Download Growth Rings in the Roots of Temperate Forbs are Robust Annual

Research Paper
224
Growth Rings in the Roots of Temperate Forbs
are Robust Annual Markers
G. von Arx and H. Dietz
Institute of Integrative Biology ETH, Swiss Federal Institute of Technology, ETH Zentrum CHN, G 35.1, 8092 Zürich, Switzerland
Received: June 10, 2005; Accepted: November 12, 2005
Abstract: There is increasing interest in the analysis of annual
growth rings in the secondary root xylem of perennial forbs
(herb-chronology). Therefore, we need to verify whether these
growth rings are always formed annually. To investigate the formation of root rings we performed common garden experiments at two distinct sites in Switzerland. We grew nine unrelated forb species from seed and subjected them to competition
and clipping treatments. Anatomical developments in the roots
of the individuals were tracked during five growing seasons.
Across all species and treatments at least 94% of the expected
growth rings associated with full growing seasons were identifiable and the development of the anatomical patterns was consistently seasonal. While the distinctness of annual rings varied
somewhat between species and sites, the treatments had no effect on the presence of annual rings. In no case were false rings
developed. The results of this study demonstrate that the
growth rings in the roots of northern temperate forbs represent
robust annual growth increments and, hence, can reliably be
used in herb-chronological studies of age- and growth-related
questions in plant ecology.
Key words: Age determination, anatomical pattern, annual ring,
growth increment, herb-chronology, perennial forbs.
Introduction
A thorough understanding of the processes underlying plant
population and community development requires not only
knowledge on the present state of vital parameters like plant
size, fitness, and population density but also on their change
over time. Limited access to understanding such changes is
possible by interpreting static, age-classified life tables (e.g.,
Dietz and Ullmann, 1998; Brys et al., 2004; Freville and Silvertown, 2005). However, age-related parameters such as birth
and death rate, the age structure of the population or age-dependent size and fitness are relatively difficult to obtain, especially for perennial forbs (cf. Harper, 1977; Vorontzova and
Zaugolnova, 1985; Menges, 2000).
Plant Biol. 8 (2006): 224 – 233
© Georg Thieme Verlag KG Stuttgart · New York
DOI 10.1055/s-2005-873051
ISSN 1435-8603
A promising methodological approach to investigate age-related patterns in forbs is herb-chronology (i.e., the ecologicallyoriented analysis of annual rings in the secondary root xylem
of perennial forbs), which has been developed during the past
decade (e.g., Dietz and Ullmann, 1997, 1998; Dietz and Fattorini, 2002; Dietz and von Arx, 2005; von Arx and Dietz, 2005).
The anatomical basis of annual growth increments (hereafter
referred to as “annual rings”) in the roots of forbs is the formation of earlywood vessels with large lumina in spring and latewood vessels with smaller lumina in late summer and autumn
(Dietz and Ullmann, 1997; von Arx and Dietz, 2005). Similar to
the better known tree rings (e.g., Larson, 1994; Schweingruber,
2001), discernible annual rings in the secondary root xylem
have proven to be a common phenomenon in many perennial
forb species with persistent main roots (Zoller, 1949; Bakshi
and Coupland, 1960; Werner, 1978; Humulum, 1981; Boggs
and Story, 1987; Dietz and Ullmann, 1998; Schweingruber and
Dietz, 2001; Dietz and Fattorini, 2002; Dietz and Schweingruber, 2002; Dietz and von Arx, 2005).
Counting the number of annual rings in the roots of forbs has
been used for age determination, demographic analyses, and
investigations into population development (e.g., Boggs and
Story, 1987; Dietz and Ullmann, 1998; Dietz et al. 1999; Dietz,
2002). Furthermore, determination of plant age along environmental gradients allows us to make inferences on how plant
life histories respond to different growth conditions (Erschbamer and Retter, 2004; Rixen et al., 2004; von Arx et al.,
2006). Patterns of ring width have been used to study how
plant growth responds to variation in environmental conditions (Dietz and Ullmann, 1998; Dietz et al., 2004; von Arx et
al., 2006) or unpredictable events such as climatic fluctuations
(Dietz and von Arx, 2005).
Although the annual nature of growth rings could be verified
for one or several individuals of known age in about twenty
species (Dietz and Schweingruber, 2002), so far, it has not
been tested whether distinct anatomical implementations of
growth rings in unrelated species all represent true annual
rings and whether annual rings are reliably formed under a
wide range of growth conditions (Schweingruber and Dietz,
2001). For example, an individual may fail to form one or several annual rings when its aboveground growth is suppressed
by strong competition (Dietz and Ullmann, 1998) or permanent snow cover (Wijk, 1980). Furthermore, under unfavourable growth conditions growth rings may become indistinct
Annual Root Rings in Forbs
Plant Biology 8 (2006)
Table 1 Characterization of the study species. All species occur in Europe (native area) and in North America (introduced area; Tutin et al., 1964 –
1983; Natural Resources Conservation Service, 2005). Characterization of plant size and habitat preference follows Oberdorfer, 1994. Table modified after Dietz and Schweingruber, 2001, courtesy of the Bulletin of the Geobotanical Institute ETH
Species
Family
Life-form
Habitat
Root architecture
Type of
species*
Anchusa officinalis L.
