J. For. Res. (2015) 26(3):663–671 DOI 10.1007/s11676-015-0074-4
ORIGINAL PAPER
Differential expression of zinc accumulation during two growing seasons of Acacia victoriae Ali Mahdavi1 • Khadijeh Khermandar2
Received: 27 February 2014 / Accepted: 27 August 2014 / Published online: 9 May 2015 Ó Northeast Forestry University and Springer-Verlag Berlin Heidelberg 2015
Abstract It is important to understand seasonal heavy metal accumulation in different parts of plants in order to develop the best phytoremediation practices for contaminated soils. For this purpose we exposed, 1 year old A. victoriae seedlings to ZnSO4 in 4 different concentrations: 0, 50, 250 and 500 mg Zn L-1 for 45 days over two growing seasons. Subsequently, bioaccumulation of Zn in different plant tissues (roots, shoots and leafs) was assessed by Atomic Absorption Spectroscopy (AAS) for two periods. In addition, various growth attributes (dry biomass, shoot and root lengths, plant appearance) and functional traits (leaf area, chlorophyll a, b and total) were measured. The accumulation of Zn was influenced by the Zn concentration in the growth medium and the number of growing seasons. The amounts of Zn concentrated in the root tissues might indicate A. victoriae as a good option for phytostabilization of soils contaminated by Zn. We recommend that if A. victoriae is used for phytoextraction purposes, then it should be harvested at the end of the first growing season (fall) because at this time the concentrations of Zn in the above-ground parts will be maximal.
The online version is available at http://www.springerlink.com Corresponding editor: Zhu Hong & Ali Mahdavi
[email protected] Khadijeh Khermandar
[email protected] 1
Department of Forest Science, Faculty of Agriculture, University of Ilam, P.O. Box 69315-516, Ilam, Iran
2
Department Range and Watershed Management, Faculty of Agriculture, University of Ilam, P.O. Box 69315-516, Ilam, Iran
Keywords Heavy metal Phytoremediation Phytoextraction Phytostabilization
Introduction Heavy metal contamination issues are becoming increasingly common (Fernandes and Henriques 1991). Soil is the vital medium in the natural environment and heavy metal contamination is one of the most serious environmental problems in soil (Fernandes and Henriques 1991). The idea of using plants that hyperaccumulate metals to selectively remove and recycle excessive soil metals was introduced in 1983, gained public exposure in 1990, and has increasingly been examined as a potential practical and more cost-effective technology than soil replacement, solidification and washing strategies presently used (Chaney et al. 1997). Phytoremediation is best applied at sites with shallow contamination of organic, nutrient, or metal pollutants. It is well-suited for use at very large field sites where other methods of remediation are not cost-effective or practicable (Brooks 1998). To date, about 400 hyperaccumulators have been identified (Qishlaqi et al. 2009). The remediation potential of hyperaccumulators relies upon their growth rates (i.e., biomass production) and metal accumulation rate (g metal per kg of plant tissue). In fact, most hyperaccumulators produce little biomass. This could be compensated by high biomass plants even if they are usually not metal-specific and accumulate low to average heavy metal concentrations (Roosens et al. 2003). Thus, phytoremediation could be a promising technique for removing soil pollutants where hyperaccumulators or accumulators are used to take up large quantities of pollutant metals (Salt et al. 1995; Roosens et al. 2003). Both essential and unessential metals ruin the balance of the ecosystem, with
