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Immortelle (Helichrysum italicum /Roth./ G. Don) is a perennial plant species of the genus Helichrysum and the family Asteraceae. It is spread in the Mediterranean region inhabiting the sea coast as well as the mountains up to 2, 200 meters above sea level (Galbany-Casals et al., 2011). Immortelle is used in the cosmetic, pharmaceutical, and food industries. It is an economically important medicinal and aromatic plant in Croatia, and in recent years the demand for H. italicum essential oil has increased. This study aimed to determine the biochemical diversity of 18 immortelle populations in Croatia.

Studying the population-genetic and phylogeographic structures of a representative species of a particular geographical region can not only provide us with information regarding its evolutionary history, but also improve our understanding of the evolutionary processes underlying the patterns of species diversity in that area. By analysing eight highly polymorphic microsatellite loci and two chloroplast DNA regions, we have investigated the influence of Pleistocene climate fluctuations on the evolutionary history of Salvia officinalis L. (common sage). The populations with the highest genetic diversity were located in the central parts of the Balkan distribution range. A large group of closely related haplotypes was distributed throughout the Balkans and the central Apennines, while the private lineage occupied the southern Apennines. In addition, two highly differentiated lineages were scattered only over the Balkans. The results suggest that a single refugium of the studied species from the last glacial period was located in the central part of the range in the Balkans. Numerous microrefugia, probably spanning several glaciation cycles, were scattered across the Balkans, while colonisation of the Apennines from the Balkans occurred at least on two occasions.

The Adriatic islands in Croatia, usually divided into two archipelagos – the Kvarner and Dalmatian islands – is one of the largest groups of islands in Europe. Over 40 islands are still inhabited. Unfortunately the traditional use of medicinal plants was never properly documented there. Our data comes from 343 interviews carried out in 36 islands, including the 15 largest islands of the archipelago. The medicinal plants are mainly used to make herbal infusions or decoctions, occasionally medicinal liqueurs, syrups, compresses, or juices squeezed out of raw plants. We recorded the use of 146 taxa, among them 131 with at least one medicinal purpose and 15 only for tea. The frequency curve of use is relatively steep – several plants are used very frequently and most are reported only by one or two informants, which can be explained both by the large geographical spread of the area, and even more so by the devolution of local knowledge and disappearance of gathering practices due to specialization in tourism, modernization and depopulation. Most of the gathered plants already occur in ancient and medieval herbals and are a part of the pan-Mediterranean pharmacopoeia.

Salvia officinalis is a perennial species, native and endemic to the Western Balkans and the Apennine Peninsula. Due to its medicinal and aromatic properties, it is used in pharmaceutical, cosmetic, and food industries. The main objectives of the study were to infer the genetic structure of S. officinalis populations in the northern and central parts of the eastern Adriatic coast, to detect the phylogeographical barriers among the putative microrefugia and to assess the genetic diversity among the resulting ancestral clusters. Twenty-five populations were assessed using amplified fragment length polymorphism markers. High polymorphism and high diversity within populations were typical for this outcrossing long-lived species. The Fitch– Margoliash tree based on Nei’s genetic distance matrix showed that most of the populations tended to group in accordance with the geographical position of their collecting sites. Spatial analysis of the genetic diversity revealed a typical pattern of isolation by distance. Very low overall among-population differentiation and detection of only three private alleles indicate that there has been high gene flow among populations. By using Bayesian Analysis of Population Structure on population level, two distinct ancestral clusters were obtained. It is likely that these two ancestral clusters were separated for a longer period by Pleistocene glaciation, although the subsequent fast recolonization resulted in diminished genetic differences. High rarity of northern and southern populations of the investigated area indicates that S. officinalis presumably survived in both northern and southern microrefugia and expanded from there resulting in secondary contact zones, characterized by lower rarity and equal genetic diversity.

An ethnobotanical survey was carried out in the Knin area (northern Dalmatia, Croatia) with the aim of recording traditional plant use by the local (native) people and contributing to the knowledge of plant biodiversity in the investigated area. Ethnobotanical data were collected by interviewing 40 local people at 17 locations. Data on 123 plant taxa (on average 21 taxa per interview) and 122 unique medicinal and 18 food uses in the local community were recorded. Prevalent medicinal uses of the recorded taxa were the treatment of digestive, respiratory, cardiovascular disorders, and injuries. The most commonly used plant parts gathered were leaves (35%) and flowers (20%). The most commonly collected plants in the investigated area were: Urtica dioica, Thymus longicaulis, Sambucus nigra, and Hypericum perforatum.

