Morten T. Limborg

Effects of Crossovers Between Homeologs on Inheritance and Population Genomics in Polyploid-Derived Salmonid Fishes

A whole genome duplication occurred in the ancestor of all salmonid fishes some 50-100 million years ago. Early inheritance studies with allozymes indicated that loci in the salmonid genome are inherited disomically in females. However, some pairs of duplicated loci showed patterns of inheritance in males indicating pairing and recombination between homeologous chromosomes. Nearly 20% of loci in the salmonid genome are duplicated and share the same alleles (isoloci), apparently due to homeologous recombination. Half-tetrad analysis revealed that isoloci tend to be telomeric. These results suggested that residual tetrasomic inheritance of isoloci results from homeologous recombination near chromosome ends and that continued disomic inheritance resulted from homologous pairing of centromeric regions. Many current genetic maps of salmonids are based on single nucleotide polymorphisms and microsatellites that are no longer duplicated. Therefore, long sections of chromosomes on these maps are poorly represented, especially telomeric regions. In addition, preferential multivalent pairing of homeologs from the same species in F1 hybrids results in an excess of nonparental gametes (so-called pseudolinkage). We consider how not including duplicated loci has affected our understanding of population and evolutionary genetics of salmonids, and we discuss how incorporating these loci will benefit our understanding of population genomics.

Signatures of natural selection among lineages and habitats in Oncorhynchus mykiss.

Recent advances in molecular interrogation techniques now allow unprecedented genomic inference about the role of adaptive genetic divergence in wild populations. We used high-throughput genotyping to screen a genome-wide panel of 276 single nucleotide polymorphisms (SNPs) for the economically and culturally important salmonid Oncorhynchus mykiss. Samples included 805 individuals from 11 anadromous and resident populations from the northwestern United States and British Columbia, and represented two major lineages including paired populations of each life history within single drainages of each lineage. Overall patterns of variation affirmed clear distinctions between lineages and in most instances, isolation by distance within them. Evidence for divergent selection at eight candidate loci included significant landscape correlations, particularly with temperature. High diversity of two nonsynonymous mutations within the peptide-binding region of the major histocompatibility complex (MHC) class II (DAB) gene provided signatures of balancing selection. Weak signals for potential selection between sympatric resident and anadromous populations were revealed from genome scans and allele frequency comparisons. Our results suggest an important adaptive role for immune-related functions and present a large genomic resource for future studies

An integrated linkage map reveals candidate genes underlying adaptive variation in Chinook salmon (Oncorhynchus tshawytscha)

Salmonids are an important cultural and ecological resource exhibiting near worldwide distribution between their native and introduced range. Previous research has generated linkage maps and genomic resources for several species as well as genome assemblies for two species. We first leveraged improvements in mapping and genotyping methods to create a dense linkage map for Chinook salmon Oncorhynchus tshawytscha by assembling family data from different sources. We successfully mapped 14 620 SNP loci including 2336 paralogs in subtelomeric regions. This improved map was then used as a foundation to integrate genomic resources for gene annotation and population genomic analyses. We anchored a total of 286 scaffolds from the Atlantic salmon genome to the linkage map to provide a framework for the placement 11 728 Chinook salmon ESTs. Previously identified thermotolerance QTL were found to colocalize with several candidate genes including HSP70, a gene known to be involved in thermal response, as well as its inhibitor. Multiple regions of the genome with elevated divergence between populations were also identified, and annotation of ESTs in these regions identified candidate genes for fitness related traits such as stress response, growth and behaviour. Collectively, these results demonstrate the utility of combining genomic resources with linkage maps to enhance evolutionary inferences.

Identification of Multiple QTL Hotspots in Sockeye Salmon ( Oncorhynchus nerka ) Using Genotyping-by-Sequencing and a Dense Linkage Map

Understanding the genetic architecture of phenotypic traits can provide important information about the mechanisms and genomic regions involved in local adaptation and speciation. Here, we used genotyping-by-sequencing and a combination of previously published and newly generated data to construct sex-specific linkage maps for sockeye salmon (Oncorhynchus nerka). We then used the denser female linkage map to conduct quantitative trait locus (QTL) analysis for 4 phenotypic traits in 3 families. The female linkage map consisted of 6322 loci distributed across 29 linkage groups and was 4082 cM long, and the male map contained 2179 loci found on 28 linkage groups and was 2291 cM long. We found 26 QTL: 6 for thermotolerance, 5 for length, 9 for weight, and 6 for condition factor. QTL were distributed nonrandomly across the genome and were often found in hotspots containing multiple QTL for a variety of phenotypic traits. These hotspots may represent adaptively important regions and are excellent candidates for future research. Comparing our results with studies in other salmonids revealed several regions with overlapping QTL for the same phenotypic trait, indicating these regions may be adaptively important across multiple species. Altogether, our study demonstrates the utility of genomic data for investigating the genetic basis of important phenotypic traits. Additionally, the linkage map created here will enable future research on the genetic basis of phenotypic traits in salmon.

