Exploring diversity in cryptorhynchine weevils (Coleoptera) using distance-, character- and tree-based species delineation.

Species boundaries are studied in a group of beetles, the western Palaearctic Cryptorhynchinae. We test for congruence of 'traditionally' identified morphospecies with species inferred through parsimony networks, distance-based clustering and the ultrametric tree-based generalized mixed yule-coalescent (GMYC) approach. For that purpose, we sequenced two variable fragments of mitochondrial DNA (CO1 and 16S) for a total of 791 specimens in 217 species of Cryptorhynchinae. Parsimony networks, morphology-calibrated distance clusters and the different tree-based species inferences all achieved low congruence with morphospecies, at best 60%. Although the degree of match with morphospecies was often similar for the different approaches, the composition of clusters partially varied. A barcoding gap was absent in morphospecies-oriented distances as well as for GMYC species clusters. This demonstrates that not only erroneous taxonomic assignments, incomplete lineage sorting, hybridization, or insufficient sampling can compromise distance-based identification, but also differences in speciation rates and uneven tree structure. The initially low match between morphospecies and the different molecular species delineation methods in this case study shows the necessity of combining the output of various methods in an integrative approach. Thereby we obtain an idea about the reliability of the different results and signals, which enables us to fine-tune sampling, delineation technique and data collection, and to identify species that require taxonomic revision.