Strukturell variasjon som påvirker genetisk miljøtilpasning i laksefisk

Structural variations (SVs), e.g. deletions, insertions, inversions and duplications of sequences, are a major source of genomic variation affecting more base pairs in the genome than single nucleotide polymorphisms (SNPs). Despite their increasingly recognised importance in adaptive evolution and species diversification, SVs are vastly understudied in most species. Long-read sequencing, together with recently developed bioinformatic tools, have provided step-change improvements in the precision and recall of SV detection and allow us to increase the detected SVs manyfold across the species range. In addition, long-reads represent a major shift in our ability to build continuous genome assemblies as fundamental resources for most genome wide studies. The work in this thesis utilises long-read data to generate multiple genome sequences for the two salmonid species Atlantic salmon (Salmo salar) and lake whitefish (Coregonus clupeaformis). We present the first pan-genome for Atlantic salmon, comprising 11 long-read-based assemblies across the species range. Among these, the highest quality genome has 2.55 Gbp assembled into chromosome sequences, 259 Mbp more sequence than in the previous Atlantic salmon reference genome. The genome has a highly improved continuity with contig N50 increasing from 58 kbp to 28.06 Mbp (484-fold). The detection of SVs in these 11 individuals, revealed 1,061,452 SVs, with an average of ~77.4 Mbp of sequence differing per sample. The Atlantic salmon has adapted to different river environment across a large geographical distribution. To investigate genomic variation underlying these adaptations, we associated SVs and environmental data in a dataset of 366 short-read samples genotyped using genome graph analyses. These analyses highlighted multiple SVs contributing to environmental adaptations, including an 18 kbp deletion encompassing a polymorphic segmental duplication of three genes associated with annual precipitation. Next, we use the Atlantic salmon pan-genome to study the emergence of supergenes. Because supergenes can be maintained over millions of years by balancing selection and typically exhibit strong recombination suppression, their underlying functional variants and how they are formed are largely unknown. Inversions are type of rearrangement commonly associated with supergenes, and by directly comparing multiple highly continuous genome assemblies we were able to detect a number of large inversions in Atlantic salmon. A 3 Mb inversion, estimated to be ~15,000-year-old, and segregating in North American populations, displayed supergene signatures with adaptive variation captured within the standard arrangement of the inversion, as well as other adaptive variation accumulating after the inversion occurred. Characterization of other inversions with matched repeat structures at the breakpoints did not show any supergene signatures, suggesting that shared breakpoint repeats may obstruct the supergene formation. Lastly, we created long-read based genome assemblies for sympatric species pairs (Dwarf and Normal) belonging to lake whitefish (Coregonus clupeaformis). The species pairs offer a suitable model system for studying genomic patterns of differentiation and in particular the role of SVs in speciation. By combining long-reads, direct assembly, and short-read methods we detect 89,909 high-confidence SVs in the species pair across two lakes, covering five times more sequence in the genome compared to SNPs. In the study, we highlight shared outliers of differentiation between the lakes, indicating that they contribute to speciation. Interestingly, we find that more than 70% of SVs differentiating between the Normal and Dwarf species pairs of lake whitefish are overlapping transposable elements. This work demonstrates that SVs may play an important role for the differentiation and speciation of sympatric species pairs in lake whitefish.Strukturell variasjon (SVer), for eksempel delesjoner, insersjoner, inversjoner og duplikasjoner av sekvens, er en viktig kilde til genomisk variasjon som samplet sett påvirker flere basepar i genomet enn punktmutasjoner (SNPs). Til tross for en økende annerkjennelse for at SVer spiller en viktig rolle i genetisk tilpassing til ulikt miljø og artsdannelse har denne typen variasjon vært lite studert i mange arter. Ny DNA-sekvenseringsteknologi med lengre leselengder (long-read sequencing), samt utvikling av nye bioinformatiske verktøy, har ført til drastiske forbedringer i deteksjonen av SVer. ‘Long-read’ sekvensering gjør det også mulig å lage mer komplette og sammenhengende genomsekvenser enn tidligere. I denne avhandlingen benytter vi oss av ‘long-read’ data til å lage flere genomsekvenser av høy kvalitet for to ulike laksefiskarter: Atlanterhavslaks (Salmo salar) og en Nordamerikansk type sik ‘lake whitefish’ (Coregonus clupeaformis). Her rapporterer vi det første pan-genomet for Atlanterhavslaks. Det består av 11 assemblier basert på ‘long- read’ sekvensering av individer fra fire ulike fylogeografiske grupper av villaks. Assembliet av høyest kvalitet inkluderer 2,55 Gbp sekvens i kromosomer, 259 Mbp mer enn det forrige referansegenomet til Atlanterhavslaks. I tillegg ble andelen sammenhengende sekvens, målt som contig N50, økt fra 58 kbp til 28,06 Mbp (484 ganger høyere). Vi fant 1.061.452 SVer på tvers av de 11 individene med ~77,4 Mbp gjennomsnittlig sekvensforskjell per prøve. Atlanterhavslaksen har over tid tilpasset miljøet i ulike elver. For å studere underliggende genetisk variasjon for denne tilpasningen assosierte vi SVer med ulike miljøvariabler i et datasett bestående av 366 ‘short-read’ sekvenserte prøver ved bruk av en genom-graf. Ved hjelp av disse analysene fant vi flere SVer som bidrar til miljøtilpasning, blant annet en 18 kbp lang delesjon som inneholder tre gener assosiert med mengden nedbør i området. Vi brukte så pan-genomet for Atlanterhavsaks til å studere dannelsen av ‘supergener’. Supergener er en sammenkobling av genetisk variasjon i koblingsulikevekt som for eksempel kan oppstå ved hjelp av store inversjoner. Her utnyttet vi 11 genomassemblier til å identifisere og karakterisere en rekke store inversjoner i Atlanterhavslaks. En av inversjonene på 3 Mbp, estimert til å være ~15.000 år gammel, viste signaturer for utvikling som supergen. For de andre inversjonene som var flankert av repetert DNA fant vi ikke karakteristiske trekk på supergener, noe som tyder på at det repetitive DNA forhindrer en dannelse av supergener. Til slutt lagde vi genomsekvenser for ulike former (‘Normal’ og ‘Dwarf’) av ‘lake whitefish’ (Coregonus clupeaformis) som lever i de samme innsjøene i Nord-Amerika. Genomsekvensene muliggjør studier av genomiske mekanismene bak artsdannelse i denne laksefisken. Ved å kombinere ‘long-read’ data, direkte sammenlikning av assemblier, og ‘short-read’ data fant vi 89,909 SVer som skilte de to formene av ‘lake whitefish’ i to innsjøer. SVene omfatter mer enn fem ganger flere basepar i genomet sammenlignet med SNPs. I studiet fant vi flere SVer med avvikende forekomst (‘outliers’) i de to formene av ‘lake whitefish’, noe som indikerer at disse SVene bidrar til artsdannelse. Videre fant vi at 70 % av SVene overlappet en form av repetert DNA kalt transposable elementer. Dette arbeidet understreker at SVer kan spille en viktig rolle for artsdannelse i ’lake whitefish’

