Horizontal Transmission and Recombination Maintain forever Young Bacterial Symbiont Genomes

Bacterial symbionts bring a wealth of functions to the associations they participate in, but by doing so, they endanger the genes and genomes underlying these abilities. When bacterial symbionts become obligately associated with their hosts, their genomes are thought to decay towards an organelle-like fate due to decreased homologous recombination and inef- ficient selection. However, numerous associations exist that counter these expectations, especially in marine environments, possibly due to ongoing horizontal gene flow. Despite extensive theoretical treatment, no empirical study thus far has connected these underlying population genetic processes with long-term evolutionary outcomes. By sampling marine chemosynthetic bacterial-bivalve endosymbioses that range from primarily vertical to strictly horizontal transmission, we tested this canonical theory. We found that transmission mode strongly predicts homologous recombination rates, and that exceedingly low recombination rates are associated with moderate genome degradation in the marine symbionts with nearly strict vertical transmission. Nonetheless, even the most degraded marine endosym- biont genomes are occasionally horizontally transmitted and are much larger than their ter- restrial insect symbiont counterparts. Therefore, horizontal transmission and recombination enable efficient natural selection to maintain intermediate symbiont genome sizes and sub- stantial functional genetic variation

The Marvelous World of tRNAs: From Accurate Mapping to Chemical Modifications

Since the discovery of transfer RNAs (tRNAs) as decoders of the genetic code, life science has transformed. Particularly, as soon as the importance of tRNAs in protein synthesis has been established, researchers recognized that the functionality of tRNAs in cellular regulation exceeds beyond this paradigm. A strong impetus for these discoveries came from advances in large-scale RNA sequencing (RNA-seq) and increasingly sophisticated algorithms. Sequencing tRNAs is challenging both experimentally and in terms of the subsequent computational analysis. In RNA-seq data analysis, mapping tRNA reads to a reference genome is an error-prone task. This is in particular true, as chemical modifications introduce systematic reverse transcription errors while at the same time the genomic loci are only approximately identical due to the post-transcriptional maturation of tRNAs. Additionally, their multi-copy nature complicates the precise read assignment to its true genomic origin. In the course of the thesis a computational workflow was established to enable accurate mapping of tRNA reads. The developed method removes most of the mapping artifacts introduced by simpler mapping schemes, as demonstrated by using both simulated and human RNA-seq data. Subsequently, the resulting mapping profiles can be used for reliable identification of specific chemical tRNA modifications with a false discovery rate of only 2%. For that purpose, computational analysis methods were developed that facilitates the sensitive detection and even classification of most tRNA modifications based on their mapping profiles. This comprised both untreated RNA-seq data of various species, as well as treated data of Bacillus subtilis that has been designed to display modifications in a specific read-out in the mapping profile. The discussion focuses on sources of artifacts that complicate the profiling of tRNA modifications and strategies to overcome them. Exemplary studies on the modification pattern of different human tissues and the developmental stages of Dictyostelium discoideum were carried out. These suggested regulatory functions of tRNA modifications in development and during cell differentiation. The main experimental difficulties of tRNA sequencing are caused by extensive, stable secondary structures and the presence of chemical modifications. Current RNA-seq methods do not sample the entire tRNA pool, lose short tRNA fragments, or they lack specificity for tRNAs. Within this thesis, the benchmark and improvement of LOTTE-seq, a method for specific selection of tRNAs for high-throughput sequencing, exhibited that the method solves the experimental challenges and avoids the disadvantages of previous tRNA-seq protocols. Applying the accurate tRNA mapping strategy to LOTTE-seq and other tRNA-specific RNA- seq methods demonstrated that the content of mature tRNAs is highest in LOTTE-seq data, ranging from 90% in Spinacia oleracea to 100% in D. discoideum. Additionally, the thesis addressed the fact that tRNAs are multi-copy genes that undergo concerted evolution which keeps sequences of paralogous genes effectively identical. Therefore, it is impossible to distinguish orthologs from paralogs by sequence similarity alone. Synteny, the maintenance of relative genomic positions, is helpful to disambiguate evolutionary relationships in this situation. During this thesis a workflow was computed for synteny-based orthology identification of tRNA genes. The workflow is based on the use of pre-computed genome-wide multiple sequence alignment blocks as anchors to establish syntenic conservation of sequence intervals. Syntenic clusters of concertedly evolving genes of different tRNA families are then subdivided and processed by cograph editing to recover their duplication histories. A useful outcome of this study is that it highlights the technical problems and difficulties associated with an accurate analysis of the evolution of multi-copy genes. To showcase the method, evolution of tRNAs in primates and fruit flies were reconstructed. In the last decade, a number of reports have described novel aspects of tRNAs in terms of the diversity of their genes. For example, nuclear-encoded mitochondrial-derived tRNAs (nm-tRNAs) have been reported whose presence provokes intriguing questions about their functionality. Within this thesis an annotation strategy was developed that led to the identification of 335 and 43 novel nm-tRNAs in human and mouse, respectively. Interestingly, downstream analyses showed that the localization of several nm-tRNAs in introns and the over-representation of conserved RNA-binding sites of proteins involved in splicing suggest a potential regulatory function of intronic nm-tRNAs in splicing

