 0.38 ± 0.186b 0.68 ± 0.403a 0.024 ± 0.005a 0.10 ± 0.137a 0.21 ± 0.627a 0.25 ± 0.043b 0.37 ± 0.310b 0.82 ± 0.055a

0.58 0.51 0.79 0.15 0.24 0.38 0.17 0.32 0.42

Cd ± ± ± ± ± ± ± ± ±

0.051b 0.027b 0.016a 0.097c 0.031b 0.027a 0.040c 0.040b 0.001a

0.019 0.026 0.098 0.004 0.008 0.028 0.018 0.016 0.063

Cr ± ± ± ± ± ± ± ± ±

0.035b 0.136b 0.138a 0.068a 0.298a 0.186a 0.154a 0.191a 0.218a

3.51 3.75 8.83 1.32 1.81 2.33 3.98 5.35 5.64

± ± ± ± ± ± ± ± ±

0.004b 0.015b 0.005a 0.008c 0.020b 0.007a 0.007c 0.016b 0.011a

T1, no exposure; T2, soil exposure; and T3, soil and atmospheric exposure. Data are means ± S.E. (n = 4). Values in the same column followed by different letters differ significantly (p < 0.05).

Fig. 4. Subcellular distribution of heavy metals in tissues of wheat shoots grown under different soil and atmospheric conditions. (A) no exposure, (B) soil exposure, and (C) soil and atmospheric exposure.

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Fig. 5. Micrographs of longitudinal sections of wheat root tips grown in different conditions. Numerous dark inclusions can be distinguished in the cell. (A) Root in no exposure, in which regularly arranged cell rows were observed. (B) Root in soil and atmosphere exposure. Notably, enlarged roots and disordered, voluminous, and misshapen epidermal and cortical cells were observed. Numerous large dark inclusions were recognizable in the cell. ( × 400 magnification).

Fig. 6. Transmission electron micrographs of wheat root grown in various conditions. (A) control; (B) deformation of cells and nucleus (no exposure); (C) vacuoles in the nucleus and plasmodesma (soil exposure); (D) vesicle formation and rupture of the nuclear membrane (soil and atmospheric exposure); (E) swelling mitochondria (soil and atmosphere exposure). Scale bars: 1 μm.

4. Heavy metal-induced cellular damage in wheat seedlings

and cytoplasm (Fig. 6D), and structural damage and swelling of the mitochondria, which then formed a globular shape (Fig. 6E). The ultrastructural alterations in the root cells of plants exposed to polluted soil and air pollution included the degradation and formation of cell wall folds, swelling of the mitochondria, rupture of the nuclear membrane, and increased number of vesicles in the nucleus (Fig. 6). Structural damage, such as formation of folds and protuberances in the cell walls (Kaur et al., 2013), rupture of cell membranes (Sanchez-Pardo et al., 2012), swelling and disintegration of mitochondria, and damage of the nucleoli (Vitoria et al., 2006), have been ascribed to heavy-metal stress. The precipitates are generally adsorbed on the cell walls and are stored in the vacuoles (Lozano-Rodriguez et al., 1997) to prevent them from rendering toxic effects while entering the cytoplasm; however, organelles become stressed with increasing heavy-metal concentrations (Yu et al., 2012). In the present study, more damage to the nucleus and greater amounts of dark precipitates were found in cytoplasm and nucleus of the root cells from plants grown in the coal-fired area compared with the atmospheric control; this suggests that airborne heavy metals were absorbed by wheat shoots and transported into the roots, thereby affecting seedling growth.

