Methodology

Work Package 1 (WP1) identifies the areas where native and naturalized tulips (Tulipa spp.) occur in Greece, in order to organize targeted botanical expeditions for the authorized collection of original propagating material, leaf, root (mycorrhiza) and soil samples.

For the authorized collections, a special collection permit is used which is issued by the Greek Ministry of Energy and Environment. WP1 also includes the process of speeding up procedures by disposing of plant material from the existing seed bank and mother plantations of the Institute of Plant Breeding and Genetic Resources (IPBGR) of the Hellenic Agricultural Organization Demeter, in order to limit the necessary collections from the natural environment and to save time and costs. The original propagating material collected from the wild (seeds, dormant bulbs, and live plants) is maintained under ex situ conservation at the grounds and facilities of the Balkan Botanic Garden of Kroussia and the IPBGR (altitude 650 m and 0 m, respectively) for acclimatization and will be regenerated in various ways to create new mother plants for the 15 species of Greek tulips.

In the WP2, additional botanical expeditions are carried out to ensure ex situ conservation of Greek Tulipa spp. and enhanced natural genetic variability of samples taken in the wild from wild-growing populations. In WP2 all soil, leaf and root (mycorrhiza) samples are analyzed for macro- and micronutrients, they are characterized, and mycorrhizal colonization events are isolated under the microscope and are analysed with molecular techniques. A phylogenetic tree will be generated with all the resulting DNA sequences to assess their genetic affinities and their association with the species and/or regions of origin. Leaf samples will also be analyzed by foliar diagnostics, and the results will be used in the selection of substrates and fertilization regimes. Plant samples from each Greek tulip species will be taxonomically examined and genetically characterized by molecular markers. The nucleotide sequences will be analyzed and will be submitted to the World Gene Bank to obtain special codes (accession numbers) for each species. Their genetic fingerprint will also be used to create a phylogenetic dendrogram documenting genetic affinities between species. The above are aimed to provide solid documentation-characterization of the tulips’ propagation material and, thus, will facilitate the subsequent WPs.

For the Greek tulip species that will be successfully adapted under ex-situ conservation in the IPBGR, special species-specific propagation protocols (WP3) will be developed, in collaboration with the company Athena Nurseries (AN), using precision felitilizers of innovative composition supplied by the project partner Theofrastos. For threatened tulip species (or if it is not possible to develop effective propagation protocols in conventional ways), in vitro propagation techiques will be recruited.

Based on the special propagation protocols that will be piloted in IPBGR, sufficient plant material will be mass-produced for 5 species (200-500 bulbs / species) aimed for the establishment of pilot crop cultivation in the field and/or greenhouses (IBPGR and AN).

For the 5 selected tulip species, precision fertilizers of special composition (foliar sprayed or watered in the soil) will be applied for plant nutrition produced by Theofrastos company from organic plant extracts, rich in amino acids, sugars, B vitamin complexes, nitrogen (non-nitrate, nitrite and ammonia), organic calcium and alternative forms of inorganic copper. These innovative products will be compared with conventional agrochemical fertilizers. During plant development, measurements of photosynthetic parameters will be performed at the grounds of the IPBGR, along with growth measurements and measurements of absorption and concentration of macro- and microelements in different plant parts and in the rhizosphere by the Aristotle University team (AUTH). The soils of the selected plant samples will be analyzed by AUTH in order to simulate them with commercially available means aided by previous experience of IPBGR in the conservation-propagation-cultivation of Greek native species. The analysis of this data will allow the formulation of good practice cultivation protocols. The developed propagation and cultivation protocols will be delivered to the company AN, so they can be applied on a pilot basis under the instructions and supervision of IPBGR.

In WP4, post-harvest treatments of cut tulip flowers will be performed at the University of Thessaly (UT), and the effect of environmental factors (light, temperature, relative humidity) on their life time as cut flowers will be studied. Based on the results obtained, a protocol will be created for the post-harvest treatment of cut tulip flowers that will maximize their life time in the vase and therefore increase their commercial value and their usability as ornamental flowers.

During the project implementation, the necessary conditions for the sustainable management of Greek tulip species will be established leading to 5 new Greek tulip products that will be aimed to enter the market. For the dissemination of results (WP5), the project involves the creation of a custom-made website, social media posts, popularizing of research results in magazines, publications in national and international conferences, as well as publications in reputable peer-reviewed scientific journals (SCI) and participations in flower expositions.