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BiologicalJoumal ofthe hnnean Sociep (1998), 63: 221 -232.

Multiple paternity in the c o m m o n fkog

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(Rana temporaria): genetic evidence &om
tadpole kin groups
ANSSI LAURILA' AND PERTTU SEPPA'"
'Division o f Population Biology, Department o f Ecology and $stematics, and 'Division o f
Genetics, Department o f Biosciences, EO. Box 17, FW-00014 Unioersip o f Helsinki, Finland

Received I1 April 1997; accepkdforpublicatiun 28 August 1997

Very few studies have investigated the occurrence of multiple paternity and sperm competition
in amphibians. We studied genetic relatedness within kin groups of tadpoles of an aquatically
breeding anuran Rana lempuraria using allozymes. We collected samples from 52 naturally
fertilized spawn clumps produced by single females at three breeding sites in two populations.
We estimated relatedness (r) within kin groups, and compared the observed genotype
distributions of the tadpoles (on average 23 individuals in each group) with the expected
distributions based on single mating. Average relatedness over five polymorphic loci was
0.44 and 0.43 in the two populations, the latter being significantly smaller than that expected
by single mating (0.5). The number of patrilines, calculated from relatedness estimates, was 1.3
in one population and 1.4 in the other. Genotype distributions deviated significantly from the
expected in half of the kin groups and at all breeding sites. The results show that egg clutches
of R. tnnpwm;l commonly contain multiply sired offspring. We suggest that communal breeding
may afiect paternity patterns in R. fanpa& as well as in anurans in general.
0 19911 The Iinnran Soriefy of London

ADDITIONAL KEY WORDS:-Amphibia - Anura - allozymes - genetic relatedness -


multiple paternity - sperm competition.
CONTENTS

Introduction . . . . . . . . . . . . . . . . . . . . . . . 222
Material and methods . . . . . . . . . . . . . . . . . . . 223
Study object and sample collection . . . . . . . . . . . . . . 223
Electrophoresis . . . . . . . . . . . . . . . . . . . . 223
Estimation of paternity patterns from the tadpole kin groups . . . . . 223
Results . . . . . . . . . . . . . . . . . . . . . . . . 225
Genetic composition of the populations . . . . . . . . . . . . 225
Relatedness in the tadpole kin groups . . . . . . . . . . . . . 226
Genotypic composition of tadpole kin groups . . . . . . . . . . 226
Discussion . . . . . . . . . . . . . . . . . . . . . . . 227
Multiple paternity in the common frog . . . . . . . . . . . . 227
Behavioural mechanisms and multiple paternity in anurans . . . . . . 228
Acknowledgements . . . . . . . . . . . . . . . . . . . . 230
Refcrences . . . . . . . . . . . . . . . . . . . . . . . 230
Appendix . . . . . . . . . . . . . . . . . . . . . . . . 232
Correspondence to A. Laurila. E-mail: anssi.laurila@helsi.fi
Present address P. Seppa: Department of Genetics, Uppsala University, Box 7003, S-75007 Uppsala, Sweden.
22 1
0024-4066/98/020221 + 12 $25.00/0/bj970180 0 1998 The Linnean Society of London
222 A. LAURIL4 AND P. SEPPA

INTRODUCTION

Mating with multiple partners should generally be advantageous for males due
to their ability to produce gametes at a higher rate (Trivers, 1972). In females, single
mating is usually enough to fertilize all the offspring in a brood (but see e.g.
Robertson, 1990; Elmberg, 1991; Bourne, 1993 for exceptions), and, consequently,