The molecular basis of high duty-cycle echolocation in bats, and its role in the divergence of populations and species

PhD thesisHow populations diverge and form new species in the face of gene flow is a key question in evolutionary biology. Recent research suggests this may be possible where the same traits affect the ecological niche and are involved in assortative mating, and that a small number of genes could be involved in driving speciation in these cases. Echolocation call frequency in bats has roles in ecology and social communication. Bats using HDC echolocation have hearing tuned to specific frequencies, with frequency shifts impacting ecological niche and mate recognition, meaning this is a good candidate trait to drive speciation. HDC echolocation has evolved independently in two highly divergent groups of bats, providing a unique opportunity to study the molecular basis of a trait potentially driving speciation. I have combined selection testing of specific loci with genomewide divergence scans to test hypotheses concerning the evolution of HDC echolocation. Members of the yangochiropteran genus Pteronotus use low duty-cycle echolocation, except for the subgenus Phyllodia. Selection tests on coding sequence data revealed loci associated with hearing under positive selection in Phyllodia and in Pteronotus, including eleven shared with a yinpterochiropteran HDC echolocator, Rhinolophus sinicus. Three size and acoustic morphs of Rhinolophus philippinensis exist in sympatry on Buton Island. Phylogenetic reconstructions revealed population structure between the morphs, though with conflicting topologies based on mitochondrial and nuclear data. Species delimitation identified at least two separate taxa. Genomewide scans of divergence indicated low background FST between the morphs, punctuated with highly diverged islands featuring an overrepresentation of genes associated with body size and hearing. 3 This thesis represents the first genome-wide investigation of HDC echolocation, highlighting candidate genes related to this trait. It additionally describes a rarely observed mammalian ecological speciation, providing support for the claim that species designated R. philippinensis represent a complex across their range

A guinea fowl genome assembly provides new evidence on evolution following domestication and selection in Galliformes

The helmeted guinea fowl Numida meleagris belongs to the order Galliformes. Its natural range includes a large part of sub-Saharan Africa, from Senegal to Eritrea and from Chad to South Africa. Archaeozoological and artistic evidence suggest domestication of this species may have occurred about 2,000 years BP in Mali and Sudan primarily as a food resource, although villagers also benefit from its capacity to give loud alarm calls in case of danger, of its ability to consume parasites such as ticks and to hunt snakes, thus suggesting its domestication may have resulted from a commensal association process. Today, it is still farmed in Africa, mainly as a traditional village poultry, and is also bred more intensively in other countries, mainly France and Italy. The lack of available molecular genetic markers has limited the genetic studies conducted to date on guinea fowl. We present here a first-generation whole-genome sequence draft assembly used as a reference for a study by a Pool-seq approach of wild and domestic populations from Europe and Africa. We show that the domestic populations share a higher genetic similarity between each other than they do to wild populations living in the same geographical area. Several genomic regions showing selection signatures putatively related to domestication or importation to Europe were detected, containing candidate genes, most notably EDNRB2, possibly explaining losses in plumage coloration phenotypes in domesticated populations

Multi-Platform Next-Generation Sequencing of the Domestic Turkey (Meleagris gallopavo): Genome Assembly and Analysis

The combined application of next-generation sequencing platforms has provided an economical approach to unlocking the potential of the turkey genome

Chromosomal-level assembly of the Asian Seabass genome using long sequence reads and multi-layered scaffolding

We report here the ~670 Mb genome assembly of the Asian seabass (Lates calcarifer), a tropical marine teleost. We used long-read sequencing augmented by transcriptomics, optical and genetic mapping along with shared synteny from closely related fish species to derive a chromosome-level assembly with a contig N50 size over 1 Mb and scaffold N50 size over 25 Mb that span ~90% of the genome. The population structure of L. calcarifer species complex was analyzed by re-sequencing 61 individuals representing various regions across the species' native range. SNP analyses identified high levels of genetic diversity and confirmed earlier indications of a population stratification comprising three clades with signs of admixture apparent in the South-East Asian population. The quality of the Asian seabass genome assembly far exceeds that of any other fish species, and will serve as a new standard for fish genomics

Multiple rearrangements and low inter- and intra-species mitogenome sequence variation in the Heterobasidion annosum s.l. species complex

IntroductionMitochondria are essential organelles in the eukaryotic cells and responsible for the energy production but are also involved in many other functions including virulence of some fungal species. Although the evolution of fungal mitogenomes have been studied at some taxonomic levels there are still many things to be learned from studies of closely related species. MethodsIn this study, we have analyzed 60 mitogenomes in the five species of the Heterobasidion annosum sensu lato complex that all are necrotrophic pathogens on conifers. Results and DiscussionCompared to other fungal genera the genomic and genetic variation between and within species in the complex was low except for multiple rearrangements. Several translocations of large blocks with core genes have occurred between the five species and rearrangements were frequent in intergenic areas. Mitogenome lengths ranged between 108 878 to 116 176 bp, mostly as a result of intron variation. There was a high degree of homology of introns, homing endonuclease genes, and intergenic ORFs among the five Heterobasidion species. Three intergenic ORFs with unknown function (uORF6, uORF8 and uORF9) were found in all five species and was located in conserved synteny blocks. A 13 bp long GC-containing self-complementary palindrome was discovered in many places in the five species that were optional in presence/absence. The within species variation is very low, among 48 H. parviporum mitogenomes, there was only one single intron exchange, and SNP frequency was 0.28% and indel frequency 0.043%. The overall low variation in the Heterobasidion annosum sensu lato complex suggests a slow evolution of the mitogenome

