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Balkan Peninsula is one of the diversity centres for the morphologically highly variable and taxonomically inconsistently treated Dianthus sylvestris Wulfen. The current taxonomic concept provided in the Flora Europaea is based on names published by Hayek in 1924. One of the overall aims of our comprehensive study combining phylogeographic methods (RADseq), morphometry, environmental niche analyses and genome size estimates, is to explore the intraspecific relationships within the Balkan populations of D. sylvestris, and to provide the revised taxonomy. Our results suggest two morphological entities, the north-western and the south-eastern group of populations, distinct by the epicalyx scales shape, calyx length and petal denticulation. These entities correspond to the two identified genetic clusters, with the split occurring around the Neretva River valley. The genetic structure does not support current taxonomy, except for D. sylvestris subsp. tergestinus (Rchb.) Hayek. Given that the type specimen of D. sylvestris subsp. nodosus (Tausch) Hayek is not known, and its diagnostic character, puberulent lower part of the stem, turned out to be variable within the population level, we suggest leaving this form of variability within D. sylvestris s.l. without taxonomic designation. The holotypes of two south Balkan subspecies, D. sylvestris subsp. alboroseus F.K. Mey. and D. sylvestris subsp. kozjakensis Micevski, are highly similar and based on our results they belong in the same morphometric and genetic clusters. Several populations from the south eastern group show morphologically intermediate forms between D. sylvestris subsp. alboroseus and D. sylvestris subsp. bertisceus Rech. f. and cannot be clearly assigned to subspecies. Finally, D. sylvestris subsp. bertisceus showed the greatest genetic admixture, as well as problematic morphological distinction, especially when populations from the Alps are considered.

Aurinia is an exclusively diploid genus comprising of seven species with the centre of origin and diversity in south-eastern Europe. Aurinia saxatilis is the most widely distributed species ranging from the central, eastern and southern Balkan Peninsula and adjacent shores of the western Asia Minor and the southern Apennine Peninsula, over the Carpathians to central Europe and growing on rocky grounds and dry soils. Due to its morphological variability it has been divided into three subspecies: typical A. saxatilis subsp. saxatilis is widespread in central and south- eastern Europe, subsp. orientalis is distributed in the Balkan Peninsula, the southern Carpathians and the western Anatolia, whereas subsp. megalocarpa is limited to the southern Apennine Peninsula and the Aegean region. The aim of this study was to provide insights into spatiotemporal diversification of the species by exploring its evolutionary dynamics and phylogeographic patterns using a combination of phylogenomic (RADseq), and phylogenetic (plastid ndhF) data as well as species distribution models. The inferred phylogenies retrieved three main geographically distinct lineages. The southern lineage is distributed throughout the Aegean area, southern Balkan Peninsula and southern Apennine Peninsula, and corresponds to the species main distribution area during the LGM. The eastern lineage extends from the eastern Balkan Peninsula over the Carpathians towards central Europe, while the central lineage occupies the central Balkan Peninsula. Molecular dating places diversification of all three lineages into the early to middle Pleistocene, indicating their long term independent evolutionary trajectories. Moreover, our data suggest an early divergence and stable in situ persistence of the southernmost populations, whereas the central Balkan and Carpathian populations experienced more complex range dynamics triggered by Pleistocene climatic oscillations. Finally, the extant genetic variation is congruent with the taxonomic separation of subsp. saxatilis and subsp. orientalis, whereas the taxonomic value of subsp. megalocarpa remains doubtful.

