David R. Minkley

The Atlantic salmon genome provides insights into rediploidization

The whole-genome duplication 80 million years ago of the common ancestor of salmonids (salmonid-specific fourth vertebrate whole-genome duplication, Ss4R) provides unique opportunities to learn about the evolutionary fate of a duplicated vertebrate genome in 70 extant lineages. Here we present a high-quality genome assembly for Atlantic salmon (Salmo salar), and show that large genomic reorganizations, coinciding with bursts of transposon-mediated repeat expansions, were crucial for the post-Ss4R rediploidization process. Comparisons of duplicate gene expression patterns across a wide range of tissues with orthologous genes from a pre-Ss4R outgroup unexpectedly demonstrate far more instances of neofunctionalization than subfunctionalization. Surprisingly, we find that genes that were retained as duplicates after the teleost-specific whole-genome duplication 320 million years ago were not more likely to be retained after the Ss4R, and that the duplicate retention was not influenced to a great extent by the nature of the predicted protein interactions of the gene products. Finally, we demonstrate that the Atlantic salmon assembly can serve as a reference sequence for the study of other salmonids for a range of purposes.

The Genome and Linkage Map of the Northern Pike (Esox lucius): Conserved Synteny Revealed between the Salmonid Sister Group and the Neoteleostei

The northern pike is the most frequently studied member of the Esociformes, the closest order to the diverse and economically important Salmoniformes. The ancestor of all salmonids purportedly experienced a whole-genome duplication (WGD) event, making salmonid species ideal for studying the early impacts of genome duplication while complicating their use in wider analyses of teleost evolution. Studies suggest that the Esociformes diverged from the salmonid lineage prior to the WGD, supporting the use of northern pike as a pre-duplication outgroup. Here we present the first genome assembly, reference transcriptome and linkage map for northern pike, and evaluate the suitability of this species to provide a representative pre-duplication genome for future studies of salmonid and teleost evolution. The northern pike genome sequence is composed of 94,267 contigs (N50 = 16,909 bp) contained in 5,688 scaffolds (N50 = 700,535 bp); the total scaffolded genome size is 878 million bases. Multiple lines of evidence suggest that over 96% of the protein-coding genome is present in the genome assembly. The reference transcriptome was constructed from 13 tissues and contains 38,696 transcripts, which are accompanied by normalized expression data in all tissues. Gene-prediction analysis produced a total of 19,601 northern pike-specific gene models. The first-generation linkage map identifies 25 linkage groups, in agreement with northern pikes diploid karyotype of 2N = 50, and facilitates the placement of 46% of assembled bases onto linkage groups. Analyses reveal a high degree of conserved synteny between northern pike and other model teleost genomes. While conservation of gene order is limited to smaller syntenic blocks, the wider conservation of genome organization implies the northern pike exhibits a suitable approximation of a non-duplicated Protacanthopterygiian genome. This dataset will facilitate future studies of esocid biology and empower ongoing examinations of the Atlantic salmon and rainbow trout genomes by facilitating their comparison with other major teleost groups.

Genomics of sablefish (Anoplopoma fimbria): expressed genes, mitochondrial phylogeny, linkage map and identification of a putative sex gene

BackgroundThe sablefish (order: Scorpaeniformes) is an economically important species in commercial fisheries of the North Pacific and an emerging species in aquaculture. Aside from a handful of sequences in NCBI and a few published microsatellite markers, little is known about the genetics of this species. The development of genetic tools, including polymorphic markers and a linkage map will allow for the successful development of future broodstock and mapping of phenotypes of interest. The significant sexual dimorphism between females and males makes a genetic test for early identification of sex desirable.ResultsA full mitochondrial genome is presented and the resulting phylogenetic analysis verifies the placement of the sablefish within the Scorpaeniformes. Nearly 35,000 assembled transcript sequences are used to identify genes and obtain polymorphic SNP and microsatellite markers. 360 transcribed polymorphic loci from two sablefish families produce a map of 24 linkage groups. The sex phenotype maps to sablefish LG14 of the male map. We show significant conserved synteny and conservation of gene-order between the threespine stickleback Gasterosteus aculeatus and sablefish. An additional 1843 polymorphic SNP markers are identified through next-generation sequencing techniques. Sex-specific markers and sequence insertions are identified immediately upstream of the gene gonadal-soma derived factor (gsdf), the master sex determinant locus in the medaka species Oryzias luzonensis.ConclusionsThe first genomic resources for sablefish provide a foundation for further studies. Over 35,000 transcripts are presented, and the genetic map represents, as far as we can determine, the first linkage map for a member of the Scorpaeniformes. The observed level of conserved synteny and comparative mapping will allow the use of the stickleback genome in future genetic studies on sablefish and other related fish, particularly as a guide to whole-genome assembly. The identification of sex-specific insertions immediately upstream of a known master sex determinant implicates gsdf as an excellent candidate for the master sex determinant for sablefish.

