Biologxal Journal of the Linnean Society (2001), 74: 475487. With 4 figures
doi:lO. 1006/bij1.2001.0600, available online at httpj//www.idealibrary.com on
ID E b
l@
c
Impacts of floral gender and whole-plant gender on
floral evolution in EebaZZium elaterium
(Cucurbitaceae)
@
DENISE E. COSTICH’ and THOMAS R. MEAGHER2*
‘Boyce Thompson Institute for Plant Research, Tower Road, Ithaca, NY 14853, U S A
‘Division of Environmental & Evolutionary Biology, University of S t Andrews, S t Andrews, Fife
KY16 9TH
Received 23 February 2001; accepted for publication 11 July 2001
Investigation of gender specialization in plants has led to several theories on the evolution of sexual dimorphism:
reproductive compensation, based on enhanced reproductive efficiency with gender specialization (flowers should
be larger on dioecious plants); Bateman’s Principle, based on sex-specific selection (display for pollinator attraction
in males and seed set in females); and intersexual floral mimicry, based on mimicry of a reward-providing gender
by a non-reward providing gender (reduced dimorphism in dioecious plants due to increased spatial separation of
male and female flowers). These theories were evaluated in Ecballium elaterium, which contains two subspecies,
elaterium (monoecious)and dioicum (dioecious). Our results show that flowers of the dioecious subspecies are larger
and allocate more to reproductive organs than do flowers of the monoecious subspecies. Both subspecies are sexually
dimorphic (male flowers larger than female flowers). Variance in flower size among populations is greater in the
dioecious subspecies. Finally, there is sufficient genetic variation to enable ongoing response t o selection; genetic
correlation constraints on independent response of female and male flowers may be stronger in the monoecious
subspecies. Our findings provide support for aspects of all three theories, suggesting that the evolution of floral
dimorphism is based on a complex interplay of factors.
0 2001 The Linnean Society of London
ADDITIONAL KEY WORDS: monoecy - dioecy - Bateman’s Principle - sexual dimorphism - reproductive
compensation - deceit pollination - Cucurbitaceae - Mediterranean flora.
INTRODUCTION
Unisexual flowers provide insight into selection pressures associated with either male or female function.
Sex-specificselection can lead to sexual dimorphism in
floral features, such as display for pollinator attraction
and resource allocation to sex organs that are optimized
differently for male and female reproductive success.
Monoecious species, in which unisexual flowers of both
genders occur on individual plants, provide a model
for the examination of sex-specific selection on floral
evolution a t the individual flower level. Dioecious species, in which whole plants are unisexual, provide a
model for examination of the consequences of sexspecific selection on floral evolution a t the whole-plant
level. In order to examine the consequences of sexspecific selection on floral morphology in these two
* Corresponding author. E-mail: [email protected]
0024-4066/01/120475 + 13 $35.00/0
different selective environments, one where sexes are
combined on individuals versus one where sexes are
separate, we chose as our study organism Ecballium
elaterium (Cucurbitaceae), comprising two subspecies,
elaterium, which is monoecious, and dioicum, which
is dioecious. The co-occurrence of these two breeding
systems within one species provides a n unusual opportunity to conduct a comparative study of their respective evolutionary consequences.
To provide a theoretical framework to predict how
unisexual flowers might be expected to diverge due
to both breeding-system and sex-specific patterns of
selection, we outline three conceptual models for natural selection (Table 1).‘Reproductive compensation’
predicts that the evolution of unisexuality should be
accompanied by increased resource allocation to the
remaining sex function. Compensation may occur
through the production of larger flowers, more flowers,
or both (Darwin, 1877; Charnov, 1982). In Ecballium,
475
0 2001 The Linnean Society of London
476
L).
E. COSTICH and T. R. MEAGHER
Table 1. Theories, mechanisms and predicted consequences for breeding-system- and sex-specificpatterns of selection
in floral evolution in Ecballium eluteriurn
Theoretical basis
for selection
Mechanism
Predicted consequences
Monoecy vs dioecy
Reproductive
compensation
Unisexual plants compensate for the loss
of one sex function through increase
allocation to the remaining sex function.
Hateman’s Principle
Male reproductive success is limited by
Conspecific floral
mimicry
pollinator attraction and concomitant
mating opportunities; female reproductive
success is limited by resources available
for seeqfruit production.
The rewardless floral morph (female)
mimics the rewarding floral morph (male).
this leads to the prediction t h a t flowers on the unisexual plants of the dioecious subspecies should be
larger, or more numerous, than those of the monoecious
subspecies. ‘Bateman’s Principle’ is a concept derived
from the study of sexual selection in fruit flies (Bateman, 1948; Rivers, 1972) t h a t has been embraced by
many plant reproductive biologists to explain floral
evolution in the context of sex-related differences in
the limits to reproductive success (reviewed by Arnold,
1994; Wilson et al., 1994; Campbell, 2000). Under
Bateman’s Principle, male function should be limited
by mating opportunities, whereas resources available
for seed production should limit female function. In
general, this leads to the prediction t h a t male flowers
will be larger to enhance pollinator attraction (Bell,
1986). The extent to which male and female flowers
evolve towards dimorphism will be constrained by
genetic correlation between male and female flowers
(Meagher, 1994,1999;Ashman, 1999). Since both floral
morphs co-occur on the same individuals in monoecious
species, it is expected t h a t this constraint would be
more pronounced in monoecious species (e.g. Agren
& Schemske, 1995). In Ecballium, this leads to the
prediction that the complete separation of the sexes in
dioecious populations should lead to increased sexual
dimorphism. ‘Intersexual Floral Mimicry’ acts in
animal-pollinated systems where a rewardless floral
morph engages in deceit by mimicking a rewarding
morph to ensure pollinator visits (Baker, 1976; Bawa,
1977, 1980b; Ferdy et al., 1998; Willson & Agren, 1989;
Schemske. Agren & Le Corff, 1996). In Ecballium,
female flowers do not produce nectar or pollen, whereas
male flowers produce both (Dukas, 1987; Fahn &
Shiniony, 2001). Thus, the prediction here would be
t.hat the floral morphs will not differ in either monoecious o r dioecious populations.
Since these different models make slightly different
Male vs female
Flowers of the unisexual
plants are larger or
more numerous than
those of cosexual plants.
