Water Air Soil Pollut (2009) 197:23–34
DOI 10.1007/s11270-008-9788-7
Metal Tolerance, Accumulation and Translocation in Poplar
and Willow Clones Treated with Cadmium in Hydroponics
Massimo Zacchini & Fabrizio Pietrini &
Giuseppe Scarascia Mugnozza & Valentina Iori &
Lucia Pietrosanti & Angelo Massacci
Received: 14 February 2008 / Accepted: 22 June 2008 / Published online: 9 July 2008
# Springer Science + Business Media B.V. 2008
Abstract To evaluate the phytoremediation capability of some poplar and willow clones a hydroponic
screening for cadmium tolerance, accumulation and
translocation was performed. Rooted cuttings were
exposed for 3 weeks to 50 μM cadmium sulphate
in a growth chamber and morpho-physiological
parameters and cadmium content distribution in
various parts of the plant were evaluated. Total leaf
area and root characteristics in clones and species
were affected by cadmium treatment in different
ways. Poplar clones showed a remarkable variability
whereas willow clones were observed to be more
homogeneous in cadmium accumulation and distribution. This behaviour was further confirmed by the
calculation of the bio-concentration factor (BCF) and
the translocation factor (Tf). Mean values of all the
clones of the two Salicaceae species showed that
M. Zacchini : F. Pietrini : V. Iori : L. Pietrosanti :
A. Massacci (*)
Institute of Agro-Environmental and Forest Biology,
The National Council of Research,
Via Salaria Km 29,300 Monterotondo Scalo,
00015 Rome, Italy
e-mail: [email protected]
G. Scarascia Mugnozza
DISAFRI, University of Tuscia,
Via San Camillo De Lellis,
01100 Viterbo, Italy
willows had a far greater ability to tolerate cadmium
than poplars, as indicated by the tolerance index (Ti),
calculated on the dry weight of roots and shoots of
plants. As far as the mean values of Tf was concerned,
the capacity of willows to translocate was double that
of poplars. On the contrary, the mean values of total
BCF in poplar clones was far higher with respect to
those in willows. The implications of these results in
the selection of Salicaceae clones for phytoremediation
purposes were discussed.
Keywords Bioconcentration factor . Cadmium .
Hydroponic culture . Phytoremediation . Poplar .
Translocation factor . Willow
1 Introduction
The enhanced level of pollutants in soil and water due
to industrialisation is one of the major environmental
problems on a global scale. In particular, cadmium is
considered one of the most widespread pollutant having
toxic effects on plants and animals. Cadmium enters the
environment from industrial processes, heating systems,
urban traffic, phosphate fertilizers and the mineralisation
of rocks (Rauser and Muwly 1995). Plants exposed to
toxic cadmium concentration undergo a stress condition, revealed by harmful symptoms such as chlorosis,
growth inhibition, a reduction in water and nutrient
uptake, alteration of enzyme activity and photosynthe-
24
sis impairment (Sanità di Toppi and Gabbrielli 1999;
Pietrini et al. 2003). To remove cadmium and other
pollutants from contaminated areas, unconventional
techniques involving biological processes have been
successfully applied. In particular, plants can be used
to remove heavy metals from soil and accumulate
them in the harvestable parts. This technology, called
phytoextraction (Kumar et al. 1995; Raskin et al. 1997;
Padmavathiamma and Li 2007), is less expensive and
less damaging to the environment than conventional
remediation systems that consist mainly in the excavation and incineration of soil (Cunningham and Ow
1996). Another advantage of this technology is the
production of biomass, which can eventually be used in
producing energy and other commodities. The efficiency of phytoextraction depends largely on the metal
bioavailability present in the contaminated matrix as
well as on several characteristics of the plant such as the
capability to hyperaccumulate essential and unessential
metals, a fast growth, a deep and extended root system
and the ability to translocate metals to the aerial parts.
Over the last few years, forest trees have been
studied to assess how potential they are in remediating heavy-metal contaminated sites (Rosselli et al.
2003; Pulford and Watson 2003; Unterbrunner et al.