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increasing economic loss and human health damage (Qishlaqi et al. 2009). The most important toxic elements belong to a group of metals, including Copper (Cu), Zinc (Zn) and Lead (Pb) and more rarely Cadmium (Cd), Chromium (Cr), Cobalt (Co), Nickel (Ni) and Mercury (Hg). A few metals, including Cu, Mn and Zn, are required by plants in trace amounts. It is only when metals are presented in bioavailable forms at excessive levels where they have the potential to become toxic to plants (Reichman 2002). Zinc is an essential nutrient for plants. This element is a co-factor requirement for the structure and function of numerous proteins (Grotz and Guerinot 2006), energy production and structural integrity of membranes (Hansch and Mendel 2009). Zn deficiency is a common problem in plants grown in high pH, calcareous soils (as also for Fe deficiency) (Cakmak et al. 1996). Zn can be toxic, and affected plants can show symptoms similar to those resulting from other heavy metal toxicities, such as those of Cd or Pb (Foy et al. 1978). High concentrations of Zn inhibit many plant metabolic functions resulting in retarded growth and senescence. Zn toxicity in plants limits the growth of both roots and shoots and produces young leaf chlorosis. Even though Zn is not redox active, high levels of this metal are toxic because it can displace other metals (e.g. Fe, Mn and Cu) in the cell (Pilon et al. 2009; Yadav 2010). Because of this, Zn homeostasis is also strongly regulated in plant cells. Human activities releasing Zn to the environment include fossil fuel combustion and the use of sewage sludge, manure and lime. In contaminated and acid soils some crops and species have a high Zn uptake capacity (Broadley et al. 2007). The range of hyperaccumulators of Zn is [1.0 % of the dry weights of leafs and stems irrespective of the metal concentration in the soil (Raskin et al. 1994). Several studies showed that Zn concentrations in the leafs of Thlaspi calaminare and Viola calaminaria can reach 3.5 and 1.0 %, respectively (Reeves and Brooks 1983; Lasat 2000). Ernst in (1986) reported Cardaminopsis halleri (Brassicaceae) as a hyperaccumulator of Zn. This study is focused on the phytoremediation potential of Zn heavy metal contaminated soils by A. victoriae. Our assumptions for this study were that the accumulator plants take up heavy metals according to seasonal growth patterns. Therefore, matching the phytoextraction practices to those patterns can maximize the remediation efficiency of contaminated soils, and increase metal uptake and accumulation. However there is not much knowledge about the pattern of heavy metal uptake and accumulation into the tissues during the growing seasons. This understanding is important to schedule phytoremediation practices to coincide with periods of high metal accumulation in aboveground tissues thereby maximizing the phytoextraction
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efficiency. In addition, the objective of the study is to investigate the effects of high concentrations of Zn in the soil medium on growth and photosynthetic characteristics of A. victoriae. The results of this study show the potential use of A. victoriae trees for phytoremediation purposes in Zn contaminated soils.
Materials and methods Experiments were performed under natural conditions using 24 seedlings (12 seedlings for each growing season) of one year old A. victoriae that were replanted into 30 cm blackened plastic pots, filled with 2.5 kg (dry weight) of loamy-silt soils. A loamy-silt soil was prepared for filling all the pots (including the control treatments) by proportions of 1:1:2 for manure (dry animal dung), sand and soil, respectively (Table 1). The seedlings were exposed to Zn (SO4) solution in 4 different concentrations: 0 (control), 50, 250 and 500 mg Zn L-1 by supplementing them into irrigation water each time (the seedling were irrigated 20 times) during 45 days in both seasons. For each Zn concentration three replicates were evaluated. The seedlings were irrigated based on 60 % of Field Capacity (FC). Plant growth and biomass measurements At the time of harvesting (45 days after being exposed to Zn2? treatments in both growing seasons), A. victoriae seedlings were removed from the pots, separated into leaf, shoot and root portions, rinsed with deionized water, dried to constant weight at 70 °C for 48 h, and weighed for biomass determination. Plant height from the soil level to the top was measured before uprooting. Growth measurements were used to estimate the lead-tolerance index (TI). The root length can be used as an important TI (Piechalak et al. 2002). TI was calculated as follows (MaldonadoMagana et al. 2011; Mahdavi et al. 2014). Ti ð% Þ ¼