More floral dimorphism
in the dioecious
subspecies.
Less floral dimorphism
in the dioecious
subspecies.
Female flowers differ
from male flowers.
Female flowers are
similar to male flowers.
predictions about relative flower morph sizes on monoecious and dioecious plants, we address the following
specific questions about floral evolution in Ecballium.
In terms of reproductive compensation: are flowers on
plants in dioecious populations larger than those in
monoecious populations? In terms of sex-specific selection versus intersexual floral mimicry: are flowers in
dioecious populations more (or less) sexually dimorphic
than those in monoecious populations? How is sexual
dimorphism distributed within and between monoecious and dioecious populations? Finally, what is the
genetic potential for ongoing floral response to selection?
MATERIAL AND METHODS
Ecballium elaterzum (L.) A. Rich. (Cucurbitaceae) is a
perennial plant native to the Mediterranean region.
As mentioned above, the species comprises two subspecies, elaterium and dioicum, which have different
breeding systems. These two subspecies are interfertile, and crosses between monoecious and dioecious plants have been used to demonstrate that
sex determination is genetically based (Galan, 1946;
reviewed in Westergaard, 1958).In populations of subspecies elaterium, all individuals possess, a t some point
in their lifetime, both male and female flowers. I t is
common for both flower types to be open on the same
day on a n individual, making geitonogamy possible.
The subspecies is self-compatible, and there is strong
evidence from our allozyme diversity study (Costich &
Meagher, 1992) t h a t the selfing rate is high in the
usually small (less t h a n 100 individuals) populations.
In subspecies diozcunz, in contrast, male and female
flowers are found on separate individuals, population
sizes are generally larger, and the enforced outcrossing
FLORAL EVOLUTION IN ECBALLIUM
477
Ecballium Populations
o
0
Dioecious
Monoecious
AA Alhama de Aragon
AT Atocha
CA Calicasas
CN Cordovin
CS Castuera Station
CT Castuera Town
CY Calatavud
IB Ibiza. "
JA Jaraiceio
LP Los PaIacios
PA Puebla de Acocer
RF Ribafrecha
SA Salamanca
S P Sevilla
TM Turre-Mojacar
TO Totana
TR Torrelodones
VA Valchillon
VL Valladolid
VN Villanueva
VS Villanueva de la Serena
0 100 200 300 400 500
Figure 1. Geographic distribution of Ecballium elaterium source populations in Spain. (.)Ecballium
eluteriurn (monoecious),(0)
=Ecballium eluteriurn subsp. dioicum (dioecious).
appears to have acted to maintain high levels of diversity and heterozygosity within populations. In addition to breeding-system differences, the two species
have different geographic distributions in Spain, with
the monoecious subspecies being found in relatively
mesic sites to the north, and the dioecious subspecies
in relatively xeric sites in the south (Costich & Galan,
1988). More details about Ecballium life history, taxonomy, distribution and genetics can be found elsewhere (Costich, 1989; Costich & Meagher, 1992;
Costich, 1995; Saavedra, 2000).
POPULATIONS SAMPLED
During the summer of 1986, a n extensive collecting trip
was made to locate Ecballium populations throughout
Spain (Fig. 1). Ten populations of each subspecies
were included in this data set, representing all the
populations mapped in Figure 1, with the exception of
SP (Sevilla), which was one of the source populations
for the common garden (see below). Samples of flowers,
leaves, and seeds were collected from 20 to 30 individuals from each population. Two to four fully open
flowers per individual were preserved together in vials
containing 70% ethanol.
Floral measurements were made with a digital calliper to the nearest millimeter (see Fig. 2). On all
flowers, the width and length of one petal and one
sepal and the width of the pedicel were taken. Due to
the complex morphology of the androecium, we decided
to use total dry weight to provide the best overall
measure of size for this part of the flower. The same
was true for the stigma plus style portion of the gynoecium of the female flowers. The androecium/gynoecium dry weights are considered approximate prereproductive sex allocation measures because we could
elaterium subsp.
not guarantee that all of the pollen was retained within
the anthers (androecium) and because, owing to the
morphology of female Ecballium flowers (inferior
ovary), the ovary was not considered homologous t o the
androecium and thus was excluded from the gynoecium
measurement.
COMMON-GARDENPLANTATIONS
Seeds collected from plants measured in five of the
monoecious [CT, CY, RF, SA, VL] and five of the dioecious populations [AT, CA, CS, SP, TO] were planted
in the greenhouse of the Department of Biological
Sciences, Rutgers University, Piscataway, New Jersey,
in Spring 1990 as part of an allozyme survey (Costich &
Meagher, 1992). They overwintered in the greenhouse
and were transplanted to a field plot adjacent to the
Waksman Institute on Busch Campus, Rutgers University, Piscataway, New Jersey, in May 1991.
The same measurements as described above were
made on fresh flowers collected from plants in this
field plot, so that parental measurements taken in
Spain were complemented by offspring measurements
taken in New Jersey. In addition to the ten characters
measured in the preserved flowers, we also measured
corolla diameter (Fig. 2) prior to detaching and transferring flowers to the lab for the remaining measurements. For each of the five monoecious populations
represented, six to eight individual plants were measured, two male plus two female flowers per plant. For
each of the five dioecious populations, six to ten male/
female sibling pairs were measured, two flowers per
plant.
We assume throughout this analysis that measurements based on preserved flowers are comparable
to measurements based on fresh flowers. Since those
478
D. E. COSTICH and T. R. MEAGHER
I PW I
I
Y
STIGMAS + STYLE
3Ide view
I!
STAMENS
MALE
Figure 2. Top and side views of female and male Ecballium elaterium flowers illustrating morphological features
common to dioecious and monoecious plants. (SL = sepal length; SW = sepal width; PL = petal length; PW = petal width;
STL = stamen length; STW = stamen width; PDW =pedicel width.)
measurements were made on different generations
grown under different conditions and subject to different statistical treatment, we assume that consequences of any differences between preserved and
fresh flowers will be negligible in our final interpretation.
SFATISTICAL ANALYSIS
Flower measurements were subjected to analysis of
variance (ANOVA) using the SAS software package
(Release 8.00, 1999, SAS Institute, Cary, NC, USA).