2007). With regards to the phytoremediation strategy
some aspects of forest tree biology and cultivation,
i.e. the large biomass yield that can be used to
produce energy, an extended and deep root apparatus,
a low impact on trophic chains and the adaptability of
some tree species to grow in marginal soils, are all
very interesting. Metal uptake by trees is reported to
be small but, on a hectare scale, the removal of heavy
metals from soil could be more effective respect to
hyperaccumulating plants due to a greater yield of
biomass (Greger and Landberg, 1999; Fischerová
et al. 2006).
Several studies have focused their attention on the
potentiality of willows and poplars in phytoextraction
(Riddell-Black 1994; Punshon and Dickinson 1999;
Robinson et al. 2000; Pulford et al. 2002; Laureysens
et al. 2004a; Kuzovkina and Quigley 2005). In fact,
these Salicaceae are reported not only to be adaptable
to growing in severe soil conditions, characteristic of
contaminated areas, but also to be capable of accumulating heavy metals (Pulford and Watson 2003).
Cultural management of willows and poplars by means
of short rotation coppice cultures (SRC) is another
interesting aspect to be considered in phytoremediation
Water Air Soil Pollut (2009) 197:23–34
strategies (Ceulemans et al. 1992; Scarascia-Mugnozza
et al. 1997; Perttu 1999; Rockwood et al. 2004). In this
context, Dickinson and Pulford (2005) have reported
that willow SRC can be utilised as an efficient and
cost-effective method in removing cadmium contamination from agricultural soil. Moreover, heavy metal
accumulation in poplar and willow clones varied
significantly (Landberg and Greger 1996; Watson et
al. 1999; Laureysens et al. 2004b).
Most of the studies carried out on trees reported
that the heavy metal accumulation pattern shows a
predominant compartmentalisation in the roots and a
low translocation to the shoots. This is probably the
major constraint to overcome for a more efficient
utilization of these species to rid soils of metal
contamination. Then, it is necessary to screen forest
plant material to establish what plants are most adapt
to translocate the absorbed metal to the aerial parts,
especially to the stem tissues that are not renewable
like foliage and that can be harvested and utilised in
energy production.
Many authors have reported differences between
willow and poplar clones in the partitioning of heavy
metals within the tree organs (Mills et al. 2000;
Robinson et al. 2000, 2005; Lunáčková et al. 2003a;
Fischerová et al. 2006; Unterbrunner et al. 2007).
Nevertheless, few studies have compared the
responses of willow and poplar clones to the presence
of cadmium in a hydroponic system (Šottníková et al.
2003; Lunáčková et al. 2003b; Dos Santos Utmazian
et al. 2007). Hydroponic culture is a very useful tool
for selecting from a considerable number of individuals. In fact, it reduces not only the period of growth
and the length of time of treatment of the plants but
also the space required to carry our the experiment. In
addition variability due to the environmental factors is
also reduced. In general, data obtained by a hydroponic
screening need to be confirmed by field performance
trials, even if Watson et al. (2003) have pointed out that
results obtained in hydroponics and in field experiments are similar.
This study was aimed at evaluating the response of
cadmium tolerance, accumulation and translocation in
different poplar and willow clones in a hydroponic
culture. The characterisation of several Salicaceae
clones to assess their effectiveness in tolerating and
bio-concentrating cadmium is very important in
specifying the potential of these plant species to
phytoremediate cadmium-polluted soils.
Water Air Soil Pollut (2009) 197:23–34
2 Materials and Methods
25
Table 1 Populus and Salix species tested in the experiment
Clone
Species/hybrid
Origin
A4A
Luisa Avanzo
I-214
Lux
11-5
Poli
58-861
6K3
14P11
Nisqually
SS5
SP3
6-03
2-03
Quirani
Cretone
Populus×canadensis Mönch.
Populus×canadensis Mönch.
Populus×canadensis Mönch.
Populus deltoides Bartr.
Populus×generosa A. Henry
Populus nigra L.
Populus nigra L.
Populus alba L.
Populus alba L.
Populus trichocarpa Torr. & A.Gray
Salix alba L.
Salix alba L.
Salix alba L.
Salix alba L.