Festuca L. represents one of the largest known paraphyletic genera systematically anchored in subtribe Loliinae whose distribution has been proven globally. In this study, we focused on F. varia complex within fine-leaved fescues and with emphasis on F. bosniaca populations, in order to provide insights into its diversification according to ploidy level and geographical affiliation. We sampled a total of 612 individuals divided in 169 populations from various locations in north-western and south-eastern Balkan Peninsula, Apennines and the Alps and complemented with samples from 135 herbarium vouchers. Nuclear ITS and two plastid trnT-trnL and trnL-trnF regions were sequenced and phylogenetically analysed. The inferred phylogenies support the placement of F. varia complex in basal position within fine-leaved fescues and reveal several supported groups within the complex. Festuca bosniaca populations are divided into five major lineages showing a geographical pattern. Two established lineages largely correspond to the F. bosniaca subsp. bosniaca, and F. bosniaca subsp. pirinensis, while the other three require additional research. The relationships of different taxa in the complex remain, however, uncertain due to incongruences between nuclear and plastid trees. This study contributes to knowledge on F. varia complex and indicates that at this level the traditional morphologically based taxonomy is of limited use for recognizing and defining monophyletic groups.

The mustard family (Brassicaceae) is a scientifically and economically important family, containing the model plant Arabidopsis thaliana and numerous crop species that feed billions worldwide. Despite its relevance, most published family phylogenies are incompletely sampled, generally contain massive polytomies, and/or show incongruent topologies between datasets. Here, we present the most complete Brassicaceae genus-level family phylogenies to date (Brassicaceae Tree of Life, or BrassiToL) based on nuclear (>1,000 genes, almost all 349 genera and 53 tribes) and plastome (60 genes, 79% of the genera, all tribes) data. We found cytonuclear discordance between nuclear and plastome-derived phylogenies, which is likely a result of rampant hybridisation among closely and more distantly related species, and highlight rogue taxa. To evaluate the impact of this rampant hybridisation on the nuclear phylogeny reconstruction, we performed four different sampling routines that increasingly removed variable data and likely paralogs. Our resulting cleaned subset of 297 nuclear genes revealed high support for the tribes, while support for the main lineages remained relatively low. Calibration based on the 20 most clock-like nuclear genes suggests a late Eocene to late Oligocene ‘icehouse origin’ of the family. Finally, we propose five new or re-established tribes, including the recognition of Arabidopsideae, a monotypic tribe to accommodate Arabidopsis. With a worldwide community of thousands of researchers working on this family, our new, densely sampled family phylogeny will be an indispensable tool to further highlight Brassicaceae as an excellent model family for studies on biodiversity and plant biology., bioRxiv

Dianthus sylvestris includes ten subspecies, whose delimitation is challenging and taxonomically inconsistent. Its main diversity centres are the Balkan and the Apennine Peninsulas where the taxa grow in various habitats from the Mediterranean coast to the alpine belt. This study is focused on the Balkan populations where six subspecies occur. In total 162 populations were sampled across the Balkans: 97 were included in detailed morphometric analyses, for 162, including 28 populations from the Alps, relative genome size (RGS) analysed was estimated using flow cytometry, while all Balkan populations were used in the environmental niche analyses. The integrated results of these analyses represent a first step toward the revised taxonomic treatment for D. sylvestris on the Balkan Peninsula. The results reveal that morphological variation gradually changes with environment and geography and does not correspond to described subspecies. The results suggest two morphologically distinct entities, the north-western and the south-eastern groups of populations, distinct by the shape of epicalyx scales, calyx length and petal denticulation. The quantitative assessment of niche overlap, equivalency and similarity of D. sylvestris subspecies indicated, that niche similarity was more common than niche divergence. Longitude was a major geographic predictor associated with morphological variability, where the changes in cauline leaf length and shape of epicalyx scales reflected a gradual transition from the coastal Mediterranean to the alpine environment. RGS analyses revealed that the majority of populations is diploid (2n=2x=30), but also spatially restricted occurrence of tetraploid populations (2n=4x=60) disclosed polyploidization in the NW Balkans. The tetraploid populations are morphologically similar to their diploid counterparts occurring in the same area indicating their autopolyploid origin. The populations at the southern distribution margin in the Balkan Peninsula, as well as those in the central and western parts of the Alps, exhibit higher RGS, likely corresponding to discrete evolutionary lineages.

