This study concerns a population of the Pied Flycatcher (Ficedula hypoleuca) living in nestboxes in an area of woodland in Mid-Wales. The occupants of 180 nestboxes were monitored during 1988 and 1989. In addition to behavioural observations and records of breeding performance, individual adults and pulli were caught and measured, and a blood sample taken. In the Pied Flycatcher, polygyny is a common mating strategy in which the two or more females mated to a single male nest in discrete territories up to 500m apart This behaviour has been interpreted in two ways, firstly as the result of female choice for the quality of the male or his territory, and secondly, as a consequence of male deception, by which already-mated males attract secondary females who suffer reduced breeding success as a result. In this population polygyny was a rare occurrence; only 3 of 240 breeding males were recognised to be polygynous. These males defended two adjacent nestboxes. The breeding success of the three secondary females was not unusually low. These results suggest that a model of male- or territory quality might better explain the situation in this population. The occurrence of extra-pair mating has being noted in a number of species, including the Pied Flycatcher. In this study it was found to account for 2.7% of the offspring screened by genetic fingerprinting. Another common method for detecting extra-pair paternity uses the heritability of a skeletal measurement.The results from the two methods are shown to be incompatible. A number of weaknesses with the heritability method are described and discussed. The increasing number of studies on the Pied Flycatcher throughout Europe reveal that the frequency of mating strategies such as polygyny and extra-pair mating differ from area to area This suggests that environmental factors may play a major part in determining the costs and benefits of such strategies.

This study involves two village chicken populations sampled from Horro and Jarso regions of Western and Eastern Ethiopia respectively. This study maps the phenomic and genomic landscape of the two chicken populations using morphological markers and a high density (600K) SNP array. Although the two chicken populations tend to display nondescript morphological characteristics, they show a subtle variation except for rare morph variants that have been in most instances scored on Jarso chickens. Morphological analysis uncovers a vast array of intrapopulation variation. Genetic diversity and population structure analyses assign the two chicken populations to two distinct genepools representing their population of origin. A high intrapopulation genetic diversity is uncovered, which shows a broad genetic base (high genetic diversity) of the two chicken populations. We hypothesized that a clearly evident genetic divergence observed between the two chicken populations may be attributed to difference in demographic history, origin (routes of introduction to Africa), breeding history of the two chicken populations and demographic structure of subsistence farmers. Absence of gene flow owing to their distant geographic location and ecological variation may have also contributed to this divergence. A population structure analysis performed on a random subset of the two Ethiopian chicken populations along with village chickens sampled from other African countries, Asia and Latin America, commercial populations and the junglefowl species reveals a unique genetic structure of Ethiopian chickens, which implicates the need for further study on the genetic landscape of the latter. To infer the extent of inbreeding we performed a run of homozygosity analysis (ROH). Our analysis indicates that ROH is more intense in Jarso than Horro chickens and in macrochromosomes than microchromosomes. The extensive ROH mapped in some chickens implicates the need to restructure the existing traditional breeding practice of subsistence farmers. Our analysis confirms the commonness of ROH in genic regions. For the first time, we detect twenty three putative uniparental disomy in twenty two Ethiopian village chickens. Signature of selection analysis detects divergently selected genomic regions in the two chicken populations indicating a considerable divergent selection imposed on the two populations. Genes involving in melanogenesis pathway are among those subjected to a divergent selection. However, some overlapping regions were also mapped in the two chicken populations implicating the ubiquitous impact of natural selection on genes regulating vital biological processes. A genome-wide association study performed on pigmentation (earlobe, plumage and shank) traits and variants of crest, comb and a lightly feathered shank maps a number of putative loci that may underlie variations in these traits. Our GWAS analysis on pigmentation traits produced a long list of loci than that have been known to involve in the genetic control of pigmentation in the chicken, with most of these have been mapped in the mouse. We also refined further the causative variants underlying a lightly feathered shank mutation. Our GWAS analysis map a number of putative novel loci that may underlie the genetic control of the traits analysed and this has laid a foundation for subsequent work that would involve targeted sequencing and a candidate gene approach. This study is the first of its kind in Africa that uses a large number of samples and a high density SNP array to unlock phenomic and genomic landscape of the true type village chickens.

