Download High Levels of Multiple Paternity in Littorina saxatilis: Hedging the

Journal of Heredity 2007:98(7):705–711
doi:10.1093/jhered/esm097
Advance Access publication December 4, 2007
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High Levels of Multiple Paternity in
Littorina saxatilis: Hedging the Bets?
TUULI MÄKINEN, MARINA PANOVA,
AND
CARL ANDRÉ
From the Department of Marine Ecology, Tjärnö Marine Biological Laboratory, Göteborg University, S-452 96 Strömstad,
Sweden (Mäkinen, Panova, and André).
Address correspondence to T. Mäkinen at the address above, or e-mail: [email protected].
Abstract
The mating system of a species can have great effects on its genetic structure and evolution. We studied the extent of
multiple paternity in a gastropod with internal fertilization, the intertidal snail Littorina saxatilis. Paternal genotype
reconstruction based on microsatellite markers was performed on the offspring of wild, naturally fertilized females from 2
populations. The numbers of males contributing to the offspring per female were among the highest detected in
invertebrates so far, with the exception of social insects. No reproductive skew in favor of males that were genetically more
distant from the females was detected, and the pattern of fertilization appeared random. The result fits a hypothesis of
indiscriminate mating, with genetic bet hedging as the most likely explanation. Bet hedging may have evolved as a form of
inbreeding avoidance, if the snails are not able to recognize relatives. However, nutritional benefits from sperm or sexual
conflict with males are additional possibilities that remain to be assessed in this species. Whatever the causes, such high
levels of multiple paternity are remarkable and are likely to have a large impact on population structure and dynamics in
a species in which migration between populations is spurious.
Observations of multiple paternity in single clutches or broods
have been commonplace since Darwin’s time (Birkhead 2000).
However, a classic paradigm in evolutionary biology stated
that females benefit little from mating with more than one
male (Bateman 1948). This view changed when modern
genotyping methods revealed polyandry to be common in
a wide variety of animal groups (Birkhead 2000; Jennions and
Petrie 2000). Mating with different males usually entails costs
but does not always bring obvious benefits for females.
Various models have therefore been proposed to explain the
occurrence of multiple mating, including ‘‘trading up’’ to a
more desirable partner, or ‘‘bet hedging’’ to avoid committing
all resources to reproduction with a possibly inferior partner
(Jennions and Petrie 2000). Factors that may contribute to
some males being genetically more desirable than others
include sexually selected ‘‘good genes’’ (Pai and Yan 2002),
inbreeding avoidance (Stockley et al. 1993), and genetic
incompatibility from the interaction of selfish genetic elements
(Zeh JA and Zeh DW 1996). On the other hand, evidence of
the ubiquity of sexual conflict, often leading to increased costs
of mating, is also emerging (Arnqvist and Rowe 2005). Fertility
assurance may be another reason why females in many species
need to mate with multiple males. In theory, a male can
contribute enough sperm in a single mating to last a female’s
lifetime in most species; but in practice, sperm use is inefficient, and sperm stores may degrade over time. Sperm can be
energetically costly to produce, so males may choose to invest
less sperm in females of lower reproductive quality, or when
there is a low chance of fertilization per mating (Bonduriansky
2001; Wedell et al. 2002).
The evidence for multiple mating is extensive but comes
largely from vertebrates and insects (Birkhead 2000). Here, we
present genetic evidence for high levels of multiple mating in
the rough periwinkle (Littorina saxatilis), a marine prosobranch
gastropod with internal fertilization. In this intertidal, rocky
shore snail, reproduction is ovoviviparous, and dispersal
capability is therefore expected to be low. Multiple paternity in
the wild has previously been suggested in a related species,
Littorina obtusata (Paterson et al. 2001), which is oviparous. In
that study, females were mated in the laboratory to known
males, but a genetic analysis revealed that some offspring
derived from previous matings in the wild. Littorina saxatilis
has been observed to be highly promiscuous: when a few
males and females are placed in a dish with some moisture,
several matings per hour can often be observed. In this study,
we investigated the effect of promiscuity by estimating
multiple paternity rates exclusively from natural matings.
