Li_2026_PhytoKeys_274_31

Huperzia serrata is an endangered perennial medicinal lycophyte, primarily distributed across most regions of China, eastern and southern Asia, Oceania, and the Americas. It produces Huperzine A (HupA), a potent natural acetylcholinesterase (AChE) inhibitor with significant therapeutic potential for treating Alzheimer's disease. To elucidate the genetic diversity and phylogenetic relationships of this species and facilitate its conservation, we performed full-length transcriptome sequencing of H. serrata using the PacBio Sequel II platform and developed expressed sequence tag-simple sequence repeat (EST-SSR) markers. Transcriptome analysis identified 16,511 SSR loci from the transcriptome, with an average density of one locus per 4.11 kb. These loci exhibited diverse repeat motifs (ranging from mono- to hexanucleotides), predominantly containing 5-20 repeats. Genotyping of 47 Huperzia accessions (representing 8 different species) using 21 polymorphic primers identified 98 alleles, yielding mean Nei's gene diversity (H) and Shannon's information index (I) values of 0.3171 and 0.4819, respectively, indicating a moderate level of genetic diversity. Principal coordinate analysis (PCoA) revealed no significant correlation between genetic and geographic distance. Cluster analysis based on genetic similarity coefficients (ranging from 0.41 to 0.82) delineated distinct genetic groupings. However, SSR cluster analysis was inconsistent with taxonomic identification based on morphology to a great extent. At a similarity coefficient of 0.51, the accessions were classified into four major groups. Notably, accessions from Guizhou (GZ) and Yunnan (YN) provinces tended to cluster together, with accessions GS_1_GL and GZ_6_GY exhibiting the highest genetic similarity. This study provides the first comprehensive set of genomic resources and insights into the genetic diversity of Huperzia across a wide range, revealing its fine-scale genetic structure. These findings establish a crucial scientific foundation for the conservation and sustainable utilization of this valuable medicinal species.