Genome-wide signatures of complex introgression and adaptive evolution in the big cats.

The great cats of the genus Panthera comprise a recent radiation whose evolutionary history is poorly understood. Their rapid diversification poses challenges to resolving their phylogeny while offering opportunities to investigate the historical dynamics of adaptive divergence. We report the sequence, de novo assembly, and annotation of the jaguar (Panthera onca) genome, a novel genome sequence for the leopard (Panthera pardus), and comparative analyses encompassing all living Panthera species. Demographic reconstructions indicated that all of these species have experienced variable episodes of population decline during the Pleistocene, ultimately leading to small effective sizes in present-day genomes. We observed pervasive genealogical discordance across Panthera genomes, caused by both incomplete lineage sorting and complex patterns of historical interspecific hybridization. We identified multiple signatures of species-specific positive selection, affecting genes involved in craniofacial and limb development, protein metabolism, hypoxia, reproduction, pigmentation, and sensory perception. There was remarkable concordance in pathways enriched in genomic segments implicated in interspecies introgression and in positive selection, suggesting that these processes were connected. We tested this hypothesis by developing exome capture probes targeting ~19,000 Panthera genes and applying them to 30 wild-caught jaguars. We found at least two genes (DOCK3 and COL4A5, both related to optic nerve development) bearing significant signatures of interspecies introgression and within-species positive selection. These findings indicate that post-speciation admixture has contributed genetic material that facilitated the adaptive evolution of big cat lineages

Comparative pan-genome analysis of Piscirickettsia salmonis reveals genomic divergences within genogroups