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the benefits of multiple mating are not obvious (e.g. Hunter el al., 1993).In addition
to fertility assurance, direct benefits to females include acquisition of nutrients and
paternal care as well as avoidance of male harassment (see Birkhead & Parker, 1997
for a recent review). Females may also gain indirect benefits like improved genetic
quality of offspring by mating with multiple partners (Birkhead & Parker, 1997).
Consequently, mating with multiple partners is a potential factor driving sexual
selection in both sexes (Meller, 1992; M ~ l l e r& Birkhead, 1994), and may S e c t
patterns of parental care (Trivers, 1972; Westneat & Sherman, 1993).
Among vertebrates, multiple paternity within a brood has been found in several
bird species (reviewed by Birkhead & Meller, 1992),mammals (e.g. Birdsall & Nash,
1973; Hanken & Sherman, 1981; Keane et al., 1994), reptiles (Stille et al., 1986,
Schwartz et al., 1989; Olsson et al., 1994) and fish (Hutchings & Myers, 1988; Philipp
& Gross, 1994). Among amphibians, females produce multiply sired clutches in
terrestrially breeding salamanders with internal fertilization and prolonged sperm
storage (Tilley & Hausman, 1976). Studies on multiple paternity have mostly
concentrated on species with internal fertilization, and there has been less interest
in species with external fertilization (but see e.g. Hutchings & Myers, 1988; Philipp
& Gross, 1994; Shapiro et al., 1994; D’Orgeix & Turner, 1995; Levitan & Petersen,
1995).
In most anurans (frogs and toads) fertilization is external. Male anurans compete
strongly for mating opportunities, and numerous papers have reported mating
advantage of large males (see Anderson, 1994 for review). However, none of those
studies has addressed how this advantage is realized in the number of fertilized
offspring. In many species, males attempt to displace each other from the female’s
back and when males are unable to solve their dispute over a female, several males
may be attached to her at the moment of fertilization. These multi-male breeding
groups are common in some terrestrially breeding frogs (Kusano et al., 1991;Jennions
& Passmore, 1993; D’Orgeix & Turner, 1995), and high fertilization success among
the competing males has been reported in the treefrog Aga&vchnis callidyas (D’Orgeix
& Turner, 1995). In aquatically breeding anurans it is not unusual to see many
males fighting for a female at the spawning site, but, possibly for practical reasons,
there are no observations of simultaneous sperm release by several males. In addition,
it has been suggested that sperm competition may occur in communal breeding
areas where amplectant pairs gather to spawn (Berger & Rybacki, 1992).
T o our knowledge, the study by D’Orgeix & Turner (1995) is the only one giving
genetic evidence for sperm competition and multiple paternity in anurans. Studies
on amphibians breeding in aquatic environment are lacking. In the present paper
we report on paternity patterns in two populations of an aquatically breeding anuran,
the common frog Rana temfiorariu. We analysed the genetic composition of tadpole
groups derived from naturally fertilized egg clumps produced by single females. We
chose to use allozymes as genetic markers, because they provide rapid information
on population-level phenomena and a large number of individuals can be screened
within a reasonable time span. We analysed paternity patterns by estimating
MULTIPLE PATERNITY IN RANA TEh4PORARL4 223

relatedness among kin group members and by deriving effective mate numbers
based on these estimates. We also compared the observed genotype distributions of
the offspring with the expectation based on single mating.

MATERIAL AND METHODS

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Stu@ object and sample collection

