Department of Animal Ecology
Evolutionary Biology Centre
Uppsala University
Gametes and Speciation: from prezygotic to
postzygotic isolation
Murielle Ålund
Introductory Research Essay No. 100
Uppsala 2012
ISSN 1404 - 4919
ISRN UU - ZEK - IRE -- 100 -- SE
Gametes and Speciation: from prezygotic to
postzygotic isolation
Murielle Ålund
Department of Animal Ecology
Evolutionary Biology Centre
Uppsala University
Norbyvägen 18D
SE-752 36 Uppsala
Sweden
ISSN 1404 - 4919
ISRN UU - ZEK – IRE -- 100 -- SE
Contents
Abstract ............................................................................................................................................... - 4 1.
Speciation and the evolution of reproductive barriers ................................................................. - 5 -
2.
Postmating prezygotic barriers or gametic isolation ................................................................... - 7 -
3.
4.
2.1
Sperm competition and conspecific sperm precedence ....................................................... - 9 -
2.2
Reproductive proteins and sperm-egg interactions ........................................................... - 11 -
2.3
Cryptic female choice and sexual conflict......................................................................... - 13 -
Intrinsic postzygotic isolation: hybrid sterility .......................................................................... - 15 3.1
The Dobzhansky-Muller model......................................................................................... - 16 -
3.2
Haldane’s rule.................................................................................................................... - 18 -
3.3
Spermatogenesis and the genetics of hybrid male sterility................................................ - 19 -
A model system for the study of pre- and postzygotic isolation: the Ficedula flycatchers....... - 21 -
Conclusions ....................................................................................................................................... - 24 Acknowledgements ........................................................................................................................... - 25 References ......................................................................................................................................... - 25 -
Abstract
Speciation lies at the heart of evolutionary biology and researchers have been trying to understand the
mechanisms leading to the evolution of reproductive isolation since over 250 years. Premating barriers
(i.e. barriers preventing heterospecific individuals to mate with each other) and extrinsic postzygotic
isolation (i.e. environmental factors affecting the fitness of hybrid individuals) have been studied in
many taxa. However, little is known about what is happening at the gametic level, both before
heterospecific fertilization (i.e. postmating prezygotic or gametic isolation) and in hybrid individuals
(i.e. intrinsic postzygotic incompatibilities). In this essay, I will give an overview of the role gametes
play in the evolution of reproductive isolation. I conclude that gametes and reproductive proteins
evolve quickly, under strong influence of sexual and sexually antagonistic selection. Gametes are very
diverse between species and sperm competition and female cryptic choice can lead to higher
fertilization success of sperm from conspecific males. In the hybrid offspring, spermatogenesis can be
easily disturbed by small differences in gene expression and this leads to a greater number of genes
causing hybrid sterility compared to hybrid inviability among taxa. Following Haldane’s rule, the
heterogametic sex is the first to be affected by hybrid incompatibilities, but different mechanisms
seem to cause inviability and sterility and taxa with heterogametic males or heterogametic females
might be affected differently. I end this review by focusing on one particular model system for
studying speciation: the Ficedula flycatchers. Much is known about the ecological factors affecting
speciation and hybridization between pied and collared flycatchers and new molecular data give
insights into the genetics of speciation, but the role of gametes has not been studied in this system.
Studies on gamete divergence and hybrid gamete production in the flycatchers will allow us to get a
better idea of the role of gametes in speciation in a wild organism with homogametic males.
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1.
Speciation and the evolution of reproductive barriers
The existence, formation and maintenance of species as different entities have puzzled evolutionary
biologists for over a century. A quick look at the number of publications including “The Origin of
Species” in their title since Darwin’s book in 1859 gives an idea on the variety of fields working on
the subject, including systematics (Mayr, 1942; Avise, 1997), genetics (Dobzhansky, 1937), molecular
biology (Manwell & Baker, 1970; De la Cruz & Davies, 2000), ecology (Dieckmann & Doebeli, 1999;
Schluter, 2001; Gavrilets, 2004) and behaviour (Van Doorn et al., 2009), among many others. The
problem with studying how new species are formed is that speciation can rarely be observed directly
since the process can take millions of years to be completed. As a result, researchers have to focus
their studies on diverging populations or incipient species that might never become separate species,
or work in the laboratory with species that diverged long ago (Rice et al., 2011).
The definition of a species has been debated for decades and there is no uniformly accepted definition
but at least 25 different “species concepts”. Scientists use different species concepts depending on the
organism they are working with (sexual or asexual) and their questions of interest (systematics,
genetics or ecology). The most widely used definition when describing sexually reproducing
organisms is the Biological Species Concept, which states that individuals belonging to the same
species can interbreed and produce viable and fertile offspring, while being reproductively isolated
from individuals of other species (Mayr, 1942). Following this definition, research in speciation
generally focuses on understanding the evolution of reproductive isolation (Coyne & Orr, 2004).
Reproductive barriers, which include anything that impedes the exchange of genes between members
of different populations, are classified in different categories, depending on when in the reproductive
cycle they are acting. Prezygotic isolation mechanisms prevent individuals from choosing each other
as mates (premating barriers) or impair fertilization success (postmating prezygotic barriers).
Premating barriers can be caused by ecological isolation, when populations use different habitats or
resources (habitat isolation) or when their breeding time is different (temporal isolation). Divergence
in a secondary sexual character (e.g. colour, song or pheromones) can also reduce mate attractiveness
between populations (behavioural isolation) or the morphology of the reproductive structures might
simply be incompatible (mechanistic isolation). Finally, postmating prezygotic barriers or gametic
isolation include all reproductive barriers acting between copulation and fertilization (see chapter 2).
Postzygotic isolation mechanisms include different forms of selection against hybrids, either caused
by extrinsic factors reducing their fitness (e.g. difficulties to find an ecological niche or to acquire a
mate because of their intermediate phenotype) or intrinsic genetic incompatibilities leading to hybrid
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sterility or inviability (Coyne & Orr, 2004, see chapter 3). Figure 1 below schematically summarizes
the different forms of reproductive barriers between heterospecific individuals.
Figure 1: Schematic representation of the reproductive barriers, starting with premating barriers preventing
individuals from choosing each other as mates, then gametic isolation (postmating prezygotic) inside the female
reproductive tract and finally picturing different forms of selection against hybrids (postzygotic barriers).
Reproductive isolation is often thought to evolve as a by-product of ecological divergence in allopatry
(Schluter, 2001). When there is a physical barrier to the exchange of migrants between populations,
the populations will diverge over time, simply by genetic drift or by adaptation to different
environments (resources, predators, competitors, etc.). Given enough time, this will lead to the
evolution of reproductive isolation (Schluter, 2001). Premating barriers and behavioural isolation in
particular can evolve relatively quickly and are reversible, as is extrinsic postzygotic isolation. Genetic
incompatibilities on the other hand might take millions of years to appear. Mammals are generally still
capable of producing hybrids after 2-3 millions, frogs after 21millions and birds after 22 million years
(Coyne & Orr, 2004). Secondary contact, where populations are found in sympatry again after the
disappearance of a physical barrier, is a crucial point in the speciation process. If there was too little
divergence between the populations in allopatry, they might merge into one species again.
Alternatively, premating barriers might be already so strong that there is no interbreeding at all: the
species were formed in allopatry. Finally, hybridization might lead to the formation of unfit hybrids, in
which case there should be natural selection for enhanced prezygotic barriers, a process called
reinforcement (Grant et al., 1996; Schluter, 2001). The outcome of secondary contact will also be
strongly influenced by environmental factors, including resources and territory availability,
competition and predation (Schluter, 2001).
In the more controversial scenario of sympatric speciation, reproductive isolation evolves without any
geographical barrier while there is still free exchange of genes, due to a divergence in resources use
coupled with strong assortative mating. The problem with this scenario is that hybridization and gene
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flow between the diverging populations is likely to break down the associations between divergent
traits and preference (Coyne & Orr, 2004). For sympatric speciation to occur, a linkage between the
diverging trait and the associated preference, so-called “magic-trait” is needed, either through linkage
disequilibrium at the molecular level or by having one trait simultaneously involved in ecological
functions and in mate preference or species recognition (Servedio et al., 2011).
