Tree Physiology 27, 251–260
© 2007 Heron Publishing—Victoria, Canada
Temperature effects on wood anatomy, wood density, photosynthesis
and biomass partitioning of Eucalyptus grandis seedlings
D. S. THOMAS,1,2,3 K. D. MONTAGU 4,5 and J. P. CONROY1
1
Centre for Horticulture and Plant Sciences, University of Western Sydney, Locked Bag 1797 South Penrith, NSW 1797, Australia
2
Present address: Forests NSW, Plantation Improvement, Land management and Technical Services, PO Box J19 Coffs Harbour Jetty, NSW 2450,
Australia
3
Corresponding author ([email protected])
4
State Forests of NSW, Research and Development Division, PO Box 100 Beecroft, NSW 2119, Australia
5
Present address: Cooperative Research Centre for Irrigation Futures, c/o School of Environment and Agriculture, University of Western Sydney,
Locked bag 1797 South Penrith, NSW 1797, Australia
Received January 20, 2006; accepted March 11, 2006; published online November 1, 2006
Summary Wood density, a gross measure of wood mass relative to wood volume, is important in our understanding of
stem volume growth, carbon sequestration and leaf water supply. Disproportionate changes in the ratio of wood mass to volume may occur at the level of the whole stem or the individual
cell. In general, there is a positive relationship between temperature and wood density of eucalypts, although this relationship
has broken down in recent years with wood density decreasing
as global temperatures have risen. To determine the anatomical
causes of the effects of temperature on wood density, Eucalyptus grandis W. Hill ex Maiden seedlings were grown in controlled-environment cabinets at constant temperatures from 10
to 35 °C. The 20% increase in wood density of E. grandis seedlings grown at the higher temperatures was variously related to
a 40% reduction in lumen area of xylem vessels, a 10% reduction in the lumen area of fiber cells and a 10% increase in fiber
cell wall thickness. The changes in cell wall characteristics
could be considered analogous to changes in carbon supply.
Lumen area of fiber cells declined because of reduced fiber cell
expansion and increased fiber cell wall thickening. Fiber cell
wall thickness was positively related to canopy CO2 assimilation rate (Ac), which increased 26-fold because of a 24-fold increase in leaf area and a doubling in leaf CO2 assimilation rate
from minima at 10 and 35 °C to maxima at 25 and 30 °C. Increased Ac increased seedling volume, biomass and wood density; but increased wood density was also related to a shift in
partitioning of seedling biomass from roots to stems as temperature increased.
Keywords: biomass partitioning, CO2 assimilation, fiber cell,
photosynthesis, xylem vessel.
Introduction
Density is an important attribute of wood that contributes to
the quality and therefore economic value of timber (MacDon-
ald and Hubert 2002). Wood density, a gross measure of wood
biomass relative to wood volume, is an integration of the processes involved in stem volume growth (i.e., cell division and
cell enlargement) and the processes involved in biomass accumulation (i.e., cell wall thickening of the existing and newly
formed cells). The arrangement of biomass in stems and the remaining voids has important implications for both solute
movement (Hacke et al. 2001, Roderick and Berry 2001) and
inferring historical late-summer temperatures (Briffa et al.
1998).
Increased growth temperatures can be related to increased
wood density of mature Eucalyptus dunnii Maiden trees
(Muneri et al. 2004), but the anatomical causes responsible for
the changes in wood density of eucalypts are unknown. Furthermore, seasonal variation in wood density has been used as
a surrogate for temperature in interpreting historical climates.
However, in recent years, the general positive relationship between temperature and wood density has broken down with
wood density decreasing as global temperatures have risen
(Briffa et al. 1998). This divergence is unusual because partitioning of biomass to shoots often increases as temperature
rises (Wardlaw 1979, Bruhn et al. 2000, Weih and Karlsson
2001), a factor expected to increase wood density.
Plant biomass accumulation typically increases with increasing temperature before declining at supra-optimal temperatures. An increase in stem biomass would be expected to
increase the biomass in fiber cell walls and, if this process occurs faster than increases in stem volume, then wood density
will increase—a situation that can occur when carbon supply
is increased (Richardson and Dinwoodie 1960, Larson 1964,
Richardson 1964, Creber and Chaloner 1984, Conroy et al.
1990, Lindström 1996, Deleuze and Houllier 1998, Barber et
al. 2000). However, when stem volume increases as fast as, or
faster than, stem biomass then wood density may remain stable or decline (Telewski et al. 1999, Ceulemans et al. 2002,
Thomas et al. 2006). Therefore the independent effects of tem-
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THOMAS, MONTAGU AND CONROY
perature on the processes of biomass accumulation of the stem
and stem volume growth will determine the impact of
temperature on wood density.
