Reproductive Capacity and Scanning Electron Microscopy (SEM) Analyses of the Micromorphological Surfaces of Three Endemic Satureja Species from Bulgaria.

Satureja pilosa Velen., S. coerulea Janka and S. kitaibelii Wierzb. ex Heuff. are Balkan endemic species, and they are distributed in restricted territories, primarily found in dry grasslands, rocky slopes, and stony habitats. This study presents the results of the first embryological and micromorphological analyses of three Satureja species ( S. pilosa , S. kitaibelii , and S. coerulea ) from the Bulgarian flora. The aim of this study was to establish the features of the male and female reproductive sphere, as well as surface characteristics of leaves, stem, and calyx in order to understand the mode of reproduction, character, size and state of species populations and delimitation. For the embryological study, flowers and flower buds in different developmental stages were collected from plants of natural populations and treated with the classic paraffin method. Reproductive capacity was assessed using the following approaches: (1) acetocarmine test for pollen viability, (2) tetrazolium test (TTZ) for seed viability, and (3) germination test. The surfaces characteristics of leaves, stem, and calyx were analyzed by scanning electron microscopy (SEM). As a result, the study revealed the structures of the male (M) and female (F) generative spheres as well as the processes of gamete development, pollination, and endosperm and embryo formation. It was found that the three Satureja species exhibited a high pollen viability but low seed viability and germination. The SEM analysis showed both common and distinct micromorphology features regarding epidermis, calyx and stem surfaces among the three Satureja species. Notably, the S. coerulea surfaces (leaves, calyx, stem) were clearly distinguishable from the other two Satureja species. Regarding the nutlet surfaces, it was observed that the exocarp surfaces of S. coerulea and S. kitaibelii have a reticulate convex type surface and comprise two types of cells: (1) highly bulging, bubble-like cells; and (2) flat cells with numerous striations. On the other hand, the exocarp of S. pilosa displayed rectangular or polygonal shapes without bubble-like cells, and it had a tabular to slightly convex type surface. Additionally, nutlets (seeds) of both S. coerulea and S. pilosa exhibited distinct papilla formations resembling non-glandular trichomes seen on the ends of the nutlets for S. coerulea and over the entire surface for S. pilosa . The characteristics of the embryological structures and processes, along with the absence of apomixis, characterized the three studied Satureja species as sexually reproducing. The established balanced processes and stable structures contribute to their high reproductive potential and population stability. However, these traits may also decrease their adaptability to environmental changes.