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ORIGINAL ARTICLE
Teiko Kato ÆKiyoshi Ishida ÆHiroaki Sato
The evolution of nettle resistance to heavy deer browsing
Received: 21 August 2006 / Accepted: 4 April 2007 / Published online: 5 June 2007
The Ecological Society of Japan 2007
Abstract We examined whether heavy browsing by sika
deer, Cervus nippon Temminck, changed morphological
characteristics of a Japanese nettle, Urtica thunbergiana
Sieb. et Zucc., in Nara Park, where a large population of
sika deer has been maintained for more than
1,200 years. Wild nettles of Nara Park exhibited smaller
leaf area, 11–223 times more stinging hairs per leaf, and
58–630-times higher stinging hair densities than those of
other areas where there was no evidence of sika deer
browsing. There were no significant differences in
stinging hair length between the areas. Nettles from
Nara Park that were cultivated from seeds in a green-
house retained a larger number and higher density of
stinging hairs. In the field, nettles of Nara Park were less
frequently browsed by sika deer and showed higher
survivorship than nettles that were transplanted from an
unbrowsed area into Nara Park. These results indicate
that: (1) the U. thunbergiana population of Nara Park
has an extremely high stinging hair density compared
with those of unbrowsed areas; (2) this characteristic has
a genetic basis, and (3) stinging hairs serve as a defensive
structure against sika deer, contributing to an increase in
survivorship. Thus, we conclude that a U. thunbergiana
population in Nara Park, with extremely high stinging
hair densities, has evolved through natural selection due
to heavy browsing by sika deer.
Keywords Genetic variation ÆPlant defense Æ
Plant–herbivore interactions ÆRegional variation Æ
Urtica thunbergiana
Introduction
Plants exposed to heavy herbivory by mammals often
have more or sturdier spinescence, which includes
thorns, spines, prickles and stinging hairs, than plants
not exposed to heavy herbivory. For instance, brambles
(Rubus hispidus), angelica trees (Aralia spinosa) and
spiny shrubs (Damnacanthus indicus) bear longer, shar-
per and larger thorns in areas with high activities of
mammalian herbivores than in areas with low activities
(Abrahamson 1975; White 1988; Takada et al. 2001).
Plants with spinescence are less damaged by mammalian
herbivores than plants whose spinescence is experimen-
tally removed, and, therefore, spinescence has been
considered as a defensive structure against herbivores
(Cooper and Owen-Smith 1986; Obenso 1997; Takada
et al. 2003).
If spinescence characteristics are influenced by heavy
herbivory, changes may result from ecological or evo-
lutionary responses by the plants to the herbivory. The
former is exemplified by a damage-induced response
(Karban and Baldwin 1997); that is, plants regrow with
more or sturdier spinescence on shoots newly produced
following damage by herbivores. The latter involves
selection for plants with more or sturdier spinescence; as
a result, such characteristics are genetically fixed within
plant populations. Many studies have demonstrated that
real or simulated herbivory induces spinescence in vari-
ous plants, including Glochidion (Okuda 1987), acacia
(Young 1987), nettle (Pullin and Gilbert 1989; Muti-
kainen and Walls 1995), cactus (Myers and Bazely
1991), alder (Bauer et al. 1991), bramble (Bazely et al.
1991) and mustard (Agrawal 1999; Traw and Dawson
2002). However, few studies have established the evo-
lutionary basis of spinescence as a response to herbivory.
Danell and Bergstro
¨m(2002) give two reasons for this: it
is difficult to separate genotypic and phenotypic varia-
tion, and some environmental processes can cause
physical damage and the removal of biomass in ways
similar to herbivory. As far as we know, only Pollard
T. Kato (&)ÆH. Sato
Department of Biological Sciences,
Faculty of Science, Nara Women’s University,
Kitauoyanishimachi, Nara 630-8506, Japan
E-mail: yt-kato@gaea.ocn.ne.jp
Tel.: +81-744-220468
Fax: +81-744-220468
K. Ishida
Forestry and Forest Products Research Institute,
Kansai Research Center, Kyoto, Japan
Ecol Res (2008) 23: 339–345
DOI 10.1007/s11284-007-0387-7
and Briggs (1982,1984) and Mutikainen and Walls
(1995) examined the adaptive significance and genetic
basis of spinescence using the stinging nettle Urtica
dioica.