Floral characters common t o flowers of both sexes were
analysed to determine whether there were differences
between subspecies, differences among populations as
i-’ nested effect within subspecies, differences between
male and female flowers, o r differences between subspecies or among populations in the extent of sexual
dimorphism, which would be indicated by significant
interactions between subspecies and flower sex or
population and flower sex.
Heterogeneity in variance among the various levels
of analysis outlined above was assessed by performing
a Levene’s test (Brown & Forsythe, 1974), based on an
ANOVA of‘ the absolute values of the residuals from
the ANOVAs described above. ANOVA is robust to
moderate levels of heteroscedasticity (Sokal & Rohlf,
1981), so a detection of heterogeneity in variances does
not invalidate the results of our primary ANOVA.
In order t o summarize the overall impacts of various
characters acting in combination as sexually dimorphic
traits, we performed a principal components analysis
of the six floral traits, treating androecium and gynoecium (style plus stigma) dry weight as analogous
characters in male and female flowers. To illustrate the
interrelationship among the different floral characters,
we also did a correlation analysis.
Finally, 0ffspring:parent regressions (Falconer &
Mackay, 1996) of floral characters measured from
preserved flowers collected in the field in Spain, and
fresh flowers collected from their progeny raised in
New Jersey were used t o assess genetic parameters
using the following model: [offspring]= p[female
parent] +error. Heritability (h’) is equivalent to twice
the single (maternal) parent regression coefficient (13)
within characters (e.g. offspring sepal length and maternal sepal length); similarly, 0ffspring:parent regressions across characters (e.g. offspring sepal length
and maternal sepal width) were used to assess the
impact of genetic correlations among characters. 5‘ince
FLORAL EVOLUTION IN ECBALLIUM
479
Table 2. Summary of floral measurement data for natural populations of Ecballium elaterium (linear measurements
in mm; dry weights in mg); mean standard deviation (sample size)
Character
Sepal length
Sepal width
Petal length
Petal width
Gynoecium dry weight
Androecium dry weight
Pedicel width
Ecballium elaterium subsp. elaterium
Ecballium elaterium subsp. diocium
(monoecious)
(dioecious)
Male
Female
Male
Female
7.62 k 1.68 (98)
1.75 0.25 (98)
16.37i2.30 (98)
6.12+ 1.01 (98)
5.90f 1.38 (121)
1.29k0.22 (121)
13.62k2.25 (121)
5.48 f 1.05 (120)
1.06k0.33 (121)
10.35k2.20 (104)
1.87k0.44 (104)
17.09 f 3.33 (104)
7.48f 1.73 (104)
8.68k1.90 (86)
1.54 f0.43 (86)
14.76f3.02 (86)
6.92 i2.02 (86)
2.13+ 1.02 (86)
1.15i0.32 (98)
1.26f0.23 (87)
1.93k0.29 (121)
these characters were measured on mature plants in
both generations, we assume that maternal effects are
negligible. Given that offspring and parent measurements took place in different environments, these
regression coefficients are only approximate and are
used here as an indication of overall evolutionary
potential and not for specific estimation of h2 and
genetic correlation per se. For monoecious plants, maternaI parents in the field had both male and female
flowers, so that we could obtain heritabilities for both
male and female floral characters. For dioecious plants,
we could similarly obtain estimates of heritability only
for female floral characters. Genetic correlations include both relationships between different characters
and relationships between the same characters expressed in either males or females (Meagher, 1999).
The extent of genetic correlation between the sexes
can be assessed by means of female progeny:male
adult character (monoecious subspecies only) and male
progeny:female adult character regressions.
RESULTS
FLOWER SIZE
In terms of differences between subspecies, flowers
from dioecious plants were consistently larger and
allocated more resources (dry weight) to reproductive
structures than did flowers from monoecious plants
(Tables 2,3, Fig. 3). There was also extensive variation
among populations within subspecies for both flower
size and reproductive dry weight allocation, indicating
potential for evolutionary modification of these characters.
SEXUAL DIMORPHISM
The overall difference betweenmale and female flowers
was even more pronounced than subspecies differences, with male flowers having a larger corolla,
1.82i0.64 (104)
1.41k0.21 (92)
2.21 k0.45 (87)
(the display area for pollinator attraction) and females
having a more substantial pedicel (presumably for
supporting the fruit as it develops). We could only
measure corolla diameter in the fresh flowers collected
in our common-garden plots, but in those plants we
found a high correlation between petal length and
corolla diameter (Table 4). Thus, we can consider petal
length to be a n appropriate measure of floral display
for the field-collected preserved flowers. The larger
petal length in males indicates a more substantial
floral display for flowers of that sex. In general, sexual
dimorphism was consistent between male and female
flowers from monoecious and dioecious plants, i.e. there
were no characters that showed a significant subspecies x flower sex interaction.
One character that was very similar for the two
sexes was dry weight allocation to reproduction (Table
3, Fig. 3), excluding the inferior ovary. Thus, it appears
that male and female flowers allocate similar resources
to pre-pollination reproductive structures excluding
the inferior ovary. Other than the lack of sex difference
in dry weight allocation, this character was highly
variable among populations, with a significant difference between subspecies (flowers on plants of the
dioecious subspecies have a higher dry weight allocation to reproduction per flower).
DISTRIBUTION OF SEXUAL DIMORPHISM IN
MONOECIOUS AND DIOECIOUS POPULATIONS
The two subspecies showed differences in levels of
variance, with the dioecious subspecies being more
variable for most traits than the monoecious subspecies
(Fig. 3, Table 5). Dry weight allocated to reproductive
structures also showed higher variance in the dioecious
subspecies as well.
The principal components (PC) analysis yielded two
predominant axes that accounted for 73% of the overall
variance in floral structures (Table 6, Fig. 4). The first
480
D. E. COSTICH and T. R. MEAGHER
Table 3. Analysis of variance of floral characters. In most cases, the pattern of significant effects called for the use of
an error MP other than Error; in those cases, the appropriate error MS that was used is as indicated in the second
column. npe-I11 sums of squares were used throughout. Data are summarized in Table 2 and Figure 3
Source
MS,,,,,,
Sepal length
df
F
1 ) hubspecles
(2) Population [subsp 1
(3) Flower sex
( 4 ) Subsp x Fl sex
(5) Pop x F1 sex [subsp.]