Background: Archipelagos of islands have played an important role in shaping some of the paradigms of biology, including the theory of the evolution of species. Later, their importance in biology was further emphasised by the theory of island biogeography, which contributed to a better understanding of the shaping of species richness not only on real islands, but on isolated habitat islands as well. Although ethnobotany is a well- established discipline, patterns of knowledge about plant uses in archipelagos have never been quantitatively analysed, and the whole concept has been only briefly mentioned in the ethnobiological context. The aim of our study was to record which taxa of wild vegetables have been consumed in the Adriatic islands and to establish if such variables as island size, population size, flora or its isolation are correlated with the number of wild vegetables used. Methods: We interviewed 225 people (15 from each island). Results: Altogether, the use of 89 species of wild vegetables has been recorded. The largest number of wild vegetables is eaten on the islands of Korčula, Vis and Šolta, and the lowest on Ugljan, Cres and Dugi Otok. The studied independent variables had a small and statistically not significant effect on the wild vegetable list length. The most visible effect was an increasing trend from north-west to south- east, overrunning the typical biogeographical island patterns. Moreover, one of the large and well-populated islands, Korčula, showed an ‘unusually’ high level of wild vegetable use. We hypothesise that the current use of so many species on this island has been maintained by the inhabitants’ awareness that they are the holders of relic knowledge, an awareness reiterated by ethnographic and popular publications, as well as a strong history of famine. The most interesting edible species used in the Adriatic islands are Bunium alpinum, Cytinus hypocystis (both mainly on Pašman), Lotus edulis (on Vis) and Posidonia oceanica (on Vis and Korčula). Conclusions: The recorded relationships between the demographic and geographical features of the islands were statistically not significant. We assume that cultural and historical factors diversifying the use of plants in particular islands are stronger than the above-mentioned measurable variables.

Helichrysum italicum (Roth) G. Don is a thermophilus perennial from family Asteraceae. Due to versatile biological activities and applications, the interest in Helichrysum italicum scientific research and use has increased in the last couple of decades. Consequently, due to the increased industrial demand its plantation farming experienced a huge expansion in Eastern European Mediterranean countries. The aim of this review article was to present the available data on Helichrysum italicum and to identify the gaps in the knowledge, and openings for future potential investigations and contributions to the topic. The review of the Scopus database was performed and scientific studies from 1990 to the most recent publications in 2019 were summarized. The article includes the last reviewed taxonomy, detailed description of morphological characteristics, geographical distribution, uses, chemical diversity, biological activity (antimicrobial, antiviral, antioxidant, anti-inflammatory, repellent, and insecticidal activity), and genetic diversity. Additionally, some other potential uses are described. The majority of the available scientific data is focused on the chemical composition of the essential oil, however, little is known about the cultivation management of Helichrysum italicum that includes sowing requirements, seedling production, plant density, fertilization and nutrition requirements, harvesting, diseases, insects, and weed control. Genetic information on Helichrysum italicum is also scarce, but needed for future breeding programs of Helichrysum italicum and selection of new cultivars.

Dalmatian pyrethrum (Tanacetum cinerariifolium (TREVIR.) SCH.BIP.) is a plant species endemic to the east Adriatic coast. The bioactive substance of Dalmatian pyrethrum is a natural insecticide, pyrethrin, a mixture of six active components (pyrethrins I and II, cinerins I and II, and jasmolins I and II). The insecticidal potential of pyrethrin was recognized decades ago, and dried and ground flowers have traditionally been used in Croatian agriculture and households. A total of 25 Dalmatian pyrethrum populations from Croatia were studied to determine the pyrethrin content and composition, and to identify distinct chemotypes. The total pyrethrin content ranged from 0.36 to 1.30% (dry flower weight; DW) and the pyrethrin I/pyrethrin II ratio ranged from 0.64 to 3.33%. The statistical analyses revealed that the correlations between the percentage of pyrethrin I and of all the other components were significant and negative. The total pyrethrin content was positively correlated with the percentage of pyrethrin I and negatively correlated with cinerin II. The multivariate analysis of the chemical variability enabled the identification of five chemotypes among 25 Dalmatian pyrethrum populations. The chemical characterization of indigenous Dalmatian pyrethrum populations may serve as a good background for future breeding and agricultural exploitation.

The aim of this study was to document the use of wild foods in two locations in the coastal part of Croatia – on the island of Krk and in the Poljica area, near Split. We chose these places as they have historical data on plant use (1900 and 1903 respectively). We carried out 67 interviews in Poljica and 55 interviews in Krk to estimate the current use and knowledge of wild foods. Altogether, 80 species of wild food and herbal tea species of plants were recorded in Poljica and 76 in Krk. On average, 13.2 species were listed by per interview in Poljica, including 7.1 species of wild vegetables, and in Krk 14.6 species, including 7.9 species of wild vegetables. Out of the list of plant names recorded in the past, in Krk, we identified 82% of the taxa and in Poljica 86%.

