Rhizosphere characteristics of indigenously growing nickel

hyperaccumulator and excluder plants on serpentine soil

W.W. Wenzel
a,
*, M. Bunkowski
a,b
, M. Puschenreiter
a
, O. Horak
b
a
Institute of Soil Science, University of Agricultural Sciences ViennaBOKU, Gregor Mendel Str., 33, A-1180 Vienna, Austria
b
Austrian Research Center Seibersdorf, Division of Life Sciences, A-2444 Seibersdorf, Austria
Received 2 May 2002; accepted 16 August 2002
Capsule: Field study reinforces that root exudates may contribute to nickel hyperaccumulation in
Thlaspi goesingense Ha la csy.
Abstract
The role of rhizosphere processes in metal hyperaccumulation is largely unexplored and a matter of debate, related eld data are
virtually not available. We conducted a eld survey of rhizosphere characteristics beneath the Ni hyperaccumulator Thlaspi goe-
singense Ha la csy and the metal-excluder species Silene vulgaris L. and Rumex acetosella L. growing natively on the same serpentine
site. Relative to bulk soil and to the rhizosphere of the excluder species, we found signicantly increased DOC and Ni concentra-
tions in water extracts of T. goesingense rhizosphere, whereas exchangeable Ni was depleted due to excessive uptake of Ni. Che-
mical speciation analysis using the MINTEQA2 software package revealed that enhanced Ni solubility in Thlaspi rhizosphere is
driven by the formation of Ni-organic complexes. Moreover, ligand-induced dissolution of Ni-bearing minerals is likely to con-
tribute to enhanced Ni solubility. Increased Mg and Ca concentrations and pH in Thlaspi rhizosphere are consistent with ligand-
induced dissolution of orthosilicates such as forsterite (Mg
2
SiO
4
). Our eld data reinforce the hypothesis that exudation of organic
ligands may contribute to enhanced solubility and replenishment of metals in the rhizosphere of hyperaccumulating species.
#2003 Elsevier Science Ltd. All rights reserved.
Keywords: Rhizosphere; Hyperaccumulation; Nickel; Thlaspi goesingense; Serpentine soil; Metal excluder plants
1. Introduction
Plants have adopted dierent strategies to cope with
extremely large (toxic) metal concentrations in their
growth medium: (1) hyperaccumulation and (2) exclu-
sion (Baker, 1981). Metal hyperaccumulator plants
comprise species that accumulate (in mg kg
1
)>10 000
(Mn or Zn), >1000 (Cu, Co, Cr, Ni, Pb) or >50 (Cd) in
their shoots (Baker and Brooks, 1989; Wenzel and
Jockwer, 1999). These plants have attracted the interest
of plant and soil scientists because of their role in the
development of phytoremediation technologies for the
treatment of metal-polluted soils, sediments and water
resources (Wenzel et al., 1999; Lombi et al., 2000b).
Thlaspi goesingense Ha la csy, based on the analysis of
herbarium specimen, has been rst described as a
hyperaccumulator of Ni by Reeves and Brooks (1983)
and conrmed in hydroponic experiments (Kra mer et
al., 1997) and in a eld survey (Wenzel and Jockwer,
1999).
Some progress has been made towards an under-
standing of plant-internal processes associated with
metal hyperaccumulation (Kra mer et al., 1997; Brooks,
1998; Salt and Kra mer, 2000; Kra mer et al., 2000; Lasat
and Kochian, 2000; Lombi et al., 2000a, 2001), how-
ever, little is known about the role of rhizosphere pro-
cesses (Wenzel et al., 1999b; McGrath et al., 2001).
Bernal et al. (1994) found no dierence between rhizo-
sphere pH of Ni hyperaccumulator Alyssum murale
Waldstein and Kitaibel and radish (Raphanus sativus
L.). McGrath et al. (1997) found that pH in the rhizo-
sphere of the Zn hyperaccumulators Thlaspi ochroleu-
cum and T. caerulescens J&C Presl decreased only
slightly, suggesting that increased metal uptake was not
related to rhizosphere acidication. Luo et al. (2000)
found even a slight increase in rhizosphere pH of
Thlaspi caerulescens. Knight et al. (1997) suggested, that
the uptake of free Zn
2+
from a well buered soil solu-
0269-7491/03/$ - see front matter # 2003 Elsevier Science Ltd. All rights reserved.
PI I : S0269- 7491( 02) 00341- X
Environmental Pollution 123 (2003) 131138
www.elsevier.com/locate/envpol
* Corresponding author. Tel.: +43-1-47654-3119; fax: +43-1-
47654-3105.