R1 100 R2
ð1Þ
where Ti is the tolerance index, R1 is the root elongation with Zn and R2 is the root elongation without Zn. Determination of pigment content Chlorophyll content in A. victoriae leaf samples was determined on fresh weight basis. Fresh leafs weighing 100 mg were pummeled and placed in 10 mL 80 % acetone in a sealed, dark bottle. The bottle was placed into a laboratory tabletop centrifuge at 3000 rotations/min for 15 min. Then, 1 mL of the surface solution was removed and adjusted with 4 mL 80 % acetone. After that, the
Differential expression of zinc accumulation during two growing seasons of Acacia victoriae
665
Table 1 Physico-chemical characterizations of control soil Soil investigation
pH
Control soil
7.2
EC (ds m-1)
Total N (%)
OM (%)
3.59
0.164
2.48
OC (%)
1.44
optical density of the solution was measured by PD-303 UV spectrophotometer at different wavelengths i.e., 645, 663 nm and chlorophyll content was calculated using relevant formulae (Strain and Svec 1966; Mahdavi et al. 2014). Estimation of Zn accumulation The dried samples (roots, shoots, and leafs) were digested in a microwave oven at high temperature (up to 235 °C) using 8 mL of HNO3 (65 %), 2 mL of H2SO4, 1 mL of HClO4 (70 %) (Moreira et al. 2011; Mahdavi et al. 2014). Zn concentrations were determined by an Atomic Absorption Spectrophotometer (AAS) (CTA-2000 AAS, Chem Tech Analytical) and the values were expressed as mg per Kg dry weight. Zn concentrations were used to estimate the bio-concentration factor (BCF) and the translocation factor (TF). Bioconcentration factor (BCF) Bioconcentration factor (BCF) was defined as the ratio of Zn concentrations in plant tissues (roots, shoots and leafs) ([Zn]plant) to Zn concentration in the soil medium ([Zn]medium) (Maldonado-Magana et al. 2011; Mahdavi et al. 2014). That is a dimensionless number representing how much of a chemical is in a tissue relevant to how much of that chemical exists in the medium. BCF ¼
Zp Zs
ð2Þ
tF ¼
Available K (mg kg-1)
Available Na (mg kg-1)
Soil texture (%) Sand
Silt
Clay
42
12
41
54
5
ZSL ZR
ð3Þ
where tF is the translocation factor, ZSL is the metal concentrations in shoots and leafs and ZR is the Zn concentration in roots. To test the phytotoxicity of Zn, the Grade of Growth Inhibition (GGI) was calculated as follows. GGI ¼ ½ðC T Þ=C
ð4Þ
where, C and T represent the dry weight of tissues of control (C) and the metal-treated plants (T) (Leita et al. 1993; Mahdavi et al. 2014). Statistical analysis Combined analysis of variance was used to estimate the average response to given treatments and to test consistency of the responses from summer season to fall season i.e. interaction of the treatment effects with seasons. Combined analysis of variance provided an overview of the magnitude of variance between the experimental seasons, the variation between treatments (Zn concentrations) and especially the treatment 9 season interaction. We used a randomized block design and the least significant difference test (LSD) for comparing means. The level of statistical significance was set at P \ 0.01 and \ 0.05. All the results are expressed as means and letters indicate statistical differences between means. The software used for the combined analysis of variance was SAS for windows program (SAS-9.1-portable).
where BCF is the Bioconcentration factor, Zp is Zn concentrations in plant tissues (roots, shoots and leafs) and Zs is the Zn concentration in the soil medium.
