Directory UMM :Data Elmu:jurnal:T:Tree Physiology:Vol15.1995:
Tree Physiology 15, 775--782
© 1995 Heron Publishing----Victoria, Canada
Growth and ABA responses of maple seedlings to aluminum
ANNICK BERTRAND,1 GILLES ROBITAILLE,1 ROBERT BOUTIN1 and PAUL NADEAU2
1
Natural Resources Canada, Canadian Forest Service-Québec Region, P.O. Box 3800, Sainte-Foy, Québec G1V 4C7, Canada
2
Agriculture and Agri-Food Canada Research Station, 2560 Hochelaga Blvd., Sainte-Foy, Québec G1V 2J3, Canada
Received November 17, 1994
Summary We assessed the impacts of low pH and 2.0 mM
aluminum (Al) on the growth of sugar maple (Acer saccharum
Marsh.) seedlings over a 13-week period. At Week 9, total leaf
area of Al-treated seedlings was reduced by 27%; however, by
Week 13, leaf area was similar for seedlings in all treatments.
None of the other growth parameters examined were negatively
affected by the treatments at either Week 9 or Week 13. The
ABA concentration in the xylem sap, which is an indicator of
tree stress in the field, was not affected by any of the treatments
and was highest during periods of high evaporative demand in
June and August. We conclude that the duration of exposure to
Al is critical when assessing a threshold concentration for Al
toxicity because plants can acclimate to an Al concentration
previously considered toxic. Although Al stress did not appear
to reduce the vigor of sugar maple seedlings directly, it could
facilitate an inciting factor such as winter frost to induce tree
decline.
Keywords: abscisic acid, Acer saccharum, acidic deposition,
soil acidity.
Introduction
Sugar maple stands (Acer saccharum Marsh.) have shown
symptoms of decline in eastern North America since the early
1980s (Lachance 1985), but the mechanisms underlying this
decline have not been precisely defined. Acidic deposition is
considered a contributing factor that weakens the trees (Foster
1989), and extreme climatic events are thought to be inciting
factors that trigger tree decline (Robitaille et al. 1995). Others
have postulated that the detrimental effects of acidic deposition on trees are caused by mobilization of aluminum (Al) in
soil solution (Ulrich et al. 1980).
Aluminum can affect plant growth by causing an imbalance
in plant mineral nutrition (Kelly et al. 1990). High Al concentrations in the growth medium usually reduce concentrations
of magnesium, calcium and often phosphorus in tree foliage
below those required for normal metabolism. Aluminum is
strongly adsorbed on cell walls of the root cortex and blocks
the normal uptake of calcium and magnesium required in the
aerial parts of the tree (Stienen and Bauch 1988). Aluminum
can also lower phosphorus availability to the shoot by enhancing its precipitation in the soil or on the plant root (Cumming
et al. 1985). In greenhouse studies, a high Al concentration in
the nutrient solution has detrimental effects on the growth and
mineral nutrition of many tree species (Thornton et al. 1986a,
1989, Kruger and Sucoff 1989).
Hormonal imbalances may also be involved in the Al effects
on sugar maple seedling growth. According to Chapin (1990),
the growth response to an inadequate nutrient supply cannot be
explained by simple changes in the nutrient status of the plant
but probably involves complex changes in hormonal balance,
particularly in abscisic acid (ABA). Leaf ABA concentration
increases in response to various mineral deficiencies (Goldbach et al. 1975, Haeder and Beringer 1981, Radin 1984). High
ABA concentrations are also a typical response to ion toxicity,
but the increased ABA concentrations are generally explained
by an indirect salinity effect induced by a water deficit
(Marschner 1986). Few studies report a direct effect of high ion
concentration, particularly Al, on the concentrations of growth
regulators (Edwards et al. 1976).
We tested the hypothesis that low pH and high Al concentration lower the vigor of sugar maple seedlings and are contributing factors to sugar maple stand decline. The effects of these
stresses on tissue mineral concentrations and various growth
parameters were measured in roots and shoots over a 13-week
period. We also measured the concentration of ABA in the
xylem sap in response to Al over time to determine whether it
could be used as an indicator of Al stress in sugar maple
seedlings.
Materials and methods
Plant material and stress treatments
On April 20, 1993, 2-year-old sugar maple seedlings (50--90
cm in height), produced from seeds of a Saint-Georges-deBeauce provenance (Québec, 46°10′ N, 70°37′ W), were
planted in 16-liter pots containing washed silica (silica 16,
Indusmin, Saint-Canut, Québec) and acclimated in a greenhouse at the Laurentian Forestry Centre, Sainte-Foy, Québec,
for 7 weeks (day/night temperature of 23/20 °C and a 16-h
photoperiod). The seedlings were fertilized weekly with halfstrength Hoagland solution (Hoagland and Arnon 1938). At the
beginning of June 1993, a shade cloth was installed in the
greenhouse 2 m above the trees. The five treatments (n = 9 per
treatment) were started on June 8, 1993, and consisted of
irrigations with: (1) complete nutrient solution at pH 5.0 (con-
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BERTRAND, ROBITAILLE, BOUTIN AND NADEAU
trol), (2) complete nutrient solution at pH 4.0 (pH 4.0), (3)
complete nutrient solution at pH 4.0 with the addition of 2.0
mM Al (pH 4.0 + Al), (4) complete nutrient solution at pH 3.5
(pH 3.5), and (5) complete nutrient solution at pH 3.5 with the
addition of 2.0 mM Al (pH 3.5 + Al). The composition of the
nutrient solution was similar to that described by Thornton et
al. (1986a): 0.25 mM Ca(NO3)2, 0.5 mM KNO3, 0.5 mM
MgSO4, 0.5 mM NH4NO3, 0.04 mM NH4H2PO4, 10 µM Fe
EDTA, 0.8 µM Zn, 9.0 µM Mn, 0.3 µM Cu, 46 µM B, and 0.05
µM Mo. Aluminum was added as AlCl3 for Treatments 3 and
5. The Cl− concentrations of all treatments were equalized with
NaCl. The pH of the solutions was adjusted with HCl. Irrigations with 0.7 l of each solution took place twice weekly during
the 13-week experiment from June 8 (Day 0) to September 8
(Day 91), 1993. Once a week, all pots were also irrigated by a
spray system with demineralized water. Four destructive harvests were made: June 8 (Day 0), July 7--8 (Week 4), August
10--11 (Week 9) and September 7--8 (Week 13).
Growth parameters
At each harvest, the total weight and height (root collar to
terminal bud) of each seedling were measured. We also determined the weight of stems, leaves, and large (diameter > 1 mm)
and fine (diameter < 1 mm) roots. The roots were thoroughly
washed with demineralized water to remove adhering particles. After blotting, the total volume of the root system was
measured by a water displacement technique described by
Burdett (1979). After fine root removal, the volume of roots
larger than 1 mm was measured, as previously described, and
the volume of fine roots (diameter < 1 mm) was calculated by
the difference between the total root volume and the large root
volume. The total leaf area of each seedling was measured with
an image analyzer (AGVision, Pullman, WA). Each type of
tissue was then dried for 72 h at 70 °C. The root/shoot ratio
was calculated on the basis of tissue dry weight. The root
relative growth rate (RGR) was calculated on the basis of root
volume and expressed as a percentage: ((final root volume −
initial root volume)/initial root volume) × 100.
Xylem sap extraction and ABA quantification
The terminal twig (approximately 12 cm long), including five
to ten leaves (or the lateral twig when the terminal twig was
missing), was removed and immediately placed in a pressure
chamber (PMS Instruments Co., Corvallis, OR) with the bottom covered with a damp paper towel. A gradual pressure was
exerted on the twig, and the pressure was then maintained
between 2.0 and 3.0 MPa for approximately 5 min for the
collection of 100 µl of xylem sap. The vials containing the sap
were immediately put on ice and stored at −80 °C within 2 h
of collection.
After a 30-min incubation with polyvinylpolypyrrolidone
(0.1 mg per 100 µl), xylem sap was assayed for free ABA
content by radioimmunoassay (RIA) as described by Bertrand
et al. (1994). Antibodies specific to free (+) cis-trans ABA
were prepared according to Weiler (1980). The antibody used
showed a linear response from 0.1 to 10.0 pmol under RIA
conditions. The antiserum was highly specific with a cross-reactivity of 53% for (±) cis-trans ABA, 32% for (±) racemic
ABA and 7% for methylated (±) ABA. The affinity constant for
the antiserum was 2.47 × 10 −9 l mol −1. All standards and
samples were assayed in triplicate. Samples were diluted 1/10
in phosphate buffered saline solution (Weiler 1980).
