Blagay's daphne (Daphne blagayana) is a small decumbent bush from the family Thymelaeaceae. Due to its rarity it is protected by national legislation in almost all countries within its range. The aim of this work was to assess the conservation status of Blagay's daphne (Daphne blagayana) throughout its distribution range, using chloroplast DNA (cpDNA) and AFLP markers. The present distribution of D. blagayana is fragmented and comprises the Balkan Peninsula and the southern Romanian Carpathians. However, the mechanism leading to such fragmented distribution range is not well understood. According to one interpretation, D. blagayana had a smaller distribution range during the interglacial, after which it expanded north and northwest. According to another interpretation, the fragmented areal is the result of fragmentation during the glacial and is still preserved today. The mechanisms leading to isolated populations play an important role in within-and between-population genetic variability, which in turn is relevant to species conservation.Sampling of D. blagayana was conducted at 21 locations across the entire distribution range in 2009 and 2010. cpDNA and AFLP markers were used as a tool for assessing its conservation status. The phylogeographic analysis included 95 plants from 21 populations. The study was based on the analysis of five spacer regions of chloroplast DNA: rpl20-rps12, atp-BrbcL, trnL-F, psbA-trnH and trnK%matK with the gene matK. The results suggest the existence of three clusters: the northern cluster (SLO), including Slovenian and Italian populations, the southern cluster (J), including populations from Macedonia, Montenegro and Stolac (BiH), and the central cluster (SR), including the remaining populations. Both haplotype and nucleotide diversity were highest in the central cluster and lowest in the northern cluster.In the second part of the study we assessed genetic structure and gene flow between 13 selected populations using dominant AFLP markers. The results showed a relatively strong genetic differentation (FST = 0.4) and absence of gene flow among distant populations. Using programs STRUCTURE and BAPS populations were subdivided into 2 and 7 groups, respectively. The groupings proposed by STRUCTURE and BAPS are not fully congruent with cpDNA clusters. This incongruence might be attributed to different reproduction strategies within populations, and to different modes of inheritance of chloroplast and nuclear DNA.Based on both marker systems we can suggest the identification of three evolutionarily significant units (ESU) and one additional management unit (MU), on which conservation efforts should be focused in the future.
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