(6) Error
(
(2)
(6)
(5)
(5)
(6)
1
18
1
1
16
19.64
31.49
32.48
0.26
5.79
Petal length
F
(1) Subspecies
(2) Population [subsp.]
( 3 ) Flower sex
(4) Subsp. x Fl sex
( 5 ) Pop > Fl sex [subsp.]
(6) Error
P
2.24
0.1516
24.70
<0.0001
57.63
<0.0001
0.19
0.6621
3.22
<0.0001
df,,,,,, = 370
P
8.98
16.11
6.71
0.01
3.64
0.0077
<0.0001
0.0197
0.9253
<0.0001
df,,,,,= 370
PC seems to be based on overall flower size and reflects
the overall positive correlations among floral characters (Table 7). The first PC also partially separates
monoecious plants from dioecious plants as well as
male flowers from female flowers. The second PC appears largely based on pedicel diameter, partially distinguishing male flowers from female flowers in both
breeding systems (Fig. 4).
GENETIC POTENTIAL FOR RESPONSE TO SELECTION
0ffspring:parent regressions showed significant heritability (ma1e:male and fema1e:femaleregressions) for
sepal length, petal length, and reproductive dry weight
(Table 8). There was also some indication of significant
genetic correlation between the sexes (ma1e:female and
fema1e:male regressions) for sepal length and petal
length, though not for reproductive dry weight. The
ma1e:female regressions allow comparison of genetic
correlation between male and female flowers in the
two subspecies. Overall, the magnitudes of the male:
female regressions were higher in the monoecious subspecies, suggesting that genetic correlations are
stronger in this group. Estimation of genetic correlations is inherently less precise than estimation
of heritability; thus, the presence of relatively few
statistically significant regression estimates is likely
to reflect a stronger underlying correlation structure.
Therefore, we conclude that there is sufficient genetic
variation for floral traits to enable evolutionary divergence, but subject to constraints imposed by genetic
correlations.
5.05
11.79
80.83
0 68
2.39
~
Petal width
F
F
0.0003
<0.0001
<0.0001
0.6152
<0.0001
df,,,, = 37 1
~
Source
Sepal width
P
~~
P
0.0374
<0.0001
<0.0001
0.4321
0.0021
df,,, = 37 1
~~~~~~~~
Pedicel width
Gyno/androeciuin
F
P
37.63
<0.0001
9.17
<0.0001
0.21
0.6526
1.14
0.3022
10.84
<0.0001
df,,,,,, = 37 1
F
P
7.9i
0.0113
8.85
<0.0001
206.57
<0.0001
1.23
0.2832
3.14
<0.0001
dfvrFc= 349
DISCUSS ION
The specific questions set out above can now be readily
addressed. Flowers on plants of the dioecious subspecies of Ecballium are larger overall and allocate
more to reproduction than flowers on plants of the
monoecious subspecies. In both subspecies, male
flowers tend to be larger than female flowers. Sexual
dimorphism is no more pronounced in the dioecious
subspecies than in the monoecious subspecies, though
the variance in flower size among populations is greater
in the former. Finally, there is ample heritability in
floral variation to provide for ongoing response t o
selection, with genetic correlation constraints on independent response of female and male flowers that
may be stronger in the monoecious subspecies. In
the following sections, we review these results in the
context of the three conceptual models for sex-specific
patterns of selection outlined above (Table 1).
REPRODUCTIVE COMPENSATION
Ultimately, the reproductive capacity of a plant is
limited or governed by the acquisition and deployment
of resources. Resource constraints can lead to various
trade-offs in relation to gender expression and underlie
reproductive compensation, in which there is a tradeoff in investment in female versus male functionality.
Our results are consistent with reproductive compensation in that male and female plants of the dioecious subspecies, which were more specialized with
FLORAL EVOLUTION IN ECBALLIUM
2
4
6
8 10 12 14 16
0.5
Male sepal length
c
5
2
7
c,
d
a
+
d
1.5
2.0
2.5
3.0
Male sepal width
26
24
22
20
18
16
14
12
10
8
1.0
481
12
11
10
9
8
7
6
5
4
10 12 14 16 18 20 22 24 26
3
Male petal length
u
4 5 6 7 8 9101112
Male petal width
3.5
2
-"
%
3.0
2.5
2.0
a
2 1.5
8
&
0
1
2
3
4
5
Male androecium weight
1.0
0.5
1.0 1.5 2.0 2.5 3.0 3.5
Male pedicel width
Figure 3. Summary of floral measurement data for natural populations of Ecballium elaterium subsp. elaterium
(monoecious: 0 )and Ecballium elaterium subsp. dioicum (dioecious: 0).
Population means and standard deviations
are indicated; overall sample sizes are reflected in Table 1. Diagonal lines indicate values for which male flower
measurements would equal female flower measurements. (A), sepal length; (B), sepal width; (C), petal length; (D),
petal width; (E), androecium/gynoecium dry weight; (F), pedicel width.
Table 4. Correlations between petal length and corolla
diameter in the common-garden population of Ecballium
elateriurn. All correlations are significant at P<O.OOOl
Flower type
N
R
Monoecious females
Monoecious males
Dioecious females
77
71
86
73
0.75
0.74
0.87
0.90
Dioecious males
respect to gender t h a n plants of the monoecious subspecies, produced larger flowers.
Another manifestation of reproductive compensation
in the evolution of dioecious taxa is to increase the
number of flowers produced. In previous work by Costich (1995) it was observed t h a t males of the dioecious
subspecies do produce more flowers on average than
plants of the monoecious subspecies. Moreover, females
of t h e dioecious subspecies ultimately produced more
fruits and more seeds than monoecious plants. Enhanced gender-specific fitness h a s also been invoked
to account for maintenance of breeding system
482
D. F:. COSTICH and T. R. MEAGHER
Table 5. Levene‘s test on variance in floral characters. In most cases, the pattern of significant effects called for the
use of an error MS other than the Emor; in those cases, the appropriate error MS that was used is as indicated in the
second column. Type-I11sums of squares were used throughout. Data are suminarized in Table 2 and Figure 3; analysis
of variance on data is presented in Table 3
Sourcc
MS,,,.,.,,, df
Sepal length
F
__~_____
(11 Subspecies
(2) Population [subsp.]
