Tree Physiology 24, 1295–1302
© 2004 Heron Publishing—Victoria, Canada
Wood density and anatomy of water-limited eucalypts
MATTHEW J. SEARSON,1,2 DANE S. THOMAS,3 KELVIN D. MONTAGU 4 and JANN
P. CONROY 1
1
Centre for Horticulture and Plant Sciences, University of Western Sydney, Locked Bag 1797, Penrith South DC, NSW 1797, Australia
2
Corresponding author ([email protected])
3
State Forests of NSW, Research and Development Division, P.O. Box J19, Coffs Harbour Jetty, NSW 2450, Australia
4
State Forests of NSW, Research and Development Division, P.O. Box 100, Beecroft, NSW 2119, Australia
Received December 22, 2003; accepted April 18, 2004; published online September 1, 2004
Summary We hypothesized that seedlings grown under water-limited conditions would develop denser wood than seedlings grown under well-watered conditions. Three Eucalyptus
species (E. grandis Hill (ex Maiden), E. sideroxylon Cunn. (ex
Woolls) and E. occidentalis Endl.) were grown in a temperature-controlled greenhouse for 19 weeks with watering treatments (well-watered and water-limited) applied at six weeks.
The water-limitation treatment consisted of four drought cycles. Wood density increased by between 4 and 13% in the water-limited seedlings, but this increase was mainly due to
extractive compounds embedded in the cell wall matrix. Once
these compounds were removed, the increase was 0 –9% and
was significant for E. grandis only. Water-limitation significantly reduced mean vessel lumen area, however, this was balanced by a trend toward greater vessel frequency in waterlimited plants, and consequently there was no difference in the
proportion of stem area allocated to vessels. Conduit efficiency
value was lowest in the water-limited plants, indicating that
there was a cost in terms of stem hydraulic conductivity for decreasing vessel lumen area. Wood density was negatively correlated with vessel lumen fraction in well-watered plants, but
this relationship broke down in the water-limited plants, possibly because of the significantly larger proportion of the stem
taken up by pith in water-limited seedlings. Diurnal variation in
leaf water potential was positively correlated with wood density in well-watered plants. This relationship did not hold in the
water-limited plants owing to the collapse of the pressure gradient between soil and leaf. We conclude that drought periods
of greater than 1 month are required to increase wood density in
these species and that increases in wood density appear to result in diminished capacity to supply water to leaves.
Keywords: conduit efficiency, extractives, pith, vessels, water
stress.
Introduction
The wood density of a stem is a gross measurement of its internal anatomy. The proportion of wood volume taken up by cell
lumens determines the reduction in wood density below the
theoretical maximum density of 1530 kg m –3 (Bamber and
Burley 1983). Smaller cell lumens or thicker cell walls will,
therefore, result in increased wood density. For some tree species, there is a trade-off between wood density and hydraulic
conductivity (Stratton et al. 2000, Barbour and Whitehead
2003, Thomas et al. 2004). Central to this link between stem
tissue density and hydraulic conductivity are the xylem conduits, which in the case of angiosperms are the vessels. The hydraulic conductivity of a tree stem is proportional to the sum of
all conduit radii raised to the fourth power, as described by
the Hagen-Poiseuille law (Tyree and Ewers 1991). Therefore,
when conduits are small in diameter or present at low frequency, hydraulic conductivity is low because more wood volume is taken up by cell wall material, resulting in increased
density. Meinzer (2003) showed that a potential consequence
of smaller diameter or less frequent vessels, or both, in highly
dense wood is that there is a positive correlation between wood
density and diurnal variation in leaf water potential (∆Ψleaf).
This indicates that the conduits pose a greater resistance to water flow in high-density wood than in low-density wood.
Several studies have attempted to relate the effects of water
stress or osmotic stress to wood density in the genus Eucalyptus. An increase in wood density has been associated with
decreased soil water supply in Eucalyptus globulus Labill.
(Wimmer et al. 2002) and E. nitens (Deane & Maiden) Maiden
trees (Beadle et al. 2001). In contrast, Macfarlane and Adams
(1998) and Catchpoole et al. (2000) found no relationship between wood density and the severity of water stress or soil salinity, respectively, in E. globulus. This lack of a consistent
trend may be associated with site factors such as temperature
or nutrient supply, but it may also be related to extractives (i.e.,
those components that can be removed by solvent) embedded
in the cell wall matrix of the wood. Ona et al. (1997, 1998) reported that the density of E. camaldulensis Dehnh. and E. globulus wood declined if the extractives were removed. When
the extractives are removed, stem wood density more accurately represents the anatomy of the wood within.