Extraction and analysis of the silica solution
At each harvest date, 1 day after a fertilization treatment, 600 g
of silica was centrifuged at 1000 g for 5 min to extract the silica
solution. The samples were then placed in a cold room (2 °C)
until they were prepared for chemical analysis. The samples
were filtered at 0.45 µm, and the conductivity and pH measured. Total monomeric Al (with chrome azurol S) was measured by colorimetry (Flow Injection Analysis, Tecator 5020,
Sweden) with a detection limit of 2.0 µmol l −1; total Ca and
Mg were measured by atomic absorption spectrophotometry.
Analysis of the mineral composition of plant tissues
The dried plant material was ground to pass through a 2-mm
mesh. Subsamples of 0.5 g of leaf and root material from four
replicates per treatment were analyzed according to the dry
ashing method (NCASI 1983). Total P content of samples was
determined by colorimetry (Colorimeter PC 800, Brinkman
Instruments, Westbury, NY). Total tissue content of Al, Ca, Mg
and Mn was analyzed by atomic absorption spectrophotometry
with nitrous oxide for Al, and flame emission spectrophotometry for K and Na. The detection limits for the ions (in
mg l −1) were: Ca, 0.1; Mn, 0.06; K, 0.05; Mg, 0.01; and Al, 1.1.
The accuracy of tissue analysis was confirmed by comparison
with the Quality Assurance Subgroup of the Federal-Provincial Research and Monitoring Coordinating Committee reference material. For each ion under study, the results from our
laboratory were less than ± 6% of the median results from 18
laboratories.
Statistical analysis
A total of 135 seedlings (9 blocks (replicates) × 5 treatments
3 sampling dates) were arranged in the greenhouse in a randomized complete block design. The data were analyzed by
the GLM procedure of SAS (SAS Institute Inc., Cary, NC)
after the confirmation of homogeneity of variances through
residual analysis. The assumption of variance homogeneity
was not satisfied for ABA, and a logarithmic transformation
was used to stabilize the variance. The initial root volume was
determined before treatments were initiated and the effect of
this covariate was accounted for in the statistical analysis of
‘‘final root volume’’ after verification of the homogeneity of
slopes. The differences between means for treatments and for
dates of data collection were compared by orthogonal contrasts
or the LSD test. When the effects of two or more treatments
were not significantly different for a given variable, we present
one curve representing the mean of the results obtained.
RESPONSE OF MAPLE SEEDLINGS TO ALUMINUM STRESS
Results
Silica solution composition
Before the treatments began, the pH of the solution extracted
from the silica growth medium was 6.9. At Week 4, the pH
values of the silica solutions were less for all treatments than
at Day 0, and the reductions were significantly greater in the
Al treatments than in the other treatments (P ≤ 0.05) (Figure 1A). The acidity of the silica solutions did not differ significantly between the two Al treatments (Treatments 3 and 5)
or between the two pH treatments (Treatments 2 and 4) (Figure 1A). The pH of the silica solutions for treatments without
Al (Treatments 1, 2 and 4) decreased sharply to 4.5--5.2 between Weeks 9 and 13. The conductivities of the silica solu-
Figure 1. (A) pH, (B) conductivity of the silica solution, and (C) soil
water content (SWC) at Weeks 4, 9 and 13 (n = 9). Symbols: d,
control; n, pH 4.0; h, pH 4.0 + Al; m, pH 3.5; j, pH 3.5 + Al. Each
curve represents the mean for nonsignificantly different treatments.
777
tions were similar for all treatments (Figure 1B). The low
conductivity of the silica solution on Day 0 corresponded to
the nutrient solution used before the treatments began, i.e.,
before the addition of AlCl3 or NaCl. The highest conductivity
(Figure 1B) was reached at Week 9 and corresponded with a
period of high evaporative demand in the greenhouse when soil
water content was at a minimum (Figure 1C).
In the pH 4.0 + Al and pH 3.5 + Al treatments, the Al
concentration of the silica solutions increased from 1.25 to
2.30 mM between Weeks 4 and 9, and then stabilized (Figure 2A). The Al/Mg ratio followed the same trend and stabilized at approximately 3.4 (Figure 2B). The Al/Ca molar ratio
doubled from 2 to 4 between Weeks 4 and 9, and then increased
more gradually to reach about 5 by Week 13 (Figure 2C). There
were no significant differences between the two Al treatments
for these variables. Throughout the study, the Mg2+ and Ca2+
concentrations in the silica solution remained higher in the Al
treatments than in the other treatments (Figures 3A and 3B).
Figure 2. (A) Al concentration, (B) Al/Mg molar ratio, and (C) Al/Ca
molar ratio of the silica solution at Weeks 4, 9 and 13 (mean ± SE,
n = 9). Symbols: j, 2.0 mM Al at pH 4.0; h, 2.0 mM Al at pH 3.5.
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BERTRAND, ROBITAILLE, BOUTIN AND NADEAU
Seedling growth and ABA concentration
Tissue mineral concentration
The 2.0 mM Al treatment stimulated root growth. By Week 13,
the root volume of Al-treated seedlings was approximately 75
cm3 compared with a mean of ≈65 cm3 for seedlings in the
other treatments (P ≤ 0.05) (Figure 4A). In response to the
presence of Al, the relative root growth rate ranged from 17 to
22% compared with 3% in the control seedlings (Figure 4B).
The Al treatment significantly (P ≤ 0.05) enhanced the
root/shoot ratio compared with the other treatments (mean
value = 0.95 versus 0.81) (Figure 4C).
Leaf area was the only parameter negatively affected by the
Al treatments (P ≤ 0.05). At Week 9, leaf area had decreased
by about 27% in Al-treated seedlings compared with seedlings
in the other treatments (Figure 5A); however, by Week 13, the
total leaf area of seedlings in all treatments was about 1000
cm2. Between Weeks 4 and 13, the fine root volume/large root
volume ratio decreased progressively and similarly for seedlings in all treatments (Figure 5B).
The ABA concentration in the xylem sap was highest (403
pmol ml −1) at the beginning of the experiment when the soil
water content was low. It decreased sharply to reach 144 pmol
ml −1 at Week 4, and then increased to 210 pmol ml −1 between
Weeks 4 and 9 as the soil dried (cf. Figures 1C and 5C). None
of the treatments had a significant effect on xylem sap ABA
concentration.
The mean Al concentration of roots of Al-treated seedlings
increased from Week 4 to Week 13 reaching a mean of 1.1 mg
gDW−1, which was three times the concentration found in roots
of control and pH-treated seedlings (Figure 6A). In leaves, a
maximum Al concentration of ≈0.6 mg gDW−1 was found in
Al-treated seedlings at Week 9 (Figure 6B). Concentrations of
Ca, P and Mn in roots and leaves were not affected by any of
the treatments (Table 1). The Mg concentration was significantly lower in roots of Al-treated seedlings (P ≤ 0.05) than in
roots of seedlings in the other treatments, whereas leaf Mg
concentration was not affected by any of the treatments (6.7-7.0 mg gDW−1) (Table 1). Root K concentration was slightly
higher (P ≤ 0.05) in Al-treated seedlings than in seedlings in
the other treatments (≈4.5 versus ≈3.5 mg gDW−1), whereas
foliar K concentration was lowest in Al-treated seedlings (≈4.0
Figure 3. (A) Mg, and (B) Ca concentrations in the silica solution at
Weeks 4, 9 and 13 (n = 9). Symbols: d, control; n, pH 4.0; h, pH 4.0
+ Al; m, pH 3.5; j, pH 3.5 + Al. Each curve represents the mean for
nonsignificantly different treatments.
Figure 4. (A) Final root volume, (B) relative root growth rate (volumetric RGR), and (C) root/shoot ratio in response to five treatments.
Treatments with the same letter are not significantly different at P =
0.05; mean ± SE for n = 27.
RESPONSE OF MAPLE SEEDLINGS TO ALUMINUM STRESS
779
Figure 6. (A) Root Al concentration, (B) leaf Al concentration, and (C)
soil water content (SWC) at Weeks 4, 9 and 13 (n = 9). Symbols: d,
control; n, pH 4.0; h, pH 4.0 + Al; m, pH 3.5; j, pH 3.5 + Al. Each
curve represents the mean for nonsignificantly different treatments.