Rana temporaria breeds in a wide range of freshwater habitats across Central and
Northern Europe. Our study population breeds in rock-pools on rocky islands along
the Baltic coast in southwestern Finland. The common frog is an ‘explosive’ breeder
(sensu Wells, 1977). In southern Finland breeding occurs in late April and normally
lasts less than 3 weeks (Elmberg, 1990; A. Laurila, pers. observ.). However, within
a single rock-pool breeding occurs during a considerably shorter time, usually less
than a week (A. Laurila, pers. observ.). In our study population of R. taporaria
mating occurs almost exclusively at night, and the behaviour of mating individuals
is susceptible to disturbance. Consequently, we were not able to carry out direct
observations of mating behaviour. Because each female lays a single distinct clump
of eggs per breeding season and fresh clumps can reliably be told apart from the
clumps laid by other females (Savage, 1961), analyses of paternity patterns were
possible.
In late April 1994, we collected freshly laid R. temporaria eggs on two islands near
Tvarminne Zoological Station, Hanko. O n the island of Lgngskar, a total of 121
females bred in 2 1 different rock-pools in 1994. In rock-pool A, we collected samples
from every deposited egg clump (20 breeding females in 1994). In pool B (24
breeding females), samples were collected from nine egg clumps. O n the island of
Porskar (182 breeding females in 23 rock-pools in 1994), samples were collected
from 23 egg clumps in a pool with 47 breeding females in 1994 (pool C in the
Appendix). O n both islands, we collected samples from the rock-pools where the
numbers of breeding R. taporaria females were the highest.
A sample of approximately 50 eggs was taken from each egg batch and brought
to the laboratory. The eggs from each clump were then placed in separate plastic
pans filled with 2 1 of water. The eggs hatched in laboratory, and after reaching
developmental stage 25 (Gosner, 1960), the tadpoles were preserved at -8OoC until
laboratory analysis.

Electrophoresis

O n average 23 (range 20-36) offspring from each clump were analysed using
horizontal starch-gel electrophoresis. Twenty-two enzyme systems were stained using
standard formulations (Harris & Hopkinson, 1976). Five loci were polymorphic in
one or both of the populations to estimate relatedness and the mating system reliably
(see Table 1). The total number of tadpoles analyzed in 52 groups was 1189.

Estimation ofpatmnig patternshm the tadpole kin group^

Paternity patterns and the general mating structure of the populations (panmixis)
were examined separately in the two populations. We studied paternity patterns by
224 A. LAURIL4 AND P. SEPPA

TABLE1. The enzymes and loci studied, Enzyme Commission (E.C.) numbers, buffers used, and the
number of alleles (no,,)in each locus.

Enzyme Locus E.C. Buffer ndl


~~ ~ ~~

Mannose phosphate isomerase Mpi 5.3.1.8 I 2


Phosphogluconate dehydrogenase PEd 1.1.1.44 I1 2
Isocitrate dehydrogenase Idh 1.1.1.42 I1 3

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Aconitase Aco 4.2.1.3 I 2
Phosphoglyceratekinae PQk 2.7.2.3 I1 2

Buffers: (I): Gel: 42.9mmol I-' TRIS, 4.2mmol I-' citrate, pH 8.4; Tray: 135mmol I-' TRIS, 42.9mmol I - '
citrate, pH 7.1 (Varvio-Ah0 & Pamilo 1980).
(11): Gel: 9mmol I-'mM TRIS, 3mmol I-'rnM citrate, 1.2rnmol I-'mM EDTA, pH 7; Tray: 135rnmol I - ' TRIS,
44.5mmol I - ' citrate, 1.2mmol I - ' EDTA, pH 7 (Ayala ef al., 1974).

estimating genetic relatedness within the kin groups from the genotype frequency
data, and examining multilocus genotype patterns in the individual kin groups.
Relatedness (r) was estimated as genotypic correlation among group members
(Pamilo, 1990). This method was originally tailored for describing colony kin
structures in haplo-diploid social insects (see Crozier & Pamilo, 1996), but it is
perfectly applicable for diploid organisms living in social groups as well (McCauley
& O'Donnell, 1984; Avise & Shapiro, 1986; Schwartz et al., 1989; Costa & Ross,
1993).
Panmixis was estimated using the inbreeding coefficient F= (1 -H J H J , where
H, is the observed and He is the Hardy-Weinberg expected heterozygosity (Wright,
1951). Both relatedness and inbreeding estimates were calculated using computer
software provided by P. Pamilo, and jackknifed over groups and loci to obtain
standard errors (Efron & Tibshirani, 1993). These were then used to test the
differences of the estimates from expected values (assuming t-distribution). For
relatedness, we used r= 0.5 as the expected value, which is the expected relatedness
among full siblings. The expected value for the inbreeding coefficient is zero, which
indicates total panmixis (e.g. Nei, 1987).
Relatedness among the members of a group is determined by the number,
relatedness and breeding success of the individuals in the parental generation (Ross,
1993; Queller, 1993). Therefore, relatedness not only describes the kin structure of
the groups, but can also be used to characterize the breeding system of the species
(the number of males and females contributing to the group). When groups of
diploid organisms belong to the same generation, and mating in the population is
random, genetic relatedness within the group (9)can be used to calculate the
number of matrilines (A4) and patrilines (P)that have contributed to this group:

