The dirty north: Evidence for multiple colonisations and Wolbachia infections shaping the genetic structure of the widespread butterfly Polyommatus icarus in the British Isles

The paradigm of differentiation in southern refugia during glacial periods followed by expansions during interglacials, producing limited genetic diversity and population sub-division in northern areas, dominates European phylogeography. However, the existence of complex structured populations in formerly glaciated areas, and on islands connected to mainland areas during glacial maxima, call for alternative explanations. Here, we reconstruct the mtDNA phylogeography of the widespread Polyommatus icarus butterfly over its native range, with an emphasis on the formerly glaciated and connected British Isles. We found distinct geographical structuring of CO 1 mitotypes, with an ancient lineage restricted to the marginal European areas, including Northern Scotland and Outer Hebrides. We detected perfect mtDNA- Wolbachia associations in Northern Britain that support the possibility of at least two post-glacial Wolbachia -mediated sweeps, suggesting a series of sequential replacement of mtDNA in the British Isles and potentially in Europe. Population genomic analysis, using ddRADSeq genomic markers, also reveal unexpected genetic structuring within Britain. However, weak mito-nuclear concordance suggests the potential for independent histories of nuclear versus mitochondrial genomes. We found clustering of genomic SNPs of French samples, with respect to those in the British Isles, is not consistent with a scenario of a single recolonisation. Taken together our mtDNA and ddRADseq observations are consistent with a history of at least two distinct colonisations, a phylogeographic scenario previously put forth to explain diversity and structuring in other British flora and fauna. Additionally, we also present preliminary evidence that Wolbachia- induced feminization may be occurring in the isolated population in the Outer Hebrides. ### Competing Interest Statement The authors have declared no competing interest.