Sorting duplicated loci disentangles complexities of polyploid genomes masked by genotyping by sequencing

Many plants and animals of polyploid origin are currently enjoying a genomics explosion enabled by modern sequencing and genotyping technologies. However, routine filtering of duplicated loci in most studies using genotyping by sequencing introduces an unacceptable, but often overlooked, bias when detecting selection. Retained duplicates from ancient whole‐genome duplications (WGDs) may be found throughout genomes, whereas retained duplicates from recent WGDs are concentrated at distal ends of some chromosome arms. Additionally, segmental duplicates can be found at distal ends or nearly anywhere in a genome. Evidence shows that these duplications facilitate adaptation through one of two pathways: neo‐functionalization or increased gene expression. Filtering duplicates removes distal ends of some chromosomes, and distal ends are especially known to harbour adaptively important genes. Thus, filtering of duplicated loci impoverishes the interpretation of genomic data as signals from contiguous duplicated genes are ignored. We review existing strategies to genotype and map duplicated loci; we focus in detail on an overlooked strategy of using gynogenetic haploids (1N) as a part of new genotyping by sequencing studies. We provide guidelines on how to use this haploid strategy for studies on polyploid‐origin vertebrates including how it can be used to screen duplicated loci in natural populations. We conclude by discussing areas of research that will benefit from better inclusion of polyploid loci; we particularly stress the sometimes overlooked fact that basing genomic studies on dense maps provides value added in the form of locating and annotating outlier loci or colocating outliers into islands of divergence.

The effects of Medieval dams on genetic divergence and demographic history in brown trout populations

BackgroundHabitat fragmentation has accelerated within the last century, but may have been ongoing over longer time scales. We analyzed the timing and genetic consequences of fragmentation in two isolated lake-dwelling brown trout populations. They are from the same river system (the Gudenå River, Denmark) and have been isolated from downstream anadromous trout by dams established ca. 600–800 years ago. For reference, we included ten other anadromous populations and two hatchery strains. Based on analysis of 44 microsatellite loci we investigated if the lake populations have been naturally genetically differentiated from anadromous trout for thousands of years, or have diverged recently due to the establishment of dams.ResultsDivergence time estimates were based on 1) Approximate Bayesian Computation and 2) a coalescent-based isolation-with-gene-flow model. Both methods suggested divergence times ca. 600–800 years bp, providing strong evidence for establishment of dams in the Medieval as the factor causing divergence. Bayesian cluster analysis showed influence of stocked trout in several reference populations, but not in the focal lake and anadromous populations. Estimates of effective population size using a linkage disequilibrium method ranged from 244 to > 1,000 in all but one anadromous population, but were lower (153 and 252) in the lake populations.ConclusionsWe show that genetic divergence of lake-dwelling trout in two Danish lakes reflects establishment of water mills and impassable dams ca. 600–800 years ago rather than a natural genetic population structure. Although effective population sizes of the two lake populations are not critically low they may ultimately limit response to selection and thereby future adaptation. Our results demonstrate that populations may have been affected by anthropogenic disturbance over longer time scales than normally assumed.

Parallel signatures of selection in temporally isolated lineages of pink salmon

Studying the effect of similar environments on diverse genetic backgrounds has long been a goal of evolutionary biologists with studies typically relying on experimental approaches. Pink salmon, a highly abundant and widely ranging salmonid, provide a naturally occurring opportunity to study the effects of similar environments on divergent genetic backgrounds due to a strict two‐year semelparous life history. The species is composed of two reproductively isolated lineages with overlapping ranges that share the same spawning and rearing environments in alternate years. We used restriction‐site‐associated DNA (RAD) sequencing to discover and genotype approximately 8000 SNP loci in three population pairs of even‐ and odd‐year pink salmon along a latitudinal gradient in North America. We found greater differentiation within the odd‐year than within the even‐year lineage and greater differentiation in the southern pair from Puget Sound than in the northern Alaskan population pairs. We identified 15 SNPs reflecting signatures of parallel selection using both a differentiation‐based method (BAYESCAN) and an environmental correlation method (BAYENV). These SNPs represent genomic regions that may be particularly informative in understanding adaptive evolution in pink salmon and exploring how differing genetic backgrounds within a species respond to selection from the same natural environment.

