Natural hybrid origin of a controversial "species", Clematis pinnata (Ranunculaceae), based on multidisciplinary evidence

Hybridization is common and has often been viewed as a driving force of plant diversity. However, it may raise problems in taxonomy, biodiversity estimation, and biological conservation. Although previous molecular phylogenetic studies suggested that hybridization may be rather common in Clematis, and artificial hybridization has been widely applied to produce new Clematis cultivars for nearly two centuries, the issue of natural hybridization of Clematis has never been addressed in detail. In this study, we tested the hybrid origin of Clematis pinnata, a rare and taxonomically controversial species distributed in northern China. Using field observations, morphological statistics, flow cytometry, niche modelling, and phylogenomic analysis, we tested the hypothesis of the possible hybrid origin of C. pinnata. The results show morphologically intermediacy of C. pinnata between the putative progenitors. Homoploidy among all the tested species is confirmed by flow cytometric analysis. Niche modelling results demonstrate that C. pinnata had not been adapted to a novel ecological niche independent of its putative parents. The plastome phylogeny indicates that C. pinnata plants in different sampling sites originated by different hybridization events. Phylonet and HyDe analyses based on transcriptome data corroborate the hybrid origins of C. pinnata from C. brevicaudata×C. heracleifolia/C. tubulosa. Rare introgression event between C. brevicaudata and C. heracleifolia/C. tubulosa was also detected in this study. These findings show that C. pinnata is representing F1 progeny or early generation hybrids of its parental species and evolved independently in different sites. The present study also indicates that interspecific or even intersectional hybridization is a common mechanism that the genus Clematis uses to generate diversity and variation, and it may play an important role in the evolution and diversification of this genus. Our study implies that morphological diversity caused by natural hybridization may overstate the real species diversity in Clematis.