Dianthus sylvestris s.l. is a group comprising morphologically highly variable taxa distributed in central Mediterranean, with the highest taxonomic diversity described in Italy. Another diversity centre is the Balkan Peninsula, from where six taxa mostly treated as subspecies of D. sylvestris have been described: subsp. alboroseus, subsp. bertisceus, subsp. kozjakensis, subsp. nodosus, subsp. sylvestris and subsp. tergestinus. Their identification is challenging due to great morphological variability and complex patterns of variation with respect to ecological preferences. In an ongoing study, we are combining an array of methods ranging from nomenclatural revision and morphometrics, over cytological and genome size estimation, to environmental niche analyses and population genetic analyses to disentangle relationships within D. sylvestris s.l. on the Balkan Peninsula. The morphometric analyses revealed a continuous variability of quantitative morphological characters and an absence of clear-cut qualitative morphological differences among the subspecies. In addition, the niche overlap assessment indicated that niche similarity is more common among the six subspecies, than niche divergence. Population genetics analyses suggest that D. sylvestris subsp. tergestinus presents a separate evolutionary lineage, while other populations on the Balkan Peninsula form a cline of genetic differentiation in the SE-NW direction. Finally, the relative genome size estimation using flow cytometry, with confirmatory chromosome counts, revealed polyploidisation in the NW Balkan Peninsula, as the tetraploid populations (2n =4x =60) are restricted to the Istrian Peninsula and Kvarner in Croatia and Italy. All the remaining Balkan populations were diploid (2n =2x =30).

Contemporary patterns of genomic variation in plant species often reflect past demographic processes linked to climate events such as Pleistocene glaciations. Dianthus sylvestris s.l. comprises a number of taxa distributed across southern and south- eastern Europe. We here used RAD-seq SNP data to unravel genomic variation in 165 populations, including all six currently described subspecies occurring in the Balkan Peninsula. Population genetic structure using Bayesian clustering revealed a north- western (NW) and south-eastern (SE) group, while an alternative model-free approach identified five genetic groups. We detected significant isolation by distance (IBD) within our populations, though with a visible discontinuity, and additional genetic connectivity analyses confirmed a sharp break between the two main groups of populations from NW and SE, suggesting that these groups derive from separate evolutionary lineages. Migration analyses revealed highest rates of gene flow between two subgroups of the NW cluster, while lower rates of gene flow were inferred between three subgroups of the SE cluster. The SE group showed higher inbreeding values compared to the NW group. Species distribution modelling (SDM) pointed to suitable habitats during the last glacial maximum (LGM) along the whole eastern Adriatic coast with higher suitability predicted in the south of the Balkans compared to the north. Overall, patterns of genomic variation and SDM suggest that D. sylvestris in the Balkans survived the Pleistocene glaciations in two separate glacial refugia located along the eastern Adriatic coast, with secondary contact of the two lineages in the area of Montenegro where admixed populations occur.

Wide-range species often exhibit morphological variability that is mostly shaped by ecological and evolutionary processes, and the resulting diversity is frequently reflected in infraspecific taxonomy. The Dianthus sylvestris complex on the Balkan Peninsula encompasses six subspecies: Dianthus sylvestris subsp. alboroseus, subsp. bertisceus, subsp. kozjakensis, subsp. nodosus, subsp. sylvestris and subsp. tergestinus. Their identification is challenging due to great morphological variability between subspecies and complex patterns of variation with respect to ecological preferences. Here, 97 populations of D. sylvestris were evaluated regarding the variability of 27 morphological characters, and 20 environmental variables were used to identify patterns of morphological variability along environmental gradients. Number and shape of epicalyx scales, calyx length and petal denticulation turned out to capture the greatest extent of morphological variability between populations. Dianthus sylvestris subsp. nodosus, subsp. sylvestris and subsp. tergestinus have similar morphologies and share parts of their geographic distributions and cannot be clearly differentiated using morphological characters. However, a different flowering period differentiates subsp. tergestinus. By quantitatively assessing niche overlap, equivalency and similarity of the subspecies, our results indicate that niche similarity is more common than divergence. Environment, rather than geography, plays a potentially more important role in shaping the morphological variability of this complex.

