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Key message Lowered pollen and seed quality of B. humilis compared to its widespread tree relatives may threaten its populations at the southwestern edge of the species range. Abstract Inability to produce vital propagules is the main threat to the persistence of rare plants at the edges of species distribution. In the present investigation, factors responsible for the low germinability of shrub birch (Betula humilis) seeds in the peripheral populations were analysed. As shrub birch populations are found in phosphorus (P)-limited peatlands with high groundwater levels, the impact of groundwater PO43− ion contents and groundwater table for B. humilis flowering was also studied. Qualities of the B. humilis seeds and pollen were compared to those of tree congeners silver birch (B. pendula) and downy birch (B. pubescens), which usually represent a high reproductive performance. We revealed that the poor germination ability of B. humilis seeds was caused by the high proportions of empty seeds (44.8%) and seeds with partially developed embryos (47.3%). The insufficient numbers of viable seeds may be a consequence of low-quality shrub birch pollen, as only 1.6% of pollen grains sprouted. The germinability of shrub birch pollen was significantly lower than that of Betula tree pollen; thus, there is a risk of pollination of B. humilis female flowers by pollen grains of tree birches. Positive relationships between the P concentrations in seeds and seed germinability were revealed in tree birches, but not in B. humilis. The availability of PO43− ions in the groundwater did not affect B. humilis flowering, but male inflorescences were more numerous in stands with higher groundwater levels. Consequently, the maintenance of peatlands is crucial for the efficient sexual reproduction of shrub birches.

Trees and shrubs belonging to the Betula L. genus seem to represent one of the most confused interspecific relationships, and hybridization is considered to be one of the main factors responsible for the substantial genetic and morphological variation of birches. In the present investigation, the internal transcribed spacer ITS1 and ITS2 regions of nuclear ribosomal DNA, nuclear alcohol dehydrogenase ( ADH ) gene sequences and amplified fragment length polymorphisms (AFLPs) as well as the Atkinson discriminant function (ADF) of leaf morphology were used to verify hypotheses concerning the origin of the dark barked Betula obscura and B. pendula var. carelica with the patterned wood. Both plants were considered by different authors to be distinct species, intraspecific forms of common B. pendula or B. pubescens , or hybrid taxa between B. pendula and B. pubescens . In the phylogenetic trees, the ITS and ADH gene sequences of both B. obscura and B. pendula var. carelica clustered with those of B. pendula , whereas B. pubescens trees were somewhat distinct. In turn, the AFLPs revealed genetic similarity of B. pendula var. carelica to both frequent species, whereas B. obscura was clearly distinct from other birches. Values of the ADF indices of the rare birches were typical for B. pendula . In the light of the results obtained, we imply that B. obscura and B. pendula var. carelica represent an intraspecific variation of B. pendula , they are neither intraspecific taxa of B. pubescens nor hybrid species between the common birches. Different grouping of B. obscura in the AFLPs and DNA sequences analyses is likely a result of an unreliable phylogenetic signal of the former molecular markers.

Juniperus excelsa s. str. (Greek juniper) in Crimea is a relic species on the limits of its range, and represents the Mediterranean flora in the Sub-Mediterranean part of the peninsula. Its origin and history in this area remains unresolved. We aimed to analyze phylogeny and potential demographic expansion of the juniper in the Crimea as well as to study its morphological differentiation. We analyzed plant material from 59 trees inhabiting eight populations. Genetic variation assessments were based on the four non-coding chloroplast DNA (cpDNA) fragments and the nuclear internal transcribed spacer region ITS1-5,8S-ITS2 (ITS). To examine the morphological differentiation, eight measured/counted traits of cones, seeds, and shoots were chosen and eight ratios were calculated. Morphological parameters were compared using ANOVA, Student’s t test, discrimination analysis and Kruskal-Wallis and U Mann-Whitney tests. Two cpDNA fragments were polymorphic and, in total, 10 cpDNA haplotypes were found. Haplotype diversity (Hd) ranged from 0.0 to 0.9. Based on both cpDNA and ITS sequences variation, phylogenetic analyses revealed a close relationship of the Crimean junipers to the individuals from other parts of the species range. In general, our molecular results confirmed the low level of genetic differentiation of J. excelsa individuals inhabiting different parts of the species range, likely resulting from a common ancestry. Only slight morphological differences were found between populations with different geographic location or habitat. The analyzes showed the distinctness of the populations from the southern part of the coast. Some unique morphological and molecular features of southern coastal populations imply that they are remnants of Late Pleistocene abundant forests. We suggest that the recent fragmentation of the Juniperus populations in the Crimean Peninsula could have arisen during the Atlantic period of the Holocene.