Results
Translocation factor
Analysis of variance
The translocation factor (TF) indicates the ability of plant to translocate heavy metals from roots to the harvestable aerial part (Waranusantigul et al. 2008; Mahdavi et al. 2014). It was calculated on a dry weight basis by dividing the metal concentration in shoots and leafs by the metal concentration in roots (Waranusantigul et al. 2008; Mahdavi et al. 2014)
In the combined analysis of variance over seasons, the effects of Zn concentrations were compared to the Zn accumulations in different plant tissues and grwoth characteristics. Some traits such as Zn contents in different tissues, TF rations and BCF had different responses related to treatments in two seasons (Table 2). In addition, all traits including root length, plant height, leafs, shoots and roots
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Table 2 Combined analysis of variance from the evaluation of Acacia victoriae traits over two seasons for 4 Zn concentrations Source of variation
df
Mean square Root length
Diameter
Plant height
Leaf area
Leaf dry matter
Shoot dry matter
Root dry matter
TI
Seasons
1
73.50**
19.54**
40.04**
0.001
4.95**
5.87**
0.55**
23.01
Error 1
4
2.50
0.02
1.17
0.05
0.07
0.02
0.01
10.17
Treatments
3
104.50**
1.65**
2.40**
1.12**
1.71**
944.18**
Treatment versus season
3
0.08ns
0.24ns
0.01ns
0.05ns
23.21
Error 2
12
C.V Source of variation
df
1.79**
160.26**
4.50ns
0.01ns
0.26ns
1.00
0.08
2.88
2.91
4.96
4.86
0.051 11.45
0.10
0.09
0.05
5.99
13.71
11.45
8.12
2.35
Root
S/R
Mean square Coll a
Coll b
Coll total
Error 1
4 0.002
0.035
0.192
4210.70
8255.05
37582.28
Treatments
3 0.06**
0.79**
3.88**
615013.01**
1404045.12**
17805572.17**
Treatment versus season
3 0.004ns
0.004ns
0.022ns
85685.54**
138134.15**
1333966.26**
1124.00
7209.87
37262.20
9.31
15.41
13.72
12
0.002 9.59
0.135
Stem
1 0.001
C.V
0.10
Leaf
Seasons
Error 2
ns
0.031
201041.98**
0.043
11.47
10.22
386732.86**
2693721.61**
L/R
L/S
BCF
0.238**
0.147**
0.001**
0.005
0.007
0.007
0.645**
0.359**
0.784**
388.93**
0.081**
0.035**
0.011**
25.05**
0.011 16.12
0.005 13.81
0.007 16.36
3.47* 0.73
0.67 8.04
* Significant at the 0.05 probability level, ** Significant at the 0.01 probability level, ns is non-significant respectively
dry matter and so on varied significantly between treatments at 1 and 5 % probability levels (Table 2). Growth and biomass responses of seedlings There was a significant different between the treatments in each season but the trend of effect of Zn concentrations on root length, plant height and dry matter of different tissues for both seasons were the same (Figs. 1, 2). The increase of Zn concentration from 50 to 250 mg L-1 significantly
a
increased the root length of seedlings (5 to 20 % higher than control seedlings) and TI for both seasons. In all treatments, TI ranged from 120 % the highest value (in seedlings exposed to 250 mg Zn L-1) to 85 % the lowest value (in seedlings exposed to 500 mg Zn L-1). Therefore, when Zn concentration in the soil medium was increased to 500 mg L-1, the TI declined significantly (Fig. 1). Plant height and dry biomass of different parts of seedlings increased significantly with increasing Zn concentration from 50 to 250 mg L-1 in the medium as compared to the
b
140
40
100 80
100
106.2
35
115.6 84.3
60 40 Summer
20 0
Root length (cm)
Tolerance index(%)
120
45
30 c
25
50
d
15 10
Fall 250
Zn Concentrations (mg L-1)
a
20
Summer
5 Control
b
500
Fall
0 Control
50 250 Zn Concentrations (mg L-1)
500
Fig. 1 Effect of Zn concentrations on the Zinc-tolerance index (TI) (a) and root length (b) of A. victoriae seedlings in two seasons for 45 days. (For any given Zn concentration during the same growing season, values followed by the same letter do not differ at p = 0.05)
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Differential expression of zinc accumulation during two growing seasons of Acacia victoriae
b
4.5 4 3.5 3 2.5 2 1.5 1 0.5 0
a
a
b
Stem dry matter (g)
Leaf dry matter (g)
a
c
Summer Control
Fall
50
250
4.5 4 3.5 3 2.5 2 1.5 1 0.5 0
500
b
Root dry matter (g)
4.5 4 3.5 3 2.5 2 1.5 1 0.5 0
b
b
a
ab
c
Summer
Control
Fall
50
250
500
Zn Concentrations (mg L-1)
Zn Concentrations (mg L-1)
c
667
a c
Summer
Control
50
Fall
250
500
Zn Concentrations (mg Zn L-1)
Fig. 2 Effect of Zn concentrations on the Leaf dry matter (a) Stem dry matter (b) and Root dry matter (c) of A. Victoria seedlings in two seasons for 45 days. (For any given Zn concentration during the same growing season, values followed by the same letter do not differ at p = 0.05)
controls without Zn (Table 3). The maximum dry biomass (10.86 g plant-1 for the fall season and 8.5 g plant-1 for the summer season) was recorded for seedlings exposed to 250 mg Zn L-1. In seedlings exposed to 50 and 250 mg Zn L-1, growth was stimulated, reaching total dry biomass between 13 and 30 % higher for fall and 8 to 16 % higher for summer than the controls (Table 3; Fig. 2). However, plant height, root length, leafs, shoots and roots dry matter treated with the highest concentration of 500 mg Zn L-1 declined significantly (Figs. 1, 2; Table 2). However, the seedlings exposed to a Zn concentration of 500 mg Zn L-1 did not show visible symptoms of Zn toxicity such as chlorotic spots or necrotic lesions at the leaf surface. But we recorded reductions in root length and
plant height (up to 25 % reduction as compared with controls), reduction of total chlorophyll (30 and 27 % reduction, respectively, as compared with controls for summer and fall), and reduction in growth and dry biomass (to 27 % reduction as compared with controls for the summer season) (Fig. 2; Table 3).