Delimiting species only on morphology may lead to taxonomic inflation, biasing conservation studies and actions, particularly in organisms recently diversified. For instance, the central Mediterranean Dianthus virgineus L. complex belongs to a lineage that evolved rapidly in Eurasia, and is characterized by high phenotypic variation. In the central Mediterranean, 21 taxa are currently accepted, albeit solely described according to qualitative morphological and phenological variation. Our objective is to apply an integrative taxonomic approach, including morphometry, karyology, and genetics, to test these taxonomic hypotheses. We genotyped thousands of genome-wide biallelic, unlinked SNPs from 123 populations from southern France, peninsular Italy, Corsica, Sardinia, and Sicily, through ddRAD sequencing and conducted an explorative PCA and STRUCTURE analysis to detect genetic differentiation. All across the study area there is an overall low genetic variability, albeit structured according to geographical distribution of populations. In particular, we detected a NW-SE cline from southern France to Calabria, and a second, orthogonal cline from central Italy to Sardinia through the Tuscan Archipelago and Corsica. Conversely, plants from Sicily form a genetic cluster that does not appear to follow clinal patterns recovered in the Italian Peninsula. We deduce that the number of genetically recognizable groups of the D. virgineus complex in central Mediterranean is much lower than the current taxonomic hypotheses, highlighting the importance of an integrative taxonomic approach to better circumscribe systematic units. However, taxonomic conclusions await thorough morphometric and karyological analyses, which are currently underway.

The genus Festuca L. (Poaceae) comprises perennial grasses with leaf blades mostly rolled or conduplicate ; inflorescence in form of a panicle composed of two- to several-flowered spikelets ; with specific upper glume, lemma and awn morphology. Festuca bosniaca and its closely related species within F. varia complex occupy large area and diverse habitats in southern Europe, thus exhibiting a variety of morphological characters in leaf and panicle morphology. Total of 112 individuals divided through 34 populations representing seven species (F. adamovicii, F. bosniaca, F. calva, F. cyllenica, F. eskia, F. varia and F. versicolor) were collected over the Balkan and the Apennine Peninsulas and the Alps. Most important diagnostic morphometric characters (anatomical and habitus) considering leaf and panicle segment were measured. Quantitative morphometric characters between closely related species were examined with exploratory data analysis and ANOVA, Tukey post hoc and Kruskal- Wallis were performed in order to obtain significant (p

Festuca L. is a highly diverse genus of grasses and its perennial species constitute important grassland vegetation worldwide. Recent phylogenetic studies revealed two main evolutionary lineages corresponding to broad- leaved and fine-leaved species. Within fine-leaved fescues, the earliest diverging group is F. sect. Eskia, known also as F. varia complex, which includes approximately 20 species. The taxonomic complexity of the group is especially evident in the Balkan Peninsula, from where several narrowly endemic taxa and a number of lower taxonomic entities have been described. The aim of the present study is to explore genome size variation and the incidence of polyploidy within the representatives of F. varia complex on the Balkan and Apennine Peninsulas based on comprehensive geographic sampling. Our relative genome size measurements conducted with flow cytometry revealed that the majority of populations is diploid (2n=14) and they are distributed across most of the distribution area. Tetraploids are more restricted and were found in southern Dinaric Mountains, whereas hexaploids were found only on Mt. Pirin (Bulgaria). Our detailed screening of cytotype diversity and genome size variation in F. varia complex will contribute to the integrative approach combining phylogenomic and morphological analyses in order to resolve relationships within this intricate group.

