Download Speciation - What Can be Learned from a Flycatcher Hybrid Zone?

Digital Comprehensive Summaries of Uppsala Dissertations
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Speciation - What Can be Learned
from a Flycatcher Hybrid Zone?
CHRIS WILEY
ACTA
UNIVERSITATIS
UPSALIENSIS
UPPSALA
2006
ISSN 1651-6214
ISBN 91-554-6737-7
urn:nbn:se:uu:diva-7358
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List of papers
This thesis is based on the following five papers, which will be referred to in
the text by their Roman numerals.
I
Wiley, C., Fogelberg, N., Sæther, S. A., Veen, T., Svedin, N.,
Vogel Kehlenbeck, J. & Qvarnström, A. Direct benefits for
hybridizing Ficedula flycatchers (submitted).
II
Wiley, C., Qvarnström, A. & Gustafsson, L. Effects of hybridization on the immunity of flycatchers and their infection
by blood parasites (submitted).
III
Wiley, C., Andersson, G., Qvarnström, A., Borge, T. &
Sætre, G.-P. Multi-generation assessments of selection
against hybridization in a wild bird (Manuscript).
IV
Wiley, C., Svedin, N., Weissing, F. J. & Qvarnström, A.
Asymmetrical hybridization: simulations and tests in a natural
hybrid zone (submitted).
V
Wiley, C., Bengtson, J. M., Svedin, N. & Qvarnström, A.
(2005) Hybridization cost of delayed maturation of secondary
sexual traits in the collared flycatcher. Evolution 59, 27112716.
Paper V was reproduced with permission from the publisher, The Society for
the Study of Evolution.
For each of the chapters included in this thesis, I was heavily involved in the
planning of experiments and the collection of field data, performed much
(III) or all (II) of the molecular work, performed all of the statistical analyses (except the assignment of genotypes in III), and prepared all manuscripts. In all papers, co-authors contributed intellectually to the planning of
experiments and final stages of manuscript preparation, as well as with field
data and laboratory work (III only).
Contents
Introduction......................................................................................................... 7
The Ficedula hybrid zone .................................................................................. 9
How maladaptive is hybridization?................................................................. 13
Are there direct benefits of hybridization? ................................................ 14
Are hybrids more or less resistant to parasites?......................................... 16
Summarizing selection against hybridization ............................................ 17
Why does hybridization occur? ....................................................................... 20
A lack of conspecific mates ........................................................................ 20
Errors in species recognition....................................................................... 23
Conclusions and future directions ................................................................... 25
Summary in Swedish (Sammanfattning) ........................................................ 27
Acknowledgements .......................................................................................... 32
References......................................................................................................... 34
Abbreviations
MHC
PHA
SNP
F1
Major histocompatibility complex
Phytohaemagglutinin
Single nucleotide polymorphism
First-generation hybrid
Introduction
The enormous diversity of life on Earth faces us with a number of important
and challenging questions. How have all these species arisen? What barriers
prevent incipient species from interbreeding with each other? And why
aren’t new species out-competed by the old? Many of these questions are not
merely philosophical, but also dictate how we should manage this biodiversity in a way that preserves the variation as well its potential for selfperpetuation. However, such fundamental questions are by no means easily
answered, owing largely to the timescales over which the process of speciation typically occurs. For these reasons, despite the subject of speciation
having attracted considerable attention from the scientific world, our understanding of the process remains fragmentary (Coyne & Orr, 2004).
Natural hybrid zones, geographical regions where closely related species
meet and occasionally interbreed, provide important insights into the evolutionary processes characterizing the final stages of speciation. This is because, by focusing on young pairs of sister-species, we can better understand
the types of reproductive barriers that are important for speciation to occur.
For example, we can examine the relative strengths of post- and pre-zygotic
isolation between populations that maintain their integrity in sympatry versus those that fuse. We can also investigate how post-zygotic isolation (i.e.
unfit hybrids) influences the evolution of pre-zygotic isolation (species recognition). Overall, hybrid zones provide us with a valuable tool for investigating the conditions favouring the evolution of new species, and allow us to
test predictions from theoretical models of speciation in nature.
Although interspecific hybridization is a widespread phenomenon (e.g.
Johnson, 1939; Grant & Grant, 1992; Dowling & DeMarais, 1993), by definition it is infrequent. Populations are defined as different species when gene
flow between them is sufficiently small that each remains distinct (modified
version of Mayr’s [1995] biological species concept, as advocated by Coyne
& Orr [2004]). Unless there are severe post-zygotic barriers to reproduction
(e.g. hybrid sterility), gene flow between populations through hybridization
ultimately leads to the breakdown of reproductive barriers and fusion into a
single species (Felsenstein, 1981). On the other hand, in the face of severe
post-zygotic isolation, individuals evolve to mate only with their own species (a process termed reinforcement) (Dobzhansky, 1937; Blair, 1955), thus
reducing the frequency of hybridization. However, it is this transitory nature
7
of many hybrid zones that makes them ideal for studying the conditions under which speciation is prevented or completed.
8
The Ficedula hybrid zone
Pied (Ficedula hypoleuca) and collared flycatchers (F. albicollis) are both
well-studied model organisms, and much is known regarding their breeding
biology and reproductive isolation. Because of limited space within each
chapter of this thesis for summarizing this knowledge, I have attempted to
outline important aspects of their biology in some detail here.
Pied and collared flycatchers are small, short-lived, migratory passerine
birds that breed in mixed and deciduous forests in Europe. They winter in
sub-Saharan Africa, and arrive at the breeding grounds in late April or early
May. Soon after arrival, males aggressively occupy and defend breeding
holes (Alatalo et al. 1994; Qvarnström 1997), and females subsequently inspect a number of males/territories before selecting their partner (Dale and
Slagsvold 1996). Their ready acceptance of nest-boxes makes them ideal
study objects. Females typically lay five to seven eggs, and are the sole incubator (when they are caught and measured). Both sexes assist in the feeding of nestlings (when males are caught and measured), and only a single
brood is produced each year. Monogamy prevails, although at least 9% of
males are polygynous (Qvarnström & Gustafsson, 2006), and approximately
15% of nestlings are sired by extra-pair males (Sheldon & Ellegren, 1999).
