DOI: 10.5061/dryad.x0k6djhvh
title: Data from: Genomic footprints of hybridization in North Atlantic eels (Anguilla anguilla and A. rostrata)

description: Understanding interspecific introgressive hybridization and the biological
 significance of introgressed variation remains an important goal in
 population genomics. European (Anguilla anguilla) and American eel (A.
 rostrata) represent a remarkable case of hybridization. Both are panmictic
 and spawn in partial sympatry in the Sargasso Sea, occasionally producing
 viable, fertile hybrids, primarily found in Iceland. We studied
 introgressive hybridization from American into European eel using
 whole-genome sequences of 78 individuals, including European, American,
 and 21 putative hybrid eels. Previous studies using few genetic markers
 could not resolve whether hybridization involved simple unidirectional
 backcrossing or a more complex hybrid swarm. However, local ancestry
 inference along individual chromosomes revealed that Icelandic hybrids
 were primarily F1 or first-generation backcrosses toward European eel,
 with some showing more complex backcrossing. All European eels outside
 Iceland contained short chromosomal blocks from American eel, indicating a
 porous genome. We found no evidence for previously hypothesized
 geographical gradients of introgression in European eel outside Iceland.
 Several chromosomal regions showed high interspecific divergence, but
 haplotype blocks introgressed from American eel were identified both
 within and outside these regions. There was little correspondence between
 regions of high relative (FST) and absolute divergence (dXY), with the
 former reflecting selective sweeps within species or reduced recombination
 rather than barrier loci. A single genomic region showed evidence of
 repeated introgression from American into European eel under positive
 selection in both species. The study illustrates that species can maintain
 genetic integrity despite porous genomes, and that standing variation in
 one species can potentially be available for future adaptive responses in
 the other species.
description: <strong>Sampling</strong> We
 analyzed a total of 50 European<em> </em>eels (of which one
 later turned out to be a hybrid), 7 American eels, and 21 individuals of
 admixed ancestry. We note that the admixed individuals do not represent a
 random selection of hybrids; 10 individuals had beforehand been identified
 as F1 hybrids and 11 as different types of backcrosses: bAA
 (first-generation backcross to European eel, N = 3), bAAxAA
 (second-generation backcross to European eel, N = 3) and bAR x AA
 (second-generation backcross, with backcrossing first to American eel and
 subsequently to European eel), N = 5). This initial hybrid identification
 was based on 68 species-diagnostic SNPs and analyses using Structure 2.3.4
 (Falush et al., 2003; Pritchard et al., 2000) and NewHybrids 1.1
 (Anderson, E. C. & Thompson, 2002) as detailed in Pujolar et al.
 (2014a). As demonstrated in this paper using both simulated and empirical
 data there is high power for distinguishing “pure” American and European
 eel from each other and from recently admixed individuals.
 The analyzed individuals were collected between 2001 and 2017
 using electrofishing or net fishing and encompassed both juvenile glass
 eels and adult eels within the continental ranges of American and European
 eel (see Fig. 1). Two eel larvae collected by ring net in the Sargasso Sea
 were also included. For individual sampling details, see Table S1. A total
 of 22 individuals, including 17 hybrids and 5 American eels, were
 sequenced for the present study. The remaining 56 individuals,
 encompassing primarily European eels, had previously been analyzed using
 whole-genome sequencing for the purpose of analyzing speciation between
 European and American eel and genome-wide methylation in European eel
 (Liu, S. et al., 2022; Nikolic et al., 2020).
 <strong>Mapping and variant
 calling</strong> Genomic DNA was extracted using
 a standard phenol-chloroform extraction or E.Z.N.A. purification columns
 (Omega Bio-Tek, Norcross, Georgia, USA). Whole-genome sequencing was
 outsourced to BGI (Beijing Genomics Institute, Hongkong, China) (nine
 individuals) and NOVOGENE (Hong Kong, China) (all other individuals).
 Sequencing libraries were constructed using the NEBNext® DNA Library Prep
 Kit (New England Biolabs, MA, USA). Genomic DNA was randomly fragmented to
 a size of 350bp by shearing, and PCR amplification was conducted.