Salix sp. autochthonous clonea
Salix sp. autochthonous clonea
Italy
Italy
Italy
USA
USA
Italy
Italy
Italy
Italy
USA
Italy
Italy
Italy
Italy
Italy
Italy
2.1 Plant Material and Growth Conditions
Previously rooted stem cuttings (20-cm-long) taken
from poplar and willow clones (listed in Table 1) were
divided into two stocks to be treated in hydroponics
with 0 (control) or 50 μM of cadmium sulphate
(Sigma, St. Louis, USA) for 3 weeks. Particular
attention was paid to choose homogenous rooted
cuttings to introduce randomly in the experimental
treatment that consisted in pots filled with third-strength
Hoagland’s nutrient solution, pH 6.5 (mol l−1): 1.34×
10−3 Ca(NO3)2 4H2O, 1.74×10−4 NH4H2PO4, 2.01×
10−3 KNO3, 0.66×10−3 MgSO4 7H2O, 0.41×10−4
NaOH, 2.97×10−5 EDTA, 2.98×10−5 FeSO4 7H2O,
3.22×10−6 H3BO3, 0.67×10−6 MnCl2 4H2O, 1.04×
10−7 ZnSO4 7H2O, 0.7×10−7 CuSO4 5H2O, 0.46×
10−7 MoO3, 2.86×10−8 Co(NO3)2 6H2O (Arnon and
Hoagland 1940). Cuttings were grown in a controlled
climate chamber equipped with metal halide lamps
(Powerstar HQI-TS; Osram, Munich, Germany) providing a photon flux density of 300 μmol m−2 s−1 for
14 h at 25°C. During the 10 h dark period the
temperature was 20°C. The relative humidity was 70–
80%. The nutrient solutions were replaced entirely
twice a week to prevent depletion of metals and
nutrients and to expose plants to a constant metal
concentration. An aeration system based on pumps was
used to avoid lack of oxygen. Each treatment group
consisted of five cuttings from each clone. The
diameters of the cuttings ranged from 1.2 to 1.7 cm
and no significant differences among clones were
observed. At the end of the experimental period,
control and treated plants were harvested and washed
with 0.05 M calcium chloride for 30 min in slow
agitation without damaging the roots. After leaf and
root measurements, plants were separated into aerial
parts (leaves, secondary stems and original cutting) and
roots. The leaf area was measured using a leaf area
meter Li 3000 (Licor, Nebraska, USA) and then each
plant part was dried in an oven at 80°C until a constant
weight was reached.
2.2 Cadmium Determination
Metal concentration was measured by an atomic
absorption spectrophotometer (Perkin Elmer, Norwalk, CT, USA) on digested samples of aerial parts
a
Collected near sulphurous springs 30 Km N-E of Rome.
and roots. Dried material was milled to a fine powder
(Tecator Cemotec 1090. Sample Mill; Tecator, Hoganas,
Sweden), then accurately weighed and mineralised.
Mineralisation was performed by treating 250 mg of
powdered samples with 6 ml of concentrated HNO3 and
2.5 ml of H2SO4 and by heating (TMD20 Heater
System, Velp Scientifica, Milano, Italy) in a two step
procedure: 120°C for 25 min followed by 250°C for
15 min.
2.3 Bio-concentration Factor (BCF), Translocation
Factor (Tf) and Tolerance Index (Ti) Calculation
According to Zayed et al. (1998), the cadmium bioconcentration factor (BCF) of root system and aerial
part (stem+leaves and secondary stems) was calculated as follows:
BCF ¼
cadmium concentration in the harvested plant material mg kg1
1
cadmium concentration in the solution mg kg
The translocation factor (Tf) was calculated to
evaluate the capability of plant to accumulate the
metal, absorbed by roots, in the aerial part:
Tf ¼
cadmium concentration in the aerialparts mg kg1
100
cadmium concentration in the the roots mg kg1
26
Water Air Soil Pollut (2009) 197:23–34
The tolerance index (Ti) was calculated to measure
the ability of the plant to grow in the presence of a
given concentration of metal, according to Wilkins
(1978):
Ti ¼
Dry weight of the plants grown in cadmium solution
100
Dry weight of the plants grown in control solution
Dry weight of plant was referred to roots, secondary
stems and leaves.