The stinging nettle (U. dioica) bears many stinging
hairs on stems and leaves. These hairs contain a toxic
liquid containing histamine, acetylcholine and serotonin
(Emmelin and Feldberg 1949; Collier and Chesher
1956). The stinging nettle is known to have higher
stinging hair densities in grazed areas than in ungrazed
areas (Pullin and Gilbert 1989). Rabbits and sheep
prefer nettles with few or no hairs to nettles with many
stinging hairs (Pollard and Briggs 1984). These facts
indicate that stinging hairs provide a defense against
mammalian herbivores. Intraspecific variation of sting-
ing hair density has a genetic basis, with a heritability of
0.3–0.4 (Pollard and Briggs 1982). Stinging hair density
is also influenced to some degree by shading (Pollard
and Briggs 1982). Furthermore, U. dioica increases its
stinging hair density on leaves newly produced following
simulated herbivore damage, which suggests that a
damage-induced increase in stinging hairs occurs in the
field. It is uncertain whether production of stinging hairs
is costly for the plant in terms of fitness; Mutikainen and
Walls (1995) found that there is a trade-off between
reproduction and stinging hair density, whereas Puusti-
nen et al. (2004) found no such trade-off. However, be-
cause U. dioica is a highly nitrophilous and
phosphorophilous species (Olsen 1921; Rorison 1968),
stinging hairs may compromise plant fitness under
nutrient-poor conditions (Pullin and Gilbert 1989).
Therefore, in U. dioica, both ecological and evolutionary
responses to herbivory are influenced by the availability
of soil nutrients and combine to determine the stinging
hair density.
Two nettles, U. thunbergiana and U. platyphylla, are
found in Japan. They often occur in areas with high
activities of sika deer (Cervus nippon) (Takatsuki 1980;
Kaji et al. 1991). Of these nettles, U. thunbergiana occurs
in Nara Park, Nara, central Japan. This park has an
area of 660 ha in which there are Buddhist temples,
Shinto shrines, open grasslands and evergreen woods,
and it serves as a habitat for about 1,200 sika deer, de-
spite being in the vicinity of an urban area. According to
the Man’yoshu—the earliest anthology of Japanese
verse—sika deer had already inhabited the park by
about AD 750. Because sika deer were regarded as
sacred animals, they have been protected for more than
1,200 years in the park (Ohigashi et al. 2003). If stinging
hairs reduce damage from sika deer, and if stinging hair
variation has a genetic basis, the nettles in Nara Park
might have higher stinging hair densities as an adaptive
(i.e., evolutionary) response to heavy browsing.
In this study, we investigated whether length and
density of stinging hairs differ among U. thunbergiana
populations in the absence and presence of sika deer
browsing, whether these characteristics have a genetic
basis, and whether stinging hairs serve as a defensive
structure against sika deer. We first examined leaf area
and the number, density and length of stinging hairs of
nettles in Nara Park and in other areas where there was
no evidence of sika deer browsing. Second, we cultivated
nettles from these areas from seeds in a greenhouse to
compare leaf and stinging hair characteristics. Third, we
transplanted nettles from an unbrowsed area into Nara
Park and compared their vulnerability to deer browsing
with that of nettles from Nara Park.
Materials and methods
Study organism
Urtica thunbergiana is distributed in central and south-
ern Japan. It is a perennial plant that grows in nutrient-
rich habitats mainly at the edges of woods (Kitamura
and Murata 1961; Kato 2001). Like U. dioica, it bears
stinging hairs on its stems and the upper and lower
surfaces of its leaves. These stinging hairs contain a toxic
liquid with chemical compounds that have not been
identified but that probably include histamine, acetyl-
choline and serotonin, as found in U. dioica (Emmelin
and Feldberg 1949; Collier and Chesher 1956).
In Japan, the mammals that can browse nettles are
hares, Japanese serows and sika deer. However, we did
not observe any nettles browsed by hares in this study.
Because Japanese serows inhabit mountainous regions,
their browsing would not affect our study area. Sika deer
are widely distributed from the lowlands to highlands
and from northern to southern Japan. Thus, only sika
deer browsing was taken into consideration in this study.
Study sites
Nara Park (3441¢N, 13551¢E) is situated at the center
of the Kansai District. It has an area of 660 ha, with
open grasslands and evergreen woods (Fig. 1). Sika deer
have been protected in the park for more than
1,200 years because they are regarded as sacred animals
(Ohigashi et al. 2003). As of 2005, the population of sika
deer in the park was about 1,200 (The Nara Foundation
for Protection of Deer, unpublished data). The study site
in the park [110 m above sea level (a.s.l.)] is sparsely
planted with Japanese cedars.
