Speciation and adaptation in Southern Ocean sea spiders

Marine shelf habitats in the Southern Hemisphere have been drastically impacted by glacial periods especially during the Plio- and Pleistocene. Large parts of the sea floor were at least temporarily covered by grounded ice, thereby reducing habitat availability for benthic organisms. Nowadays, an astonishingly high number especially of endemic species has been reported for this shelf fauna. Many of the species have radiated during the Plio- and Pleistocene. Allopatric speciation in independent and isolated refugia has often been postulated as the driving speciation mechanism, especially in the Antarctic, but also at higher latitudes. However alternative driver for speciation have rarely been considered, and especially ecological speciation in sympatry (e.g. shared refugia) due to adaptive divergence seems to be a promising mechanism to address with morphological and genetic data. Hence, one aim of this thesis is to explore evidence for both speciation scenarios focussing on two sea spider species complexes, namely Pallenopsis patagonica (Hoek, 1881) and Colossendeis megalonyx Hoek, 1881. The first study addresses genetic diversity within the P. patagonica species complex. Previously analysed mitochondrial cytochrome c oxidase subunit I (COI) data of Patagonian and Antarctic specimens was expanded by adding further samples from Patagonia, sub- Antarctica and the Eastern Weddell Sea. Furthermore, sequence data for the nuclear internal transcribed spacer (ITS) were added to obtain more information about the species complex. In fact, a higher number of distinct lineages were detected. Some lineages detected by mitochondrial data were not supported by nuclear data probably due to a lack of resolution (too few substitutions) in the ITS region rather than hybridization and speciation reversal events as reported in an earlier study about the sea spider species complex C. megalonyx. While the first study was based on two single-markers only, in the second study of this thesis an unprecedentedly large genomic data set for specimens of the P. patagonica species complex was generated based on a new high-throughput sequencing method, called target hybrid enrichment. Out of 1607 targeted EOGs (Eukaryotic Ortholog Groups, i.e. imputative single- copy genes), 821 EOGs were successfully captured for the P. patagonica species complex. Results of phylogenomic analyses supported previously reported lineages but further resolved the species phylogeny and added much higher statistical support to branches. In combination with extensive morphological and morphometric analyses, the large data set enabled to delineate at least eight species. On the basis of this, two new species, namely P. aulaeosmanorum sp. nov. Dömel & Melzer, 2019 and P. obstaculumsuperavit sp. nov. Dömel, 2019 were formally described. Neither genomic nor morphological data revealed consistent evidence for positive selection, rendering speciation in allopatric glacial refugia, due to genetic drift, as the most likely scenario. 7 Abstract In the third study the target hybrid enrichment approach was applied to the second sea spider species complex C. megalonyx. All 1607 targeted EOGs were recovered and a phylogenetic tree with high branch support values was generated based on the genomic data. Phylogenomic analyses supported already reported recent divergences as well as mito-nuclear discordances within this taxon, but resolution and statistical support were improved. Using SNP data inferred from EOGs and their more variable flanking regions, also intraspecific divergence patterns could be resolved. The results indicate restricted gene flow between geographically distinct populations. Morphometric analyses revealed multiple significant differences between lineages, but a clear separation was difficult. Evidence for positive selection was found for four genes associated with structural and neuronal functions. Hence, there is indication for positive selection on genes of the C. megalonyx species complex, but its specific contribution to the speciation process remains to be explored further. In the last study the phylogeny of the group of the longitarsal Colossendeidae which is especially abundant in the Southern Ocean and includes the species complex C. megalonyx, was investigated using target hybrid enrichment. Here, also all 1607 targeted EOGs were recovered. A well-resolved phylogeny of the group, which is mostly consistent with morphological data, was obtained. The data supported an Antarctic origin of the longitarsal Colossendeidae and multiple dispersal events to other regions, which occurred at different timescales. This scenario highlights the role of the Southern Ocean as a centre of origin also for non-Antarctic species. The comparison of phylogenetic trees based on single and target hybrid enrichment marker approaches underlines the benefit of the latter when resolving both ancient and recent speciation events. Analyses of morphological characters performed in this thesis revealed high variation even within lineages of both species complexes. Although distinct morphological characters suitable for species description and characters with significant differences between lineages were found, none of those seem to have distinct differences of lineages. Hence, no evidence for ecological character displacement and adaptive divergence of lineages that occur in sympatry as found. However, for some genes studied for lineages of the C. megalonyx species complex, dN/dS tests revealed codons and branches under selection. Differences in the detection of genes under selection between the two sea spider species complexes are likely based on methodological limitations as the design for target hybrid enrichment approach was based on a transcriptome of C. megalonyx. The reduced enrichment of only about half the number of EOGs for the P. patagonica species complex, limited the analyses. However, also biological differences, e.g. in reproduction strategies, can be considered as a factor for differences in speciation scenarios. The reproductive mode remains unclear for the C. megalonyx species complex but it most likely differs from the brooding strategy of the P. patagonica species complex. This can lead to differences in dispersal 8 Abstract capability and reproduction possibility of distinct populations, that are potentially higher in the C. megalonyx species complex. Based on the results of this thesis it is likely that different mechanisms drove speciation events (and speciation reversal) in Southern Ocean sea spiders and future studies should furthermore test different options for drivers of speciation rather than considering that only the impact of glacial cycles formed the species-rich benthic communities in the Southern Hemisphere. While analyses can be improved by adding more genomic regions and improved gene annotations in order to more holistically analyse targeted genes of selection, using target hybrid enrichment enabled to generate the largest genomic data sets available for sea spiders, so far.