E-mail address: wwenzel@edv1.boku.ac.at (W.W. Wenzel).
tion induces further dissociation of Zn
2+
into soil solu-
tion to maintain the initial free Zn
2+
concentration. In
addition, hyperaccumulator species may release root
exudates containing chelators with the potential to
enhance heavy metal uptake, translocation and resis-
tance. Among the compounds that have been proposed
to participating in Ni chelation in hyperaccumulation
are citrate (Lee et al., 1977a) and free histidine (Kra mer
et al., 1996). However, Salt et al. (2000) have shown in a
hydroponic experiment that the release of citrate and
histidine did not appear to be involved in Ni-hyper-
accumulation in T. goesingense. They suggested that
there may have been other Ni-chelating exudates that
could not be detected. Also for T. caerulescens no evi-
dence was found that root exudates are involved in
active mobilisation of metals (Zhao et al., 2001).
One approach to understand whether rhizosphere
processes contribute to the excessive metal uptake by
hyperaccumulator plants employs the assessment of
root exudation or of derived microbial metabolites in
rhizosphere soil solutions (Wenzel et al., 1999b). How-
ever, exudation rates and chemical composition of exu-
dates of metal hyperaccumulator species are virtually
unknown and dicult to collect from soil-grown plants.
It has been proposed that hyperaccumulating species
may enhance metal solubility in the rhizosphere via root
exudation as it is known for other plants (Knight et al.,
1997). It is still a matter of debate whether root exuda-
tion by hyperaccumulating and non-accumulating spe-
cies are dierent in terms of quality and exudation rate.
If yes, such dierences may signicantly contribute to
the excessive metal uptake by hyperaccumulators. The
few data available have been obtained in hydroponic
conditions (Salt et al., 2000; Zhao et al., 2001) and do
not account for plantsoil interactions and the potential
role of microorganisms in the rhizosphere. Moreover
the rhizosphere of metal hyperaccumulator and metal-
excluding plant species yet has not been studied in eld
conditions.
Therefore, the objective of this study was to investi-
gate rhizosphere characteristics of indigenously eld
growing Thlaspi goesingense and of the two excluder
species Silene vulgaris and Rumex acetosella growing on
the same serpentine site in eastern Austria.
2. Materials and methods
Soil and plant samples were collected at a serpentine
site near Redlschlag, eastern Austria (Fig. 1), previously
described by Kra mer et al. (1997) and Wenzel and
Jockwer (1999). The soil is low in NaHCO
3
-extractable
P (<3 mg kg
1
). Further soil characteristics are pre-
sented in Table 1. Fig. 2 displays the distribution of the
collected plant and soil samples in the eld plot.
Individual plant specimen were collected along with the
soil adhering to their roots. Operationally dened rhizo-
sphere soil was separated immediately after sampling by
gentle shaking of the roots, lled in polyethylene bags
and frozen using liquid nitrogen to minimize microbial
changes of organic compounds. Operationally dened
bulk soil, collected from non-vegetated spots between
the plants (Fig. 2) was subjected to the same proce-
dure. Roots and shoots were separated and trans-
ferred in polyethylene bags without freezing. Soil and
plant samples were immediately transported to the
laboratory in transportable coolers (about 4

C). As
the eld site and laboratory were located within 2 h
driving distance, samples were virtually not aected
by thawing.
In the laboratory, soil samples were stored in a freezer
(20

C) until further treatment. After thawing in a
refrigerator (4

C), the soil samples were homogenized
by mixing, passed through a 2-mm stainless steel mesh
Fig. 1. Photograph of the experimental plot in Redlschlag, east Austria.
Table 1
Soil characteristics and total (T) and labile (1 M NH
4
NO
3
-extactable; E) metal concentrations of the soil Redlschlag (adopted from Wenzel and
Jockwer, 1999)
pH CaCO
3
OC Cd Co Cr Cu Ni Pb Zn
(CaCl
2
) g kg
1
mg kg
1
T 3.70 172.2 1910 51.9 2580 16.3 61.1
6.55 19.4 13.3 E <0.13 <0.13 <0.05 <0.05 5.81 <0.25 <0.15
132 W.W. Wenzel et al. / Environmental Pollution 123 (2003) 131138
and split in two batches. Batch 1 was air-dried and a
subsample of 2 g was digested in aqua regia under reux
(Blum et al., 1996).