Males of L. saxatilis actively seek out mating partners by
following the mucus trails of other snails (Johannesson K,
Sundin A, Lindegarth M, Jonsson P, Havenhand J and
Hollander J, in preparation). Matings between snails of
different size or shell shape are interrupted more frequently,
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Journal of Heredity 2007:98(7)
possibly indicating assortative mating and partial reproductive isolation between ecotypes (Hollander et al. 2005). Such
findings have been interpreted in terms of male mate choice
(Johannesson K, Sundin A, Lindegarth M, Jonsson P,
Havenhand J and Hollander J, in preparation). Although
much more ubiquitous in nature, mate choice by females has
not been studied in this species. Here, we examine one aspect
of female choice: whether the pattern of sperm use by females
indicates postcopulatory sexual selection. Such mechanisms
are termed sperm competition if male driven and cryptic
female choice if female driven (Birkhead and Pizzari 2002).
Sometimes the distinction between them is artificial: providing opportunities for sperm competition within the
female can be a form of cryptic female choice (Birkhead
and Pizzari 2002). On the other hand, sperm competition can
also have negative consequences for the female, such as the
well-known example of seminal fluid in Drosophila having
toxic effects on the female (Chapman et al. 1995).
Littorinid snails commonly store sperm for long periods.
Sperm is received in the bursa copulatrix and later transferred
to the seminal receptacle for storage (Buckland-Nicks et al.
1999). Storage longer than 1 year has been observed in
L. saxatilis (Johannesson K, personal communication). This
provides ample opportunity for multiple paternity and sperm
competition. In this study, wild L. saxatilis females were
maintained in the laboratory, where they produced offspring
from stored sperm. The mothers and a sample of their
offspring were genotyped for microsatellite DNA markers at
5 loci, and the pattern of paternity was reconstructed from
their genotypes.
Materials and Methods
Collection and Incubation of Snails
In Sweden, 2 ecotypes of L. saxatilis occur: a larger and
thicker shelled morph (S) on sheltered shores and a smaller,
thinner shelled morph (E) on wave-exposed shores (Janson
1982). To control for ecotype differences, L. saxatilis
individuals were collected in July 2004 from one sheltered
and one exposed shore separated by 100 m on the island
Saltö (N58°53#, E11°10#) on the Swedish west coast. The
snails were sexed and measured, and 9 females of each
ecotype, with shell lengths of approximately 9 mm for the
E ecotype and 12 mm for the S ecotype (over the minimum
size for sexual maturity; cf. Erlandsson and Rolán-Alvarez
1998), were chosen for incubation. They were placed
individually in 11 jars with a constant flowthrough of
seawater. The jars were covered with fine mesh to prevent
escape and kept illuminated during daytime to promote the
growth of microalgae for the snails to feed on. The families
were incubated for a total of 2 and a half months, and at the
end of the experiment, all snails were collected on the same
date. One S female died and 2 did not produce any offspring
most likely owing to sterility caused by trematode infection,
which is not uncommon in Swedish populations, particularly
in the S ecotype (Granovitch and Johannesson 2000). The
remaining 15 females produced 23–302 juveniles, which
706
corresponded well to the number of embryos recorded
earlier in uninfected females of L. saxatilis: mean (range)
S ecotype 163 (1–526), E ecotype 64 (8–200); data from
Janson (1985). Out of these females, we randomly chose 4
families from each population for parental analysis. These
families contained 23, 24, 34, and 87 offspring for E ecotype
and 34, 40, 69, and 71 offspring for S ecotype, respectively.
From each family, we sampled 23 offspring randomly. The
largest diameter of each juvenile was measured to the
nearest millimeter, and the snails were frozen and kept at
70 °C until DNA extraction.
Genetic Analysis
DNA was extracted with the cetyltrimethylammonium
bromide protocol (Murray and Thompson 1980). For the
juveniles, the whole animal was used for extraction, and
about 1 mm3 of muscle tissue was taken from the females.
The snails were genotyped at 5 microsatellite loci: Lsub62,
Lsub32, Lsub8 (Tie et al. 2000), Lsax6, and Lx23 (Sokolov
et al. 2002). The polymerase chain reaction (PCR) protocol
was as follows: 2 ll DNA, 0.6 U Taq polymerase (TaKaRa
Inc., Otsu, Japan), 1 supplied buffer, 1.5 mM MgCl2, 0.2
mM of each diethylnitrophyenyl thiophosphate, 0.5 lM of
fluorescence-labeled forward primer (Sigma Proligo AB,
The Woodlands, TX) and 0.5 lM of reverse primer, and
ddH20 to a total volume of 12 ll. The amplification was
done in Eppendorf thermal cyclers in conditions described
by the primers’ authors, with the exception of different
annealing temperatures for Lsub62 (54 °C) and Lsub32 (55
°C). The DNA from some juveniles failed to amplify at all
loci probably due to poor extraction quality, resulting in
final sample sizes of 20–23 offspring per mother. The PCR
products were run on a Beckman Coulter CEQ 8000
automatic sequencer using standard chemical kits (Beckman
Coulter GmbH, Krefeld, Germany), and the results were
analyzed with the CEQ Fragment Analysis software.