Here I investigate, using autosomal whole-genome sequence data, the signature of positive selection and/or introgression in the indigenous domestic village chickens from three countries (Ethiopia, Saudi Arabia, Sri Lanka), three fancy birds, three junglefowl species (red junglefowl Gallus gallus, grey junglefowl G. sonneratii, Ceylon Junglefowl G. lafayettii) and the Javan red junglefowl G. g. bankiva. All the new sequencing data were obtained from Illumina HiSeq 2000/2500 DNA sequencers with an individual bird depth of genome coverage ranging from 10 X to 30 X. The analyses in this thesis have been completed using the reference genome Galgal 4.0. For the detection of signatures of positive selection, this analysis excluded the three fancy birds and the grey junglefowl due to small sample size. Using the pool heterozygosity and SweeD composite likelihood selection signature methods, I identified two candidate selected regions shared between all the three indigenous domestic village chicken populations and the red junglefowl (chapter 2). These regions contain genes that are associated with the development of the central nervous system and adaptation to hypoxic environments. Five candidate regions were shared among the three indigenous village chicken populations, and they represent candidate domestication regions. Unique regions in each domestic chicken population were also identified. Functional genes have not been assigned to most of these regions but in those where the genes have been annotated, the gene function may be related to production and reproductive traits as well as adaptation to cold/hot temperatures and hypoxia. In chapter 3, I analysed only the Ceylon and green junglefowl whole genome sequences for the detection of candidate signatures of positive selection using both the pool heterozygosity and Tajima’s D. In both species, I identified candidate selected regions that contained genes which may be linked to adaptation to different environmental challenges e.g disease resistance, stress, thermoregulation and hypoxia. In the genome of green junglefowl, candidate selected regions associated with skeletal formation and ovarian follicle development were significantly detected. In chapter 4, I identified introgressed candidate regions from the grey and Ceylon junglefowls in domestic chicken (including the three indigenous chicken populations and fancy birds) using the ABBA – BABA four taxon method. Our result shows that, domestic chickens shared 75.8% of their genome with the red junglefowl, 4% with the grey junglefowl and 1.1% with the Ceylon junglefowl. I observed introgression in both directions, namely from the grey/Ceylon junglefowls into domestic chickens and vice versa. While from the grey junglefowl, introgression was present in all the domestic chicken populations as well as interestingly in the red junglefowl, for the Ceylon junglefowl, introgression was more restricted to the domestic chicken from Sri Lanka. From the ABBA – BABA analysis between the grey junglefowl and the domestic chicken, I also identified a single candidate introgressed region from the green junglefowl G. varius in two domestic birds from Sri Lanka. Future study should therefore consider investigating the genome-wide analysis of introgression from the green junglefowl into the domestic chicken. In chapter 5, I ended our introgression study by investigating if a distantly related subspecies of red junglefowl, the Javan red junglefowl Gallus gallus bankiva, has contributed to the gene pool of the domestic chicken. Alongside the three indigenous domestic chicken populations, I included the genome sequences of three domestic chickens sampled from the Java island of Indonesia. Our result shows a significant 10.6% genome admixture between the domestic chicken and the Javan red junglefowl. Overall, our results indicate that the genetic make-up of the domestic chicken is rather complex with multiple species and subspecies influences. These introgression events have contributed to the genetic diversity of these domesticates. Our results also support the geographic difference of introgression and indicate that these introgression events may have contributed to the adaptive traits of the domestic chicken. However, this requires further investigation.

‘Eco-evolutionary feedbacks’ occur when evolution of organismal traits causes environmental change that drives further evolution. Predator and prey interactions provide good examples of eco-evolutionary feedbacks. Here I examine the potential for eco-evolutionary feedbacks between three-spined sticklebacks (Gasterosteus aculeatus, hereafter ‘stickleback’) and their zooplankton prey in lochs (lakes) on the Scottish island of North Uist. Many lochs on the island were colonised by sticklebacks after the last glaciation, approximately 10,000-15,000 years ago. Previous work has shown that sticklebacks in different lochs have diverged greatly from each other in response to local environments. On the other hand, apart from several very old studies on the occurrence of some zooplankton species, there have been no previous in depth studies on the population dynamics of zooplankton on North Uist. I investigated first the diversity and abundance of zooplankton groups and the most common species across all North Uist lochs. Thirty-nine species were classified from three main groups (Rotifera, Cladocera and Copepoda). Species abundance in the presence of fish was relatively more even, while the abundance of zooplankton groups was not generally related to fish presence except for Cladocera in a subset of locations. The effects of predation on the life-history of zooplankton were also examined by comparing reproductive traits of the dominant cladoceran species (Bosmina and Daphnia) in lochs with and without fish. Cladocerans in lochs with fish have more rapid reproductive cycles and higher fecundity parameters, probably in response to the increased threat of predation. These effects remain evident in Bosmina after they have been raised through three generations in the laboratory, supporting the hypothesis that they have a genetic basis and are not a plastic response to predator presence. Life-history variables of cladocerans were also related to abiotic and biotic variation among lochs, including depth, pH, chlorophyll levels and the concentration of alkaline metals. I also demonstrate that stickleback have diversified greatly among lochs in functional trophic traits that determine the efficiency of feeding on different types of prey, which could affect total primary production and the structure of prey communities. Ancestral stickleback populations have adapted according to the type of habitat colonised. Fish feed on benthic prey in shallow lochs, which requires greater effort for successful foraging than that required by fish that feed on planktonic prey. I found that variation in stickleback trophic traits was related to both abiotic and biotic variation among lochs, including loch depth and the mean contributions of planktonic and benthic prey to diet. The results presented in this thesis suggest the possibility of eco-evolutionary feedbacks in these simple ecosystems on North Uist based on the significant responses in fish and zooplankton communities. The thesis also provides a basis for further studies on fish-zooplankton interactions on North Uist and contributes to the wider body of knowledge concerning the relevance of natural variation in shaping the foraging mechanisms of animals.