Figure 5. (A) Total leaf area per seedling, (B) fine/large root ratio, and
(C) xylem sap ABA concentration at Weeks 4, 9 and 13 (n = 9).
Symbols: d, control; n, pH 4.0; h, pH 4.0 + Al; m, pH 3.5; j, pH
3.5 + Al. Each curve represents the mean for nonsignificantly different
treatments.
mg gDW−1) (Table 1). Root Na concentration in Al-treated
seedlings (≈ 0.7 mg gDW−1) was approximately half the concentration found in the roots of seedlings in the other treatments.
Foliar Na concentrations were lowest in Al-treated seedlings
(0.4 mg gDW−1) and highest in pH-treated seedlings (1.6 mg
gDW−1) (Table 1).
Discussion
The negative effects of Al were more pronounced in the aerial
part (shoots and leaves) than in the roots of sugar maple
seedlings. The finding that the aerial part was more sensitive
to Al than the roots may be a common feature of deciduous tree
species (Thornton et al. 1986a, Kelly et al. 1990).
In our study, the Al concentration of the roots was much
lower than the concentration reported by Thornton et al.
(1986a) in response to a similar Al treatment (2.0 mM) and
probably accounts for the absence of toxic Al effects on the
root system. Reductions in root growth rate, root branching, or
both that are generally associated with Al stress are usually
explained by a decrease in Ca2+ or Mg2+ uptake by the root
system in the presence of Al3+, resulting in a nutritional deficiency of these elements (Edwards et al. 1976, Thornton et al.
1987). The concentrations of Ca2+ and Mg2+ measured in the
silica solution of Al-treated seedlings were higher than in the
silica solution of control seedlings, which indicates that Al
effectively blocked the entry of these ions into the root. We
conclude that, under our experimental conditions, the blocking
effect did not cause Ca or Mg deficiencies, because these
elements were supplied in sufficient amounts in the nutrient
medium (cf. Kelly et al. 1990). The absence of an Al effect on
Ca concentration in leaves and roots differs from the observations made on honeylocust, peaches and coniferous trees in
which the tissue concentrations of Ca were depleted in the
presence of Al (Edwards et al. 1976, Sucoff et al. 1990, Raynal
et al. 1990). Kelly et al. (1990) concluded that the molar ratio
of Al/Ca of the nutrient solution determines the interaction
between the two elements. The Al/Ca ratio of our silica solution was about 2 at Week 4 and increased to about 5 at Week
13. However, Kelly et al. (1990) observed that a wide range of
Al/Ca ratios was ineffective in modifying the growth of red oak
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BERTRAND, ROBITAILLE, BOUTIN AND NADEAU
Table 1. Mineral concentration of sugar maple roots and leaves (mg gDW−1) in response to five treatments for the pooled sampling dates. Different
letters indicate significant differences among treatments at P ≤ 0.05; mean (± SE) for n = 27.
Element
Control
pH 4.0
pH 3.5
pH 4.0 + Al
pH 3.5 + Al
Roots
Ca
Mg
K
P
Mn
Na
5.68 (0.47)
1.60 (0.09) a
3.07 (0.20) b
1.30 (0.07)
0.09 (0.01)
1.41 (0.12) a
5.91 (0.42)
1.84 (0.09) a
3.94 (0.22) b
1.62 (0.12)
0.14 (0.02)
1.34 (0.10) a
5.81 (0.61)
1.79 (0.10) a
3.68 (0.34) b
1.56 (0.13)
0.12 (0.02)
1.37 (0.11) a
4.75 (0.40)
1.45 (0.06) b
4.47 (0.30) a
1.22 (0.10)
0.09 (0.006)
0.78 (0.08) b
5.12 (0.34)
1.56 (0.10) b
4.66 (0.21) a
1.36 (0.14)
0.11 (0.006)
0.90 (0.10) b
Leaves
Ca
Mg
K
P
Mn
Na
18.84 (1.16)
6.71 (0.34)
4.86 (0.60) ab
1.69 (0.11)
0.21 (0.04)
0.85 (0.20) b
22.07 (1.63)
6.92 (0.31)
6.39 (0.70) a
2.07 (0.18)
0.19 (0.03)
1.85 (0.39) a
21.41 (2.01)
6.90 (0.30)
5.53 (.55) a
1.97 (0.16)
0.28 (0.05)
1.18 (0.36) a
21.69 (1.94)
7.03 (0.36)
4.06 (0.28) b
1.72 (0.20)
0.28 (0.05)
0.40 (0.06) c
21.30 (1.22)
6.89 (0.37)
4.20 (0.31) b
1.81 (0.15)
0.28 (0.06)
0.40 (0.04) c
and suggested that the observed differences in Al sensitivity
exhibited by tree species were associated with experimental
conditions resulting in differences in either Ca2+ availability or
ionic strength of the solution. Our results support the suggestion that Al sensitivity can be offset by the presence of suitable
amounts of plant-available Ca.
We observed a stimulation of relative root growth rate by 2.0
mM Al3+. Thornton et al. (1986a) reported a similar observation at 1.0 mM Al3+; however at 2.0 mm Al3+, these workers
reported detrimental effects of Al3+ on roots growing in solution culture, suggesting that the results obtained from plants in
solution culture are not directly comparable with those obtained from plants in silica culture. Although a steady-state
nutrient supply, as is possible in solution culture, cannot be
achieved when silica is used as the growing medium, silica
culture provides information about plant responses to stress
such as exclusion mechanisms and root exudates (Taylor
1991). It has been shown for Glycine max (L.) Merill. that the
mechanical impedance caused by sand culture increased tolerance to Al stress as a result of reduced Al uptake into the root
tips compared with uptake from solution culture (Horst et al.
1990). These authors suggest that enhanced release of exudates
might account for the higher Al tolerance in sand culture than
in solution culture. Plants could exclude Al from the symplasm
if chelate ligands were released into the rhizosphere and these
ligands formed stable complexes with Al, thus reducing the
absorption of the metal ion across the plasma membrane (Taylor 1991). In addition to root exudates, there is the possibility
that genotype may also strongly influence responses.
The Al treatments caused decreases in leaf area between
Weeks 4 and 9 so that at Week 9, total leaf area per tree was
reduced by more than 27% in Al-treated seedlings relative to
seedlings in the other treatments. By Week 13, the total leaf
area was similar for all treatments even though the leaf Al
concentration was twice as high in Al-treated seedlings as in
untreated seedlings. Thornton et al. (1986b) observed an absolute, but small (≤ 10% over a 30-day period), increase in
damage with increasing exposure time to Al. Our results show
that plants can acclimate to a concentration of Al previously
considered toxic and indicate that the duration of exposure to
Al should therefore be taken into account when assessing a
critical concentration for Al toxicity in silica culture. Possible
mechanisms of acclimation include entrapment of excess Al in
roots, reducing Al concentration in seedling tops (Foy et al.
1978) as observed at Week 13, and an enzyme reaction to Al
stress in the leaves (Bowler et al. 1992).
Leaf K concentration decreased in response to the Al treatments. Translocation of K from roots to leaves may have been
impaired in the presence of Al because the root K concentration was significantly higher in Al-treated seedlings than in
control seedlings. In our study, the leaf K concentration (≈4.0
mg gDW−1) of all the seedlings was below the critical concentration of 6.0 mg g−1 needed for adequate nutrition of A. saccharum (Bernier and Brazeau 1988) and lower than the
concentration reported for mature sugar maple trees (Bernier
et al. 1989) or for sugar maple seedlings growing in solution
culture (Thornton et al. 1986a). We note that deficiencies in
leaf K in sugar maple trees growing in the Beauce region
(Québec, Canada), which is the provenance of our seedlings,
have been associated with incidences of stand decline in this
region (Ouimet and Fortin 1992). The numerous differences in
nutrition requirements among varieties or lines of plants suggest genetic control of inorganic plant nutrition (Gerloff and
Gabelman 1983). Moreover, a large range of variation in sensitivity to Al has been observed among various provenances of
paper birch (Steiner et al. 1980), among Populus hybrids (Steiner et al. 1984), and among genotypes of loblolly pine (Raynal
et al. 1990). The differential intraspecific sensitivity to toxic
ions also reflects genetic differences in mineral nutrition,
which could prove useful in breeding trees for tolerance to
nutritional stress.
