Population structure and diversification of , a plant species with extremely small populations endemic to eastern China.

Background: is the only species distributed in the subtropical region within the spring ephemeral genus . Extensive human exploitation and habitat destruction have resulted in a rapid shrink of populations. This study utilizes microsatellite markers to analyze the genetic diversity and structure and to deduce historical population events of extant populations of .

Methods: A total of 143 individuals from eight extant populations of , including two populations from Anhui Province and six populations from Zhejiang Province, were analyzed with using 21 pairs of microsatellite markers. Genetic diversity indices were calculated using Cervus, GENEPOP, GenALEX. Population structure was assessed using genetic distance (UPGMA), principal coordinate analysis (PCoA), Bayesian clustering method (STRUCTURE), and molecular variation analysis of variance (AMOVA). Population history events were inferred using DIYABC.

Results: The studied populations of exhibited a low level of genetic diversity ( = 0.179, = 0.286), but a high degree of genetic differentiation ( = 0.521). The mean value of gene flow ( ) among populations was 1.082, indicating prevalent gene exchange pollen dispersal. Phylogeographic analyses suggested that the populations of were divided into two lineages, Zhejiang (ZJ) and Anhui (AH). These two lineages were separated by the Huangshan-Tianmu Mountain Range. AMOVA analysis revealed that 36.59% of total genetic variation occurred between the two groups. The ZJ lineage was further divided into the Hangzhou (ZJH) and Zhuji (ZJZ) lineages, separated by the Longmen Mountain and Fuchun River. DIYABC analyses suggested that the ZJ and AH lineages were separated at 5.592 ka, likely due to the impact of Holocene climate change and human activities. Subsequently, the ZJZ lineage diverged from the ZJH lineage around 2.112 ka. Given the limited distribution of and the significant genetic differentiation among its lineages, both and conservation strategies should be implemented to protect the germplasm resources of .