2.4 Statistical Analysis
The data reported refer to a single typical experiment
with five replicates. Normally distributed data were
processed with a two-way analysis of variance
(ANOVA) using the SPSS software tool.
3 Results
Effects due to cadmium were detected in all parameters but not in the mean root number. Differences in
all parameters were found both in poplar or willow
clones. A significant interaction between poplar or
willow clones and cadmium treatment was found in all
the morpho-physiological parameters analysed but not
in the mean root number in willow clones (Table 2).
In Fig. 1 a comparison between representative
willow and poplar plants whether treated or not with
cadmium is reported. No chlorosis symptoms were
revealed in both plant species. Shoot and root growth
in willows was not particularly affected by cadmium
exposition while a significant reduction of the same
occurred in poplars.
Damage exerted by cadmium at leaf level is an
important aspect to evaluate in plants screened for
phytoremediation. In fact, an efficient photosynthetic
apparatus allows plants to maintain an effective
transpiration flux that drives metals from roots to
aerial parts. Total leaf area is a parameter sensitive to
cadmium presence in the growth medium. Figure 2
shows the total leaf area of poplar and willow clones
whether subjected or not to 21 days of cadmium
treatment in a hydroponic experiment. All poplar
clones revealed a dramatic reduction in total leaf area
caused by cadmium exposure. The heavy metal treatment affected especially the clones 11-5, I-214, L.
Avanzo and 14P11 that showed a greater reduction in
leaf area. A4A and Nisqually, that showed contrasting
behaviour under control condition, revealed less inhibition than the other clones under metal treatment. In
willows, 6-02 and 2-03 clones resulted particularly
affected by cadmium treatment, while the other clones
showed no reduction in total leaf area.
The root system plays a key role in the interaction
between contaminants and plant. In poplar and willow
clones the effect of cadmium on the root system was
analysed measuring some morphological parameters
such as the mean number of roots per plant, the mean
root length per plant and total root length. Cadmium
treatment reduced the mean number of roots per plant
only in five poplar clones while no effect was observed
in willow clones (data not shown). The mean root
length per plant (Fig. 3) was noticeably affected by
cadmium treatment. In poplars, eight clones out of ten
showed a significant reduction in length. The greatest
reduction resulted in clone 11-5. Clones Lux and I214 were not affected by metal treatment. In willows,
clones SS5 and Quirani showed no inhibition in mean
root length as a result of cadmium treatment. On the
contrary, the mean root length of the other clones was
affected more or less dramatically by such exposure.
The entire root system, expressed as total root length
(Fig. 4), was negatively influenced by metal treatment
in six out of ten poplar clones with a dramatic
reduction in clones 14P11, 6K3 and 58-861, and a
Table 2 Observed significance levels (P values) for effects of clone, of cadmium treatment and their interaction from ANOVA for
some morpho-physiological parameters in poplar and willow cuttings grown in hydroponic solution
Factor
Poplar clone
Cadmium treatment
Interaction
Willow clone
Cadmium treatment
Interaction
Total leaf area
Mean root length
Mean root number
Total root length
<0.001
<0.001
<0.001
0.009
0.431
<0.001
<0.001
<0.001
0.004
<0.001
<0.001
0.049
<0.001
0.205
0.036
<0.001
0.505
0.347
<0.001
<0.001
0.008
<0.001
0.018
0.045
Water Air Soil Pollut (2009) 197:23–34
27
Fig. 1 Morphological
aspect of willow (a) and
poplar (b) plants exposed to
50 μM cadmium sulphate
(T) compared to control (C)
consistent reduction in clones Luisa Avanzo, 11-5 and
Poli. Clones Nisqually, A4A, I-214 and Lux were
statistically not affected by cadmium treatment. Clone
Lux showed the longest root system under metal
treatment among the tested poplar clones. In willow
clones (Fig. 4), total root length was negatively
affected by metal treatment in Cretone, SP-3 and 203 while no effect was observed in SS5, Quirani and
6-03. SP-3 and 6-03 showed the longest root extension
among the willow clones grown in cadmium added
solution.