The treatments had no effect on ABA concentration in the
xylem sap. Maximum ABA concentrations in the xylem sap
were reached at the onset of the experiment and at Week 9,
RESPONSE OF MAPLE SEEDLINGS TO ALUMINUM STRESS
which corresponded to low soil water contents. A direct increase in ABA concentration of the xylem sap in response to
soil water depletion has been observed in maize (Zhang and
Davies 1990). At Week 9, when the soil water content was at
its lowest value, the ABA concentration in the xylem sap was
comparable to the concentration measured in mature sugar
maple trees during a physiological drought stress caused by the
freezing of soil water, whereas at Weeks 4 and 13, the ABA
concentration in the xylem sap was similar to that in untreated
trees with an adequate water supply (Bertrand et al. 1994).
Based on this comparison of field and laboratory results, we
conclude that water stress was the principal cause of increases
in ABA concentration in the xylem sap and that Al had little or
no effect on ABA concentration of the xylem sap.
Aluminum stress or other nutritional imbalances may facilitate an inciting factor such as early or late winter frost to induce
sugar maple tree decline. The impacts of high concentrations
of Al and acid rain acting alone and in combination on the cold
tolerance of sugar maple seedlings are under investigation.
Acknowledgments
We thank Mr. Pascal Aubé and Mr. Clément Aubin for their technical
assistance, Mr. Alain LePage for his contribution to the chemical
analysis, and Ms. Michèle Bernier-Cardou for discussions on statistical analysis. This work was supported by the Long Range Transport of
Atmospheric Pollutants (LRTAP).
References
Bernier, B. and M. Brazeau. 1988. Foliar nutrient status in relation to
sugar maple dieback and decline in the Québec Appalachians. Can.
J. For. Res. 18:754--761.
Bernier, B., D. Par and M. Brazeau. 1989. Natural stresses, nutrient
imbalances and forest decline in southeastern Québec. Water Air
Soil Pollut. 48:239--250.
Bertrand, A., G. Robitaille, P. Nadeau and R. Boutin. 1994. Effects of
soil freezing and drought stress on abscisic acid content of sugar
maple sap and leaves. Tree Physiol. 14:413--425.
Bowler, C., M. Van Montagu and D. Inz. 1992. Superoxide dismutase
and stress tolerance. Annu. Rev. Plant Physiol. Plant Mol. Biol.
43:83--116.
Burdett, A.N. 1979. A nondestructive method for measuring the volume of intact plant parts. Can. J. For. Res. 9:120--122.
Chapin III, F.S. 1990. Effects of nutrient deficiency on plant growth:
evidence for a centralized stress-response system. In Importance of
Root to Shoot Communication in the Response to Environmental
Stress. Eds. W.J. Davies and B. Jeffcoat. British Society for Plant
Growth Regulation, Lancaster, pp 135--148.
Cumming, J.R., R.T. Eckert and L.S. Evans. 1985. Effect of aluminum
on 32P uptake and translocation by red spruce seedlings. Can. J. For.
Res. 16:864--867.
Edwards, J.H., B.D. Horton and H.C. Kirckpatrick. 1976. Aluminum
toxicity symptoms in peach seedlings. J. Am. Soc. Hortic. Sci.
101:139--142.
Foster, N.W. 1989. Acidic deposition: what is fact, what is needed?
Water Air Soil Pollut. 48:299--306.
Foy, C.D., R.L. Chaney and M.C. White. 1978. The physiology of
metal toxicity in plants. Annu. Rev. Plant Physiol. 29:511--566.
781
Gerloff, G.C. and W.H. Gabelman. 1983. Genetic basis of inorganic
plant nutrition. In Encyclopedia of Plant Physiology. Vol. 15B. New
Series. Eds. A. Lauchli and R.L. Bieleski. Springer-Verlag, Berlin,
pp 453--480.
Goldbach, E., H. Goldbach, H. Wagner and G. Michael. 1975. Influence of N-deficiency on the abscisic acid content of sunflower
plants. Physiol. Plant. 34:138--140.
Haeder, H.E. and H. Beringer. 1981. Influence of potassium nutrition
and water stress on the abscisic acid content in grains and flag
leaves during grain development. J. Sci. Food Agric. 32:552--556.
Hoagland, D.R. and D.I. Arnon. 1938. The water culture method for
growing plants without soil. Calif. Agric. Exp. Stn. Circ. 347, Univ.
of California, Berkeley, CA, 39 p.
Horst, W.J., F. Klotz and P. Szulkiewicz. 1990. Mechanical impedance
increases aluminum tolerance of soybean (Glycine max) roots. In
Plant Nutrition----Physiology and Applications. Ed. M.L. van
Beusichem. Kluwer Academic Publishers, Dordrecht, pp 351--355.
Kelly, J.M., M. Schaedle, F.C. Thornton and J.D. Joslin. 1990. Sensitivity of tree seedlings to aluminum: II. Red oak, sugar maple, and
European beech. J. Environ. Qual. 19:172--179.
Kruger, E. and E. Sucoff. 1989. Growth and nutrient status of Quercus
rubra L. in response to aluminum and calcium. J. Exp. Bot. 40:653-658.
Lachance, D. 1985. Répartition géographique et intensité du dépérissement de l’érable à sucre dans les érablières au Québec. Phytoprotection 66:83--90.
Marschner, H. 1986. Mineral nutrition in higher plants. Academic
Press Inc., London, 674 p.
NCASI. 1983. Field study program elements to assess the sensitivity
of soils to acidic deposition induced alterations in forest productivity. Technical Bulletin No. 404. Nat. Counc. of the Paper Ind. for
Air and Stream Improvement, New York, 88 p.
Ouimet, R., and J.-M. Fortin. 1992. Growth and foliar nutrient status
of sugar maple: incidence of forest decline and reaction to fertilization. Can. J. For. Res. 22:699--706.
Radin, J.W. 1984. Stomatal responses to water stress and to abscisic
acid in phosphorus-deficient cotton plants. Plant Physiol. 76:392-394.
Raynal, D.J., J.D. Joslin, F.C. Thornton, M. Schaedle and G.S. Henderson. 1990. Sensitivity of tree seedlings to aluminum: III. Red
spruce and loblolly pine. J. Environ. Qual. 19:180--187.
Robitaille, G., R. Boutin and D. Lachance. 1995. Effects of soil
freezing stress on sap flow and sugar content of mature sugar
maples (Acer saccharum). Can. J. For. Res. 25:577--587.
Steiner, K.C., J.R. Barbour and L.H. McCormick. 1984. Response of
Populus hybrids to aluminum toxicity. For. Sci. 30:404--410.
Steiner, K.C., L.H. McCormick and D.S. Canavera. 1980. Differential
response of paper birch provenances to aluminum in solution culture. Can. J. For. Res. 10:25--29.
Stienen, H. and J. Bauch. 1988. Element determination in tissues of
spruce and fir seedlings from hydroponic and soil cultures simulating acidification and deacidification. Plant Soil. 106:231--238.
Sucoff, E., F.C. Thornton and J.D. Joslin. 1990. Sensitivity of tree
seedlings to aluminum: I. Honeylocust. J. Environ. Qual. 19:163--171.
Taylor, G.J. 1991. Current views of the aluminum stress response; the
physiological basis of tolerance. Curr. Top. Plant Biochem. Physiol.
10:57--93.
Thornton, F.C., M. Schaedle and D.J. Raynal. 1986a. Effect of aluminum on the growth of sugar maple in solution culture. Can. J. For.
Res. 16:892--896.
Thornton, F.C., M. Schaedle and D.J. Raynal. 1986b. Effects of aluminum on growth, development, and nutrient composition of honeylocust (Gleditsia triacanthos) seedlings. Tree Physiol. 2:307--316.
782
BERTRAND, ROBITAILLE, BOUTIN AND NADEAU
Thornton, F.C., M. Schaedle and D.J. Raynal. 1987. Effects of aluminum on red spruce seedlings in solution culture. Environ. Exp. Bot.
27:489--498.
Thornton, F.C., M. Schaedle and D.J. Raynal. 1989. Tolerance of red
oak and American and European beech seedlings to aluminum. J.
Environ. Qual. 18:541--545.
Ulrich, B., R. Mayer and P.K. Khanna. 1980. Chemical changes due to
acid precipitation in a loess-derived soil in central Europe. Soil Sci.
130:193--199.
Weiler, E.W. 1980. Radioimmunoassays for the differential and direct
analysis of free and conjugated abscisic acid in plant extract. Planta
148:262--272.
Zhang, J. and W.J. Davies. 1990. Changes in the concentration of ABA
in xylem sap as a function of changing soil water status can account
for changes in leaf conductance and growth. Plant Cell Environ.