The two species probably remained geographically isolated in separate
glacial refuges during the Pleistocene, and have since expanded their breeding ranges northward (Sætre et al., 2001). Today, they co-occur (are sympatric) within a broad hybrid zone that includes much of central and eastern
Europe, with an isolated hybrid zone on the Swedish islands of Gotland and
Öland in the Baltic Sea. The two species have remained morphologically
distinct in the face of hybridization. The work included in this thesis was
carried out within the Swedish hybrid zone, which constitutes a more recent
contact between the two species (around 100 years) (Lundberg and Alatalo,
1992). On the Swedish islands (as in much of Europe), collared flycatchers
have gradually expanded their range, excluding pied flycatchers from optimal breeding habitats (Alatalo et al., 1982, 1994; Sætre et al., 1999; see Fig
1). The Swedish hybrid zone is surrounded on all sides by allopatric pied
flycatchers, which is a likely source of migrants into the hybrid zone. Immigration into the Swedish collared population, when it occurs, most likely
involves individuals arising from other hybrid zones in eastern Europe.
9
Figure 1. (A) Breeding distribution of pied (red), collared (yellow) flycatchers. Hybrid zone is shown in orange (adapted from Haavie et al., 2004). (B) Typical mixed
forest inhabited by pied flycatchers. (C) Typical, high quality, deciduous forest
dominated by collared flycatchers (both B and C taken in northern Öland). (D) Location of nest-box areas monitored within the Swedish flycatcher hybrid zone.
Figure 2. (A) Adult male collared flycatcher, (B) subadult male collared flycatcher,
(C) male hybrid, and (D) male pied flycatcher. Photos kindly provided by N. Svedin and
M. Hjernquist.
The males of both species have bold, black-and-white plumage patterns,
which are involved in intraspecific sexual selection (both male-male compe10
tition and female choice) (Pärt & Qvarnström, 1997; Qvarnstöm, 1999;
Qvarnström et al., 2000). However, they delay maturation of these traits until
they are two years old. Subadults (one year old) are sexually mature, but are
less boldly marked than older birds, either to avoid conflicts with dominant,
older males (Qvarnström, 1997), or through developmental constraints. The
two species differ in the size of their white patches and the blackness of their
pigmented plumage, which not only permits their identification by humans,
but may also be important for females to recognize conspecific mates (Sætre
et al., 1997a). Females of the two species are both dull grey-brown, and differ only in subtle differences in the tone of their upperparts and the amount
of white at the base of their neck feathers (Svensson 1992). Previous matechoice trials have demonstrated that females display clear preferences for
males of their own species (Sætre et al., 1997b). Males, on the other hand,
appear to be less selective towards whom they display (Dale and Slagsvold
1994; Sætre et al., 1997b). Plumage, however, is unlikely to be the only cue
used for mate-recognition, as the two species also have distinct songs. Song
is thought to be an important sexual signal, isolating many species that otherwise do not differ in secondary sexual traits (Martens, 1996). However, the
use of song by female flycatchers for locating conspecific mates is complicated by the fact that in areas of sympatry many male pied flycatchers learn
to sing like collared flycatchers (Gelter, 1987; Alatalo et al., 1990; Haavie et
al., 2004; Qvarnström et al., 2006). Whether this ‘mixed singing’ has
evolved in males to signal territory occupancy to individuals of both species,
or whether it is a maladaptive behaviour persisting from allopatry is currently unclear. However, mixed singing is associated with high rates of hybridization (Qvarnström et al. 2006).
The above-mentioned pre-mating barriers between the two species are
quite strong. Heterospecific pairs typically constitute only 5.1% of the study
population on Öland, and 2.4% on Gotland. When hybridization does occur,
it results in the production of largely unfit F1 offspring. Females are sterile,
and F1 males also have reduced fitness (Alatalo et al. 1990; Gelter et al.
1992; Veen et al., 2001). Postzygotic barriers acting on female hybrids are
mostly intrinsic, although males appear to suffer mostly through their inability to compete for mates (sperm inviability is another likely possibility)
(Svedin, 2006). F1 hybrids are intermediate in morphology between the two
parental species (Sætre et al., 2003). They frequently resemble collared flycatchers, although their white collar is incomplete, they have more grey in
the plumage, and they possess little white in their primaries. However, past
studies into the fitness of hybrids have been fraught with problems arising
from the lack of genetic tools to clearly separate the different hybrid generations (i.e. F1 and backcrosses). Furthermore, some aberrant collared flycatchers (especially subadults) closely resemble hybrids, and genetic tools
are required to distinguish true hybrids. Prior to this thesis, nothing was
11
known about the fitness of later generation hybrids, a situation common to
almost all natural hybrid zones.
Despite the low fitness of hybrids, one previous study suggested that heterospecific pairing is sometimes adaptive (Veen et al., 2001). Female collared flycatchers can reduce the indirect costs of heterospecific pairing by
copulating with extra-pair conspecific males and biasing the sex ratio of their
offspring towards sons (Veen et al., 2001). Furthermore, the fact that such
heterospecific pairs produce more fledged young (perhaps a result of direct
benefits of heterospecific pairing) during the food-limited later half of the
breeding season suggests that heterospecific pairing may be adaptive for
late-arriving female collared flycatchers (Veen et al., 2001).
12
How maladaptive is hybridization?
Estimating the fitness of hybridizing individuals relative to those paired with
conspecific mates not only provides a measure of the strength of selection
against hybridization, but also of the reproductive isolation between sympatric populations. It is therefore important for predicting the amount of gene
flow between two taxa, and is vital for inferring the likelihood that complete
reproductive isolation will ultimately arise through reinforcement. When the
F1 hybrids produced are sterile, hybridizing taxa are fully reproductively
isolated. Because gene flow is prevented, they can be considered separate
species. Furthermore, there is expected to be strong selection on individuals
to avoid interspecific mating (Dobzhansky, 1937), such that eventually the
two species are expected to rarely, if ever, interbreed. However, when the
resultant hybrids are not fully sterile or inviable, a small amount of gene
flow results. Gene flow can effectively disrupt the process of reinforcement
by preventing linkage disequilibrium between genes causing prezygotic isolation and those causing postzygotic isolation, leading to an amalgamation of
the two genomes (Barton & Hewitt, 1981; Felsenstein, 1981). Whether low
levels of gene flow prevent reinforcement has been the topic of much theoretical discussion, and depends greatly on the details of the models employed
to test this (Liou & Price, 1994; Kelly & Noor, 1996; Servedio & Kirkpatrick, 1997; Kirkpatrick & Servedio, 1999; Servedio, 2000). This is perhaps
a question that is best answered by empiricists. Surveying the strengths of
selection against hybrids in hybrid zones where species-boundaries easily
break down versus those in which the two taxa remain distinct and display
character displacement would provide the best tool for investigating the conditions favouring speciation via reinforcement. Unfortunately, there are very
few systems for which we currently have good estimates for the strength of
selection against hybridization (Arnold & Hodges, 1995; Rieseberg & Carney, 1998).