GO Trimming: Systematically reducing redundancy in large Gene Ontology datasets

BackgroundThe increased accessibility of gene expression tools has enabled a wide variety of experiments utilizing transcriptomic analyses. As these tools increase in prevalence, the need for improved standardization in processing and presentation of data increases, as does the need to guard against interpretation bias. Gene Ontology (GO) analysis is a powerful method of interpreting and summarizing biological functions. However, while there are many tools available to investigate GO enrichment, there remains a need for methods that directly remove redundant terms from enriched GO lists that often provide little, if any, additional information.FindingsHere we present a simple yet novel method called GO Trimming that utilizes an algorithm designed to reduce redundancy in lists of enriched GO categories. Depending on the needs of the user, this method can be performed with variable stringency. In the example presented here, an initial list of 90 terms was reduced to 54, eliminating 36 largely redundant terms. We also compare this method to existing methods and find that GO Trimming, while simple, performs well to eliminate redundant terms in a large dataset throughout the depth of the GO hierarchy.ConclusionsThe GO Trimming method provides an alternative to other procedures, some of which involve removing large numbers of terms prior to enrichment analysis. This method should free up the researcher from analyzing overly large, redundant lists, and instead enable the concise presentation of manageable, informative GO lists. The implementation of this tool is freely available at: http://lucy.ceh.uvic.ca/go_trimming/cbr_go_trimming.py

Sex-specific expression, synthesis and localization of aromatase regulators in one-year-old Atlantic salmon ovaries and testes

Transcripts for dax1, foxl2, mis and sf1 are co-expressed in the somatic companion cells of teleost germ cells. These regulatory factors function, in part, to modulate the transcription of aromatase, particularly cyp19a, the terminal enzyme of estrogen biosynthesis. At least two separate aromatase loci exist in teleost fish that encode distinct isoforms. The activity of two forms, cyp19a and cyp19b1, is predominantly associated with the ovary and the brain, respectively. We isolated sequences that compose the proximal promoters of cyp19a, cyp19b1 and foxl2a, to identify potential transcription factor binding motifs to define sex-specific regulatory profiles for each gene. We also provide evidence for the translation and immunological localization of DAX-1, FOXL2 and MIS to the endoplasmic reticulum and accumulation within secretory vesicles of the salmon oocyte. We found no evidence for the expression of CYP19A or CYP19B1 in the oocyte at the one-year-old stage. However, synthesis of both aromatases was localized to testicular germ and soma cells at this early stage of development. Production of these regulatory factors in the germ cells may serve to modulate the transcription and activity of endogenous aromatase and/or contribute to the differentiation of the neighbouring companion cells through secretory signaling.

Chinook salmon (Oncorhynchus tshawytscha) genome and transcriptome.

When unifying genomic resources among studies and comparing data between species, there is often no better resource than a genome sequence. Having a reference genome for the Chinook salmon (Oncorhynchus tshawytscha) will enable the extensive genomic resources available for Pacific salmon, Atlantic salmon, and rainbow trout to be leveraged when asking questions related to the Chinook salmon. The Chinook salmon’s wide distribution, long cultural impact, evolutionary history, substantial hatchery production, and recent wild-population decline make it an important research species. In this study, we sequenced and assembled the genome of a Chilliwack River Hatchery female Chinook salmon (gynogenetic and homozygous at all loci). With a reference genome sequence, new questions can be asked about the nature of this species, and its role in a rapidly changing world.

