Ecography 31: 731740, 2008
doi: 10.1111/j.0906-7590.2008.05509.x
# 2008 The Authors. Journal compilation # 2008 Ecography
Subject Editor: Francisco Pugnaire. Accepted 2 June 2008
What shapes the altitudinal range of a high mountain Mediterranean
plant? Recruitment probabilities from ovule to seedling stage
Luis Giménez-Benavides, Adrián Escudero and José M. Iriondo
L. Giménez-Benavides (luis.gimenez@urjc.es), A. Escudero and J. M. Iriondo, Área de Biodiversidad y Conservación, Univ. Rey Juan CarlosESCET, Tulipa´n s/n. ES28933 Mo´stoles, Madrid, Spain
Recruitment is a complex process consisting of sequential stages affected by biotic interactions and abiotic factors.
Assessment of these sequential stages and corresponding subprocesses may be useful in identifying the most critical stages.
Accordingly, to assess the factors that may determine the altitudinal range limits of the high mountain Mediterranean
plant Silene ciliata, a set of demographic stages, from flower production to establishment of 2-yr-old plants, and their
influence on recruitment probability were examined using a step-by-step approach. We integrated florivory, pollination
and pre-dispersal seed predation as pre-dispersal factors, and seedling emergence and survival as post-dispersal
determinants of recruitment. Three populations were monitored at the southernmost margin of the species along its local
altitudinal range. Previous studies suggest that seediness is strongly limited by summer drought especially at the lower
boundary of the species, a situation that may worsen under current global warming.
Our results showed that recruitment was mainly limited by low seed production in the pre-dispersal stage and low
seedling emergence and survival in the post-dispersal stage, probably due to environmental harshness in summer. By
contrast, biotic factors responsible for propagule loss, such as flower and fruit predation, had a minor effect on the
probability of plant recruitment. Although the relative importance of transition probabilities was similar among
populations along the altitudinal range, comparatively lower flower production significantly reduced the number of
recruited plants at the lowest altitude population. This demographic bottleneck, together with increased competition with
other species favoured by climate warming, might collapse population growth and limit persistence at the lower
altitudinal range of the species, raising its low local altitudinal edge.
Plant species are not expected to occur outside their
geographical range due to environmental and/or biotic
constraints. Similarly, decreased recruitment is considered a
primary mechanism forcing changes in distributional ranges
and driving plant populations to local extinction (Bond
1995). As seed recruitment is one of the most critical
processes in the life-history of plants (Kitajima and Fenner
2000), knowledge of this process may help predict species
range constraints and trends in range shifts. However,
recruitment is an extremely complex process involving
successive life stages and biological mechanisms (Herrera
2000a). Therefore, to fully understand the ecological limits
of plant distributions, we need to examine the multiple
subprocesses involved, such as flower production, pollination, fruit development, seed dispersal, germination and
seedling survival. All of these subprocesses are affected
primarily by the physiological status of the plant and also by
a complex set of abiotic factors and biotic interactions, such
as flower and seed predation, and pollinator and seed
dispersal mutualisms. However, most studies have only
focused on single effects of a small number of abiotic or
biotic factors (Louda 1982, Horvitz and Schemske 1984,
Galen 1990, Galen and Butchart 2003) which may lead to a
biased perspective of this complex process. Few of them
have provided an integrated, sequential view of recruitment
probabilities from the ovule to the seedling stage (Herrera
2000a, Gulias et al. 2004). Comprehensive, sequential
assessment of recruitment may be the best approach for
addressing regeneration constraints, as harsh abiotic impacts
and/or biotic stress acting in just one step can be responsible
for hindering a species’ presence at a particular site.
Furthermore, factors that are favourable for a certain
recruitment stage may be unfavourable for another (e.g.
discrepancies between factors affecting emergence versus
those affecting seedling survival, de la Cruz et al. in press).
In this context, special interest has recently arisen on how
plants respond to climate warming. Many studies have
explored reproductive stages and processes along environmental gradients and their response to climate warming
(Louda 1982, 1983, Scherff et al. 1994, Louda and Potvin
1995, Dunne et al. 2003, Kudo and Hirao 2006), but they
lack a sequential, integrated approach.
High mountain habitats are especially useful for evaluating the impacts of climate change because they provide
731
sharp altitudinal gradients for testing ecological responses of
biota to geophysical influences (Körner 2007). Among
them, Mediterranean mountain environments impose both
harsh summer drought and cold winter conditions that
severely limit plant reproduction and growth (Cavieres et al.
2005, 2007, Ramı́rez et al. 2006, Giménez-Benavides et al.
2007a). High mountain plants are also presumably less
exposed to biotic pressures, like those exerted by hervibores
or pathogens, as biological activity has been found to
decrease with increasing altitude (Molau et al. 1989, Galen
1990, Kelly 1998, Scheidel et al. 2001).
The purpose of our study was to assess the factors that
determine the altitudinal range limits of Silene ciliata, a
long-lived high mountain Mediterranean plant, in its
southernmost distribution limit, and to identify the most
critical life stages in the recruitment process. Thus, we
evaluated the factors affecting each stage of the regeneration
process from the ovule to the established seedling, and
calculated transition probabilities along the altitudinal range
of the species in central Spain for two years. Comparison of
marginal and central populations along an altitudinal
gradient may help us to determine the factors that limit
the altitudinal range of the species and predict trends in
altitudinal shifts due to global warming. Specifically, we
hypothesized that the biotic and abiotic factors affecting
recruitment would vary along the altitudinal range of the
species. We expected recruitment to be increasingly controlled by biotic factors and water deficit as we move down
the mountain, reaching a maximum at the lower altitudinal
edge, which would eventually force local upward displacement.
