Pronounced mito-nuclear discordance and various Wolbachia infections in the water ringlet Erebia pronoe have resulted in a complex phylogeographic structure.

Several morphological and mitochondrial lineages of the alpine ringlet butterfly species Erebia pronoe have been described, indicating a complex phylogenetic structure. However, the existing data were insufficient and allow neither a reconstruction of the biogeographic history, nor an assessment of the genetic lineages. Therefore, we analysed mitochondrial (COI, NDI) and nuclear (EF1α, RPS5) gene sequences and compared them with sequences from the sister species Erebia melas. Additionally, we combined this information with morphometric data of the male genitalia and the infection patterns with Wolbachia strains, based on a WSP analysis. We obtained a distinct phylogeographic structure within the E. pronoe-melas complex with eight well-distinguishable geographic groups, but also a remarkable mito-nuclear discordance. The mito-nuclear discordance in E. melas and E. pronoe glottis can be explained by different ages of Wolbachia infections with different Wolbachia strains, associated selective sweeps, and hybridisation inhibition. Additionally, we found indications for incipient speciation of E. pronoe glottis in the Pyrenees and a pronounced range dynamic within and among the other high mountain systems of Europe. Our results emphasize the importance of combined approaches in reconstructing biogeographic patterns and evaluating phylogeographic splits.

First microsatellite markers for the pine catkin sawfly Xyela concava (Hymenoptera, Xyelidae) and their application in phylogeography and population genetics.

Microsatellites are widely used as powerful markers in population genetics because of their ability to access recent genetic variation and to resolve subtle population genetic structures. However, their development, especially for non-model organisms with no available genome-wide sequence data has been difficult and time-consuming. Here, a commercial high-throughput sequencing approach (HTS) was used for the very first identification of microsatellite motifs in the genome of Xyela concava and the design of primer pairs flanking those motifs. Sixteen of those primer pairs were selected and implemented successfully to answer questions on the phylogeography and population genetics of X. concava. The markers were characterized in three geographically distinct populations of X. concava and tested for cross-species amplification in two additional Xyela and one Pleroneura species (Xyelidae). All markers showed substantial polymorphism as well as revealing subtle genetic structures among the three genotyped populations. We also analyzed a fragment of the nuclear gene region of sodium/potassium-transporting ATPase subunit alpha (NaK) and a partial mitochondrial gene region coding for cytochrome oxidase subunit I (COI) to demonstrate different genetic resolutions and sex-biased patterns of these markers, and their potential for combined use in future studies on the phylogeography and population genetics of X. concava. Although a limited number of populations was analyzed, we nevertheless obtained new insights on the latter two topics. The microsatellites revealed a generally high gene flow between the populations, but also suggested a deep historical segregation into two genetic lineages. This deep genetic segregation was confirmed by NaK. While the high gene flow was unexpected, because of assumed restricted dispersal ability of X. concava and the discontinuous distribution of the host trees between the populations, the segregation of two lineages is comprehensible and could be explained by different refuge areas of the hosts during glacial times. The COI results showed a discordant strong genetic structure between all populations, which might be explained by the smaller effective population size of the mitochondrial genome. However, given the frequent evidence of a similar nature in recent studies on sawflies, we also consider and discuss mitochondrial introgression on population level as an alternative explanation.