It is generally accepted that speciation occurs as a combination of different reproductive barriers given
enough divergence time, but it is difficult to know the relative importance of the different barriers and
their order of appearance over time when observing complete species or species that just started
diverging (Coyne & Orr, 2004; Rice et al., 2011). A solution to that may be to observe hybrid zones
with species that came into secondary contact after a long period of divergence in allopatry, since it
allows us to study the mechanisms evolving to avoid costly hybridization (Barton & Hewitt, 1985;
Rice et al., 2011). In the next chapters, I will concentrate on two types of reproductive barriers
involving gametes (i.e. sperm and egg): postmating prezygotic and intrinsic postzygotic isolation. In
reviewing the existing literature, I will mostly concentrate on studies on animals, since it is the focus
of my own work, although reproductive barriers have also been extensively studied in plants.
2.
Postmating prezygotic barriers or gametic isolation
Postmating prezygotic isolation, also called gametic isolation, includes any reproductive barrier acting
between copulation and fertilization (Coyne & Orr, 2004; Howard et al., 2009). Postcopulatory
prezygotic barriers are physiological and often difficult to investigate and as a result they are
unrepresented among the studies on reproductive isolation (Coyne & Orr, 2004; Birkhead & Brillard,
2007; Martín-Coello et al., 2009). As discussed above, premating barriers and extrinsic postzygotic
mechanisms evolve faster but are also easier to study, since they mostly depend on ecological factors.
However, when premating isolation mechanisms are not efficient enough to prevent heterospecific
individuals from mating (i.e. hybridization), postmating prezygotic reproductive barriers might play an
important role in preventing the costly production of unfit hybrid offspring (Ludlow & Magurran,
2006; Immler et al., 2011). It is also likely to be more important in external than in internal fertilizers,
particularly in broadcast spawners, since they are releasing gametes into the water without any
premating interaction (Landry et al., 2003).
Several factors can contribute to a reduced fertilization success in heterospecific mating compared to
mating between individuals of the same species (i.e. conspecifics). First, there could be too few sperm
transferred or a poor storage of the sperm in the female tract. A reduced gamete transfer and loss of
sperm from the female tract were both observed in heterospecific mating in Drosophila (Price et al.,
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2001; Sagga & Civetta, 2011). Furthermore, male gametes may have lower survival or an abnormal
behaviour in a foreign reproductive tract, as was observed in the ground cricket where sperm was
immotile in the reproductive tract of heterospecific females (Gregory & Howard, 1994). The proteins
of the seminal fluid might also fail to stimulate ovulation or oviposition, as observed in Drosophila
(Price et al., 2001). There may also be a lack of chemical attraction between gametes (e.g. in marine
broadcast spawners, Miller, 1997) or incompatibilities between sperm and egg-recognition proteins
(Vacquier, 1998, see section 2.3). Finally, when females mate both with conspecific and heterospecific
males, conspecific sperm precedence is likely to occur (Servedio, 2001; Coyne & Orr, 2004, see
section 2.1).
Gametes and sperm in particular can evolve very quickly. Sperm morphology has been found to vary
widely between closely related species in several taxa including mammals, insects and marine
invertebrates (Howard et al., 2009). Even within populations, sperm morphology has been shown to
vary between individuals in response to their social environment (Immler et al., 2010). A rapid change
in morphology could contribute to differential fertilization success between species and thus to the
rapid evolution of gametic isolation. Moreover, reproductive proteins are known to evolve quickly and
several studies showed a rapid differentiation and strong selection pressures on genes coding for
reproductive proteins in Drosophila and marine invertebrates (see section 2.3). Similarly, genitalia and
female sperm storage organs also vary greatly across species (Howard et al., 2009).
Gametic isolation has been observed throughout the animal kingdom, from corals and molluscs to
mammals and birds. However, there is an overrepresentation of insects and marine invertebrates
among studies on postmating prezygotic barriers. The later can be explained by the fact that broadcast
spawners do not have any other form of premating barriers and there is thus strong selection on
gamete compatibility. Insects on the other hand represent an extremely species-rich taxa and are very
well suited for laboratory experiments, hence the abundance of data available for insect species
(Howard et al., 2009). Gametic isolation involves many different barriers that can be more or less
effective. If there is potentially strong pressure to avoid heterospecific mating in particular situations,
it might also be costly to develop too strong barriers that would even impair conspecific mating. If
gametic isolation has already been observed between populations at the intraspecific level (Coyne &
Orr, 2004), it can also take millions of years to be totally effective, in birds for example (Birkhead &
Brillard, 2007). In the following sections, I will in more detail describe two specific mechanisms of
gametic isolation: conspecific sperm precedence and interaction between reproductive proteins. I will
also discuss the role of female cryptic choice and sexual conflict in the evolution of reproductive
isolation.
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2.1
Sperm competition and conspecific sperm precedence
Sperm competition occurs whenever ejaculates from several males compete for the fertilization of the
same set of ova (Parker, 1970). Polyandry is known to occur in most taxa, and as females copulate
with several males, sperm of different males might co-occur in the female reproductive tract at a given
time and compete for fertilization. There will be selection for traits that increase the fertilization
success of sperm and females can exert sperm selection or female cryptic choice and bias the
fertilization success in favour of certain males (Eberhard, 1996; Pizzari & Parker, 2009, see section
2.3). Sperm number and sperm quality are the two major determinants of an ejaculate’s fertilization
success. Sperm size, morphology, swimming velocity, metabolic performance, longevity and seminal
fluid peptides are all thought to contribute to sperm quality (Pizzari & Parker, 2009). The relationship
between sperm size or morphology and the outcome of sperm competition is still not well understood
(Pizzari & Parker, 2009). Over 20 years ago, Gomendio and Roldan proposed that longer sperm swim
faster (Gomendio & Roldan, 1991) and this has been confirmed by numerous studies since then
(reviewed in Helfenstein et al., 2009; Pizzari & Parker, 2009). The relationship between sperm size
and longevity is still debated and contradicting results have been reported in different studies
(Helfenstein et al., 2009). A trade-off between sperm velocity and longevity is often assumed, the idea
being that fast sperm expend all their energy quickly and thus suffer earlier senescence (Pizzari &
Parker, 2009). If long sperm swim faster and also expend their energy earlier, small sperm are likely to
live longer and there should be a trade-off between the production of short, long-lived sperm and long,
fast-swimming ones. When there is little sperm competition, it might be more beneficial to have longliving sperm that will remain long enough in the reproductive tract to fertilize a high number of eggs
(Pizzari & Parker, 2009). If sperm competition is high on the other hand, there will not be any benefit
of having long-lived sperm, as the number of competitors will increase over time and the priority will
be to invest in highly competitive ejaculates containing sperm, which are able to reach the ovum first
directly after insemination (before the female re-mated) (Engqvist, 2012). Different selection pressures
or different strategies are likely to be found in different populations and this could have consequences
for the outcome of sperm competition at secondary contact after heterospecific mating. Hybridization
will be more likely to occur between a male from a population with high sperm competition and a
female from a population with low sperm competition than the opposite, leading to asymmetric or
unidirectional hybridization, as observed in three species of mouse for example (Birkhead & Brillard,
2007; Martín-Coello et al., 2009).
“Conspecific sperm precedence” is defined as the capacity of sperm from conspecific males to gain
more fertilizations than sperm from heterospecific males in situations of sperm competition (Howard,
1999). This can happen either because conspecific sperm outcompetes heterospecific sperm in the race
to the ovum (at any stage from the entry in the female reproductive tract to the deposition in storage
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organs to the actual fusion of the sperm with the ovum), or because heterospecific sperm fails to
fertilize the egg. In both cases, the end result is the production of fewer hybrid offspring than would be
expected given the proportion of heterospecific mating (Howard, 1999). Conspecific sperm
precedence is potentially a very strong reproductive barrier when females randomly mate with
conspecific and heterospecific males. However, as one species becomes rarer and the rate of encounter
of a conspecific male decreases, more hybrid offspring will be produced and gene flow will increase if
postzygotic barriers are not strong enough (Howard, 1999).