We describe an experiment to test the hypotheses that waterlimited greenhouse-grown Eucalyptus seedlings have greater
stem wood density than well-watered seedlings once the extractives are removed. Additionally, we investigated the rela-
1296
SEARSON, THOMAS, MONTAGU AND CONROY
tionship between wood density and ∆Ψleaf in these greenhouse-grown seedlings. We tested these hypotheses in three
Eucalyptus species: E. grandis Hill (ex Maiden), E. sideroxylon Cunn. (ex Woolls) and E. occidentalis Endl.
Materials and methods
Plant culture
Seeds of the three species was obtained from the Australia
Tree Seed Centre, CSIRO, ACT, Australia. The seed source for
E. grandis (Seedlot 13019: 28°40′ S, 152°90′ E) was the Coffs
Harbour region of northern New South Wales (NSW), Australia. Eucalyptus sideroxylon (seedlot 19557; 31°20′ S, 148°
50′ E) seed was from the dryer region of mid-western NSW,
Australia. Eucalyptus occidentalis (seedlot 13638; 33°10′ S,
and 119°60′ E) was from the Mallee physiographic region of
southwest, Western Australia.
Seedlings were germinated in trays containing 1:1 (v/v)
soil:pearlite. After growing in the germination trays for 7 weeks,
the seedlings were transplanted to pots (height 42.5 cm, diameter 15.5 cm, 8-l volume and lined with plastic bags to prevent
water loss) containing soil from the Robertson region of NSW,
Australia that was supplemented with mineral nutrients as described by Sefton et al. (2002). The potted seedlings were
grown under well-watered conditions for 6 weeks before the
commencement of the watering treatments, designated Week 0.
Experimental and growth conditions
The experiment was carried out over 13 weeks (from Week 0)
in a single room of a naturally illuminated, climate-controlled
greenhouse on the Hawkesbury campus of the University of
Western Sydney. Pots were arranged in a randomized block
design. Day/night temperature was controlled at 25/20 °C, and
mean daytime relative humidity was 55%, ranging from 24 to
89% at midday during the study. Greenhouse air temperature
and humidity were monitored with microprocessor data loggers (TGP-1500, Tiny-tag Plus, Gemini Data loggers, Chichester, U.K.).
The experiment was a 3 × 2 factorial design with three Eucalyptus species and two watering treatments replicated five
times. The watering treatments were well-watered or waterlimited. In the well-watered treatment, soil was brought to
100% of field capacity (FC) every third day, whereas the soil
in the water-limited treatments was allowed to dry to 50% FC
before it was re-watered to 90% FC. The FC of the soil was determined to be 0.35 cm 3 cm –3. Soil water content declined to
50% FC four times during the experiment and these drought
cycles lasted for 33, 27, 18 and 19 days, decreasing as the seedlings grew larger and consumed water more rapidly. The experimental design was duplicated to accommodate two harvests, one after 4 weeks and the other at 13 weeks. Five
replicates of each species were germinated for a preliminary
harvest at Week 0.
ment, Santa Barbara, CA) during Week 13 when soil water
content was at its lowest value in the water-limited treatment.
Sampled leaves were the youngest fully expanded leaf attached to the main stem of each replicate. Measurements were
made before dawn (0500 – 0600 h), at midday (1200 –1300 h)
and early afternoon (1500 h) on the same day. Diurnal variation in Ψleaf (∆Ψleaf ) was calculated as the difference between
predawn Ψleaf and the most negative of the two afternoon measurements.
Harvests
There were two main harvests, one at 4 weeks and the other
13 weeks after the introduction of the watering treatments, and
there was also a preliminary harvest at Week 0. At the preliminary harvest, five replicates of each species were randomly
chosen and harvested. Stem diameter and wood density were
measured (see description of methods below). The Week 4
harvest was for the determination of plant dry mass and the
harvest at 13 weeks was for the more extensive measurements.
At Week 4, the roots were cut from the shoots and washed free
of soil. Both plant parts were dried at 70 °C for 48 h and total
dry mass determined. At Week 13, plant height and diameter
were measured. The shoots were cut from the roots at soil
level. A 6-cm-long sample of the main stem was taken from
near the base of the seedling and kept for wood density measurements. The leaf area of each seedling was measured with a
digital area measurement system (DIAS, Delta-T Devices,
Cambridge, U.K.). Leaf petioles were included in the leaf
sample. The roots were washed free of soil and then all plant
parts were dried at 70 °C for 48 h and dry mass measured. Relative growth rate (RGR) was calculated from the difference in
the natural log of dry matter increment between the two harvests (Atwell et al. 1999).