Batch 2 was used in eld-moist condition to extract a
labile heavy metal fraction using 1 M NH
4
NO
3
(2 h, 2.5
ml g
1
soil) extraction (Deutsches Institut fu r Nor-
mung, 1995), and water-soluble metals (2 h, 5 ml deio-
nised H
2
O g
1
soil) according to Blum et al. (1996).
Soluble organic acids and sugars in the soils were
extracted using an aceton water mixture (24 h, 200
rpm) according to Bachmann and Kinzel (1992). The
plant samples were oven-dried at 80

C and digested in
a mixture of nitric and perchloric acid (5:1 v/v).
Element concentrations in the soil extracts, soil and
plant digests were determined using inductively coupled
plasma emission spectrometry (ICPAES, Varian
Saturn Liberty II). Soluble organic acids were deter-
mined by HPLC (Dionex DX 500, Ionpac AS 11 col-
umn), sugars were measured using HPLC-PAD (Dionex
DX 500, Carbopac PA 100 column).
Assuming equilibrium, measured pH and total ion
concentrations of Ni, Ca, Mg, K, SO
4
, Cl, NO
3
and
DOC in the water extracts were used to calculate che-
mical speciation of Ni using the PRODEFA2/MIN-
TEQA2 software package (Allison, 1991). Measured
DOC concentrations in the water extract were con-
verted to molar site concentration (mol l
1
) of reactive
functional groups associated with dissolved organic
matter (DOM), assuming an average site density of 5
mmol mg
1
DOC (Wenzel et al., 1996).
3. Results
3.1. Plant characteristics
Shoot and root biomass of the investigated plants are
listed in Table 2. Mean shoot biomass of T. goesingense
exceeded that of S. vulgaris and R. acetosella, whereas
root biomass decreased in the order
S. vulgaris>T. goesingense>R. acetosella.
Fig. 3 shows metal concentrations in shoots and roots
of the investigated plant species. Average Ni concentra-
tions were 3180 mg kg
1
in shoots and 1610 mg kg
1
in
roots of T. goesingense. Nickel concentrations >1000
mg kg
1
in shoots and a shoot: root concentration ratio
>1 conrm Ni hyperaccumulation (Reeves and Brooks,
1983; Wenzel and Jockwer, 1999). No hyperaccumula-
tion was found for the elements Cr, Zn and Cd,
although Cr in the soil was present at large concentra-
tions (Table 1). However, Zn and Cd concentrations in
soil did not exceed background concentrations of non-
polluted soils. These ndings are consistent with those
reported by Wenzel and Jockwer (1999) for the same
site.
Nickel concentrations in shoots and roots of S. vul-
garis and R. acetosella were well below those of T. goe-
singense (Fig. 3), conrming that the former plants tend
to exclude this metal from uptake. Furthermore, the
shoot:root concentration ratio was <1, indicating lim-
ited Ni transport into shoots. The abundance (Figs. 1
and 2) and obvious health of both species growing on
the experimental site suggests that they tolerate large
metal concentrations in soil. Chromium and Cd con-
centrations in roots and shoots of both excluder species
were similar to T. goesingense, whereas Zn concentra-
tions were signicantly smaller (Fig. 3).
Macronutrient concentrations in the investigated plant
species are shown in Fig. 3. Magnesium concentrations
in shoots were not signicantly dierent among plant
species, whereas Mg concentrations in roots were sig-
nicantly smaller in T. goesingense compared to Silene
vulgaris. The Ca concentrations in roots and shoots
decreased in the order S. vulgaris>T. goesingense>R.
acetosella. Potassium concentrations in shoots decreased
in the order S. vulgaris>T. goesingense>R. acetosella; K
in roots decreased in the order
T. goesingense>R. acetosellaS. vulgaris.
Fig. 2. Schematic representation of the experimental plot, showing the
locations of individual plants and spots where root-free bulk soil was
collected.
W.W. Wenzel et al. / Environmental Pollution 123 (2003) 131138 133
3.2. Soil characteristics
Bulk soil and rhizosphere soil characteristics deter-
mined in the water extract are presented in Table 3.
Signicantly larger pH, EC and DOC were found in the
rhizosphere of T. goesingense compared to the bulk soil,
while for R. acetosella and S. vulgaris rhizosphere only
DOC was signicantly enhanced. The concentrations of
soluble Ni tended to be generally larger in rhizosphere
Fig. 3. Total concentrations of Ca, Cd, Cr, K, Mg, Ni and Zn in roots (black column) and shoots (white column) of the experimental plants (means
and SE, n=7). TG=Thlaspi goesingense, SV=Silene vulgaris, RA=Rumex acetosella.