Studies on conspecific sperm precedence are often needed to detect a barrier at the gametic level
between species. Some studies failed to find any incompatibility between species until sperm
precedence was analysed, as heterospecific sperm may be able to fertilize the egg in absence of
competition with conspecific sperm (Howard et al., 2009). This is the case for the ground cricket
Allonemobius fasciatus and its sister species A. socius, which produce fertile hybrid offspring when a
female is mated to a heterospecific male only, while fertilization by heterospecific sperm rarely occurs
when females are mated to both conspecific and heterospecific males (Howard et al., 1998).
Conspecific sperm or pollen precedence seems to be a wide-spread mechanism reducing hybridization
and has been identified in Drosophila (Price et al., 2000), mouse (Martín-Coello et al., 2009), flour
beetles (Wade et al., 1994), grasshoppers (Hewitt et al., 1989), mussels , Iris, monkey flowers,
sunflowers (reviewed in Howard, 1999 and Coyne & Orr, 2004) and even at the intraspecific level
between populations of guppies separated for two million years (Ludlow & Magurran, 2006). Among
external fertilizers, conspecific sperm precedence was observed in one type of heterospecific crossing
but not in the other in two hybridizing sunfish species (Immler et al., 2011) and in sea urchins, only
very few eggs are fertilized by heterospecific sperm compared to conspecific (Lillie, 1921). The latter
is due to the specificity of sperm and egg binding proteins, which will be discussed in the following
section.
When females are in contact with both conspecific and heterospecific males, multiple mating and
mechanisms of conspecific sperm precedence allow the female to minimize gamete waste in the
production of unfit hybrid offspring or eggs that will not hatch. This was observed in two closely
related species of ladybirds which show only 5% of egg hatching in heterospecific compared to
conspecific mating, but achieve normal hatching rates when females were mated to both
heterospecifics and conspecifics (Nakano, 1985). However, if mechanisms of conspecific sperm
precedence are likely to evolve as by-products of differential sperm competition pressures in different
populations and will be advantageous for females when premating barriers are incomplete, selection
for postmating prezygotic barriers is unlikely in males since it contributes to the waste of gametes they
already inseminated in the female (Coyne & Orr, 2004). The potential sexual conflict resulting from
this will be discussed in section 2.3.
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2.2
Reproductive proteins and sperm-egg interactions
Reproductive proteins are usually defined as proteins acting after mating by influencing gamete
transfer, usage, storage and fertilization (Swanson & Vacquier, 2002b). Reproductive genes are among
the 10% most rapidly evolving genes and a high divergence in reproductive proteins has been found in
many different organisms, including marine ciliates, unicellular green algae, fungi, Arabidopsis,
abalones, sea urchins, Drosophila, crickets, rodents and primates (Swanson & Vacquier, 2002a; b;
Clark et al., 2006; Turner & Hoekstra, 2008; Marshall et al., 2011). The most well-known cases of
rapid evolution of reproductive proteins are found in abalones and sea urchins, both marine
invertebrates and broadcast spawners. These external fertilizers lack any form of behavioural
interaction before mating, and gamete recognition proteins are thus likely to be an important barrier to
heterospecific mating. In the abalone, the sperm contains lysin in its acrosomal capsule, which
dissolves the vitellin envelope of the egg. Lysin has to bind to a receptor on the vitellin envelope
named VERL in order to create a hole in which the sperm can swim (Turner & Hoekstra, 2008). This
species-specific protein complex has been shown to evolve 50 times faster than the fastest evolving
mammalian proteins (Swanson & Vacquier, 2002a). In sea urchins, bindin, a protein binding to a
receptor on the egg surface is also evolving fast and is under positive selection (Turner & Hoekstra,
2008).
In internal fertilizers, the sperm has to pass multiple barriers before reaching and fertilizing the ovum
(see Figure 1). Numerous proteins play an important rule at different stages of this process (reviewed
in Clark et al., 2006) When sperm enters the reproductive tract, it has to face different pathogens and
resist against the female immune system. Antibacterial proteins and prostaglandin decreasing female
immune response have been observed in Drosophila and mammals. Chemotaxis (chemical attraction)
between the egg and the sperm can be found in humans, facilitating the navigation of sperm towards
the egg. Just after ovulation, both sperm and egg have to go through maturation for fertilization to be
possible, a mechanism that is likely to be controlled by specific substances released in the oviduct.
Sperm is often stored in the female reproductive tract, from a few days to several months or even over
years. Seminal proteins have been shown to affect sperm storage in Drosophila. When reaching the
ovum, the sperm first has to open a hole in the egg coat and then bind to the egg membrane and fuse
with the nucleus, a process that is controlled by species specific molecules and enzymes present in the
sperm head and on the egg membrane (Clark et al., 2006). In mammals, one of these proteins is the
zona pellucida glycoprotein 3 (ZP3), one of the egg-coat proteins, which is binding sperm and has
been shown to be under positive selection (Turner & Hoekstra, 2008).
Protein involved in all the steps mentioned above as well as seminal proteins inducing ovulation or
controlling females remating rates and ion-channels influencing sperm motility have all been shown to
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be rapidly diverging between closely related species in different taxa (Howard, 1999; Swanson &
Vacquier, 2002a; b; Clark et al., 2006; Turner & Hoekstra, 2008). In Drosophila, over 100 accessory
gland proteins (Acps) have been identified in the seminal fluid and affect various processes in the
female reproductive tract and they are twice as diverse as non-reproductive proteins (Swanson &
Vacquier, 2002a; Turner & Hoekstra, 2008). Recently, proteomic analyses have allowed the study of
protein contents of ejaculates of non-model species as well. This was the case in crickets for example,
where a comparison between the proteomes of two closely related species showed high divergence and
positive selection on ejaculates, indicating that the rapid evolution of reproductive isolation through
postmating, prezygotic isolation is possible in 30’000 years only (Marshall et al., 2011). The use of
molecular methods also allows broader comparisons of selective forces acting on gamete recognition
genes. Berlin and colleagues showed evidence of positive selection on six gamete-recognition genes
across several species of birds and concluded that the selection mechanisms were similar to those
found in mammals (Berlin et al., 2008).
Different hypotheses have been proposed to explain the fast evolution of reproductive proteins
(Howard, 1999; Swanson & Vacquier, 2002b; Findlay & Swanson, 2010). One common explanation
for fast divergence of reproductive proteins is pathogen avoidance. Both eggs of marine invertebrates
and the reproductive tract of internal fertilizers are exposed to all kinds of pathogens and female egg
coat proteins are thought to diverge rapidly in order to counter-act rapid pathogen evolution (Howard,
1999; Swanson & Vacquier, 2002b; Findlay & Swanson, 2010). Another hypothesis is reinforcement
or selection against hybridization, where reproductive proteins evolve faster in sympatry than in
allopatry to prevent costly heterospecific fertilization. This will most likely act on egg proteins to
avoid fertilization by the wrong sperm, since egg production is costlier than the production of
numerous sperm. Furthermore, once sperm has been released, any mechanism reducing the chance of
fertilization will induce a waste of this sperm. Reinforcement of gamete isolation has been observed in
sea urchins (Palumbi, 2009) and in Drosophila (Matute, 2010). In the latter example, a lower
proportion of fertilized eggs following heterospecific mating were observed in a hybrid zone compared
to similar crosses between allopatric populations. Furthermore, heterospecific sperm loss was more
pronounced between sympatric than between allopatric populations (Matute, 2010). Interestingly,
populations experimentally kept in sympatry showed signs of enhanced gametic isolation after four
generations only (Matute, 2010). Finally, sexual selection on gametes (or cryptic female choice) and
sexual conflict have been hypothesized to drive the fast evolution of reproductive proteins (Howard,
1999; Swanson & Vacquier, 2002b; Findlay & Swanson, 2010). These two mechanisms are described
in more details in the following section.