Boraginaceae
semi-rosette forb,
deep-rooting
ruderal and other
disturbed sites
branched taproot
1A
Cichorium intybus L.
Asteraceae
semi-rosette forb,
deep-rooting
ruderal and other
disturbed sites
branched taproot
1A
Trifolium pratense L.
Fabaceae
low-growing forb
ruderal sites, meadows,
pastures
branched taproot
1A
Digitalis grandiflora Mill.
Scrophulariaceae
tall-growing forb
tall herb communities,
forest edges
fibrous roots
2A
Sanguisorba minor L.
Rosaceae
semi-rosette forb
open vegetation, often
on raw, calcareous soils
branched taproot
2A
Silene vulgaris Garcke
Caryophyllaceae
semi-rosette forb
open vegetation, often
on raw soils
branched taproot
2A
Euphorbia esula L.
Euphorbiaceae
clonal forb with lateral
roots, deep-rooting
ruderal sites with relatively low soil moisture
branched roots
3Z
Galium verum L.
Rubiaceae
clonal forb with lateral
roots, deep-rooting
meadows and pastures
on calcareous soils
fibrous roots
3Z
Hypericum perforatum L.
Hypericaceae
medium-sized forb
meadows, pastures,
road verges etc. on
nutrient-poor soils
branched roots
3Z
* 1, short-lived species with distinct growth rings; 2, long-lived species with distinct growth rings; 3, long-lived species with diffuse ring pattern; A, used in all sites and
treatments; Z, only used in control plots at Zurich.
because of missing or scarce latewood between successive earlywood zones (Dietz and Fattorini, 2002). Conversely, if a plant
is mown or heavily grazed by herbivores it may respond with
regeneration growth and formation of large, earlywood-like
vessels in the roots that may look like the beginning of a new
(annual) ring. Besides resulting in inaccurate determination of
plant age, such anatomical responses would blur the presence
of ring width signals preserved in specific calendar years that
were characterized by particularly good or bad growth conditions (Dietz and von Arx, 2005) by shifting the reading frame
of the chronologies.
Since short-lived forb species are generally more responsive to
short-term changes in growth conditions (Grime, 2001), they
may be particularly prone to forming false rings, for instance
when root growth resumes after disturbance (e.g., Martinkova
et al., 2004) or a temporary interruption due to unfavourable
growth conditions. Conversely, in long-lived species rings
may be hard to identify (pseudo-missing rings) when growth
rings become narrow and tightly packed (Dietz and Fattorini,
2002). In species that show less distinct patterns of annual
rings, environmental fluctuations may result in anatomical responses in the roots that prevent accurate identification of annual growth rings. In addition, patterns may vary depending
on the seasonality of the climate (Schweingruber and Dietz,
2001).
Here we report on the first experimental verification of the annual formation of growth rings in the secondary root xylem of
perennial forbs under contrasting growing conditions. We addressed the following main questions: (i) How does the anato-
my of annual rings develop over time and are there unambiguous criteria for detecting transitions between two consecutive
annual rings? (ii) Are growth rings formed on a truly annual
basis, irrespective of contrasting microsite conditions and anatomical and life history characteristics of the species? (iii) Is
the distinctness of the annual ring anatomy related to variation in the seasonality of climate and associated local site factors?
To investigate these questions we selected nine perennial forb
species that represent a wide range of phylogeny, longevity,
and distinctness of ring anatomy. Two common garden experiments were established, one located in the temperate lowland
north of the Swiss Alps and the other one in the insubric southern foothills of the Swiss Alps. Plants were subjected to control, competition, and clipping treatments and tracked over
five growing seasons.
Materials and Methods
Study species
Nine species of perennial forbs, representing a wide phylogenetic and ecological range, were selected for the study
(Table 1). Most of the species are widespread and abundant in
Switzerland, and all develop persistent main roots that allow
analysis of the development of annual rings over several years
(Fig. 1). The species were selected so as to include three species from each of three types: i) short-lived species with clearly demarcated growth rings (hereafter referred to as “shortlived species”), ii) long-lived species with clearly demarcated
225
226
Plant Biology 8 (2006)
Fig. 1 Examples of anatomical patterns in the secondary root xylem
of the study species. (a) Anchusa officinalis, (b) Cichorium intybus, (c) Trifolium pratense, (d) Digitalis grandiflora, (e) Sanguisorba minor, (f) Silene
vulgaris, (g) Euphorbia esula, (h) Galium verum, and (i) Hypericum perforatum as viewed through a dissecting microscope. Lignified structures
are stained with phloroglucinol/HCl and appear dark in the images. All
shown plants were grown from seed in summer 2000 and were har-
G. von Arx and H. Dietz
vested in early May 2004, except E. esula and H. perforatum which were
harvested in late April 2003. Black markers denote transitions from
latewood of the previous to earlywood of the following growing season, white markers denote presumable positions of uncertain annual
ring borders that were partly classified as missing in the analysis. Scale
bars = 500 μm.