Stem volume is a function of the volumes of the various cell
types (Kramer and Kozlowski 1979, Malan and Hoon 1992,
Zhang and Zhong 1992, Zobel and Jett 1995, Denne and Hale
1999). If we assume that the stem comprises only fiber cells
and xylem vessels, then the relative volumes of cell wall material and cell lumen will affect wood density (Roderick and
Berry 2001). Temperature affects the relationship between fiber cell wall thickness and fiber cell lumen, and the number
and size distribution of xylem vessels in seedlings and mature
trees (Larson 1964, Richardson 1964, Roderick and Berry
2001, Thomas et al. 2004). In a simple model of wood production in gymnosperms, Deleuze and Houllier (1998) assumed
temperature was the most limiting factor for initial fiber cell
division. In their model, higher temperatures result in increased cell production, but the rate of cell production declines
at supra-optimal temperatures (Deleuze and Houllier 1998).
These modeling results are supported by the finding that
cambial activity, measured as rate of cell division in larch
(Larix siberica Ldb.) and Scots pine (Pinus sylvestris L.), increased with increasing temperature, and then declined when
mean temperatures were greater than about 20 °C (Antonova
and Stasova 1993, 1997); the effects of temperature on stem
biomass accumulation and wood density were not determined
in these studies.
To test the hypothesis that Eucalyptus grandis W. Hill ex
Maiden seedlings have higher wood density when grown at
higher temperature, we grew seedlings in controlled-environment cabinets at constant temperatures from 10 to 35 °C. Specifically, we determined if the positive effect of temperature on
wood density is correlated with an overall increase in seedling
carbon supply and biomass allocated to stems. Additionally,
we assessed if the positive correlation of temperature with
stem wood density is associated with enhanced production of
denser thick-walled cells, and decreased lumen areas of the
fiber cells and xylem vessels.
Materials and methods
Plant culture
Soil was collected from the A horizon at Belanglo State Forest
near Moss Vale, NSW (152°13′ E, 34°30′ S). The soil is
podsolic derived from Triassic shales and sandstones. Total N
is less than 0.09% and exchangeable Ca less than 0.4me%
(Turner 1982). Batches of soil were mixed with CaCO3 (7 g
kg –1) and MgCO3 (1.8 g kg –1) to adjust the pH to 6.7 (1:5 w/v
in 0.01 M CaCl2). Phosphorus as CaHPO4 was added to the
soil, which had an available P concentration of less than 1 mg
kg –1 dry soil (Bray method 1, Bray and Kurtz 1954), at the
rate of 1000 mg P kg –1 dry soil. Basal nutrients were added to
the soil (mg kg –1 dry soil): K (90 + 360); B (5); Cu (5); Zn
(10); Mo (0.1); Mn (50); and Fe (50) as K2SO4 + K2CO3;
H3BO3; CuSO4; ZnSO4; Na2MoO4; MnSO4; and FeSO4. Nitrogen was added weekly at 67 mg kg –1 as KNO3, Ca(NO3)2,
Mg(NO3)2, (NH4)2SO4 and NH4NO3 in the ratio (by weight of
salts) of 1:2:1:1:1.
Eucalyptus grandis seeds were germinated in a 1:1 (v/v)
mix of perlite and amended soil at 25 °C in a Eurotherm
Chessell 392 plant growth chamber (Thermoline, Australia)
located at the University of Western Sydney, Hawkesbury
campus, Richmond, NSW (150°45′ E, 33° 36′ S). The vapor
pressure deficit in the chamber was maintained at 1 kPa. Photosynthetic photon flux (PPF) was maintained at 1000 µmol
m – 2 s – 1 at mid-canopy height with 1000-W metal halide lamps
during the 12-h photoperiod.
When seedlings were 6 weeks old they were 2 ± 0.02 (SE)
cm high, had a stem diameter at soil height of 0.4 ± 0.005 mm,
a leaf area determined by a Delta T leaf area meter (Burwell,
Cambridge, U.K.) of 3.5 ± 0.36 cm2 and biomass after drying
at 70 °C for 48 h of 0.04 ± 0.004 g. At this time, 75 seedlings
were each transplanted to a 6.9-l pot (PVC pipe: diameter
15 cm, height 39 cm) containing 7.5 kg of air dried, amended
soil. This allowed for five replicate seedlings in each of five
growth temperature treatments at each of three harvests during
the growth period. The growth temperatures were constant
day/night temperatures of 10, 20, 25, 30 and 35 °C with vapor
pressure deficit maintained at 1 kPa. Harvests were made
when the seedlings were aged 11 weeks (Harvest 1), 15 weeks
(Harvest 2) and 19 weeks (Harvest 3).