The main reason for this lack of knowledge stems not from a lack of work
into hybrid zones, but from the difficulty of obtaining accurate estimates of
fitness. Fitness is defined as an individual’s contribution to the gene pool of
future generations. In practice, however, it is typically estimated through a
range of indirect measures, without knowing how they reflect the true currency of fitness; the number of gene copies (descendents) in later generations
(Benton & Grant, 2000). Such measures often include seed germination,
13
larval survival, body mass, growth rates, immunocompetence, success at
attracting mates, or fertility. Situations where some fitness components are
high and others are low in the same hybrids highlight the need for a holistic
approach to estimating selection against hybrids.
Ficedula flycatchers highlight the problem with inferring selection from
single fitness components. Initial investigations into the system revealed low
fertility among hybrids (Gelter et al. 1992), which resulted in the conclusion
that hybridization was maladaptive. The fertility of hybrids was 45.9% of
that of pure individuals. However, a later study revealed that one combination of heterospecific pair (collared females pied males) produced more
chicks than conspecific pairs during certain times in the season (Veen et al.,
2001), even though many of these were unfit hybrids. This suggested that
there were either direct benefits to female collared flycatchers of hybridizing, or hybrids possessed enhanced vigour, which was only expressed in
stressful environments. Either explanation represents a fitness component
that acts to counter low fertility, at least to some extent. This raised some
doubt over previous suggestions that hybridization was always maladaptive
in flycatchers.
Are there direct benefits of hybridization?
In Chapter I, I explore the origin of the pattern that late-breeding heterospecific pairs often produce more offspring than conspecific pairs. In
particular, I investigate whether females gain direct benefits from pairing
with males of the other species. These direct benefits might include when
heterospecific males possess superior territories, or utilize their territory in a
way that benefits the nestlings (e.g. by providing more food or complementary food-types). I found that the two species fed similar items to nestlings,
and that the proportion of the parental workload adopted by males was not
different between the two species. Therefore, there were no indications that
heterospecific males utilize their territories in ways that benefit the nestlings
more than conspecific fathers. However, the territories occupied by heterospecific pairs did differ in quality from those occupied by conspecific
pairs. The effects of territory quality and having heterospecific parents were
disentangled by looking at the performance of nests in the same box as heterospecific pairs, but in other years. I found that late-breeding female collared flycatchers that pair with pied flycatchers have better territories than
those that do not hybridize (see Figure 3). This pattern closely matched previously reported fledgling rates in nests reared by such heterospecific pairs
(Veen et al. 2001).
Direct benefits of mate-choice are widely appreciated as an important
component of intraspecific sexual selection (Thornhill, 1976; Searcy, 1979;
Gwynne, 1984; Reynolds & Gross, 1990; Kirkpatrick & Ryan, 1991). De14
spite this, few studies have considered direct benefits to females of hybridization. Most, instead, focus on the indirect costs/benefits of producing hybrid offspring. This trend no doubt reflects the, often severe, nature of indirect costs of hybridization (e.g. sterility of hybrid offspring). Also, direct
benefits can explain the evolution of intraspecific mate choice, while hybridization is normally viewed as maladaptive, and therefore unlikely to
evolve in response to the existence of direct benefits. My findings do not
argue against such logic. What they do suggest is that, despite the existence
of indirect costs, there may be direct benefits to females of hybridization.
Such benefits may be important in reducing selection against hybridization
and thereby hindering the evolution of reproductive isolation. Furthermore,
among hybrid zones there exists a continuum of postzygotic isolation, from
complete sterility to hybrid vigour. In those hybrid zones where postzygotic
isolation is weak, direct benefits of hybridization may dictate selection for
/against hybridization.
Territory quality
(total mass of chicks)
90
47
80
23
63
70
59 64
16
3
60
50
2
40
30
-10
-5
-5
0
0
5
5
1 0
1 5
10
Laying date (0 = yearly mean)
Figure 3. Comparison of data on fledging success of heterospecific pairs with male
pied flycatchers (left panel, curve 1: from Veen et al., 2001), and the quality of their
territories (right panel, triangles: from Chapter I) with those of collared flycatchers
(negative slope in left panel; squares in right panel). Heterospecific pairs tend to
have better territories and produce more fledglings than pairs of collared flycatcher
after two days before the mean laying date of each year.
In Chapter I, I also discuss another important aspect of speciation;
namely, for the coexistence of two sister taxa, they must not only be reproductively isolated, but should diverge sufficiently in niche to prevent competitive exclusion (Gause, 1934; Lack, 1946; Macarthur and Levins, 1964,
1967). Collared and pied flycatchers prefer the same breeding habitat (Alatalo et al., 1994), compete over the same nest-holes, and also have highly
overlapping diets (Chapter I). Previous studies have indicated that pied flycatchers have a broader tolerance of environmental stress (Sætre et al., 1999;
Qvarnström et al., 2005), which allows them to persist in habitats of lowest
15
quality, while being excluded from those of the highest quality (Alatalo et
al., 1994). Nevertheless, the overlap in their realized niches (including spatial segregation resulting from competition) is extensive, and long-term persistence of the two species on the Baltic islands is unlikely without further
divergence in ecology. Undoubtedly, the persistence of pied flycatchers on
the two Swedish study islands is made possible through extensive immigration from surrounding allopatric source populations.
Are hybrids more or less resistant to parasites?
Another component of fitness potentially enhanced by hybridization is resistance against parasites. A high level of heterozygosity across the genome,
and in particular among parts coding for the major histocompatibility complex (MHC) may promote overall immunity (Hughes & Hughes, 1995;
Wegner et al. 2003; Westerdahl et al., 2005). Furthermore, highly hostspecific parasites may be unable to colonise and infect the novel host environment offered by hybrids (Whitham et al., 1994). In Chapter II, I investigate whether hybrid flycatchers are more or less susceptible to infection by
blood parasites (Haemoproteus and Plasmodium) and whether this susceptibility is correlated with their immune-response to a novel antigen, phytohaemagglutinin (PHA). F1 hybrids had parasite infection rates that were intermediate between, but not significantly different from, collared and pied
flycatchers (see Figure 4). Furthermore, their immune response to PHA was
similar to that of the parents. Later generation hybrids, however, had significantly stronger immune responses than the parental species (Figure 4). They
also had the lowest malarial infection rate, although this was not significantly lower than the parental species. More samples are required before it is
known whether this pattern is robust.
16
Figure 4. Rates of infection by avian malarial parasites (left panel) and immune
response to PHA (right panel) of collared and pied flycatchers and their first- (F1)
and later-generation (RECOM) hybrids (see Chapter II).