Dalmatian sage (Salvia officinalis L., Lamiaceae) is a well-known aromatic and medicinal Mediterranean plant that is native in coastal regions of the western Balkan and southern Apennine Peninsulas and is commonly cultivated worldwide. It is widely used in the food, pharmaceutical and cosmetic industries. Knowledge of its genetic diversity and spatiotemporal patterns is important for plant breeding programmes and conservation. We used eight microsatellite markers to investigate evolutionary history of indigenous populations as well as genetic diversity and structure within and among indigenous and cultivated/naturalised populations distributed across the Balkan Peninsula. The results showed a clear separation between the indigenous and cultivated/naturalised groups, with the cultivated material originating from one restricted geographical area. Most of the genetic diversity in both groups was attributable to differences among individuals within populations, although spatial genetic analysis of indigenous populations indicated the existence of isolation by distance. Geographical structuring of indigenous populations was found using clustering analysis, with three sub-clusters of indigenous populations. The highest level of gene diversity and the greatest number of private alleles were found in the central part of the eastern Adriatic coast, while decreases in gene diversity and number of private alleles were evident towards the northwestern Adriatic coast and southern and eastern regions of the Balkan Peninsula. The results of Ecological Niche Modelling during Last Glacial Maximum and Approximate Bayesian Computation suggested two plausible evolutionary trajectories: 1) the species survived in the glacial refugium in southern Adriatic coastal region with subsequent colonization events towards northern, eastern and southern Balkan Peninsula; 2) species survived in several refugia exhibiting concurrent divergence into three genetic groups. The insight into genetic diversity and structure also provide the baseline data for conservation of S. officinalis genetic resources valuable for future breeding programmes.

Nine new microsatellite markers (SSR) were isolated from Salvia officinalis L. A total of 125 alleles, with 8 to 21 alleles per locus, were detected in a natural population from the east Adriatic coast. The observed heterozygosity, expected heterozygosity, and polymorphic information content ranged from 0.46 to 0.83, 0.73 to 0.93 and 0.70 to 0.92, respectively. New microsatellite markers, as well as previously published markers, were tested for cross-amplification in Salvia brachyodon Vandas, a narrow endemic species known to be present in only two localities on the Balkan Peninsula. Out of 30 microsatellite markers tested on the natural S. brachyodon population, 15 were successfully amplified. To obtain evidence of recent bottleneck events in the populations of both species, observed genetic diversity (HE) was compared to the expected genetic diversity at mutation-drift equilibrium (HEQ) and calculated from the observed number of alleles using a two-phased mutation model (TPM). Recent bottleneck events were detected only in the S. brachyodon population. This result suggests the need to reconsider the current threat category of this endemic species.

Grapevine ( Vitis vinifera L.) and two selected weeds from Mediterranean Croatian vineyards (Plantago lanceolata L. and Tanacetum cinerariifolium (Trevir.) Sch.Bip.) were examined in pot culture experiments, individually or when combined, to see whether multiple hosts influenced the formation of the symbiosis with arbuscular mycorrhizal fungi (AMF). The results after six-month period showed that plant identity and density significantly influenced development of mycorrhizal intra- and extraradical mycelium and/or sporulation. Grapevine and T. cinerariifolium individually and in combination resulted in a greater development of arbuscular mycorrhizae in terms of spore production, extraradical mycelium length and root colonization compared with pots containing P. lanceolata. Herbaceous weed species seemed to promote a different set of dominant AMF, potentially providing a wider spectrum of AMF for colonising grapevine roots. This indicates the value of encouraging host plant diversity in vineyards. AMF sequences obtained in this study are the first data reported for soils in Croatia.

Nine new microsatellite markers (SSR) were isolated from Salvia officinalis L. A total of 125 alleles, with 8 to 21 alleles per locus, were detected in a natural population from the east Adriatic coast. The observed heterozygosity, expected heterozygosity, and polymorphic information content ranged from 0.46 to 0.83, 0.73 to 0.93 and 0.70 to 0.92, respectively. New microsatellite markers, as well as previously published markers, were tested for cross-amplification in Salvia brachyodon Vandas, a narrow endemic species known to be present in only two localities on the Balkan Peninsula. Out of 30 microsatellite markers tested on the natural S. brachyodon population, 15 were successfully amplified. To obtain evidence of recent bottleneck events in the populations of both species, observed genetic diversity (H$_E$) was compared to the expected genetic diversity at mutation-drift equilibrium (H$_{EQ}$) and calculated from the observed number of alleles using a two-phased mutation model (TPM). Recent bottleneck events were detected only in the S. brachyodon population. This result suggests the need to reconsider the current threat category of this endemic species.