Five other study sites are located at the edges of cedar
forests, 20–50 km from Nara Park (Fig. 1): Kouchidani
(3434¢N, 1366¢E, 280 m a.s.l.), Takatori (3425¢N,
13549¢E, 550 m), Arashiyama (3500¢N, 13540¢E,
60 m), Sakurai (3432¢N, 13551¢E, 138 m), and
Yoshino (3419¢N, 13558¢E, 320 m). Sika deer have
increased in number since the 1980s in the Kansai Dis-
trict, including these sites, and they have now spread
throughout the district (Biodiversity Center of Japan
2004). However, these sites are situated near residential
areas or beside pathways leading to a main road, and,
thus, few sika deer may have migrated there at least
before the 1980s. Indeed, local woodland officers have
340
said that they had seen no sika deer around the sites. We
also found neither fecal pellets of sika deer nor evidence
of browsing by sika deer during the period of this study.
Therefore, it can safely be assumed that densities of sika
deer at these sites are much lower than that at Nara
Park.
Field survey
We examined the stinging hairs of U. thunbergiana be-
tween 6 and 20 October 2002 in Nara Park, Kouchidani,
Takatori and Yoshino. At each site, we randomly se-
lected ten nettles and sampled one leaf from each of the
2nd, 4th, and 6th or 7th nodes. These leaves are referred
to as upper, middle and lower, respectively. Leaf area
was measured with an automatic area meter (Model
AAM-7, Hayashi Denko Co. Ltd., Tokyo, Japan).
Stinging hairs on the lower and upper surfaces of the
leaves were counted, using photocopies of leaves en-
larged two or four times. The density of stinging hairs on
a leaf was calculated as the number of stinging hairs
divided by leaf area. Five stinging hairs were randomly
selected on each leaf surface, and their lengths were
measured to 0.01 mm under a binocular microscope.
The mean value was used to represent the stinging hair
length for the leaf.
Light intensity at the study sites was expressed as the
relative photosynthetically active photon-flux density
(RPPFD). RPPFD is defined as the ratio of the photo-
synthetically active photon-flux density (PPFD) at a
spot in question to the PPFD at an adjacent unshaded
place without vegetation. At each study site in mid-
October 2002, the RPPFD was measured ten times using
the LI-190SA Quantum sensor connected to the LI-1400
data logger (LI-COR, Lincoln, USA).
Greenhouse experiment
In October 2001 we collected seeds from four nettles that
were more than 3 m apart, to avoid the progeny of one
maternal plant or clones at four of the study sites: Nara
Park, Sakurai, Arashiyama and Kouchidani. The low
number of selected plants was due to small population
sizes, ranging from 50 to 200 individuals, and the small
proportions of plants bearing seeds at the sites, except
Nara Park. Seeds from the four plants were kept to-
gether at room temperature in a well-ventilated room
until the following spring. On 20 April 2002, the seeds
were put on moistened cellulose mats in petri dishes.
They were kept at 4C for 10 days in an incubator.
Afterwards, they were kept at 25C for 10 days to pro-
mote synchronous germination. Twelve seedlings with
two intact cotyledons were selected to represent each
location. Seedlings were planted into plastic pots (two
seedlings per pot; upper caliber, 16 cm; lower caliber,
10 cm; depth, 12.5 cm) filled with 1 l of perlite, and were
cultivated in a greenhouse, with each pot receiving
100 ml of a nutrient solution containing 1.0 mM
NaNO
3
every 2 days (for details of the solution, see
Koyama et al. 2001). The nitrate ion concentration in
the solution was considerably higher than those in the
soil at the study sites, which ranged from 0.3 mM to
0.8 mM (Kato et al. unpublished). Pots were randomly
moved weekly within the greenhouse, to avoid position
effects. On 22 November 2002, leaf area and the number
and length of stinging hairs were measured as described
previously.
Transplantation experiment
Before the transplantation experiment, we examined
stinging hair densities of nettles in Nara Park and Sak-
urai. In July 2002 we collected ten nettles randomly
within a quadrat with an area of 60 m
2
at both sites and
measured stinging hair densities of those nettles in the
same way as described previously. As a result, nettles
from Nara Park had significantly higher stinging hair
densities than nettles from Sakurai (means ± SDs for
Nara Park and Sakurai, n= 10: upper leaves, 12.12 ±
7.51 cm
2
and 0.023 ± 0.031 cm
2
; middle leaves,
6.08 ± 2.57 cm
2
and 0.030 ± 0.044 cm
2
; lower
leaves 6.43 ± 4.00 cm
2
and 0.139 ± 0.187 cm
2
)
[analysis of variance (ANOVA) F= 128.113, d.f. = 1,
P< 0.001].
