Phylogenetic analysis of Chinese Enidae: Inference from genome-wide analyses and morphological studies on three selected genera

Enidae (Gastropoda, Stylommatophora, Pupilloidea) land snails exhibit specific regional distributions mainly in southern Gansu and northwestern Sichuan, China. Despite many morphological studies, molecular and integrative evidence remains insufficient for classification of doubtful species. More rigorous investigations into phylogeny are necessary to comprehend the evolutionary biology of Chinese Enidae in this biodiversity hotspot region. To address this issue, we focused on three Enidae genera—<i>Subzebrinus</i>, <i>Pupinidius</i>, and <i>Petraeomastus</i>—from 19 localities around Wenxian and Wenchuan in the adjoining regions of Gansu and Sichuan, China. We conducted genome-wide ddRAD-seq and employed a machine learning-based image clustering method, aiming to better integrate phylogenetic and morphological analyses of Enidae. Phylogenetic trees constructed by genome-wide SNP data revealed species-level monophyly but genus-level polyphyly, indicating the current taxonomy should be revised. Image clustering analysis revealed structured shell morphology clusters closely associated with phylogeny, suggesting both phylogeny and morphology were conservative within species, which was also demonstrated by geographical differentiation and localization within species. However, diversifications of molecular and shell morphology were also existed in particular sepcies of <i>Pupinidius</i> and <i>Petraeomastus</i>. The integrative analysis of genome-wide phylogeny and shell image clustering for selected genera will establish a solid foundation for evolutionary studies of Chinese Enidae.