The use of medicinal and aromatic plants (MAP) in Croatia has a very long tradition. Commercial gathering was economically important source of income in many Croatian regions. However, gathering in the wildness has a negative impact on biodiversity, collected plant material is not uniform and its price is significantly lower. Current production of medicinal and aromatic crops in Croatia is very limited. The MAP cultivation on family farms is usually marginal activity. MAP producers and processors generally agree that marketing opportunities do exist in case of a number of species. Natural MAP populations in Croatia show great biodiversity in morphological, biochemical and genetic level. The assessment of biodiversity is a starting point for efficient conservation of plant genetic resources and its use in plant breeding programmes. The main aim of the management of plant genetic resources is conservation, characterization, evaluation and documentation of the existing genetic biodiversity. For the purpose of conservation of these valuable genetic resources, The Collection of Medicinal and Aromatic Plants has been established at the Faculty of Agriculture, University of Zagreb, Croatia. Characterization and evaluation of accessions in plant genebanks is traditionally based on morphological traits. Currently, the analysis of target species including Dalmatian sage (Salvia officinalis L.), Dalmatian pyrethrum (Tanacetum cinerariifolium/Trevir./Sch. Bip.) and basils (Ocimum spp.) has been carried out on morphological, biochemical and genetic level.

Essential oils of 25 indigenous populations of Dalmatian sage (Salvia officinalis L.) that represent nearly half of native distribution area of the species were analyzed. Plantlets collected from wild populations were grown in the same field under the same environmental conditions and then sampled for essential-oil analysis. The yield of essential oil ranged from 1.93 to 3.70% with average of 2.83%. Among the 62 compounds detected, eight (cis-thujone, camphor, trans-thujone, 1,8-cineole, beta-pinene, camphene, borneol, and bornyl acetate) formed 78.1387.33% of essential oils of individual populations. Strong positive correlations were observed between camphor and beta-pinene,beta-pinene and borneol, as well as between borneol and bornyl acetate. The strongest negative correlation was detected between camphor and trans-thujone. Principal component analysis (PCA) on the basis of eight main compounds showed that first main component separated populations with high thujone content, from those rich in camphor, while the second component separated populations rich in cis-thujone from those rich in trans-thujone. Cluster analysis (CA) led to the identification of three chemotypes of S. officinalis populations: cis-thujone; trans-tujone, and camphor/beta-pinene/borneol/bornyl acetate. We propose that differences in essential oils of 25 populations are mostly genetically controlled, since potential environmental factors were controlled in this study.

The multiplication rate of eleven gerbera cultivars was tested on a MS medium that contained 4.5% sucrose, 80 mg L-1 adenine sulphate, 100 mg L-1 L-tyrosine. 0.5 mg L-1 IAA and 2.0 mg L-1 N6-benzyladenipe (BA). The highest multiplication rates (8 shoots) per inoculum in a 4 week period were shown by cvs. Ansofie, Terracerise and Lablinel; cvs. Maria, Shanghai and Fresultane had moderate rates (6—7 shoots), and cvs. Croduro, Raisa, Fredigor, Terramaxima and Fredibel had the lowest rates (4—5 shoots). Plant- lets rooted effectively in three weeks on MS salts with 3% sucrose and 2.0 mg L-1 IAA, and successfully acclimatized to the greenhouse conditions., Istražen je utjecaj genetske osnove na stopu umnažanja jedanaest sorti gerbere na hranidbenoj podlozi MS s dodatkom 4,5% saharoze, 80 mg LT1 adenin sulfata, 100 mg L-1 L-tirozina, 0,5 mg L-1 IAA i 2,0 mg L-1 N6-benziladenina (BA). Najvišu stopu multiplikacije po presadnici (8 izdanaka) u vremenu od 4 tjedna imale su sorte Ansofie, Terracerise i Lablinel; sorte Maria, Shanghai i Fresultane umjereno su se multiplicirale (6—7 izdanaka) a sorte Croduro, Raisa, Fredigor, Terramaxima i Fredibel imale su najslabiju stopu umnažanja (4—5 izdanaka). Biljčice su se u tri tjedna dobro zakorijenile na podlozi MS-soli s 3°/o saharoze i 2,0 mg L-1 IAA i uspješno prilagodile uvjetima u stakleniku.