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2.3
Cryptic female choice and sexual conflict
Cryptic female choice is defined as any mechanism acting after copulation that biases paternity
towards one male (Eberhard, 1996). The female reproductive tract consists of a variety of strong
barriers that allow only sperm of vigorous or so-called “good quality” males to fertilize the egg
(Pitnick et al., 2009). Through polyandry and extra-pair copulations, situations will occur where sperm
of different males compete for the fertilization of the same egg and this should allow female to
“select” the sperm of certain males over others. There are three major competing hypotheses
describing the benefit of such choice. According to the sexually selected sperm hypothesis,
fertilization by males with sperm of superior competitive ability will result in sons with highly
competitive sperm and hence higher reproductive success. The good sperm hypothesis on the other
hand proposes that males with good sperm are also of higher overall genetic quality (Pitnick et al.,
2009). Finally, the genetic compatibility hypothesis suggests that cryptic female choice functions as a
mechanism to avoid fertilization by sperm of genetically incompatible males, possibly through antisperm immune reactions (Zeh & Zeh, 1997). A positive relationship between sperm competitive
ability and offspring fitness was found in the yellow dung fly and in a marsupial, while links between
condition and sperm characteristics were observed in red deer, guppies, Atlantic cod, dung beetle and
Drosophila (Pitnick et al., 2009). A recent study showed biased fertilization towards sperm carrying
particular MHC alleles in a socially monogamous bird (Alcaide et al., 2012), a mechanism that could
both be linked to genetic compatibility or superior genetic quality, but no data on offspring fitness
were analysed. Fertilization bias against inbreeding has been more extensively studied and was
reported in mice, lizard, field cricket, Drosophila and C. elegans (Pitnick et al., 2009).
The exact mechanisms involved in female cryptic choice are difficult to study and still poorly
understood. Females can bias fertilization towards a particular genotype either by using physical or
chemical barriers or by actively ejecting sperm (Zeh & Zeh, 1997; Pitnick et al., 2009). In birds for
example, less than 2% of the sperm ever reaches the storage organs (Pitnick et al., 2009). These strong
barriers select for offensive adaptations in males, evolving aggressive ejaculates with higher quantity
or faster sperm, more efficient at finding fertilization sites or at depletion of previous sperm, among
other characteristics (Arnqvist & Rowe, 2005). However, a highly fertile ejaculate will in turn enhance
the risk of polyspermy (the penetration of the egg by several sperm at the same time). Polyspermy is
often detrimental to the embryo (it is lethal in mammals for example), thus it is in the interest of the
female to evolve resistance against it (Palumbi, 2009). This situation of conflict between the
evolutionary interests of the two sexes (here males willing to enhance their fertilization success and
females avoiding polyspermy and selecting for sperm quality) is more generally termed sexual conflict
(Parker, 1979). It will usually result in sexually antagonistic coevolution or coevolutionary arm race
between the sexes with the evolution of persistence adaptations in males and enhanced resistance in
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females (Arnqvist & Rowe, 2005, see Figure 2). This can lead to extreme cases of indirect costs to
females with for example the evolution of harmful genitalia or injection of sperm directly into the
haemolymph in insects. On the other hand, the evolution of too strong barriers by the females might
result in no fertilization at all (Arnqvist & Rowe, 2005).
Figure 2: Schematic representation of sexually antagonistic selection at the gametic level or the co-evolutionary arm
race occurring between gametes of the two sexes in polyandrous organisms.
Sexual conflict and sexually antagonistic evolution are thought to drive the evolution of a variety of
reproductive proteins in both sexes. The best described system here again is Drosophila, where male
accessory gland proteins have been found to increase egg production, laying and ovulation, influence
sperm competition, storage and utilization, enhance courtship receptivity, decrease food intake and
suppress immune response of females. Some proteins also induce changes in the conformation of the
female reproductive tract (e.g. induced muscle contraction to push sperm towards the storage organs).
These accessory proteins have been shown to decrease longevity in female Drosophila (Arnqvist &
Rowe, 2005). Female proteins can in turn induce changes in the ejaculate, including sperm maturation
and activation of certain proteins (Arnqvist & Rowe, 2005; Pitnick et al., 2009).
This continuous arm race between males and females leads to a very fast evolution of reproductive
traits and the maintenance of a high diversity despite their evolutionary importance. This in turn will
eventually lead to reproductive isolation between allopatric populations (Arnqvist et al., 2000). In a
comparative study, Arnqvist and colleagues showed a four-fold increase in speciation rates in
polyandrous insect clades, where there is more opportunity for sexual selection and sexual conflict,
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compared to monoandrous clades where sperm competition and female cryptic choice is unlikely to
occur (Arnqvist et al., 2000). This demonstrates the importance of polyandry, sexual selection and
sexual conflict in the evolution of reproductive isolation and hence the formation of new species.
3.
Intrinsic postzygotic isolation: hybrid sterility
In the previous section of this essay, I explored several mechanisms preventing mating between
individuals of different species. Prezygotic reproductive barriers are numerous and diverse, however,
also often incomplete. When individuals of different populations lack complete reproductive isolation
at secondary contact or in sympatry, this can lead to hybridization. Hybridization is quite common in
nature, according to a survey by Mallet, it occurs in 10% of all animal species and 25% of all plant
species at least (Mallet, 2005). Similarly, 10% of all bird species hybridize or have hybridized (Grant
& Grant, 1992). In general, hybrids perform poorly as compared to their parents (Burke & Arnold,
2001) and different form of selection against hybrid individuals are grouped under the term
“postzygotic isolation”. Extrinsic postzygotic isolation refers to lower hybrid fitness due to a poor
match with its environment (e.g. difficulties to acquire resources or mates due to intermediate
phenotype), while intrinsic postzygotic isolation generally describes hybrid sterility and/or inviability
due to genetic incompatibilities (Turelli et al., 2001; Coyne & Orr, 2004).
Intrinsic postzygotic isolation can be caused by different ploidy levels, different chromosomal
rearrangements (chromosomal speciation), divergent alleles not functioning together (DobzhanskyMuller incompatibilities, see section 3.1) or an infection by different endosymbionts causing
cytoplasmic incompatibilities (Coyne & Orr, 2004). Chromosomal speciation is more common in
plants than animals since self-fertilization increases the probability of fixation of underdominant
rearrangements and fertility can be restored in plants through higher ploidy levels (e.g. tetrapoloidy for
diploid plants). In animals, the presence of degenerated sex chromosomes (Y or W) enhances the
probability of the evolution of genetic incompatibilities (Coyne & Orr, 2004). Cytoplasmic
incompatibilities can be caused by different infectious agents, of which Wolbachia is a well-studied
example. This bacterial parasite is maternally transmitted and induces the disruption of paternal
chromosome processing /and or mitosis, causing hybrid inviability when populations adapted to
different strains of Wolbachia meet at secondary contact (Coyne & Orr, 2004).
Postzygotic isolating mechanisms are generally less studied than premating barriers and most of the
literature on postmating isolation is biased towards Drosophila and other invertebrates (Birkhead and
Brillard, 2007). The first studies of hybrid sterility were performed at the beginning of the last century,
- 15 -
generally in domestic animals. These experiments consisted of crossing different species of equids and
bovids (Crew & Koller, 1936; Craft, 1938), ducks (Rigdon & Mott, 1965) and phasianids (Yamashina,
1943) and performing cytological analyses on the hybrid offspring. The outcome of numerous
interspecific crosses have been described since then, mostly in captive (Johnsgard, 1970; Price &
Bouvier, 2002; Lijtmaer et al., 2003) and wild derived populations crossed in the laboratory (e.g.
Close et al., 1992; Kopp & Frank, 2005; Britton-davidian et al., 2005; Borodin et al., 2006; Good et
al., 2008). More recently, studies have focused on the genetics of intrinsic postzygotic
incompatibilities, in the search of “speciation genes”, mainly in mice and Drosophila, typically
crossing laboratory species that diverged long ago (reviewed in Turelli et al., 2001; Orr, 2005;
Presgraves, 2010). A general pattern emerging from these studies is that genes causing hybrid sterility
seem to be affecting one sex only and they are ordinary genes having normal functions within species
(Coyne & Orr, 2004).