4.1. Cellular damage to roots The results of morphological analyses demonstrated that heavy metals in the soil and air have a negative impact on the cell structure and root arrangement of wheat seedlings. Using light microscopy, we compared the longitudinal section of the root tip of the control (Fig. 5A) with a plant grown in the coal-fired area (Fig. 5B). The results revealed disordered rows of considerably enlarged and misshapen cells of the epidermis and the outer cortical layers of plants exposed to polluted soil and air. Furthermore, the enlarged cells were not lengthened, but demonstrated atrophy. In both roots, numerous dark inclusions were distinguishable in most cells, although much larger dark inclusions were seen in the cells of the plants of T3 (Fig. 5A and B). Plants grown in atmosphere control showed deformation of the cell and nucleus, the presence of vesicles in the nucleus, rupture of the nuclear membrane, vacuolation of the plasmodesma, and the presence of dark precipitates of metals in the nucleus of plants grown in the coalfired area (Fig. 6B and C) compared with control plants. Compared with the atmospheric control, exposure to both polluted soil and polluted air resulted in the presence of more vesicles in the nucleus, more lesions in the nuclear membrane, the presence of dark precipitates in the cell wall 6

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Fig. 7. Transmission electron micrographs of wheat leaf cells grown in various conditions. (A) control; (B) swollen mitochondria (atmospheric control); (C) chloroplast containing dark precipitates (atmospheric control); (D) globular chloroplasts and ruptured membrane (coal-fired area); (E) dark precipitates in chloroplasts (coal-fired area). Scale bars: 1 μm.

deposition of Cr fluxes in the coal-fired region was higher than those in other areas worldwide. The airborne heavy metals significantly increased heavy-metal accumulation in the cell wall of wheat seedlings, which in turn deformed and severely damaged the nucleus of the cells of the root tip and altered the ultrastructure of the chloroplast. Further studies should be performed to elucidate the impact of airborne heavy metals on various physiological and biochemical processes during wheat growth and on the quality of this and other crops.

4.2. Cellular damage to shoots In the shoot cell of the control, the chloroplasts were closely distributed underneath the plasma membrane of cells, exhibiting ellipsoid structures. The stroma and thylakoid of the chloroplast were also well developed, and the lamellar structure of the thylakoid in the chloroplast was clear and intact (Fig. 7A). Swollen mitochondria with disordered cristae (Fig. 7B), an integral chloroplast containing dark precipitates, and thylakoids with a loose lamellar structure (Fig. 7C) were observed in the leaf cells of the atmospheric control. When exposed to both polluted soil and polluted air, the chloroplast was obviously deformed and contained an abundance of stroma and thylakoids (Fig. 7D). Moreover, the chloroplast membrane was ruptured, and large dark precipitates were observed in the membrane or in the chloroplast (Fig. 7E). Photosynthesis occurs in the chloroplasts of plant cells. Therefore, normal photosynthesis depends on the integrity of the chloroplast ultrastructure. Ultrastructural investigation of the leaf cells revealed that the airborne heavy metals damaged the ultrastructure of chloroplasts by deforming and dilating the thylakoid membranes (Fig. 7D and E). These ultrastructural alterations indicate that heavy metals disrupt the metabolic functions and affect the lipid composition of chloroplast membranes (Cai et al., 2011; Wang et al., 2009). Larger dark precipitates were also observed in the chloroplast and in the cytoplasm of plants grown in the coal-fired area (Fig. 7D and E). Thus, greater metal accumulation occurred on exposure to atmospheric particles, and the chloroplast was damaged by multiple heavy metals in wheat seedlings.