3000
Poplar clones
Willow clones
2500
2
Total leaf area (cm )
Fig. 2 Total leaf area (cm2)
measured at the end of the
experiment on poplar and
willow clones grown in the
presence of 0 (control, black
bars) and 50 μM (grey
bars) cadmium sulphate.
Values are the mean of five
replicates. Error bars indicate standard error
In Fig. 5 the concentration of cadmium detected in
roots and the aerial parts of poplar and willow
cuttings exposed for 3 weeks to 50 μM cadmium
sulphate is reported. Metal concentration in control
cuttings was below the threshold of detection. A
considerable concentration of cadmium was found in
the roots of each poplar clone, varying greatly one
from the other. Root accumulation represented approximately 95% of the total cadmium accumulated
by the whole plants. P. nigra (clones 58-861 and Poli)
showed the highest metal root concentration while
2000
1500
1000
500
0
-5
14 A4A anzo Lux 861
11 I-2
v
58
L.A
li
3
11
lly S5 irani tone P3 -03 -03
Po 6K 14P qua
6
S u
S
2
e
Q
Cr
Nis
28
60
Willow clones
Poplar clones
50
Mean root lenght (cm)
Fig. 3 Mean root length
(cm) measured at the end of
the experiment on poplar
and willow clones grown in
the presence of 0 (control,
black bars) and 50 μM
(grey bars) cadmium sulphate. Values are the mean
of five replicates. Error bars
indicate standard error
Water Air Soil Pollut (2009) 197:23–34
40
30
20
10
0
-5 14 A4A anzo
11 I-2
v
L.A
li
Po
1
i
3
3
ly
e
3
5
6K 14P1 qual SS uiran eton SP3 6-0 2-0
Q
Cr
Nis
To evaluate the capability of poplar and willow
clones to extract and accumulate cadmium in the
plant, the bio-concentration factor (BCF) was calculated. In Fig. 6, the BCF of poplar and willow clones,
referring to the root system and the aerial part of the
plant, is reported. In poplars, the root BCF represented approximately 97% of the whole plant BCF,
with the highest root BCF resulting in clones 58-861
and Poli and the lowest in 6K3. The aerial part BCF
indicated Poli and Lux as the poplar clones with the
greatest capability to accumulate cadmium in leaf and
stem tissues. In willows, the clone 6-03 resulted, from
root BCF, as the most efficient cadmium bioaccumulator among the screened willow clones. The
clones Lux and 6K3 the lowest. Accumulation of
cadmium in the aerial parts of the plant among poplar
clones varied noticeably. Clones Poli and Lux showed
the highest metal concentration while L.Avanzo, 58861 and 6K3 the lowest.
In willows (Fig. 5), root cadmium concentration
was very homogeneous among clones with the highest
content detected in clone 6-03. Root cadmium concentration on average was approximately 87% of the total
concentration measured in the whole plant. On the
contrary, in the aerial part of the plant cadmium
concentration varied much more with the highest metal
accumulation found in clones 6-03 and Quirani and the
lowest in 2-03.
1200
Poplar clones
Willow clones
1000
Total root lenght (cm)
Fig. 4 Total root length
(cm) measured at the end of
the experiment on poplar
and willow clones grown in
the presence of 0 μM (control, black bars) and 50 μM
(grey bars) cadmium sulphate. Values are the mean
of five replicates. Error bars
indicate standard error
x
1
Lu 8-86
5
800
600
400
200
0
-5
14 A4A anzo
11 I-2
v
L.A
i
3
e
y
i
x
3
1
1
5
Lu 8-86 Pol 6K3 4P1 quall SS uiran eton SP3 6-0 2-0
1
5
Q
Cr
Nis
Water Air Soil Pollut (2009) 197:23–34
20000
-1
Cadmium concentration (mg . Kg )
Fig. 5 Cadmium concentration (mg kg−1) in roots
(black bars) and aerial parts
(grey bars) of poplar and
willow clones grown in hydroponics for 3 weeks in
50 μM cadmium sulphate.