Seedlings were maintained at 25 °C for one week after
transplanting, then allocated to one of the five temperature
treatments in one of five replicated growth chambers. To reduce temperature shock to the seedlings, the temperature
within the growth cabinets was altered in daily steps no greater
than 1 °C until the desired constant day/night temperatures
were achieved. This meant that when seedlings were harvested, the seedlings grown at 30 and 20 °C had experienced
these constant temperatures for about 1 week less than seedlings maintained at 25 °C, and seedlings grown at 35 and 10 °C
experienced these constant temperatures for about 2 weeks
less than seedlings maintained at 25 °C. Therefore, although
the comparisons were made among seedlings of similar age
(11 weeks at Harvest 1, 15 weeks at Harvest 2 and 19 weeks at
Harvest 3), the seedlings were exposed to the treatment temperatures for different periods of time depending on the treatment.
During the period from seedling transplanting to harvest,
seedlings were randomly repositioned within a growth chamber on a weekly basis. Growth chambers were randomly reallocated to different treatments every 2 weeks. Temperature
and vapor pressure deficit inside the growth chambers stabilized to the new conditions within 2 h of relocating the seedlings. The seedlings were irrigated daily throughout the
experiment.
Determination of CO2 assimilation rate
Leaf CO2 assimilation rate (Al) was measured on the youngest
fully expanded leaf between Harvests 2 and 3 when the seedlings were 18 weeks old. Measurements were made with an
LI-6400 portable photosynthesis system (Li-Cor, Lincoln,
NE) equipped with a light source (6400-02B LED, Li-Cor).
TREE PHYSIOLOGY VOLUME 27, 2007
TEMPERATURE EFFECTS ON EUCALYPT GROWTH AND WOOD DENSITY
Measurements of Al were completed between 0900 and 1500 h
as it had previously been determined that Al showed little diurnal variation during this period. Measurements were made at
the seedlings’ growth temperature and at a PPF of 1200 µmol
m – 2 s –1 to ensure the response was fully light saturated. Ambient CO2 was maintained at 360 µmol mol –1 during measurements of Al.
Canopy CO2 assimilation capacity (Ac) was calculated by
Equation 1 which is based on equations of gross plant photosynthetic production (Atwell et al. 1999):
Ac = Al(1 – exp(–kSLAWl))
(1)
where the extinction coefficient, k, was assumed to be 0.5
(Atwell et al. 1999), and values of specific leaf area (SLA) and
dry biomass of leaves (Wl) at the final harvest were used. Calculation of Ac by Equation 1 was used as a representation of the
seedlings’ carbon source strength at the final harvest, but it is
acknowledged that this approach assumes Al did not vary either over time or within the seedling, and does not account for
carbon losses through respiration of the various organs.
Seedling growth and harvests
Seedling height from the stem–root interface to the apical
meristem was measured every 2 weeks. At each harvest, stem
diameter over bark and wood diameter, i.e., stem diameter under bark, were measured at the first internode. At Harvest 3
only, the next 5 mm of stem wood material was collected for
microscopic examination of stem anatomy, and the next
20–40 mm of stem wood material was collected to measure
stem wood density. The volume of the entire main stem wood
was calculated from seedling wood diameter at the first
internode and height assuming a conical stem form.
At each harvest, the areas of the two youngest fully expanded leaves on the main stem were measured with a Delta T
leaf area meter (Burwell, Cambridge, United Kingdom) before determining their dry mass. At the final harvest, the thickness of the youngest fully expanded leaves was measured with
electronic callipers. The number and area of the remaining
leaves were measured and their dry mass was determined after
drying at 70 °C for 48 h. Specific leaf area (SLA) and mean
area of a leaf were calculated from these data. The mass of the
remaining main stem, branches and washed roots was determined after drying at 70 °C for 48 h and seedling biomass was
calculated after inclusion of the sample used to determine
wood density. Leaf area ratio (LAR) was calculated from these
data as the ratio of seedling leaf area to seedling biomass
(cm2 g –1).
Wood density was calculated for the 20–40 mm stem segment as dry mass/fresh volume. Fresh wood volume of the
sample was measured by immersing the sample in a beaker of
distilled water of known mass on a balance sensitive to 0.1 mg
and observing the change in balance reading. The mass of water displaced in grams was taken to correspond closely to sample volume in cm3. Each sample was lanced with a probe and
placed just under the surface of the water. A clamp attached to
a retort stand adjacent to the breaker was used to support the
253
probe and minimize movement of the sample. Measurements
of volume were complete within 10 s of immersion to minimize entry of water into the cut ends of the wood sample. Mass
of the sample was then determined after drying for 48 h at
105 °C.