Wood density
The periderm was removed from all stem segments before
wood density measurements were made. Wood density was
measured on the basal 6-cm stem segment. Fresh wood volume (Vfs ) of the 6-cm stem segment was measured using the
Archimedes principal by immersing the segment in a beaker
containing distilled water of known mass on a four-place balance. Each sample was lanced with a probe and placed just under the surface of the water. A clamp attached to a retort stand
supported the probe, minimizing any movement of the sample.
Once Vfs was measured, the sample was freeze-dried and dry
mass (Wds) determined. Total density was measured on a mass/
volume basis, Dt (kg m –3) as:
Dt =
(1)
Because some of the Wds is composed of extractives (Wdse) embedded within the non-extractible cell wall matrix (Wdsne), this
can be included as:
Water potential
Diurnal leaf water potential (Ψleaf) was measured with a Scholander-type pressure chamber (Mk 3005, Soil Moisture Equip-
Wds
Vfs
Dt =
Wds Wdse + Wdsne
=
Vfs
Vfs
TREE PHYSIOLOGY VOLUME 24, 2004
(2)
DROUGHT EFFECTS ON WOOD DENSITY AND ANATOMY IN EUCALYPTUS
Once D t had been measured, all extractives in the wood samples
were removed. The freeze-dried wood samples were placed in
plastic vials with screw-top lids that had one small perforation
to act as a pressure release. The vials were filled with 100%
ethanol and incubated for 5 days in a water bath held at 70 °C.
Each day, the ethanol was discarded and replaced with fresh
ethanol. After 5 days, the ethanol was replaced with 50% (v/v)
ethanol in water and the vials were agitated for 2 days. The
50% ethanol solution was then discarded and replaced with
100% distilled water and the vials incubated for 5 days in a
water bath at 90 °C. The distilled water was replaced each day.
We then measured Vfs as described previously. Samples were
freeze-dried and their Wdsne recorded. Residual density (D r )
was calculated as:
Dr =
Wdsne
Vfs
(3)
1297
with the factorial ANOVA algorithm of the STATISTICA software package (Version 6, StatSoft, Tulsa, OK). Tukey’s HSD
post-hoc test made individual comparisons between species/
watering combinations. The effect of water-limitation within
species was tested with a Students t-test. Where data were
nonhomogeneous, data were log- or square root-transformed,
with data presented as non-transformed means. Significance
level was set at 0.05.
Results
Preliminary harvest
Mean stem diameter and Dt at Week 0 were respectively:
1.73 mm and 390 kg m –3 for E. grandis, 1.22 mm and 280 kg
m –3 for E. sideroxylon and 1.26 mm and 320 kg m –3 for E. occidentalis. Diameter was significantly greater in E. grandis than
in the other species, but Dt did not differ between species.
Wood anatomy
Water potential and growth
After the basal stem segment was removed to measure wood
density, a 1-cm sample was taken from the remaining stem to
examine wood anatomy. This sample was immediately fixed
in 4% paraformaldehyde in 0.1 M phosphate buffer and dehydrated in an ethanol series before embedding it in LR White
resin. Sections of each sample (1 µm thick) were made with a
Leica microtome with glass knives and stained with 0.5%
(v/v) toluidine blue for 5 s. The sections were mounted on
glass slides with a cover slip. Microscopic digital images were
captured at 10× magnification with an Olympus BX60 compound microscope (Olympus, Tokyo, Japan) interfaced with a
Jenoptik ProRes C14 digital camera (Jenoptik, Jena, Germany). Digital images were taken on a radial path from the
pith to the bark. All image enhancement and analyses were
performed with Image Pro Plus Version 3.0 (Media Cybernetics, Silver Spring, MD). Binary versions of these images were
created. The optimal threshold setting for each binary image
was selected visually for each image as described by Hacke et
al. (2001). The area of the pith was measured with an Image
Pro Plus area measuring function. A custom written Image Pro
macro, designed to detect xylem vessels, estimated the number
and dimensions of xylem vessels in each binary image. The
data were weighted mathematically to reflect the proportion of
the stem that an image represented, determining the proportion
of stem area taken up by vessels, vessel frequency and mean
vessel lumen area. The conduit efficiency value (CEV) was
calculated as the sum of all vessel radii increased to the fourth
power extrapolated up to the whole stem area divided by the
leaf area supplied by this stem. The CEV is an estimate of the
efficiency with which a stem supplies a unit leaf area; a low
CEV indicates that the stem has lower hydraulic conductivity.