A major problem when studying species that are completely reproductively isolated is that it is often
impossible to determine the origin of hybrid sterility or inviability, because many genetic
incompatibilities will have accumulated over time (Orr, 1995; Turelli et al., 2001; Rice et al., 2011). A
solution to this problem is to study incipient speciation or hybrid zones where species started
diverging in allopatry and have come into secondary contact before being completely reproductively
isolated (Barton & Hewitt, 1981; Grant & Grant, 1992; Rice et al., 2011). Hybrid sterility has been
studied in some wild-derived populations of house mice and Drosophila (Kopp & Frank, 2005;
Britton-davidian et al., 2005; Good et al., 2008) but it remains difficult to detect in nature, since
hybrids need to be identified and their fitness has to be quantified accurately (The Marie Curie
Speciation Network, 2012). As a result, very few studies describe intrinsic genetic incompatibilities in
hybrids resulting from natural hybridization (see Rhymer & Simberloff, 1996) and these studies are
usually based on single or scarce observations of hybrid individuals (e.g. Reifová et al., 2011).
3.1
The Dobzhansky-Muller model
When hybridization occurs at secondary contact between populations that have diverged in allopatry
(i.e. geographically isolated), some alleles that have never been in contact before might interact in the
hybrid genome, provoking genetic incompatibilities and possible dysfunctions, since there was no coevolution between those alleles for many years (Moehring, 2011). This is known as “DobzhanskyMüller incompatibilities” (Dobzhansky, 1936; Müller, 1940). Figure 3 below describes this
phenomenon in more details.
- 16 -
Figure 3: Dobzhansky-Muller model: different mutations occur in different populations in allopatry and the new
alleles, which have never co-occurred in the same genetic background before, cause incompatibilities in the hybrids.
When populations are isolated from each other for a long time, even if they experience similar
environmental conditions, they will accumulate divergent mutations just by chance. This is because
genomes are so complex that there are many different routes to the same adaptation (Turelli et al.,
2001). New alleles emerging in each separate population will be well adapted to their own background
genome, but may cause incompatibilities when they occur together with genes that they have not coevolved with in hybrid individuals. Through this process, alleles that have positive effects in one
population can be detrimental in a hybrid background, and hybrid incompatibilities can arise without
any fixation of “negative”-effect allele and without maladaptive intermediates (Turelli et al., 2001).
The model above describes a very simple case where only one allele diverges in each population.
Dobzhansky-Muller incompatibilities typically follow a “snowball effect”: genetic divergence
increases linearly with diverging time and incompatibilities contributing to postzygotic isolation
increase at least to the square of the divergence time (Turelli et al., 2001). This is because each new
mutation increases the number of possible combinations resulting in incompatibilities (Coyne & Orr,
2004). The Dobzhansky-Muller model for the evolution of hybrid incompatibilities describes
incompatibilities between diverging regions of the nuclear genome, but it is also possible that
interactions between cytoplasmic (from the chloroplast or the mitochondria) and nuclear genes
produce similar hybrid incompatibilities (Burke & Arnold, 2001). Hybrid sterility and inviability
evolving as by-products of divergence of genomes in allopatry have been confirmed mostly in
Drosophila and in plants (reviewed in Coyne & Orr, 2004), but also in hybrids between the platyfish
and the swordtail (Wittbrodt et al., 1989) and between Nasonia wasp species (Gadau et al., 1999).
- 17 -
3.2
Haldane’s rule
Haldane’s rule describes a pattern observed among hybrid offspring of many taxa regarding hybrid
incompatibilities: “When in the F1 offspring of two different animal races one sex is absent, rare, or
sterile, that sex is the heterozygous sex” (Haldane, 1922). What Haldane described as “heterozygous”
is now commonly termed “the heterogametic sex”, i.e. the sex having one single copy of each sex
chromosome. In mammals and many insects, males are heterogametic (XY) and females are
homogametic (XX), while in birds and Lepidoptera, the opposite pattern is the rule (heterogametic
females (WZ) and homogametic males (ZZ)). Haldane’s rule is obeyed is all taxa surveyed so far, for
both inviability and sterility (Wu & Davis, 1993). Independently of which sex is the heterogametic
one, it is the first one to be affected by incompatibilities (Coyne & Orr, 2004). In birds, female sterility
evolves first, but male sterility generally appears before female inviability (Price & Bouvier, 2002), as
opposed to Drosophila where the heterogametic sex (males) loses both fertility and viability long
before the homogametic sex (females) is affected (Coyne & Orr, 1997). In Lepidoptera, Haldane’s rule
is obeyed equally for sterility and inviability (the female is affected first in both cases) (Presgraves,
2002). Hybrid sterility evolves faster than inviability and in 38% of the crosses observed by
Presgraves, complete female inviability evolves before male sterility in hybrids.
One of the most widely accepted explanations for Haldane’s rule is the dominance theory (Muller &
Pontecarvo, 1942). If genetic incompatibilities exist between sex chromosomes and autosomes, the
heterogametic sex will suffer from recessive and dominant incompatibilities, while the homogametic
sex will only be affected by dominant incompatibilities, having a second copy of the sex chromosome
to compensate for recessive incompatibilities (Muller & Pontecarvo, 1942). Another possible cause for
the observed phenomenon (in species with heterogametic males) is the faster X theory, based on the
idea that genes on the X chromosome evolve faster than autosomes, as they are exposed to selection in
the heterogametic sex (Charlesworth et al., 1987). Moreover, the faster male theory argues that
stronger sexual selection on males might cause faster evolution of male expressed genes, and that
spermatogenesis might be more sensible than female gamete production (Wu & Davis, 1993, see next
section for more details on spermatogenesis and hybrid male sterility). The faster X and the faster male
theories only seem to be valid when describing hybrid sterility, and not hybrid inviability, and do not
explain Haldane’s rule in taxa where the female is the heterogametic sex (Coyne & Orr, 2004). Finally,
meiotic drive, or the distortion of segregation away from Mendelian ratios, has been proposed to
explain hybrid male sterility (Frank, 1991). Alleles containing a drive will be transmitted to more than
50% of the offspring and will distort sex ratio if they are found on a sex chromosome (elimination of
all sperm carrying a Y chromosome if the drive is found on the X chromosome for example). This will
lead to a rapid coevolution of drivers and suppressors, increasing the rate of divergence between
species. Hybrid offspring might end up with a driver but lose the corresponding suppressor through
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recombination (McDermott & Noor, 2010). This would lead to hybrid male sterility if a male carries
different drivers present on different sex chromosomes, which would lead to the destruction of both
sperm carrying X and Y chromosomes (Coyne & Orr, 2004).
All in all, none of the theories mentioned above seem to explain both the observed rates of hybrid
inviability versus sterility among crosses and the evolution of hybrid incompatibilities in both taxa
with heterogametic males and taxa with heterogametic females (Wu & Davis, 1993). It therefore
seems likely that Hadane’s rule is a composite phenomenon, with several different mechanisms
affecting the evolution of hybrid inviability and hybrid sterility (Wu & Davis, 1993). Hybrid
inviability is probably easier to explain because lethal mutations are likely to affect both sexes in the
same way, and Muller’s dominance theory (i.e. recessive interactions between autosomes and
hemizygous sex chromosomes) seem to be enough to explain Haldane’s rule for inviability among all
taxa (Wu & Davis, 1993). The faster-male hypothesis coupled to the dominance theory could explain
the higher occurrence of hybrid sterility versus hybrid inviability, particularly in taxa with
heterogametic males (Coyne & Orr, 2004). Different sets of genes affect male and female sterility and
incompatibilities are likely to accumulate at different rates in the different sexes (Turelli et al., 2001).
This could explain why the homogametic sex suffers from sterility before the evolution of inviability
of the heterogametic sex in birds for example, or the observation of male sterility first in species where
males have two full sets of genes on their sex chromosomes (e.g. Aedes mosquitoes having a fully
functional Y chromosome, Presgraves & Orr, 1998) or where both sexes are functionally hemizygous
(e.g. marsupials, Watson & Demuth, 2012). In the following section, I will explore the fast evolution
of hybrid male sterility in more details.