Anatomy
Stem samples were fixed in 2.5% paraformaldehyde in 0.1 M
Na-phosphate buffer (pH 7.0) for two days. Stem samples
were washed in 0.1 M Na-phosphate buffer (pH 7.0), dehydrated through an ethanol series and embedded in London
White resin. Four of the five replicates per treatment collected
at Harvest 3 were randomly selected for sectioning and anatomical observation. One µm thick sections of the sample were
cut with glass knives with a Leica RM2165 microtome (Leica
Microtomes, Nassloch, Germany). Mounted sections were
stained with 0.5% (v/v) toluidine blue for 5 s before the digital
images were captured with an Olympus BX60 compound microscope (Olympus corporation, Tokyo, Japan) interfaced
with an MTI 3CCD digital camera (DAGE MTI Inc., Michigan). All analyses of digital images were performed using Image Pro Plus (Media Cybernetics). The total cross-sectional
area of wood, and the area of pith within the wood was calculated. The number of fiber cells along radial sections (from the
pith to the cambium), average fiber cell size, fiber cell wall
thickness, the number of xylem vessels and the proportion of
stem cross-sectional area occupied by xylem vessels were also
determined. Over 75% of the growth in wood diameter and
over 95% of the growth in biomass of the seedling and stem
occurred in the final 8 weeks of growth, i.e., within the period
during which constant growth temperatures were maintained.
The anatomical data were weighted to reflect the proportion of
the stem’s entire cross-sectional area that the image represented. Thus, the microscopy comparisons largely reflect the
anatomical responses accountable for the treatment differences of plant production.
The density of xylem vessels per area of wood and the size
distribution of the xylem vessels were calculated. The diameters of fiber cell lumens were calculated from the difference
between fiber cell diameter and fiber cell wall thickness. The
cross-sectional areas of the fiber cell lumens (Fv) and of the fiber cell wall material (Fw) were calculated assuming the fiber
cells were circular. The ratio Fv:Fw is therefore a measure of
the amount of voids within a fiber cell in relation to the amount
of carbon deposited in the fiber cell walls.
Statistical analysis
Where variance in data was heterogeneous, arcsin or log transformation were used, with data presented as non-transformed
means. Effects of growth temperature were analyzed with the
analysis of variance (ANOVA) algorithm of STATISTICA
software (Version 6, StatSoft, Oklahoma, USA). Fishers LSD
test was used for comparisons between treatments. The multiple regression algorithm of STATISTICA was used to determine linear relationships between factors. The effect of plant
ontogeny on biomass partitioning relationships was examined
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THOMAS, MONTAGU AND CONROY
by analysis of covariance (ANCOVA) of log-transformed data.
The slopes of these relationships were used to explore the partitioning of biomass between different plant parts independent
of plant size (Hunt 1990).
Results
Partitioning of seedling biomass – growth temperature or
ontogeny?
We compared seedlings of a similar age but grown for periods
ranging between 10 and 12 weeks at constant temperature
(Harvest 3). Therefore, it is necessary to consider the effect of
plant ontogeny on the results. Seedlings grown at different
temperatures differed markedly in size; however, use of plants
of similar size but different age did not alter the effect of temperature on wood density, namely, wood density increased
with increasing growth temperature (Figure 1). Seedlings of
similar size were grown for either 7–8 or 10–12 weeks at constant temperature (i.e., Harvest 2 and Harvest 3). In these analyses, we compared the slower-growing seedlings grown at 10
and 35 °C at Harvest 3 with the faster-growing seedlings
grown at 20, 25 and 30 °C at Harvest 2. These groups of seedlings were of similar biomass and had similar numbers of
branches, indicating they were at an approximately similar developmental stage. We were unable to make a similar comparison between slower-growing seedlings at Harvest 2 with
faster-growing seedlings at Harvest 1 because these groups of
seedlings differed considerably in size and because of the rela-
Figure 1. Effects of growth temperature on: (a) wood density; (b) stem
biomass; and (c) stem wood volume in 19-week-old seedlings grown
at a constant temperature for the final 10–12 weeks (䊏). In (a) the effects of growth temperature on wood density are also shown for
15-week-old seedlings grown at a constant temperature for the final
7–8 weeks (䊐). Each value represents the mean and SE of five replicates.
tively short time the seedlings had grown at constant temperatures (between 4 and 6 weeks). Unfortunately, because Al,
which is required for Ac, was measured only at Week 18, i.e.,
one week before Harvest 3, and data on stem anatomy at Harvest 2 were not collected, we are able to compare the effects of
growth temperature on Al, Ac or stem anatomy only in seedlings of similar age but not in seedlings of similar size.
The allometric relationships for seedlings of all harvests
show that growth temperature influenced partitioning of seedling biomass between seedling organs (Figure 2). Thus we feel
confident in exploring the relationships between biomass partitioning and wood density. At Harvest 3 (10–12 weeks growth
at constant temperature), partitioning of total seedling biomass
in the roots declined from 35 to 20% as growth temperature increased (Figure 2). Partitioning of seedling biomass in stems
increased with growth temperature from 0.5 g to a maximum
of 23 g at 25 °C before declining, representing an increase
from 4 to 13% of seedling biomass. Biomass partitioning to
Figure 2. Effects of growth temperature on: (a) total seedling biomass;
(b) the proportion of total biomass in leaves; (c) wood in branches
(䊐,䊏) and main stems (䊊,䊉); and (d) roots of 19-week-old seedlings
(filled symbols) or of similar-sized seedlings (open symbols). Similar
aged seedlings (19 weeks old) were grown at a constant temperature
for the final 10–12 weeks. Seedlings, which were of similar size, were
19 weeks old if grown at 10 or 35 °C, but 15 weeks old if grown at 20,
25 or 30 °C. The 15-week-old seedlings were grown at a constant temperature for the final 7–8 weeks. Each value is the mean of five replicates. Bars represent SE.