Additionally, Image Pro Plus was used to make a single transect of vessel diameters from the edge of the pith to the cambium for each stem. These measurements were made on color
images, not the binary images.
There was no difference in the degree of water stress experienced by each species based on predawn Ψleaf (Factorial
ANOVA) (Table 1). Predawn Ψleaf was lower in water-limited
plants than in well-watered plants. Seedling height was reduced 17–37% by the water-limitation treatment (Table 1).
Stem diameter was also reduced by water-limitation in all species, with the greatest reduction in E. grandis (39%) and
E. sideroxylon (38%) and least in E. occidentalis (10%). Seedling dry mass was significantly affected by the water-limitation treatment, with reductions ranging from 51–70%. There
was no significant effect of watering treatment or species on
RGR, but there was a trend for RGR to be lower in water-limited plants than in well-watered plants.
Statistical analyses
The effects of species and watering treatment were analyzed
Total and residual wood density
Compared with well-watered seedlings, Dt was higher in water-limited seedlings of E. grandis and E. sideroxylon, but not
E. occidentalis (Figure 1A). Eucalyptus sideroxylon had higher
D t than E. grandis and E. occidentalis (P < 0.01) (Figure 1).
When ethanol and water-soluble extractives were removed,
there was no difference in residual wood density (Dr) between
the well-watered and water-limited E. sideroxylon plants (Figure 1B); however, water-limitation had a significant effect on
the D r of E. grandis (Figure 1B). The decline in D r of water-limited plants was, by definition (see Equation 2), a result
of the 10−20% decrease in dry mass of the residual wood
samples (Figure 2) because there was no difference in Vfs when
D t and Dr were measured (data not shown). Wood density was
higher in E. sideroxylon than in E. grandis and E. occidentalis
even when the soluble components were removed (Figure 1).
Wood anatomy
Vessels comprised a greater proportion of wood cross-sectional area in E. grandis and E. occidentalis than in E. sideroxylon (Table 2). This was because of a greater number of
vessels per unit area of wood in E. occidentalis and a trend for
larger vessel lumen areas in E. grandis (Table 2). Water-limita-
TREE PHYSIOLOGY ONLINE at http://heronpublishing.com
1298
SEARSON, THOMAS, MONTAGU AND CONROY
Table 1. Predawn leaf water potential (Ψleaf ), plant dimensions, biomass and relative growth rate (RGR) of well-watered (WW) and water-limited
(WL) E. grandis, E. sideroxylon and E. occidentalis seedlings. Data are the mean of five replicates (± standard error). Significance values are indicated by asterisks: * = P < 0.05; ** = P < 0.01; *** = P < 0.001; and ns = not significant. Letters denote factors in two-way ANOVA: S = species;
W = water; and S × W = interaction.
Parameter
E. grandis
E. sideroxylon
E. occidentalis
Significance
WW
WL
WW
WL
WW
WL
S
W
S×W
Predawn Ψleaf (MPa)
– 0.37
(0.04)
–1.36
(0.10)
– 0.39
(0.08)
–1.61
(0.12)
– 0.62
(0.13)
–1.30
(0.14)
ns
***
ns
Height (cm)
24.60
(2.89)
18.00
(2.28)
31.50
(3.80)
26.30
(3.69)
37.10
(7.49)
23.70
(4.24)
***
**
ns
Stem diameter (mm)
8.38
(0.57)
5.07
(0.13)
7.61
(0.34)
4.72
(0.36)
5.87
(0.34)
5.28
(0.38)
ns
***
*
Dry mass (g)
49.38
(9.15)
15.11
(2.08)
43.77
(5.31)
14.19
(1.50)
25.98
(11.96)
12.84
(2.72)
***
***
ns
RGR (g g–1 day–1)
0.028
(0.001)
0.020
(0.005)
0.042
(0.005)
0.030
(0.004)
0.034
(0.010)
0.031
(0.006)
ns
ns
ns
tion did not affect the overall proportion of wood taken up by
vessels. The water-limitation treatment did however reduce
mean vessel lumen area by 7–30 % (Table 2). This probably
did not result in a change in the proportion of wood taken up by
vessels because there was a trend for vessel frequency to increase in water-limited stems (Table 2). The drought cycles did
not result in a clear pattern of vessel diameters increasing
when seedlings were re-watered and decreasing as soil water
was depleted (Figure 3). There was a significant negative relationship between vessel lumen fraction and wood density for
well-watered seedlings; however, this relationship broke down
Figure 1. Wood density measured on a volumetric basis in well-watered (filled bars) and water-limited (open bars) E. grandis, E. sideroxylon and E. occidentalis seedlings. (A) Samples before soluble components were extracted (Dt ). (B) Samples after soluble components
were extracted (Dr ). The results of within species t-tests for the effect
of water-limitation are shown for each species. Data are the mean of
five replicates ± SE.
