DOI: 10.5061/dryad.bvq83bkks
title: History of speciation inferred from genomic analysis of a species complex of north temperate fishes

description: Comparative biogeographic analyses have provided key insights into the
 history of divergence among closely-related species. We collected genomic
 data across much of the range of two salmonid fishes, Arctic char (AC,
 Salvelinus alpinus) and Dolly Varden (DV, S. malma) that comprise
 a complex of lineages whose relationships and evolutionary interactions
 have remained uncertain. A time-calibrated phylogeny indicated reciprocal
 monophyly of AC and DV, that the species diverged an estimated ~1.4
 million years ago, and that eastern Pacific southern Dolly Varden (S. m.
 lordi) is the basal lineage within DV. Historical and contemporary gene
 flow was detected between species and regional groups within species. We
 found strong evidence for a model of divergence without gene flow between
 AC and DV followed by secondary contact about 14,500 years ago with
 subsequent gene flow. Our geographic and genomic investigation within the
 AC-DV complex clarifies the origin and inter-relationships of the species
 and further highlight the North Pacific and Arctic as dynamic areas of
 evolution of regional faunas
description: Genotyping by sequencing analysis using Pst1 enzyme. It has been
 aligned to the <em>Salvelinus</em> sp. reference genome and
 consists of a variant call format file (vcf) with 350 samples that
 contains biallelic single nucelotide polymorphims that have a
 heterozygosity of less than 0.6 (N = 6,601,213).
description: # History of speciation inferred from genomic analysis of a species
 complex of north temperate fishes Our submission consists of two files:
 (1) a CSV file of supporting information and (2) a VCF file of
 single-nucleotide polymorphisms. **(1)** **CSV** **Supporting
 information** A file of 21 columns consisting of: **Sample name**: the
 name of each sample corresponding to samples in the VCF file, with the 5
 code variables corresponding to: 1. **Taxon**: AC=Arctic char, SC = Stone
 char, NDV = Northern Dolly Varden, EPSDV = Eastern Pacific Southern Dolly
 Varden, WPSDV = Western Pacific Southern Dolly Varden, xNDVEPSDV = Admixed
 NDV and EPSDV, x NDVWPSDV = admixed NDV and WPSDV, WSC = Whitespotted
 char, BT = Bull trout, LKTR = lake trout, BRTR = brook trout.  2.
 **Geographic area:** AK = Alaska, CARC = Canadian Arctic, YT = Yukon
 Territory, ATL = western Atlantic Basin, JP = Japan, RU = Russia, KUR =
 Kuril Islands, BC = British Columbia, WA = Washington State, NA = Not
 applicable 3. **Life history applicable to DV and AC:** AF = Allopatric,
 freshwater resident, AS = Allopatric, sea-run, SF = Sympatric, freshwater
 resident, NN/AN = not applicable. 4. **Three letter/number code
 corresponding to sequencing lane:** A01, A02, A04, LBF, BWA, CHI 5.
 **Individual sample number** For example: AC_AK_AF_A04_031 Refers to an
 Arctic char from Alaska that is allopatric (with respect to Dolly Varden)
 and a permanent resident of freshwater, It was in sequencing lane A04 and
 is sample number 31 from that population. Five columns (2-6) indicated
 whether (Y = yes, N = No) the sample was used in one of five analyses
 (SplitsTree, BEAST/SNAPP, PCA/fastSTRUCTURE/Admixture, Dsuite,
 Fastsimcoal2) **Columns 7-21 consist of:** The barcode of each sample used
 in each sequencing run The barcode assigned to each plate if more than one
 per sequencing run Name of the sequencing lane: A01, A02, A04, LBF, BWA,
 CHI The taxonomic name of each sample The original lab sample geographic
 location code and number The name of the location code given immediately
 above Latitude of sample site (Decimal degrees) Longitude of sample site
 (Decimal degrees) Collection year Population number code shown in
 Supplementary table 1 all_reads: Total number of reads after
 demultiplexing with process_radtags bam2_mapped: Number of reads mapped to
 the  reference genome with bwa as reported by samtools flagstat reads
 mapped with q&gt;20: Number of reads mapped to the genome with mapping
 quality (MAPQ) af 20 or higher as reported by samtools view -q20 depth
 q&gt;20: Number of positions in the genome with at least one read mapped
 with mapping quality of at least 20, obtained by counting the number of
 lines in the output of command samtools depth -Q 20 FMISS350: Proportion
 of missing data Missing data code: NA **(2) VCF file**
 (MASTER350revised.recode.vcf): Single nucleotide polymorphisms in VCF
 format for 350 individuals from six species of *Salvelinus* (Pisces) from
 throughout the North Pacific, Canadian Arctic, and Canadian northwest
 Atlantic basins. The SNPs were called after aligning raw sequence reads to
 the *Salvelinus malma* reference genome (assembly ASM291031v2; Christensen
 et al., 2018) across 36 non-sex liked chromosomes. This file is the
 dataset used in our article, "History of speciation inferred from
 genomic analysis of a species complex of north temperate fishes" The
 article investigates the history of speciation and gene flow in two
 species of Salvelinus which a checkered taxonomic history and produces a
 well-supported phylogeny, phylogeographic structure within species and
 strong evidence for historical introgression as the explanation for a
 history of taxonomic confusion. This Variant Call Format (VCF) file
 contains Single Nucleotide Polymorphism (SNP) data from 350 individuals
 across 98 sampling locations. The average number of reads of minimum
 genotype quality of 20 was 6.8 million. The data were generated using
 genptyping-by-sequencing with the enzyme Pst1. We used VCFtools v0.1.11
 (Danecek et al., 2011) to estimate the amount of missing data per sample
 and used only those with no more than 70% missing data (average of 0.51).