+
r/=0.25/M+ (rM/4)[(M- 1)/M] 0.25/P+ ( ~ p / 4 ) [ ( P1)/pl
- (1)
(Queller, 1993), where rM is relatedness among females and rp relatedness among
males that have contributed to this group. In our case, equation (1) can be
considerably simplified, because the egg clumps of R. temporaria are produced by a
single female (M= 1). If multiple males contributing to the offspring of a single
female are related (rp>O), our r/ values are underestimates. However, our study
populations turned out to be panmictic (see results), and multiple males contributing
to the same group were therefore assumed to be unrelated (rp=O). Thus, the
MULTIPLE PATERNITY IN RANA 72NPORARk4 225

2. Allele frequencies of the Rana temporaria populations (estimated weighing groups equally) of
TABLE
the loci studied.

Mpi I& Aco Pgk


100 F 100 pgd F 100 F s 100 s I00 s
Ungskar 0.548 0.452 0.891 0.109 0.497 0.205 0.299 0.632 0.368 0.936 0.064

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Porskar 0.677 0.323 0.990 0.010 0.597 0.142 0.262 0.730 0.270 0.873 0.127

relatedness among the tadpoles in a kin group depends only on the number of males
fathering the brood. By substituting M= 1, rp= 0, and solving for P gives:

P= 1 /(4?71) (2)
Matrilines and patrilines in this context are genetically effective numbers. Therefore
they refer to the harmonic means across groups, reflecting the genetic effects of
multiple females or males contributing to the groups (Wade, 1985).
Multiple paternity was also analysed directly from the genotype distributions of
each offspring.Deviation from single mating by females was determined by comparing
the observed genotypes in all loci to the ones expected from a single pair using x2
tests (with the Yates correction when appropriate). The parental genotypes were
deduced from the offspring genotypes by always choosing the ones requiring the
smallest number of fathers according to simple Mendelian rules. For example, in
tadpole group C-1 in enzyme Mpi, there were ten offspring of the genotype ZOO/
ZOO, and ten of F/ZOO (Appendix).The deduced parental genotypes were ZOO/ZOO,
and F/ZOO,although the mother being ZOO/ZOO, she could have had two mating
partners, Z OO/ZOO and F/F. Consequently, the estimates of paternity frequency
estimated from genotype distributions were conservative.

RESULTS

Genetic composition of the populations

Genotype frequencies of individual offspring are shown in the Appendix, and


allele frequencies (estimated weighing groups equally) of the two populations in
Table 2. Enzymes Pgd, Aco and Pgk were all bi-allelic systems. The more common
allele was designated ZOO, and the rare allele either Fast (4 or Slow (4 depending
on the relative mobility compared to the common allele. Enzyme Zdh was tri-allelic,
with both a fast and a slow rare allele. In enzyme Mpi, two fast alleles (with relative
mobilities 10.5 and 107) were first separated besides the common allele (ZOO). The
fast alleles were, however, difficult to separate, and because there may have been
ambiguities in scoring them, they were combined for the data analysis (as allele 4.
The inbreeding estimates per locus, and overall loci for the two populations are
presented in Table 3. The single-locus estimates varied slightly around zero, but
none deviated significantly from zero. Consequently, the population estimates were
close to zero (Table 3) indicating random mating within our study populations. The
Lbgskar population was sampled from two distinct rock-pools; thus the zero
inbreeding coefficient suggests that they belong to the same breeding population.
226 A. LAURILA AND P. SEPPA