Figure 2. Reduction in stem sample dry mass of well-watered (filled
bars) and water-limited (open bars) E. grandis, E. sideroxylon and
E. occidentalis seedlings after the extraction of soluble wood components. The results of within species t-tests for the effect of water-limitation are shown for each species. Data are the mean of five replicates
± SE.
TREE PHYSIOLOGY VOLUME 24, 2004
DROUGHT EFFECTS ON WOOD DENSITY AND ANATOMY IN EUCALYPTUS
1299
Table 2. Stem anatomy (vessels and pith fraction, vessel frequency and mean vessel lumen area) and conduit efficiency value (CEV) of well-watered (WW) and water-limited (WL) E. grandis, E. sideroxylon and E. occidentalis seedlings. Microscopy samples were taken from the base of the
stem samples measured for wood density. Data are the mean of five replicates (± standard error). Significance values are indicated by asterisks: * =
P < 0.05; ** = P < 0.01; and ns = not significant. Letters denote factors in two-way ANOVA: S = species; W = water; and S × W = interaction.
Parameter
E. grandis
E. sideroxylon
E. occidentalis
Significance
WW
WL
WW
WL
WW
WL
S
W
S×W
Vessel lumen fraction (%)
11
(0.08)
14
(2.0)
7
(1.0)
8
(0.5)
9
(0.5)
10
(0.2)
*
ns
ns
Vessel frequency (no. mm –2 )
156
(5.50)
201
(18.5)
116
(22.1)
137
(10.3)
175
(17.5)
224
(58.0)
*
ns
ns
Vessel lumen area (µm2 )
778
(46.4)
549
(28.3)
629
(62.6)
586
(48.0)
623
(55.0)
508
(100)
ns
*
ns
Pith fraction (%)
1.19
(0.54)
5.43
(2.24)
10.1
(2.14)
19.6
(6.65)
4.00
(2.19)
7.23
(3.43)
**
*
ns
CEV (m2 × 10 –10 )
60.6
(13.6)
39.2
(12.5)
45.0
(13.7)
24.8
(10.3)
82.6
(22.0)
49.6
(11.0)
ns
*
ns
in the water-limited plants (Figure 4). The proportion of the
stem occupied by pith was highest in E. sideroxylon, and
within each species, pith occupied a greater proportion of the
stem in the water-limited plants than in the well-watered plants
(Table 2). Water-limitation significantly reduced CEV, and
this was lowest in E. sideroxylon. There was no overall effect
of species on CEV (Table 2).
the pith was slightly less than 200 –300 kg m –3 given that some
Relationship between wood density and diurnal variation in
Ψleaf
There was a significant positive relationship between Dr and
∆Ψleaf for the well-watered plants (Figure 5), but not for the
water-limited plants. There was no correlation between ∆Ψleaf
and CEV (data not shown).
Discussion
We tested the hypotheses that water-limited Eucalyptus seedlings have higher wood density than well-watered seedlings
when grown in a common environment and that there is a positive correlation between wood density and ∆Ψleaf. We found
that D r of E. sideroxylon and E. occidentalis was unaffected by
water-limitation, whereas E. grandis produced higher Dr in response to water-limitation. There was a positive correlation
between D r and ∆Ψleaf for the well-watered plants but not for
the water-limited plants.
Effect of water limitation on wood density
Based on measurements of Dr, which is a coarse measure of
wood anatomy, we found that water limitation modified the
stem wood anatomy of E. grandis but had no effect on that of
E. sideroxylon and E. occidentalis. Differences in the effects of
water limitation on Dr across the species may have been associated with a dilution effect of the pith. In the transverse section, the pith appeared as a small group of cells about 10 –15
cells in the x/y dimension with thin walls and large lumens.
From our preliminary harvest, we estimated that the density of
Figure 3. Transect of vessel diameter from pith to cambium of (A)
E. grandis, well-watered (䊉), water-limited (䊊); (B) E. sideroxylon,
well-watered (䊏), water-limited (䊐); and (C) E. occidentalis, wellwatered (䉱), water-limited (䉭). Distance from pith is expressed relative to the total distance from pith to cambium. For each species, data
are a combination of five replicates from each watering treatment.