Methods
Study species and study sites
Silene ciliata (Caryophyllaceae) is a long-lived high mountain plant which occurs in the main mountain ranges in the
northern half of the Mediterranean basin. This species
forms cushions of up to 2 cm in height and 15 cm in
diameter. It flowers in late summer, with a peak in early
August (Giménez-Benavides et al. 2007a). Flowering stems
have 15 flowers and fruit capsules have up to 100 seeds
that are passively dispersed by stalk vibration in August
September. Flowers are mainly predated by the blister beetle
Mylabris sobrina (Coleoptera: Meloidae) which nibbles
petals and carpels preventing pollination, and occasionally
by nectar-thieving ants (Hymenoptera: Formicidae)
(Giménez-Benavides unpubl.). Fruits are almost exclusively
consumed by the larvae of Hadena consparcatoides (Lepidoptera: Noctuidae) and, rarely, by weevils (Coleoptera:
Curculionidae). Hadena consparcatoides is an Iberian endemic moth that lays its eggs in S. ciliata flowers when
pollinating (the so-called nursery pollination, GiménezBenavides et al. 2007c). Therefore, this moth species can act
as a mutualist or parasite depending on the tradeoff between pollination and seed predation (GiménezBenavides et al. 2007c). Silene ciliata plants are rarely
grazed by domestic cows, and dispersed seeds may be
predated by harvester ants (Giménez-Benavides unpubl.).
732
We selected three populations at different altitudes in
Sierra de Guadarrama (Peñalara Natural Park, 408N, 38W),
a mountain range located 50 km north of the city of
Madrid where the species reaches its southernmost limit.
The lowest altitude population was located at 1976 m in a
moraine at Hoya de Peñalara at the Pinus sylvestris treeline
and constitutes the lower regional boundary of the species,
where it occurs in very fragmented, small subpopulations.
The medium altitude population was located at 2256 m on
the Dos Hermanas summit, composed by Festuca curvifolia
fellfield and Cytisus-Juniperus shrub formations, where
S. ciliata becomes abundant and forms large populations.
The third population was located at 2428 m on the
Peñalara summit, the highest peak of Sierra de Guadarrama.
Here, Cytisus-Juniperus clumps are relatively scarce and
the Festuca fellfield with S. ciliata dominates the landscape.
This altitudinal gradient covers the overall altitudinal range
of S. ciliata in Sierra de Guadarrama. As these populations
represent the southernmost limit of this species’ distribution, they may be especially sensitive to changes in climate
and land use. Sierra de Guadarrama is characterized by a
Mediterranean-type climate. Mean annual precipitation in
the Navacerrada Pass (1890 m) is 1350 mm, concentrated
from early October to late May followed by a pronounced
drought season from June to September. Temperature
and rainfall varied considerably in the two years of study.
Temperatures and summer rainfall were average in 2002,
while an extreme heatwave in 2003 caused a relevant
increase in maximum temperatures throughout the summer
(Giménez-Benavides et al. 2007a). In the past 45 yr average
annual temperatures have increased by ca 1.88C, while
annual precipitation has remained constant. However, snow
cover duration during the growing season has decreased
19.7 d (Giménez-Benavides et al. 2007a). As a consequence,
the distribution and fitness of animal and plant species in
this area have been significantly affected (Sanz et al. 2003,
Sanz-Elorza et al. 2003, Wilson et al. 2005).
Sampling design
Flower predation, pollination and fruit predation
We randomly established three study plots (1012 m2)
spaced 1050 m in each population (altitude). In each plot,
5560 adult individuals (2 cm in diameter) were
randomly tagged. Cushions located at a minimum distance
of 2 cm from each other were considered different individuals. As the species does not propagate vegetatively, each
labelled plant was considered a genet. Every week throughout two growing seasons (2002 and 2003) we recorded the
number of open flowers and mature fruits on each plant, as
well as the number of flowers and fruits with signs of
predation. Predated flowers presented nibbled petals and
styles, and the gynoecium was also occasionally affected.
Predated fruits were in most cases apparently intact from
the outside but had a larva developing inside when opened.
Sometimes fruits presented a hole produced by the exit of
the larva after eating the seeds. All mature fruits were
collected from each plant before opening to avoid seed
dispersal, and seed number, non-fertilized ovules and
larval predation were recorded in the laboratory. Fruit set
(proportion of total flowers setting fruit) and seed set
(proportion of total ovules setting seed) were calculated for
each plant.