(2)
(6)
( 3 ) Flower sex
(4)Subsp. x Fl sex
(5) Pol) 14 s ~ [subsp.]
x
( 6 )Error
(6)
(-5)
(6)
Petal length
sourcr
F
1) Subspecies
( 2 )Population [subsp.]
( 3 ) Flower sex
(4)Suhsp. K Fl sex
(5) f’op * F1 StfX Isubsp.1
(6)ICrror
28.01
1.92
0.40
0.09
0.66
1
18
1
1
16
P
F
8.23
1.44
0.27
0.21
1.36
0.0277
0.1115
0.6011
0.6494
0.1578
df,,,,,, = 370
0.0002
0.0490
0.0640
0.8257
0.2577
df,,,, = 370
Table 6. Factor pattern from a principal components
analysis of Ecbrcllium elrcteriurn flower measurements
~
(‘h ariict c r
~
Factor pattern
2nd PC
1st PC
Sep.ii length
S e p d width
P e t J length
Pet,il width
Androrc iurn/gynoecium drj weight
l’cdicel *idth
Proportion of variance explained
0.89250
0 82477
0 77727
0 80721
0 68789
0 00940
0 53
<0.0001
0.0137
0.4468
0.7213
0.8344
df,,,,, = 37 1
Petal width
P
21.45
1.64
3.45
0.05
1.21
-0 08135
-0 27426
- 0 13960
0 10480
0 45636
0 93509
0 20
polymorphism in other species (e.g. Fleming et al.,
1994; Elle & Meagher, 2000; Vassiliadis et al., 2000).
Another manifestation of resource compensation is
increased gender specialization under stress. Costich
(1995) found this to be the case in Ecballium: monoccious plants showed a tendency towards specialization
as either males or females in dryer sites. This phenomenon has also been invoked to account for gender
shifts under stress in Mercurialis (Pannell, 1997),suggesting that when plants are subjected to resource
stress there is a gendey trade-off. Stress has also been
claimed t o account for an increased incidence of dioecy
in cwtain txtreme habitats (e.g. Arroyo & Squeo, 1990;
Rarrett. 1992).
Sepal width
P
F
8.16
3.47
0.01
0.93
1.65
Gyno/androecium
F
P
29.02
<0.0001
<0.0001
3.71
0.0157
7.30
0.0350
5.31
2.08
0.0086
df,.,,,, = 37 1
P
0.0105
<o,0001
0.9141
0.3497
0.0547
df,,,,,, = 371
Pedicel width
F
P
0.57
2.87
24.66
4.22
0.96
0.4614
<0.0001
<0.0001
0.0418
0.492
BATEMAN’S PRINCIPLE
The key issue relating to Bateman’s Principle is that
floral selection favouring male reproductive success is
likely to be different from floral selection favouring
female reproductive success. The general expectation
is that male success is more limited by pollinator
visitation than female success, so that ’male’selection
should be geared towards increased floral display. On
the other hand, female success is expected to be more
limited than male success by the resource cost of fruit
and seed production. Although we have not measured
selection per se, our findings can be interpreted as a
consequence of past selection on floral characters in
Ecballium.
Perhaps the most global prediction of Bateman’s
Principle is that male display for pollinator attraction
should be greater than female display, leading t o larger
male flower size (Bell, 1985). Indeed, the common
pattern of sexual dimorphism in flower size is that of
males larger than females (Bell, 1985; Ilelph, 1996;
Delph, Galloway & Stanton, 1996). Our findings in
Ecballium are consistent with this trend. A s noted
above, male plants and cosexual plants produce an
excess of male flowers, which is also consistent with
response to past selection for enhanced floral display.
CONSPECIFIC FLORAL MIMICRY
Differential selection pressures on male and female
reproductive success lead to secondary differentiation
FLORAL EVOLUTION IN ECBALLIUM
483
Table 7. Correlations among floral characters for the two subspecies of Ecballium elaterium (sample sizes in
parentheses). Male flower correlations are in the upper diagonal and female flower correlations are in the lower
diagonal. Levels of statistical significance are indicated by superscripts (0 - not significant; 1 - B 0 . 0 5 ; 2 - RO.01;
3 - P<0.005;4 - P<O.OOl; 5 - P<0.0005; 6 - P<O.OOOl)
Character
Sepal length
Sepal width
Ecballium elaterium subsp. elaterium (monoecious)
+0.36'j (294)
Sepal length
Sepal width
+0.3@ (326)
Petal length
+0.676 (324) +0.22'j (324)
Petal width
+0.44'j (324) +0.24'j (324)
Gynoeciudandroecium
+0.34'j (320) +0.16* (320)
Pedicel width
+0.33'j (315) +0.163 (315)
Ecballium elaterium subsp. dioicum (dioecious)
Sepal length
+0.52fi(197)
Sepal width
+0.51' (174)
Petal length
+0.79' (174) +0.45fi(174)
Petal width
+0.45'j (174) +0.54'j (174)
Gynoeciudandroecium
+0.286 (173) +0.326 (173)
Pedicel width
+0.36'j (171) +0.264 (171)
2
1
hl
0
a
0
-1
-2
-4
-2
0
2
4
PC 1
Figure 4. PCA graph based on the factor pattern shown
in Table 5 [circles= females; squares =males; filled symbols =Ecballium elaterium subsp. elaterium (monoecious),
open symbols= Ecballium elaterium subsp. dioicum (dioecious)].
of male and female flowers. Yet, at the same time,
male and female flowers are limited in the extent of
differentiation because they need to be similar enough
in appearance t o attract the same pollinators. Charlesworth (1993) has pointed out that this could be achieved
most easily with generalist pollinators that might not
be as discriminating as specialist pollinators. A wellknown correlate of unisexuality is relatively small,
Petal length
Petal width
Gynd
androecium
Pedicel width
+0.66'j (294)
+ 0.25' (294)
+0.23'j (294)
+0.26'j (294)
+O.2ls (294)
+0.316 (292)
+0.25fi (292)
+0.1g4 (292)
+0.25'j (292)
+0.233 (190)
+0.07' (190)
+0.32' (190)
+0.02" (190)
+0.09" (188)
+0.536 (325)
+0.3g6 (319)
+0.306 (314)
10.41' (196)
+0.3g6 (196)
+0.48'j (174)
+0.32'j (173)
+0.5Z6 (171)
+0.376 (319)
+0.1g4 (314)
+0.406 (196)
+0.416 (196)
+0.316 (196)
+0.45j6 (173)
+0.3g6 (171)
+ 0.24'j (314)
+0.44'j (197)
+ 0.54'j (197)
+ 0.346 (196)
+0.223 (196)
+0.90° (143)
+0.222 (143)
+0.15' (143)
+0.16" (142)
+0.15" (143)
+0.253 (172)
unspecialized flowers and pollination by small bees
and bee-flies, 'opportunists' (Bawa, 1980a).