3.3
Spermatogenesis and the genetics of hybrid male sterility
Spermatogenesis seems to be one of the first mechanisms to be affected by hybrid incompatibilities in
species where the heterogametic sex is the male (Haldane, 1922). Low sperm numbers have been
observed in hybrid males of the parasitic wasp Nasonia (Clark et al., 2010) and between two species
of Lepomis sunfish (Immler et al., 2011), whereas a loss of sperm motility and other alteration of
sperm functions were described in hybrids Drosophila mojavensis x D. arizonae (Reed & Markow,
2004). A failure of hybrid males to produce mature sperm has been repeatedly observed in Drosophila
(Wu & Davis, 1993; Clancy et al., 2011), sticklebacks (Takahashi et al., 2005), mice (Britton-davidian
et al., 2005; Jeffrey M Good et al., 2008; Oka et al., 2010) and other rodents (Borodin et al., 2006).
The mechanisms leading to the failure of producing mature and functional sperm were already
analysed in detail by Dobzhansky in hybrids between two “races” (as he called them) of Drosophila
pseudoobscura (Dobzhansky, 1934). In a comparison of the spermatogenesis of pure and hybrid
- 19 -
individuals, Dobzhansky described abnormalities in the first meiotic division and the absence of the
second meiotic division, resulting in the production of “large worm-like spermatids”. In birds, where
males are the homogametic sex, the absence of meiotic division and the consequent lack of gamete
production was found to be the cause of sterility in hybrids between the domestic fowl and the
common pheasant (Yamashina, 1943) and in hybrid ducks (Crew & Koller, 1936; Rigdon & Mott,
1965), where the presence of abnormal spermatids was also observed.
The general pattern seems to be that spermatogenesis stops at early stages of the process (typically
before or during the first meiosis) in sterile males, leading to the failure of producing mature sperm
(Dobzhansky, 1934; Yamashina, 1943; Britton-davidian et al., 2005; Borodin et al., 2006; Good et al.,
2008; Oka et al., 2010; Clancy et al., 2011). Some studies have found multiple genes on the X
chromosome to be at the origin of spermatogenesis disruption in Drosophila (e.g. Tao et al., 2003) and
mice (e.g. Dzur-Gejdosova et al., 2012), while others report an effect of mitochondrial elements, the
cytochrome C in particular (Clancy et al., 2011). A recent study on Drosophila identified segregation
distorters on the X-chromosome to be the origin of hybrid sterility (only sperm carrying Xchromosomes are viable in those hybrids that are partially fertile) (Phadnis & Orr, 2009). Several
authors also mention an asymmetry in the sterility of hybrid males depending on the direction of the
cross at early stages of speciation (Reed & Markow, 2004; Kopp & Frank, 2005; Britton-davidian et
al., 2005; Good et al., 2008; Moehring, 2011), suggesting that incompatibilities might depend on the
origin of the X-chromosome or the maternally inherited element and can be caused by a relatively
small number of loci.
Spermatogenesis seems to be particularly sensitive to any perturbation in gene expression. Sun and
colleagues demonstrated that genes that only have minor influence on the fertility of pure-species
Drosophila males can have detrimental consequences for hybrid fertility (Sun et al., 2004). Similarly,
in a comparison between pure species and hybrid Drosophila males, Michalak and Noor observed
different patterns of gene expressions between them, particularly for genes affecting spermatogenesis
(Michalak & Noor, 2003). This may be due to the fact that spermatogenesis lacks any postmeiotic
transcription regulation and therefore is very sensible to any slight misregulation (Wu & Davis, 1993;
Sun et al., 2004). Wu and Davis noted that a delicate balance between X-linked and autosomal genes
is required for spermatogenesis, since the X-chromosome follows a particular pattern of condensation
during this process, and perturbations in the timing or level of gene expression can cause hybrid
sterility (Wu & Davis, 1993). If changes in a few, minor genes is enough to cause strong hybrid
sterility, it is not surprising to find numerous genes involved in hybrid sterility among species that
diverged long ago and are likely to have accumulated many of those incompatibilities (Wu et al.,
1996; Sun et al., 2004). As an example, 120 genes were found to be involved in hybrid sterility
between Drosophila simulans and mauritania, which is many more than genes for hybrid female
- 20 -
sterility or hybrid inviability (Wu et al., 1996). The reasons behind this very fast evolution of male
infertility in hybrid offspring is likely to involve the exact same mechanisms as discussed in the
previous part of this essay: namely selection for fertilization efficiency and rapid evolution of
reproductive features (Wu et al., 1996). If this is the case and if strong selection on spermatogenesis to
improve fertilization success results in hybrid incompatibilities, we would expect to find more genes
involved in hybrid male sterility in males than in females in taxa with homogametic males as well (i.e.
birds and Lepidoptera) (Wu et al., 1996), despite the fact that females might be affected first due to
their unbalanced sex chromosomes. Unfortunately, no study has confirmed this prediction among
neither birds nor Lepidoptera so far. In the next section I will describe how the Ficedula flycatchers, a
model system for the study of speciation, might help us to further understand the patterns of gametic
isolation and hybrid sterility in the wild and in birds in particular.
4.
A model system for the study of pre- and postzygotic
isolation: the Ficedula flycatchers
The collared (Ficedula albicollis) and the pied flycatcher (F. hypoleuca) are two closely related
passerine species that diverged less than two million years ago and probably went through repeated
events of allopatry and secondary contact during the cycles of the last glaciation events in Europe
(Saetre et al., 2001; Ellegren et al., 2012). The two species have very similar phenotypes and ecology.
The males are black and white, the collared flycatchers have a white collar on their neck and a more
pronounced white patch on their forehead (see Figure 4) and are also slightly bigger than the pied
flycatcher males, while females of both species are grey-brownish (Vallin et al., 2011). Both species
have co-occurred during the reproductive season in a large hybrid zone in central Europe for 12’000
years and more recently on the Baltic islands of Gotland (for 150 years) and Öland (since 1960), in
Sweden (Qvarnström et al., 2009). The pied and the collared flycatchers have similar diets and habitat
preferences and males of the two species are thus competing over territories at the start of the breeding
season (Vallin et al., 2011). Females prefer mates of their own species, but hybridization still occurs
and 4% of the breeding pairs on the island of Öland are mixed pairs between collared and pied
flycatchers (Qvarnström et al., 2010; Saetre & Saether, 2010). Hybridization is not without
consequences, as reflected by the strong postzygotic isolation acting against hybrid offspring. All
hybrid females are sterile, following Haldane’s rule (females are the heterogametic sex in birds, see
section 3.2), and lay eggs that never hatch. There is strong sexual selection on hybrid males due to
their intermediate plumage (see Figure 4) and song and they achieve only 47% of the reproductive
success of pure-species males due to their difficulty of finding mates and elevated levels of nestlings
issued from extra-pair copulations in their nests (Svedin et al., 2008). Backcross individuals are
- 21 -
extremely rare and consequently, the fitness of heterospecific pairs calculated over three generations
drops to less than 3% of that of conspecific pairs (Wiley et al., 2009).
Figure 4: Left: collared flycatcher male, middle: pied flycatcher male, right: hybrid flycatcher male.
Several pre-mating barriers help preventing hybridization and facilitate the coexistence of the two
species in sympatry. As mentioned before, the males differ in their plumage coloration, but also in
their song and females choose their mate accordingly, with a strong preference for conspecific males
(Veen et al., 2001). Pied and collared flycatchers have different wintering grounds in Africa and take
different migration routes to their breeding areas in the spring (Veen et al., 2007). Both species feed
their nestlings caterpillar and thus prefer deciduous forest as breeding sites, but pied flycatchers are
more tolerant to harsh conditions and can breed in pine forests, where the food is less abundant
(Qvarnström et al., 2009). Males of the two species also differ in their level of aggressiveness:
collared flycatchers are more dominant and better at defending territories, resulting in the pied
flycatchers being pushed towards fewer, lower quality territories at the Northern tip of the island
(Vallin et al., 2012). There is also divergence in breeding time, possibly as a consequence of the
differences in habitats and wintering grounds, and pied flycatchers have a tendency to breed later than
the collareds (Alatalo et al., 1990). On the other hand, the strong competition between the two
flycatcher species enhances the risk of hybridization, since pied flycatcher males have difficulties
establishing territories and pied females might be forced to hybridize as a result of the low density of
conspecific males (Vallin et al., 2012). Furthermore, some pied flycatchers raised as nestlings in the
neighborhood of collared flycatchers include part of the heterospecific song in theirs, which can cause
females to fail in recognizing conspecific mates. Indeed, pied flycatcher singing a so-called “mixedsong” have a 30% probability of hybridizing (Haavie et al., 2004; Qvarnström et al., 2006).