To assess whether the reproductive output of S. ciliata
was affected by pollen limitation, we conducted a handpollination experiment in the field. In August 2003 we
randomly chose 90 plants from a single population located
in the middle of the local altitudinal gradient (2100 m,
1 km away from the Dos Hermanas population). The
following treatments were randomly assigned to each plant:
1) passive autogamy: complete flowers were bagged before
anthesis, 2) geitonogamy: pollen collected from one flower
was applied to other flowers of the same plant with a fine
paintbrush, 3) intrapopulational xenogamy: flowers from
one plant were pollinated with a pollen mixture from 10
plants of the same population, located at a minimum
distance of 10 m, 4) interpopulational xenogamy: flowers
from one plant were pollinated with a pollen mixture from
10 plants of the Dos Hermanas population within 2 h after
pollen collection. Pollen mixtures were stored in 2 ml
plastic vials until use. Non-manipulated flowers were used
as free-pollinating controls. In the geitonogamy and
xenogamy treatments, anthers were removed before the
onset of flowering and flowers were bagged with small
pieces of anti-pollen mesh (3M† , St. Paul, MN) to exclude
floral visitors. Flowers were monitored until withering or
fruit development. Fruit set and seed number per capsule
were obtained for each treatment.
Seedling emergence and survival
In 2002 and 2003, we collected mature seeds from 20 to 25
randomly selected mother plants in each population. Each
year, seeds from each population were pooled together and
stored at room temperature until sowing experiments were
carried out. Seeds from 2002 were sown in May 2003
(immediately after snowmelt), whereas seeds from 2003
were sown in September 2003 (before the winter season).
On each sowing date, four 1616 cm quadrats were
randomly established in small bare ground areas in each
population. Fifty full-sized seeds, randomly selected from
the pooled seeds, were slightly buried at 3 cm intervals in
each quadrat in their population of origin. Seedling
emergence and survival were monitored every 10 d during
the snow-free season in 2004, and at the beginning and end
of the season in 2005. Summer drought was assigned as the
cause of death to dried-out seedlings without any other
visible damage, herbivore/pathogen attack to dead seedlings
with external signs of predation such as cotyledon removal,
and winter frost to those seedlings that disappeared during
winter or appeared uprooted after snowmelt. Additional
details on the sowing method are provided in GiménezBenavides et al. (2007b).
Calculation of transition probabilities (TPs)
We evaluated the probability of an ovule becoming a 2-yrold seedling by decomposing it into a series of sequential
subprocesses and intermediate stages (Fig. 1) and estimating
the transition probabilities of each subprocess. ‘‘Ovules’’
were considered those present in the flowers of a mature
plant, and ‘‘non-predated ovules’’ those remaining after
flower predation. Similarly, ‘‘seeds’’ were considered those
found in full-sized fruits after successful pollination, while
‘‘non-predated seeds’’ were those remaining after pre-
Figure 1. Stages (rectangles), subprocesses (ovals) and some factors
influencing subprocesses (left-hand side) from the ovule to the
seedling stage of the high mountain cushion plant S. ciliata. TP:
transition probabilities; OPR: overall probability of recruitment.
dispersal seed predation. Finally, seedlings were classified
into three categories: ‘‘emerged seedlings’’, ‘‘1st summer
seedlings’’ and ‘‘2nd summer seedlings’’, corresponding to
seedlings initially emerging from the ground, and those
surviving after the first summer and second summer,
respectively.
Then, transition probabilities were defined as follows:
1) TP1 was the probability of an ovule escaping flower
predation. Since ovules are packed in flowers and predated
flowers do not develop any seed, this probability is
equivalent to the probability of a flower escaping predation.
We calculated this probability assuming that the number of
ovules did not differ between predated and non-predated
flowers. 2) TP2 was the probability of a non-predated ovule
being successfully fertilized and developing into a seed, i.e.
the proportion of non-predated ovules that set seed. It thus
combines fruit set and seediness (Herrera 2000a). 3) TP3
was the probability of a seed escaping predispersal predation
by H. consparcatoides larvae. As each larva needs more than a
single fruit to complete its development, it consumes all the
seeds contained in one fruit before moving to the next
733
(Giménez-Benavides et al. 2007c). Therefore, the probability of a seed escaping predation was equivalent to the
probability of a fruit escaping predation, assuming that seed
number did not differ between predated and non-predated
fruits. 4) TP4 was the probability of seedling emergence (i.e.
the ratio of the number of emerged seedlings to the number
of seeds sown in each population). 5) TP5 was the
probability of a seedling surviving from emergence to the
end of the first summer. 6) TP6 was the probability of a
seedling surviving from the end of the first summer to the
end of the second summer, thereby becoming a 2-yr-old
plant.
The first three TPs were calculated at the plant level, and
mean values per plot (three per population) were used in the
statistical analysis. The last three TPs were estimated at the
sowing quadrat level (four per population). The product of
the stage-specific transition probabilities (TPs) from the
ovule to a given stage at plot level provided the cumulative
probability of recruitment (CP) whereas the overall probability of recruitment (OPR), that is, the probability of an
ovule becoming a 2-yr-old plant was obtained by multiplying all the transition probabilities involved (TPs). It
should be noted that TP4, TP5 and TP6 were estimated
only from the cohort of seeds produced in 2003, as
emergence was negligible in the sowing trials carried out
in 2002 due to an inadequate sowing date.