TREE PHYSIOLOGY VOLUME 27, 2007
TEMPERATURE EFFECTS ON EUCALYPT GROWTH AND WOOD DENSITY
255
woody branches increased more steadily from 15 to 22% as
growth temperature increased (Figure 2). Leaf biomass remained constant at 46% of seedling biomass at all growth temperatures (Figure 2). Values were similar when seedlings of
similar size were compared (Figure 2). Wood density was
higher when a larger component of seedling biomass was partitioned to the woody main stem and branches (r = 0.69; P <
0.05) (Figure 3).
Wood density and anatomy
Wood density increased 20% with increasing growth temperature to 436 kg m – 3 at 30 °C before declining to 410 kg m – 3
when seedlings were grown at 35 °C (P < 0.05) (Figure 1). The
changes in wood density were related to changes in stem anatomy (Figures 4 and 5). Wood density was positively correlated
to fiber cell wall thickness (r = 0.62 ; P < 0.05), and negatively
correlated to both the diameter of fiber cell lumen (r = –0.60;
P < 0.05) and the proportion of the stem occupied by xylem
vessels (r = –0.76 ; P < 0.05) (Figure 4). Together these three
variables explained 76% of the variation in wood density (r =
0.87). Seedlings grown at 10 °C had a larger proportion of
stem allocated to xylem vessels and a higher abundance of xylem vessels than seedlings grown at higher temperatures (Table 1, Figure 5). Nearly 70% of xylem vessels in seedlings
grown at 10 °C, like those present in seedlings grown at 35 °C,
had diameters of less than 30 µm, and less than 5% of xylem
vessels had diameters greater than 45 µm. In comparison,
about 50% of xylem vessels in seedlings grown at temperatures between 20 and 30 °C had diameters less than 30 µm and
20% had diameters greater than 45 µm (Table 1). Temperature
marginally decreased fiber cell diameter, and this, together
with the changes in fiber cell wall thickness, reduced the fiber
cell lumen diameter, and altered the spatial arrangement between fiber cell lumen and fiber cell walls (Table 1). The radius of the pith was similar in seedlings grown at 10 and 35 °C,
and smaller than in seedlings grown at the other temperatures
(Table 1); consequently, the proportion of stem area occupied
by the pith was higher in seedlings grown at 10 and 35 °C. In
seedlings in all treatments, however, the pith accounted for
less than 3% of stem area (Table 1).
Figure 3. Wood density as a function of the proportion of total seedling biomass partitioned to the woody stems and branches. Seedlings
grown at a constant temperature for the final 2–4 weeks (䊏),
6–8 weeks (䊉) and 10–12 weeks (䉱) were 11, 15 and 19 weeks old,
respectively. Each value represents the mean and SE of five replicates.
Figure 4. Relationships between wood density and (a) fiber cell wall
thickness, (b) diameter of fiber cell lumen and (c) the cross-sectional
area of the stem occupied by xylem vessels. Data are for 19-week-old
seedlings grown for the final 10–12 weeks at constant temperatures of
10 °C (䊏), 20 °C (䊉), 25 °C (䉱), 30 °C (䉬) and 35 °C (夹). The regression line and correlation coefficient are for all data.
Seedling growth
The temperature optimum for maximum growth (i.e., biomass
accumulation, stem volume, leaf area) of E. grandis differed
from the temperature for maximum wood density (Table 1,
Figures 1 and 6). Stem radial growth and stem volume were
principally related to the production of new cells rather than
cell expansion (Table 1, Figure 1). Fiber cell diameter varied
by only 20% with changes in temperature, whereas fiber cell
number increased by 290% (Table 1). Seedling height, diameter and biomass increased by up to 2000% with increasing
growth temperature to a maximum at 25 °C before declining
(Table 1, Figure 2). As a result, overall seedling size in the 10
and 35 °C growth temperature treatments were similar (Table 1, Figure 2). In contrast, leaf-scale parameters (number of
leaves at final harvest, leaf area, leaf thickness, SLA, LAR, Al)
had the same temperature optimum (30 °C) as observed for
wood density (Table 1, Figures 1, 6). However, in all these
cases, the 30% to greater than 80% decline in leaf scale parameters at 35 °C relative to 30 °C contrast to the less than 10% decline in wood density as growth temperature increased from 30
to 35 °C (Figures 1 and 6). Leaf CO2 assimilation rate increased with temperature to a maximum of 22 µmol m – 2 s – 1 at
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THOMAS, MONTAGU AND CONROY
Table 1. Characteristics of E. grandis seedlings aged 19 weeks and grown at constant temperature for the final 10–12 weeks. Values are means of
five replicates for growth measurements and means of four replicates for anatomical determinations. Standard errors are shown in parenthesis.
