Taxonomic revision of the Primulina secundiflora complex based on morphological and molecular evidence

the species complex represents one of the most challenging issues in taxonomic research, and the genus Primulina Hance is particularly prone to such complexes due to intricate morphological variation and incomplete lineage sorting. Five morphologically similar species, namely Pri. secundiflora , Pri. vestita , Pri. tenuituba , Pri. flexusa , and Pri. huahaii , collectively form the Pri. secundiflora complex, within which species boundaries remain ambiguous. In this study, we conducted a comprehensive integrative analysis of all five species, combining detailed morphological observations with multiple molecular approaches, including chloroplast genome comparison, nucleotide diversity analysis, population genetic diversity analysis, population genetic structure analysis, and phylogenetic reconstruction. Our results revealed several key findings. First, chloroplast genomes within the complex are highly conserved, with nucleotide diversity far below the average level of the genus Primulina. Second, uncorrected sequence divergence (Dxy) based on both ITS and chloroplast genomes showed that intraspecific differentiation often exceeds interspecific differentiation. Third, population genetic structure analysis indicated extensive genetic admixture among all populations, with no species represented by a single genetic component. Fourth, phylogenetic analyses based on chloroplast genomes failed to resolve any of the five species as monophyletic. Integrative evidence thus strongly supports treating all five species as a single species. However, Pri. secundiflora , Pri. flexusa , and Pri. huahaii exhibit distinct and stable corolla morphological differentiation across populations, which we interpret as a selfing syndrome. resulting from mating system shifts rather than evidence of independent speciation. Accordingly, we propose the following taxonomic revisions: Pri. tenuituba is reduced to synonymy under Pri. vestita , and Pri. vestita , Pri. flexusa , and Pri. huahaii are each treated as varieties of Pri. secundiflora . This study highlights the necessity of integrating multiple lines of evidence in species delimitation and cautions against taxonomic oversplitting based solely on morphological discontinuities in Primulina .