In Nara Park, five plots (90 cm ·90 cm) were placed
at least 10 m apart, and each of them was divided into
two subplots. On 2 April 2004, four young nettles were
transplanted from the quadrat at Nara Park to one
subplot and from that of Sakurai to the other subplot in
each plot, for fear that tall nettles from Sakurai
(6.0 ± 1.97 cm) would shade short nettles from Nara
Park (3.7 ± 1.37 cm) (the difference was significant; t-
test, P= 0.008, d.f. = 38). Nettles were arranged at
least 25 cm apart. Immediately after being transplanted,
Fig. 1 Locations of study sites in the Kansai District (AR
Arashiyama, KO Kouchidani, NA Nara Park, SA Sakurai, TA
Takatori, YO Yoshino)
341
the nettles were left for exposure to sika deer browsing.
Browsed nettles, which were identified by teeth marks on
their shoots, were counted twice each month from April
to August 2004. The surviving nettles were counted the
following spring.
Statistical analysis
To analyze the differences in leaf area and the number,
density and length of stinging hairs between nettles from
Nara Park and from each of the other sites, we per-
formed multiple comparisons using the Dunnett meth-
od. In the transplantation experiment, we analyzed
differences in vulnerability to deer browsing between
nettles from Nara Park and Sakurai by applying the log-
rank test (Krebs 1998) to cumulative percentage curves
of browsed nettles. All statistical analyses were carried
out with SPSS version 9.01J (SPSS 1999). A probability
of 0.05 was chosen as the level of statistical significance.
Results
Field survey
The RPPFDs did not differ significantly between study
sites (F= 0.192, d.f. = 2, P= 0.826). Values for
RPPFDs (means ± SDs) were as follows: Nara Park,
10.7 ± 3.3%; Kouchidani, 12.2 ± 7.7%; Takatori,
12.5 ± 1.1%. The site at Yoshino was submerged after
the construction of a dam, and so the RPPFD could not
be obtained.
Nettles in Nara Park had smaller leaf area than
nettles at all other sites at any leaf position (Fig. 2a),
although statistical significance of the differences
depended on leaf position along shoots. Middle leaves of
Nara Park nettles exhibited significantly smaller leaf
area than middle leaves collected at any other site. Up-
per leaves were significantly smaller only when com-
pared with those at Takatori, and lower leaves were
significantly smaller than those from both Takatori and
Yoshino.
The number of stinging hairs per leaf in Nara Park
was significantly higher than that at any other site at
each leaf position by a factor of 11–223 (Fig. 2b). Sim-
ilarly, the density of stinging hairs in Nara Park was 58–
630 times as high as those at other sites (Fig. 2c).
In contrast, there were no significant differences in
the length of stinging hairs between Nara Park and any
other sites at any leaf position except for the middle
leaves at Yoshino (Fig. 2d).
Greenhouse experiment
The leaf area of nettles grown from seeds did not differ
significantly between Nara Park and any other sites at
any leaf position except for the upper leaves at Sakurai
(Fig. 3a). This was obviously inconsistent with the
results from comparisons of wild plants.
Like wild nettles, however, nettles from Nara Park
had significantly more stinging hairs per leaf and higher
stinging hair densities on leaves than those from any
other site at each leaf position (Fig. 3b, c). Compared
with wild nettles, cultivated nettles exhibited high
stinging hair densities at any site and a small variation in
stinging hair density among study sites.
Stinging hair length did not differ significantly be-
tween Nara Park and any other sites at any leaf posi-
tions except for the middle and lower leaves at Sakurai
(Fig. 3d). Such small variation in stinging hair length
among sites was common to both cultivated and wild
nettles.
Fig. 2 Leaf area (a), number of
stinging hairs per leaf (b),
density of stinging hairs
(number per square centimeter)
(c), and length of stinging hairs
(d) of a Japanese nettle, Urtica
thunbergiana, growing in the
wild. An asterisk indicates a
significant difference between
Nara Park and one of the other
sites (P< 0.05). Upper,middle
and lower leaves represent the
2nd, 4th, and 6th or 7th leaves
along shoots, respectively. The
error bar is SD. ND, no data.
