<p>In this thesis, different genetic tools are used to investigate pre- and postzygotic barriers to gene exchange and their role in speciation in the pied flycatcher (<i>Ficedula hypoleuca</i>) and the collared flycatcher (<i>F. albicollis</i>). This species complex consists of four genetically distinct clades that apparently diverged in allopatry (I). Sequencing of introns from autosomal and Z-linked genes from the two species reveals signs of selection on the Z-chromosome. Sexual selection acting on Z-linked genes might explain this pattern (II). By using large-scale genotyping of single nucleotide polymorphisms (SNPs), introgression is observed at autosomal- but not Z-linked loci, mostly from the pied- to the collared flycatcher. Male plumage characters and genes involved in hybrid fitness are largely mapped to the Z-chromosome (III). By studying mate choice of female hybrids I show that there is a link between female preferences and the Z chromosome (IV). The rate of introgression in island versus clinal hybrid zones is consistent with regional differences in hybrid fertility. Asymmetric gene flow from allopatry on the islands may oppose reinforcement, leading to introgression and a partial breakdown of postzygotic isolation. Adaptive introgression may explain the high rate of introgression observed at one of the genetic markers (V). For late breeding female collared flycatchers it appears to be adaptive to use pied flycatchers as social fathers but conspecific males as genetic fathers. Additionally, females in mixed species pairs may reduce hybridization costs by producing an excess of male hybrid offspring that are more fertile than females (VI).</p><p>In conclusion, the Z-chromosome appears to play a major role in flycatcher speciation. Sexual selection and reinforcement are important mechanisms in the divergence of these birds. However, gene flow from allopatry, introgression of adaptive genes and adaptive hetrospecific pairing by late breeding collared flycatcher females may work in the opposite direction.</p>

This thesis presents results from genetic studies conducted in the chicken (Gallus gallus). The domestication of chicken is believed to have been initiated approximately 7,000 – 9,000 years ago in Southeast Asia. Since that time, selective breeding has altered the appearance of the wild ancestor, creating highly specialized chicken lines developed for egg and meat production. The first part of this thesis describes a detailed genetic analysis conducted on an F2 intercross between two phenotypically diverse chicken lines. The two parental lines used in this experiment originated from the same base population and have been developed by divergent selection for juvenile body-weight. Selection during forty generations has resulted in an eight-fold difference in body-weight between the High-Weight Selected (HWS) and Low-Weight Selected (LWS) line. In an attempt to identify the genetic factors differentiating the two lines, a large intercross population was bred to map Quantitative Trait Loci (QTL) affecting body-weight traits. A linkage map was constructed which included 434 genetic markers covering 31 of the 38 chicken autosomes. Although there is a dramatic phenotypic difference between the two founder lines, the QTL analysis for marginal effect could only identify seven QTL, each with small additive effects, influencing body-weight. We extended the genetic analysis to also include a model testing for pair-wise interactions between loci (epistasis). The analysis revealed 15 QTL pairs that affect body-weight and several of those formed a network of interacting loci. These results suggest that the genetic basis for the large difference in body-weight is most likely a result of a combined effect of multiple genetic factors, including QTL with small additive effects in combination with pair-wise interactions between QTL. The second part of this thesis presents two linkage maps. The first map constructed was of the chicken Z chromosome, the second used a genome-wide marker set, including 12,945 SNP markers, to build an updated consensus map of the chicken genome. The resulting consensus map includes 9,268 genetic markers and covers 33 chromosomes, still leaving five microchromosomes without marker coverage. The genome average rate of recombination was estimated to 3.1 cM/Mb, but varied considerably between and within chromosomes. A general trend of elevated recombination rates towards telomeric ends and lower rates near centromeres was observed. This was in concordance to previous reports from mammalian species. Recombination rates in chicken were also found to be highly positively correlated with GC-rich sequences.