Values are the mean of five
replicates. Error bars indicate standard error
29
Poplar clones
Willow clones
15000
10000
5000
1000
800
600
400
200
0
o
-5 14 A4A anz Lux -861 Poli 6K3 4P11 ually S5 irani tone SP3 6-03 2-03
v
S Qu
11 I-2
e
1
q
58
A
.
Cr
L
Nis
BCF of the aerial part showed that clones Quirani and
6-03 were the most efficacious in bio-accumulating
cadmium in the above ground tissues while the clone
2-03 was the least efficient.
The capability of poplar and willow clones to
accumulate cadmium in the above ground tissue was
further confirmed by calculating the translocation
factor (Tf), that indicated the percentage of the
absorbed metal that reached the aerial part of the
plant respect to that present in the roots (Fig. 7). This
percentage varied greatly among poplar clones. The
highest Tf values were observed in clones Lux and
Poli while the lowest were found in 58-861. Calculation of the Tf in willow plants showed a similar
4 Discussion
The availability of selected plant material is a key
factor if phytoremediation is to be efficiently applied
to different types of contaminated substrate (soil,
water, sludge etc.).
Poplar and willow clones, due to their growth,
genetic and cultural characteristics, are potential candidates in the remediation of contaminated substrates. In
this work the behaviour of a significant number of
350
300
Poplar clones
Willow clones
250
Bio-concentration factor (BCF)
Fig. 6 Bio-concentration
factor (BCF) in roots (black
bars) and aerial parts (grey
bars) of poplar and willow
clones grown in hydroponics for 3 weeks in 50 μM
cadmium sulphate. Values
are the mean of five replicates. Error bars indicate
standard error
translocation capability among clones whereas in the
clone 2-03 this value was slightly lower.
200
150
100
50
15
10
5
0
A
4
zo ux 861 oli K3 P11 ally
-5
ni
5
ne P3 -03 -03
6 4
11 I-21 A4 van
L 8S
P
2
6
SS Quira reto
1
qu
5
C
L.A
Nis
30
Poplar clones
Willow clones
30
Translocation factor (Tf)
Fig. 7 Translocation factor
(Tf) calculated at the end of
the experiment on poplar
and willow clones grown in
the presence of 50 μM cadmium sulphate. Data angle
pffiffiffiffi
transformation (arcsin %)
was performed. Values are
the mean of five replicates.
Error bars indicate standard
error
Water Air Soil Pollut (2009) 197:23–34
20
10
0
4
A
zo ux 861 oli
-5
L 811 I-21 A4 van
P
A
5
.
L
clones with regards to cadmium tolerance, accumulation
and translocation to aerial parts were compared. These
fundamental aspects should form the criteria to be
followed in screening plants for selection in phytoremediation. Metal tolerance, and consequently the
protection of the integrity and functionality of the
primary physiological and metabolic processes (Pietrini
et al. 2003), is an essential pre-requisite for a plant to
be utilised in phytoremediation. Nevertheless, this
characteristic should be the result of a combination of
metal absorption and reduction of damaging effects,
and not be merely due to metal exclusion. Tolerance at
root level, that means the preservation of the selective
property of the cell membrane, represents the first step
in metal absorption and loading into the xylem vessels.
In the present work, the root system of poplar and
willow clones, analysed by following morphological
parameters such as mean root length per plant and total
root length, showed remarkable differences in tolerance
to cadmium (Figs. 3 and 4). Total root length did not
significantly result reduced in 40% of poplar clones
and in 50% of willow clones, and in the latter case
proved to be more tolerant than poplar clones to
cadmium at root level.
Absorbed metal can be loaded onto xylem vessels by
binding it to organic acids, thiol and amine compounds
(Kramer et al. 1996; Keltjens and van Beusichem 1998;
Rauser 1999), to be transported to the shoots. In leaves,
cadmium can represent a very toxic agent, since it can
destroy thylacoidal membranes and alter enzyme
activities, hindering photosynthesis (Becerril et al.
1
3
lly S5 rani one P3 -03 -03
6K 14P1 qua
6
S
S ui
2
et
s
i
Q
Cr
N
1988; Pietrini et al. 2005). In the present work, the
effect of cadmium on leaves of poplar and willow
clones was evaluated by measuring the total leaf area
(Fig. 2). Poplar clones showed, on average, a more
evident reduction of the leaf area than willow clones.