Paternity Inference and Statistical Analyses
Initially, we used the GERUD1.0 program (Jones 2001) for
paternity inference, but because our data sets exceeded the
maximum limit of fathers in this program, we repeated the
analyses with the likelihood-based COLONY (Wang 2004).
COLONY has no maximum limit of contributing parents
and estimates the most probable number of fathers instead
of the minimum. The number of alleles in a family at the
most polymorphic locus, divided by 2, was calculated as
another measure of the minimum number of fathers. All
analyses other than estimating the number of fathers were
done exclusively in COLONY. COLONY provided the
most probable configuration of paternity, including assignations of every offspring to one of the estimated paternal
genotypes. If the program provided 2 options, the most
probable genotype was always chosen.
The genetic data were checked for occurrence of null
alleles by mother–offspring comparison. Two of the loci
(Lsax6 and Lsub8) were found to contain possible null
alleles (when half of the offspring of an apparently
Mäkinen et al. Multiple Paternity in a Snail
homozygous mother appeared to be homozygous for the
other allele). These loci were excluded for the analyses on
GERUD1.0. The results from COLONY were controlled
by doing 2 separate estimations: first, by allowing the
program to treat the null alleles as genotyping errors, and
second, manually assigning a value for the suspected null
alleles where a mother and her offspring did not match. The
latter is a more conservative approach according to Jones
and Ardren (2003). The subfamilies reconstructed with the
2 methods were identical, except for a few offspring in one
family that were assigned to different fathers by the 2
methods. In those cases, the analysis with unspecific
genotyping errors suggested several alternative paternal
genotypes with approximately equal posterior probabilities,
whereas the analysis with indicated null alleles still produced
one paternal genotype with much higher posterior probabilities than others (data not shown). The latter estimate was
chosen for the analysis. Therefore, it can be assumed that
null alleles were dealt with adequately in our analysis.
The average numbers of contributing males in the 2
populations (7 and 8 for S and E, respectively) were not
significantly different (t-test, P 5 0.56), and the data for the
2 ecotypes were pooled, resulting in a sample size of 8
families. After the families had been divided into subfamilies
by likelihood, a truncated Poisson distribution (Cohen 1960)
was fit to the distribution of offspring numbers per father.
The expected distribution was then compared with the
observed one by a chi-squared test. This tested the null
hypothesis that the sampling of sperm at fertilization was
random from a relatively large pool. Genetic similarity
between the maternal and estimated paternal genotypes,
expressed as the proportion of shared alleles Ps (Bowcock
et al. 1994), was calculated with Microsatellite analyzer
(Dieringer and Schlötterer 2003) and correlated to the
estimated fertilization success of the male (i.e., number of
offspring). To determine if the sperm from different males
was used sequentially or simultaneously (indicating sperm
mixing), we used offspring size at the end of the experiment
as a proxy for age, with the assumption that the time from
fertilization to emergence and the growth rate of juveniles
were relatively constant. Nested analysis of variance
(ANOVA) was used to test whether the offspring of
different males by the same female were significantly
different in size. The males that had less than 4 offspring
assigned to them were excluded from this test.
Results
The total numbers of alleles detected per locus were 8 in
Lsub62, 11 in Lsub32, 18 in Lsub8, 15 in Lsax6, and 27 in
Lx23 (see also Mäkinen et al. forthcoming). The paternal
allele count, divided by 2, in the most polymorphic locus
(Lx23) is a conservative estimate of the minimum number of
fathers and resulted in estimates of 3–5 sires for our samples
of up to 23 offspring. Minimum numbers of fathers, estimated from all loci in GERUD1.0, ranged from 3 to 8 fathers,
8 being the maximum limit of the program. The estimates of
Figure 1. The numbers of sires in 8 half-sib families of
Littorina saxatilis estimated by 3 different methods: allele count:
an estimation of minimum number of sires as the number of
paternal alleles at the most polymorphic locus (Lx23) divided by
2, GERUD1.0 software: estimation of minimum number of
sires based on multilocus genotypes (maximum limit 8), and
COLONY software: estimation of the most probable number
of sires based on multilocus genotypes (no maximum limit).