13:277--285.
© 1995 Heron Publishing----Victoria, Canada
Growth and ABA responses of maple seedlings to aluminum
ANNICK BERTRAND,1 GILLES ROBITAILLE,1 ROBERT BOUTIN1 and PAUL NADEAU2
1
Natural Resources Canada, Canadian Forest Service-Québec Region, P.O. Box 3800, Sainte-Foy, Québec G1V 4C7, Canada
2
Agriculture and Agri-Food Canada Research Station, 2560 Hochelaga Blvd., Sainte-Foy, Québec G1V 2J3, Canada
Received November 17, 1994
Summary We assessed the impacts of low pH and 2.0 mM
aluminum (Al) on the growth of sugar maple (Acer saccharum
Marsh.) seedlings over a 13-week period. At Week 9, total leaf
area of Al-treated seedlings was reduced by 27%; however, by
Week 13, leaf area was similar for seedlings in all treatments.
None of the other growth parameters examined were negatively
affected by the treatments at either Week 9 or Week 13. The
ABA concentration in the xylem sap, which is an indicator of
tree stress in the field, was not affected by any of the treatments
and was highest during periods of high evaporative demand in
June and August. We conclude that the duration of exposure to
Al is critical when assessing a threshold concentration for Al
toxicity because plants can acclimate to an Al concentration
previously considered toxic. Although Al stress did not appear
to reduce the vigor of sugar maple seedlings directly, it could
facilitate an inciting factor such as winter frost to induce tree
decline.
Keywords: abscisic acid, Acer saccharum, acidic deposition,
soil acidity.
Introduction
Sugar maple stands (Acer saccharum Marsh.) have shown
symptoms of decline in eastern North America since the early
1980s (Lachance 1985), but the mechanisms underlying this
decline have not been precisely defined. Acidic deposition is
considered a contributing factor that weakens the trees (Foster
1989), and extreme climatic events are thought to be inciting
factors that trigger tree decline (Robitaille et al. 1995). Others
have postulated that the detrimental effects of acidic deposition on trees are caused by mobilization of aluminum (Al) in
soil solution (Ulrich et al. 1980).
Aluminum can affect plant growth by causing an imbalance
in plant mineral nutrition (Kelly et al. 1990). High Al concentrations in the growth medium usually reduce concentrations
of magnesium, calcium and often phosphorus in tree foliage
below those required for normal metabolism. Aluminum is
strongly adsorbed on cell walls of the root cortex and blocks
the normal uptake of calcium and magnesium required in the
aerial parts of the tree (Stienen and Bauch 1988). Aluminum
can also lower phosphorus availability to the shoot by enhancing its precipitation in the soil or on the plant root (Cumming
et al. 1985). In greenhouse studies, a high Al concentration in
the nutrient solution has detrimental effects on the growth and
mineral nutrition of many tree species (Thornton et al. 1986a,
1989, Kruger and Sucoff 1989).
Hormonal imbalances may also be involved in the Al effects
on sugar maple seedling growth. According to Chapin (1990),
the growth response to an inadequate nutrient supply cannot be
explained by simple changes in the nutrient status of the plant
but probably involves complex changes in hormonal balance,
particularly in abscisic acid (ABA). Leaf ABA concentration
increases in response to various mineral deficiencies (Goldbach et al. 1975, Haeder and Beringer 1981, Radin 1984). High
ABA concentrations are also a typical response to ion toxicity,
but the increased ABA concentrations are generally explained
by an indirect salinity effect induced by a water deficit
(Marschner 1986). Few studies report a direct effect of high ion
concentration, particularly Al, on the concentrations of growth
regulators (Edwards et al. 1976).
We tested the hypothesis that low pH and high Al concentration lower the vigor of sugar maple seedlings and are contributing factors to sugar maple stand decline. The effects of these
stresses on tissue mineral concentrations and various growth
parameters were measured in roots and shoots over a 13-week
period. We also measured the concentration of ABA in the
xylem sap in response to Al over time to determine whether it
could be used as an indicator of Al stress in sugar maple
seedlings.
Materials and methods
Plant material and stress treatments
On April 20, 1993, 2-year-old sugar maple seedlings (50--90
cm in height), produced from seeds of a Saint-Georges-deBeauce provenance (Québec, 46°10′ N, 70°37′ W), were
planted in 16-liter pots containing washed silica (silica 16,
Indusmin, Saint-Canut, Québec) and acclimated in a greenhouse at the Laurentian Forestry Centre, Sainte-Foy, Québec,
for 7 weeks (day/night temperature of 23/20 °C and a 16-h
photoperiod). The seedlings were fertilized weekly with halfstrength Hoagland solution (Hoagland and Arnon 1938). At the
beginning of June 1993, a shade cloth was installed in the
greenhouse 2 m above the trees. The five treatments (n = 9 per
treatment) were started on June 8, 1993, and consisted of
irrigations with: (1) complete nutrient solution at pH 5.0 (con-
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BERTRAND, ROBITAILLE, BOUTIN AND NADEAU
trol), (2) complete nutrient solution at pH 4.0 (pH 4.0), (3)
complete nutrient solution at pH 4.0 with the addition of 2.0
mM Al (pH 4.0 + Al), (4) complete nutrient solution at pH 3.5
(pH 3.5), and (5) complete nutrient solution at pH 3.5 with the
addition of 2.0 mM Al (pH 3.5 + Al). The composition of the
nutrient solution was similar to that described by Thornton et
al. (1986a): 0.25 mM Ca(NO3)2, 0.5 mM KNO3, 0.5 mM
MgSO4, 0.5 mM NH4NO3, 0.04 mM NH4H2PO4, 10 µM Fe
EDTA, 0.8 µM Zn, 9.0 µM Mn, 0.3 µM Cu, 46 µM B, and 0.05
µM Mo. Aluminum was added as AlCl3 for Treatments 3 and
5. The Cl− concentrations of all treatments were equalized with
NaCl. The pH of the solutions was adjusted with HCl. Irrigations with 0.7 l of each solution took place twice weekly during
the 13-week experiment from June 8 (Day 0) to September 8
(Day 91), 1993. Once a week, all pots were also irrigated by a
spray system with demineralized water. Four destructive harvests were made: June 8 (Day 0), July 7--8 (Week 4), August
10--11 (Week 9) and September 7--8 (Week 13).
Growth parameters
At each harvest, the total weight and height (root collar to
terminal bud) of each seedling were measured. We also determined the weight of stems, leaves, and large (diameter > 1 mm)
and fine (diameter < 1 mm) roots. The roots were thoroughly
washed with demineralized water to remove adhering particles. After blotting, the total volume of the root system was
measured by a water displacement technique described by
Burdett (1979). After fine root removal, the volume of roots
larger than 1 mm was measured, as previously described, and
the volume of fine roots (diameter < 1 mm) was calculated by
the difference between the total root volume and the large root
volume. The total leaf area of each seedling was measured with
an image analyzer (AGVision, Pullman, WA). Each type of
tissue was then dried for 72 h at 70 °C. The root/shoot ratio
was calculated on the basis of tissue dry weight. The root
relative growth rate (RGR) was calculated on the basis of root
volume and expressed as a percentage: ((final root volume −
initial root volume)/initial root volume) × 100.
Xylem sap extraction and ABA quantification
The terminal twig (approximately 12 cm long), including five
to ten leaves (or the lateral twig when the terminal twig was
missing), was removed and immediately placed in a pressure
chamber (PMS Instruments Co., Corvallis, OR) with the bottom covered with a damp paper towel. A gradual pressure was
exerted on the twig, and the pressure was then maintained
between 2.0 and 3.0 MPa for approximately 5 min for the
collection of 100 µl of xylem sap. The vials containing the sap
were immediately put on ice and stored at −80 °C within 2 h
of collection.
After a 30-min incubation with polyvinylpolypyrrolidone
(0.1 mg per 100 µl), xylem sap was assayed for free ABA
content by radioimmunoassay (RIA) as described by Bertrand
et al. (1994). Antibodies specific to free (+) cis-trans ABA
were prepared according to Weiler (1980). The antibody used
showed a linear response from 0.1 to 10.0 pmol under RIA
conditions. The antiserum was highly specific with a cross-reactivity of 53% for (±) cis-trans ABA, 32% for (±) racemic
ABA and 7% for methylated (±) ABA. The affinity constant for
the antiserum was 2.47 × 10 −9 l mol −1. All standards and
samples were assayed in triplicate. Samples were diluted 1/10
in phosphate buffered saline solution (Weiler 1980).
