Epidermal morphology and leaflet anatomy of Dioon (Zamiaceae) with comments on climate and environment

Anatomical descriptions can be effective for solving systematic issues, but these studies are relatively scarce for cycads. Therefore, we present here a leaflet and cuticle anatomical study on the genus Dioon, to provide a set of epidermal traits that clarify species delimitation and relationships between species and their habitats. We used standard micro-technique for leaflet sectioning, and cuticular peel preparation for light microscopy. Also, we used the chromium trioxide method for scanning electron microscope observations on cuticles. Measurements were taken on 10 randomly chosen replicates of each cell or tissue type, for each of the leaflets sampled per taxon. Micromorphological variation among species was calculated for each trait. Finally, we reconstructed the ancestral states of the observed epidermal fibre-like cell and pore shapes, by tracing the characters on the species phylogenetic tree of Dioon. We were able to describe the leaflet anatomy, cuticles, and epidermal features for 14 Dioon species. The quantitative analysis was useful to reveal five geographically structured species groups. Character tracing on the phylogenetic tree of Dioon has amplified our current understanding on species relationships with respect to habitats. The presence/absence data suggest that the evolutionary acquisition-deletion of structural shapes is phylogenetically independent, thus climate seems to play a very important role in the variation of cuticular and stomatal traits. Many epidermal traits, especially adaxial cuticle thickness and epistomatal pore width and depth, might be adaptations resulting from a long-term influence of climate, since they appear to have correlation with climatic conditions in relation to their biogeography. We conclude that the variation of all traits are mostly sustained and intrinsic to the species, and are of promising taxonomic value. The combination of the epidermal traits with other characters has potential for taxonomic resolution at species level.