O presente projeto representa uma extensão dos paradigmas que criamos com estudos de simulação molecular para compreender as frequentes mudanças de estrutura e função das Aldeído Desidrogenases (ALDHs) durante a evolução. As ALDHs formam uma superfamília de proteínas que catalisam a oxidação de vários aldeídos, mas as origens evolutivas das preferências pelos seus substratos são pouco conhecidas. Apesar de possuírem uma elevada identidade sequencial, duas destas ALDHs, a ALDH1 e a ALDH2, exibem distintos papéis funcionais de sinalização celular e detoxicação, respectivamente. Através de prévia análise computacional e logenética, identicamos que, curiosamente, as ALDH1s de organismos invertebrados, Branchiostoma oridae e Ciona intestinalis apresentam características estruturais mais semelhantes às de suas ALDH2s do que das ALDH1 típicas. Isto sugere que essas ALDH1s divergentes podem ter evoluído na direção da atividade de degradação de aldeídos pequenos e tóxicos, que parecem representar a função ancestral das ALDH2s eucarióticas. Nossa análise identicou três assinaturas de aminoácidos localizadas na área interna do canal de entrada do substrato (CES) que distinguem as ALDH1 das ALDH2. Desta forma, constatamos que as ALDH1s possuem um CES amplo e desobstruído, consistente com o fato destas enzimas catalisarem aldeídos de cadeia longa como o retinaldeído, que é um precursor de vias de sinalização por retinóides. Em contraste, as ALDH2s possuem o CES pouco volumoso e constrito, consistente com sua função na degradação de pequenos aldeídos tóxicos e reativos, como o acetaldeído. Neste projeto o nosso objetivo é analisar a correlação funcional e estrutural entre as ALDH1 e ALDH2 presentes em B. oridae e C. intestinalis para desvendar e compreender seus papéis funcionais e evolutivos em cordados. Especicamente, testaremos a hipótese de que as três assinaturas descritas constituem o núcleo fundamental da preferência por substratos, e que sua inversão por mutações sitio-dirigidas entre ALDH1 e ALDH2 modicará a preferência de substrato de acordo com a origem da assinatura. Se conrmado experimentalmente, este será um exemplo pioneiro de reversão evolutiva molecular, que terá impacto direto sobre as interpretações atuais sobre controversa lei de Dollo da irreversibilidade evolutiva. / This project represents an extension of the paradigms that we created from our molecular simulation studies to understand the frequent structure and function changes of aldehyde dehydrogenases (ALDH) during evolution. The ALDHs form a superfamily of proteins that catalyze the oxidation of several aldehydes, but the evolutionary origins of their substrate preference are unknown. Despite having a high sequence identity, two of these ALDHs, the ADLH1, and ALDH2, exhibit distinct functional roles of cellular signaling and detoxication, respectively. Through previous computational and phylogenetic analysis, we found that, interestingly, the ALDH1s of invertebrate organisms (Branchiostoma oridaeand Ciona intestinalis) show structural features more similar to their ALDH2s than the typical ALDH1. It suggests that these divergent ALDH1sevolved to provide small aldehydes detoxication pattern, what seems to represent the ancestral eukaryotic ALDH2s function. Our analysis also identied three aminoacidsignatures, located internally in the substrate entry channel (SEC), that distinguishes the ALDH1 from ALDH2. Thus, we nd that ALDH1s have a wide, open and unobstructed SEC, consistent with the fact that these enzymes catalyze bulky long-chainaldehydes, like retinaldehyde, a precursor of important signaling pathways. In contrast, the ALDH2s have a small and constricted SEC, consistent with the degradation function of small aldehydes, like the toxic metabolite acetaldehyde. In this project, our objective is to understand the functional and structural correlation between ALDH1 and ALDH2 found in B. oridaeand C. intestinalisto discover and comprehend their functional and evolutionary roles in chordates. Specically, we will test the hypothesis that the three signatures described above are the fundamental core of the substrate preference, based on the signature origin. If conrmed experimentally, this will be a pioneeringexample of evolutionary molecular reversion, impacting directly on the current interpretations of the controversial Dollos Law of Irreversibility.