The Arctic charr (Salvelinus alpinus) genome and transcriptome assembly

Arctic charr have a circumpolar distribution, persevere under extreme environmental conditions, and reach ages unknown to most other salmonids. The Salvelinus genus is primarily composed of species with genomes that are structured more like the ancestral salmonid genome than most Oncorhynchus and Salmo species of sister genera. It is thought that this aspect of the genome may be important for local adaptation (due to increased recombination) and anadromy (the migration of fish from saltwater to freshwater). In this study, we describe the generation of a new genetic map, the sequencing and assembly of the Arctic charr genome (GenBank accession: GCF_002910315.2) using the newly created genetic map and a previous genetic map, and present several analyses of the Arctic charr genes and genome assembly. The newly generated genetic map consists of 8,574 unique genetic markers and is similar to previous genetic maps with the exception of three major structural differences. The N50, identified BUSCOs, repetitive DNA content, and total size of the Arctic charr assembled genome are all comparable to other assembled salmonid genomes. An analysis to identify orthologous genes revealed that a large number of orthologs could be identified between salmonids and many appear to have highly conserved gene expression profiles between species. Comparing orthologous gene expression profiles may give us a better insight into which genes are more likely to influence species specific phenotypes.

A 200K SNP chip reveals a novel Pacific salmon louse genotype linked to differential efficacy of emamectin benzoate

Antiparasitic drugs such as emamectin benzoate (EMB) are relied upon to reduce the parasite load, particularly of the sea louse Lepeophtheirus salmonis, on farmed salmon. The decline in EMB treatment efficacy for this purpose is an important issue for salmon producers around the world, and particularly for those in the Atlantic Ocean where widespread EMB tolerance in sea lice is recognized as a significant problem. Salmon farms in the Northeast Pacific Ocean have not historically experienced the same issues with treatment efficacy, possibly due to the relatively large population of endemic salmonid hosts that serve to both redistribute surviving lice and dilute populations potentially under selection by introducing naïve lice to farms. Frequent migration of lice among farmed and wild hosts should limit the effect of farm-specific selection pressures on changes to the overall allele frequencies of sea lice in the Pacific Ocean. A previous study using microsatellites examined L. salmonis oncorhynchi from 10 Pacific locations from wild and farmed hosts and found no population structure. Recently however, a farm population of sea lice was detected where EMB bioassay exposure tolerance was abnormally elevated. In response, we have developed a Pacific louse draft genome that complements the previously-released Atlantic louse sequence. These genomes were combined with whole-genome re-sequencing data to design a highly sensitive 201,279 marker SNP array applicable for both subspecies (90,827 validated Pacific loci; 153,569 validated Atlantic loci). Notably, kmer spectrum analysis of the re-sequenced samples indicated that Pacific lice exhibit a large within-individual heterozygosity rate (average of 1 in every 72 bases) that is markedly higher than that of Atlantic individuals (1 in every 173 bases). The SNP chip was used to produce a high-density map for Atlantic sea louse linkage group 5 that was previously shown to be associated with EMB tolerance in Atlantic lice. Additionally, 478 Pacific louse samples from farmed and wild hosts obtained between 2005 and 2014 were also genotyped on the array. Clustering analysis allowed us to detect the apparent emergence of an otherwise rare genotype at a high frequency among the lice collected from two farms in 2013 that had reported elevated EMB tolerance. This genotype was not observed in louse samples collected from the same farm in 2010, nor in any lice sampled from other locations prior to 2013. However, this genotype was detected at low frequencies in louse samples from farms in two locations reporting elevated EMB tolerance in 2014. These results suggest that a rare genotype present in Pacific lice may be locally expanded in farms after EMB treatment. Supporting this hypothesis, 437 SNPs associated with this genotype were found to be in a region of linkage group 5 that overlaps the region associated with EMB resistance in Atlantic lice. Finally, five of the top diagnostic SNPs within this region were used to screen lice that had been subjected to an EMB survival assay, revealing a significant association between these SNPs and EMB treatment outcome. To our knowledge this work is the first report to identify a genetic link to altered EMB efficacy in L. salmonis in the Pacific Ocean.