It has been well established that there are potentially
differential costs of reproduction through male and
female function in plants, and this is particularly
pronounced in monoecious and dioecious species where
sex-specificreproductive effort is isolated into different
flowers or different individual plants. Overall investment towards male and female reproductive effort
is expected to evolve towards equality, either through
a shift in allocation or a shift in sex ratio (Fisher,
1930; Charnov, 1982). Thus, costs of fruit and seed set
incurred by female flowers will preempt additional
resource investments that may be possible for males,
such as nectar production.
In a number of monoecious and dioecious species, it
has been determined that male flowers provide rewards
for pollinators and female flowers do not. Thus, female
flowers attract pollinators by mimicking male flowers,
leading to what has been termed 'deceit pollination'
(Baker, 1976; Bawa, 1977, 1980b; Ferdy et al., 1998;
Willson & Agren, 1989; Schemske et al., 1996). Under
this scenario, additional male investment in pollinator
reward plays a n important role in pollinator attraction
for both sexes, counterbalancing additional female investment in fruit and seed set.
In a series of pollination studies of four monoecious
species in the genus Begonia that vary in degree of
sexual dimorphism in flower size and shape, it was
shown that the pollinators could discriminate between
the larger, rewarding male flow:rs and the smaller,
non-rewarding female flowers (Agren & Schemske,
1991; Schemske et al., 1996; Le Corff, Agren &
Ecballium eluteriurn subsp. dioicum (dioecious)
Sepal length
Sepal width
Petal length
Petal width
Gynoecium/androecium dry weight
Pedicel width
Ecbullium eluteriurn subsp. eluteriurn (monoecious)
Sepal length
Sepal width
Petal length
Petal width
Gynoecium/androecium dry weight
Pedicel width
Character
P
ma1e:male ( N = 32)
0.332 k 0.079 0.0002
0.266k0.209 0.2127
0.459k0.126 0.0010
0.011 k0.220 0.9620
-0.455 k0.354 0.2091
0.392 k0.130 0.0540
PkSD
Table 8. 0ffspring:parent regression analysis of floral characters
P
fema1e:female ( N = 23)
0.652 k0.292 0.0367
-0.057k0.281 0.8404
0.657 k0.248 0.0151
0.278 k0.291 6.3491
0.771 k0.252 0.0006
0.082 k0.196 0.6791
fema1e:female ( N = 35)
0.431 k0.088 0.0001
0.117k0.199 0.5610
0.151 i0.103 0.1511
-0.316k0.179 0.0872
0.624k0.263 0.0238
0.037 k0.130 0.7778
PkSD
P
fema1e:male ( N = 31)
0.271 k0.079
0.0018
0.330k0.176
0.0708
0.208k0.108
0.0633
0.241 k0.278
0.3936
0.180k0.427
0.6762
0.327 k0.181
0.0829
PkSD
P
ma1e:female ( N = 28)
0.382k0.221 0.0948
-0.357 k0.257 0.1765
0.261 k0.262 0.3285
-0.067k0.231 0.7753
0.028+0.158 0.8608
0.034 i0.089 0.7063
ma1e:female ( N = 36)
0.449k0.087 0.0001
0.685 k0.244 0.7300
0.382+0.122 0.0037
-0.092k0.133 0.4938
-0.453k0.269 0.1014
0.118k0.094 0.2166
PkSD
FLORAL EVOLUTION IN ECBALLIUM
Schemske, 1998). Seed set was pollen limited in three
of the four species; the exception was the species with
the most similar male and female flowers, where the
visitation rate to female flowers was higher than in
the other species (Le Corff et al., 1998).
Ecballium is a potential example of deceit pollination, in that male flowers reward flower visitors
with nectar and pollen, while female flowers do not. In
a detailed study of pollinator behaviour on monoecious
Ecballium, Dukas (1987) found evidence that the three
bee species observed did discriminate between male
and female flowers: the level of visitation to the female
flowers was only about 30% of that to male flowers,
and the duration of visits to female flowers was also
shorter than that to male flowers, especially for honeybees. More recently, Fahn & Shimony (2001) observed
an abundance of small hemipteran insects in monoecious Ecballium flowers, finding almost twice as many
on average in male flowers, where they appeared to
be foraging on nectar. Thus, conspecific floral mimicry
may be an important limiting factor in the evolution
of floral sexual dimorphism in Ecballium elaterium.
OTHERHYPOTHESES
In an extensive comparative analysis of sexual dimorphism in flower size in plants with unisexual
flowers, Ecballium fell in with the majority (51 of
74) of the temperate zone animal-pollinated group, in
which the perianths of male flowers are larger than
those of female flowers (Delph et al., 1996).The authors
examined a number of hypotheses regarding patterns
of floral sexual dimorphism, three of which we can
explicitly test in Ecballium. First, the pollinator attraction function of the perianth is obviously important
in Ecballium andis the subject of much of the preceding
discussion. Second, a strong developmental correlation
between stamens and petals could lead to larger petal
size in male flowers. The specific prediction for Ecballium would be a positive correlation between androecium dry weight and petal size in male flowers,
which is what we found. However, correlations between
gynoecium dry weight and petal size (length and width)
are of equal o r higher significance in female flowers
compared to correlations between androecium dry
weight and petal size in male flowers. Thus, as concluded by Delph et al. (1996) for the multi-species data
set, the evidence is not compelling that a developmental constraint imposed by the relative size
of stamens is a major factor in floral evolution in
Ecballium. Third, the perianth is presumed to play a n
important role in enclosing reproductive structures in
the bud stage. Ecballium flowers have a n inferior
ovary; thus the petals of female flowers only enclose
the stigma + style portion of the gynoecium in bud. One
of the striking features of our results is the equality of
485
androecium and gynoecium (excluding the ovary) dry
weights, providing no support for the enclosing function as a n underlying mechanism for sexual dimorphism in petal size.