More recently, studies on the genetics of speciation in the Ficedula flycatchers have demonstrated that
both genes coding for characters used in species recognition (e.g. male plumage coloration) and for
female preference are situated on the Z chromosome, of which females inherit one copy only from
their father (Saether et al., 2007). The recent sequencing and comparison of the flycatcher genomes
- 22 -
confirms a high divergence of the Z chromosome compared to autosomes, since 35% of all fixed
differences between both genomes are found on this chromosome (Ellegren et al., 2012). Meiotic
drive was proposed as a major mechanism behind the divergence of the two species, which could have
strong consequences on the genetics of hybrid incompatibilities.
The mixed population of the hybrid zone on the island of Öland has been monitored in nest-box areas
during 1981-1985 and since 2002 and a large amount of data on pre- and post-zygotic ecological
processes as well as molecular data have been accumulated. However, the mechanisms involving
gametes production and divergence, both at the postmating prezygotic and postzygotic levels, have
never been investigated so far. The production of hybrid offspring is very costly and premating
reproductive barriers are obviously not completely efficient. Mechanisms allowing females to avoid
maladaptive heterospecific fertilization at the gametic level could potentially be very advantageous.
Furthermore, the strongly reduced fitness of hybrid males and the low level of introgression observed
between species hints towards reduced fertility of hybrid males, which could potentially be observed
by analyzing their gametes. As mentioned earlier, the collared and pied flycatcher males differ in
levels of aggressiveness and might therefore adopt different mating strategies. Does this result in
different levels of extra-pair copulations between species? Does it have an influence on the level of
sperm competition between heterospecific males and a consequent divergence in sperm traits between
species? Do we observe conspecific sperm precedence when females mate with both conspecifics and
heterospecifics? Is there divergence in gamete recognition proteins between both species? Do hybrids
have intermediate sperm traits or reproductive strategies that could explain their reduced fitness?
These are few of the numerous questions that could potentially be answered by investigating the
gametes of the collared and the pied flycatchers in the mixed population of Öland. Preliminary results
suggest that there is no major divergence in sperm morphology between species (Podevin, 2011, see
Figure 5). However, the levels of extra-pair copulations differ, since 17.2% of collared offspring are
issued from extra-pair copulations while there are 22.4% of extra-pair offspring in the nests of pied
flycatchers (Podevin, 2011). Despite any obvious divergence in sperm morphology between species,
many other mechanisms could contribute to a differential fertilization success of sperm of the two
species (conspecific sperm precedence), including differences in the ability of sperm to swim in the
reproductive tract of females of the two species and divergences in reproduction proteins influencing
sperm transport, storage and/or fertilization success. The analysis of ejaculates of hybrid males
collected over the three last years suggest that they are strongly affected by genetic incompatibilities
resulting in the absence of sperm production in most hybrid males and the production of immature
spermatids in some of them (Figure 5), a sign of spermatogenesis dysfunction (Ålund et al., under
review). Moreover, extreme rates of egg hatching failures and extra-pair paternities were observed in
the nests of hybrid males, and no hybrid male breeding in the population between 2010 and 2012 sired
- 23 -
any offspring. Hatching failures could be the result of absence of fertilization as well as early
embryonic death due to developmental incompatibilities (Birkhead et al., 2008). The extreme rate of
extra-pair paternity (100%) indicates that if any sperm was produced by those males, it was either nonfunctional or performed poorly compared to the sperm of pure-species males. The advance of
molecular tools and the recent publication of the flycatcher genome (Ellegren et al., 2012) might allow
us to get more insights into the genetic basis of hybrid male incompatibilities between the collared and
the pied flycatchers.
Figure 5: Left: pied flycatcher sperm, middle: collared flycatcher sperm, right: immature spermatids from a hybrid male.
Conclusions
In this essay, I gave an overview of the role of gametes as reproductive isolation barriers between
heterospecific individuals, both before and after fertilization. We have seen that gametes and
reproductive proteins evolve extremely quickly and their divergence constitutes a potentially strong
prezygotic isolation barrier, particularly in organisms with external fertilization. Furthermore,
competition between sperm of different species and mechanisms of cryptic female choice allow sperm
from conspecifics or “preferred” males to be used first for fertilization. Sexual conflict resulting from
the different interests of sexes over fertilization might play a strong role in accelerating the evolution
of reproductive barriers, through a continuous arm race between adaptations in females and males.
When hybridization nevertheless occurs, the next level of reproductive barrier between species is
incompatibilities in the hybrid offspring. We have seen that hybrid incompatibilities can arise as a byproduct of genetic divergences in allopatry, and alleles having a positive effect within species can
cause dysfunctions when found together in the hybrid genomic background. Hybrid sterility caused by
spermatogenesis dysfunctions is pretty common in species with heterogametic males and genes
causing hybrid sterility are more numerous than genes for hybrid inviability.
- 24 -
In general, postmating prezygotic isolation and intrinsic postzygotic incompatibilities are less wellstudied than extrinsic ecological factors and are almost entirely restricted to model or laboratory
organisms. Furthermore, the best studied cases of gamete isolation are species with external
fertilization while hybrid incompatibilities are studied in taxa with heterogametic males. The Ficedula
flycatchers have been studied since decades and still not much is known about gamete interactions
between the two hybridizing species and gamete production in the hybrids. They potentially constitute
a good model for the study of gamete isolation in the wild in internal fertilizers and for hybrid
incompatibilities in homogametic males. The comparison of gametes, reproductive proteins and
fertilization patterns between pied, collared and hybrid flycatchers might help us understanding some
of the key-questions in the study of speciation.
Acknowledgements
I would like to thank my supervisor Anna Qvarnström for her useful comments and Johan Ålund for
his precious contribution to the graphical parts of this essay.
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The evolution of life history traits.
Anders Berglund. 1979
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Predation in marine littoral communities.
Carin Magnhagen. 1979
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Insular biogeographic theory, developments in the past decade.
Stefan Åhs. 1979
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Theories and tests of optimal diets.
Juan Moreno. 1979
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Ecological characters in insular communities.
Mats Malmquist. 1980
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Plant/insect relationships: a synopsis.
Ola Jennersten. 1980
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Sexual size dimorphism in birds of prey.
Per Widén. 1980
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The evolution of avian mating systems.
Per Angelstam. 1980
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Dispersal, dispersion and distribution in small rodent populations.
Fredric Karlsson. 1980
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Age-related differences of competence in birds.
Karin Ståhlbrandt. 1980
13
Respiration measurements as a tool in fish ecology.
Bengt Fladvad. 1981
14
Intraspecific variation; its adaptive value and consequences.
Lars Gustafsson. 1981
15
Cooperative breeding in birds: theories and facts.
Mats Björklund. 1982
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Some aspects of parasitism.
Göran Sundmark. 1982
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Evolution and ecology of migration and dispersal.
Jan Bengtsson. 1982
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Are animals lazy? - Time budgets in ecology.
Bodil Enoksson. 1983
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Ecology of island colonization.
Torbjörn Ebenhard. 1983
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Reproductive habits and parental care in some lacustrine African Cichlids (Pisces)
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Ulrich Jessen. 1983
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Monogamy or polygyny? - Mating systems in passerine birds.
Björn Westman. 1983
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Distributional patterns in reptiles and amphibians.
Per Sjögren. 1983
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Some theoretical considerations of dispersal.
Gunnar Nilsson. 1983
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Interactions between animals and seeds.
Urban Wästljung. 1984
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Reproductive modes and parental care in fishes.
Ingrid Svensson. 1984
26
Reproductive strategies in fishes.
Peter Karås. 1984
27
Sexual selection and female choice based on male genetic qualities.
Jacob Höglund. 1984
28
The timing of breeding in birds with special reference to the proximate control.
Mats Lindén. 1985
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Philopatry and inbreeding versus dispersal and outbreeding.
Tomas Pärt. 1985
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Evolution of sociality in Hymenoptera.