Annual Root Rings in Forbs
Plant Biology 8 (2006)
Table 2 Description of the two study sites. Table modified after Dietz
and Schweingruber, 2001, courtesy of the Bulletin of the Geobotanical
Institute ETH
Zurich (north
of the Alps)
Oggio (southern
foothills of the Alps)
Location
47826′N, 8830′E
46805′N, 8859′E
Altitude
520 m asl
590 m asl
Climatic conditions
temperate
insubric (warm summers with high precipitation, relatively dry
and mild winters)
Mean annual
temperature
9 8C
11 8C
Mean annual
precipitation
1 100 mm
1 800 mm
Site characteristics
flat, sun-exposed
area
sun-exposed terrace
on south-facing slope
Soil characteristics
loamy
rich organic forest
soil with very good
drainage
Treatments
control, competition, clipping
control, clipping
growth rings (hereafter referred to as “long-lived species”),
and iii) long-lived species that show less clear, deviating anatomical patterns (hereafter referred to as “diffuse species”).
Experimental sites and setup
The experiment was established at a temperate site north of
the Swiss Alps (Zurich) and at an insubric site south of the
Swiss Alps (Oggio; Table 2). Zurich has a colder climate than
Oggio and the moisture of the loamy soil at Zurich is usually
higher than that of the well-drained, south-facing forest soil
at Oggio, although mean annual precipitation is higher south
of the Alps (Table 2). Furthermore, the Oggio site hardly experiences any frost during the vegetation period from March
to October or any day with a mean temperature below freezing, even in the winter months (MeteoSchweiz, 2005). At the
Zurich site, blocks with control plots (n = 12), clipping plots
(n = 10), and competition plots (n = 9) were established, whereas at Oggio, 12 plots each of control and clipping treatments
were established and randomized within a block. The three
diffuse species were only grown at the Zurich site.
Plants were raised from seeds obtained from different botanical gardens in central Europe and grown in the greenhouse for
one month before planting into the plots in late September
2000. Within plots, plants of the same species were grouped
together to minimize interspecific competition. More competitive species were grouped together within a plot to prevent
them from overgrowing smaller species. In the control and
clipping plots three individuals per species were planted at a
distance of 25 cm from each other to maintain competition
between individuals at low levels. In the clipping treatment,
plants were cut 2 cm above the ground in September 2001
and May 2002. In the competition treatment, eight individuals
per species were planted at a distance of 10 cm from each other to result in considerable, predominantly intraspecific com-
petition. Plots were not weeded, except for elimination of
strong spontaneous competitors in control and clipping plots
about three to five times per growing season. The plants were
not watered except occasionally during the summer heatwave
of 2003 (Schaer et al., 2004) to avoid excessive drought/mortality. The experiment was run until May 2004 and covered
three complete (2001 – 2003) and two incomplete (2000,
2004) growing seasons.
Data collection
Beginning in autumn 2001, one plot each per treatment and
site was randomly selected and completely harvested three
times per year (April, June, and October) to track the developing root xylem anatomy during the growth periods. Between
autumn 2001 and spring 2004, a total of eight plots were harvested per treatment and site. There was a total mortality of
40 % across all plots and years. At Oggio mortality in both treatments was higher (60 %) than at Zurich (20 % in control and
clipping plots and 40 % in competition plots). We therefore
combined all remaining plots in the last harvest at Oggio to obtain a sufficient number of plants for analysis. Differences in
mortality between species were pronounced, ranging from
13 % (Hypericum perforatum) to 57 % (Cichorium intybus).
Thin cross-sections of the proximal ends of the main roots
were obtained with a sledge microtome and stained with
phloroglucinol-HCl to render lignified structures of the secondary xylem (vessels and, where present, lignified parenchyma cells) red. Cross-sections were photographed through the
phototube of a dissecting microscope (Leica MZ8) using a digital camera (Nikon CoolPix 990) and the obtained images were
used for further analysis (Dietz and Fattorini, 2002).