Extraction and analysis of the silica solution
At each harvest date, 1 day after a fertilization treatment, 600 g
of silica was centrifuged at 1000 g for 5 min to extract the silica
solution. The samples were then placed in a cold room (2 °C)
until they were prepared for chemical analysis. The samples
were filtered at 0.45 µm, and the conductivity and pH measured. Total monomeric Al (with chrome azurol S) was measured by colorimetry (Flow Injection Analysis, Tecator 5020,
Sweden) with a detection limit of 2.0 µmol l −1; total Ca and
Mg were measured by atomic absorption spectrophotometry.
Analysis of the mineral composition of plant tissues
The dried plant material was ground to pass through a 2-mm
mesh. Subsamples of 0.5 g of leaf and root material from four
replicates per treatment were analyzed according to the dry
ashing method (NCASI 1983). Total P content of samples was
determined by colorimetry (Colorimeter PC 800, Brinkman
Instruments, Westbury, NY). Total tissue content of Al, Ca, Mg
and Mn was analyzed by atomic absorption spectrophotometry
with nitrous oxide for Al, and flame emission spectrophotometry for K and Na. The detection limits for the ions (in
mg l −1) were: Ca, 0.1; Mn, 0.06; K, 0.05; Mg, 0.01; and Al, 1.1.
The accuracy of tissue analysis was confirmed by comparison
with the Quality Assurance Subgroup of the Federal-Provincial Research and Monitoring Coordinating Committee reference material. For each ion under study, the results from our
laboratory were less than ± 6% of the median results from 18
laboratories.
Statistical analysis
A total of 135 seedlings (9 blocks (replicates) × 5 treatments
3 sampling dates) were arranged in the greenhouse in a randomized complete block design. The data were analyzed by
the GLM procedure of SAS (SAS Institute Inc., Cary, NC)
after the confirmation of homogeneity of variances through
residual analysis. The assumption of variance homogeneity
was not satisfied for ABA, and a logarithmic transformation
was used to stabilize the variance. The initial root volume was
determined before treatments were initiated and the effect of
this covariate was accounted for in the statistical analysis of
‘‘final root volume’’ after verification of the homogeneity of
slopes. The differences between means for treatments and for
dates of data collection were compared by orthogonal contrasts
or the LSD test. When the effects of two or more treatments
were not significantly different for a given variable, we present
one curve representing the mean of the results obtained.
RESPONSE OF MAPLE SEEDLINGS TO ALUMINUM STRESS
Results
Silica solution composition
Before the treatments began, the pH of the solution extracted
from the silica growth medium was 6.9. At Week 4, the pH
values of the silica solutions were less for all treatments than
at Day 0, and the reductions were significantly greater in the
Al treatments than in the other treatments (P ≤ 0.05) (Figure 1A). The acidity of the silica solutions did not differ significantly between the two Al treatments (Treatments 3 and 5)
or between the two pH treatments (Treatments 2 and 4) (Figure 1A). The pH of the silica solutions for treatments without
Al (Treatments 1, 2 and 4) decreased sharply to 4.5--5.2 between Weeks 9 and 13. The conductivities of the silica solu-
Figure 1. (A) pH, (B) conductivity of the silica solution, and (C) soil
water content (SWC) at Weeks 4, 9 and 13 (n = 9). Symbols: d,
control; n, pH 4.0; h, pH 4.0 + Al; m, pH 3.5; j, pH 3.5 + Al. Each
curve represents the mean for nonsignificantly different treatments.
777
tions were similar for all treatments (Figure 1B). The low
conductivity of the silica solution on Day 0 corresponded to
the nutrient solution used before the treatments began, i.e.,
before the addition of AlCl3 or NaCl. The highest conductivity
(Figure 1B) was reached at Week 9 and corresponded with a
period of high evaporative demand in the greenhouse when soil
water content was at a minimum (Figure 1C).
In the pH 4.0 + Al and pH 3.5 + Al treatments, the Al
concentration of the silica solutions increased from 1.25 to
2.30 mM between Weeks 4 and 9, and then stabilized (Figure 2A). The Al/Mg ratio followed the same trend and stabilized at approximately 3.4 (Figure 2B). The Al/Ca molar ratio
doubled from 2 to 4 between Weeks 4 and 9, and then increased
more gradually to reach about 5 by Week 13 (Figure 2C). There
were no significant differences between the two Al treatments
for these variables. Throughout the study, the Mg2+ and Ca2+
concentrations in the silica solution remained higher in the Al
treatments than in the other treatments (Figures 3A and 3B).
Figure 2. (A) Al concentration, (B) Al/Mg molar ratio, and (C) Al/Ca
molar ratio of the silica solution at Weeks 4, 9 and 13 (mean ± SE,
n = 9). Symbols: j, 2.0 mM Al at pH 4.0; h, 2.0 mM Al at pH 3.5.
778
BERTRAND, ROBITAILLE, BOUTIN AND NADEAU
Seedling growth and ABA concentration
Tissue mineral concentration
The 2.0 mM Al treatment stimulated root growth. By Week 13,
the root volume of Al-treated seedlings was approximately 75
cm3 compared with a mean of ≈65 cm3 for seedlings in the
other treatments (P ≤ 0.05) (Figure 4A). In response to the
presence of Al, the relative root growth rate ranged from 17 to
22% compared with 3% in the control seedlings (Figure 4B).
The Al treatment significantly (P ≤ 0.05) enhanced the
root/shoot ratio compared with the other treatments (mean
value = 0.95 versus 0.81) (Figure 4C).
Leaf area was the only parameter negatively affected by the
Al treatments (P ≤ 0.05). At Week 9, leaf area had decreased
by about 27% in Al-treated seedlings compared with seedlings
in the other treatments (Figure 5A); however, by Week 13, the
total leaf area of seedlings in all treatments was about 1000
cm2. Between Weeks 4 and 13, the fine root volume/large root
volume ratio decreased progressively and similarly for seedlings in all treatments (Figure 5B).
The ABA concentration in the xylem sap was highest (403
pmol ml −1) at the beginning of the experiment when the soil
water content was low. It decreased sharply to reach 144 pmol
ml −1 at Week 4, and then increased to 210 pmol ml −1 between
Weeks 4 and 9 as the soil dried (cf. Figures 1C and 5C). None
of the treatments had a significant effect on xylem sap ABA
concentration.
The mean Al concentration of roots of Al-treated seedlings
increased from Week 4 to Week 13 reaching a mean of 1.1 mg
gDW−1, which was three times the concentration found in roots
of control and pH-treated seedlings (Figure 6A). In leaves, a
maximum Al concentration of ≈0.6 mg gDW−1 was found in
Al-treated seedlings at Week 9 (Figure 6B). Concentrations of
Ca, P and Mn in roots and leaves were not affected by any of
the treatments (Table 1). The Mg concentration was significantly lower in roots of Al-treated seedlings (P ≤ 0.05) than in
roots of seedlings in the other treatments, whereas leaf Mg
concentration was not affected by any of the treatments (6.7-7.0 mg gDW−1) (Table 1). Root K concentration was slightly
higher (P ≤ 0.05) in Al-treated seedlings than in seedlings in
the other treatments (≈4.5 versus ≈3.5 mg gDW−1), whereas
foliar K concentration was lowest in Al-treated seedlings (≈4.0
Figure 3. (A) Mg, and (B) Ca concentrations in the silica solution at
Weeks 4, 9 and 13 (n = 9). Symbols: d, control; n, pH 4.0; h, pH 4.0
+ Al; m, pH 3.5; j, pH 3.5 + Al. Each curve represents the mean for
nonsignificantly different treatments.
Figure 4. (A) Final root volume, (B) relative root growth rate (volumetric RGR), and (C) root/shoot ratio in response to five treatments.
Treatments with the same letter are not significantly different at P =
0.05; mean ± SE for n = 27.
RESPONSE OF MAPLE SEEDLINGS TO ALUMINUM STRESS
779
Figure 6. (A) Root Al concentration, (B) leaf Al concentration, and (C)
soil water content (SWC) at Weeks 4, 9 and 13 (n = 9). Symbols: d,
control; n, pH 4.0; h, pH 4.0 + Al; m, pH 3.5; j, pH 3.5 + Al. Each
curve represents the mean for nonsignificantly different treatments.
Figure 5. (A) Total leaf area per seedling, (B) fine/large root ratio, and
(C) xylem sap ABA concentration at Weeks 4, 9 and 13 (n = 9).
