Target Enrichment Data Uncovers Rapid Radiation, Whole Genome Duplication, and Extensive Hybridization in Slipper Orchid Genus Cypripedium L.

Cypripedium is the most widespread and morphologically diverse genus of slipper orchids. Despite several published phylogenies based on Sanger sequencing data, the topology and monophyly of its infrageneric taxa remained uncertain. Here, we aimed to reconstruct a robust section-level phylogeny of Cypripedium and explore its evolutionary history using target capture data for the first time. We used the orchid-specific bait set "Orchidaceae963" to reconstruct the phylogeny of Cypripedium based on 614 nuclear loci, covering 11 out of 13 sections. Subsequently, we investigated tree discordance, estimated divergence times and ancestral ranges, searched for anomaly zones, polytomies, and diversification rate shifts, and identified gene duplication and hybridization events. All sections were recovered as monophyletic, contrary to the subsections within sect. Cypripedium . Although the two subclades within this section did not correspond to its two subsections, they matched the geographic distribution of their species. Additionally, we discovered high levels of discordance in the short backbone branches of the genus and within sect. Cypripedium , which can be attributed to gene duplication and hybridization events, a potential whole genome duplication, and incomplete lineage sorting caused by rapid radiation. Our biogeographic analysis suggested a Neotropical origin of the genus during the Early Miocene (~20 Ma). The rapid radiations at the backbone likely occurred in Southeast Asia around the Middle Miocene Climatic Transition (~15-13 Ma), followed by several independent dispersals back to the New World. Moreover, the Pliocene-Quaternary glacial cycles may have contributed to further speciation and reticulate evolution, giving rise to a hybrid swarm within sect. Cypripedium . Our study provided novel insights into the evolutionary history of Cypripedium based on high-throughput molecular data, shedding light on the dynamics of its distribution and diversity patterns from its origin to the present. ### Competing Interest Statement The authors have declared no competing interest.