Biogeography and ecology of geographically distant populations of sibling Cryptocephalus leaf beetles

Different populations of two closely related species, Cryptocephalusflavipes and C. bameuli, from western (Alps, Apennines and Pyrenees) and central Europe (Poland, Ukraine and Pannonia) were analysed. On the basis of DNA sequences from two genes, cox1 and ef1-alpha, distinctiveness of both species was confirmed. Nevertheless, possible hybrids were identified in Carpathian mountains. We found a significant genetic differentiation among populations of C. flavipes and C. bameuli from distant regions but a high genetic similarity between populations of C. bameuli from north and south of the Carpathians. Demographic estimates suggest a past population expansion in the case of C. bameuli and a recent one for C. flavipes, possibly occurred during Pleistocene and Holocene, respectively. Distribution modelling showed that C. flavipes is typically present in the mountain systems, whereas C. bameuli is associated with hilly areas of central and eastern Europe. Based on the present data, Last Glacial Maximum refugia of both species were located in the Alpine region and Black Sea coasts, but on different elevations. The characterization of the insect diet, through a DNA metabarcoding approach targeting the trnL plant intron, demonstrated a significant differentiation of food preferences between the two species, as well as between geographic populations within the species