N: number of genotyped offspring/total number of offspring
per female.
most likely numbers of fathers from COLONY ranged from
4 to 10 contributing males per female with an average of 7.6
(Figure 1). The estimates of contributing fathers by different
methods matched relatively well, considering that a single locus
will result in lower estimates than multiple loci, and the first 2
methods estimate only the minimum number of fathers,
whereas the most probable number of sires given by
COLONY can be higher than the minimum (Figure 1). Further
results refer only to those from COLONY, and males or sires
refer to male genotypes reconstructed by the same program.
Female fertility (total offspring number) was not correlated
with the number of sires (Figure 1; R2 5 0.24, P 5 0.22).
The division of offspring among different males is
presented in Figure 2. The mean size of offspring of
different males within the same females was not significantly
different (nested ANOVA; between subgroups comparison:
degrees of freedom [df] 5 [16, 139], F 5 1.04, P 5 0.42),
suggesting either that the sperm from different fathers was
mixed during fertilization or that the size of juvenile snails is
not a good indicator of their age and/or fertilization time.
The distribution of paternal offspring numbers was not
significantly different from an expected distribution resulting from a random fertilization process, approximated by
a truncated Poisson distribution (Figure 3; df 5 5, v2 5
11.1, P 5 0.82), indicating that there was no apparent
reproductive skew. Only in one family had a single male
sired more than half of the offspring (F1; 57%). Genetic
similarity between the estimated male genotypes and
females, measured by the proportion of shared alleles
between them, had no effect on male fertilization success
(Figure 4; P . 0.05).
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Journal of Heredity 2007:98(7)
Figure 2. The most probable division of offspring into fullsib families estimated by COLONY. Each bar represents the
offspring of one female. The bars are divided and numbered
according to the number of offspring attributed to each male.
Discussion
The number of sires per brood was among the highest
detected in invertebrate species so far, with the exception of
social insects: the army ant and the honeybee (Table 1;
species that to our knowledge have relatively high levels of
multiple paternity in their respective groups were chosen for
this comparison). Moreover, our estimate is likely to be
a conservative one because only a subset of offspring was
sampled, and many sires were estimated to have only one
offspring in the sample. Such a high level of multiple
paternity suggests a high rate of multiple mating in the
species and is in need of evolutionary explanation because
mating rates are usually assumed to be the result of balancing
costs and benefits of mating (Arnqvist and Rowe 2005).
Mating can be costly for females in many ways, for
example, by taking time and energy or creating a higher risk
of predation (Watson et al. 1998). In L. saxatilis, the male
mounts the shell of the female during mating, which makes
Figure 3. Observed offspring number per estimated male
genotype compared with the expected offspring number,
approximated by a truncated Poisson distribution. The
distribution of offspring numbers was not significantly different
from the one expected from a random fertilization process.
708
Figure 4. The genetic similarity (proportion of shared alleles,
Ps) between a female and the estimated genotypes of males that
had sired her offspring, compared with the number of offspring
attributed to a male genotype. The genetic similarity had no
effect on fertilization success.
the pair more susceptible to being dislodged by waves
(Erlandsson and Rolán-Alvarez 1998; Johannesson K,
Sundin A, Lindegarth M, Jonsson P, Havenhand J and
Hollander J, in preparation). Multiple mating may also
deplete energy resources through reduced time for feeding,
which has been suggested to be the most generally occurring
cost of mating (Daly 1978). Littorina saxatilis males follow
the slime tracks of other snails but unlike L. littorea, a species
with pelagic larvae, seem unable to use chemical cues from
the tracks to differentiate between males and females
(Johannesson K, Sundin A, Lindegarth M, Jonsson P,
Havenhand J and Hollander J, in preparation). It has been
suggested that the loss of sexual identification in L. saxatilis
is a female adaptation: whereas females of a species with
a shorter reproductive season, like L. littorea, would advertise
their identity by pheromones, in a species with continuous
reproduction like L. saxatilis, it pays to avoid excess matings
(Erlandsson and Rolán-Alvarez 1998). In many species,
suboptimal matings result from male coercion, for example,
in marine turtles (Lee and Hays 2004), damselflies (van
Gossum et al. 2001), and dung flies (Mühlhäuser and
Blanckenhorn 2002). Such behavior, where multiple mating
may not be beneficial to the female, but the cost of
resistance is higher than the cost of mating, has been termed
convenience polyandry (Cordero and Andres 2002). In the
face of intense sexual conflict, females more often than not
evolve some countermeasures (Arnqvist and Rowe 2005).