TREE PHYSIOLOGY ONLINE at http://heronpublishing.com
1300
SEARSON, THOMAS, MONTAGU AND CONROY
Figure 4. Relationship between mean vessel lumen fraction and wood
density (Dr ) of E. grandis, well-watered (䊉), water-limited (䊊); E. sideroxylon, well-watered (䊏), water-limited (䊐); and E. occidentalis;
well-watered (䉱), water-limited (䉭). Line of best fit is through the
well-watered sample data. Density was measured on wood samples
with soluble components removed.
secondary xylem would have surrounded the pith at this early
stage and that these wood samples probably contained extractives. Wood with a greater proportion of pith has its overall Dr
‘diluted’ by the presence of these low tissue-density cells. The
Figure 5. Relationship between wood density (Dr ) and diurnal variation in leaf water potentail (∆Ψleaf ) of E. grandis, well-watered (䊉),
water-limited (䊊); E. sideroxylon, well-watered (䊏), water-limited
(䊐); and E. occidentalis, well-watered (䉱), water-limited (䉭). Density was measured on wood samples with soluble components removed.
pith occupied a greater proportion of the stem in water-limited
plants than in well-watered plants (Table 2), and was produced
when the seedlings were growing under well-watered conditions before the start of the experimental treatments. The effect
of water on the proportion of the stem taken up by pith was
therefore a consequence of less wood being produced around
the pith in the water-limited plants during the experiment
rather than water-limitation causing an increase in pith diameter. Without this artifact of the pith on D r, there may have been
a clearer difference in D r between the well-watered and water-limited plants. Also, E. sideroxylon would probably have
had even greater D r, because this species developed a larger
pith than the other two species and so its negative effect on
wood density was greater. The increase in Dr in water-limited
E. grandis seedlings may have been associated with the trend
for this species to be shorter and have a larger stem diameter
than the other species (Table 2). The effect of the pith on the D r
of water-limited plants may also explain why there was no relationship between the proportion of wood occupied by vessels and D r (Figure 4).
Effect of water limitation on wood anatomy
Image analysis of the wood indicated an effect of water limitation on wood anatomy (Table 2). Differences in vessel diameter in response to water-limitation were evident from pith to
cambium for E. grandis and E. sideroxylon (Figure 3); however, this difference was consistent across the transect and was
unrelated to the changing soil water conditions through the
four drought cycles. The lack of an increasing and decreasing
pattern in vessel diameters may have been related to the time
taken for a xylem vessel to develop. The longest drought cycle
was about 4 weeks with shortest cycle being closer to 2 weeks.
Ridoutt and Sands (1994) estimated that fiber maturation (cell
division, enlargement and wall deposition) took an average of
42.8 days in 6-year-old field-grown E. globulus, and they observed that vessel development was faster than fiber development. From this information, we estimate that the time during
which morphology of individual vessels could be affected during our experiment was less than 43 days. Assuming that
vessel development was twice as fast as fiber development,
various degrees of water limitation rather than a single stressful effect may have influenced the environment of the cambium during vessel maturation. Further investigations should
include water-limitation treatments lasting for more than 1
month.
Despite uncertainty about the effect of the pith on wood density, our anatomical data showed that it is difficult to change
the proportion of wood occupied by vessels in water-limited
Eucalyptus seedlings. Mean vessel lumen area declined and
there was a trend for increased vessel frequency in the water-limited plants of each species (Table 2). The net effect of
these changes was that there was no significant difference in
the vessel lumen fraction between well-watered and waterlimited seedlings (Table 2). Atwell et al. (2000) also reported
that the proportion of the stem occupied by vessels was conserved in water-limited E. teretecoirnis Sm. because of an increase in vessel frequency despite a decline in mean vessel
TREE PHYSIOLOGY VOLUME 24, 2004
DROUGHT EFFECTS ON WOOD DENSITY AND ANATOMY IN EUCALYPTUS
cross-sectional area. A similar pattern was observed by February et al. (1995) in a comparative study of well-watered and
water-limited E. grandis and E. grandis × camaldulensis and
E. grandis × nitens cuttings. Outside the Eucalyptus genus,
Lovisolo and Schubert (1998) reported a similar effect in
well-watered and water-limited Vitis vinifera L. Apart from
water-limitation, Gartner et al. (2003) found that the proportion of stem occupied by vessels was conserved in the experimentally induced tension wood of Quercus ilex L. seedlings
because of an increase in vessel frequency despite a decline in
mean vessel diameter. Vessel diameter decreased whereas frequency increased in downwardly oriented V. vinifera plants
(Lovisolo et al. 2002). The same effect on vessel diameter and
frequency was shown in a transgenic Populus hybrid (Populus
tremula L. × Populus tremuloides Michx.) that over-produced
indoleacetic acid (Tuominen et al. 1995). This hormone may
have caused decreased vessel diameter and increased vessel
frequency in both our study and in studies by other workers.