TABLE
3. Genetic relatedness (r+ SE) and inbreeding (F+ SE), estimated from the tadpole kin groups
of Rana temporaria. tF refms to the deuiation of the inbreeding coeficients fmm <em (one-sample (two-sided) t-tests).
None ofthe tf-values were significant at the level -0.05.
r SE F SE b
Ungskar Mpi 0.389 0.085 -0,058 0.107 0.54

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PSd 0.426 0.064 -0.003 0.112 0.03
Idh 0.467 0.063 -0.043 0.074 0.58
Aco 0.42 I 0.071 0.037 0.075 0.49
P& 0.463 0. I30 0.041 0.06 1 0.67
Total 0.431 0.030 -0.018 0.049 0.37
Porskar Mpi 0.230 0.073 -0.055 0.089 0.62
Pgd 0.450 0.526 -0.010 0.967 0.0 I
I& 0.495 0.059 0.070 0.067 1.04
Aco 0.534 0.094 0.076 0.111 0.68
PSk 0.496 0.129 -0.109 0.055 1.98
Total 0.439 0.052 0.013 0.037 0.35

Relatedness in the tadpole kin groups

Single locus and population estimates of relatedness are shown in Table 3.


Generally, single locus estimates gave similar results about relatedness and although
there was slight variation among loci, the estimates did not significantly differ within
the populations. Relatedness estimates in the two populations were almost identical.
However, the LHngskar estimate was significantly smaller than 0.5, because of the
lower standard errors (one-sample t-tests: LHngshr: t= 2.3, df= 28, P= 0.0146;
Porskar: t = 1.17, df=22, P=O.1273).
Similar relatedness values also gave similar numbers of patrilines contributing to
the offspring. From equation 3, the number of patrilines was calculated to be 1.4
in LBngskar, and 1.3 in Porskar. Because these values refer to the harmonic means
of the mate number (Wade 1985), such figures would result, for instance, if about
55% of the females produce offspring sired by two males in LHngskar, and about
50% in Porskar.

Genotypic composition of tadpole kin groups

In two kin groups from LHngskr (A-54, B-23), and in one group from Porskar
(C-ll), the observed genotypes could not have been produced by a single pair
(Appendix). In addition, on LHngskr the observed genotype frequencies were
significantly different from the expected based on single mating in one or more loci
for 14 kin groups, and on Porskar for nine groups (Table 4). These figures add up
to 16 kin groups in LBngskar (55%), and 10 groups in Porskar (43%), as the
proportion of females producing kin groups fathered by more than one male. In
ten cases, the deviation from single mating in a single kin group was corroborated
by a significant value at more than one locus.
MULTIPLE PATERNITY IN RANA 7EMPORARL.4 227

TABLE 4. Deviations of genotype distributions from the expected


based on single mating in individual tadpole groups produced by x2
tests. Statistically significant deviations are indicated with asterisks:
*0.05<P<0.01; **0.01<P<0.001;***RO.OOl. NC =genotypes of
the brood are not compatible with a single reproducing pair. Blank
cell = ns

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Group Mpi Pgd Idh Aco Pgk
A- 1 **
A-53 * *
A-54 NC
A-55 * ***
A-57 *** *** *
A-6 I ***
A-73 *
A-79 ***
A-83 * *
A-88 **
B-2 1 ** ***
8-23 * NC
B-77 * *
B-83 * *
B-84 ***
B-86 **
c-4 **
c-5 **
C-6 ***
c-7 *** ***
c-lI NC
C-13 *
C:-14 **
Cl-18 *
C-23 * *
C-24 **