For location abbreviations, see
Fig. 1
342
Transplantation experiment
Most of the nettles transplanted from Sakurai to the
study site in Nara Park had been browsed by sika deer
by mid-May, and by the end of the study all of these
nettles had suffered damage (Fig. 4). Nettles trans-
planted from within Nara Park were rarely browsed
until late May, with the percentage showing damage
gradually increasing to 60% in early August. The two
cumulative percentage curves differed significantly (log-
rank statistic = 29.07, P< 0.001), indicating that net-
tles from Sakurai had suffered damage earlier and more
frequently than those from Nara Park.
Rates of survival till the following spring for trans-
planted nettles were 60% for those from Nara Park and
25% for those from Sakurai. The difference in survival
was only nearly significant (Fisher’s exact test,
P= 0.054).
Discussion
Three major findings resulted from this study of U.
thunbergiana in Nara Park, where a large population of
sika deer has been maintained for more than
1,200 years. First, the wild nettles of Nara Park have
extremely high stinging hair densities compared with
those in areas with no sika deer browsing (Fig. 2). The
stinging nettle U. dioica also has a higher stinging hair
density in grazed areas than in ungrazed areas (Pullin
and Gilbert 1989). However, differences in stinging hair
density of U. thunbergiana between Nara Park and other
areas are much greater (58–620 times) than those re-
ported for U. dioica between grazed and ungrazed hab-
itats (1.4–10.3 times).
The second major finding was that the number and
density of stinging hairs have a genetic basis. Both wild
and cultivated nettles from Nara Park exhibited a sig-
nificantly higher number and density of stinging hairs
than those from other areas (Figs. 2and 3). Pollard and
Briggs (1982) reported that stinging hair density in U.
dioica also has a genetic basis, estimating the heritability
at 0.3–0.4 by parent–offspring correlation analysis. One
might have some doubts about the soundness of our
result because of small sample sizes, particularly in the
greenhouse experiment. Cultivated seedlings from each
area were descended from only four plants. Further-
more, there is a possibility that those seedlings were the
progeny of one maternal plant, because they were se-
lected from bulked seeds of the four plants. Hence,
among-site variation in stinging hair traits may have
merely reflected an inter-individual variation. However,
the differences in the number and density of stinging
hairs between Nara Park and the other sites were very
distinct. This is hardly attributable to small sample size,
because results from small sample sizes would exhibit a
Fig. 3 Leaf area (a), number of
stinging hairs per leaf (b),
density of stinging hairs
(number per square centimeter)
(c), and length of stinging hairs
(d) of a Japanese nettle, Urtica
thunbergiana, cultivated from
seed in a greenhouse. For
abbreviation and symbols, see
Figs. 1and 2
Fig. 4 Curves showing the cumulative percentages of nettles
browsed by sika deer at a study site in Nara Park. Nettles were
transplanted from within Nara Park and from Sakurai into the
study site
343
large variation among the sites. Therefore, it is a safe
assumption that the result was largely unaffected by
sample sizes.
The third major finding was that nettles from Nara
Park were browsed less frequently by sika deer than were
nettles from Sakurai with lower stinging hair densities
(Fig. 4), so that they had a trend of higher survivorship.
This indicates that the stinging hairs of U. thunbergiana
served as a defensive structure against sika deer brows-
ing and contributed to an increase in survivorship.
Possibly, the difference in plant height between nettles
from Nara Park and those from Sakurai may have re-
sulted in the difference in damage and survivorship.
Because nettles from Nara Park were significantly
shorter than those from Sakurai, the former might be
less apparent to sika deer than those from Sakurai.
However, this possibility can be ignored because nettles
from the two sites were transplanted into different sub-
plots within a plot for fear that Nara Park nettles would
be shaded by Sakurai nettles.
U. dioica shows increased stinging hair density on
leaves newly produced following damage (Pullin and
Gilbert 1989). It is uncertain whether U. thunbergiana
shows such a damage-induced increase in stinging hair
density. However, it is unlikely, because cultivated net-
tles from Nara Park exhibited an increase in stinging
hair density compared with wild ones; the result was the
opposite of what was expected.