Discussion Chlorophyll In higher plants the contents of chlorophyll a and chlorophyll b are usually in the ratio 3:1 (chlorophyll
Table 3 Growth parameters of A.victoriae seedlings, grown in pots and exposed to four concentrations of Zn over two seasons for 45 days Treatments
Seasons
Root length (cm)
Diameter (mm)
Plant height (cm)
Leaf area (cm2 plant-1)
Leaf dry matter (g plant-1)
Shoots dry matter (g plant-1)
Roots dry matter (g plant-1)
TI
Zn
Summer
36a
5.09b
36.25a
1.98a
1.96b
2.17b
3.13a
102.86a
b
a
33.66
b
a
a
a
b
104.82a
Fall
32
6.90
1.99
Control 50 (mg Zn/L)
c
33 35b
b
6.03 6.18ab
c
33 36b
a
2.54 2.22b
250 (mg Zn/L)
39.5a
6.53a
41a
500 (mgZn/L)
d
c
d
29
5.24
28
2.86
3.16
2.82
2.26 2.74a
b
2.62 2.87ab
b
2.92 3.09b
100.00a 106.08b
1.86c
3.05a
3.08a
3.59a
119.66c
d
c
c
c
89.63d
1.32
b
1.60
2.08
2.30
Means with the same letter are not significantly different
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A. Mahdavi, K. Khermandar
a dominates) (Porra 2002). Changes in the ratio of chlorophyll a:b can reflect the negative influence of heavy metals to the photosynthetic apparatus of plants (Porra 2002). A typical phytotoxicity critical concentration is about 500 lg Zn g-1 (Reichman 2002). The results of chlorophyll pigment analysis showed that in treatments exposed to 250 mg Zn L-1, the Chlorophyll (a, b and total) contents were significantly higher than for the other treatments. The lowest chlorophyll (a, b and total) contents were found in 500 mg Zn L-1 treatments and reduced total chlorophyll to about 25 % of the control (or increased the ratio of chlorophyll a:b) in both studied growing seasons. A significant increase was seen in the chlorophyll a:b ratio for Zn alone, suggesting that the chlorophyll b pool is more sensitive to Zn exposure (Macfarlane and Burchett 2001). There was no significant difference in chlorophyll (a, b and total) contents between two seasons in all treatments (Fig. 3). Zn content in the tissues Metal uptake was dependent upon the metal concentrations in the soil medium. Significant positive correlations existed between Zn concentrations in the soil medium and Zn contents in root shoot and leaf tissues (Table 4). Zn accumulation in all treatments was significantly different between the two seasons (Table 2). A. victoriae accumulated large concentrations of Zn during the two growing seasons. Concentrations of Zn in leafs and shoots of A. victoriae seedlings in fall growing season were significantly larger than summer (in all treatments, Fig. 4a, b). While in all treatments, Zn concentrations in root tissues of A. victoriae seedlings in summer growing season were greater than fall season (Fig. 4c). In all treatments, Zn was mainly located in the roots and most Zn was transported to aboveground tissues in the shoots. The concentration gradients of Zn in different parts of seedlings ranked as follows: roots [ shoots [ leafs (Fig. 4). The highest Zn
Chlo b
3
Chlo a
2.5 2
b
b
Chlo total
b
a c
1.5 1 0.5 0 Control
50
250
Zn Concentrations (mg Zn L-1)
500
Chlorophyll pigments(mg g -1)
Chlorophyll pigments(mg g -1)
a