In general, this study failed to support the idea that parasite resistance is
enhanced in F1 hybrids. This finding agrees with a number of recent studies
indicating that parasite resistance of animal hybrids does not tend to be
higher than that of the parental species (Moulia, 1999; Derothe et al., 2001;
Parris, 2004; Wolinska et al., 2004). This is a comparable pattern to that
observed in plants (Moulia, 1999; Fritz et al., 1999), despite vastly different
immune systems.
Summarizing selection against hybridization
In flycatchers, individual components of fitness are differently affected by
hybridization. Fertility is reduced in F1 hybrids, but their immunity against
malarial parasites is not impaired (Chapter II). Furthermore, growing up in a
territory possessed by heterospecific parents is sometimes associated with
direct benefits (Chapter I). Selection on flycatchers to avoid hybridization is
therefore unclear, and may be lower than previously thought, based on fertility alone. In Chapter III, I attempt to resolve this confusion by producing a
single estimate of the selection against hybridization; a composite of all individual fitness components. As mentioned earlier, estimating the fitness of
hybridizing individuals is a complex problem. Not only is it difficult to find
proxies of fitness that accurately reflect genetic representation in later generations, but it is unclear how many generations in the future we should consider. Because of the frequent indirect costs/benefits of hybridization (i.e.
hybrid offspring with especially unfit or vigorous hybrid offspring), esti17
mates of a hybridizing individual’s fitness as their genetic contribution to the
next generation are unsuitable, as this may not represent their contribution to
‘future’ generations. This is widely recognized, and is why selection against
hybridization is typically inferred from the reproductive output of F1 hybrids,
rather than the reproductive output of hybridizing individuals themselves
(see recent examples by Parris et al. 1999; Bierne et al., 2002; Pertl et al.,
2002; Ramsey et al., 2003; Bleeker & Matthies, 2005; Kirk et al. 2005; Peterson et al., 2005). However, the fitness consequences of hybridization (hybrid vigour or outbreeding depression) may be expressed over several generations (Rieseberg & Carney, 1998; Barton, 2001; Burke & Arnold, 2001).
As a result, the indirect costs or benefits of hybridization may be underestimated when only F1 offspring are considered. Due to the difficulty of estimating fitness in nature, as well as correctly identifying late-generation hybrids, very little is currently known about for how many generations of hybrid offspring the indirect costs of hybridization typically persist. Yet, this
information is important if we are to understand how well single- and twogeneration estimates of fitness (i.e. based on the reproductive success of
hybridizing pairs and their F1 offspring) represent overall selection against
hybridization.
Until recently, the lack of genetic tools for accurately identifying lategeneration hybrid flycatchers has prohibited the estimation of selection
against hybridization using multiple generations of hybrid descendents.
However, recently identified SNPs (Borge et al., 2005) now allow us to identify F1 hybrids, first-generation backcrosses, and second-generation backcrosses. In Chapter III, I use the number of descendents three generations
after hybridization to estimate selection against hybridization. Two independent methods for estimating this (comparing frequencies of the hybrid
descendents and combining reproductive data from all hybrid generations)
revealed similar results. Hybridizing individuals produce approximately 2%
of the number of descendents arising from non-hybridizing individuals. Furthermore, this severe postzygotic isolation between collared and pied flycatchers is likely to be a conservative estimate. This is because I also observed a high rate of ‘re-hybridization’ (mating with the species that constitutes the minority of the individual’s genome) among second-generation
backcrosses. This suggests that the depressed fitness of hybrids persists beyond first-generation backcrosses.
Strong postzygotic isolation between collared and pied flycatchers is not
surprising considering they have remained distinct in the face of extensive
hybridization. Still, this was only detected once multiple generations of hybrid descendents were considered. Chapter III highlights the importance of
taking a holistic approach when estimating fitness. This study suggests a
crucial role for post-zygotic isolation as a reproductive barrier between
newly evolved bird species. This opposes previous suggestions that the evolution of behavioural barriers to interbreeding typically precedes the evolu18
tion of postmating isolation in birds (Grant & Grant, 1997). Continued developments in the genetic tools available for identifying later-generation
hybrids (backcrosses and ‘re-hybrids’) will greatly assist in quantifying
postzygotic isolation in other study-systems, allowing more educated conclusions to be drawn regarding the evolution of reproductive barriers.
19
Why does hybridization occur?
Given that selection against hybridization between flycatcher species is so
strong, it is worth considering why complete pre-mating isolation has not
evolved. Perhaps the two taxa have simply not spent a sufficient amount of
time in sympatry for this to occur. Alternatively, other factors may have
prevented the evolution of complete pre-mating isolation though reinforcement. Whether reinforcement is common has been a topic of much debate
(Bigelow, 1965; Spencer et al., 1986; Butlin, 1987, 1989; Noor, 1999; Coyne
& Orr, 2004). Coyne and Orr (2004) highlight the main objections, and I
summarise these below. First, gene flow causes recombination among genes
coding for assortative mating and genes coding for low hybrid fitness. This
often leads to a breakdown in isolation barriers. Second, if one of the two
taxa is rare, they suffer higher rates of hybridization. When this is costly, as
is required for reinforcement, the rare species is likely to go extinct before
pre-mating isolation has evolved. Third, gene flow from outside the hybrid
zone prevents local adaptation to sympatry (including the evolution of species recognition). Fourth, as reinforcement progresses, hybridization becomes more infrequent, and this relaxed selection slows further evolution of
traits influencing hybridization.
To gain insight into which of these might be important in flycatcher hybrid zones, it is important to examine which factors currently affect hybridization rates. Given that hybridization is costly, there are two main reasons
why an individual might still hybridize. First, it may be poorly able to distinguish conspecific mates from heterospecific mates (i.e. there are errors in
species recognition). Such a scenario arises if reinforcement has not yet occurred. Alternatively, there may be no errors in species recognition, but conspecific mates are simply not available, forcing hybridization. Thus, hybridization may result even if reinforcement has occurred. Chapters IV and V
explore these possibilities in flycatchers.
A lack of conspecific mates
Even if both species display perfect preferences for their own kind, hybridization may still occur when conspecific mates are lacking. If this is the pri20
mary reason for hybridization, we can make a number of predictions. First,
the risk of hybridization experienced by individual females increases as their
species constitutes a smaller proportion of the population. Second, maximum
hybridization rates occur when one species is rare (Randler, 2002). Third,
because females are thought to be the choosy sex, heterospecific pairs should
mostly contain a female of the rare species (Wirtz, 1999). In Chapter IV, I
test each of these predictions in flycatchers. Because the study islands consist of a large number of separate forests with differing ratios of the two species (see Figure 1) and females generally sample potential partners within
the one woodlot (Dale & Slagsvold, 1996), I was able to determine how
hybridization patterns varied with the relative abundance of the two species.