Genomic islands of divergence linked to ecotypic variation in sockeye salmon

Regions of the genome displaying elevated differentiation (genomic islands of divergence) are thought to play an important role in local adaptation, especially in populations experiencing high gene flow. However, the characteristics of these islands as well as the functional significance of genes located within them remain largely unknown. Here, we used data from thousands of SNPs aligned to a linkage map to investigate genomic islands of divergence in three ecotypes of sockeye salmon (Oncorhynchus nerka) from a single drainage in southwestern Alaska. We found ten islands displaying high differentiation among ecotypes. Conversely, neutral structure observed throughout the rest of the genome was low and not partitioned by ecotype. One island on linkage group So13 was particularly large and contained six SNPs with FST > 0.14 (average FST of neutral SNPs = 0.01). Functional annotation revealed that the peak of this island contained a nonsynonymous mutation in a gene involved in growth in other species (TULP4). The islands that we discovered were relatively small (80–402 Kb), loci found in islands did not show reduced levels of diversity, and loci in islands displayed slightly elevated linkage disequilibrium. These attributes suggest that the islands discovered here were likely generated by divergence hitchhiking; however, we cannot rule out the possibility that other mechanisms may have produced them. Our results suggest that islands of divergence serve an important role in local adaptation with gene flow and represent a significant advance towards understanding the genetic basis of ecotypic differentiation.

Linkage mapping reveals strong chiasma interference in Sockeye salmon: Implications for interpreting genomic data

Meiotic recombination is fundamental for generating new genetic variation and for securing proper disjunction. Further, recombination plays an essential role during the rediploidization process of polyploid-origin genomes because crossovers between pairs of homeologous chromosomes retain duplicated regions. A better understanding of how recombination affects genome evolution is crucial for interpreting genomic data; unfortunately, current knowledge mainly originates from a few model species. Salmonid fishes provide a valuable system for studying the effects of recombination in nonmodel species. Salmonid females generally produce thousands of embryos, providing large families for conducting inheritance studies. Further, salmonid genomes are currently rediploidizing after a whole genome duplication and can serve as models for studying the role of homeologous crossovers on genome evolution. Here, we present a detailed interrogation of recombination patterns in sockeye salmon (Oncorhynchus nerka). First, we use RAD sequencing of haploid and diploid gynogenetic families to construct a dense linkage map that includes paralogous loci and location of centromeres. We find a nonrandom distribution of paralogs that mainly cluster in extended regions distally located on 11 different chromosomes, consistent with ongoing homeologous recombination in these regions. We also estimate the strength of interference across each chromosome; results reveal strong interference and crossovers are mostly limited to one per arm. Interference was further shown to continue across centromeres, but metacentric chromosomes generally had at least one crossover on each arm. We discuss the relevance of these findings for both mapping and population genomic studies.

Recombination patterns reveal information about centromere location on linkage maps

Linkage mapping is often used to identify genes associated with phenotypic traits and for aiding genome assemblies. Still, many emerging maps do not locate centromeres – an essential component of the genomic landscape. Here, we demonstrate that for genomes with strong chiasma interference, approximate centromere placement is possible by phasing the same data used to generate linkage maps. Assuming one obligate crossover per chromosome arm, information about centromere location can be revealed by tracking the accumulated recombination frequency along linkage groups, similar to half‐tetrad analyses. We validate the method on a linkage map for sockeye salmon (Oncorhynchus nerka) with known centromeric regions. Further tests suggest that the method will work well in other salmonids and other eukaryotes. However, the method performed weakly when applied to a male linkage map (rainbow trout; O. mykiss) characterized by low and unevenly distributed recombination – a general feature of male meiosis in many species. Further, a high frequency of double crossovers along chromosome arms in barley reduced resolution for locating centromeric regions on most linkage groups. Despite these limitations, our method should work well for high‐density maps in species with strong recombination interference and will enrich many existing and future mapping resources.