Table 2
Shoot and root biomass [g DM plant
1
] of the investigated plants
(n=7) from the eld experiment
a
T. goesingense S. vulgaris R. acetosella
Shoots Roots Shoots Roots Shoots Roots
Mean 5.73 1.23 2.14 1.64 2.98 0.73
SE 1.50 0.25 0.46 0.44 1.09 0.29
a
SE, standard error.
Table 3
pH, EC, DOC and the concentration of macronutrients and heavy metals in the water extract of the eld study (P<0.05; ANOVA, n=7)
pH EC (mS cm
2
) DOC (mg L
1
) Ni Cr Zn Cd Mg Ca K NO
3
-
SO
4
2
Cl

Thlaspi goesingense 6.76 b 43.5 b 91.0 c 0.38 c 0.014 b 0.03 a 0.001 a 30.5 c 2.85 b 2.03 b 0.57 a 14.7 b 10.9 a
Silene vulgaris 6.39 a 30.6 a 69.2 b 0.24 b 0.009 a 0.04 a 0.001 a 21.8 b 1.72 a 0.58 a 2.43 a 11.8 ab 10.8 a
Rumex acetosella 6.39 a 28.9 a 54.7 b 0.19 ab 0.008 a 0.03 a 0.001 a 16.2 ab 1.35 a 1.20 a 2.70 a 9.73 a 10.8 a
Bulk soil 6.37 a 23.1 a 32.8 a 0.12 a 0.007 a 0.02 a 0.001 a 11.2 a 0.85 a 0.65 a 12.4 b 8.13 a 7.44 b
134 W.W. Wenzel et al. / Environmental Pollution 123 (2003) 131138
soil, but the dierence was only signicant for T. goe-
singense. Similar results were obtained for Cr, whereas
for Cd and Zn no dierences were found. Water soluble
Mg, Ca and K were signicantly increased in the rhizo-
sphere of T. goesingense. The increase of Mg, Ca and Kin
the rhizosphere of S. vulgaris and R. acetosella was less
pronounced, while K in the S. vulgaris rhizosphere was
slightly decreased. Nitrate concentrations in the bulk soil
were substantially larger compared to the rhizosphere of
all experimental plants, with the smallest concentrations
in T. goesingense rhizosphere. In turn, the largest sulfate
concentrations were found in T. goesingense rhizosphere
whereas less pronounced insignicant increases relative
to bulk soil were found for the other plants. Chloride
was signicantly lower in bulk soil compared to its
concentration in the rhizosphere of all experimental
plants, with no dierences among the plant species.
Table 4 shows the concentration of soluble sugars in
the rhizosphere compared to bulk soil. While glucose,
trehalose and ranose concentrations were signicantly
enhanced in the rhizosphere of all investigated plant
species, no signicant dierence to bulk soil was found
for fructose and sucrose. No signicant dierence was
observed among plant species. Organic acids were gen-
erally below the detection limit of 50 mg l
1
.
The concentrations of labile (1 M NH
4
NO
3
-extrac-
table) Ni, Cr, Zn and Cd are shown in Table 5. Com-
pared to the water-extractable fraction (Table 3), labile
Ni in the rhizosphere of all investigated plants was
smaller than in bulk soil, with the most pronounced
decrease in T. goesingense rhizosphere. In contrast to
Ni, labile Cr was signicantly increased in T. goe-
singense rhizosphere relative to bulk soil and the rhizo-
sphere of S. vulgaris and R. acetosella. Labile Zn in the
rhizosphere of T. goesingense and R. acetosella was sig-
nicantly smaller than in the rhizosphere of S. vulgaris
and bulk soil. Labile Cd was signicantly decreased in
the rhizosphere of all investigated plant species com-
pared to bulk soil.
4. Discussion
4.1. Role of macronutrients in alleviation of Ni toxicity
Large concentrations of Mg in all investigated plant
species are related to the fraction of dissolved Mg in the
serpentine soil (Table 3), corresponding to Mg satura-
tion of the exchange complex of >50% (Wenzel and
Jockwer, 1999). Similarly, Horak (1971) observed high
concentrations of Mg in members of Brassicaceae, Car-
yophyllaceae and Polygonaceae growing on serpentine
soil. Various Brassicaceae species accumulate Mg in
plant tissue even on non-serpentine soil (Rattenbo ck,
1978). Although K concentrations in soil were small and
Mg:K ratio in the water extract were large (3060), the
Mg:K ratio in plants was considerably smaller (1.53.2).