Data analysis
For each individual, the anatomical pattern, in particular vessel size at the growing periphery of the secondary xylem, was
inspected for each harvest date and classified as consistent
or inconsistent with the expected phenological development
(Dietz and Ullmann, 1997). Consistency with the postulated
phenological development was noted if the following criteria
for the outer margin of the secondary xylem were met at the
different harvest times: (i, April) earlywood zone present with
large vessels fully developed (cf. Figs. 1 a – g, i, Fig. 2, spring);
(ii, June) earlywood zone complete, latewood zone with small
vessels developing (cf. Fig. 2, summer); or (iii, October) latewood zone fully developed (cf. Fig. 2, autumn). Furthermore,
the number of detectable rings and, if possible, the calendar
year of false and missing rings were determined for each individual to assess the reliability of annual ring formation. The incomplete rings of 2000 and 2004 were included in ring counting. The age of the plant samples was known to the observer,
which might raise concern whether ring counting might have
been biased. However, the objectivity of the applied criterion
(see Introduction) was proven by an automated image analysis
system (ROXAS, see von Arx and Dietz, 2005). To examine
whether clipping in autumn 2001 or spring 2002 resulted in
any irregularity in the developing anatomical patterns, the individuals from all treatments were classified as showing either
i) no irregularities, ii) conspicuous irregularities in parenchyma lignification, or iii) intra-annual fluctuations in vessel size.
227
228
Plant Biology 8 (2006)
Fig. 2
G. von Arx and H. Dietz
Annual Root Rings in Forbs
3
Plant Biology 8 (2006)
Fig. 2 Chronology of anatomical root development of the short-lived
Cichorium intybus (a) and the long-lived Sanguisorba minor (b) harvested between autumn 2001 (second growth period) and spring 2004
(fifth growth period). Note varying degrees in the lignification of parenchyma cells among individuals, especially in C. intybus, while the
pattern and arrangement of vessels are quite consistent within species.
Markers as explained in Fig. 1. Scale bars = 500 μm.
To test for dependence of the response variables on the factors
site, growth treatment, and type of species (cf. Table 1), nominal logistic regression was performed on proportion data and
log-linear analysis on count data (see Table 3 for details; Crawley, 2005). Significant relationships were assumed at p ≤ 0.05.
Data of the different harvest dates were pooled prior to statistical analysis. For factors with more than two levels, Bonferroni-adjusted α levels were used for pairwise comparisons. Results for the three species with diffuse anatomical patterns
were only used in comparisons between types of species. Differences between growth treatments were not statistically
tested for the Zurich data to avoid problems of pseudo-replication (Hurlbert, 1984). Nominal logistic regressions were performed in SAS JMP 5.1 and log-linear analysis in StatSoft Inc.
Statistica 5.5.
Results
Between autumn 2001 and spring 2004 a total of 686 individual plants (514 from Zurich, 172 from Oggio) were harvested
and analyzed; these were expected to contain overall 2257 annual rings in the main roots (depending on harvest date, individuals were expected to show between two and five annual
rings per plant).
Phenology of annual ring formation
Phenological development of annual rings corresponded to the
expected, strongly seasonal pattern in 99 % of all plants, i.e.,
wide earlywood vessels were only formed in spring and vessel
size decreased towards autumn (Figs. 1, 2). Only in H. perfora-
tum (Zurich, control plots) was there a considerable proportion
of individuals (38 %) that showed inconsistencies in the anatomical development.
Although there were occasional intra-annual fluctuations in
vessel size, these were generally easily discriminated against
true annual ring borders, which were characterized by a sharp
increase of vessel size between latewood of the previous and
first earlywood of the following growing period. Deviations
from this pattern were most pronounced in the diffuse species
(cf. Table 1). H. perforatum developed comparably wide vessels
within a band of weakly lignified parenchyma in the distal portion of the latewood (Fig. 1 i). In Euphorbia esula, annual ring
borders were only weakly demarcated but corresponded to a
distinct band of unlignified parenchyma (Fig. 1 g), whereas reliability and distinctness of annual rings varied considerably
within individual plants in Galium verum (Fig. 1 h). The onset
of earlywood development varied among species: while Trifolium pratense and Sanguisorba minor showed well-developed
earlywood by April, the other species had only started to develop the earlywood by this time (Fig. 1).
Presence of annual rings
In 74 % of individuals, the number of observed rings was identical to the number of expected rings, corresponding to 91 % of
all expected annual rings being observed across all species and
treatments. The first annual ring, representing the short initial
autumn growth period in 2000, was missing or indistinct
(hereafter referred to as “missing”) in 20% of all plants, whereas annual rings other than the first one were missing in 6 % of
the plants. Of these, about half had more than one missing
ring. False rings were generally not observed.