DISCUSSION

Multiple p a h i p in the common f i g

We have demonstrated that multiple paternity among the offspring of single


females is a common phenomenon in R. temporaria. Using two independent methods,
we estimated that roughly half the egg batches had multiply sired offspring.
Furthermore, our result was qualitatively confirmed by the tadpole kin groups,
which could not have been fathered by a single male. Multiple paternity was detected
in both populations and in all three study pools. Because R. temporaria females release
their eggs very rapidly in a single batch (Savage, 1961), the occurrence of multiple
paternity means that spermatozoa from several males are competing. Earlier, Berger
& Rybacki (1992) showed that sperm competition occurs between two species of
aquatically breeding water frogs (R.esculata-complex) under laboratory conditions,
but evidence from nature has been lacking.
The exact numbers of males fathering the offspring of single females should,
however, be interpreted with caution. First, when relatedness is estimated from
genetic data, different kinds of sampling error may occur. Even small changes in
the point estimates of relatedness would translate to fairly large changes in the
228 A. LAURILA AND P.SEPPA

number of patrilines calculated from these estimates. Second, the estimate in Porskar
was not significantly lower than the expected value for full sibs. Finally, the number
of patrilines estimated from the genotype frequencies of the offspring may have
been exaggerated, because of the large number of x2 tests. If a standard Bonferroni
correction is made for the significance levels of the tests (separately for each locus),
the values showing P<O.OOl in Table 4 remain significant. This would leave

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significant evidence for eight kin groups (28%) in LAngskar and three (13%) in
Porskar having been fathered by more than one male.
On the other hand, our methods also tend to underestimate the number of
patrilines. First, when analysing multiple paternity directly from genotype dis-
tributions, the parental genotypes were chosen so as to minimize the mating
frequency. Second, multiple matings may go undetected if males have an identical
multilocus genotype. The probability of this depends on the allele frequencies in
the population, but it is fairly low when several loci are studied.
Both the number of mates calculated from relatedness estimates and the evaluation
of the genotype distributions within the offspring gave remarkably similar results.
Depending on the method and population, between 48 and 55% of the tadpole kin
groups were fathered by at least two males. The Bonferroni correction results in
lower estimates. It is also worth remembering that the number of patrilines calculated
from relatedness estimates is a genetically effective number. If different males father
the offspring unequally, the true number of patrilines is much larger. Therefore we
believe that although we were not able to reliably estimate exact paternity frequencies,
our results emphasize the underlining biological phenomenon of multiple paternity
in R. temporaria.

Behavioural mechanims and multiple paternip in anurans

Our results suggest that in explosively breeding anurans, primary (amplectant)


males are not as successful as has previously been presented. However, because we
were not able to carry out behavioural observations,we can currently only hypothesize
about the behavioural strategies leading to multiple paternity in R. temporaria. In
anuran breeding sites only a small fraction of the males are paired at any given
time, and there is commonly an excess of males (Wells, 1977). Also in R. temporaria
the operational sex ratio in a breeding pond is usually male-biased (Elmberg, 1990;
1991). Theoretically, surplus males might simply gather near an amplectant pair
about to spawn, and release sperm as the female starts to lay eggs. Several males
may also grasp a female at the moment of fertilization. In R. temporaria, several males
sometimes grasp a female shortly before spawning (J. Elmberg, pers. comm.), but
observations of male behaviour at the moment of fertilization are lacking. Among
some terrestrially breeding anurans (e.g. family Rhacoporidae) multi-male breeding
groups are common (Kusano et al., 1991;Jennions & Passmore, 1993), and sperm
competition occurs (Jennions & Passmore, 1993; D’Orgeix & Turner, 1995). Since
proximity to the female has been assumed to affect fertilization success (Gross, 1985),
the lack of observations of multi-male breeding groups in aquatic breeders has been
explained by the priority of primary male’s sperm (Jennions & Passmore, 1993).
However, in the treefrog A. callidy.s, D’Orgeix & Turner (1995) reported high
fertilization success by both competing males in the two documented cases. Un-
fortunately, we do not have data on parental genotypes to perform a similar analysis.
MULTIPLE PATERNITY IN M A TEMPORARIA 229