Le but de toute étude en biologie évolutive du développement est l’étude des mécanismes développementaux à l’origine des diversifications morphologiques. Dans ce contexte, j’ai décidé de me focaliser sur l’émergence des Vertébrés au cours de l’évolution, par la mise en œuvre d’études comparatives entre différents modèles de Deutérostomiens. Le travail réalisé durant ma thèse est subdivisé en trois projets: (i) j’ai abordé le lien entre l’évolution du cerveau chez les Chordés et les modifications de la signalisation à l’acide rétinoïque (AR) au cours du développement. En particulier, j’ai examiné les rôles de l’AR au cours du développement du cerveau chez la lamproie Lampetra fluviatilis, et j’ai comparé les résultats obtenus chez cette espèce aux mécanismes développementaux agissant chez l’amphioxus, un Chordé invertébré, et chez les modèles gnathostomes classiques. Les données obtenues lors de ces analyses comparatives ont permis une meilleure compréhension de l’évolution de la régionalisation cérébrale chez les Vertébrés. (ii) j’ai étudié l’évolution des séquences régulatrices présentes au sein des clusters de gènes hox, connus pour agir dans la régionalisation du système nerveux des Chordés. L’identification d’éléments non-codants conservés ainsi que d’éléments de réponse à l’AR (RARE) potentiels dans des clusters hox de Chordés, combinée à la caractérisation de RAREs in vivo en cellules murines a permis une vision intégrée de l’évolution du contrôle des gènes hox par l’AR, chez les Chordés. (iii) j’ai analysé l’évolution des microARNs chez les Chordés en comparant les répertoires microARN chez plusieurs espèces de Deutérostomiens. Cette étude a permis d’émettre de nouvelles hypothèses quant à l’émergence des microARNs chez les animaux. Toutes ces analyses ont abordé différents aspects de l’évolution des Chordés avec pour objectif la proposition d’une vision intégrée des mécanismes moléculaires à l’origine de l’émergence des Vertébrés. / The aim of the evolutionary developmental biology is to study the developmental mechanisms at the base of morphological diversification. In this context, I decided to focus my attention on the emergence of vertebrates during evolution by carrying out comparative analyses in several deuterostome models. The work carried out during of my thesis can be subdivided into three major projects: (i) I addressed the link between brain evolution and modifications in retinoic acid (RA) signaling during chordate development. In particular, I investigated the roles of RA signaling in brain development in a jawless vertebrate, the lamprey Lampetra fluviatilis, and compared the results with developmental mechanisms in the invertebrate chordate amphioxus and classical developmental model systems in jawed vertebrates. Data obtained from these comparative studies provided insights into the evolution of brain patterning in vertebrate evolution. (ii) I investigated the evolution of the regulatory landscape of hox gene clusters that are known to be fundamental for the patterning of the chordate central nervous system. The identification of conserved non-coding elements and putative RA response elements (RAREs) in hox clusters of different chordate species combined with the in vivo characterization of functional RAREs in mouse F9 cells provided an integrated view of the evolution of RA-dependent hox cluster regulation in chordates. (iii) I studied the roles of microRNAs (miRNAs) in chordate evolution by comparing the miRNA complements of different deuterostome species. This analysis provided novel insights about the general mechanisms of miRNA emergence in animals and highlighted species-specific miRNA complement amplifications in different deuterostome lineages. In sum, these studies have tackled different aspects of chordate evolution from an evo-devo perspective, aiming at an integrated view of the molecular mechanisms underlying vertebrate diversification.

Light detection is one of the crucial abilities of all animals. The light cues are important e.g. for maintaining of circadian rhythms, regulation of spawning cycles, changes of pigmentation and arguably most importantly for vision. Most animals detect light by opsins, members of the G protein coupled receptors superfamily. Amphioxus belongs to earliest branching chordate clade, cephalochordates. Thanks to their phylogenetic position, physiology and morphology, cephalochordates became the most relevant model organism for understanding the evolutionary origins of vertebrate specific traits. Amphioxus evince various reactions to light throughout its development. In the presented thesis light detecting systems of amphioxus were studied thoroughly. More specifically characterization of the opsin gene repertoire of two amphioxus species Branchiostoma floridae and Branchiostoma lanceolatum and their comparison with opsins from other animals is presented. In addition, remarkable similarity on the gene expression level between one of amphioxus visual organs, so called frontal eye, and neurons and retinal pigmented epithelium in vertebrate retina was shown. These data confirm the long time ago proposed homology between amphioxus frontal eye and vertebrate lateral eyes. Taken together all the presented data...