Increases in vessel frequency when vessel diameters decrease
in response to stress could be viewed as a compensatory response to loss of water transport capacity as a result of reductions in vessel diameter; however, increased vessel frequency
may not restore stem hydraulic conductivity to its value prior
to the application of stress because of the fourth-power relationship between vessel radius and conductivity. In our experiment, CEV was significantly lower in the water-limited plants
compared with the well-watered plants, indicating that the increase in vessel frequency did not restore the efficiency of xylem water supply to the leaves to pre-stress values.
Relationship between Dr and diurnal Ψleaf
Meinzer (2003) found a positive relationship between Dr and
∆Ψleaf in a range of tree species and suggested that it was associated with the low hydraulic conductivity of the high-density
wood. Our well-watered seedlings also showed a positive correlation between Dr and ∆Ψleaf ; however, Dr and ∆Ψleaf were
not correlated under water-limited conditions (Figure 5). The
positive relationship between D r and ∆Ψleaf under well-watered conditions implies that hydraulic conductivity is more
limited in high density wood than in lower density wood because more energy is required to transport water from the soil
to the leaves. The breakdown of this relationship in the water-limited seedlings probably occurred because of stomatal
closure (cf. Searson et al. 2004). Stomatal closure regulates
water loss from the leaf and hence restricts ∆Ψleaf, making
∆Ψleaf less a function of stem hydraulic conductivity.
In our study, the primary driver of the relationship between
Dr and ∆Ψleaf was E. sideroxylon, which had the highest wood
density of the three species and the lowest mean vessel area
and vessel frequency. As a result of these differences in xylem
anatomy, CEV was lowest in E. sideroxylon (Table 2). This
trend for lower CEV in E. sideroxylon than in the other species
supports our finding that ∆Ψleaf is greater in trees with higher
density wood. We cannot explain why the CEV of E. sideroxylon was not significantly different from the other two species, or why there was no relationship between CEV and
1301
∆Ψleaf, given the relationship between Dr and ∆Ψleaf. Measurements of the hydraulic conductivity of the stem segment used
to measure D r may help resolve this issue.
Conclusion
We were unable to establish unequivocally that water limitation increases wood density in the three Eucalyptus species
studied. The stem wood of water-limited Eucalyptus seedlings
had similar Dr to that of well-watered seedlings. Although
changes in vessel diameter and frequency were observed in the
water-limited plants, the proportion of the stem cross-sectional area allocated to vessels remained relatively stable between well-watered and water-limited plants, which may explain why differences in D r were not large. However, the
apparent lack of an effect of water limitation on Dr may also
have been associated with the dominance of the pith in the
stems of water-limited plants or because of the relatively short
duration of the drought treatments. Water may have to be withheld continuously for longer than one month to cause a change
in wood anatomy sufficient to cause a significant change in
wood density in response to water limitation. The positive correlation between Dr and ∆Ψleaf indicates that increasing wood
density results in decreasing hydraulic conductivity; however,
this method could not assess the hydraulic conductivity of the
water-limited plants because of stomatal regulation of Ψleaf.
Acknowledgments
We greatly thank Liz Darley (Centre for Horticulture and Plant Sciences - CHAPS) for assistance with microscopy and Mark Emanuel
(UWS) for freeze-drying the wood samples. Michael Roderick helped
to focus and clarify this paper. We also thank Penny Searson for reading and commenting on early drafts of this paper and Philip Groom for
proofreading the final versions. While carrying out this research Matthew J. Searson was supported by an Australian Postgraduate Award
and a CHAPS top-up scholarship.
References
Atwell, B.J., P.E. Kreidemann and C. Turnbull. 1999. Plants in action.
MacMillan, Melbourne, 664 p.
Atwell, B.J., S. Barlow, G. Rogers and J.P. Conroy. 2000. Farm trees
for the 1990s and beyond. Rural Industries Research and Development Corporation, Canberra, 60 p.
Bamber, R.K. and J. Burley. 1983. The wood properties of radiata
pine. Commonwealth Agricultural Bureau, Slough, U.K., 84 p.