In this thesis, different genetic tools are used to investigate pre- and postzygotic barriers to gene exchange and their role in speciation in the pied flycatcher (Ficedula hypoleuca) and the collared flycatcher (F. albicollis). This species complex consists of four genetically distinct clades that apparently diverged in allopatry (I). Sequencing of introns from autosomal and Z-linked genes from the two species reveals signs of selection on the Z-chromosome. Sexual selection acting on Z-linked genes might explain this pattern (II). By using large-scale genotyping of single nucleotide polymorphisms (SNPs), introgression is observed at autosomal- but not Z-linked loci, mostly from the pied- to the collared flycatcher. Male plumage characters and genes involved in hybrid fitness are largely mapped to the Z-chromosome (III). By studying mate choice of female hybrids I show that there is a link between female preferences and the Z chromosome (IV). The rate of introgression in island versus clinal hybrid zones is consistent with regional differences in hybrid fertility. Asymmetric gene flow from allopatry on the islands may oppose reinforcement, leading to introgression and a partial breakdown of postzygotic isolation. Adaptive introgression may explain the high rate of introgression observed at one of the genetic markers (V). For late breeding female collared flycatchers it appears to be adaptive to use pied flycatchers as social fathers but conspecific males as genetic fathers. Additionally, females in mixed species pairs may reduce hybridization costs by producing an excess of male hybrid offspring that are more fertile than females (VI). In conclusion, the Z-chromosome appears to play a major role in flycatcher speciation. Sexual selection and reinforcement are important mechanisms in the divergence of these birds. However, gene flow from allopatry, introgression of adaptive genes and adaptive hetrospecific pairing by late breeding collared flycatcher females may work in the opposite direction.

Theory predicts that sexually antagonistic mutations will be over- or under-represented on the X and Z chromosomes, depending on the average dominance coefficient of the mutations. However, as little is known about the dominance coefficients for new mutations, the effect of sexually antagonistic selection is difficult to predict. To elucidate the role of sexually antagonistic selection in the evolution of Z chromosome gene content in chicken, we analyzed publicly available microarray data from several somatic tissues as well as somatic and germ cells of the ovary. We found that the Z chromosome is enriched for genes showing preferential expression in ovarian somatic cells, but not for genes with preferential expression in primary oocytes or non-sex-specific somatic tissues. Our results suggest that sexual antagonism leads to higher abundance of female-benefit alleles on the Z chromosome. No bias towards Z-linkage of oocyte-enriched genes can be explained by lower intensity of sexually antagonistic selection in ovarian germ cells compared to ovarian somatic cells. An alternative explanation would be that meiotic Z chromosome inactivation hinders accumulation of oocyte-expressed genes on the Z chromosome. Our results are consistent with findings in mammals and indicate that recessive rather than dominant...

Unambiguous gender identification (ID) is needed to assess parameters in studies of population dynamics, behavior, and evolutionary biology of Caribbean Flamingo (Phoenicopterus ruber ruber) and other birds. Due to its importance for management and conservation, molecular (DNA-based) avian gender ID assays targeting intron-size differences of the Chromosome Helicase ATPase DNA Binding (CHD) gene of males (CHD-Z) and females (CHD-W) have been developed. Male (ZZ) and female (WZ) genotypes are usually scored as size polymorphisms through agarose or acrylamide gels. For certain species, W-specific restriction sites or multiplex polymerase chain-reaction (PCR) involving CHD-W specific primers are needed. These approaches involve a minimum of three steps following DNA isolation: PCR, gel electrophoresis, and photo-documentation, which limit high throughput scoring and automation potential. In here, a short amplicon (SA) High-resolution Melting Analysis (HRMA) assay for avian gender ID is developed. SA-HRMA of an 81-Base Pair (bp) segment differentiates heteroduplex female (WZ) from homoduplex male (ZZ) genotypes by targeting Single-nucleotide Polymorphisms (SNPs) instead of intron-size differences between CHD-Z and CHD-W genes. To demonstrate the utility of the approach, the gender of Caribbean Flamingo (P. ruber ruber) (17 captive from the Dallas Zoo and 359 wild from Ria Lagartos, Yucatan, Mexico) was determined. The assay was also tested on specimens of Lesser Flamingo (P. minor), Chilean Flamingo (P. chilensis), Saddle-billed Stork (Ephippiorhynchus senegalensis), Scarlet Ibis (Eudocimus ruber), White-bellied Stork (Ciconia abdimii), Roseate Spoonbill (Platalea ajaja), Marabou Stork (Leptoptilos crumeniferus), Greater Roadrunner (Geococcyx californianus), and Attwater's Prairie Chicken (Tympanuchus cupido attwateri). Although the orthologous 81 bp segments of Z and W are highly conserved, sequence alignments with 50 avian species across 15 families revealed mismatches affecting one or more nucleotides within the SA-HRMA forward or reverse primers. Most mismatches were located along the CHD-Z gene that may generate heteroduplex curves and thus gender ID errors. For such cases, taxon and species-specific primer sets were designed. The SA-HRMA gender ID assay can be used in studies of avian ecology and behavior, to assess sex-associated demographics and migratory patterns, and as a proxy to determine the health of the flock and the degree by which conservation and captive breeding programs are functioning.