Genomics of clownfish adaptive radiation

How species diversify, creating the astonishing biodiversity observed on Earth, has been a central question since Darwin’s On the Origin of Species. Thanks to the sequencing revolution, this question can be approached today from a genomic perspective, examining how intrinsic genomic architecture and extrinsic biological and ecological factors interplay to shape the diversification of organisms. In this sense, clownfishes, which experienced an adaptive radiation following the acquisition of mutualism with sea anemones, represent a fascinating system. Thus, in this thesis, I combine comparative and population genomics approaches to study the genomic architecture underlying the diversification of this group. In the first two sections, I generate genomic resources for ten closely related but ecologically divergent clownfish species and the damselfish Pomacentrus moluccensis. Using this data, I question the genetic mechanisms underlying the acquisition of mutualism. I identify several candidate genes that experienced positive selection at the basis of clownfish radiation and show functions associated with sea anemones toxins discharge, thus likely involved in the evolution of clownfishes' ability to live unharmed within their otherwise-toxic hosts. In the last two sections, I dive into the diversification process of clownfishes. Through comparative genomics approaches, I show that the group experienced bursts of transposable elements, overall accelerated molecular evolution, and ancestral hybridization events, which likely facilitated the radiation of the group by generating the genomic variations necessary for natural selection to act on. I identify genes undergoing differential selective pressures linked with ecological divergence, suggesting that parallel evolution is shaping clownfish diversification, and I pinpoint candidate genes involved in the evolution of the particular size- based hierarchical social structure observed in the group. I finally focus on the mechanisms underlying the evolution of a clownfish clade, the skunk complex. Through population genomics approaches, I demonstrate that gene flow occurred throughout the diversification of the group. Indeed, the species experienced moderate ancestral gene flow, which lessened but still persists in sympatry. Moreover, contrary to what was previously suggested, I demonstrate that A. sandaracinos did not originate from hybrid speciation. I finally pinpoint candidate regions of introgression between species that likely played a role in the diversification of the complex. Overall, my work provides the first insights into the genomic mechanisms underlying clownfish adaptive radiation. -- Comprendre comment les espèces se diversifient et créent l'étonnante biodiversité observée sur Terre sont des questions centrales depuis l’écriture de l’Origine des espèces par Darwin. Grâce à la révolution du séquençage, ces questions peuvent être abordées aujourd'hui en examinant comment l'architecture génomique et les facteurs biologiques et écologiques interagissent et mènent à la diversification des organismes. En ce sens, les poissons-clowns, qui ont connu une radiation adaptative suite à l'acquisition du mutualisme avec les anémones de mer, représentent un système fascinant. Ainsi, dans cette thèse, j’étudie l'architecture génomique qui sous-tend la diversification de ce groupe. Dans les deux premières sections, je génère des ressources génomiques pour dix espèces de poissons-clowns ainsi que pour l’espèce de demoiselle Pomacentrus moluccensis. À partir de ces données, je questionne les mécanismes génétiques qui sous-tendent l'acquisition du mutualisme. J’identifie plusieurs gènes ayant subi une sélection positive à la base du rayonnement des poissons-clowns. Ces gènes ont des fonctions associées à la décharge des toxines des anémones, suggérant donc une implication dans l'évolution de la capacité des poissons-clowns à vivre au sein de leurs hôtes normalement toxiques. Dans les deux dernières sections, je me plonge dans le processus de diversification des poissons-clowns. Grâce à des approches de génomique comparative, je montre non seulement que ce groupe compte une importante quantité d'éléments transposables au sein de son génome, mais qu’il a également subi une évolution moléculaire accélérée ainsi que des événements d'hybridation ancestrale. La combinaison de ces différents éléments a probablement facilité leur diversification en générant les variations génomiques nécessaires à l'action de la sélection naturelle. J’identifie également des gènes ayant subi des pressions de sélection différentielles en lien avec des divergences écologiques, suggérant donc un processus d’évolution parallèle impliquée dans la diversification des poissons-clowns. Finalement, je me suis concentrée sur les mécanismes liés à l'évolution d'un clade de poisson- clown - le clade “skunk”. Je démontre qu’un flux de gènes modéré s'est produit tout au long de la diversification de ce groupe, et - bien que son amplitude ait diminué avec le temps - il persiste encore en sympatrie. De plus, j’identifie des régions candidates d'introgression entre espèces qui ont probablement joué un rôle dans la diversification du complexe. Dans l'ensemble, mon travail fournit les premières informations concernant les mécanismes génomiques impliqués dans la radiation adaptative des poissons-clowns