Silene ciliata shows considerable interpopulational differences in the percentage of adult plants that flower each
year, mainly due to environmental conditions (GiménezBenavides et al. 2007a). This trait may be relevant in
recruitment dynamics since the ratio of established seedlings
per adult plant will be very low in years or populations with
few flowering individuals. Therefore, mean recruitment per
plant (MRP) at each plot and year was calculated as follows:
MRP Fp FlowersOvulesOPR
where Fp is mean flowering probability per plot, Flowers is
the mean number of flowers per plant (at the plot level),
Ovules is the mean number of ovules per flower (at the
plot level), and OPR is the overall probability of recruitment. To assess to what extent the estimated values of
recruitment per plant fit real recruitment values at the field
sites, we carried out an additional census. In September
2003 (at the end of the growing season), we established five
1-m2 plots at each population and counted the number of
adult plants and seedlings (isolated rosettes with cotyledons
or less than five pairs of leaves). Recruitment values in the
field were then calculated as the number of seedlings per
adult plant.
Data analysis
Differences in transition probabilities and cumulative
probability of recruitment between populations, considering three plots per population, were analyzed by nonparametric Kruskal-Wallis tests, followed by Nemenyi
post-hoc tests to perform multiple comparisons among
populations (Zar 1999). The Bonferroni method was used
to adjust the significance level for multiple comparisons.
Differences among treatments in the hand-crossing experiment were also analyzed by a Kruskal-Wallis test, followed
734
by a Nemenyi post-hoc test. Interannual differences in
transition probabilities with two years of data were analyzed
by Mann-Whitney tests. Statistical analyses were performed
using the SAS (SAS Inst., Cary, NC) and SPSS (SPSS,
Chicago) statistical packages.
Results
We observed significant altitudinal differences in flowering
probability, with high flowering rates at the medium and
high-altitude sites and low flowering rates at the low site,
especially in the second year (Fig. 2). Flower, fruit and seed
production per plant varied significantly among sites and
between years. In general, reproductive output was the
greatest at the high-altitude population and the lowest at the
population located at the treeline, especially in the second
year of the study due to the severity of summer drought
(Giménez-Benavides et al. 2007a). Statistical differences
among populations for each transition probability and each
stage are shown in Table 1, whereas post-hoc tests are
depicted in Fig. 2 and 4.
Interannual differences (p B0.05) were only found in
the flowering probability at the lowest altitude population,
the probability of an ovule escaping flower predation (TP1)
at the highest altitude population, and the probability of a
seed escaping predispersal predation (TP3) at the intermediate population.
Flower predation, pollination and fruit predation
Of the 502 tagged plants, 17.1 and 9.2% were attacked by
florivores in 2002 and 2003, respectively, with some
differences among sites (Table 2). However, 94.8% of
produced flowers escaped predation by insects (TP1 in Fig.
2). The proportion of non-predated ovules setting seed
(TP2) ranged between 40 and 60%, and did not differ
significantly among sites (Fig. 2). Regarding pre-dispersal
seed predation, 93.5% of produced fruits escaped from
Hadena larvae across populations (TP3). The incidence of
both flower and fruit predation was always higher at the
low-altitude population (Table 2), but significant differences were only detected between populations at the low
and high-altitude sites in 2003 (Fig. 2).
Fruit set and seed set of the pollination experiment are
shown in Table 3. Non-manipulated bagged flowers
(passive autogamy) produced both the lowest fruit set and
seed set values. Interpopulational xenogamy had significantly higher fruit set than the geitonogamy and control
treatments, but intrapopulational xenogamy did not differ
from control flowers. However, no significant differences in
seed set were found among treatments.
Seedling emergence and survival
Results were obtained just for the year 2003. Seedling
emergence (TP4) started soon after snowmelt (MayJune)
and lasted until early July. Emergence was significantly
greater at the medium-altitude site than at the lower site,
whereas intermediate values were obtained at the high site as
revealed by the post-hoc test (Fig. 2). On average, only 26%
Figure 2. Partial fitness values of all subprocesses involved in the regeneration from the ovule to the seedling stage at low, medium and
high-altitude S. ciliata populations in Sierra de Guadarrama. Post-dispersal subprocesses were assessed for only one season (2003). Vertical
lines denote standard errors. Different letters denote significant differences between populations within each year (p B0.05).
of emerged seedlings survived until the end of the first
summer (TP5), and barely 10% reached the 2-yr-old stage
(TP6). The causes of seedling mortality were summer
drought (85.8% of dead plants), winter frost (10.7%) and
herbivore/pathogen attack (3.5%). The low-altitude population recorded the lowest seedling survival both after the
first and the second summer (Fig. 2).
Overall probability of recruitment and seedling
density in the field
The cumulative probability of recruitment per plant is
represented in Fig. 3. The Kruskal-Wallis test by ranks
found significant differences among populations, mainly in
the post-dispersal stages of the recruitment process corresponding to the sowing experiment performed the second
year. The most significant differences in the probability of
setting an established seedling were found between the low
and the medium-altitude populations. However, the high
and medium-altitude populations did not differ significantly at any stage. The cumulative probability of recruitment declined sharply during fruit set, emergence and first
summer survival, denoting the most critical processes (Fig.
3). As a result, the overall probability of recruitment (OPR)
showed that the ovules produced in the medium-altitude
population had the highest chance of reaching the 2-yr-old
seedling stage, followed by those produced in the highaltitude population. However, when we estimated mean
recruitment per plant, the high and medium-altitude
populations yielded similar numbers of seedlings (Fig. 3).
