The molecular mechanisms underlying major phenotypic changes that have evolved repeatedly in nature are generally unknown. Pelvic loss in different natural populations of threespine stickleback fish has occurred by regulatory mutations deleting a tissue-specific enhancer of the Pituitary homeobox transcription factor 1 (Pitx1) gene. The high prevalence of deletion mutations at Pitx1 may be influenced by inherent structural features of the locus. Although Pitx1 null mutations are lethal in laboratory animals, Pitx1 regulatory mutations show molecular signatures of positive selection in pelvic-reduced populations. These studies illustrate how major expression and morphological changes can arise by single mutational leaps in natural populations, producing new adaptive alleles via recurrent regulatory alterations in a key developmental control gene.

Hindlimb loss has evolved repeatedly in many different animals by means of molecular mechanisms that are still unknown. To determine the number and type of genetic changes underlying pelvic reduction in natural populations, we carried out genetic crosses between threespine stickleback fish with complete or missing pelvic structures. Genome-wide linkage mapping shows that pelvic reduction is controlled by one major and four minor chromosome regions. Pitx1 maps to the major chromosome region controlling most of the variation in pelvic size. Pelvic-reduced fish show the same left-right asymmetry seen in Pitx1 knockout mice, but do not show changes in Pitx1 protein sequence. Instead, pelvic-reduced sticklebacks show site-specific regulatory changes in Pitx1 expression, with reduced or absent expression in pelvic and caudal fin precursors. Regulatory mutations in major developmental control genes may provide a mechanism for generating rapid skeletal changes in natural populations, while preserving the essential roles of these genes in other processes.

Next-generation sequencing and the collection of genome-wide data allow identifying adaptive variation and footprints of directional selection. Using a large SNP data set from 259 RAD-sequenced European eel individuals (glass eels) from eight locations between 34 and 64(o) N, we examined the patterns of genome-wide genetic diversity across locations. We tested for local selection by searching for increased population differentiation using F(ST) -based outlier tests and by testing for significant associations between allele frequencies and environmental variables. The overall low genetic differentiation found (F(ST) = 0.0007) indicates that most of the genome is homogenized by gene flow, providing further evidence for genomic panmixia in the European eel. The lack of genetic substructuring was consistent at both nuclear and mitochondrial SNPs. Using an extensive number of diagnostic SNPs, results showed a low occurrence of hybrids between European and American eel, mainly limited to Iceland (5.9%), although individuals with signatures of introgression several generations back in time were found in mainland Europe. Despite panmixia, a small set of SNPs showed high genetic differentiation consistent with single-generation signatures of spatially varying selection acting on glass eels. After screening 50 354 SNPs, a total of 754 potentially locally selected SNPs were identified. Candidate genes for local selection constituted a wide array of functions, including calcium signalling, neuroactive ligand-receptor interaction and circadian rhythm. Remarkably, one of the candidate genes identified is PERIOD, possibly related to differences in local photoperiod associated with the >30° difference in latitude between locations. Genes under selection were spread across the genome, and there were no large regions of increased differentiation as expected when selection occurs within just a single generation due to panmixia. This supports the conclusion that most of the genome is homogenized by gene flow that removes any effects of diversifying selection from each new generation.

The outcome of natural hybridization is highly variable and depends on the nonexclusive effects of both pre- and post-mating reproductive barriers. The objective of this study was to address three specific questions regarding the dynamics of hybridization between the American and European eels (Anguilla rostrata and Anguilla anguilla). Using 373 AFLP loci, 1127 eels were genotyped, representing different life stages from both continents, as well as multiple Icelandic locations. We first evaluated the extent of hybridization and tested for the occurrence of hybrids beyond the first generation. Second, we tested whether hybrids were randomly distributed across continents and among Icelandic sampling sites. Third, we tested for a difference in the proportion of hybrids between glass eel and yellow eel stages in Iceland. Our results provided evidence for (i) an overall hybrid proportion of 15.5% in Iceland, with values ranging from 6.7% to 100% depending on life stages and locations; (ii) the existence of hybrids beyond the first generation; (iii) a nonrandom geographic distribution of hybrids in the North Atlantic; and (iv) a higher proportion of first and later generation hybrids in yellow eels compared to glass eels, as well as a significant latitudinal gradient in the proportion of hybrids in Icelandic freshwater. We propose that the combined effect of both differential survival of hybrids and variation in hybridization rate through time best explain these patterns. We discuss the possibility that climate change, which is impacting many environmental features in the North Atlantic, may have a determinant effect on the outcome of natural hybridization in Atlantic eels.

The two North Atlantic eel species, the European eel (Anguilla anguilla) and the American eel (Anguilla rostrata), spawn in partial sympatry in the Sargasso Sea, providing ample opportunity to interbreed. In this study, we used a RAD (Restriction site Associated DNA) sequencing approach to identify species-specific diagnostic single-nucleotide polymorphisms (SNPs) and design a low-density array that combined with screening of a diagnostic mitochondrial DNA marker. Eels from Iceland (N ¼ 159) and from the neighboring Faroe Islands (N ¼ 29) were genotyped, along with 94 larvae (49 European and 45 American eel) collected in the Sargasso Sea. Our SNP survey showed that the majority of Icelandic eels are pure European eels but there is also an important contribution of individuals of admixed ancestry (10.7%). Although most of the hybrids were identified as F1 hybrids from European eel female  American eel male crosses, backcrosses were also detected, including a first-generation backcross (F1 hybrid  pure European eel) and three individuals identified as second-generation backcrosses originating from American eel  F1 hybrid backcrosses interbreeding with pure European eels. In comparison, no hybrids were observed in the Faroe Islands, the closest bodies of land to Iceland. It is possible that hybrids show an intermediate migratory behaviour between the two parental species that ultimately brings hybrid larvae to the shores of Iceland, situated roughly halfway between the Sargasso Sea and Europe. Only two hybrids were observed among Sargasso Sea larvae, both backcrosses, but no F1 hybrids, that points to temporal variation in the occurrence of hybridization.