Distinguishing eight Polygonatum species using morphological, anatomical, and SCoT markers

Polygonati Rhizoma, a traditional Chinese medicine, holds significant value in the pharmaceutical and food industries. However, adulteration with morphologically similar congeneric species often compromises product quality and efficacy. Current identification methods, primarily reliant on single approaches, struggle to accurately distinguish processed rhizomes. Therefore, developing a comprehensive and reliable identification method is essential for standardizing Polygonati Rhizoma products. This study identified eight Polygonatum species using a multidimensional approach integrating phenological, morphological, anatomical, and molecular characteristics. Significant interspecific differences were observed in phenological and morphological traits, with P. hookeri having the shortest growth cycle (206 days) and P. zanlanscianense the longest (254 days). The greatest variation was observed in the number of florets and fruits, with coefficients of variation (CV) of 135.92 % and 117.82 %, respectively. This study provides the first descriptions of leaf epidermal, stomatal, and leaf and stem cross-sectional features for P. kingianum var. grandifolium, P. punctatum, and P. hookeri, as well as leaf and stem cross-sectional features for P. filipes. Significant anatomical variation was observed, with stem vascular bundle diameters ranging from 65.60 to 421.06 µm (CV 66.74 %), followed by stomatal width (CV 48.99 %) and aspect ratio (CV 48.16 %). Molecularly, optimized SCoT-PCR identified 12 polymorphic primers, showing abundant polymorphism (96.56 %) in Polygonatum species. Both SCoT-PCR-based and anatomical trait-based clustering showed that P. kingianum var. grandifolium is closely related to P. punctatum, supporting the recognition of P. kingianum var. grandifolium as a separate species. These findings provide a robust basis for species identification and classification of Polygonatum species, improve methodological accuracy, and ensure the quality, safety, and sustainable utilization of Polygonati Rhizoma products.