Circumpolar phylogeography of the northern pike (Esox lucius) and its relationship to the Amur pike (E. reichertii)

Frontiers in Zoology Circumpolar phylogeography of the northern pike (Esox lucius) and its relationship to the Amur pike (E. reichertii) Anna Skog 0 L Asbjrn Vllestad 0 Nils Chr Stenseth 0 Alexander Kasumyan Kjetill S Jakobsen 0 0 Centre for Ecological and Evolutionary Synthesis (CEES), Department of Biosciences, University of Oslo , P. O. Box 1066 Blindern, N-0316 Oslo , Norway Background: Freshwater fishes of the genus Esox are found throughout the Holarctic region. The northern pike (E. lucius) has a circumpolar distribution whereas the assumed sister species the Amur pike (E. reichertii) is only found in the Amur region. The genetic structure and post-glacial dispersal of these species are not well known. Here, we use sequence variation at two mitochondrial DNA regions (cytb and D-loop) to investigate the phylogeography, infer location of glacial refugia and investigate the time of divergence and potential demographic expansion of the various clades detected. Results: The two species did not share haplotypes implying long-term isolation with no gene flow, and divergence of the taxa were estimated at 4.55 Myr. The northern pike mtDNA haplotypes revealed three main lineages. One of the northern pike mtDNA lineages was found throughout the entire Holarctic region suggesting transcontinental dispersal from a single refugium. The three lineages exhibited a star phylogeny, indicating population expansion following isolation in separate glacial refugia. Estimated time of divergence of these lineages was 0.18 - 0.26 Myr. Conclusions: The precise location of the glacial refugia is uncertain, but our data suggests an Asian origin. The expansion of the circumpolar lineage is estimated to be around the end of the second glacial, implying that the current distribution is due to a recent recolonization from an east-Asian refugium. All three northern pike mtDNA lineages occurred sympatrically in Europe, possibly due to secondary contact. Two of the lineages probably originated from different European refugia, one in the Danube-region and one in Western Europe, the latter seems to be the primary source for recolonization of northern Europe. cyt b; D-loop; Glacial refugia; Phylogeny - Background Present-day distribution of temperate species of plants and animals are strongly influenced by geological and climatic processes such as glacial cycles. During glaciation periods populations were displaced and population size reduced, and as a consequence genetic diversity was lost [1,2]. During interglacial periods aquatic species such as freshwater fishes had access to large lakes formed by meltwater, providing a diversity of dispersal opportunities. There are a number of phylogeographic studies investigating the geographic distribution of genealogical lineages of freshwater fish, but patterns differ between species [3], and there are many potential biases. Freshwater fishes of the genus Esox are found throughout the Holarctic region. The northern pike (E. lucius) is the only species within the genus with a circumpolar distribution, whereas E. americanus, E. masquinongy and E. niger are only found in North America and two species only in Eurasia (E. reichertii, and E. cisalpinus (synonymous with E. flaviae)) [4-6]. The geographic origin of the genus Esox is unclear. The relatively higher number of species in North America, together with fossil evidence [7], has led to the suggestion that the genus originated on this continent [8]. While northern pike is one of the freshwater fish species with the widest Holarctic distribution range, the closely related Amur pike (E. reichertii) [9,10] has a narrow distribution essentially being restricted to the Amur River basin in northeastern Asia. Some recent genetic evidence suggests that the northern pike originated in North American subsequently dispersing to Eurasia over the Bering land bridge [8]. This is contrary to what has been suggested earlier [11]. Further, studies indicate that North American northern pike populations have less genetic diversity and structure than the European populations [8,12-19]. Lack of genetic diversity and structure are signs often associated with recently colonized regions, while more variability and genetic structuring are seen in populations that have been existing for longer periods of time [20]. During the different glacial periods Holarctic freshwater fishes survived in a number of geographically separated refugia, and this isolation and the subsequent dispersal from these refugia have been important for shaping the intraspecific genetic variability currently seen for many freshwater fish species in the Holartic [1,3]. Generally, the geographic distribution of a species and the association with distinct phylogeographic groups (clades) are useful data for identifying glacial refugia [21-23]. Here, we assess the level and geographic distribution of genetic variation of the northern and the Amur pike. Amur pike has so far only been investigated in context of higher level phylogeny [9,10]. Maes et al. [24] have suggested, on the basis of short fragments of mitochondrial DNA, that the northern pike experienced a recent expansion across the Holarctic from a single refugium. The geographic location of this refugium could not be inferred. A later study [25] found more structuring (in the distribution of hapotypes) based on a longer mitochondrial DNA fragment, implying that larger fragments are needed to test this hypothesis. In this study, we used two mitochondrial DNA markers, a 1133 bp sequence of the cytochrome b (cyt b) region and a 443 bp sequence of the D-loop, to evaluate the origin and dispersal of northern pike and Amur pike using a circumpolar dataset for a northern pike. Results Mitochondrial diversity in northern and Amur pike We obtained samples from 24 northern pike populations representing a total of 12 drainages across the Holarctic, and three Amur pike populations from the Amur River (sampling information and GenBank accession numbers are provided in Additional file 1). Details on diversity is given in Additional file 2, but is summarized here. The final mtDNA alignments included 443 bp of the D-loop region, and 1133 bp of the cyt b region for both northern and Amur pike. There were, respectively, 39 and 17 segregating sites in the cyt b and D-loop region for northern pike. Nucleotide diversity for this species ranged from 0.0052 (D-loop) to 0.0031 (cyt b). The alignments of Amur pike showed 10 segregating sites for cyt b and 3 for D-loop. The nucleotide diversity ranged from 0.0012 (D-loop) to 0.0020 (cyt b). Haplotype distribution The phylogenetic tree of concatenated D-loop and cyt b sequences revealed 38 haplotypes, 28 in northern pike and 10 in Amur pike (Figure 1). The northern and Amur pike haplotypes each formed two strongly supported clades and the two species did not share any haplotypes. Thus, the minimum spanning network revealed two separate, unconnected clusters (Figure 2). (...truncated)