This is consistent with ndings that plants can maintain
normal K concentrations in tissues even on soils with
limited K supply (Lee, 1977b). Corresponding to the
small Ca concentrations in the water extract, accumu-
lation of this element in plant tissues was generally
small. The largest Ca concentrations were found in
Silene vulgaris. Crooke and Inkson (1955) observed
decreased Ni toxicity in oat when larger concentrations
of Ca, Mg, K and N were applied. Therefore, larger Ca
concentrations in S. vulgaris may be associated with
decreased Ni toxicity.
4.2. Rhizosphere processes involved in Ni hyperaccumu-
lation and tolerance
The data of our eld study indicated a signicant
increase of soluble Ni in the rhizosphere of the hyper-
accumulator T. goesingense compared to the bulk soil
(Table 3). Signicantly larger DOC concentrations in
the rhizosphere of T. goesingense indicate enhanced root
exudation. However, concentrations of individual
organic acids did not exceed the detection limit of 50 mg
l
1
. Relative to bulk soil, the concentrations of some
sugars were signicantly increased in T. goesingense
rhizosphere, but did not dier signicantly from those
measured in the rhizosphere of the excluder species
(Table 5).
Linear regression and correlation analysis using all
individual plantsoil pairs (n=27) of this study reveals
Ni
sol
0:00447 DOC0:0455 R
2
0:84

Table 4
Sugar concentrations in the water/acetone extract of the rhizosphere
of Thlaspi goesingense, Silene vulgaris and Rumex acetosella compared
to non-rooted bulk soil (P<0.05; ANOVA, n=7, values in mg kg
1
)
in the eld study
Glucose Fructose Sucrose Ranose Trehalose
T. goesingense 9.74 b 0.50 a 0.66 a 0.32 b 10.2 b
S. vulgaris 7.79 b 0.43 a 0.76 a 0.37 b 10.3 b
R. acetosella 9.02 b 0.40 a 0.39 a 0.30 b 10.6 b
Bulk soil 5.10 a 0.29 a 0.39 a 0.19 a 8.27 a
Table 5
Labile (1 M NH
4
NO
3
-extractable) Ni, Cr, Zn and Cr in the rhizo-
sphere of Thlaspi goesingense, Silene vulgaris and Rumex acetosella
compared to non-rooted bulk soil (P <0.05; ANOVA, n=7, values in
mg kg
1
) in the eld study
Ni Cr Zn Cd
T. goesingense 5.06 a 0.0048 b 0.03 a 0.0007 a
S. vulgaris 6.63 ab 0.0008 a 0.04 b 0.0006 a
R. acetosella 6.94 b 0.0004 a 0.03 a 0.0012 a
Bulk soil 7.72 b 0.0008 a 0.04 b 0.0072 a
W.W. Wenzel et al. / Environmental Pollution 123 (2003) 131138 135
with water extractable Ni (Ni
sol
) and DOC in mg l
1
.
Calculations using PRODEFA2/MINTEQ2A (Allison,
1991) indicate that this relation can be explained by the
formation of Ni-organic complexes, rendering Ni more
soluble (Fig. 4). At small DOC concentrations, repre-
sentative of bulk soil and Rumex acetosella rhizosphere,
free Ni and Ni-DOM complexes are present at almost
equal amounts. As DOCincreases, MINTEQA2 predicts
up to 6-fold enhanced Ni-DOM and 2-fold increased
Ni
2+
activities (Fig. 4). The increase in Ni
2+
appears to
be an artifact of the calculation procedure as DOC was
assumed to exhibit equal molar site density and com-
plexing power of functional groups in all samples irre-
spective of the plant species involved. However, the
nature of DOC in the rhizosphere is likely to dier
among plant species. Interestingly, virtually all
enhanced Ni
2+
activities in Fig. 4 are derived from
T. goesingense rhizosphere. Our data suggest that DOM
in the rhizosphere of this hyperaccumulator may form
stronger complexes with Ni than DOM derived from
bulk soil or from the rhizosphere of excluder S. vulgaris
or R. acetosella.
The eect of increased DOC concentrations in the
rhizosphere on metal mobilization involves several
mechanisms. In addition to the shift of equilibrium
towards more total dissolved Ni as revealed by chemical
speciation (Fig. 4), DOM can be sorbed onto surfaces of
Ni-bearing minerals, weakening metal-oxygen bonds.
The rate of mineral dissolution (R
L
) has been shown to
be proportional to the surface concentration C
L
S
of
organic ligands,
R
L
k
L
C
S
L
1
where k
L
represents a reaction constant. The rate-limiting
step is typically the detachment of the metal-organic com-
plex from the mineral surface (Furrer and Sticher, 1999).