In accordance with expected effects of lacking conspecific mates, female
pied flycatchers had much higher probabilities of hybridizing in sites where
conspecifics were rare. A lack of data on female collared flycatchers in pieddominated sites prevented a similar comparison being done for them, but the
available data was consistent with such a pattern (see Figure 5).
Figure 5. The effect of relative abundance on the hybridization risk of individual
females of either species (see Chapter IV). Each pair of points (collared and pied)
represents one or several forest sites with similar species composition.
However, the overall hybridization rate was highest in sites where the two
species were at equal frequencies (Figure 6a). This pattern resulted from the
fact that, while individual females had lower hybridization risks when they
became more common, there were more of them. Simulations confirmed that
21
such a pattern is consistent with hybrid zones in which there are moderate
levels of error in species recognition. This implies that a lack of conspecific
mates, while important, is not the sole explanation of hybridization in flycatchers.
This conclusion was further supported by asymmetries in hybridization
towards heterospecific pairs involving females of the rare species. Flycatcher
hybrid zones in fact showed the opposite pattern; that heterospecific pairs
more often contained female pied flycatchers as pied flycatchers were increasingly common (figure 6b). Such a pattern implies that rarity can influence hybridization in ways other than through limiting conspecific mates.
Rarity can affect the error rate of choosy females, either through increasing
rates of mixed singing (false male signals) or as a plastic response of females
to their perceived hybridization risk.
Figure 6. The relationships between relative abundance of pied flycatchers within a
forest site and (a) the overall hybridization rate within a site, and (b) the bias in the
direction of hybridization towards heterospecific pairs involving female pied flycatchers (see Chapter IV for details).
Overall, much of the hybridization occurring in flycatchers results from a
lack of conspecific mates. However, even when the choice between a heterospecific and conspecific is available, errors do occur with notable frequency.
22
Errors in species recognition
Species recognition depends upon the existence of traits that characterize
each species, and preferences of individuals for those traits that signal conspecifics. Errors in mate choice (i.e. hybridization) can therefore result if the
traits are not sufficiently divergent, and/or if the preferences for the trait are
weak. Chapters IV and V examine components of the species recognition
process in flycatchers.
In flycatchers, there is a slight but significant bias towards hybridization
mostly involving female pied flycatchers and male collared flycatcher
(Chapter IV). From figure 6b, I conclude that this bias probably doesn’t result from pied flycatchers being the generally rarer species across the field
sites. I therefore tested whether differences between the females of the two
species in their discriminatory ability are a likely reason for this asymmetrical hybridization. In Chapter IV, I present results from a song-broadcast
experiment, which indicated that collared flycatchers respond more strongly
to conspecific song than pied song. Pied flycatchers, on the other hand, do
not display preferences for conspecific song. These differences in discriminatory ability are a likely cause of asymmetrical hybridization. However, the
selection on such preferences to be refined within the hybrid zone is dependent on the reliability of the information contained within the song-signal. A
large proportion of male pied flycatchers within the hybrid zone copy the
songs of male collared flycatchers (Gelter, 1987; Alatalo et al., 1990; Haavie
et al., 2004), increasing their probability of being chosen by female collared
flycatchers (Qvarnström et al., 2006), and reducing the selection on female
pied flycatchers to narrow their preference.
Pied flycatchers within the Swedish hybrid zone have probably lived
alongside collared flycatchers for a shorter time than Swedish collared flycatchers have lived alongside pieds. Furthermore, there is probably substantial immigration into the sympatric population of pied flycatchers from allopatric areas (Alatalo et al., 1982). Both factors make it likely that if one of
the two species were less adapted to sympatry, it is expected to be the pied
flycatcher. Indeed, they show weaker preferences for their own kind, and
more often falsely signal their specific identity through song. Comparisons
with allopatric populations of collared flycatchers are required to confirm
whether their apparent adaptation to sympatry is a result of reinforcement.
Migration from allopatric populations is typically implicated as a factor inhibiting reinforcement, and this may be true for pied flycatchers. However, it
can also enhance the process in collared flycatchers, by prolonging the time
before the local extinction of the competitively least fit taxon.
However, some taxa that have spent a long time in sympatry and experience very little gene flow from allopatry may continue to possess traits that
increase their hybridization rate, especially if lacking those traits is associated with costs. In Chapter V, I show that delaying the maturation of secon23
dary sexual characters until two years of age in collared flycatchers is associated with an increased risk of hybridization among one-year-olds (subadults). This arises because subadults approach pied flycatchers in morphology (see figures 2 and 7), and they are therefore chosen by female pied
flycatchers faced with a lack of conspecific mates. Despite this cost of delayed maturation, it persists within the hybrid zone. The maturation of sexual
ornaments in flycatchers may be constrained developmentally (as appears to
be the case in shorebirds: Chu, 1994), such that it cannot be avoided without
substantial re-organization of the developmental process. Alternatively, the
benefits associated with reduced aggressive confrontation by older, dominant
males (as indicated by Qvarnström, 1997) may override the risk of hybridization in a hybrid zone where heterospecific females are scarce. In either
case, the trait increasing hybridization (delayed maturation) is maintained
because the costs of lacking it are too great. This highlights an important
criticism of speciation via reinforcement; namely, once hybridization becomes rare, further selection on traits that prevent it is weak, and such traits
are unlikely to evolve if there are costs or constraints associated with them.
Figure 7. Principal components analysis of four plumage traits, showing phenotypic
overlap between male pied flycatchers (triangles), subadult male collared flycatchers
(squares) and adult male collared flycatchers (crosses) (see Chapter V).
24
Conclusions and future directions
This study provides one of the first comprehensive estimates of the
strength of selection against natural hybridization. It also highlights that
different components of fitness can be differently affected by hybridization,
such that it is rarely possible to know a priori if any particular component
acts as an appropriate proxy for overall selection. By estimating fitness in a
context that is biologically relevant (i.e. in nature), studies such as presented
in this thesis provide a basis for inferring the range of conditions under
which populations fuse or remain separate in the face of hybridization. This
knowledge is fundamental for understanding and managing the diversity of
life on Earth. With continuing advances in the genetic tools available for
identifying later-generation hybrids and monitoring gene flow, estimating
the strength of selection against hybridization may soon be an achievable
goal in many other hybrid zones.
I found that post-mating barriers play a crucial role in maintaining species
integrity in flycatchers, while pre-mating barriers are incomplete. This contrasts with previous suggestions that pre-mating barriers typically evolve
fastest in birds (Grant & Grant, 1997). Clearly, the relative importance for
speciation of pre- versus post-mating barriers to gene flow remains unclear.