Festuca L. is the most species-rich genus of the grasses in Europe. Festuca bosniaca (belonging to F. varia complex) is an Apennine-Balkan endemic species occurring in alpine and subalpine grasslands. We here investigated current and past (Last Glacial Maximum - LGM) habitat suitability of F. bosniaca using Species distribution modelling (SDM). Our models were built based on the collected occurrence points for F. bosniaca from various sources (a total of 103 points after filtering) and a set of eight selected environmental variables which included climatic and geomorphological predictors available for both periods. In addition, current habitat suitability was also predicted based on 13 selected predictors available exclusively for the current period, including additional variables that were not available for the LGM period. SDM was carried out using the Maximum Entropy method with 70% of the occurrences used for model training, and 30% of occurrences used for model evaluation. Projections for the LGM period were based on four general circular models (GCMs) (NCAR-CCSM4, MRI-CGCM3, MPI-ESM-P, MIROC-ESM) which were averaged into a consensus LGM projection. All the resulting models showed very good performance (AUC > 0.9) and the main environmental variables influencing habitat suitability of F. bosniaca were mean daily mean air temperatures of the wettest quarter (bio8), mean monthly precipitation amount of the warmest quarter (bio 19), mean daily mean air temperatures of the coldest quarter (bio11) and slope. In addition, current habitat suitability was also influenced by soil organic carbon content and number of snow days. Overall, predicted current distribution corresponded well to the known occurrences of F. bosniaca on the Balkan and Apennine peninsulas, with some potential undiscovered populations in the area of southern part of the Scardo-Pindic mountain range and the north-western part of Apennine peninsula. LGM predictions pointed to larger suitable areas in the past, suggesting that F. bosniaca might have had a wider distribution during LGM compared to present.

The Dianthus sylvestris complex is a group comprising morphologically highly variable taxa distributed in central Mediterranean, with the highest taxonomic diversity described in Italy. Another diversity centre is the Balkan Peninsula, from where six taxa mostly treated as subspecies of D. sylvestris have been described: subsp. alboroseus, subsp. bertisceus, subsp. kozjakensis, subsp. nodosus, subsp. sylvestris and subsp. tergestinus. Their identification is challenging due to great morphological variability and complex patterns of variation with respect to ecological preferences. In an ongoing study, we are combining an array of methods ranging from nomenclatural revision and morphometrics, over karyological and genome size estimation, to environmental niche modelling and population genetic analyses to disentangle relationships within D. sylvestris s.l. on the Balkan Peninsula. The morphometric analyses revealed a continuous variability of quantitative morphological characters and an absence of clear-cut qualitative morphological differences among the subspecies. In addition, the niche overlap assessment indicated that niche similarity is more common among the six subspecies, than niche divergence. Preliminary SNPs analyses suggest that D. sylvestris subsp. tergestinus presents a separate evolutionary lineage, while other populations on the Balkan Peninsula form a cline of genetic differentiation in the SE-NW direction. Finally, the relative genome size estimation using flow cytometry, with confirmatory chromosome counts, revealed polyploidisation in the NW Balkan Peninsula, as the tetraploid populations (2n=4x=60) are restricted to the Istrian Peninsula and Kvarner in Croatia and Italy. All the remaining Balkan populations were diploid (2n=2x=30).