Acknowledgments This work was supported by the Fundamental Research Funds for the Central Universities of China (2014QNA32). References Allan, D.L., Jarrell, W.M., 1989. Proton and copper adsorption to maize and soybean root cell walls. Plant Physiol. 89, 823–832. Amato, F., Pandolfi, M., Viana, M., Querol, X., Alastuey, A., Moreno, T., 2009. Spatial and chemical patterns of PM10 in road dust deposited in urban environment. Atmos. Environ. 43, 1650–1659. Avila, A., Rodrigo, A., 2004. Trace metal fluxes in bulk deposition, throughfall and stemflow at two evergreen oak stands in NE Spain subject to different exposure to the industrial environment. Atmos. Environ. 38, 171–180. Bari, M.A., Kindzierski, W.B., Cho, S., 2014. A wintertime investigation of atmospheric deposition of metals and polycyclic aromatic hydrocarbons in the Athabasca Oil Sands Region, Canada. Sci. Total Environ. 485, 180–192. Bermudez, G.M.A., Jasan, R., Pla, R., Luisa Pignata, M., 2011. Heavy metal and trace element concentrations in wheat grains: assessment of potential non-carcinogenic health hazard through their consumption. J. Hazard Mater. 193, 264–271. Bermudez, G.M.A., Jasan, R., Pla, R., Pignata, M.L., 2012. Heavy metals and trace elements in atmospheric fall-out: their relationship with topsoil and wheat element composition. J. Hazard Mater. 213, 447–456. Boussen, S., Soubrand, M., Bril, H., Ouerfelli, K., Abdeljaouad, S., 2013. Transfer of lead, zinc and cadmium from mine tailings to wheat (Triticum aestivum) in carbonated Mediterranean (Northern Tunisia) soils. Geoderma 192, 227–236. Burkhardt, J., 2010. Hygroscopic particles on leaves: nutrients or desiccants? Ecol. Monogr. 80, 369–399. Cai, Y., Cao, F., Wei, K., Zhang, G., Wu, F., 2011. Genotypic dependent effect of exogenous glutathione on Cd-induced changes in proteins, ultrastructure and antioxidant defense enzymes in rice seedlings. J. Hazard Mater. 192, 1056–1066.

5. Conclusion This study investigated the influence of the heavy metals present in the soil and atmosphere on wheat grown in a coal-fired electricityproducing region. Results revealed severe atmospheric pollution in the coal-fired electricity-producing region compared with the non-coalfired region in the same city. In particular, the daily element bulk 7