accumulations in leafs, shoots and roots of A. victoriae seedlings (755, 1141 and 3843 mg kg-1 dry matter, respectively) were found in the seedlings exposed to 500 mg Zn L-1 (Table 4). BCF values were between 9.63 and 19.60 for treatments at 500 and 50 mg Zn L-1, respectively. These high BCF values show the higher amount of metal accumulated in the roots than shoots and leafs. In all treatments an average of 42–67 % of the Zn accumulated in the entire plant tissues was retained in the roots. These results were associated with values (0.23 B TFs C 0.79) of the TFs from roots in all the treatments (Table 4). Toxic metals usually cause reduction of plant growth (Chaoui et al. 1997). This can be expressed as reduced growth rate, leaf area and root biomass, and can be followed by reduction of total biomass production (Begonia et al. 1998). The first symptom of Zn toxicity in most species is a general chlorosis of younger leafs (Fontes and Cox 1995). Plants exhibiting Zn toxicity have smaller leafs than control plants (Ren et al. 1993). The present study revealed that A. victoriae did not reduce growth or biomass, but actually increased growth and biomass when seedlings were treated with 50 and 250 mg Zn L-1. These results can be expressed because of A. victoriae tolerance to relatively high amounts of Zn. Kabata-Pendias (2011) determined the mean content of Zn in plants leafs of a series of plant species and divided this content of Zn into three groups: deficient content (10–20 mg Zn/kg DW), sufficient content (27–150 mg Zn/kg DW) and toxic content (100–400 mg Zn/kg DW). Of course, Zn is an essential nutrient for plants and the threshold of Zn toxicity varies among plant species, time of exposure to Zn stress and composition of the nutrient growth medium (Soares et al. 2001). Some studies reported low levels of heavy metals such as cadmium (Arduini et al. 2004), lead (Begonia et al. 1998; Kadukova et al. 2008; Mahdavi et al. 2014) and arsenic and cadmium (Fayiga et al. 2004) that had positive effects on plant growth and biomass
Summer
3 2.5
b
b
Fall
a
2 c 1.5 1 0.5 0 Control
50
250
500
Zn Concentrations (mg Zn L-1)
Fig. 3 Chlorophyll pigments (a, b and total) (mg/g) in A. victoria leaves (a) and comparison of total Chlorophyll between two seasons (b). (For any given Zn concentration during the same growing season, values followed by the same letter do not differ at p = 0.05)
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Differential expression of zinc accumulation during two growing seasons of Acacia victoriae Table 4 Zn contents (mg Zn Kg-1 dry biomass) in roots, shoots and leaves, Zn translocation factors(TFs, leaf/root Zn ratios(L/R), leaf/ shoot Zn ratios(L/S) and shoot/root Zn ratios (S/R), Bio-concentration
669
factor (BCF), of A.victoriae seedlings grown in pots and exposed to four concentrations of Zn over two seasons for 45 days
Treatments
Seasons
Leaf
Zn
Summer
268.44b
390.48b
1741.47a
0.30b
0.21b
0.53a
a
a
b
a
a
a
Fall
Stem
451.49
644.36
d
Root
1071.43
d
0
S/R
0.50
d
L/R
L/S
BCF
0.37
0.54
d
b
9.79b 10.56a
Control
0
0
0
0
0
0
50 (mg Zn/L) 250 (mg Zn/L)
240.37c 443.79b
324.51c 603.49b
415.5c 1366.4b
0.79a 0.48b
0.58a 0.38b
0.73a 0.76a
19.60a 11.47b
500 (mg Zn/L)
755.70a
1141.66a
3843.9a
0.34c
0.23c
0.65a
9.63c
1200 1000
b Summer
Fall
mg Zn Kg -1 stem dry biomass
a mg Zn Kg -1 leaf dry biomass
Means with the same letter are not significantly different
a
800 600
b
400
c
200 0
Control
50 250 Zn Concentrations (mg L-1)
500
1600 1400
Summer
1200
Fall a
1000
b
800 600
c
400 200 0 Control
50
250
500
c
6000
mg kg -1 root dry biomass
Zn Concentrations (mg L-1)
5000
a Summer
Fall
4000 3000 b
2000 c
1000 0 Control
50
250
Zn Concentrations (mg
500 L-1)
Fig. 4 Zn concentrations in Leaves (a), Shoots (b) and Roots (c) of A. victoria seedlings subject to different concentration of Zn treatments in two seasons