Moreover, mean recruitment per plant estimated directly in
the field plots showed the same altitudinal pattern of
recruitment (Fig. 4).
735
7.261
2
0.027
7.200
2
0.027
7.200
2
0.027
2.756
2
0.252
1.156
2
0.561
6.252
2
0.044
5.600
2
0.061
5.956
2
0.051
6.489
2
0.039
5.956
2
0.051
5.067
2
0.079
1.422
2
0.491
0.356
2
0.837
7.200
2
0.027
2003
x2
DF
Sig.
5.067
2
0.079
0.356
2
0.837
1.689
2
0.430
0.157
2
0.925
0.800
2
0.670
7.200
2
0.027
1.067
2
0.587
5.956
2
0.051
2002
x2
DF
Sig.
2-summer
seedling
1-summer
seedling
Emerged
seedling
Non-pred.
seed
Seed
Non-pred.
ovule
MRP
OPR
TP6
TP5
TP4
TP3
TP2
TP1
Flow. prob.
Table 1. Kruskal-Wallis test for differences between populations in each transition probability and each stage, including flowering probabilty (Flow. prob.), overall probability of recruitment (OPR) and
mean recruitment per plant (MRP). Nemenyi post-hoc tests are depicted in Fig. 2 and 3.
736
Discussion
Our results show that the overall probability of recruitment
in S. ciliata is determined by a series of factors acting at
sequential stages, with effects on each of them varying from
negative to positive. Although many previous studies have
explored the importance of seedling establishment in alpine
environments (Chambers 1995, Galen and Stanton 1999,
Niederfriniger and Erschbamer 2000, Forbis 2003, Cavieres
et al. 2007) none of them have assessed the ultimate causes
of variation among populations by using a multiphase
approach. For instance, the most critical subprocesses
limiting recruitment mainly rely on abiotic factors affecting
seedling emergence and survival (specially water availability). However, a slightly decreasing trend in flower and seed
predation with increasing altitude might also condition
recruitment (Fig. 2), as previous works along altitudinal
gradients have suggested (Molau et al. 1989, Galen 1990,
Kelly 1998, Scheidel et al. 2001). In our case, neither flower
nor fruit predation significantly affected overall recruitment
of the species.
Florivory (i.e. herbivory focused on floral tissues) can be
especially harmful for plant reproductive success, because it
combines the direct reduction of ovules and the indirect
effect of reduced flower display for insect attraction
(Schemske and Horvitz 1988, Herrera 2000b, McCall
and Irwin 2006). In alpine environments, flower predation
is common and sometimes causes significant reductions in
fecundity (Norment 1988, Galen and Butchart 2003).
However, our results showed that flower predation by the
blister beetle M. sobrina only slightly reduced S. ciliata seed
output and flower predation was not significantly different
along the elevational gradient.
Only 50% of non-predated flowers across populations
produced seeds, suggesting certain pollinator limitation on
fruiting success. Our hand-pollination experiment revealed
that pollen from distant plants yielded a higher number of
fruits than control plants, whereas control plants did not
differ from intrapopulational crosses, suggesting some
degree of inbreeding depression rather than pollen limitation. Pollinator scarcity and the consequent pollen limitation have traditionally been considered for plant sexual
reproduction in high mountain environments (Bliss 1971,
Kevan and Baker 1983, Totland 1994). However, our
results suggest that the probability of an ovule becoming a
seed may be more dependent on factors other than
pollinator effectiveness, such as resource limitation or
environmental harshness. In a previous study, GiménezBenavides et al. (2007a) found that low fruit production
was due to the late-flowering pattern of S. ciliata, which
limited the use of snowmelt moisture during reproduction,
and the subsequent summer drought.
Pre-dispersal seed predation has also been identified as a
detrimental subprocess of plant reproductive success (Louda
1982, Louda and Potvin 1995), even in mountain
environments (Molau et al. 1989, Freeman et al. 2003,
Weppler and Stöcklin 2006). Our results suggest that most
S. ciliata seeds escape pre-dispersal seed predation, so this
factor, at least for the two years of this study, does not
actually represent a severe limitation for seedling recruitment. However, seed predation by H. consparcatoides may
be underestimated in our study because Hadena larvae
Table 2. Incidence of flower and fruit predation on low, medium and high-altitude S. ciliata populations in Sierra de Guadarama (Madrid,
Spain).
2002
2003
Low
Medium
High
Mean
Low
Medium
High
Mean
Florivory
% plants attacked (n502)
% flowers predated (n5796)
15.7
8.1
14.5
5.1
21.1
6.0
17.1
6.2
12.0
10.2
10.9
6.2
4.7
0.7
9.2
3.4
Pre-dispersal seed predation
% plants attacked (n502)
% fruits predated (n1851)
16.3
16.3
7.3
4.7
9.4
4.1
11.0
7.4
5.4
16.5
4.8
6.4
5.8
2.9
5.4
5.2
typically move to other host fruits once they have eaten the
contents of the fruit where the egg was laid (GiménezBenavides et al. 2007c). Since we collected fruits before seed
dispersal to calculate seediness, some moth larvae were
removed early from plants preventing subsequent fruit
predation. Therefore, pre-dispersal seed predation could
actually be higher and present an important source of
propagule loss in S. ciliata.