Berger & Rybacki (1 992) suggested that, in nature, sperm competition occurs at
communal oviposition sites where amplectant pairs gather to release the gametes.
In R. temporaria, as in many other explosively breeding anurans, oviposition occurs
within a very restricted area in the breeding pond (Wells, 1977; Seale, 1982;
Waldman, 1982; A. Laurila, pers. observ.).For example, in our study pools oviposition
took place in an area of approximately 1 m2 in pools A and B and 1 .5m2 in pool

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C. Furthermore, the egg clumps laid during one night are usually deposited very
close to each other (cf. Waldman, 1982; A. Laurila, pers. observ.). Consequently,
during the peak spawning time there may be plenty of free-swimming spermatozoa
in the water. The spermatozoa may be able to reach the egg clumps laid by other
pairs and fertilize a proportion of the eggs, either by active swimming or by being
passively transported by the water movements. Hence, when several females spawn
in a small area during a limited time, multiple paternity may simply be a consequence
of high spermatozoa concentration in the water. In R. temporaria this possibility is
further increased, because several females spawn within a very short time (during
the few dark hours of the night), and there are usually few active reproductive nights
in each pond (Elmberg, 1990).
When males differ in fertilization ability, females may mate with multiple partners
to maximize the number of fertilized ova (Drummond, 1984). In R. taporariu, there
is considerable variation in fertilization rate among egg clumps (5-1 00%; Gibbons
& McCarthy, 1986; Elmberg, 1991). Elmberg (199 1) showed that fertilization success
of male R. temporaria was positively correlated with male fat reserves, implying that
male nutritional status affects sperm production. Thus, the fertilization capacity of
a male may vary accordingly. In anurans, the choice of the oviposition site is
generally under female control. For example, both in R. taporaria (J.Elmberg, pers.
comm.) and in the wood frog R. sylvutica (Berven, 1981) females about to deposit
eggs have been found to move towards the center of the spawn clump mass. If free-
swimming spermatozoa released by other males can fertilize a proportion of the
eggs, female frogs may try to maximize the number of fertilized eggs by breeding
synchronously in a limited area. Previously, synchronous breeding has been shown
to accelerate development and increase hatching success of anuran eggs (Seale,
1982; Waldman, 1982).
Owing to their better fighting ability, large males have a mating advantage in
some anurans (e.g. Davies & Halliday, 1979; Howard & Kluge, 1985). Furthermore,
in some species females actively choose large mates (reviewed by Andersson, 1994).
Mating with a large male may be directly advantageous, because offspring sired by
large males may grow faster (reviewed by Andersson, 1994). Interestingly, female
mate choice has also been found to increase fertilization success in some anurans
(Robertson, 1990; Bourne, 1993). In these species fertilization success is dependent
on the relative body sizes of the breeding pair. Females select mates on the basis of
their vocalization, and preferred males have a higher fertilization rate than their rivals
(Robertson, 1990; Bourne, 1993). However, in R. temporaria opportunities for female
mate choice are usually low and random mating appears to be the rule (Elmberg,
1987, 1991). In terms of mate choice, a R. temporaria female has probably very little to
lose by allowing her offspring to be sired multiply.
Nevertheless, sophisticated mate choice has been reported in many anurans,
especially in prolonged breeders (Andersson, 1994). Therefore, it is interesting to
note that in prolonged breeders communal oviposition is rare (Wells, 1977). If
multiple paternity prevents females from controlling the identity of the fathers, a
230 A. LAUFULA AND P. SEPPA

female that has chosen the best available mating partner may avoid situations
leading to multiply sired offspring. On the other hand, if multiple paternity results
from a female’s assurance strategy against reduced fertilization success and limited
mate choice opportunities, it may be more widespread among species with explosive
synchronous breeding systems. Further studies are needed to study whether paternity
patterns covary with breeding systems in anurans.