At Zurich, 82 % of the plants showed the expected number of
annual rings, and in a further 13 % only the first (incomplete)
ring was missing. At the insubric site near Oggio the proportion of individuals that lacked the first ring was significantly
higher (23 %; df = 2, χ2 = 33.8, p < 0.001; Fig. 3 a). This site effect
was mainly due to a strong response in only two species: at
Table 3 Overview of data analysis used to test for dependence of the anatomical response variables on the factors site, growth treatment, and
type of species. Proportion data were analyzed using nominal logistic regression and count data using log-linear analysis. The diffuse species were
excluded for site comparison. Treatment effects were only tested for Oggio data, types of species were only compared for data of control plots at
Zurich (see text and Table 1 for further explanations)
Response variable
Response levels
Data type
Model tested
Phenology of root anatomy
consistent with seasonal rhythm
inconsistent with seasonal rhythm
proportion
–*
Presence of annual rings
all rings present
first ring missing
other ring(s) missing
proportion
site × type of species × species
treatment × type of species × species
type of species × species
Calendar year of missing rings
only 2000 (first ring incomplete)
other calendar years missing
count variable
site × species
treatment × species
type of species
Anatomical effects of clipping
no effect
irregularity in parenchyma lignification
intra-annual fluctuation in vessel size
proportion
treatment × type of species × species
* Data structure was inappropriate for statistical testing, see “Results”.
229
230
Plant Biology 8 (2006)
G. von Arx and H. Dietz
Fig. 3 Variation in the proportion of plant
individuals with missing rings. (a) Differences
between sites, treatments, and types of species (cf. Table 1; for the latter classification
only individuals from control plots in Zurich
were used) and (b) differences between species. Species with diffuse anatomical pattern
were omitted from site and treatment comparisons, since they were only grown in control plots at Zurich. Different letters indicate
significant differences (p = 0.05). Z, Zurich;
O, Oggio; 0, control; CP, clipping; CM, competition; S, short-lived; L, long-lived; D, diffuse
species; Ao, Anchusa officinalis; Ci, Cichorium
intybus; Tp, Trifolium pratense; Dg, Digitalis
grandiflora; Sm, Sanguisorba minor; Sv, Silene
vulgaris; Ee, Euphorbia esula; Gv, Galium verum; Hp, Hypericum perforatum.
Zurich 91 % of the Anchusa officinalis individuals showed the
expected number of annual rings compared to only 47 % at
Oggio (df = 2, χ2 = 49.8, p < 0.001). In Digitalis grandiflora the respective proportions were 43 % (Zurich) and 27 % (Oggio; df = 2,
χ2 = 6.8, p = 0.04). In the other species, there was no or a similar
trend, but the differences in the respective proportion were
small (0 – 9 %). In contrast, plants from the two sites did not differ significantly in the low proportion of missing rings at positions other than the first position (28 % at Zurich vs. 21 % at
Oggio; df = 1, χ2 = 0.8, p > 0.25).
Among the clipped plants at Oggio, there was a higher proportion of individuals with the expected number of annual rings
and a lower proportion with only the first ring missing than
among the control plants (df = 2, χ2 = 20.4, p < 0.001; Fig. 3 a).
In clipped plants at Oggio, 81 % of missing rings were the first
ring, which was not significantly different from control plants
(77 %; df = 1, χ2 = 0.0, p > 0.90). At Zurich, irrespective of treatments, 82 % of the plants showed the expected number of annual rings (Fig. 3 a), and 71 % of the remainder lacked only the
first annual ring.
There was significant variation in the proportion of individuals
showing the expected number of rings among species and
among different types of species (df = 4, χ2 = 19.4, p < 0.001;
Fig. 3 a). The short-lived species showed the highest (84 %)
and the diffuse species the lowest number of exact matches
(58 %; Fig. 3 a). In all three types of species there were species
which showed a very close match between the number of observed and the number of expected rings (Fig. 3 b). However,
Annual Root Rings in Forbs
whereas the three short-lived species had a homogeneously
high proportion of individuals showing the expected number
of rings (≥ 77 %), this proportion varied considerably in the
long-lived and the diffuse species (36 % to 95 % and 30 % to
92 %, respectively). While in the short-lived species only 10%
of all individuals lacked the first ring, both the long-lived and
diffuse species had a much higher proportion of individuals in
which the first ring was missing (≥ 25 %; Fig. 3 a). In contrast,
the short-lived species showed a higher proportion of missing
rings at other positions than the first position (49 %) than the
long-lived (12 %) and diffuse species (28 %; df = 2, χ2 = 26.8,
p < 0.001).
Effects of clipping on root anatomy
At Oggio, anatomical irregularities occurred significantly more
frequently in clipped than in control plants (df = 2, χ2 = 27.8,
p < 0.001). Whereas conspicuous changes in parenchyma lignification were more frequent in clipped than in control plants
(26 % vs. 14 %), there was no considerable difference in intraannual fluctuations of vessel size (6 % vs. 8 %). At Zurich, between 4 % (competition) and 13 % (clipping) of the plants
showed conspicuous irregularities in parenchyma lignification
and between 4 % (competition) and 9 % (control and clipping)
intra-annual fluctuations of vessel size.