Investigations into the number of genes typically involved in such barriers
may give insight into their likely rate of evolution, and therefore importance.
Strong selection against hybridization need not always result in reinforced
pre-mating isolation. I identify a number of traits (e.g. song discrimination
and copying, delayed maturation of sexual ornaments) influencing hybridization, implying that these traits are under different selection within and outside the hybrid zone. This thesis therefore provides testable predictions about
the expected directions of change in the values of these traits between allopatric and sympatric populations of each species, as predicted from reinforcement. Such comparative studies will provide an indication of the role
that reinforcement has had in causing the pre-mating isolation currently observed within the flycatcher hybrid zone.
Finally, this thesis highlights a generally neglected reason for hybridization; a lack of conspecific mates. Even when individuals express perfect
preferences for their own kind, hybridization may occur when no conspecific
mates are available to choose from. Whether this is a widespread phenomenon is not known, but some studies suggest that it is (Randler, 2002). In hybrid zones where it is more beneficial to hybridize than to not breed at all,
25
hybridization may occur as an adaptive “back-up” strategy. This may have
important repercussions for reinforcement, and yet this is rarely considered.
Overall, the Ficedula hybrid zone provides an ideal opportunity for studying a range of questions pertaining to speciation, and this thesis provides an
important early step towards answering some of these.
26
Summary in Swedish (Sammanfattning)
På vår planet finns miljontals arter, men hur alla dessa arter utvecklats och
hur de håller sig separerade har vi liten kunskap om. Enligt det biologiska
artbegreppet utgör varje art en population av individer som är reproduktivt
isolerade från andra sådana populationer. Bildandet av nya arter kan därför
ses som utvecklandet av reproduktiva barriärer mellan populationer. Sådana
barriärer brukar delas in i två huvudgrupper. Den ena gruppen, prezygotiska
barriärer (t.ex. partnerpreferenser för artspecifika karaktärer), hindrar parningar över artgränserna. Den andra gruppen, postzygotiska barriärer, orsakas
av nedsatt livsduglighet eller fortplantingsförmåga hos hybrider (avkomman
från korsningar mellan olika arter eller populationer). Den relativa betydelsen av dessa barriärer, var för sig eller i samverkan, har diskuterats mycket i
den vetenskapliga litteraturen om artbildningsprocesser. Ett exempel på en
sådan samverkan är att nedsatt fortplantingsförmåga hos hybrider kan leda
till att individer som föredrar att para sig inom den egna arten gynnas. Trots
en livlig teoretisk diskussion om hur och när olika typer av reproduktiva
barriärer utvecklas och samverkar vet vi mycket lite om hur det verkligen
går till i naturen. Svårigheterna med att studera artbilningsprocesser kommer
från den tid dessa processer kräver. Fullständig reproduktiv isolering (d.v.s.
att hybrider inte är livsdugliga) tar igenomsnitt ca 4 miljoner år hos däggdjur
och ännu längre tid hos fåglar. Hur ska man över huvudtaget kunna studera
en så långsam och utdragen process?
Hybridzoner (plaster där två nyligen bildade arter fortfarande delvis korsar sig med varandra) utgör en perfekt situation för att kunna studera de faktorer och processer som är inblandade i uppkomsten av reproduktiv isolering. Den svenska hybridzonen mellan svartvit och halsbandsflugsnappare
utgör ett sådant exempel. Svartvit flugsnappare häckar i hela norden medans
halsbandsflugnappare bara häckar på Öland och Gotland. På dessa öar förekommer korsningar mellan de båda flugsnappararterna som resulterar i honliga hybrider som är sterila och hanliga hybrider som kan fortplanta sig.
Trots att det uppenbarligen förekommer genflöde mellan de båda arterna är
de tydligt morfologiskt och genetiskt skilda från varandra. Mitt avhandlingsarbete undersöker vilka faktorer som är avgörande för att de två flugsnappararterna ska kunna behålla sina identiteter och således vilka faktorer som
är viktiga i artbilningsprocessen genom att studera naturlig hybridisering.
Jag har fokuserat på två huvudfrågeställningar:
27
(1) Hur stark är den evolutionära urvalsprocessen (selektionen) emot hybridisering och hur kan man bäst uppskatta och mäta den?
(2) Varför förkommer hybridisering när det leder till avkommor med nedsatt
fortplantningsförmåga?
ÄR HYBRIDISERING KOSTSAMT?
För individer är det oftast kostsamt, ur ett evolutionärt perspektiv, att para
sig över artgränsen. Hybrider är ofta sterila eller har nedsatt fertilitet, vilket
är fallet hos flugsnappare. Emellertid kan det även finnas vissa fördelar med
att välja en partner som tillhör en annan art och alla livsprocesser behöver
inte fungera sämre hos hybrider. Jag undersöker därför två fall där hybridisering kan vara gynnsamt i de två första kapitlen i min avhandling.
I kapitel I visar jag att hybridisering kan vara gynnsamt för honor om hanar som tillhör en närbesläktad art har tillgång till bättre resurser till
avkomman än hanar av den egna arten. Hanar av den andra arten kanske
bidrar mer till vården av avkomman eller så har de tillgång till bättre häckningsrevir. Sådana direkta fördelar kopplat till val av partner anses vara viktiga när en hona ska välja mellan hanar av den egna arten men de diskuteras
sällan i samband med hybridisering. Jag fann bevis för att
halsbandsflugsnapparhonor faktiskt tjänar på att välja en svartvit flugsnapparhane i slutet av häckningssäsongen, då dessa hanar har tillgång till bättre
revir. Därmed kan jag förklara den underliggande orsaken till att sena
halsbandsflugsnapparhonor producerar fler ungar när de häckar med svartvita flugsnapparhanar. Alltså kan direkta fördelar minska selektionstrycket
mot hybridisering och därmed till en viss del underlätta genflöde mellan
arter.
En annan tänkbar fördel kopplat till hybridisering är immunitet mot parasiter. Hybrider har mer variation i sina gener jämfört med föräldrararterna,
vilket inkluderar gener som styr immunförsvaret. Ökad variation bland dessa
gener kan leda till bättre immunförsvar genom motståndskraft mot fler typer
av parasiter. Dessutom skulle hybrider kunna undslippa många parasiter
genom att helt enkelt utgöra en miljö som den vissa parasiter inte är anpassad till. Många parasiter är nämligen ofta starkt anpassade till specifika värdorganismer. I kapitel II undersöker jag om hybridflugsnappare har relativt
färre blodparasiter och ett starkare immunförsvar. Jag fann att hybriders
immunförsvar var intermediärt och inte skiljde sig från någon av föräldraarterna. Infektionsfrekvensen av blodparasiter skiljde sig däremot mellan de
båda föräldrararterna och halsbandsflugsnappare hade färre parasiter. Detta
kan, i sin tur, påverka hur pass konkurrenskraftiga de båda arterna är och
därmed påverka deras samexistens i hybridzonen.