In fact, in willows the leaf area of only two clones out
of six was negatively affected by the presence of metal,
whereas in poplars, contrary to Pilipović et al. (2005),
all clones presented a remarkable reduction in this
parameter. A reduction in leaf area in poplar and
willow species following cadmium treatment was also
found by Lunáčková et al. (2003a). Differences
between the two species for what regards cadmium
tolerance was further confirmed by the analysis of the
tolerance index, Ti (Table 3). On the basis of the dry
biomass of the total plants, the Ti revealed that on
average willows tolerate cadmium much more than
poplars. According to the scheme proposed by Lux
et al. (2004), willow clones tested in this work can be
defined as highly tolerant (Ti>60) while poplar clones
as moderately tolerant (Ti between 35 and 60). With
respect to these data, a relatively higher Ti of root and
leaves in some willows and poplar clones was reported
by Dos Santos Utmazian et al. (2007) but in that case
the cadmium concentration was far less than that used
in our experiment. Moreover, a remarkable difference
in cadmium tolerance among willow clones was found
by Punshon and Dickinson (1999) and Kuzovkina
et al. (2004).
Cadmium accumulation differed greatly in poplar
clones while in willows it was more homogeneous
Water Air Soil Pollut (2009) 197:23–34
31
(Fig. 5). On average the amount of cadmium that
poplar clones were capable of accumulating was
double that of willow clones (Table 3) even if it was
almost entirely confined in roots. On the contrary,
willows showed a greater ability than poplars to
accumulate the metal in the aerial part. Data reported
in literature about cadmium concentration and allocation among organs are often contradictory and this is
mainly due to the different experimental conditions
adopted. In fact, metal concentration, type and length
of exposure (whether the metal solution is renewed
over time or not) and type of substrate can noticeably
modify metal availability for the plant. Moreover,
washing procedures at the end of the experiment can
affect the quantity of metal measured in roots. In the
present work, root washing was performed with
calcium chloride for a far shorter time with respect
to Dos Santos Utmazian et al. (2007) and this could
explain, together with the different metal concentration used, why the cadmium accumulation differed
from that found by those authors. Nevertheless, our
data were in accordance with those previously
reported by Robinson et al. (2000) and Lunáčková
et al. (2003b), in different experimental trials, showing a prominent cadmium accumulation in poplar
roots and a more evident capability in willows to
allocate metal in the aerial part. In this context, the
bioconcentration factor (BCF) can give further valuable information regarding the capability for these
Salicaceae to extract metal from a contaminated
matrix. Poplar and willow clones, tested in this work,
exhibited very different BCF values in root and aerial
part (Fig. 6) and, on average, poplar clones showed
double the capability to remove metal from the
solution with respect to willow clones (Table 3),
expressed as total BCF. A comparison with other
studies, as has already been stated above in the case
of cadmium accumulation, is difficult to perform due
to the entirely different conditions in substrate and
metal concentration used. However, the BCF calcu-
lated in the present work was consistently lower than
that reported in similar hydroponic studies by Ait Ali
et al. (2004) and Wang et al. (2008) even if they were
carried out on herbaceous plants with lower cadmium
concentrations.
The ability to accumulate metal in the aerial parts
with respect to roots can be better illustrated by
calculating the translocation factor (Tf). In this work
the Tf in willows was double that in poplars (Table 3).
That willows have a better capacity to translocate
cadmium (expressed as leaf:root ratio) with respect to
poplars in a hydroponic experiment was also reported
by Dos Santos Utmazian et al. (2007). It is worth
mentioning that the Tf in Salicaceae clones, measured
in our work, is lower than that found in herbaceous
plants cultivated in pots by Mattina et al. (2003) and
Marchiol et al. (2004) or in wetlands (Deng et al.
2004), though it is difficult to compare the results due
to the different experimental conditions. Evaluation of
the Ti, BCF and Tf in poplar and willow clones in the
present experiment confirms that Salicaceae plants
have a considerable potentiality to remove cadmium
from a contaminated medium that could be increased
by improving the translocation of the metal to shoots.