Festuca bosniaca is an endemic species of the Balkan and Apennine Peninsulas, occurring in alpine and subalpine grasslands. Species distribution modelling (SDMs) was used to determine the habitat suitability for current and past (Last Glacial Maximum - LGM) time periods. The occurrence points of F. bosniaca were collected in the field from 2018 to 2021 and were supplemented with additional points from the Flora Croatica database. The models were build on 103 points after filtering and of eight (one geomorphological and seven bioclimatic) selected environmental variables of initial set of 43 variables from CHELSA and exChelsa databases. Current habitat suitability was predicted based on 13 selected predictors. SDMs was conducted using the Maximum Entropy method, with 70% of the occurrences used for model training and 30% of the occurrences used for model evaluation. Projections for the LGM period were based on four general circular models (GCMs) (NCAR-CCSM4, MRI-CGCM3, MPI-ESM-P, MIROC-ESM) averaged to produce a consensus LGM projection. All resulting models performed very well (AUC > 0.9) and the main environmental variables affecting habitat suitability of F. bosniaca were: mean daily mean air temperatures of the wettest quarter (bio8), mean monthly precipitation amount of the warmest quarter (bio 19), mean daily mean air temperatures of the coldest quarter (bio11) and slope. Current habitat suitability was also influenced by soil organic carbon content and number of snow days. The habitat suitability coincides well with the known occurrences of F. bosniaca in the Balkans and Apennines. The models also showed some potentially newly discovered populations in the area of the Scardo-Pindic mountains and in the north-western part of the Apennine Peninsula. When comparing the models for the LGM and the present, a much wider range was observed in the past, which may be related to the lower temperatures favoured by F. bosniaca.

The Dianthus ciliatus complex has three morphologically very similar subspecies that are geographically well defined in the Adriatic region. The nominal subspecies D. ciliatus subsp. ciliatus occurs in northern and central Adriatic and in central Italy, while D. ciliatus subsp. dalmaticus (Čelak.) Hayek is endemic to southern Adriatic and D. ciliatus subsp. medunensis (Beck et Szyszył.) Trinajstić is endemic to Montenegro and Albania. Because of previously unclear morphological distinctions and due to the overlapping of morphological characters, it was not possible to define the subspecies unambiguously. A morphometric analysis was performed on the collected herbarium materials of the three subspecies, in total including 66 individuals from 22 populations across the Adriatic region (11 populations of D. ciliatus subsp. ciliatus, seven populations of D. ciliatus subsp. dalmaticus, and four populations of D. ciliatus subsp. medunensis). Seventeen quantitative and five qualitative morphological characters were analysed. Descriptive and multivariate statistics were performed, and seven morphological characters that describe the size and shape of epicalyx scales had statistically significant differences among subspecies. Those seven characters (epicalyx height, outer second scale width and length, outer first scale width and length, inner scale length and apex length of first outer scale) can be used to better distinguish between D. ciliatus subsp. ciliatus and the other two subspecies. Dianthus ciliatus subsp. dalmaticus and D. ciliatus subsp. medunensis can also be distinguished on the basis of petal colour and petal denticulation. Based on this morphometric analysis, an identification key for the subspecies of D. ciliatus complex was given, and a taxonomic revision of herbarium specimens from CNHM, ZA, ZAGR and ZAHO was also performed.

High-throughput, restriction site-associated DNA sequencing (RAD-seq) is widely used for research in fields of population genetics (e.g. for calculations of genetic diversity, distance and structure) and phylogenomics (integrative evolution and genomic analyses on next-generation sequencing data) which combined with spatial spans of investigated populations can provide valuable insights into their phylogeographic patterns. Originally RAD-seq was used to untangle complex infraspecific relationships (among populations of the same species), but nowadays is often used to elucidate interspecific relationships within recently derived phylogenetic groups. Here we present the overview of the original RAD-seq methodology implemented in our research with comprehensive subsequent bioinformatic analyses, pipelines and programs used. Important findings of different analysed short nucleotide polymorphisms (SNP) datasets are shown through most recent and comprehensive published articles dealing with plant species from various genera (e.g. Aurinia, Dianthus, Euphorbia and Helleborus), all conducted in south-eastern Europe. Additionally, our preliminary results from 17 sequenced populations within F. varia complex (Poaceae) from the Balkan and the Apennine Peninsulas and Alps are presented.