Discussion
Overall, our results confirm previous claims that the growth
rings in the roots of perennial forbs growing in the temperate
seasonal zone are robust annual markers. Across all species
and treatments at least 94 % of the expected growth rings associated with full growing seasons were identifiable. While
the formation of annual rings corresponded to the postulated
seasonal pattern in all but very few, exceptional cases, a small
proportion of rings were hard to identify or not identifiable.
However, most of these missing rings are attributable to problematic species or particular conditions as discussed below.
Robustness of annual ring formation in perennial forbs
In 76 % of all the plants with missing annual rings, these rings
were barely or not identifiable due to an indistinct transition
between xylem produced in the incomplete first and the second growing seasons. Plants which germinate late in the growing season may not attain a sufficient size to produce a detectable first growth ring. This is supported by the results of our
study and by the results of a separate experiment at Zurich in
which individuals of the short-lived Trifolium pratense and the
long-lived Sanguisorba minor were germinated in spring and
early summer and grown under conditions comparable to
those of the control plots (von Arx, unpublished data). In this
sample, all 70 plants used showed a clearly identifiable annual
ring marking the transition from the first growing period to
the second one. This result demonstrates that the first annual
ring may be consistently formed if plants do not germinate late
in the growing period. More critical are missing rings associated with later years, which were observed in 6 % of the individuals. However, an unknown percentage of these cases may be
ascribed to a few plants, harvested in spring, that had not yet
initiated formation of a new ring, or may be artefacts caused
by an erroneous choice of a younger root instead of the main
root, or by a spontaneous individual of the same species that
Plant Biology 8 (2006)
was harvested at the position of a dead experimental plant.
The existence of erroneous samples is also suggested by the
clarity of the other annual rings in several individuals that did
not show the expected number of rings.
Regional variation
Differences in climate and soil conditions (cf. Table 2) were
probably responsible for the higher distinctness of annual
rings at the temperate Zurich than at the insubric Oggio site.
The sun-exposed site at Oggio, which is located in a south-facing forest clearing, is more protected against strong climatic
influences such as frost than the Zurich site. The observed difference may indicate a general trend of decreasing distinctness
of annual rings along a gradient from the temperate to the subtropical zones (cf. Schweingruber and Dietz, 2001). For example, whereas individuals of Sanguisorba minor collected at
900 m asl on east-Mediterranean Cyprus (358N) showed distinct annual rings, in established individuals of the same species collected at 100 m asl on sub-tropical Tenerife (Canary
Islands, 288N) there were hardly any growth rings in the roots
(von Arx and Dietz, unpublished data). The distinctness of annual rings in response to different site conditions may also
vary considerably between species. The short-lived species
were more strongly affected by the differences between Oggio
and Zurich which may be due to their greater growth plasticity
(Grime, 2001).
Variation due to local growth constraints
Small-scale biotic and abiotic factors (vegetation density, herbivore attack, nutrient and light availability, etc.) often vary
considerably within plant populations and may strongly influence plant growth (e.g., Crawley, 1997; Silvertown and
Charlesworth, 2001). Although clipped plants or plants grown
in high-density plots were 20 – 80 % smaller than those of control plots (data not shown), and irrespective of corresponding
intra-specific differences in ring width (cf. Fig. 2), they showed
a lower proportion of missing rings than control plants. Our results indicate that regeneration growth after aboveground biomass removal or reduction of growth by strong competition is
not likely to result in missing or false rings. However, occasional intra-annual fluctuations in vessel size that may occur in regenerating plants require cautious anatomical interpretation
and comparison with the anatomical characteristics of true
ring borders in the same individual to avoid recording false
rings. Consequently, the general earlywood-latewood pattern
of annual rings proved to be a robust characteristic in forbs under a wide variety of growing conditions.
Phylogenetic variation
As assumed by our a priori classification of species, problems
in observing the expected number of annual rings were most
pronounced in the (diffuse) species with an abnormal anatomical pattern. Accordingly, tracking the formation of annual
rings during several growing seasons yielded less consistent
patterns than in the other types of species. This result supports
previous evidence that there is no generally reliable criterion
for identification of annual rings in the roots of forbs other
than patterns of vessel size (Werner 1978; Dietz and Ullmann
1997; von Arx and Dietz 2005). However, our results also show
that even in species with a problematic anatomical pattern
231
232
Plant Biology 8 (2006)
(e.g., Euphorbia esula), annual rings may be identifiable (with
the assistance of anatomical markers other than vessel size)
if the phenology of ring formation is known. The reliability of
such markers must be verified for each species. In some or
most of the species (e.g., Anchusa officinalis, Cichorium intybus, T. pratense, Silene vulgaris, Galium verum, and Hypericum
perforatum) the number of occasionally occurring bands of
(stronger) lignified parenchyma does not always correspond
to the number of annual rings and may therefore mislead the
observer (Werner, 1978).
The comparably uniformly low number of missing first rings in
the (tap-rooted) short-lived species may result from their generally faster growth that may have allowed them to attain a
sufficient size in the incomplete first growing season for producing an identifiable first annual ring, quite in contrast to
the long-lived species with fibrous roots (cf. Table 1).