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ACKNOWLEDGEMENTS

Tvarminne zoological station provided the laboratory facilities for rearing the
tadpoles. Comments by J. Elmberg, I. K. Hanski, K. Lindstrom, N. Peuhkuri, E.
Ranta and L. Sundstrom substantially improved the manuscript. This work has
been financially supported by University of Helsinki (to AL, PS), the Academy of
Finland (AL, PS), and the Emil Aaltonen Foundation (PS).

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232 A. LAURILA AND P. SEPPA

APPENDIX

Absolute geno&pej%quencies oftadpole kin p u p s in Rana temporaria

100 F
Mpi
F 100 f' F 100 F 100
Idh
F F S
Aco
100 100 S 100
Pgk
100 S

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100 100 F 100 100 F 100 100 S F S S 100 S S 100 S S

UngskPr:
A-1 9 12 21 13 7 2 7 1 1 7 2
A-2 9 11 20 4 7 3 8 12 10
A-3 4 8 8 21 3 3 1 1 4 7 3 1 0
A-4 9 9 3 21 1 4 5 6 4 13 4 21
A-52 8 14 3 24 12 9 17 8 12 I 1
A-53 10 10 11 9 8 1 1 5 5 7 3
A-54 20 20 4 6 4 1 3 5 10 5 20
A-55 5 15 20 1 8 I1 10 10 10 9
A-56 13 8 7 14 7 11 3 13 5 21
A-57 13 20 36 30 1 1 17 18 13 19 2
A-58 11 9 7 14 4 7 6 8 6 2 1
A-59 15 9 6 10 8 24 5 15 4 18 4
A-60 11 11 22 13 9 7 15 21
A-61 1 21 24 7 8 3 3 8 9 23
A-73 11 7 4 3 3 5 1 34 33
A-74 7 11 6 26 10 12 25 26
A-79 22 13 10 12 11 7 16 23 1
A-82 22 22 8 10 1 15 7 22
A-88 4 12 7 20 4 12 9 6 10 8 11
A-89 11 15 10 14 6 5 10 5 13 12 26
B-19 6 13 4 26 5 8 9 4 13 9 26
B-2 1 I1 1 25 19 2 9 3 17 3
B-23 6 16 23 2 2 10 8 9 8 24
B-77 5 16 11 10 11 8 2 2 11 5 21
B-81 11 12 23 5 4 8 5 22 22
B-83 2 10 11 14 10 9 2 8 1 4 11 6 24
B-84 17 1 9 10 11 4 13 7 12 7
B-85 11 11 15 8 10 13 12 9 23
B-86 13 I1 13 I1 1 2 9 1 2 23 19
Porskar:
c-1 10 10 20 3 3 4 12 8 6 4
C-2 8 13 1 23 9 14 23 23
c-3 I1 11 23 6 5 7 5 12 10 22
c-4 12 5 7 24 13 9 12 11 20 4
c-5 10 4 8 22 10 I1 8 13 14 6
C-6 18 2 20 14 6 14 20
C-7 14 6 1 21 8 1 6 1 1 1 9
C-8 5 12 7 24 9 4 8 3 12 12 24
c-9 4 15 2 20 9 12 7 13 8 4
c-10 2 13 7 24 4 5 3 3 9 1 3 2 2 3
C-11 8 11 1 21 7 10 1 2 12 9 21
C-12 8 13 22 22 7 15 11
(2-13 20 20 15 5 18 10
C-14 5 15 I 11 10 2 9 2 4 2 12 7 18 3
C-15 5 10 5 20 10 9 20 20
C-16 25 25 3 12 10 12 12 25
C-17 21 12 7 2 22 9
C-18 14 8 22 9 13 12 10 17 5
C-20 8 12 2 22 6 4 4 8 12 8 12 2
c-22 4 9 26 7 9 3 3 3 10 10 11 15
C-23 5 9 4 21 4 15 10 11 8 1
C-24 12 11 2 14 8 22 1 10
(2-25 11 10 21 5 4 21 6 3

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