Folke Larsson. 1985
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Bird song and sexual selection.
Dag Eriksson. 1986
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The energetics of avian breeding.
Lars Hillström. 1986
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Sex role reversal in animals.
Gunilla Rosenqvist. 1986
34
Factors affecting post-fledging survival in birds.
Kjell Larsson. 1986
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Quantitative genetics in evolutionary ecology: methods and applications.
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Body size variations in small mammals.
Anders Forsman. 1986
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Habitat heterogeneity and landscape fragmentation - consequences for the
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The evolution of predispersal seed predation systems in insects.
Mats W. Pettersson. 1987
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Animal aggregations.
Anette Baur. 1988
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Operational and other sex ratios: consequences for reproductive behaviour,
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Jan Sundberg. 1988
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Genetic variation in natural populations.
Annika Robertson. 1989
42
Adaptations for a parasitic way of life.
Reija Dufva. 1989
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Predator-prey coevolution.
Lars Erik Lindell. 1990
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Parasite impact on host biology.
Klas Allander. 1990
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Ecological factors determining the geographical distribution of animal
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Berit Martinsson. 1990
46
Mate choice - mechanisms and decision rules.
Elisabet Forsgren. 1990
47
Sexual selection and mate choice with reference to lekking behaviour.
Fredrik Widemo. 1991
48
Nest predation and nest type in passerine birds.
Karin Olsson. 1991
49
Sexual selection, costs of reproduction and operational sex ratio.
Charlotta Kvarnemo. 1991
50
Evolutionary constraints.
Juha Merilä. 1991
51
Variability in the mating systems of parasitic birds.
Phoebe Barnard. 1993. ISRN UU-ZEK-IRE--51--SE
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Foraging theory: static and dynamic optimization.
Mats Eriksson. 1993. ISRN UU-ZEK-IRE--52--SE
53
Moult patterns in passerine bird species.
Christer Larsson. 1993. ISRN UU-ZEK-IRE--53--SE
54
Intraspecific variation in mammalian mating systems.
Cheryl Jones. 1993. ISRN UU-ZEK-IRE--54--SE
55
The Evolution of Colour Patterns in Birds.
Anna Qvarnström. 1993. ISRN UU-ZEK-IRE--55--SE
56
Habitat fragmentation and connectivity.
Torbjörn Nilsson. 1993. ISRN UU-ZEK-IRE--56--SE
57
Sexual Selection. Models, Constraints and Measures.
David Stenström. 1994. ISRN UU-ZEK-IRE--57--SE
58
Cultural Transmission and the Formation of Traditions in Animals.
Henk van der Jeugd. 1995. ISRN UU-ZEK-IRE--58—SE
59
Ecological Factors Affecting the Maintenance of Colour Polymorphism.
Sami Merilaita. 1995. ISRN UU-ZEK-IRE--59--SE
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The Evolution and Maintenance of Hermaphroditism in Animals.
Anna Karlsson. 1996. ISRN UU-ZEK-IRE--60--SE
61
Costs and Benefits of Coloniality in Birds.
Tomas Johansson. 1996. ISRN UU-ZEK-IRE--61--SE
62
Nutrient reserve dynamics in birds.
Måns S. Andersson. 1996. ISRN UU-ZEK-IRE--62--SE
63
Nest-building for parental care, mate choice and protection in fishes.
Sara Östlund. 1996. ISRN UU-ZEK-IRE--63--SE
64
Immunological ecology.
Dag Nordling. 1996. ISRN UU-ZEK-IRE--64--SE
65
Hybrid zones and speciation by reinforcement.
Anders Ödeen. 1996. ISRN UU-ZEK-IRE--65--SE
66
The timing of breeding onset in temperate zone birds.
Robert Przybylo. 1996. ISRN UU-ZEK-IRE--66--SE
67
Founder speciation - a reasonable scenario or a highly unlikely event?
Ann-Britt Florin. 1997. ISRN UU-ZEK-IRE--67--SE
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Effects of the Social System on Genetic Structure in Mammal Populations.
Göran Spong. 1997. ISRN UU-ZEK-IRE--68--SE
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Male Emancipation and Mating Systems in Birds.
Lisa Shorey. 1998. ISRN UU-ZEK-IRE--69--SE
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Social behaviours as constraints in mate choice.
Maria Sandvik. 1998. ISRN UU-ZEK-IRE--70--SE
71
Social monogamy in birds.
Kalev Rattiste. 1999. ISRN UU-ZEK-IRE--71--SE
72
Ecological determinants of effective population size.
Eevi Karvonen. 2000. ISRN UU-ZEK-IRE--72--SE
73
Ecology of animals in ephemeral habitats.
Jonas Victorsson. 2001. ISRN UU-ZEK-IRE--73--SE
74
Extra-pair paternity in monogamous birds.
Katherine Thuman. 2001. ISRN UU-ZEK-IRE--74-SE
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Intersexual communication in fish.
Niclas Kolm. 2001. ISRN UU-ZEK-IRE--75--SE
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The role of postmating prezygotic reproductive isolation in speciation.
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Host parasite interactions: Local adaptation of parasites and changes in host
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Lena Sivars. 2002. ISRN UU-ZEK-IRE--77—SE
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The molecular clock hypothesis. Reliable story or fairy tale?
Marta Vila-Taboada. 2002. ISRN UU-ZEK-IRE--78--SE
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Chemical cues in mate choice.
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Does sexual selection promote speciation? A comparative analysis of the
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Sarah Robinson. 2002. ISRN UU-ZEK-IRE--80--SE
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Population bottlenecks and their effects on genetic diversity: introducing some recent
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Marnie H Demandt. 2003. ISRN UU-ZEK-IRE--1--E
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Immune response and its correlations to other traits in poultry.
Joanna Sendecka. 2003. ISRN UU-ZEK-IRE--82--SE
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Resistance of Hybrids to Parasites.
Chris Wiley. 2003. ISRN UU-ZEK-IRE--83--SE
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The Role of Genetics in Population Viability Analysis.
Johanna Arrendal. 2003. ISRN UU-ZEK-IRE--84--SE
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Sexual Selection and Speciation – The Scent of Compatibility.
Nina Svedin. 2003. ISRN UU-ZEK-IRE--85--SE
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Male-female coevolution: patterns and process.
Johanna Rönn. 2004. ISRN UU-ZEK-IRE--86—SE
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Inbreeding in wild populations and environmental interactions.
Mårten Hjernquist. 2004. ISRN UU-ZEK-IRE--87—SE
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Sympatric speciation – A topic of no consensus.
Emma Rova. 2005. ISRN UU-ZEK-IRE--88--SE
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Man-made offshore installations: Are marine colonisers a problem or an advantage?
Olivia Langhamer. 2005. ISRN UU-ZEK-IRE--89--SE
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“Isolation by distance”: – A biological fact or just a theoretical model?
Sara Bergek. 2006. ISRN UU-ZEK-IRE--90--SE
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Signal evolution via sexual selection – with special reference to Sabethine
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Sandra South. 2007. ISRN UU-ZEK-IRE--91--SE
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Aposematic Colouration and Speciation – An Anuran Perspective.
Andreas Rudh. 2008. ISRN UU-ZEK-IRE--92--SE
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Models of evolutionary change.
Lára R. Hallson. 2008. ISRN UU-ZEK-IRE--93--SE
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Ecological and evolutionary implications of hybridization.
Nicals Vallin. 2009. ISRN UU-ZEK-IRE--94--SE
95
Intralocus sexual conflict for beginners.
Paolo Innocenti. 2009. ISRN UU-ZEK-IRE--95--SE
96
The evolution of animal mating systems.
Josefin Sundin. 2009. ISRN UU-ZEK-IRE--96--SE
97
The evolutionary ecology of host-parasite interactions.
Katarzyna Kulma. 2011. ISRN UU-ZEK-IRE--97--SE
98
Sensory exploitation – trends, mechanisms and implications.
Mirjam Amcoff. 2011. ISRN UU-ZEK-IRE--98--SE
99
Evolutionary biology of aging.
Hwei-yen Chen 2012. ISRN UU-ZEK-IRE--99--SE