Sometimes L. saxatilis females seem to be ‘‘wrestling’’ with
the males sitting on top of their shells, with the male
possibly using its shell as a lever to lift the female’s shell to
enable copulation (Hollander J, personal communication).
Similar mating behavior is seen in the hermaphrodite snail
Physa: a recipient snail sometimes tries to reject a sperm
donor by swinging the shell (DeWitt 1996). This behavior
usually makes the sperm donor dismount and does not
involve a risk of injury or death for the partners (DeWitt
Mäkinen et al. Multiple Paternity in a Snail
Table 1. Numbers of males contributing to a single female’s offspring in Littorina saxatilis, compared with other studied invertebrate
species with internal fertilization.
Species
N(off)
N(fem)
Males
Eciton burchellii (army ant)
Apis dorsata (honeybee)
20–60
111–288
6
13
15.5 ± 5.8
80.1 ± 3.6
1.7
2.6
15–146
89
16
20–236
34–58
40–46
20–23
20
4
7
15
5
3
8
3.1 ± 0.3
5 ± 0.8
3.8 ± 0.9
2 ± 1.1
3 ± 1.0
5 ± 1.0
7.6 ± 2.1
12.5
17.8
4.2
30
14.3
8.6
2.9
Drosophila mojavensis (fruit fly)
Drosophila melanogaster (fruit fly)
Gryllus bimaculatus (cricket)
Orconectes placidus (crayfish)
Loligo pealeii (squid)
Littorina obtusata (marine snail)
L. saxatilis (marine snail)
Offspr./male
Method
Reference
Allele count
Allele count
Denny et al. (2004)
Wattanachaiyingcharoen et al.
(2003)
Good et al. (2006)
Imhof et al. (1998)
Bretman and Tregenza (2005)
Walker et al. (2002)
Buresch et al. (2001)
Paterson et al. (2001)
Present study
KINSHIP
Allele count
PARENTAGE
HAPLOTYPES
Allele count
CERVUS
COLONY
The selected species showed, to our knowledge, relatively high levels of multiple paternity in natural or nearly natural mating situations in their respective
groups. N(off), number of offspring sampled per female; N(fem), number of females analyzed; Offspr./male, average sample size divided by average
number of sires detected in a family.
1996). Otherwise, no mating behavior that might indicate
physical attack has been reported in L. saxatilis. In addition,
species in which male harassment is suggested to be
a sufficient explanation for female multiple mating have
much lower rates of polyandry than L. saxatilis (e.g.,
Cordero and Andres 2002; Lee and Hays 2004).
Although multiple mating is likely to impose different
costs on females, there have to be some gains; they can be
direct (e.g., collecting nutrition gifts or ensuring fertilization)
or indirect (genetic benefits; Jennions and Petrie 2000).
Sperm may provide a direct benefit for females in L. saxatilis:
in addition to normal sperm cells, littorinids produce sterile
parasperm, which is digested in the bursa copulatrix of
the female before the storage of eusperm (Buckland-Nicks
et al. 1999). The parasperm may function either in sperm
competition or as nuptial gifts for the female, but the
importance of this nutritive gain is unclear. Sperm depletion
is unlikely to explain multiple mating in L. saxatilis because
all females in the experiment were found to continuously
produce offspring in the absence of males, nor did we find
evidence that females gained fertility benefits from multiple
paternity.
Genetic benefits of multiple mating may stem from
trading up (additional mating with a genetically superior
partner), postcopulatory sexual selection (cryptic female
choice and sperm competition) or bet hedging (mating with
several males when females are unable to identify the best
viability genes for the future; Jennions and Petrie 2000). We
did not find any evidence of reproductive skew or difference
in fertilization success between males that would indicate
the occurrence of sperm competition or cryptic female
choice. Moreover, the sperm storage organs of this species
are not especially complex (Reid 1996), and thus, it is
unlikely that sperm from up to 10 different males could be
differentially stored or capacitated by the female. Although
the females could have mated with even more partners than
observed from genetic data in this study, with some males
being later excluded by postcopulatory mechanisms, the
present data nevertheless indicate that sperm from a high
number of males were used indiscriminately.
The pattern of more males gaining paternity than can
reasonably be explained by females looking for superior
males was also reported in Lake Malawi cichlids (Kellogg
et al. 1995), tree swallows (Lifjeld et al. 1993), and insects,
(Zeh et al. 1997) among others. One plausible explanation
for this is bet hedging to avoid inbreeding (Stockley et al.