Symbols: d, control; n, pH 4.0; h, pH 4.0 + Al; m, pH 3.5; j, pH
3.5 + Al. Each curve represents the mean for nonsignificantly different
treatments.
mg gDW−1) (Table 1). Root Na concentration in Al-treated
seedlings (≈ 0.7 mg gDW−1) was approximately half the concentration found in the roots of seedlings in the other treatments.
Foliar Na concentrations were lowest in Al-treated seedlings
(0.4 mg gDW−1) and highest in pH-treated seedlings (1.6 mg
gDW−1) (Table 1).
Discussion
The negative effects of Al were more pronounced in the aerial
part (shoots and leaves) than in the roots of sugar maple
seedlings. The finding that the aerial part was more sensitive
to Al than the roots may be a common feature of deciduous tree
species (Thornton et al. 1986a, Kelly et al. 1990).
In our study, the Al concentration of the roots was much
lower than the concentration reported by Thornton et al.
(1986a) in response to a similar Al treatment (2.0 mM) and
probably accounts for the absence of toxic Al effects on the
root system. Reductions in root growth rate, root branching, or
both that are generally associated with Al stress are usually
explained by a decrease in Ca2+ or Mg2+ uptake by the root
system in the presence of Al3+, resulting in a nutritional deficiency of these elements (Edwards et al. 1976, Thornton et al.
1987). The concentrations of Ca2+ and Mg2+ measured in the
silica solution of Al-treated seedlings were higher than in the
silica solution of control seedlings, which indicates that Al
effectively blocked the entry of these ions into the root. We
conclude that, under our experimental conditions, the blocking
effect did not cause Ca or Mg deficiencies, because these
elements were supplied in sufficient amounts in the nutrient
medium (cf. Kelly et al. 1990). The absence of an Al effect on
Ca concentration in leaves and roots differs from the observations made on honeylocust, peaches and coniferous trees in
which the tissue concentrations of Ca were depleted in the
presence of Al (Edwards et al. 1976, Sucoff et al. 1990, Raynal
et al. 1990). Kelly et al. (1990) concluded that the molar ratio
of Al/Ca of the nutrient solution determines the interaction
between the two elements. The Al/Ca ratio of our silica solution was about 2 at Week 4 and increased to about 5 at Week
13. However, Kelly et al. (1990) observed that a wide range of
Al/Ca ratios was ineffective in modifying the growth of red oak
780
BERTRAND, ROBITAILLE, BOUTIN AND NADEAU
Table 1. Mineral concentration of sugar maple roots and leaves (mg gDW−1) in response to five treatments for the pooled sampling dates. Different
letters indicate significant differences among treatments at P ≤ 0.05; mean (± SE) for n = 27.
Element
Control
pH 4.0
pH 3.5
pH 4.0 + Al
pH 3.5 + Al
Roots
Ca
Mg
K
P
Mn
Na
5.68 (0.47)
1.60 (0.09) a
3.07 (0.20) b
1.30 (0.07)
0.09 (0.01)
1.41 (0.12) a
5.91 (0.42)
1.84 (0.09) a
3.94 (0.22) b
1.62 (0.12)
0.14 (0.02)
1.34 (0.10) a
5.81 (0.61)
1.79 (0.10) a
3.68 (0.34) b
1.56 (0.13)
0.12 (0.02)
1.37 (0.11) a
4.75 (0.40)
1.45 (0.06) b
4.47 (0.30) a
1.22 (0.10)
0.09 (0.006)
0.78 (0.08) b
5.12 (0.34)
1.56 (0.10) b
4.66 (0.21) a
1.36 (0.14)
0.11 (0.006)
0.90 (0.10) b
Leaves
Ca
Mg
K
P
Mn
Na
18.84 (1.16)
6.71 (0.34)
4.86 (0.60) ab
1.69 (0.11)
0.21 (0.04)
0.85 (0.20) b
22.07 (1.63)
6.92 (0.31)
6.39 (0.70) a
2.07 (0.18)
0.19 (0.03)
1.85 (0.39) a
21.41 (2.01)
6.90 (0.30)
5.53 (.55) a
1.97 (0.16)
0.28 (0.05)
1.18 (0.36) a
21.69 (1.94)
7.03 (0.36)
4.06 (0.28) b
1.72 (0.20)
0.28 (0.05)
0.40 (0.06) c
21.30 (1.22)
6.89 (0.37)
4.20 (0.31) b
1.81 (0.15)
0.28 (0.06)
0.40 (0.04) c
and suggested that the observed differences in Al sensitivity
exhibited by tree species were associated with experimental
conditions resulting in differences in either Ca2+ availability or
ionic strength of the solution. Our results support the suggestion that Al sensitivity can be offset by the presence of suitable
amounts of plant-available Ca.
We observed a stimulation of relative root growth rate by 2.0
mM Al3+. Thornton et al. (1986a) reported a similar observation at 1.0 mM Al3+; however at 2.0 mm Al3+, these workers
reported detrimental effects of Al3+ on roots growing in solution culture, suggesting that the results obtained from plants in
solution culture are not directly comparable with those obtained from plants in silica culture. Although a steady-state
nutrient supply, as is possible in solution culture, cannot be
achieved when silica is used as the growing medium, silica
culture provides information about plant responses to stress
such as exclusion mechanisms and root exudates (Taylor
1991). It has been shown for Glycine max (L.) Merill. that the
mechanical impedance caused by sand culture increased tolerance to Al stress as a result of reduced Al uptake into the root
tips compared with uptake from solution culture (Horst et al.
1990). These authors suggest that enhanced release of exudates
might account for the higher Al tolerance in sand culture than
in solution culture. Plants could exclude Al from the symplasm
if chelate ligands were released into the rhizosphere and these
ligands formed stable complexes with Al, thus reducing the
absorption of the metal ion across the plasma membrane (Taylor 1991). In addition to root exudates, there is the possibility
that genotype may also strongly influence responses.
The Al treatments caused decreases in leaf area between
Weeks 4 and 9 so that at Week 9, total leaf area per tree was
reduced by more than 27% in Al-treated seedlings relative to
seedlings in the other treatments. By Week 13, the total leaf
area was similar for all treatments even though the leaf Al
concentration was twice as high in Al-treated seedlings as in
untreated seedlings. Thornton et al. (1986b) observed an absolute, but small (≤ 10% over a 30-day period), increase in
damage with increasing exposure time to Al. Our results show
that plants can acclimate to a concentration of Al previously
considered toxic and indicate that the duration of exposure to
Al should therefore be taken into account when assessing a
critical concentration for Al toxicity in silica culture. Possible
mechanisms of acclimation include entrapment of excess Al in
roots, reducing Al concentration in seedling tops (Foy et al.
1978) as observed at Week 13, and an enzyme reaction to Al
stress in the leaves (Bowler et al. 1992).
Leaf K concentration decreased in response to the Al treatments. Translocation of K from roots to leaves may have been
impaired in the presence of Al because the root K concentration was significantly higher in Al-treated seedlings than in
control seedlings. In our study, the leaf K concentration (≈4.0
mg gDW−1) of all the seedlings was below the critical concentration of 6.0 mg g−1 needed for adequate nutrition of A. saccharum (Bernier and Brazeau 1988) and lower than the
concentration reported for mature sugar maple trees (Bernier
et al. 1989) or for sugar maple seedlings growing in solution
culture (Thornton et al. 1986a). We note that deficiencies in
leaf K in sugar maple trees growing in the Beauce region
(Québec, Canada), which is the provenance of our seedlings,
have been associated with incidences of stand decline in this
region (Ouimet and Fortin 1992). The numerous differences in
nutrition requirements among varieties or lines of plants suggest genetic control of inorganic plant nutrition (Gerloff and
Gabelman 1983). Moreover, a large range of variation in sensitivity to Al has been observed among various provenances of
paper birch (Steiner et al. 1980), among Populus hybrids (Steiner et al. 1984), and among genotypes of loblolly pine (Raynal
et al. 1990). The differential intraspecific sensitivity to toxic
ions also reflects genetic differences in mineral nutrition,
which could prove useful in breeding trees for tolerance to
nutritional stress.