En viktig slutsats som man kan dra från flugsnapparna är att olika processer under deras liv kan påverkas olika av hybridisering. Hybridernas fertilitet är nedsatt men inte deras förmåga att motstå parasiter. Dessutom kan
28
hybridisering ibland medföra direkta fördelar. En viktig fråga blir då; hur ska
man mäta selektionen mot hybridisering? Hur pass evolutionärt framgångsrik en individ är mäts i dess relativa genetiska bidrag till kommande
generationer, vilket i praktiken är lättare sagt än gjort. Istället brukar forskare
mäta olika komponenter som tillsammans påverkar en individs reproduktiva
framgång så som direkta fördelar eller immunförsvar. När det gäller hybrider
kan en generations reproduktiva framgång vara mycket missvisande eftersom även hybridernas avkomma också kan påverkas. I kapitel III uppskattar
jag den andel avkomlingar tre generationer framåt i tiden som hybridisering
har bidragit med till populationen. Den typen av uppskattningar kan sällan
göras ute i naturen eftersom hybrider och speciellt återkorsningar (avkomman från hybrider eller återkorsningar som parar sig med föräldrararterna) är
sällsynta och svåra att identifiera. Därför ger min studie viktig ny information om långsiktiga effekter av hybridisering i naturliga populationer. Jag
visar att inte bara hybrider har nedsatt fertilitet utan också senare generationers återkorsningar. Återkorsningar hade också nedsatt fertilitet på grund av
en hög andel av dem parade sig med individer som tillhör den art som de
delar lägst andel gener med. Min slutliga uppskattning, som alltså tar hänsyn
till flera generationer, visar att det är en stark selektion mot hybridisering.
Flugsnappare som parar sig över artgränsen bidrar bara med 2% av antalet
avkomlingar som icke-hybriserande flugsnappare bidrar med till populationen. Det är alltså mycket mer kostsamt med hybridisering än vad tidigare
studier visat.
VARFÖR SKER HYBRIDISERING?
Eftersom det är en stark selektion mot hybridisering kan man fråga sig varför
de båda arterna inte utvecklat en bättre förmåga att undvika parningar över
artgränsen. En tänkbar förklaring är att de helt enkelt inte har tillbringat tillräcklig lång tid tillsammans. Alternativt finns det andra faktorer som leder
till att individer ibland tvingas eller bara råkar hybridisera.
Även om båda arterna har utvecklat en perfekt preferens för sin egen art
så kan individer hybridisera när det finns ont om partners som tillhör den
egna arten. Valet kanske står mellan att hybridisera eller inte fortplanta sig
alls. I Kapitel IV, undersöker jag hur fördelningen mellan halsbands- och
svartvit flugnsappare påverkar hybridisering. Som förväntat fann jag att
svartvitflugsnapparhonor löpte störst risk att hybridisera när den egna arten
var ovanlig. Däremot var den totala hybridiseringsfrekvensen högst i områden där de båda arterna förekom i lika stora antal. En tänkbar förklaring är
att trots att individuella honor löper störst risk att hybridisera när den egna
arten är ovanlig finns det fler individer som kan göra fel när arterna är mer
jämt fördelade. Alltså är relativ vanlighet en viktig faktor när det gäller att
förklara förekomst av hybridisering mellan de båda flugsnappararterna men
det är inte den enda faktorn som är viktig.
29
I kapitel IV och V identifierar jag flera karaktärer som påverkar felaktigheter i partnervalet (som orsakar hybridisering) hos flugsnapparna. Hanar
från de båda arterna sjunger olika sånger som skiljer sig tydligt när de finns
på olika geografiska platser. När de förekommer tillsammans händer det att
svartvit flugsnapparhanar kopierar element från halsbandsflugsnapparnas
sång och detta leder i sin tur till ökad hybridiseringsrisk. I kapitel IV visar
jag att svartvit flugsnappare är sämre på att särskilja mellan de två arttypiska
sångerna och att detta är en trolig förklaring till att de har en högre benägenhet att välja fel. I blandade par är honorna oftast svartvit flugsnappare.
Denna asymmetri är mindre i områden där svartvit flugsnappare är ovanligare vilket betyder att individuella halsbandsflugnapparhonor ökar sin
benägenhet att göra fel i sådana områden. Varför har de en sämre förmåga
att särskilja den egna artens hanar när det finns gott om dem? Antingen blir
honorna helt enkelt mer slarviga när risken att träffa en hane som tillhör fel
art är liten. Alternativt blir det av någon anledning svårare för dem att särskilja mellan hanarna. En tänkbar anledning till att det blir svårare att särskilja hanarna är att svartvit flugsnapparhanar oftare lär sig element från
halsbandsflugsnapparnas sång i områden där halsbandsflugsnappare utgör
den vanligaste arten. Både benägenheten att lära sig sångelement från den
andra arten och den sämre förmåga att särskilja de två arttypiska sångerna åt
avspeglar antagligen den svartvit flugsnapparens sämre anpassning till hybridiseringsrisken. Att utvecklingen av sådana anpassningar inte har kommit
längre kan förklaras av den korta tid som arterna har samexisterat på Öland
och Gotland och av den stora immigrationen av svartvit flugsnappare från
fastlandet.
Halsbandflugnapparna på Öland och Gotland härstammar däremot från
utbredningsområden där båda arterna har förekommit tillsammans under en
lång tid och dessutom tycks de flesta av individerna vara födda i öpopulationen. Halsbandsflugsnapparna förväntas därför vara relativt bättre
anpassade till att undvika hybridisering. Trots detta har den här arten också
bibehålligt karaktärer som är associerade med hybridiseringsrisk. I kapitel V
visar jag att den fjäderdräkt som ett åriga hanar har medför en ökad hybridiseringsrisk. Detta beror på att ettåriga halsbandflugsnapparhanar inte ännu har
en adult (vuxen) fjäderdräkt och därför utseendemässigt närmar sig svartvit
flugsnapparhanars utseende. Unga halsbandsflugsnapparhanar väljs därför i
första hand av de svartvit flugsnapparhonor som inte hittar en artegen partner. Trots kostnaden av avsaknaden av vuxna karaktärsdrag hos ett åriga
hanar bibehålls denna egenskap antagligen på grund av en ännu högre kostnad att anlägga en vuxen fjäderskrud för unga hanar. Kanske skulle det leda
till ökad aggression från äldre hanar. Mina resultat illustrerar en vanlig kritik
mot idén att nedsatt fortplantingsförmåga hos hybrider leder till en urvalsprocess som driver artbilningsprocessen framåt och avslutar den. När hybridisering har blivit tillräckligt ovanligt så utgör hybridiseringsrisk inte
30
längre ett viktigt selektionstryck och utvecklingen mot en fullständig reproduktiv isolering avstannar.
