Spatial genetic structure of Campanula thyrsoides across the European Alps: Indications for glaciation-driven allopatric subspeciation

Quaternary climate change had profound impacts on the geographical distribution and genetic structure of plant species which is hypothesized to have triggered allopatric speciation due to spatial isolation. However, evidence is scarce despite recent advances that indicate glacial history and substrate requirements as main drivers of spatial genetic structures. Here we built upon these studies to test the role of glaciations on the morphological and ecological differentiation within the calcicolous Campanula thyrsoides across its European Alpine distribution range. We collected samples from 51 populations (1173 plants), used five microsatellite markers, estimated diversity (Na, HE) and differentiation (Dest, GST_est, FST) and applied Bayesian clustering analysis and tessellation methods. We found support for four genetically distinct groups of populations, arranged from West to East: (i) France and Western Switzerland, (ii) Central and most of Eastern Switzerland, (iii) parts of Eastern Switzerland and Central Austria, and (iv) Southeastern Austria, Slovenia and Northeastern Italy. Among-subspecies variance was 8.4% and each subspecies was highly differentiated (C.*thyrsoides: Dest = 0.47; C.*carniolica: Dest = 0.58). Geographic structuring of within-population diversity was not related to refugia outside of previously-glaciated terrain but to subspecies: the more thermophilic C.*carniolica showed significantly lower levels of within-population diversity and higher numbers of private alleles. The location of the genetic break lines between these four groups of populations corresponds to well-known biogeographic barriers. However, the phylogeographic pattern has elements formerly found in both calcicolous and silicicolous species and thus questions the generality of substrate-related patterns. Within C.*thyrsoides, population admixture upon secondary contact may have led to high genetic diversity across the distribution range. Given the geographic and genetic differences of the subspecies we conclude that differentiation between C.*thyrsoides and C.*carniolica represents a case of glaciation-driven allopatric subspeciation reinforced by missing secondary contact due to incomplete post-glacial recolonization of potential habitats.