The comparison between poplars and willows
regarding cadmium accumulation and distribution
revealed that these two Salicaceae species could each
be used for different purposes in phytoremediation
strategies. On the one hand, in fact, poplars which
showed a remarkable ability to bio-concentrate cadmium in the root system, could be efficiently used in
the remediation of polluted water (rhizofiltration) or
contaminated sites to limit metal percolating to the
water layer (phytostabilisation). On the other hand,
willow clones proved to be potentially promising in
translocating and concentrating cadmium in the above
ground organs associated with a great metal tolerance.
Hence, this species could be very useful in removing
pollutants from the soil to the harvestable parts of
plants (phytoextraction) and, if cultured in SRC
Table 3 Comparison between poplar and willow for tolerance index (Ti), bio-concentration factor (BCF), translocation factor (Tf),
cadmium concentration (mg kg−1) in roots and aerial parts of plants
Plant species
Ti
BCF
Tf
Poplar
Willow
45 (±3.03)
73 (±3.19)
159 (±9.08)
80 (±2.92)
10 (±0.45)
23 (±0.68)
Cadmium in roots
Cadmium in aerial parts
9962 (±563)
4296 (±164)
293 (±19)
651 (±36)
Data (± E.S.) refer to mean value of cuttings
pffiffiffiffifrom all clones grown in hydroponics for 3 weeks in 50 μM cadmium sulphate. For Ti
and Tf, data angle transformation (arcsin %) was performed.
32
management, could yield also biomass for energy
production, thereby carrying out a double ecological
service.
The effective capability of these Salicaceae clones
to accumulate cadmium in the above ground organs is
currently under evaluation in an outdoor mesocosm
system covering the whole growing season. This
system is a cultural technique more similar to the
environmental conditions of an open-field than
hydroponics. Preliminary results showed only a slight
decrease in the BCF and Tf values in willow and
poplar plants cultivated in a mesocosm system with
respect to hydroponics (data not shown), confirming
their potentiality to extract cadmium from a contaminated matrix and accumulate it in the aerial part.
Metal translocation to the above ground organs is a
crucial biochemical process in an effective utilisation
of plants to remediate polluted sites. In fact, a more
efficient mobilisation of metals from root to the above
ground organs could reduce the damaging effects
exerted by these pollutants on root physiology and
biochemistry. This would improve the effectiveness of
plant metal uptake allowing metal removal from the
contaminated substrate over time. Therefore metal
tolerance, bioaccumulation and translocation capability must be considered together to evaluate species,
clones or individuals with interesting perspectives in
phytoremediation, in order to characterise the biochemical and molecular traits involved in these
processes. From results obtained in this trial poplar
clones showed contrasting behaviour in cadmium bioconcentration and allocation in plant parts. Two
clones of Populus nigra, Poli and 58-861, exhibited
particular and interesting characteristics to be better
elucidated. In fact, both clones showed the highest
level of metal accumulation among poplar clones.
However, Poli proved to have a remarkable ability
whereas 58-861 exhibited the lowest ability to bioconcentrate cadmium in the aerial part. Willow clones
were more homogeneous regarding cadmium tolerance and accumulation but no clear indications were
obtained since the most efficient cadmium accumulator clone, 6-03, was also the most affected by
cadmium exposure at leaf level. Interesting responses
were then obtained in the clone SS5 and in the
autochthonous clone Quirani. It is worth noting that
the Quirani clone was collected in a sulphurous area
and cadmium is a well known sulphur-philic metal.
Studies are still in progress to characterise poplar and
Water Air Soil Pollut (2009) 197:23–34
willow clones for what concerns the biochemical and
molecular processes involved in the accumulation and
translocation of cadmium to the above ground organs.
Acknowledgements This work was funded by MIUR (Ministry for Education, University and Research) under PRIN 2005
project no. 2005-072892. Authors also wish to thank Prof.
Paolo Sequi (CRA-RPS) for research collaboration within
PRAL research project and Antonio Barchetti (CRA-RPS) for
valuable technical assistance.
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