The subsequent stages of seedling emergence and
survival had a significant impact on the cumulative
probability of recruitment, and accounted for most of the
differences observed among populations (Fig. 3). Although
seedling emergence was only measured in the field in one
year, the obtained values were well below the potential
germination capacity of the species under laboratory
conditions (low altitude: 68%, medium altitude: 78% and
high altitude: 58%, Giménez-Benavides et al. 2005). This
suggests that local environmental factors played a major role
in emergence success. The medium-altitude population had
greater seedling emergence and, subsequently, a greater
overall probability of recruitment than the high-altitude
population, even though their seedling survival probabilities
were similar (Fig. 3). Abiotic factors, especially drought,
were the major cause of low seedling survival probability,
particularly in the low-altitude population. Several authors
have recently pointed out the existence of a severe water
shortage in high Mediterranean mountain environments
and its negative effect on plant regeneration (Castro et al.
2004, 2005, Lloret et al. 2004, Cavieres et al. 2005). In our
case, soil moisture measurements at the experimental
sowing sites showed an extreme water shortage during
mid-summer, which accounted for most S. ciliata seedling
deaths (Giménez-Benavides et al. 2007b). This summer
drought was especially intense at the low-altitude population.
Recruitment per adult plant was notably higher in
the medium and high-altitude populations compared to
the low-altitude population where recruitment values were
around five-fold lower (Fig. 3). Although the mediumaltitude population reached a higher probability of recruitment per ovule, the high-altitude population yielded a
similar number of 2-yr-old seedlings per adult plant due to
both higher flowering probability and higher flower
production (Fig. 2). Recruitment per plant values estimated
by means of transition probabilities were significantly
different from direct censuses in the field plots, although
the altitudinal gradient pattern was similar (Fig. 3, 4). This
suggests that some steps of the recruitment process that
were not directly considered in our study may also play a
relevant role. For instance, post-dispersal seed fates (e.g.
predation and seed arrival to safe sites) are decisive
subprocesses of the regeneration cycle that may also cause
significant reductions in the reproductive output of perennial plants. Some studies have assessed the importance of
sequential post-dispersal stages on plant recruitment, sowing that seed predation is one of the most important
botlenecks (Rey and Alcántara 2000, Traveset et al. 2003,
Gulias et al. 2004, Lázaro et al. 2006, Rodrı́guez-Pérez and
Traveset 2007). In fact, both subprocesses have been
reported to be important sources of seed loss in alpine
environments (Chambers 1995, Muñoz and Cavieres
2006).
This study illustrates how the combination of observational and experimental surveys may help detect the most
critical stages in the seed regeneration process, how they
change along the altitudinal range of a mountain plant and,
more specifically, what factors determine the final ouput in
each population. This approach provides a very useful,
standardized measure (i.e. the probability of an ovule
becoming an established seedling) for comparing regeneration patterns under contrasting ecological scenarios, like
peripheral vs central populations, disturbance regimes,
management practices, etc. Although the time-consuming
nature of these surveys limited our study to three populations to describe the altitudinal pattern, results were robust
and led to relevant conclusions on the limiting factors of
recruitment in the studied system.
In summary, our results showed that the most critical
subprocesses of recruitment in S. ciliata were flowering
probability of adult plants (very poor at the lower boundary), seed production, seedling emergence and seedling
Table 3. Fruit set and number of seeds per fruit after different
pollination treatments in Silene ciliata. Superscripts denote Nemenyi post-hoc test. Treatments with the same letter do not differ
significantly (p0.05).
Pollination treatment
Passive autogamy
Geitonogamy
Intrapopulation
xenogamy
Interpopulation
xenogamy
Control
Flowers
(n)
Fruit set
Seed set
49
33
33
0.3090.07a
0.3990.12b
0.4590.16cd
0.1490.15a
0.3090.22b
0.2790.21ab
30
0.6490.23c
0.2090.16ab
44
0.4190.13bd
0.2390.13ab
737
Figure 3. Cumulative probability of recruitment of low, medium and high-altitude S. ciliata populations, calculated by multiplying all
the transition probabilities at each site for each year of study. Small plot represents mean recruitment per plant (number of ovules
becoming 2-yr-old seedlings per S. ciliata adult plant). Vertical lines denote standard errors. *Differences between low and high altitude
populations at pB0.05, **differences between low and medium-altitude populations at pB0.025. Separate analyses for each year.
survival. These processes were highly dependent on local
environmental conditions, and especially on summer
drought which is exacerbated as altitude decreases
(Giménez-Benavides et al. 2007a, b). Except for pollination
which was not specifically measured in this study, biotic
Figure 4. Direct field estimation of mean recruitment per plant
(number of seedlings per adult plant) in low, medium and highaltitude S. ciliata populations. Different letters denote significant
differences between populations (pB0.05).
738
interactions such as herbivory were less restrictive for sexual
regeneration than abiotic factors. This is a common feature
of declining plant populations in their southernmost range
(Eriksson 1996, Garcı́a et al. 1999, 2000, Hampe and
Arroyo 2002). As a result, the small, isolated populations
located at the lower-altitudinal edge of the species (those
most affected by summer drought) showed extremely lower
recruitment probabilities and may, therefore, be highly
affected by a recruitment shortage. Since this species is
unable to reproduce vegetatively, the long-term persistence
at the lower-altitude edge will ultimately depend on the
longevity of adult plants. Sanz-Elorza et al. (2003) reported
increasing density and upward displacement of the CytisusJuniperus shrub community in this mountain range, as a
result of climate warming and land-use change. This
community typically occurs interspersed in the lower and
intermediate populations of S. ciliata, which is unable to
grow beneath these shrubs. Therefore, in addition to the
direct effects of climate harshness on the recruitment of
S. ciliata, competition with shrubs may also promote the
exclusion of this species from its current lower limit.
