Pollen morphology and taxonomic significance of Crataegus accessions using scanning electron microscopy

Crataegus Linnaeus (Rosaceae) is a taxonomically complex genus, where macro-morphological traits often show high plasticity, rendering species identification challenging. Evaluating pollen morphology provides valuable stable traits for taxonomic resolution. In the present study, the pollen morphological traits of 19 Crataegus accessions (comprising 13 species, 3 varieties, and 3 unidentified accessions) along with Mespilus germanica were systematically investigated using scanning electron microscopy (SEM) followed by principal component analysis (PCA) and cluster analysis to evaluate their taxonomic significance. All tested pollen grains occurred as large-to-medium monads, appearing 3-lobed circular in polar view and narrowly elliptical in equatorial view, with shapes were prolate, and surfaces displaying striate or rugulate ornamentation. Through PCA results, polar axis (P), equatorial axis (E), size grade, and P × E value were screened as the key traits driving interspecific variation. Hierarchical cluster analysis successfully segregated these accessions into five distinct groups, which is basically consistent with the traditional classification of Crataegus. However, an exceptional case was observed where C. maximowiczii and C. kansuensis were assigned to two different groups, contrasting with previous molecular phylogenies that grouped them into a single clade. Additionally, Mespilus germanica and C. crus-galli were clustered together due to their highly similar elongated large pollen grains, confirming their close intergeneric affinity. Meanwhile, the three unidentified accessions (GSSZ, RR3H, and ZWSZ) formed a completely independent group, supporting their distinct taxonomic status. This study improves the Crataegus taxonomic framework by establishing quantifiable palynological criteria. These key micromorphological drivers not only provide definitive diagnostic keys to resolve ambiguous germplasms, but also offer critical phenotypic benchmarks to reconcile existing morphological-molecular incongruences within the genus.