Dianthus sylvestris complex includes ten subspecies whose delimitation is still challenging and full of controversy. Its important diversity centre is in the central Mediterranean area. Identification of D. sylvestris subspecies continues to be problematic due to great morphological variability between subspecies, and complex patterns of variation with respect to ecological preferences and geographic distribution. This study is focused on Balkan populations of six subspecies (D. sylvestris ssp. alboroseus, D. sylvestris ssp. bertisceus, D. sylvestris ssp. kozjakensis, D. sylvestris ssp. nodosus, D. sylvestris ssp. sylvestris and D. sylvestris ssp. tergestinus), which have been treated inconsistently in previous taxonomic accounts. Qualitative and quantitative morphological characters were examined on plants collected in the field and deposited in ZA herbarium. The multivariate analyses based on 15 morphological characters (2 qualitative, 5 continuous, 6 ratios and 2 meristic) were used to describe and identify patterns of morphological variation in D. sylvestris complex, including 461 individuals from 97 populations across the Balkan Peninsula. When considering the subspecies in two-dimensional ordination space, a separation of southern Balkan and northern Balkan subspecies along PC1 (27% of variation) was detected, as well as a vast amount of overlap along PC2 (19% of variation). Calyx length, petal denticulation, and number and shape of epicalyx scales turned out to capture the greatest extent of morphological variability between individuals. However, they should not be considered as taxonomically reliable, since individual values can fall within the variability range of several subspecies. Subspecies such as D. sylvestris ssp. nodosus, D. sylvestris ssp. sylvestris and D. sylvestris ssp. tergestinus, have similar morphologies and share parts of their geographic distributions, and cannot be clearly differentiated using quantitative morphological concepts. However, a different flowering period differentiates D. sylvestris ssp. tergestinus, suggesting it to be a sympatric taxon, which is supported by our preliminary results of genetic analyses. Therefore, we suggest treating D. sylvestris complex on the Balkan Peninsula as a single morphologically variable species, while only D. sylvestris ssp. tergestinus deserves infraspecific rank.

The genus Festuca is one of the largest genera within the grass family [1]. It is characterized by a complex taxonomy resulting from morphological complexity which includes not only heteromorphism but also convergent morphology of different species inhabiting similar environments. Additional taxonomical confusion is caused by the fact that some species include populations with different degrees of ploidy, resulting in varied morphological characteristics [2]. A different degree of ploidy within the same species may cause a higher ecological adaptability [3]. The aim of our research is to determine the genome size and ploidy degree in Festuca bosniaca that is native to the high mountains of the Balkans. Chromosome number in diploids of the species is 2n=14. Thirty-seven populations of F. bosniaca were sampled across its distribution range during the summer months from 2018 to 2020. On average, five individuals were sampled in each population. Genome size measured by flow cytometry showed that in sampled populations, diploids (27 populations) and tetraploids (10 populations) are found. Relative genome size ranged from 1.138 to 1.637 for diploid and 2.255 to 2.627 for tetraploid populations. Diploid populations were found throughout the whole area of species distribution, while tetraploids are limited to the central part of the range, and probably represent relicts from former glacial refugia.