SLUTSATSER
Den här avhandlingen presenterar det hitintills bästa måttet på styrkan av
selektionen mot hybridisering i en naturlig population. Genom att undersöka
selektion mot hybridisering i en naturlig miljö kan studier som min ta reda
på vilka förhållande som gör att populationer som hybridiserar antingen slås
samman eller separerar från varandra. Den typen av kunskap är kritisk för
vår förståelse av uppkomsten och bevarandet av biodiversitet på jorden. Utvecklingen av genetiska hjälpmedel gör det realistiskt att spåra hybridisering
i flera generationer och i mer detalj studera genflöde mellan populationer i
hybridzoner även hos andra arter än flugsnappare.
Jag fann att postzygotiska barriärer mot genflöde mellan flugsnappararterna är starka medan de prezygotiska bärriärerna är förhållandevis
ofullkomliga. Den upptäckten kontrasterar med den tidigare uppfattningen
att prezygotiska barriärer generellt utvecklas snabbare hos fåglar. Jag identifierade ett antal karaktärer (t.ex. förmåga att särskilja den egna artens sång,
sångkopiering, åldersrelaterad fjäderdräkt) som ger upphov till en ofullkomlig prezygotisk barriär. I förlängningen leder mina resultat till testbara
förutsägelser om hur dessa karaktärer kommer att utvecklas i populationer
som lever inom och utanför hybridzonen. Sådana jämförelser skulle kunna
avslöja om reproduktiv isolering delvis utvecklas till följd av direkt selektion
för ökad isolering i hybridzoner eller om den uppstår som en sidoeffekt av
processer som helt sker inom arten.
Ficedula hybridzonen är ett ypperligt studiesystem för en rad
frågeställningar rörande artbildning, och min avhandling representerar ett
viktigt första steg för att besvara några av dessa centrala frågor om arters
uppkomst.
31
Acknowledgements
First, I’d like to extend thanks to my supervisors, Anna Qvarnström and Lars
Gustafsson, for allowing me the opportunity to come to Uppsala and work
on what has long been a wonderful system for studying animal ecology and
evolution. A special thanks to you, Anna, for always being supportive and
enthusiastic (even if sometimes overly optimistic!) throughout my work. I’ve
really enjoyed working with you, and hope there will be plenty more opportunities for this in the future. Being one of many in the ‘flycatcher group’ in
Uppsala, there was always someone happy to lend advice about the system
and provide great company through those hectic field months. Thank you
Thor Veen, Nina Svedin, Mårten Hjernquist, Stein Are Sæther, Joanna Sendecka, and Jukka Forsman for being a great team to work with. In particular,
thanks Mårten for proofreading the introduction to this thesis. Thanks especially to Nina, who never hesitates to drop everything to help out. It has been
great having someone else alongside ‘for the ride’, from learning how to
measure a flycatcher to writing up a thesis…not to mention the numerous
nights hanging out watching the prime of British comedy and the worst of
American reality TV!
To my collaborators at the University of Oslo, Glenn Peter Sætre, Gunilla
Andersson and Thomas Borge, thanks for being such a pleasure to work with
and for being there to fall back on when the usual lab problems arose!
Thanks especially to Glenn Peter, Maria, Helena and Camilla for kindly
providing a home-away-from-home during my many visits to Norway.
Thank you to Franjo Weissing for inviting me to spend time with the
Theoretical Group at Groningen. Your kindness and passion for science is
inspiring. To Thor, thanks for all the help during my stay in the Netherlands
(at least when you finally escaped Turkish imprisonment), and for always
being enjoyable company both during work and play. To the superprogrammer, Richel Bilderbeek, thanks for your help, and best of luck with
your welding/knitting/spicy-food-eating/music-writing/theatre-lighting/stand
-up comedy. Thanks also to Liz, Laura and Stephanie for making my stay in
the Netherlands lots of fun.
Back at Uppsala I wish to thank Mats Björklund and Göran Arnqvist for
always being happy to help out whenever I needed some statistical advice.
To Katherine Thuman Hjernquist, Dagmar Jonsson and Reija Dufva thanks
for teaching me the art of microsatellites and helping out in the lab. Thanks
especially to Reija, who not only ran many of the paternity analyses, but also
32
checked through all the parasites. For great help in the field during the various field seasons, I wish to thank Katherine, Mathilda, Isobel, Mattias, Lisa,
Janette, Kalle, and Selma. In addition, I would especially like to thank Johan
Träff for not only kindly offering access to heterospecific pairs within his
personally managed flycatcher population on Gotland, but also for sharing
his wealth of knowledge about flycatcher biology. To my collaborators on
the various papers, thank you for helping make this work possible. In addition to those co-authors I’ve mentioned earlier, thanks Jenny Bengtson, Nilla
Fogelberg and Jenny Vogel Kehlenbeck. Thanks to my many office-mates
over the years (Göran S, Cath, Mårten, Emma, Katherine, Sara, Sarah and
Mårten again), and to all the others that I had the pleasure of spending time
with at the department (Alexei, Anders Ö., Anders B., Björn, Bo W.,
Claudia, Damian, Ingrid, Johanna A., Johanna R., Mari, Marta, Martin, Marnie, Niclas, Olivia, Olle, Sandra, Ted, Urban). It’s never easy moving so far
from home, but the friendly, social environment at work and the wonderful
people I shared the workplace with has made this a thoroughly enjoyable
four years. On the same note, thanks to my non-Zooeko friends (Dick, Johiris, Annika, Veronica, Henrik) for making my time in Sweden so much
fun. I hope to see you all in Australia some time!
To my family, accepting that I would spend four years away on the other
side of the world cannot have been easy. I really appreciate that you made
the effort to be able to come to the defense. I hope you get to see what I have
been up to all this time! To Mum, Dad, Liam and Rach, I love you.
33
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Acta Universitatis Upsaliensis
Digital Comprehensive Summaries of Uppsala Dissertations
from the Faculty of Science and Technology 245
Editor: The Dean of the Faculty of Science and Technology
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