Characteristic
Growth temperature (°C)
LSD0.05
10
20
25
30
35
Seedling morphology
Height (m)
Over bark stem diameter (mm)
Under bark wood diameter (mm)
Number of branches
Root dry matter (g)
Leaf dry matter (g)
Mean area per leaf (cm2)
Number of leaves on seedling at harvest
Leaf thickness (mm)
0.14 (0.01)
6 (0.5)
4 (0.4)
9 (0.6)
4 (0.3)
6 (1.1)
30.7 (3.6)
24 (2)
0.27 (0.01)
0.61 (0.04)
14 (0.4)
11 (0.5)
15 (0.4)
31 (3.0)
61 (7.8)
37.6 (3.0)
301 (44)
0.24 (0.02)
0.99 (0.07)
16 (0.5)
12 (0.5)
18 (0.7)
36 (4.0)
79 (2.7)
40.2 (2.6)
440 (37)
0.20 (0.01)
0.70 (0.04)
13 (0.4)
10 (0.4)
17 (0.5)
30 (1.8)
67 (2.9)
25.4 (1.9)
633 (46)
0.21 (0.01)
0.21 (0.01)
5 (0.7)
3 (0.2)
11 (0.9)
2 (0.5)
5 (0.9)
7.0 (0.2)
99 (14)
0.25 (0.02)
0.15
1.5
1.5
1.8
6.8
13.9
8.4
49
0.02
Wood anatomy
Pith radius (µm)
Pith area (% of stem area)
Number of fiber cells per radial transect
Average fiber cell diameter (µm)
Fiber cell wall thickness (µm)
Fiber cell lumen diameter (µm)
Cross-sectional areas of fiber cell lumen:fiber cell wall
Xylem vessel area (% stem area)
Xylem vessel number (vessels mm – 2 stem)
Proportion of xylem vessel < 30 µm diameter
Proportion of xylem vessel from 30 to 45 µm diameter
Proportion of xylem vessel > 45 µm diameter
260 (68)
2.4 (0.6)
116 (13)
15.1 (0.4)
1.8 (0.1)
11.4 (0.4)
1.31 (0.05)
17.0 (0.3)
408 (63)
0.68 (0.004)
0.29 (0.006)
0.03 (0.008)
455 (39)
0.9 (0.2)
300 (16)
15.6 (0.1)
2.1 (0.1)
11.3 (0.3)
1.12 (0.09)
11.5 (0.5)
233 (21)
0.54 (0.005)
0.27 (0.02)
0.19 (0.04)
425 (28)
0.6 (0.1)
328 (13)
16.0 (0.1)
2.3 (0.1)
11.4 (0.1)
1.05 (0.06)
11.3 (0.4)
271 (33)
0.52 (0.009)
0.28 (0.03)
0.20 (0.03)
369 (37)
0.6 (0.2)
315 (17)
14.6 (0.4)
2.3 (0.1)
9.9 (0.4)
0.87 (0.04)
9.6 (0.2)
169 (14)
0.47 (0.02)
0.35 (0.03)
0.18 (0.01)
212 (37)
1.8 (0.4)
113 (10)
13.4 (0.3)
2.0 (0.1)
9.5 (0.5)
1.03 (0.15)
10.4 (0.4)
290 (48)
0.88 (0.05)
0.12 (0.05)
0.0 (0.0)
137
1.3
32
0.9
0.3
1.0
0.23
3.6
43
0.05
0.04
0.04
30 °C and declined sharply to 12 µmol m – 2 s –1 at 35 °C (Figure 6), whereas leaf respiration increased linearly from 1 µmol
m – 2 s –1 at 10 °C to 2 µmol m – 2 s –1 at 30 °C then increased to
3 µmol m – 2 s –1 at 35 °C. Although seedlings grown at 35 °C
had greatly reduced growth but comparatively high wood density, positive correlations existed between wood density and
several leaf-scale parameters including leaf area, LAR and Al
(Figure 7).
Extrapolation of Al to Ac based on plant leaf area showed
that Ac peaked at 25 and 30 °C and reached a minimum at 10 or
35 °C (Figure 8). As shown in Figure 8, Ac was positively correlated with wood density (r = 0.63), wood volume (r = 0.85)
and fiber cell wall thickness (r = 0.57).