 We used VCFtools to filter these 350 samples to retain only biallelic SNPs
 (no indels or sites with more than two segregating bases; N = 7,434,394
 retained). Finally, we eliminated SNPs that showed an observed
 heterozygosity of 0.6 or higher, as these are likely the result of mapping
 to paralogous regions of the genome, using a custom script (Owens et al.,
 2016). This resulted in a “master” VCF file with 6,601,213 SNPs that was
 subject to additional filters depending on the particular analysis (see
 associated manuscript). File Details File Name:
 MASTER350revised.recode.vcf File Format: VCF (Variant Call Format) Version
 4.2 Date: February 4, 2025 Source Software: Stacks v2.41 Data Description
 The VCF file includes the following columns standard to the format:
 \#CHROM: Chromosome number POS: Position of the SNP on the chromosome ID:
 Identifier of the SNP REF: Reference base ALT: Alternate base(s) QUAL:
 Quality score of the SNP FILTER: Filter status INFO: Additional
 information (e.g., allele frequency, number of samples) FORMAT: Data
 format Sample columns: One per individual, containing genotype information
 Specific Fields in INFO NS: Number of samples with data AF: Allele
 frequency DP: Total depth of reads Specific Fields in FORMAT GT: Genotype
 AD: Allele depth DP: Read depth GQ: Genotype quality GL: Genotype
 likelihood How to Open and Analyze the Data The VCF file can be opened and
 analyzed using various bioinformatics tools and software incliding: IGV
 (Integrative Genomics Viewer): Useful for visualizing genomic data.
 BCFtools: Command-line tool for working with VCF/BCF files, including
 filtering, viewing, and converting. GATK4 (Genome Analysis Toolkit):
 Provides tools to analyze high-throughput sequencing data with a focus on
 variant discovery. vcftools: Command-line program designed to work
 specifically with VCF files to perform various types of analyses.
 Sharing/Access information All sequencing data from this study is being
 deposited at NCBI Sequence Read Archive under submission SUB15066945.
 Methods Data Collection We examined 350 DNA samples from a database of 909
 fish acquired over the past 25 years in the context of various independent
 studies on char evolutionary ecology. Our samples comprised Northern (NDV,
 S. m. malma) and Southern Dolly Varden (SDV, S. m. lordi) collected from
 northwestern North America. The SDV samples were collectively referred to
 as Eastern Pacific Southern Dolly Varden, EPSDV. We also examined AC (S.
 alpinus erythrinus) from Alaska, Yukon, and east to the eastern Canadian
 Arctic and Atlantic Canada (see Supporting Information file). Samples
 included fish from allopatric and sympatric populations as well as both
 sea-run and freshwater-resident life history types. Our samples also
 included four specimens of the “stone char” a phenotypically distinct form
 of NDV from Kamchatka (see Melnick et al., 2020), five samples of other
 NDV from Russia (Kamchatka), 19 samples thought to be S. m.
 krascheninnikovi, S. m. malma, or hybrids between the two subspecies from
 the western Pacific (5 from Hokkaido, Japan and 14 from the Kuril Island
 chain). In Asia, S. m. krascheninnikovi is also referred to as Southern
 Dolly Varden; here, we refer to it as Western Pacific Southern Dolly
 Varden (WPSDV). We also included two samples each of coastal and interior
 lineages of bull trout (S. confluentus, Taylor et al., 1999), and two
 specimens each of brook trout (S. fontinalis), lake trout (S. namaycush),
 and white-spotted char (S. leucomaenis) as outgroup taxa. Initial identity
 of all taxa was based on independent information on geographic
 distribution, and morphological and genetic (nuclear DNA) data (e.g.,
 Moore et al., 2015; Taylor & May-McNally, 2015; Weinsten et al.,
 2024a,b). All fish tissues were collected under UBC Animal Care Committee
 approved protocols. Total genomic DNA was extracted from tissues using the
 DNeasy Blood & Tissue kit and eluted in 200 µL of AE buffer. We used a
 reduced representation genome sequencing approach,
 genotyping-by-sequencing (GBS; Elshire et al., 2011), to generate sequence
 data from a representative fraction of the genome. We used a modified GBS
 protocol (Alcaide et al., 2014; Toews et al., 2016; Geraldes & Taylor,
 2023) to generate pooled libraries of digested and individually barcoded
 DNA. The libraries were sequenced using Illumina HiSeq 4000, NovaSeq 6000
 SP and NovaSeq 6000 S4 always with 150 bp paired-end reads at the McGill
 University and Génome Québec Innovation Centre. Analysis of the sequence
 data followed a bioinformatics pipeline available at
 [https://doi.org/10.5061/dryad.t951d](https://doi.org/10.5061/dryad.t951d)
 (Irwin et al., 2016) for GBS read processing and mapping with
 modifications detailed below. We used process_radtags from the STACKS v2.5
 pipeline (Catchen et al., 2013; Rochette et al., 2019) to demultiplex the
 raw sequencing reads according to the barcode sequence of each sample (or
 the combination of barcode and plate sequence) and remove the barcode
 sequences. The resulting reads for each sample were then trimmed with
 Trimmomatic-0.39 (Bolger et al., 2014) with options TRAILING:3,
 SLIDINGWINDOW:4:10, MINLEN:30. We used BWA-MEM (Li & Durbin, 2009)
 with default settings to align the trimmed reads from each of the
 Salvelinus samples for which we generated GBS data to what was originally
 considered the AC (Salvelinus alpinus) reference genome sequence (assembly
 ASM291031v2; Christensen et al., 2018), but which now has been identified
 as a specimen of NDV with about 5% admixture from AC (Shedko, 2020;
 Christensen et al., 2021; authors, unpublished data). References Alcaide,
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