The Balkan Botanic Garden of Kroussia (BBGK) is dedicated to the ex situ conservation of native plants of Greece and the Balkans. The BBGK has formulated a conservation strategy for the collection of wild plant material for propagation, prioritizing mainly the endemic, rare, endangered, threatened and vulnerable plants of Europe found in different regions of Greece. Its aim is to contribute to the implementation of Target 8 of the Global and European Strategies for Plant Conservation at local, regional and international scales. In order to (i) define the ecological profile of the in situ requirements preferred and/or tolerated by each selected species, (ii) develop rapid and effective species-specific propagation protocols, and (iii) improve the cultivation of species of conservation concern in BBGK's nurseries and ex situ conservation sections, geographical coordinates and in situ collection data obtained for each taxon were imported into a Geographic Information System environment (GIS). This information was then linked with several digital GIS thematic layers, including topographic, geological, edaphic, climatic, precipitation and temperature data derived from digital databases. Based on this approach, sexual and asexual propagation of plants from the Ionian Islands were conducted and rapid and effective baseline protocols were developed for 29 taxa (species and subspecies); four are presented here in detail and Electronic supplementary material The online version of this article (species-specific ex situ propagation and cultivation guidelines are given. Most of the taxa originating from the Ionian Islands were propagated by cuttings (55.2%) or seeds (34.5%), while the rest were propagated by root division at a rate from 1.7 to 2. The first round of propagation achieved a success rate ranging from 15 to 50% for 3 taxa, from 60 to 80% for 8 taxa and from more than 80 to 100% for 16 taxa, while the ex situ cultivation of the wild and propagated plant material has, so far, been successful. The application of GIS exemplified here presents a sensible and invaluable tool with a broad-scale potential in enhancing the prospects of the ex situ conservation of priority species collected from diverse environmental conditions in man-made habitats such as botanic gardens.

Alkannin and shikonin (A/S) are enantiomeric naphthoquinones produced in the roots of certain plants from the Boraginaceae family such as Lithospermum spp. and Alkanna spp. They possess antimicrobial, anti-tumoral and wound healing properties. The production of secondary metabolites by Alkanna tinctoria might be influenced by its endomicrobiome. To study the interaction between this medicinal plant and its bacterial endophytes, we isolated bacteria from the roots of wild growing Alkanna tinctoria collected near to Athens and Thessaloniki in Greece. Representative strains selected by MALDI-TOF mass spectrometry were identified by partial 16S rRNA gene sequence analysis. In total, 197 distinct phylotypes of endophytic bacteria were detected. The most abundant genera recovered were Pseudomonas, Xanthomonas, Variovorax, Bacillus, Inquilinus, Pantoea, and Stenotrophomonas. Several bacteria were then tested in vitro for their plant growth promoting activity and the production of cell-wall degrading enzymes. Strains of Pseudomonas, Pantoea, Bacillus and Inquilinus showed positive plant growth properties whereas those of Bacteroidetes and Rhizobiaceae showed pectinase and cellulase activity in vitro. In addition, bacterial responses to alkannin and shikonin were investigated through resistance assays. Gram negative bacteria were found to be resistant to the antimicrobial properties of A/S, whereas the Gram positives were sensitive. A selection of bacteria was then tested for the ability to induce A/S production in hairy roots culture of A. tinctoria. Four strains belonging to Chitinophaga sp., Allorhizobium sp., Duganella sp., and Micromonospora sp., resulted in significantly more A/S in the hairy roots than the uninoculated control. As these bacteria can produce cell-wall degrading enzymes, we hypothesize that the A/S induction may be related with the plant-bacteria interaction during colonization.

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Crithmum maritimum (Apiaceae), a perennial halophyte native in Greece, could be used as an alternative culture at problematic soils. It presents significant economical potentials as its essential oils are in high demand from the medicinal and cosmetic industry. The response of the species on in vitro conditions was studied. MS proved to be the most effective of the basal media tested for in vitro adventitious shoot production, resulting in significantly increased number of new microshoots/explant and higher shoots. 6-Benzyladenine (BA) at 2.5 lM increased shoot proliferation. The combination of a-Naphthaleneacetic acid (NAA) (1-2.5 lM) with BA (2.5 lM) had a positive influence at simultaneous proliferation and rooting resulting in high rooting percentage (82.5-95%) and increased number of roots. Rooting percentage reached 100% and number of roots increased significantly when 0.5 lM and 1 lM IBA was combined with MS and full strength MS. The in vitro response to salinity stress (0-300 mM NaCl) was also tested. Shoot proliferation was gradually reduced at higher concentrations of NaCl but shoot height was enhanced. Acclimatization procedure was successful.

Conservation and sustainable exploitation of threatened endemic plants with medicinal and/or horticultural/ornamental value can be achieved through the development of effective propagation protocols. After unveiling the bioclimatic preferences of Carlina diae (Asteraceae) with geographic information systems (GIS), four propagation trials were conducted using seeds of this endangered local Cretan endemic for in vivo and in vitro germination, as well as seasonal vegetative propagation trials (softwood cuttings) and micropropagation (nodal explants). Seed germination was accomplished at a level of 77–90% in vivo (30 days) and 96% in vitro (10 days) using an MS medium with 2.9 μM gibberellic acid (GA3). The optimum treatments for cuttings’ rooting were 1000 and 2000 ppm indole-3-butyric acid (IBA) (11–16 roots, 2–3 cm long, 100% rooting) within 40 days in mist. In vitro shoot propagation exhibited a 2.8 proliferation rate after six successive subcultures on an MS medium with 2.9 μM GA3. Both ex vitro rooting and acclimatization were successful in 40 days, with 96% microshoot rooting and an equal survival rate. The GIS-facilitated effective species-specific propagation protocols developed in this study can consolidate the perspective of successful re-introduction of ex situ-raised material of C. diae into wild habitats and may serve its sustainable exploitation for high-added value ornamental products.