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.

Phytochemical Investigation and Reproductive Capacity of the Bulgarian Endemic Plant Species Marrubium friwaldskyanum Boiss. (Lamiaceae).

Marrubium friwaldskyanum Boiss (Lamiaceae) is a Bulgarian endemic species. Overall, the essential oil (EO) composition of M. friwaldskyanum was different from that of the other Marrubium species reported in the literature. The main EO constituents of M. friwaldskyanum were (E)-caryophyllene, germacrene D, and caryophyllene oxide. The effect of the harvest stage was significant only on α-copaene, (E)-caryophyllene, caryophyllene oxide, and τ-muurolol. The concentration of α-copaene (1.26-1.83% range of the total oil), (E)-caryophyllene (31-41%), caryophyllene oxide (6.4-11.8%), and τ-muurolol (1.3-2.8%) were the highest at 2-3 pair of leaves or before flowering and lower at flowering. The harvest stage did not significantly affect the concentrations of the other six identified EO compounds ß-bourbonene (1.1%), α-humulene (2.8%), germacrene D (23.3%), bicyclogermacrene (2.85%), δ-cadinene (1.1%), and spathulenol (2.8%). In a separate experiment, grinding of the biomass prior to EO extraction had a significant effect only on the concentrations of D-limonene (0.24-3.3%) and bicyclogermacrene (3.6-9.1%). Grinding in water or without water, maceration, and addition of Tween®20 had rather small effects on the EO profile. The identified EO constituents and their mean concentrations in this experiment were (E)-caryophyllene (25.4%), germacrene D (17.6%), caryophyllene oxide (9.1%), spathulenol (6.5%), τ-muurolol (5.0%), carvacrol (3.9%), α-copaene (2.5%), ß-bourbonene (2.5%), δ-cadinene (2.4%), α-humulene (1.8%), and Z-ß-farnesene (1.3%). Embryological studies observed anther and the development of the male gametophyte and ovule and development of the female gametophyte of M. friwaldskyanum. Furthermore, pollen and seed viability assays were conducted, and mass spectrometry-based metabolomics analysis of an extract from shoots revealed the presence of 45 natural products, identified as flavonoids, phenolic acids, and (tri)terpenoids. Overall, the phytochemistry and some of the microscopic analyses distinguished this endemic species from other species in Marrubium.

Fossil nutlets of Boraginaceae from the continental Eocene of Hamada of Méridja (southwestern Algeria): The first fossil of the Borage family in Africa.

PREMISE OF THE STUDY: The Paleogene deposits of the Hamada of Méridja, southwestern Algeria, are currently dated as lower-to-middle Eocene in age based on fossil gastropods and charophytes. Here we report the presence of fruits that can be assigned to the Boraginaceae s.str., apparently representing the first fossil record for this family in Africa, shedding new light on the historical biogeography of this group. METHODS: Microscopic studies of the fossil nutlets were carried out and compared to extant Boraginaceae nutlets, and to types reported in the literature for this family. KEY RESULTS: The fossils are strikingly similar in general size and morphology, particularly in the finer details of the attachment scar and ornamentation, to nutlets of extant representatives of the Boraginaceae tribe Echiochileae, and especially the genus Ogastemma. We believe that these nutlets represent an extinct member of this lineage. CONCLUSIONS: The Ogastemma-like fossils indicate that the Echiochileae, which are most diverse in northern Africa and southwestern Asia, have a long history in this region, dating back to the Eocene. This tribe corresponds to the basal-most clade in Boraginaceae s.str., and the fossils described here agree well with an assumed African origin of the family and the Boraginales I, providing an important additional calibration point for dating the phylogenies of this clade.

EFECTO DE LA LUZ Y DEL ÁCIDO GIBERÉLICO (AG3) EN LA GERMINACIÓN DE Minthostachys mollis KUNTH. GRISEB.(LABIATAE) / Effect of Light and Giberellic Acid (AG3) on the Germination of Minthostachys mollis Kunth. Griseb. (Labiatae)

Minthostachys mollis (Kunth) Griseb., es un arbusto perenne que crece en la región andina de Colombia; es ampliamente utilizada por las comunidades rurales donde es valorado por sus propiedades medicinales. El objetivo de este estudio fue determinar el efecto de la luz (luz roja, luz blanca y luz natural) y del ácido giberélico (0, 5, 15, 25 ppm) en el proceso de germinación de M. mollis. Se encontró inhibición de la germinación bajo condiciones de oscuridad. No se observó efecto de AG3 sobre la germinación. Los porcentajes de germinación fueron superiores a 80% para los tratamientos con luz (semilla fotoblástica positiva), donde el tratamiento con luz roja tuvo el efecto más significativo sobre el proceso de germinación.

Minthostachys mollis (Kunth) Griseb., is a perennial shrub that grows in the Andean region of Colombia; this species is widely used by rural communities where it is valued for its medicinal properties. The aim of this study was to determine the effect of light (red light, white light, and natural light) and gibberellic acid (0, 5, 15, 25 ppm) on the process of germination of M. mollis. We found a strong inhibition of germination under conditions of darkness. We do not observe effect of AG3 on germination. The germination percentages obtained were above 80% (positive photoblastic seeds) for the three proposed lighting treatments where red-light treatment had a most significant effect on the germination process.