Grupa Dianthus sylvestris smatra se jednom od taksonomski najzahtjevnijih unutar roda Dianthus. Prema dostupnoj literaturi unutar ove grupe je opisano nekoliko svojti na razini vrste ili podvrste, međutim njihov taksonomski status često je upitan. Grupu karakteriziraju svojte s malim i geografski vrlo ograničenim arealima, što upućuje na njihovu recentnu diversifikaciju. Središte raznolikosti ove skupine nalazi se u središnjem Mediteranu, gdje svojte zauzimaju uglavnom kamenita staništa i rasprostranjene su duž širokog visinskog raspona. Tijekom 2018. i 2019. godine provedeno je uzorkovanje biljaka s područja Balkanskog poluotoka i Alpa, biljni materijal je osušen pomoću silikagela te je protočnom citometrijom utvrđena relativna veličina genoma i procijenjen stupanj ploidije sakupljenih jedinki. Veličina genoma pouzdan je pokazatelj razine ploidije u rodu Dianthus, iako je endopoliploidija u ovom rodu uobičajena pojava. Od poliploida zabilježeni su tetraploidi te pojava smanjivanja monoploidnog genoma (eng. monoploid genome downsizing). Relativna veličina genoma određena je na temelju 124 izmjerene populacije grupe D. sylvestris, od kojih je 116 populacija diploidno, sedam tetraploidno, dok su unutar jedne populacije zabilježene diploidne i tetraploidne jedinke. Zabilježene tetraploidne populacije ograničene su na malo geografsko područje (Ćićarija i Kvarner). Relativna veličina genoma (vrijednost 2C) kretala se u rasponu od 1.095 do 1.228 pg za diploide, te od 2.162 to 2.217 pg za tetraploide. Vidljive su razlike u monoploidnoj relativnoj veličini genoma te da se ona povećava prema granicama uzorkovanog područja, ali za sad točan uzrok ovoj varijabilnosti ostaje nepoznat. Sličan obrazac manje veličine genoma u središtu, te njegovog povećavanja prema granicama područja rasprostranjenosti također je opaženo u rodu Knautia. Istraživanje obrazaca varijabilnosti veličine genoma i ploidije je dio integrativnog pristupa koji će uz morfološke i molekularne podatke doprinijeti poznavanju D. sylvestris grupe i razumijevanju njezinih evolucijskih putova i diversifikacije.

With over 200.000 sheets, ZA is the oldest and the largest herbarium collection in Croatia, while the ZAHO stores as many as 78.000 sheets. Digitisation was initiated in 2015 resulting in 11.5 % of digitised sheets so far. Based on 3.721 sheets analysed, we determined specific temporal biases with different peak collections, e.g. orchids from 1870’s to 1910’s, aquatic plants in 1910’s, 1960’s and 2010’s, fagetal species from 1900 to 1950, and invasive flora from 1960 to 1970 and after 2010. Moreover, the average age of those collections is 90 years. Disproportionally large percentage of specimens collected by very few individuals indicates the presence of collector biases too. The ZA keeps a dozen aquatic taxa as the only evidence of their presence in Croatia. As herbaria usually follow invasions from the first recorded colonizer onwards, we searched whether the collections hide data on the arrival of aliens in Croatia. The first record of 17 taxa occurred much earlier in herbaria than in literature, with gaps that sometimes counted more than 100 years. Based on the study of several common and easily recognizable fagetal taxa, we found that they are rarely collected and their representation in the collections is inversely proportional to their number and distribution in nature.

Dianthus sylvestris group is considered to be one of the most complex groups within the genus Dianthus (Caryophyllaceae). It includes several taxa of doubtful taxonomic value, and synonymy confusion is common. In order to initiate a taxonomical review, an inventory of historical D. sylvestris herbarium material from ZA and ZAHO collections was conducted. Herbarium specimens of D. sylvestris group from these two Croatian herbaria were digitized and the data from the original text on labels were stored within the Flora Croatica Database. A total of 319 herbarium sheets were digitized ; the ZA collection holds 182 sheets and ZAHO collection holds 137 sheets. Altogether, four taxa (D. sylvestris Wulfen in Jacq., D. sylvestris Wulfen in Jacq. ssp. sylvestris, D. sylvestris Wulfen in Jacq. ssp. tergestinus (Rchb.) Hayek and D. sylvestris Wulfen in Jacq. ssp. nodosus (Tausch) Hayek) were registered within studied collections. The exsiccata originate from eight European countries ; however, 61% are from Croatia. The majority of specimens were collected between 1910s and 1940s, originating mostly from Ivo Horvat’s fieldwork. The average age of the collected specimens was 98 years. Inventory of herbarium sheets from ZA and ZAHO historical collections provided a significant insight into historical distributional data of D. sylvestris taxa related to the area of Balkan Peninsula, which is a prerequisite for accurate taxonomic/geographic sampling for further morphological and molecular analyses. Apart from historical herbarium specimens, D. sylvestris collection from ZA herbarium has recently been growing due to extensive sampling of D. sylvestris taxa throughout the Balkans