Serpentine is partly made of orthosilicates of Mg that
may contain more than 10
3
M Ni. Orthosilicates such
as forsterite (Mg
2
SiO
4
) are rather unstable in terms of
weathering and dissolve according to (Furrer and Sti-
cher, 1999)
Mg
2
SiO
4
16 H
2
O $ 2 Mg H
2
O
2
6
H
4
SiO
0
4
4 OH

; 2
or, in more acidic conditions according to
Mg
2
SiO
4
4 H

$ 2 Mg H
2
O
2
6
H
4
SiO
0
4
: 3
Enhanced mineral dissolution in the rhizosphere of
T. goesingense is indicated by about 3-fold elevated
concentrations of Ca, Mg and K in water extracts of
T. goesingense rhizosphere relative to bulk soil (Table 3).
Increased pH in the rhizosphere of T. goesingense
(Table 3) is consistent with the release of hydroxyl ions
during mineral dissolution according to Eq. (3). Sig-
nicant correlations between water-extractable Ni and
Mg (Mg [mg l
1
]=63.9 Ni [mg l
1
]+5.07; r
2
=0.86,
n=27) or Ca (Ca [mg l
1
]=6.55 Ni [mg l
1
]+0.18;
r
2
=0.73, n=27) support that (ligand-induced) co-dis-
solution of Ni-bearing minerals appears to be involved
in Ni mobilization in the rhizosphere of T. goesingense
Fig. 4. Relationship between DOC and Ni speciation (calculated using PRODEFA2/MINTEQA2, Allison et al., 1991) in the experimental soils
(n=27), including bulk soils and rhizosphere of Thlaspi goesingense, Silene vulgaris and Rumex acetolsella. Ni-DOM denotes organic complexes of
Ni, Ni
2+
refers to free Ni in the water extracts.
136 W.W. Wenzel et al. / Environmental Pollution 123 (2003) 131138
and to a lesser extent, of the other two investigated
plant species. A similar tendency was found for K.
It has been proposed that root exudation does not
contribute to Ni hyperaccumulation in T. goesingense
(Salt et al., 2000) and T. caerulescens (Zhao et al., 2001),
however, these experiments were conducted in hydro-
ponic culture. Our results, obtained in eld conditions,
show a signicantly enhanced Ni solubility in the rhi-
zosphere of T. goesingense (Table 3) even though the
labile (1 M NH
4
NO
3
-extractable) Ni fraction was
depleted (Table 5) due to excessive Ni uptake. This
apparent contradiction can only be resolved if an addi-
tional mechanisms of Ni mobilization is assumed. Our
data indicate that enhanced DOC derived from root /
microbial exudation in the rhizosphere of T. goesingense
appears to control this mobilization process (Fig. 4).
Other evidence in literature also points to active metal
mobilization in the rhizosphere of metal hyper-
accumulating plants. McGrath et al. (1997) concluded
that mass ow cannot fully explain Zn hyperaccumula-
tion in T. caerulescens, indicating a specic mechanism
for further solubilization of Zn in the rhizosphere.
Interestingly, mobilization processes seem to be more
important on moderately contaminated soils: Brown et
al. (1994) showed that at higher soil Zn concentrations,
Zn accumulation in T. caerulescens shoots is a function
of Zn concentrations in the soil solution. Brown et al.
(1994) concluded that above a certain level of water-
extractable Zn (approximately 35 mg kg
1
) uptake by
T. caerluescens may change from an active to a passive
process. Zn concentration in T. arvense sharing its rhi-
zosphere with T. caerluescens is increased on non-con-
taminated soil, which indicates an active mobilisation
process by T. caerulescens (Whiting et al., 1997),
whereas on contaminated soil Zn concentration in
T. arvense is decreased, which is likely to be due to Zn
depletion by T. caerulescens (Whiting et al., 2001).
5. Conclusions
In contrast to results of hydroponic studies reported
in literature (Salt et al., 2000; Zhao et al., 2001), our
eld data reinforce that ligand-induced metal mobiliza-
tion in the rhizosphere of Thlaspi goesingense may con-
tribute to Ni hyperaccumulation by this species. Such
processes seem to be less eective in the rhizosphere of
excluder Silene vulgaris and Rumex acetosella growing
on the same site. This hypothesis is currently further
investigated in rhizobox experiments.
Acknowledgements
We acknowledge the nancial support from the Aus-
trian Science Foundation (FWF, P 13454) and a grant
(BOKU Priority Area Project 16) received from the
University of Agricultural Sciences ViennaBOKU.