 Paired-end Illumina sequencing was conducted using the Illumina HiSeq 2500
 platform with a read length of 150 bp and aimed for a coverage of
 ~20x. Using BWA MEM v.0.7.17 (Li, 2013; Li &
 Durbin, 2009a) with default parameter settings, the reads were mapped to a
 recent chromosome level European eel genome assembly (Rhie et al., 2021)
 (GenBank accession: GCA_013347855.1). Mapped reads were converted to BAM
 files, sorted and indexed using SAMtools v.1.9 (Li et al., 2009b). A VCF
 file of SNPs encompassing all 78 individuals was generated from the BAM
 files with BCFtools v.1.9 (Li, 2011; Li et al., 2009b) using a minimum
 mapping quality threshold of 20. Initial filtering of the SNPs was
 performed using VCFutils.pl (Li et al., 2009b) and VCFtools v.0.1.16
 (Danecek et al., 2011). Only biallelic SNPs with  minimum variant quality
 of 20, minor allele count ≥3, no missing data, and pooled read depth
 between 900 and 1500 across all individuals were kept. The thresholds were
 determined based on inspection of the SNP depth distribution (Fig. S1).
 SNPs on non-anchored scaffolds were discarded. We
 furthermore produced an ‘all sites’ data set by using BCFtools
 <em>call</em> without the <em>--variants-only</em>
 input option. Similarly to the ‘variant sites’ data set, we filtered the
 VCF to discard indels and retain only sites with minimum mapping quality
 of 20 and pooled read depth between 900 and 1500 across all individuals.
 No filtering for missing data and minor allele frequency was
 performed.
 <strong>LD-pruning</strong> The
 ‘variant sites’ data set was thinned using VCFtools, so that no two SNPs
 are within &gt;1,000 bp from one another - a distance at which linkage
 disequilibrium (LD) is virtually absent in the species (Jacobsen et al.,
 2014a). <strong>Statistical
 phasing</strong> For the purpose of inferring
 local ancestry along chromosomes and scanning for selective sweeps, we
 created a phased data set for the reference populations. The previously
 produced VCF file was split between hybrids and non-hybrids. The former
 file was not phased, as statistical phasing introduced a large number of
 switch errors (results not shown, but see e.g. Smeds et al. (2021)).
 Statistical phasing was performed using SHAPEIT v.2 (r900) (Delaneau et
 al., 2013). To increase accuracy, the number of conditioning states was
 increased from 100 states per SNP (default) to 200 states. Each chromosome
 was phased individually, and no recombination map was specified. The
 American and European eels were phased simultaneously and the phased VCF
 file was subsequently subset by species.
description: # Data from: Genomic footprints of hybridization in North Atlantic eels
 (Anguilla anguilla and A. rostrata)
 [https://doi.org/10.5061/dryad.x0k6djhvh](https://doi.org/10.5061/dryad.x0k6djhvh) ## Description of the data and file structure The data encompasses multiple VCF files generated from whole-genome resequencing of European eels *(Anguilla anguilla)*, American eels *(A. rostrata)* and their hybrids. ### Files and variables #### 1) chr01-19.vcf.gz Raw variant calls. #### 2) chr01-19.filtered.ann.mac3.max2.miss1.recode.vcf.gz Variant calls from file 1) filtered to retain only biallelic SNPs with minimum variant quality of 20, minor allele count ≥3, no missing data, and pooled read depth between 900 and 1500 across all individuals were kept. #### 3) chr01-19.filtered.ann.mac3.max2.miss1.phased.vcf.gz Variant calls from file 2) with phase information obtained by statistical phasing with SHAPEIT. #### 4) chr01-19.filtered.ann.mac3.max2.miss1.LD-thinned.recode.vcf.gz A thinned version of file 3) containing only no more than one SNP per 1,000 bp. #### 5) allSites.chr01-19.vcf.gz File containing both variant and invariant sites filtered as described for file 2. No filtering for missing data performed. ## Code/software Scripts are available at [https://github.com/atengstedt/Eel_hybrids](https://github.com/atengstedt/Eel_hybrids).