Barbour, M.M. and D. Whitehead. 2003. A demonstration of the theoretical prediction that sap velocity is related to wood density in the
conifer Dacrydium cupressinum. New Phytol. 158:477– 488.
Beadle, C.L., P.W. Banham, D. Worledge, S.L. Russell, S.J. Hetherington, J.L. Honeysett and D.A. White. 2001. Effect of irrigation
on growth and fibre quality of Eucalyptus globulus and Eucalyptus
nitens. Appita J. 54:144 –147.
Catchpoole, S.J., G. Downes and S.M. Read. 2000. The effect of salt
on wood and fibre formation in eucalypts. Report No. UM-18A.
Rural Industries Research and Development Corporation, Canberra, 55 p.
February, E.C., W.D. Stock, W.J. Bond and D. Le Roux. 1995. Relationships between water availability and selected vessel characteristics in Eucalyptus grandis and two hybrids. IAWA J. 16:269–276.
TREE PHYSIOLOGY ONLINE at http://heronpublishing.com
1302
SEARSON, THOMAS, MONTAGU AND CONROY
Gartner, B.L., J. Roy and R. Huc. 2003. Effects of tension wood on
specific conductivity and vulnerability to embolism of Quercus ilex
seedlings grown at two atmospheric CO2 concentrations. Tree Physiol. 23:387–395.
Hacke, U.G., J.S. Sperry, W.T. Pockman, S.D. Davis and K.A. McCulloh. 2001. Trends in wood density and structure are linked to prevention of xylem implosion by negative pressure. Oecologia 126:
457– 461.
Lovisolo, C. and A. Schubert. 1998. Effects of water stress on vessel
size and xylem hydraulic conductivity in Vitis vinifera L. J. Exp.
Bot. 49:693–700.
Lovisolo, C., A. Schubert and C. Sorce. 2002. Are xylem radial development and hydraulic conductivity in downwardly-growing grapevine shoots influenced by perturbed auxin metabolism? New
Phytol. 156:65–74.
Macfarlane, C. and M.A. Adams. 1998. δ13 C of wood in growth-rings
indicates cambial activity of drought-stressed trees of Eucalyptus
globulus. Funct. Ecol. 12:655– 664.
Meinzer, F.C. 2003. Functional convergence in plant responses to the
environment. Oecologia 134:1–11.
Ona, T., T. Sonoda, K. Ito and M. Shibata. 1997. Relationship between various extracted basic densities and wood chemical components in Eucalyptus camaldulensis. Wood Sci. Technol. 31:205–216.
Ona, T., T. Sonoda, K. Ito and M. Shibata. 1998. Relations between
various extracted basic densities and wood chemical components
in Eucalyptus globulus. J. Wood Sci. 44:165–168.
Ridoutt, B.G. and R. Sands. 1994. Quantification of the processes of
secondary xylem fibre development in Eucalyptus globulus at two
height levels. IAWA J. 15:417– 424.
Searson, M.J., D.S. Thomas, K.D. Montagu and J.P. Conroy. 2004.
Leaf water use efficiency differs between Eucalyptus species from
contrasting rainfall environments. Funct. Plant Biol. 31:441– 450.
Sefton, C.A., K.D. Montagu, B.J. Atwell and J.P. Conroy. 2002. Anatomical variation in juvenile eucalypt leaves accounts for differences in specific leaf area and CO2 assimilation rates. Aust. J. Bot.
50:301–310.
Stratton, L., G. Goldstein and F.C. Meinzer. 2000. Stem water storage
capacity and efficiency of water transport: their functional significance in Hawaiian dry forest. Plant Cell Environ. 23:99 –106.
Thomas, D.S., K.D. Montagu and J.P. Conroy. 2004. Changes in
eucalypt wood density due to temperature—the physiological link
between water viscosity and wood anatomy. For. Ecol. Manage.
193:157–165.
Tuominen, H., F. Sitbon, C. Jacobsson, G. Sandberg, O. Olsson and
B. Sundberg. 1995. Altered growth and wood characteristics in
transgenic hybrid Aspen expressing Agrobacterium tumefaciens
T-DNA indoleacetic acid–biosynthetic genes. Plant Physiol. 109:
1179 –1189.
Tyree, M.T. and F.W. Ewers. 1991. The hydraulic architecture of trees
and other woody plants. New Phytol. 119:345–360.
Wimmer, R., G. Downes and R. Evans. 2002. High-resolution analysis of radial growth and wood density in Eucalyptus nitens, grown
under different irrigation regimes. Ann. For. Sci. 59:519 –524.
TREE PHYSIOLOGY VOLUME 24, 2004