Abstract:

Aims: Gymnospermium kiangnanense is a rare and endangered plant species endemic to China. Its distribution is restricted to Anhui and Zhejiang provinces, and it has been catalogued in the local registry of rare and endangered flora. However, there is relatively little research on it. Therefore, to investigate the genetic structure and elucidate the endangered mechanisms of G. kianganese, we conducted a comprehensive analysis based on its chloroplast genome, leading to evidence-based conservation recommendations.

Methods: This study assembled chloroplast genome sequences from 39 individuals across 6 populations to enhance the conservation of G. kiangnanense. Based on these chloroplast genomes, comparative genomic analyses and population genetic structure analyses were conducted to explore conservation strategies for G. kiangnanense.

Results: (1) The chloroplast genome of G. kiangnanense was highly conserved in terms of sequence composition, gene structure, and gene content, among which simple sequence repeats (SSRs) types exhibited obvious population characteristics. (2) The three non-coding regions in the chloroplast genome, namely spacer between psbZ and trnG-GCC (psbZ-trnG-GCC), spacer between trnT-UGU and trnL-UAA (trnT-UGU-trnL-UAA), and spacer between ycf1 and ndhF (ycf1-ndhF), all exhibited high variability. Meanwhile, the ndhF gene also showed high nucleotide diversity, suggesting that these regions had potential as molecular markers. (3) The chloroplast genome showed high genetic diversity and high genetic differentiation among populations. (4) Analysis of 39 chloroplast genome sequences from 6 populations identified 14 haplotypes, which were classified into 3 distinct lineages through Network and Beast analyses. (5) The variation among the populations of G. kiangnanense was significant, and it had a clear geographical structure of lineages. (6) Demographic history analysis indicated a stable population size in G. kiangnanense, with no evidence of past expansion.

Conclusion: Gymnospermium kiangnanense employs an outcrossing reproductive strategy, which not only enhances genetic variation but also mitigates inbreeding depression. Moreover, the existence of glacial refugia in eastern China during the Quaternary Ice Age offered a stable habitat for this species, thereby further promoting its population genetic diversity. Nevertheless, several factors are likely contributing to its endangered status, including low seed-setting rates, limited seed dispersal capacity, and excessive human activities. Therefore, based on the analysis of the chloroplast genome and conservation genetics of G. kiangnanense, the following conservation strategies are proposed: (1) Establish three conservation units according to distinct genetic lineages, with priority given to protecting the Fenglinxia Village population in Zhuji, Zhejiang Province, through the creation of a conservation area. (2) Implement managed bee pollination during the flowering period to enhance pollination and seed set rates. (3) Reduce population density to improve light availability for seedlings and promote their growth. (4) Complement in situ conservation with ex situ efforts by establishing artificial breeding programs in suitable botanical gardens. (5) Strengthen scientific outreach to minimize anthropogenic disturbances. (6) Promote its propagation and conservation through rational exploitation of its medicinal value in pharmaceutical development.

Key words: Gymnospermium kiangnanense, chloroplast genome, comparative genomics, population genetic structure