The evolution of an adaptive trait through natural
selection requires three conditions: heritable variation in
the trait, variation in lifetime reproductive success
among individuals, and the correlation of the trait with
lifetime reproductive success (Stearns and Hoekstra
2005). The evolution of extremely high stinging hair
density in the U. thunbergiana population of Nara Park
may satisfy these conditions. Variation in stinging hair
density was shown to be heritable. Nettles with higher
stinging hair densities were browsed less frequently by
sika deer than those with lower densities, so that the
former may survive and consequently reproduce more
successfully than the latter. A large number of browsing
sika deer have existed in Nara Park for more than
1,200 years (Ohigashi et al. 2003), thus acting as an
agent in natural selection for nettles with higher stinging
hair densities over this period. A population of U.
thunbergiana with extremely high stinging hair densities
might have evolved in Nara Park as a consequence of
this selective pressure.
Two factors other than mammalian herbivory could
influence stinging hair density of nettles: light conditions
(Pollard and Briggs 1982) and soil nutrient conditions
(Pullin and Gilbert 1989). Pollard and Briggs (1982)
demonstrated that shading changes stinging hair densi-
ties in U. dioica. In our study, however, light conditions
were unlikely to have caused variations in the stinging
hair density of U. thunbergiana, because light intensity,
expressed as RPPFD, did not differ significantly between
study sites. On the other hand, Pullin and Gilbert (1989),
in greenhouse experiments, suggested that nutrient-poor
conditions can lead to reduced stinging hair densities.
This may be true for U. thunbergiana as well, because
nettles cultivated in the greenhouse had high stinging
hair densities compared with those in wild plants; nitrate
ion concentration under cultivating conditions (1.0 mM)
was considerably higher than those under natural con-
ditions (0.3–0.8 mM). Even if soil conditions influence
stinging hair density of U. thunbergiana, this alone could
not explain the extremely large differences between net-
tles from Nara Park and those from other areas. The
differences in the stinging hair traits were evident, even
in the greenhouse experiment under nutrient-rich con-
ditions.
The length of stinging hairs of nettles in Nara Park
and other areas was similar, which suggests that stinging
hair length is not influenced by sika deer browsing. This
is somewhat inconsistent with observations involving
other plants. Brambles (R. hispidus), angelica trees (A.
spinosa) and spiny shrubs (D. indicus) under high
browsing pressure bear longer, sharper, and larger
thorns than those under low browsing pressure (Abra-
hamson 1975; White 1988; Takada et al. 2001). This
difference between U. thunbergiana and other plants may
reflect the difference between chemical and physical
defensive tactics. The stinging hairs of nettles contain
toxic compounds such as histamine, acetylcholine, and
serotonin; in addition, they are fragile and penetrate the
skin of browsing mammals, introducing toxins (Leng-
genhager 1974). Therefore, nettles do not need sturdy
stinging hairs. In contrast, thorns of other plants rep-
resent a physical defensive tactic, and, thus, may be
more effective when sharper, longer and larger.
Herbivory may induce changes in the size and shape
of plants (Danell and Bergstro
¨m2002). For instance,
heavy grazing by sika deer causes the grass Sasa nippo-
nica to become dwarf, with small leaves and short tillers
(Yokoyama and Shibata 1998). Wild nettles in Nara
Park had small leaves rather than those in unbrowsed
areas, whereas cultivated nettles from the park had
leaves as large as those of unbrowsed areas (Figs. 2and
3). This suggests that U. thunbergiana might decrease
leaf size following damage by sika deer, although there is
also a possibility that leaf area varies with environmental
conditions, such as water and soil nutrients, as it does in
the mustard Sinapis arvensis (Roy et al. 1999).
In conclusion, stinging hairs of U. thunbergiana serve
as a defensive structure against sika deer, and heavy
browsing by these deer in Nara Park would have re-
sulted in selection for plants with higher stinging hair
densities over 1,200 years. As a consequence, the ex-
tremely high stinging hair density of the population of
U. thunbergiana in Nara Park might have evolved
through natural selection.
Acknowledgments We thank Y. Shimada, Y. Hara, M. Inaba, and
Y. Kato for their assistance in the field and greenhouse experi-
ments; Y. Hirano, H. Furusawa, K. Kuroda, and J. Kikuchi for
their technical support; H. Torii, E. Shibata, S. Ikeda, and M.
Ohishi for valuable information and advice; and K. Kitagawara, K.
Tutsui, the late Y. Semura, and N. Maeshima for their permission
344
to use the study sites. We are grateful to K. Matsui, K. Maeda, and
M. Ishida for their permission to use apparatus and a greenhouse
of Nara University of Education. This study was supported
financially in part by a Grant-in-Aid for Scientific Research from
the Japan Society for the Promotion of Science (no. 18380090).
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