Implications for herb-chronology
The two major concerns with herb-chronological analyses
have been that the method is destructive and that one needs
to be sure that the growth rings in the roots of forbs are developed annually. Many previous studies (see Introduction) have
demonstrated that the destructive approach of herb-chronological analysis does not seriously limit its value in plant ecology (but see Colling et al., 2002). Individuals in a small number
of species may regenerate from the roots remaining in the soil
after herb-chronological harvest and sample sizes in the order
of ten to 100 individuals per population already produce reasonable results for most questions in population ecology (cf.
Boggs and Story, 1987; Dietz and Ullmann, 1998; Dietz et al.,
1999; Dietz, 2002; Dietz et al., 2004; Erschbamer and Retter,
2004; Münzbergovà et al., 2005; von Arx and Dietz, 2005). In
most cases this sample size represents only a tiny fraction of
the population size and, hence, should neither represent an
unacceptable intervention into the viability of the population
nor preclude future resampling of the same population to
monitor changes in age or growth pattern over time (cf. Dietz,
2004).
In addition, the results of our present study have experimentally confirmed existing evidence that clear growth rings in
the roots of forbs are only induced by the seasonality of the
climate in the temperate seasonal zone but not by other factors such as competition or regrowth following disturbance. A
baseline proportion of about 1 – 2 % of full growing seasons
may lack a detectable annual growth ring in the main roots of
seasonal temperate forbs. For most studies, this small percentage will not pose serious problems. Neither the shape of population age structures nor ring width patterns will be considerably affected if an annual ring is occasionally missing. Even
studies sensitive to shifts in the reading frame of ring width
chronologies will produce acceptable results if low percentages of rings are absent, especially if one is interested in a ring
width signal that is among the most recent years of the studied
plants (Dietz et al., 2004; Dietz and von Arx, 2005). This is because the lower the number of rings one has to count (starting
from the outer border) to locate the ring of interest, the lower
the likelihood that a missing ring will result in the wrong identification of the target ring.
G. von Arx and H. Dietz
Acknowledgements
We thank Fritz H. Schweingruber for help in planning and
setting up the experiment, for technical advice and for comments on the manuscript. Sabine Güsewell is acknowledged
for assistance with data analysis and Tino Fotsch for support
in maintaining the common garden experiment. Jake M.
Alexander and two anonymous reviewers improved a former
version of the manuscript. This work was supported by a grant
of the Swiss National Science Foundation (SNF) to HD (NF 3165426.01).
References
Bakshi T. S. and Coupland R. T. (1960) Vegetative propagation in Linaria vulgaris. Canadian Journal of Botany 38, 243 – 249.
Boggs K. W. and Story J. M. (1987) The population age structure of
spotted knapweed (Centaurea maculosa) in Montana. Weed Science 35, 194 – 198.
Brys R., Jacquemyn H., Endels P., De Blust G., and Hermy M. (2004)
The effects of grassland management on plant performance and
demography in the perennial herb Primula veris. Journal of Applied
Ecology 41, 1080 – 1091.
Colling G., Matthies D., and Reckinger C. (2002) Population structure
and establishment of the threatened long-lived perennial Scorzonera humilis in relation to environment. Journal of Applied Ecology
39, 310 – 320.
Crawley M. J. (1997) Plant Ecology. Oxford: Blackwell Science.
Crawley M. J. (2005) Statistical Computing: An Introduction to Data
Analysis Using S-Plus. Chichester: John Wiley.
Dietz H. (2002) Plant invasion patches – reconstructing pattern and
process by means of herb-chronology. Biological Invasions 4, 211 –
222.
Dietz H. (2004) Plant invasion patches revisited – changes in development. Weed Technology 18, 1381 – 1385.
Dietz H. and Fattorini M. (2002) Comparative analysis of growth
rings in perennial forbs grown in an alpine restoration experiment.
Annals of Botany 90, 663 – 668.
Dietz H., Fischer M., and Schmid B. (1999) Demographic and genetic
invasion history of a 9-year-old roadside population of Bunias orientalis L. (Brassicaceae). Oecologia 120, 225 – 234.
Dietz H. and Schweingruber F. H. (2001) Development of growth
rings in roots of dicotyledonous perennial herbs: experimental
analysis of ecological factors. Bulletin of the Geobotanical Institute
ETH 67, 97 – 105.
Dietz H. and Schweingruber F. H. (2002) Annual rings in native and
introduced forbs of lower Michigan, USA. Canadian Journal of Botany 80, 642 – 649.
Dietz H. and Ullmann I. (1997) Age-determination of dicotyledonous
herbaceous perennials by means of annual rings: exception or
rule? Annals of Botany 80, 377 – 379.