OUTCROSSING, MONOECY AND DIOECY IN ECBALLIUM
In the early 1980s, a debate was raging over the
relative importance of the genetic consequences of the
evolution of separate sexes, with respect to inbreeding
avoidance, versus the advantage of sexual specialization in certain ecological contexts. Although the
field of mating-system ecology and evolution has expanded and diversified over the intervening decades,
these issues continue to be revisited in reviews and
books (Freeman et al., 1997; Richards, 1997; Geber,
Dawson & Delph, 1999). In our earlier allozyme study
of Ecballium (Costich & Meagher, 1992) we found
different levels and patterns of distribution of genetic
variation in the dioecious and monoecious subspecies:
the higher allelic diversity in the dioecious subspecies
was found within most populations, whereas the reduced genetic variation in the monoecious subspecies
was mostly distributed among the populations. A similar pattern of greater phenotypic variation in floral
traits in the dioecious subspecies was detected in the
present study. In two other studies where genetic
diversity in closely related taxa with different inbreeding levels were compared, again a pattern of
higher among-population variation in the selfing taxon
was revealed (Layton & Ganders, 1984; Liu, Zhang &
Charlesworth, 1998).
CONCLUSION
The reproductive biology of a species involves a complex
interaction among a number of factors. A great deal
of attention has been focused on sexual selection as
manifested through Bateman’s Principle, which has
been widely adopted by plant evolutionary biologists
because it is intuitively satisfying. Although our results
are consistent with expectations under Bateman’s
Principle, they can also be accounted for by other
hypotheses suggested by resource compensation or
floral mimicry. Much of the support in the literature for
Bateman’s Principle is based on floral size dimorphism,
but results that actually evaluate sex-specific patterns
of selection on floral pattern have been equivocal.
Our results show strong support for the interplay
between resource compensation and floral mimicry in
the evolution of sex-specific reproductive patterns in
Ecballium. Indeed, this interplay seems likely to be
widespread and should receive more attention in future
studies of the evolution of sexual dimorphism.
486
D. E. COSTICH and T. R. MEAGHER
ACKNOWLEDGEMENTS
This paper is dedicated t o the m e m o r y of Professor
F e r n a n d o Calan (1908-1999), whose enthusiasm for
Ecballzuin could n o t be deterred, neither b y b o m b s
d r o p p e d near h i s s t u d y sites during the Spanish Civil
War at the very beginning of his career, n o r b y infirmity
in his later y e a r s . Once he s a i d , in complete sincerity,
“Ecballium es una joya.” [“Ecballium i s a jewel.”] We
wholeheartedly agree w i t h him.
N a t i o n a l Geographic Society grants and a FulbrightH a y e s F’redoctoral Fellowship provided funding t o
DEC for the original E c b a l l i u m p o p u l a t i o n s a m p l i n g .
NSF Grant BSR9020155 t o TRM and DEC supported
s u b s e q u e n t work.
L. Dehart d i d the a r t w o r k i n Figure 2 and measured
m a n y of the p r e s e r v e d Ecballium flowers. H. L a u , a
Liberty Science Center Partner in Science in o u r lab
a t Rutgers University during summer 1991, did m o s t
of the floral measurements in the experimental population. 1)r S. C. H. Barrett and Dr C. Vassiliadis read
and r o n i m e n t e d o n an earlier draft of the m a n u s c r i p t .
We thank t h e m for their c o n t r i b u t i o n s t o this study.
REFERENCES
Agren el, Schemske DW. 1991. Pollination by deceit in a
nwtropical monoecious herb, Begonia inwolucrata. Biotmpica 23: 235-241.
Agren J , Schemske DW. 1995. Sex allocation in the monoecious herh Begoizia seniiomta. Evolution 4% 121-130.
Arnold SJ. 1994. Bateman’s Principles and the measurement of sexual selection in plants and animals. The Americnn IVaturalist 144: S126-SI49.
Arroyo M, Squeo F. 1990.Relationship between plant breedtems and pollination. In: Kawano s, ed. Biological
chcs and Evolutionary Trends in Plants. San Diego:
Academic Press, 205-227.
Ashman TL,. 1999. Quantitative genetics of floral traits
in a gynodioecious wild strawberry Fraguria virginiana:
implications for the independent evolution of female and
hermaphrodite floral phenotypes. Heredity 83: 733-741.
Baker HG. 1976. “Mistake” pollination as a reproductive
ten1 with special reference to the Caricaceae. In: Burley
.J. Stylcs HT. eds,. Tropical trees: Variation, Breeding and
Conserration. New York: Academic Press, 161-169.
Barrett SCH. 1992. Gender variation and the evolution of
dioecy in i’Vurmhea dioica (Iiliaceae). Journal of E ~ o l u tionor? Biology 5: 423-444.
Bateman AJ. 1948. Intra-sexual selection in Dmsophila.
Herrdity 2: 349-368.
Bawa KS. 1977. The reproductive biology of Cupania
gurrtrrtzalmsis Radlk. (Sapindaceae). Euolution 31: 52-63.
Bawa KS. 1980a. Evolution of dioecy in flowering plants.
Arinuol Kwieuj of Ecology and Systematics 11: 15-39.
Rawa KS. 1980b. Mimicry of‘ male by female flowers and
intrasexual competition for pollinators in Jacaratiu dolichaula (D. Smith) Woodson (Caricaceae). Evolution 34:
467474.
Bell G. 1985. On the function of flowers. Proceedings o f the
Royal Society London B 224: 223-265.
Brown MB, Forsythe AB. 1974. Robust tests for the equality of variances. Journal of the American Statistical Association 69: 364-367.
Campbell DR. 1989. Inflorescence size: test of the male
function hypothesis. American Journal of Botany 7 6 730738.
Campbell DR. 2000. Experimental tests of sex-allocation
theory in plants. Trends in Ecology and Euolution 15:
227-232 J U N 2000.