The treatments had no effect on ABA concentration in the
xylem sap. Maximum ABA concentrations in the xylem sap
were reached at the onset of the experiment and at Week 9,
RESPONSE OF MAPLE SEEDLINGS TO ALUMINUM STRESS
which corresponded to low soil water contents. A direct increase in ABA concentration of the xylem sap in response to
soil water depletion has been observed in maize (Zhang and
Davies 1990). At Week 9, when the soil water content was at
its lowest value, the ABA concentration in the xylem sap was
comparable to the concentration measured in mature sugar
maple trees during a physiological drought stress caused by the
freezing of soil water, whereas at Weeks 4 and 13, the ABA
concentration in the xylem sap was similar to that in untreated
trees with an adequate water supply (Bertrand et al. 1994).
Based on this comparison of field and laboratory results, we
conclude that water stress was the principal cause of increases
in ABA concentration in the xylem sap and that Al had little or
no effect on ABA concentration of the xylem sap.
Aluminum stress or other nutritional imbalances may facilitate an inciting factor such as early or late winter frost to induce
sugar maple tree decline. The impacts of high concentrations
of Al and acid rain acting alone and in combination on the cold
tolerance of sugar maple seedlings are under investigation.
Acknowledgments
We thank Mr. Pascal Aubé and Mr. Clément Aubin for their technical
assistance, Mr. Alain LePage for his contribution to the chemical
analysis, and Ms. Michèle Bernier-Cardou for discussions on statistical analysis. This work was supported by the Long Range Transport of
Atmospheric Pollutants (LRTAP).
References
Bernier, B. and M. Brazeau. 1988. Foliar nutrient status in relation to
sugar maple dieback and decline in the Québec Appalachians. Can.
J. For. Res. 18:754--761.
Bernier, B., D. Par and M. Brazeau. 1989. Natural stresses, nutrient
imbalances and forest decline in southeastern Québec. Water Air
Soil Pollut. 48:239--250.
Bertrand, A., G. Robitaille, P. Nadeau and R. Boutin. 1994. Effects of
soil freezing and drought stress on abscisic acid content of sugar
maple sap and leaves. Tree Physiol. 14:413--425.
Bowler, C., M. Van Montagu and D. Inz. 1992. Superoxide dismutase
and stress tolerance. Annu. Rev. Plant Physiol. Plant Mol. Biol.
43:83--116.
Burdett, A.N. 1979. A nondestructive method for measuring the volume of intact plant parts. Can. J. For. Res. 9:120--122.
Chapin III, F.S. 1990. Effects of nutrient deficiency on plant growth:
evidence for a centralized stress-response system. In Importance of
Root to Shoot Communication in the Response to Environmental
Stress. Eds. W.J. Davies and B. Jeffcoat. British Society for Plant
Growth Regulation, Lancaster, pp 135--148.
Cumming, J.R., R.T. Eckert and L.S. Evans. 1985. Effect of aluminum
on 32P uptake and translocation by red spruce seedlings. Can. J. For.
Res. 16:864--867.
Edwards, J.H., B.D. Horton and H.C. Kirckpatrick. 1976. Aluminum
toxicity symptoms in peach seedlings. J. Am. Soc. Hortic. Sci.
101:139--142.
Foster, N.W. 1989. Acidic deposition: what is fact, what is needed?
Water Air Soil Pollut. 48:299--306.
Foy, C.D., R.L. Chaney and M.C. White. 1978. The physiology of
metal toxicity in plants. Annu. Rev. Plant Physiol. 29:511--566.
781
Gerloff, G.C. and W.H. Gabelman. 1983. Genetic basis of inorganic
plant nutrition. In Encyclopedia of Plant Physiology. Vol. 15B. New
Series. Eds. A. Lauchli and R.L. Bieleski. Springer-Verlag, Berlin,
pp 453--480.
Goldbach, E., H. Goldbach, H. Wagner and G. Michael. 1975. Influence of N-deficiency on the abscisic acid content of sunflower
plants. Physiol. Plant. 34:138--140.
Haeder, H.E. and H. Beringer. 1981. Influence of potassium nutrition
and water stress on the abscisic acid content in grains and flag
leaves during grain development. J. Sci. Food Agric. 32:552--556.
Hoagland, D.R. and D.I. Arnon. 1938. The water culture method for
growing plants without soil. Calif. Agric. Exp. Stn. Circ. 347, Univ.
of California, Berkeley, CA, 39 p.
Horst, W.J., F. Klotz and P. Szulkiewicz. 1990. Mechanical impedance
increases aluminum tolerance of soybean (Glycine max) roots. In
Plant Nutrition----Physiology and Applications. Ed. M.L. van
Beusichem. Kluwer Academic Publishers, Dordrecht, pp 351--355.
Kelly, J.M., M. Schaedle, F.C. Thornton and J.D. Joslin. 1990. Sensitivity of tree seedlings to aluminum: II. Red oak, sugar maple, and
European beech. J. Environ. Qual. 19:172--179.
Kruger, E. and E. Sucoff. 1989. Growth and nutrient status of Quercus
rubra L. in response to aluminum and calcium. J. Exp. Bot. 40:653-658.
Lachance, D. 1985. Répartition géographique et intensité du dépérissement de l’érable à sucre dans les érablières au Québec. Phytoprotection 66:83--90.
Marschner, H. 1986. Mineral nutrition in higher plants. Academic
Press Inc., London, 674 p.
NCASI. 1983. Field study program elements to assess the sensitivity
of soils to acidic deposition induced alterations in forest productivity. Technical Bulletin No. 404. Nat. Counc. of the Paper Ind. for
Air and Stream Improvement, New York, 88 p.
Ouimet, R., and J.-M. Fortin. 1992. Growth and foliar nutrient status
of sugar maple: incidence of forest decline and reaction to fertilization. Can. J. For. Res. 22:699--706.
Radin, J.W. 1984. Stomatal responses to water stress and to abscisic
acid in phosphorus-deficient cotton plants. Plant Physiol. 76:392-394.
Raynal, D.J., J.D. Joslin, F.C. Thornton, M. Schaedle and G.S. Henderson. 1990. Sensitivity of tree seedlings to aluminum: III. Red
spruce and loblolly pine. J. Environ. Qual. 19:180--187.
Robitaille, G., R. Boutin and D. Lachance. 1995. Effects of soil
freezing stress on sap flow and sugar content of mature sugar
maples (Acer saccharum). Can. J. For. Res. 25:577--587.
Steiner, K.C., J.R. Barbour and L.H. McCormick. 1984. Response of
Populus hybrids to aluminum toxicity. For. Sci. 30:404--410.
Steiner, K.C., L.H. McCormick and D.S. Canavera. 1980. Differential
response of paper birch provenances to aluminum in solution culture. Can. J. For. Res. 10:25--29.
Stienen, H. and J. Bauch. 1988. Element determination in tissues of
spruce and fir seedlings from hydroponic and soil cultures simulating acidification and deacidification. Plant Soil. 106:231--238.
Sucoff, E., F.C. Thornton and J.D. Joslin. 1990. Sensitivity of tree
seedlings to aluminum: I. Honeylocust. J. Environ. Qual. 19:163--171.
Taylor, G.J. 1991. Current views of the aluminum stress response; the
physiological basis of tolerance. Curr. Top. Plant Biochem. Physiol.
10:57--93.
Thornton, F.C., M. Schaedle and D.J. Raynal. 1986a. Effect of aluminum on the growth of sugar maple in solution culture. Can. J. For.
Res. 16:892--896.
Thornton, F.C., M. Schaedle and D.J. Raynal. 1986b. Effects of aluminum on growth, development, and nutrient composition of honeylocust (Gleditsia triacanthos) seedlings. Tree Physiol. 2:307--316.
782
BERTRAND, ROBITAILLE, BOUTIN AND NADEAU
Thornton, F.C., M. Schaedle and D.J. Raynal. 1987. Effects of aluminum on red spruce seedlings in solution culture. Environ. Exp. Bot.
27:489--498.
Thornton, F.C., M. Schaedle and D.J. Raynal. 1989. Tolerance of red
oak and American and European beech seedlings to aluminum. J.
Environ. Qual. 18:541--545.
Ulrich, B., R. Mayer and P.K. Khanna. 1980. Chemical changes due to
acid precipitation in a loess-derived soil in central Europe. Soil Sci.
130:193--199.
Weiler, E.W. 1980. Radioimmunoassays for the differential and direct
analysis of free and conjugated abscisic acid in plant extract. Planta
148:262--272.
Zhang, J. and W.J. Davies. 1990. Changes in the concentration of ABA
in xylem sap as a function of changing soil water status can account
for changes in leaf conductance and growth. Plant Cell Environ.
13:277--285.