production. Although we recorded no visible symptoms of Zn toxicity in seedlings treated with 500 mg Zn L-1 we recorded reduced total dry biomass by 27 % in comparison with control seedlings. These results were compared with the results of Marchiol et al. (2004) who did not observed any obvious symptoms of metal toxicity with the mixture of Cd, Pb, Cr, Ni, Cu and Zn during treatment but recorded reduced growth and biomass production in Brassica napus and Raphanus sativus. Doncheva et al. (2001) reported plant growth inhibition extended in Eucalyptus maculate and E. urophylla by five weeks after addition of 400 lM ZnSO4.
Zn toxicity in roots is apparent as a reduction in the growth of the main root, fewer and shorter lateral roots and a yellowing of roots (Ren et al.1993). Our root length measurements (Fig. 1) showed that shorter root length was found in 500 (mg Zn L-1) treatments. This negative effect on root length was also accompanied by reduction in plant height, decrease in chlorophyll a, b and total as well as total dry biomass (Judith et al. 1977; Sagardoy et al. 2009) (Figs. 1, 2, 3; Table 3). The study showed that plant height was significantly greater after treatment with 250 mg Zn L-1 compared to controls, whereas in of 500 mg Zn L-1 treatment plant height was significantly less than in the control seedlings.
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The reduction in the plant height might have been mainly due to reduced root growth and regulation of less nutrient and water transport to the aerial parts of the plant (Shanker et al. 2005). Others showed the same results (Judith et al. 1977; Sagardoy et al. 2009). Concentrations of Zn differed between two seasons. The seedling response to soil Zn varied both in the nature and the amount in two seasons (Fig. 4a–c). Our study showed that because of the ability of A. victoriae to accumulate higher concentrations of Zn in roots in summer as compared to fall (Table 4), it has the potential to be used for Zn phytostabilisation (Rizzi et al. 2004). This study showed that, at 500 mg Zn L-1 treatments, Zn concentrations in the leafs and shoots of A. victoriae seedlings exceeded 1000 mg kg-1 in fall growing season (Table 4 and Fig. 4). In addition, the plant was also able to grow normally at Zn concentrations up to 500 mg L-1 and the symptoms of toxicity (chlorosis and necrosis on leafs and roots) were not revealed at this treatment. Therefore, the results indicate that although A. victoriae was not a Zn hyperaccumulator, it can be used for phytoextraction purposes and should be harvested at the end of the growing season (fall season) because the concentration of the metal in the above-ground tissues was maximal at this time.
Conclusions Heavy metal toxicity in soils is a significant global problem. Although at low concentrations Zn acts as micronutrient, it becomes toxic at high concentrations. In this study we described the pattern of metal (Zn) accumulation in A. victoriae over two growing seasons so that we can effectively manage Zn contaminated areas. The highest concentrations of Zn were found in roots. Therefore, A. victoriae could be considered as a root bioaccumulator species. However significant concentrations of Zn were found in both shoots and leaf, especially in fall. Finally, from the results of study, it can be concluded that due to the ability of A. victoriae to accumulate high concentrations of Zn in its tissues, produce relatively large biomass and maintain high tolerance to drought and saline soils, is a suitable and effective choice to be used as a tool for phytoremediation (phytostabilization or phytoextraction) for industrial sites in arid zones.
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