Dietz H. and Ullmann I. (1998) Ecological application of “Herbchronology”: comparative stand age structure analyses of the invasive
plant Bunias orientalis L. Annals of Botany 82, 471 – 480.
Dietz H. and von Arx G. (2005) Climatic fluctuation causes largescale synchronous variation in radial increments of the main roots
of northern hemisphere forbs. Ecology 86, 327 – 333.
Dietz H., von Arx G., and Dietz S. (2004) Growth patterns in two Alpine forbs as preserved in annual rings of the roots: the influence
of a snowbank gradient. Arctic, Antarctic, and Alpine Research 36,
591 – 597.
Erschbamer B. and Retter V. (2004) How long can glacier foreland
species live? Flora 199, 500 – 504.
Freville H. and Silvertown J. (2005) Analysis of interspecific competition in perennial plants using life table response experiments.
Plant Ecology 176, 69 – 78.
Annual Root Rings in Forbs
Grime J. P. (2001) Plant Strategies, Vegetation Processes, and Ecosystem Properties. Chichester: John Wiley.
Harper J. L. (1977) Population Biology of Plants. London: Academic
Press.
Humulum C. (1981) Age distribution and fertility of populations of
the arctic-alpine species Oxyria digyna. Holarctic Ecology 4, 238 –
244.
Hurlbert S. H. (1984) Pseudoreplication and the design of ecological
field experiments. Ecological Monographs 54, 187 – 211.
Larson P. R. (1994) The Vascular Cambium. Development and Structure. Berlin: Springer.
Martinkova J., Kocvarova M., and Klimesova J. (2004) Resprouting after disturbance in the short-lived herb Rorippa palustris (Brassicaceae): an experiment with juveniles. Acta Oecologica 25, 143 – 150.
Menges E. S. (2000) Population viability analyses in plants: challenges
and opportunities. Trends in Ecology and Evolution 15, 51 – 56.
MeteoSchweiz (2005) Norm- und Mittelwerte 1961 – 1990 (http://
www.meteoschweiz.ch /de/Daten/Messwerte/IndexMesswerte.
shtml).
Münzbergova Z., Krivanek M., Bucharova A., Juklickova V., and Herben, T. (2005) Ramet performance in two tussock plants – do the
tussock-level parameters matter? Flora 200, 275 – 284
Natural Resources Conservation Service, USDA (2005) Plants national database (http://plants.usda.gov/index.html).
Oberdorfer E. (1994) Pflanzensoziologische Exkursionsflora. Stuttgart: Ulmer.
Rixen C., Casteller A., Schweingruber F. H., and Stoeckli V. (2004) Age
analysis helps to estimate plant performance on ski pistes. Botanica Helvetica 114, 127 – 138.
Schaer C., Vidale P. L., Lüthi D., Frei C., Häberli C., Lingier M. A., and
Appenzeller C. (2004) The role of increasing temperature variability in European summer heatwaves. Nature 427, 332 – 336.
Schweingruber F. H. (2001) Dendroökologische Holzanatomie. Anatomische Grundlagen der Dendrochronologie. Bern: Paul Haupt.
Schweingruber F. H. and Dietz H. (2001) Annual rings in the xylem of
dwarf shrubs and perennial dicotyledonous herbs. Dendrochronologia 19, 115 – 126.
Silvertown J. and Charlesworth D. (2001) Introduction to Plant Population Biology. Oxford: Blackwell Science.
Tutin T. G., Heywood V. H., Burges N. A., Moore D. M., Valentine D. H.,
Walters S. M., and Webb, D. A. (1964 – 1983) Flora Europaea. Cambridge: Cambridge University Press.
von Arx G. and Dietz H. (2005) Automated image analysis of annual
rings in the roots of perennial forbs. International Journal of Plant
Sciences 166, 723 – 732.
von Arx G., Edwards P. J., and Dietz H. (2006) Evidence for life history changes in high-altitude populations of three perennial forbs.
Ecology, in press.
Vorontzova L. I. and Zaugolnova L. B. (1985) Population biology of
steppe plants. In The Population Structure of Vegetation (White,
J., ed.), Dordrecht: Dr W. Junk Publishers, pp.143 – 178.
Werner P. A. (1978) On the determination of age in Liatris aspera using cross-sections of corms: implications for past demographic
studies. The American Naturalist 112, 1113 – 1120.
Wijk S. (1980) Plants and the snow cover. Fauna and Flora 75, 32 – 36.
Zoller H. (1949) Beitrag zur Altersbestimmung von Pflanzen aus
der Walliser Felsensteppe. Berichte über das Geobotanische Forschungsinstitut Rübel in Zürich 61 – 68.
Plant Biology 8 (2006)
G. von Arx
Institute of Integrative Biology ETH
Swiss Federal Institute of Technology
ETH Zentrum CHN
G 35.1
8092 Zürich
Switzerland
E-mail: [email protected]
Editor: R. Monson
233