1993; Yasui 2001). Littorina saxatilis, unlike many marine
invertebrates, lacks free-swimming pelagic larvae, and both
juveniles and adults have limited mobility (Janson 1982).
Although the species frequently occurs at high population
densities, they are often concentrated in cracks and crevices
on rocky shores, with limited movement between them.
This increases the risk of inbreeding, and limited population
size is also a requirement for the evolution of bet hedging
(Yasui 2001). Although purging of deleterious alleles may
reduce inbreeding depression in populations with continuous inbreeding, the occurrence of deformed embryos at high
rates in some populations, even in unpolluted areas (Janson
1985), suggests that inbreeding depression is common in
L. saxatilis. Several intriguing observations of benefits of
outbreeding through mixed paternity have recently been
made in many animal groups including pythons (Madsen
et al. 2005), crickets (Tregenza and Wedell 2002), livebearing pseudoscorpions (Zeh JA and Zeh DW 2006), and
salmon (Garant et al. 2005).
Another suggested benefit of bet hedging is that higher
genetic diversity in the offspring increases the possibility
that some of them will survive in a heterogeneous or
fluctuating environment (Watson 1991; Yasui 1998).
However, simulations show that multiple mating solely for
genetic diversity can only evolve in very limited conditions,
for example, when full sibs compete more intensely than
half sibs and the environment is fine grained so that
individuals may choose between microhabitats (Yasui 1998),
or when the environment fluctuates unpredictably (Yasui
2001). The intertidal zone may be considered such a finegrained environment for L. saxatilis juveniles, but there is no
evidence for differential resource use between half sibs.
A high level of multiple paternity combined with sperm
storage may be beneficial in animals with low dispersal
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Journal of Heredity 2007:98(7)
ability and a fluctuating population size because it provides
a reservoir of genetic variability. Populations of L. saxatilis on
the Swedish west coast went through a severe bottleneck in
1988, when most of the snail populations were wiped out by
a toxic algal bloom. During the following few years, small
islands were rapidly repopulated, probably by snails occasionally rafting with algae (Johannesson K and Johannesson B
1995). These reestablished populations did not exhibit much
loss of genetic variation either in allozymes (Janson 1987) or in
microsatellite loci (Panova M, unpublished data). Moreover,
the species has managed to colonize remote locations, such as
the island of Rockall in the Atlantic, despite its limited mobility
and lack of pelagic larvae (Johannesson 1988). Colonizing
success of this species is not surprising, though, taking into
account the high fertility of females (Janson 1985), their ability
to store sperm for a year or more (Johannesson K, Sundin A,
Lindegarth M, Jonsson P, Havenhand J and Hollander J, in
preparation), and that a mated female might carry genetic
material from up to 10 males (present data). The genetic effects
of an introduction of a single multiply mated female may be
compared with a single queen establishing a colony in some ant
species. Interestingly, ant species where a single queen starts
a new colony are polyandrous, whereas in other ants, having
several foundresses per colony, queens are usually monogamous (Keller and Reeve 1994). Multiple mating and sperm
storage were also suggested to have evolved as adaptations
against genetic deterioration in the land snails Cepaea nemoralis
and Helix aspersa. As L. saxatilis, these species have limited
dispersal abilities and live in environments with frequent
stochastic disturbances (Murray 1964; Arnaud and Laval 2004).
In conclusion, multiple paternity is likely to be of great
importance for recolonization and population expansion in
L. saxatilis. Genetic bet hedging may evolve in response to
a fluctuating environment or a high risk of inbreeding in
small populations; both are plausible explanations for
multiple mating with lack of preference for a particular male
genotype in L. saxatilis females. However, the roles of direct
benefits of sperm, precopulatory female choice, and sexual
conflict remain to be experimentally assessed in this species.
Funding
European commission project, European Marine Genetic
Biodiversity (EVK3-CT-2001-00048; www.eumar.tmbl.gu.se);
Collianders stiftelse.
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Acknowledgments
Kerstin Johannesson’s and Johan Hollander’s knowledge on the mating
behavior and other peculiarities of Littorina saxatilis was invaluable to the
planning of this study. Christer Wiklund, Kerstin Johannesson, Elizabeth
Boulding, and 3 anonymous reviewers provided comments on earlier drafts.
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Received January 30, 2007
Accepted August 30, 2007
Corresponding Editor: Rob DeSalle
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