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of urban Coimbatore. Arch. Environ. Contam. Toxicol. 47, 162–167. Pacyna, E.G., Pacyna, J.M., Fudala, J., Strzelecka-Jastrzab, E., Hlawiczka, S., Panasiuk, D., Nitter, S., Pregger, T., Pfeiffer, H., Friedrich, R., 2007. Current and future emissions of selected heavy metals to the atmosphere from anthropogenic sources in Europe. Atmos. Environ. 41, 8557–8566. Pariyar, S., Eichert, T., Goldbach, H.E., Hunsche, M., Burkhardt, J., 2013. The exclusion of ambient aerosols changes the water relations of sunflower (Helianthus annuus) and bean (Vicia faba) plants. Environ. Exp. Bot. 88, 43–52. Rascio, N., Mariani, P., Tommasini, E., Bodner, M., Larcher, W., 1991. Photosynthetic strategies in leaves and stems of Egeria densa. Planta 185, 297–303. Rascio, N., Vecchia, F.D., La Rocca, N., Barbato, R., Pagliano, C., Raviolo, M., Gonnelli, C., Gabbrielli, R., 2008. Metal accumulation and damage in rice (cv. Vialone nano) seedlings exposed to cadmium. Environ. Exp. Bot. 62, 267–278. Sanchez-Pardo, B., Fernandez-Pascual, M., Zornoza, P., 2012. Copper microlocalisation, ultrastructural alterations and antioxidant responses in the nodules of white lupin and soybean plants grown under conditions of copper excess. Environ. Exp. Bot. 84, 52–60. Shahid, M., Dumat, C., Khalid, S., Schreck, E., Xiong, T., Niazi, N.K., 2017. Foliar heavy metal uptake, toxicity and detoxification in plants: a comparison of foliar and root metal uptake. J. Hazard Mater. 325, 36–58. Sharma, R.K., Agrawal, M., Marshall, F.M., 2008. Heavy metal (Cu, Zn, Cd and Pb) contamination of vegetables in urban India: a case study in Varanasi. Environ. Pollut. 154, 254–263. Song, W.-Y., Park, J., Mendoza-Cozatl, D.G., Suter-Grotemeyer, M., Shim, D., Hoertensteiner, S., Geisler, M., Weder, B., Rea, P.A., Rentsch, D., Schroeder, J.I., Lee, Y., Martinoia, E., 2010. Arsenic tolerance in Arabidopsis is mediated by two ABCCtype phytochelatin transporters. Proc. Natl. Acad. Sci. U.S.A. 107, 21187–21192. Sternbeck, J., Sjodin, A., Andreasson, K., 2002. Metal emissions from road traffic and the influence of resuspension - results from two tunnel studies. Atmos. Environ. 36, 4735–4744. Uzu, G., Sobanska, S., Sarret, G., Munoz, M., Dumat, C., 2010. Foliar lead uptake by lettuce exposed to atmospheric fallouts. Environ. Sci. Technol. 44, 1036–1042. Vitoria, A.P., Da Cunha, M., Azevedo, R.A., 2006. Ultrastructural changes of radish leaf exposed to cadmium. Environ. Exp. Bot. 58, 47–52. Wang, L., Zhou, Q., Huang, X., 2009. Photosynthetic responses to heavy metal terbium stress in horseradish leaves. Chemosphere 77, 1019–1025. Wang, X.S., Qin, Y., Sang, S.X., 2005. Accumulation and sources of heavy metals in urban topsoils: a case study from the city of xuzhou, China. Environ. Geol. 48 (1), 101–107. Wang, Y., Hopke, P.K., Chalupa, D.C., Utell, M.J., 2011. Effect of the shutdown of a coalfired power plant on urban ultrafine particles and other pollutants. Aerosol. Sci. Technol. 45, 1245–1249. Wong, C.S.C., Li, X.D., Zhang, G., Qi, S.H., Peng, X.Z., 2003. Atmospheric deposition of heavy metals in the pearl river delta, China. Atmos. Environ. 37, 767–776. Xiong, T., Austruy, A., Pierart, A., Shahid, M., Schreck, E., Mombo, S., Dumat, C., 2016. Kinetic study of phytotoxicity induced by foliar lead uptake for vegetables exposed to fine particles and implications for sustainable urban agriculture. J. Environ. Sci. 46, 16–27. Yan, X., Shi, W.Z., Zhao, W.J., Luo, N.N., 2014. Impact of aerosols and atmospheric particles on plant leaf proteins. Atmos. Environ. 88, 115–122. Yu, Z., Yong-Han, X., Hong-Yin, Y., Ji-Ming, G., 2012. Vacuolar membrane transporters OsVIT1 and OsVIT2 modulate iron translocation between flag leaves and seeds in rice. Plant J. Cell & Molecular Bio.l 72, 400–410. Zeng, F., Zhou, W., Qiu, B., Ali, S., Wu, F., Zhang, G., 2011. Subcellular distribution and chemical forms of chromium in rice plants suffering from different levels of chromium toxicity. J. Plant Nutr. Soil Sci. 174, 249–256.