Indexación: Scopus.Piscirickettsia salmonis is the etiological agent of salmonid rickettsial septicemia, a disease that seriously affects the salmonid industry. Despite efforts to genomically characterize P. salmonis, functional information on the life cycle, pathogenesis mechanisms, diagnosis, treatment, and control of this fish pathogen remain lacking. To address this knowledge gap, the present study conducted an in silico pan-genome analysis of 19 P. salmonis strains from distinct geographic locations and genogroups. Results revealed an expected open pan-genome of 3,463 genes and a core-genome of 1,732 genes. Two marked genogroups were identified, as confirmed by phylogenetic and phylogenomic relationships to the LF-89 and EM-90 reference strains, as well as by assessments of genomic structures. Different structural configurations were found for the six identified copies of the ribosomal operon in the P. salmonis genome, indicating translocation throughout the genetic material. Chromosomal divergences in genomic localization and quantity of genetic cassettes were also found for the Dot/Icm type IVB secretion system. To determine divergences between core-genomes, additional pan-genome descriptions were compiled for the so-termed LF and EM genogroups. Open pan-genomes composed of 2,924 and 2,778 genes and core-genomes composed of 2,170 and 2,228 genes were respectively found for the LF and EM genogroups. The core-genomes were functionally annotated using the Gene Ontology, KEGG, and Virulence Factor databases, revealing the presence of several shared groups of genes related to basic function of intracellular survival and bacterial pathogenesis. Additionally, the specific pan-genomes for the LF and EM genogroups were defined, resulting in the identification of 148 and 273 exclusive proteins, respectively. Notably, specific virulence factors linked to adherence, colonization, invasion factors, and endotoxins were established. The obtained data suggest that these genes could be directly associated with inter-genogroup differences in pathogenesis and host-pathogen interactions, information that could be useful in designing novel strategies for diagnosing and controlling P. salmonis infection. © 2017 Nourdin-Galindo, Sánchez, Molina, Espinoza-Rojas, Oliver, Ruiz, Vargas-Chacoff, Cárcamo, Figueroa, Mancilla, Maracaja-Coutinho and Yañez.https://www.frontiersin.org/articles/10.3389/fcimb.2017.00459/ful

Efficient Inference of Recent and Ancestral Recombination within Bacterial Populations

Prokaryotic evolution is affected by horizontal transfer of genetic material through recombination. Inference of an evolutionary tree of bacteria thus relies on accurate identification of the population genetic structure and recombination-derived mosaicism. Rapidly growing databases represent a challenge for computational methods to detect recombinations in bacterial genomes. We introduce a novel algorithm called fastGEAR which identifies lineages in diverse microbial alignments, and recombinations between them and from external origins. The algorithm detects both recent recombinations (affecting a few isolates) and ancestral recombinations between detected lineages (affecting entire lineages), thus providing insight into recombinations affecting deep branches of the phylogenetic tree. In simulations, fastGEAR had comparable power to detect recent recombinations and outstanding power to detect the ancestral ones, compared with state-of-the-art methods, often with a fraction of computational cost. We demonstrate the utility of the method by analyzing a collection of 616 whole-genomes of a recombinogenic pathogen Streptococcus pneumoniae, for which the method provided a high-resolution view of recombination across the genome. We examined in detail the penicillin-binding genes across the Streptococcus genus, demonstrating previously undetected genetic exchanges between different species at these three loci. Hence, fastGEAR can be readily applied to investigate mosaicism in bacterial genes across multiple species. Finally, fastGEAR correctly identified many known recombination hotspots and pointed to potential new ones. Matlab code and Linux/Windows executables are available at https://users.ics.aalto.fi/similar to pemartti/fastGEAR/ (last accessed February 6, 2017).Peer reviewe

Species-level functional profiling of metagenomes and metatranscriptomes.

Functional profiles of microbial communities are typically generated using comprehensive metagenomic or metatranscriptomic sequence read searches, which are time-consuming, prone to spurious mapping, and often limited to community-level quantification. We developed HUMAnN2, a tiered search strategy that enables fast, accurate, and species-resolved functional profiling of host-associated and environmental communities. HUMAnN2 identifies a community's known species, aligns reads to their pangenomes, performs translated search on unclassified reads, and finally quantifies gene families and pathways. Relative to pure translated search, HUMAnN2 is faster and produces more accurate gene family profiles. We applied HUMAnN2 to study clinal variation in marine metabolism, ecological contribution patterns among human microbiome pathways, variation in species' genomic versus transcriptional contributions, and strain profiling. Further, we introduce 'contributional diversity' to explain patterns of ecological assembly across different microbial community types

Origin matters: Using a local reference genome improves measures in population genomics.

Genome sequencing enables answering fundamental questions about the genetic basis of adaptation, population structure and epigenetic mechanisms. Yet, we usually need a suitable reference genome for mapping population-level resequencing data. In some model systems, multiple reference genomes are available, giving the challenging task of determining which reference genome best suits the data. Here, we compared the use of two different reference genomes for the three-spined stickleback (Gasterosteus aculeatus), one novel genome derived from a European gynogenetic individual and the published reference genome of a North American individual. Specifically, we investigated the impact of using a local reference versus one generated from a distinct lineage on several common population genomics analyses. Through mapping genome resequencing data of 60 sticklebacks from across Europe and North America, we demonstrate that genetic distance among samples and the reference genomes impacts downstream analyses. Using a local reference genome increased mapping efficiency and genotyping accuracy, effectively retaining more and better data. Despite comparable distributions of the metrics generated across the genome using SNP data (i.e. π, Tajima's D and FST ), window-based statistics using different references resulted in different outlier genes and enriched gene functions. A marker-based analysis of DNA methylation distributions had a comparably high overlap in outlier genes and functions, yet with distinct differences depending on the reference genome. Overall, our results highlight how using a local reference genome decreases reference bias to increase confidence in downstream analyses of the data. Such results have significant implications in all reference-genome-based population genomic analyses