Discussion
Wood density and anatomy
The increase in wood density of E. grandis seedlings grown at
high temperatures was related to reductions in lumen transverse area of both xylem vessels and fiber cells and thicker fiber cell walls (Table 1, Figures 1, 4 and 5). Changes in these
anatomical features accounted for 76% of the observed variation in wood density.
The volume of pith could influence wood density, because
the pith contains cells with thin walls and large lumens that affect overall stem density. However, the pith occupied less than
3% of stem area (Table 1) and thus contributed minimally to
wood density. The similar proportion of pith in stems of seedlings grown at 10 and 35 °C and the large differences in wood
density indicate that other factors play a more important role in
the response of wood density to growth temperature (Table 1,
Figures 4 and 5).
The decline in xylem vessel lumen diameter was most evident in comparisons between seedlings grown at 10 and 35 °C
(Table 1, Figures 1, 4 and 5). These seedlings were similar in
other aspects of growth (stem volume, Al, leaf area and LAR)
(Figures 1, 6 and 7), yet xylem vessel number, size distribution, area of xylem vessels and wood density differed considerably (Table 1, Figures 1, 4 and 5). The importance of xylem
vessel lumen diameter determining wood density of other angiosperms has been shown in studies with East-Liaoning oak
(Quercus liaotungensis Koidz) (Zhang and Zhong 1992) and
other eucalypts (Malan and Hoon 1992, Thomas et al. 2004).
Xylem vessel size and number has a degree of plasticity that
is dependent on environmental conditions. Why then do xylem
vessels respond to growth temperature independently of evaporative demand and water supply? Roderick and Berry (2001)
postulated that changes in xylem vessel size in response to
growth temperature are associated with the decrease in viscosity of water as temperature increases, making it unnecessary
for a plant to have more or larger xylem vessels to maintain hydraulic conductivity at high temperatures. This postulate is
supported by the results of a study of E. camaldulensis Dehnh.
grown at different temperatures (Thomas et al. 2004), and by
our finding that xylem vessel frequency and size of E. grandis
TREE PHYSIOLOGY VOLUME 27, 2007
TEMPERATURE EFFECTS ON EUCALYPT GROWTH AND WOOD DENSITY
Figure 5. Photographs of 1 µm wood sections stained with toluidine
blue obtained from 19-week-old seedlings grown at: (a) 10 °C; (b)
20 °C; (c) 25 °C; (d) 30 °C; and (e) 35 °C for the final 10–12 weeks.
The bar represents 100 µm.
decreased with increasing growth temperature (Table 1, Figure 4).
The arrangement of mass to volume within the fiber cells
decreased with growth temperature because of declining fiber
cell expansion and increasing cell wall thickness (Table 1, Figure 4). Both factors were important in explaining wood density. These changes in fiber anatomy are analogous to the
changes in tracheid cell anatomy of early and latewood in
gymnosperms. The less-dense earlywood tracheid cells have
thin walls and a large lumen, whereas the more dense latewood
tracheids have thick walls and small lumens (Creber and
Chaloner 1984, Zobel and Jett 1995). Wood density of gymnosperms depends on the ratio of the lumen diameter of earlywood tracheids to latewood tracheids (Creber and Chaloner
257
Figure 6. Effects of growth at constant temperature for the final 10–12
weeks on: (a) leaf area; (b) leaf area ratio; (c) specific leaf area; (d)
leaf CO2 assimilation rate (Al ); and (e) canopy CO2 assimilation capacity (Ac) of 19-week-old seedlings. Each value represents the mean
and SE of five replicates.
1984). It appears that wood density of eucalypts depends on
similar localized arrangements of fiber cell wall thickness to
fiber lumen area, and these changes are influenced by growth
temperature. The thickness of fiber cell walls was positively
related to Ac (Figure 8), which was principally affected by
changes in seedling leaf area. Cells with thicker walls had
smaller lumen diameters, but the ratio of cell wall area to cell
lumen diameter was also related to lumen fiber cell diameter
(Table 1). Thus, growth temperature affects the diameters of
fiber cells of E. grandis seedlings (Table 1) in a way that is
similar to its effect on fibers of gymnosperm seedlings
(Larson 1964, Richardson 1964). These data may not correspond with field data, because other growth effects, such as
rainfall, elevation and soil conditions, may alter the general relationship between temperature and wood density (Wilkes
1987, Horacek et al. 1999) through independent effects on volume growth and biomass accumulation or partitioning.
TREE PHYSIOLOGY ONLINE at http://heronpublishing.com
258
THOMAS, MONTAGU AND CONROY
Figure 7. Relationships between wood density and (a) leaf area, (b)
leaf area ratio and (c) leaf CO2 assimilation rate measured in
19-week-old seedlings. Each value represents the mean and SE of five
replicates at each growth temperature. Seedling growth temperatures
are indicated on graphs. Regression lines and correlation coefficients
are for all growth temperatures (solid line) and for growth temperatures between 10 and 30 °C (dashed line).