Candeias, C., Melo, R., Avila, P.F., da Silva, E.F., Salgueiro, A.R., Teixeira, J.P., 2014. Heavy metal pollution in mine-soil-plant system in S. Francisco de Assis - panasqueira mine (Portugal). Appl. Geochem. 44, 12–26. Chakrabarti, B., Singh, R., Bhatia, A., Singh, S.D., Singh, B., 2014. Impact of aerial deposition from thermal power plant on growth and yield of rice (Oryza sativa) and wheat (Triticum aestivum). Indian J. Agric. Sci. 84, 602–606. Chang, D., Song, Y., Liu, B., 2009. Visibility trends in six megacities in China 1973-2007. Atmos. Res. 94, 161–167. Cheng, S.J., Bohrer, G., Steiner, A.L., Hollinger, D.Y., Suyker, A., Phillips, R.P., Nadelhoffer, K.J., 2015. Variations in the influence of diffuse light on gross primary productivity in temperate ecosystems. Agric. For. Meteorol. 201, 98–110. Eqani, S.A.M.A.S., Kanwal, A., Bhowmik, A.K., Sohail, M., Ullah, R., Ali, S.M., Alamdar, A., Ali, N., Fasola, M., Shen, H., 2016. Spatial distribution of dust-bound trace elements in Pakistan and their implications for human exposure. Environ. Pollut. 213, 213–222. Hovmand, M.F., Kystol, J., 2013. Atmospheric element deposition in southern Scandinavia. Atmos. Environ. 77, 482–489. Huang, S., Hua, M., Feng, J., Zhong, X., Jin, Y., Zhu, B., Lu, H., 2009. Assessment of selenium pollution in agricultural soils in the xuzhou district, northwest Jiangsu, China. J.Environ. Sci.-China 21, 481–487. Jamali, M.K., Kazi, T.G., Arain, M.B., Afridi, H.I., Jalbani, N., Kandhro, G.A., Shah, A.Q., Baig, J.A., 2009. Heavy metal accumulation in different varieties of wheat (Triticum aestivum L.) grown in soil amended with domestic sewage sludge. J. Hazard Mater. 164, 1386–1391. Kanniah, K.D., Beringer, J., North, P., Hutley, L., 2012. Control of atmospheric particles on diffuse radiation and terrestrial plant productivity: a review. Prog. Phys. Geogr. 36, 209–237. Kara, M., Dumanoglu, Y., Altiok, H., Elbir, T., Odabasi, M., Bayram, A., 2014. Seasonal and spatial variations of atmospheric trace elemental deposition in the Aliaga industrial region, Turkey. Atmos. Res. 149, 204–216. Kaur, G., Singh, H.P., Batish, D.R., Kohli, R.K., 2013. Lead (Pb)-induced biochemical and ultrastructural changes in wheat (Triticum aestivum) roots. Protoplasma 250, 53–62. Kulmala, M., Suni, T., Lehtinen, K.E.J., Dal Maso, M., Boy, M., Reissell, A., Rannik, U., Aalto, P., Keronen, P., Hakola, H., Back, J.B., Hoffmann, T., Vesala, T., Hari, P., 2004. A new feedback mechanism linking forests, aerosols, and climate. Atmos. Chem. Phys. 4, 557–562. Li, Z., Wu, L., Hu, P., Luo, Y., Christie, P., 2013. Copper changes the yield and cadmium/ zinc accumulation and cellular distribution in the cadmium/zinc hyperaccumulator Sedum plumbizincicola. J. Hazard Mater. 261, 332–341. Lozano-Rodriguez, E., Hernandez, L.E., Bonay, P., Carpena-Ruiz, R.O., 1997. Distribution of cadmium in shoot and root tissues of maize and pea plants: physiological disturbances. J. Exp. Bot. 48, 123–128. Luis Marrugo-Negrete, J., Urango-Cardenas, I.D., Burgos Nunez, S.M., Diez, S., 2015. Atmospheric deposition of heavy metals in the mining area of the San Jorge river basin, Colombia (vol. 7, pg 577, 2015). Air Quality Atmos. Health 8, 149–150. Marx, S.K., Kamber, B.S., McGowan, H.A., 2008. Scavenging of atmospheric trace metal pollutants by mineral dusts: inter-regional transport of Australian trace metal pollution to New Zealand. Atmos. Environ. 42, 2460–2478. Mercado, L.M., Bellouin, N., Sitch, S., Boucher, O., Huntingford, C., Wild, M., Cox, P.M., 2009. Impact of changes in diffuse radiation on the global land carbon sink. Nature 458, 1014–U1087. Mijic, Z., Stojic, A., Perisic, M., Rajsic, S., Tasic, M., Radenkovic, M., Joksic, J., 2010. Seasonal variability and source apportionment of metals in the atmospheric deposition in Belgrade. Atmos. Environ. 44, 3630–3637. Mohanraj, R., Azeez, P.A., Priscilla, T., 2004. Heavy metals in airborne particulate matter

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