Acknowledgements We are grateful to the staff of Parque Natural
de las Cumbres, Circo y Lagunas de Peñalara for permission to
carry out this study, and to R. Garcı́a, C. Fernández, A. L.
Luzuriaga and M. J. Albert for help with the field work. M. Garcı́a
Parı́s helped to identify the blister beetle species. We also thank
M. J. Albert for helpful comments, and L. De Hond for linguistic
assistance. This work was supported by the Spanish Government
CICYT project EXTREM (CGL2006-09431).
References
Bliss, L. C. 1971. Arctic and alpine plant life cycles. Annu. Rev.
Ecol. Syst. 2: 405438.
Bond, W. J. 1995. Assessing the risk of plant extinction due to
pollinator and disperser failure. In: Lawton, J. H. and May,
R. M. (eds), Extinction rates. Oxford Univ. Press, pp. 131
146.
Castro, J. et al. 2004. Seedling establishment of a boreal tree
species (Pinus sylvestris) at its southernmost distribution limit,
consequences of being in a marginal, Mediterranean habitat.
J. Ecol. 92: 266277.
Castro, J. et al. 2005. Alleviation of summer drought boots
establishment success of Pinus sylvestris in a Mediterranean
mountain: an experimental approach. Plant Ecol. 181: 191
202.
Cavieres, L. A. et al. 2005. Nurse effect of the native cushion plant
Azorella monantha on the invasive non-native Taraxacum
officinale in the high-Andes of central Chile. Perspect. Plant
Ecol. Evol. Syst. 7: 217226.
Cavieres, L. A. et al. 2007. Microclimatic modifications of cushion
plants and their consequences for seedlings survival of native
and non-native plants in the high-Andes of central Chile.
Arct. Antarct. Alp. Res. 39: 229236.
Chambers, J. C. 1995. Relationships between seed fates and
seedling establishment in an alpine Ecosystem. Ecology 76:
21242133.
de la Cruz, M. et al. in press. Where do the seedlings go? A spatiotemporal analysis of seedling mortality in a semiarid gypsophyte. Ecography.
Dunne, J. A. et al. 2003. Subalpine meadow flowering phenology
responses to climate cange: integrating experimental and
gradient methods. Ecol. Monogr. 73: 6986.
Eriksson, O. 1996. Regional dynamics of plants: a review
of evidence for remnant, source-sink and metapopulations.
Oikos 77: 248258.
Forbis, T. A. 2003. Seedling demography in an alpine ecosystem.
Am. J. Bot. 90: 11971206.
Freeman, R. S. et al. 2003. Flowering phenology and compensation for herbivory in Ipomopsis aggregata. Oecologia 136:
394401.
Galen, C. 1990. Limits to the distribution of alpine tundra plants,
herbivores and the skypilot, Polemonium viscosum. Oikos 59:
355358.
Galen, C. and Stanton, M. L. 1999. Seedling establishment in
alpine buttercups under experimental manipulations of growing-season length. Ecology 80: 20332044.
Galen, C. and Butchart, B. 2003. Ants in your plants: effects of
nectar-thieves on pollen fertility and seed-siring capacity in the
alpine wildflower, Polemonium viscosum. Oikos 101: 521
528.
Garcı́a, D. et al. 1999. Age structure of Juniperus communis L. in
the Iberian peninsula: conservation of remnant populations in
Mediterranean mountains. Biol. Conserv. 87: 215220.
Garcı́a, D. et al. 2000. Geographical variation in seed production,
predation and abortion in Juniperus communis throughout its
range in Europe. J. Ecol. 88: 436446.
Giménez-Benavides, L. et al. 2005. Seed germination of high
mountain Mediterranean species, altitudinal, interpopulation
and interannual variability. Ecol. Res. 20: 433444.
Giménez-Benavides, L. et al. 2007a. Reproductive limits of a lateflowering high-mountain Mediterranean plant along an elevational climate gradient. New Phytol. 173: 367382.
Giménez-Benavides, L. et al. 2007b. Local adaptation enhances
seedling recruitment along an altitudinal gradient in a high
mountain Mediterranean plant. Ann. Bot. 99: 723734.
Giménez-Benavides, L. et al. 2007c. Generalist diurnal pollination
provides greater fitness in a plant with nocturnal pollination
syndrome: assessing the effects of a Silene-Hadena interaction.
Oikos 116: 14611472.
Gulias, J. et al. 2004. Critical stages in the recruitment process of
Rhamnus alaternus L. Ann. Bot. 93: 723731.
Hampe, A. and Arroyo, J. 2002. Recruitment and regeneration in
populations of an endangered south Iberian Tertiary relict tree.
Biol. Conserv. 107: 263271.
Herrera, C. M. 2000a. Flower-to-seedling consequences of
different pollination regimes in an insect-pollinated shrub.