References
Allison, J.D., Brown, D.S., Novo-Gradac, K.J., 1991. Mintequa2/
Prodefa2, a geochemical assessment model for environmental
systems. Version 3.0 users manual. EPA/600/391/021. USEPA,
Athens, GA.
Bachmann, G., Kinzel, H., 1992. Physiological and ecological aspects
of the interactions between plant roots and rhizosphere soil. Soil
Biol. Biochem. 24, 543552.
Baker, A.J.M., 1981. Accumulation and excludersstrategies in the
response of plants to heavy metals. J. Plant Nutr. 3, 643654.
Baker, A.J.M., Brooks, R.R., 1989. Terrestrial higher plants which
hyperaccumulate metallic elementsa review of their distribution,
ecology and phytochemistry. Biorecovery 1, 81126.
Bernal, M.P., McGrath, S.P., Miller, A.J., Baker, A.J.M., 1994.
Comparison of the chemical changes in the rhizosphere of the nickel
hyperaccumulator Alyssum murale with the non-accumulator
Raphanus sativus. Plant Soil 164, 251259.
Blum, W.E.H., Spiegel, H., Wenzel, W.W., 1996. Bodenzu-
standsinventur. Konzeption, Durchfu hrung und Bewertung. Uni-
versita t fu r Bodenkultur, Wien.
Brooks, R.R., 1998. Phytochemistry of hyperaccumulation. In:
Brooks, R.R. (Ed.), Plants that Hyperaccumulate Metals. CAB
International, Wallingford, UK, pp. 261287.
Brown, S.L., Chaney, R.L., Angle, J.S., Baker, A.J.M., 1994. Phyto-
remediation potential of Thlaspi caerulescens and bladder campion
for zinc- and cadmium-contaminated soil. J. Environ. Qual. 23,
11511157.
Crooke, W.M., Inkson, R.H.E., 1955. The relationship between nickel
toxicity and major nutrient supply. Plant Soil 6, 115.
Deutsches Institut fu r Normung, 1995. Soil quality extraction of trace
elements with ammonium nitrate solution. DIN 19730. Beuth Ver-
lag, Berlin, Germany.
Furrer, G., Sticher, H., 1999. Chemische Verwitterungsprozesse. In:
Blume, H.-P., Felix-Henningsen, P., Fischer, W.R., Frede, H.-G.,
Horn, R., Stahr, K. (Eds.), Handbuch der Bodenkunde, Ecomed,
Landsberg/Lech.
Horak, O., 1971. Vergleichende Untersuchungen zum Miner-
alstowechsel der Panze. Dissertation Universita t Wien, Verlag
Notring, Wien.
Knight, B., Zhao, F.J., McGrath, S.P., Shen, Z.G., 1997. Zinc and
cadmium uptake by the hyperaccumulator Thlaspi caerulescens in
contaminated soils and its eects on the concentration and chemical
speciation of metals in soil solution. Plant Soil 197, 7178.
Kra mer, U., Cotter-Howells, J.D., Charnock, J.M., Baker, A.J.M.,
Smith, J.A.C., 1996. Free histidin as a metal chelator in plants that
hyperaccumulate nickel. Nature 379, 635638.
Kra mer, U., Smith, R.D., Wenzel, W.W., Raskin, I., Salt, D.E., 1997.
The role of metal transport and tolerance in nickel hyperaccumula-
tion by Thlaspi goesingense Ha la csy. Plant Physiol. 115, 16411650.
Kra mer, U., Pickering, I.J., Prince, R.C., Raskin, I., Salt, D.E., 2000.
Subcellular localization and speciation of nickel in hyper-
accumulator and non-accumulator Thlaspi species. Plant Physiol.
122, 13431353.
Lasat, M.M., Kochian, L.V., 2000. Physiology of Zn hyperaccumula-
tion in Thlaspi caerulescens. In: Terry, N., Banuelos, G. (Eds.),
Phytoremediation of Contaminated Soil and Water. CRC Press
LLC, pp. 159169.
Lee, J., Reeves, R.D., Brooks, R.R., Jare , T., 1977a. Isolation and
identication of a citrato-complex of nickel from nickel-accumulat-
ing plants. Phytochemistry 16, 15031505.
W.W. Wenzel et al. / Environmental Pollution 123 (2003) 131138 137
Lee, J., Brooks, R.R., Reeves, R.D., Boswell, C.R., Jare , T., 1977b.
Plantsoil relationships in a New Caledonean serpentine ora. Plant
Soil 46, 675680.