Figure 8. Effects of canopy CO2 assimilation capacity on (a) wood
volume, (b) wood density and (c) fiber cell wall thickness of
19-week-old seedlings. Each value is the mean and SE of five replicates for each growth temperature. Seedling growth temperatures are
indicated on graphs. Regression lines and correlation coefficients are
for all growth temperatures (solid line) or for growth temperatures between 10 and 30 °C (dashed line).
Carbon supply and partitioning
In the stem, sink strength can be measured as the production of
new cells and cell wall thickening. As hypothesized, temperature affected sink strength by altering the overall biomass production and also by shifting the pattern of biomass partitioning
from roots to shoots. The woody components of the shoots
(stems and branches) rather than the leaves received a larger
proportion of the extra biomass (Figure 2). The extra biomass
contributed not only to greater cell production but also to
thicker fiber cell walls and higher wood density (Table 1, Figure 3). Similar changes in biomass partitioning in response to
temperature have been observed in seedlings of beech (Fagus
sylvatica L.) (Bruhn et al. 2000) and mountain birch (Betula
pubescens Ehrh. ssp czerepanovii (Orlova) Hamet-Ahti)
(Bruhn et al. 2000, Weih and Karlsson 2001).
The leaf-scale parameters contributing to Ac (i.e., Al , LAR
and SLA) peaked at a growth temperature between 25 and
30 °C (Figure 6). The same parameters also increased in
B. pubescens ssp czerepanovii seedlings grown at high air and
soil temperatures (Weih and Karlsson 2001). Although the
growth temperatures tested had no effect on biomass partitioning to the leaves (Figure 2), LAR changed with growth tem-
perature because of changes in SLA, suggesting that the link to
Al is through leaf morphology (Sefton et al. 2002). Both Al and
leaf area contributed to changes in Ac (Figure 6), with changes
in leaf area principally influencing these changes. Changes in
Ac contributed to changes in fiber cell wall thickness, wood
volume and wood density of E. grandis (Figure 8), as found in
other studies (Larson 1964, Creber and Chaloner 1984,
Lindström 1996). Compared with seedlings grown between 20
and 30 °C, seedlings grown at 10 and 35 °C had low Al and
very low leaf area resulting in low Ac, which was reflected in
thin fiber cell walls (Table 1, Figures 6 and 8). Low photosynthate supply as a result of factors, such as higher night temperatures leading to increased respiration, reduced solar irradiance or shorter photoperiods, may result in reduced cell wall
thickness of gymnosperm seedlings and saplings (Larson
1964, Richardson 1964).
Wood density: combining mass and volume
Wood density may be stable if Ac and wood volume change by
similar amounts. This occurred when growth temperature was
increased from 20 to 25 °C (Figure 8). In contrast, wood den-
TREE PHYSIOLOGY VOLUME 27, 2007
TEMPERATURE EFFECTS ON EUCALYPT GROWTH AND WOOD DENSITY
sity may increase if temperature affects wood volume independently of Ac. This occurred when growth temperature was
increased from 25 to 30 °C (Figure 8), because fiber cell expansion rather than cell division was affected (Table 1). When
the temperature sensitivity of stem volume and stem biomass
growth are similar, wood density is unaffected. Such coordination of temperature responses occurred when growth temperature increased from 20 to 25 °C. Similarly, elevated CO2
concentration, which can cause increases in Ac and stem biomass, increased stem volume but not wood density of
P. sylvestris and loblolly pine (Pinus taeda L.) (Telewski et al.
1999, Ceulemans et al. 2002).
In summary, temperature affected wood density by changing the mass of the stem and the volume of the lumen within
the stem. Stem mass changed because temperature altered
both the seedling carbon source strength—by changing Al, and
leaf area—and the partitioning of this carbon to the stems.
Temperature altered stem volume and stem radial growth by
changing the rate of cell division more than the rate of cell expansion. At a more detailed anatomical level, changes in wood
density were driven by changes in the proportion of xylem lumen volume. At the level of individual cells, the proportion of
fiber cell lumen to fiber cell wall material altered because of
declining fiber cell expansion and increasing cell wall thickness. The mechanism by which increasing temperature may
reduce the total volume of xylem vessels per unit stem volume
may be related to the decrease in the viscosity of water with
increasing temperature, which allows fewer or smaller xylem
vessels to transport similar volumes of water.
Acknowledgments
This research was funded by an ARC SPIRT grant in collaboration
with State Forests of NSW (Grant No. C00001999). We thank
Deborah Birch, Elizabeth Darley, Kate Düttmer, Mark Emanuel, David Giles, Matthew Searson and Christine Sefton for assistance during
plant harvests and microscopy analysis, and Georgina Kelley for assistance with the preparation of the manuscript.
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