Ecology 81: 1529.
Herrera, C. M. 2000b. Measuring the effects of pollinators
and herbivores: evidence for non-additivity in a perennial
herb. Ecology 81: 21702176.
Horvitz, C. C. and Schemske, D. W. 1984. Effects of ants and an
ant-tended herbivore on seed production of a neotropical herb.
Ecology 65: 13691378.
Kelly, C. A. 1998. Effects of variable life history and insect
herbivores on reproduction in Solidago macrophylla (Asteraceae) on an elevational gradient. Am. Midl. Nat. 139: 243
254.
Kevan, P. G. and Baker, H. G. 1983. Insects as flower visitors and
pollinators. Annu. Rev. Entomol. 28: 407453.
Kitajima, K. and Fenner, M. 2000. Ecology of seedling regeneration. In: Fenner, M. (ed.), Seeds: the ecology of regeneration
in plant communities. CAB International, pp. 331359.
Körner, C. 2007. The use of ‘altitude’ in ecological research.
Trends Ecol. Evol. 22: 569574.
Kudo, G. and Hirao, A. 2006. Habitat-specific responses in the
flowering phenology and seed set of alpine plants to climate
variation: implications for global-change impacts. Popul.
Ecol. 48: 4958.
Lázaro, A. et al. 2006. Spatial concordance at a regional scale in
the regeneration process of a circum-Mediterranean relict
(Buxus balearica): connecting seed dispersal to seedling
establishment. Ecography 29: 114.
Lloret, F. et al. 2004. Experimental evidence of reduced diversity
of seedlings due to climate modification in a Mediterraneantype community. Global Change Biol. 10: 248258.
Louda, S. M. 1982. Distribution ecology: variation in plant
recruitment over a gradient in relation to insect seed predation.
Ecol. Monogr. 52: 2541.
Louda, S. M. 1983. Seed predation and seedling mortality in the
recruitment of a shrub, Haplopappus venetus Blake (Asteraceae), along a climatic gradient. Ecology 64: 511521.
Louda, S. M. and Potvin, M. A. 1995. Effect of inflorescencefeeding insects on the demography and lifetime of a native
plant. Ecology 76: 229245.
McCall, A. C. and Irwin, R. E. 2006. Florivory: the intersection of
pollination and herbivory. Ecol. Lett. 9: 13511365.
Molau, U. et al. 1989. Predispersal seed predation in Bartsia
alpina. Oecologia 81: 181185.
Muñoz, A. A. and Cavieres, L. A. 2006. A multi-species
assessment of post-dispersal seed predation in the central
Chilean Andes. Ann. Bot. 98: 193201.
Niederfriniger, R. and Erschbamer, B. 2000. Germination and
establishment of seedlings on a glacier foreland in the central
Alps. Arct. Antarct. Alp. Res. 32: 270277.
Norment, C. J. 1988. The effect of nectar-thieving ants on the
reproductive success of Frasera speciosa (Gentianaceae). Am.
Midl. Nat. 120: 331336.
Ramı́rez, J. M. et al. 2006. Altitude and woody cover control
recruitment of Helleborus foetidus in a Mediterranean mountain area. Ecography 29: 375384.
739
Rey, P. J. and Alcántara, J. M. 2000. Recruitment dynamics of a
fleshy-fruited plant (Olea europaea): connecting patterns of
seed dispersal to seedling establishment. J. Ecol. 88: 622
633.
Rodrı́guez-Pérez, J. and Traveset, A. 2007. A multi-scale approach
in the study of plant regeneration: finding bottlenecks is not
enough. Perspect. Plant Ecol. Evol. Syst. 9: 113.
Sanz, J. J. et al. 2003. Climate change and fitness components of a
migratory bird breeding in the Mediterranean region. Global
Change Biol. 9: 461472.
Sanz-Elorza, M. et al. 2003. Changes in the high-mountain
vegetation of the central Iberian Peninsula as a probable sign of
climate warming. Ann. Bot. 92: 273280.
Scheidel, U. et al. 2001. Altitudinal differences in herbivory on
montane Compositae species. Oecologia 129: 7586.
Schemske, D. W. and Horvitz, C. C. 1988. Plant-animal
interactions and fruit production in a neotropical herb: a
path analysis. Ecology 69: 11281137.
740
Scherff, E. J. et al. 1994. Seed dispersal, seedling survival and
habitat affinity in a snowbed plant-limits to the distribution of
the snow buttercup, Ranunculus adoneus. Oikos 69: 405
413.
Totland, O. 1994. Intraseasonal variation in pollination intensity
and seed set in an alpine population of Ranunculus acris in
southwestern Norway. Ecography 17: 159165.
Traveset, A. et al. 2003. Transition probabilities from pollination
to establishment in a rare dioecious shrub species (Rhamnus
ludovici-salvatoris) in two habitats. J. Ecol. 91: 427437.
Weppler, T. and Stöcklin, J. 2006. Does pre-dispersal seed
predation limit reproduction and population growth in the
alpine clonal plant Geum reptans? Plant Ecol. 187: 277287.
Wilson, R. J. et al. 2005. Changes to the elevational limits and
extent of species ranges associated with climate change. Ecol.
Lett. 8: 11381146.
Zar, J. H. 1999. Biostatistical analysis. Prentice Hall.