Charlesworth D. 1993. Why are unisexual flowers associated with wind pollination and unspecialized pollinators? The American, Naturalist 141: 481-490.
Charnov EL. 1982. The T h e o q ofSex Allocaticin. Princeton,
NJ: Princeton University Press.
Costich DE. 1989. The ecology of breeding system variation
in Ecballiurn eluteriurn. Unpublished D. Phil. Thesis. The
University of Iowa.
Costich DE. 1995. Gender specialization across a clirnatic
gradient: experimental comparison of monoecious and dioecious Ecballium. Ecology 7 6 1036-1050.
Costich DE, Galan F. 1988. The ecology of the monoecious
and dioecious subspecies of Ecballiurn eluteriurn (L.) A.
Rich. (Cucurbitaceae) I. Geographic distribution and its
relationship to climatic conditions in Spain. Laga.scalia
15(Suppl.): 697-710.
Costich DE, Meagher TR. 1992. Genetic variation in Ecballiuin eluteriurn (Cucurbitaceae): breeding system and
geographic distribution. ?Journal of Euolutimary Biology
5: 589-601.
Darwin C. 1877. The different forms of flouers on plants of
the same species. London: John Murray.
Delph LF. 1996. Flower size dimorphism in plants with
unisexual flowers. In: Lloyd D, Barrett SCH, eds. Flora2
biology: studies on floral evolution in aninzal-polliizated
plants. New York: Chapman & Hall, 217-237.
Delph LF, Galloway LF, Stanton ML. 1996. Sexual dimorphism in flower size. The American Naturalist 148:
299-320.
Dukas R. 1987. Foraging behavior of three bee species in a
natural mimicry system: female flowers which mimic male
flowers in Ecballiuin eluteriurn (Cucurbitaceae). Oecologia
7 4 256-263.
Elle E, Meagher TR. 2000. Sex allocation and reproductive
success in the andromonoecious perennial Solanum cawlinense (Solanaceae). 11. Paternity and functional gender.
The American Naturalist 156: 622-636.
Fahn A, Shimony C. 2001. Nectary structure and ultrastructure of unisexual flowers of Ecballiuin eluteriurn (L.)
A. Rich. (Cucurbitaceae) and their presumptive pollinators.
Annals of Botany 87: 27-33.
Falconer DS, Mackay TFC. 1996. IntfYJdUCtlOn to quantitative genetics, 4th ed. London: Longman.
Ferdy J-B, Gouyon P-H, Moret J, Godelle B. 1998.
FLORAL EVOLUTION IN ECBALLIUM
Pollinator behavior and deceptive pollination: learning
process and floral evolution. American Naturalist 1 5 2 696705.
Fisher RA. 1930. The genetical theory of natural selection.
Oxford: Oxford University Press.
Fleming TH, Maurice S, Buchmann SL, 'httle MD. 1994.
Reproductive biology and relative male and female fitness
in a trioecious cactus, Pachycereus pringlei (Cactaceae).
American Journal of Botany 81: 858467.
Freeman DC, Lovett Doust J, El-Keblawy A, Miglia KJ,
McArthur ED. 1997. Sexual specialization and inbreeding
avoidance in the evolution of dioecy. The Botanical Review
63: 65-92.
Galan F. 1946. Sur la genetique de la monoecie e t la dioecie
zygotique chez Ecballium elaterium Rich. Comptes rendus
de l'Aca&mie des Sciences Paris 222: 1130-1131.
Geber MA, Dawson TE, Delph LF. 1999. Gender and
sexual dimorphism in flowering plants. Berlin: SpringerVerlag.
Layton CR, Ganders FR. 1984. The genetic consequences
of contrasting breeding systems in Plectritis. Evolution 38:
1308-1 325.
Le Corff J, Agren J, Schemske DW. 1998. Floral display,
pollinator discrimination, and female reproductive success
in two monoecious Begonia species. Ecology 7 9 1610-1619.
Liu F, Zhang L, Charlesworth D. 1998. Genetic diversity
in Leavenworthia populations with different inbreeding
levels. Proceedings of the Royal Society of London B 265:
293-301.
Meagher TR. 1994. The quantitative genetics of sexual
dimorphism in Silene latifolia. 11. Responses to sex-specific
selection. Evolution 48: 939-951.
Meagher TR. 1999. The quantitative genetics of sexual
487
dimorphism. In: Geber M, Dawson T, Delph L, eds. Sexual
dimorphism in plants. New York: Springer-Verlag, 275294.
Pannell J. 1997. Variation in sex ratios and sex allocation
in androdioecious Mercurialis annua. Journal of Ecology
8 5 57-69.
Richards AJ. 1997. Plant breeding systems, 2nd edition.
London: Chapman & Hall.
Saavedra MM. 2000. Caracteristicas y control del pepinillo
del diablo Ecballium elaterium (L.) Richard. [Comunicaciones I + D 28/00]. J u n t a de Andalusia. Consejeria
de Agricultura y Pesca.
Schemske DW, Agren J, Le Corff J. 1996. Deceit pollination i n the monoecious, neotropical herb Begonia oaracana (Begoniaceae). In: Lloyd D, Barrett SCH, eds. Floral
Biology: Studies on Floral Evolution in Animal-Pollinated
Plants. New York: Chapman & Hall, 292-318.
Sokal RR, Rohlf FJ. 1981. Biometry, 2nd ed. New York:
Freeman.
Trivers RL. 1972. Parental investment and sexual selection.
In: Campbell B, ed. Sexual selection and the descent of
man, 1871-1971. Chicago: Aldine, 136-179.
Vassiliadis C, Lepart J, Saumitou-Laprade P, Vernet P.
2000. Self-incompatibility and male fertilization success in
Phillyrea angustifolia (Oleaceae). Interrxational Journal of
Plant Science 161: 393402.
Westergaard M. 1958. The mechanism of sex determination
in dioecious flowering plants. Advances in Genetics 9 217281.
Willson MF, Agren J. 1989. Differential floral rewards and
pollination by deceit in unisexual flowers. Oikos 5 5 23-29.
Wilson P, Thomson JD, Stanton ML, Rigney LP. 1994.
Beyond floral Batemania: gender biases in selection for
pollination success. The American Naturalist 143: 283-296.