Lombi, E., Zhao, F.J., Dunhan, S.J., McGrath, S.P., 2000a. Cadmium
accumulation in populations of Thlaspi caerulescens and Thlaspi
goesingense. New. Phytol. 145, 1120.
Lombi, E., Wenzel, W.W., Gobran, G.R., Adriano, D.C., 2000b.
Dependency of phytoavailability of metals on indigenous and
induced rhizosphere processes: a review. In: Gobran, G.R., Wenzel,
W.W., Lombi, E. (Eds.), Trace Elements in the Rhizosphere. CRC
Press LLC, Boca Raton, pp. 324.
Lombi, E., Zhao, F.J., McGrath, S.P., Young, S.D., Sacchi, G.A., 2001.
Physiological evidence for a high-anity cadmium transporter highly
expressed in a Thlaspi caerulescens ecotype. New Phytol. 149, 5360.
Luo, Y.M., Christie, P., Baker, A.J.M., 2000. Soil solution Zn and pH
dynamics in non-rhizosphere soil and in the rhizosphere of Thlaspi
caerulescens grown in a Zn/Cd-contaminated soil. Chemosphere 41,
161164.
McGrath, S.P., Shen, Z.G., Zhao, F.J., 1997. Heavy metal uptake and
chemical changes in the rhizosphere of Thlaspi caerulescens and Thlaspi
ochroleucum grown in contaminated soils. Plant Soil 188, 153159.
McGrath, S.P., Zhao, F.J., Lombi, E., 2001. Plant and rhizosphere
characteristics involved in phytoremediation of metal-contaminated
soils. Plant Soil 232, 207214.
Rattenbo ck, H., 1978. Chemisch-physiologische Charakterisierung der
Brassicaceae. Ein Beitrag zum Physiotypenkonzept. Dissertation,
Universita t Wien.
Reeves, R.D., Brooks, R.R., 1983. European species of Thlaspi L.
(Cruciferae) as indicators of nickel and zinc. J. Geochem. Explor.
18, 275283.
Salt, D.E., Kra mer, U., 2000. Mechanisms of metal hyperaccumula-
tion in plants. In: Raskin, I., Ensley, B.D. (Eds.), Phytoremediation
of Toxic Metals: Using Plants to Clean Up the Environment. John
Wiley & Sons, Inc, pp. 231246.
Salt, D.E., Kato, N., Kra mer, U., Smith, R.D., Raskin, I., 2000. The
role of root exudates in nickel hyperaccumulation and tolerance in
accumulator and nonaccumulator species of Thlaspi. In: Terry, N.,
Banuelos, G. (Eds.), Phytoremediation of Contaminated Soil and
Water. CRC Press LLC, pp. 189200.
Wenzel, W.W., Lombi, E., Adriano, D.C., 1999. Biogeochemical pro-
cesses in the rhizosphere: Role in phytoremediation of metal-pol-
luted soils. In: Prasad, N.M.V., Hagemeyer, J. (Eds.), Heavy Metal
Stress in PlantsFrom Molecules to Ecosystems. Springer Verlag,
Heidelberg, pp. 273303.
Wenzel, W.W., Jockwer, F., 1999. Accumulation of heavy metals in
plants grown on mineralised soils of the Austrian Alps. Environ.
Pollut. 104, 145155.
Wenzel, W.W., Blum, W.E.H., Brandstetter, A., Jockwer, F., Ko chl,
A., Oberfo rster, M., Oberla nder, H.E., Riedler, C., Roth, K., Vla-
deva, I., 1996. Eects of soil properties and cultivar variety on cad-
mium accumulation in wheat grain. Z. Panzenerna hr. Bodenk.
159, 609614.
Whiting, S.N., Leake, J.R., Baker, A.J.M., McGrath, S.P., 1997.
Changes in phytoavailability of zinc to plants sharing a rhizosphere
with the zinc hyperaccumulator Thlaspi caerulescens J & C Presl. In:
Iskander, I. et al. (Eds.), Proc. 4th International Conference on the
BiogeochemistryTrace Elements. , pp. 469470.
Whiting, S.N., Leake, J.R., McGrath, S.P., Baker, A.J.M., 2001.
Hyperaccumulation of Zn by Thlaspi caerulescens can ameliorate Zn
toxicity in the rhizosphere of cocropped Thlaspi arvense. Environ.
Sci. Technol. 35, 32373241.
Zhao, F.J., Hamon, R.E., McLaughlin, M.J., 2001. Root exudates of
the hyperaccumulator Thlaspi caerulescens do not enhance metal
mobilization. New Phytol 151, 613620.
138 W.W. Wenzel et al. / Environmental Pollution 123 (2003) 131138