Trees and shrubs belonging to the Betula L. genus present highly confusing interspecific relationships. The complex evolutionary history of birches may be due to several factors: interspecific gene flow, the independent occurrence of both polyploidy and dwarf forms in different lineages, incidents of major speciation events within a very short time period and different selection pressures in the distant populations of widespread species. We assessed phylogenetic relationships between the uncommon dark-barked B. obscura and B. pendula var. carelica with patterned wood texture and widespread birch tree species: white-barked B. pendula and B. pubescens as well as dark-barked B. nigra and B. dahurica, using the internal transcribed spacer ITS1 and ITS2 regions of nuclear ribosomal DNA, nuclear ADH (alcohol dehydrogenase) gene sequences and AFLPs (amplified fragment length polymorphisms). In the phylogenetic trees based on the ITS and ADH gene sequences, B. pendula var. carelica and B. obscura clustered with B. pendula, but were distinct from B. pubescens, B. nigra and B. dahurica. In turn, both clustering and multivariate ordination methods using the AFLPs revealed clear distinctness of B. obscura from the remaining species. Betula pendula var. carelica grouped with B. pendula and B. pubescens individuals in both AFLP analyses. Values of the leaf morphology indices of B. obscura and B. pendula var. carelica were within the range of B. pendula variation. All genetic and morphological analyses confirmed the intraspecific status of B. pendula var. carelica. We also suggest that B. obscura represent intraspecific variation of B. pendula, although discrepancies between nuclear sequences and AFLP results were noted.

In many plant species sexual and asexual reproduction occur simultaneously. Clone size and clonal architecture, i.e. the spatial distribution of ramets, can significantly affect pollination and mating. In guerrilla strategy vegetative ramets belonging to one genet are dispersed and intermixed with other clones, which facilitate cross-fertilization. In turn, the mixing of ramets of different clones is significantly limited in phalanx strategy, increasing the probability of self-fertilization by geitonogamy. The shrub birch, Betula humilis, is a wind-pollinated species, which also reproduces vegetatively. The aim of the study was to define the strategy of clonal growth and investigate clonal diversity in the endangered B. humilis populations from the southwestern margin of the species range to assess their potential influence on pollination and selfing. We conducted genotyping at seven nuclear SSR (simple sequence repeats) loci of 522 ramets within 24 sampling plots in six populations differentiated in size and level of competition from other plant species. The analysis revealed 86 genets in total. Clonal and genetic diversities were substantial in all populations. Spatial distribution of ramets and number of genets differed between studied populations, but in general, the phalanx strategy dominated, as from 59% to 95% of ramets shared the same genotype with their closest neighbour. Nevertheless, clones were relatively small, most of individuals were unrelated, and plots with intermixed genets were found in all studied localities. This observation indicates that self-pollination does not seem to be a considerable threat, even in small and overgrown B. humilis stands. The plasticity of clonal growth strategies may facilitate survival of the shrub birch and allows it to persist under different environmental conditions.

Background: Hybridisation can be a threat for the survival of a rare species because, in the case of insufficient numbers of appropriate mates, a rare form is much more likely to cross with a widespread taxon.Aims: In the present study, we tested hypotheses concerning the level of hybridisation between endangered Betula humilis and its widespread congeners: B. pendula and B. pubescens as a function of habitat conditions.Methods: We genotyped 312 individuals of three species using AFLP markers. B. humilis specimens were sampled in populations with low and high groundwater levels. Morphological identification of B. pubescens and B. pendula was verified using the Atkinson discriminant function.Results: Altogether, 15 individuals (4.8%) were indicated as putative hybrids. The B. humilis hybrids were found in dry habitats and they could be classified as F1 or F2 generation. Tree hybrids could represent backcrosses to either B. pendula or B. pubescens.Conclusions: Genetic analyses contradicted the idea ...

Many plant species can reproduce by both sexual and vegetative means. Clonal diversity and degree of intermingling of clones in the vegetative reproductive mode can influence the mating and fertility of individuals. The aim of the study was to assess the clonal structure and its potential influence on gene flow and generative reproduction efficiency in six endangered Betula humilis populations from the southwestern margin of the species range. Analyses of seven microsatellite loci revealed 86 genets among 522 samples. In general, the phalanx strategy dominated in the populations considered, as 76% of ramets shared the same genotype with their closest neighbour. Nevertheless, substantial clonal and genetic diversities and high contribution of unrelated individuals in all B. humilis stands suggest that panmictic pollination prevails. On the other hand, positive and significant relationships between genetic and geographic distances in the two populations could be a consequence of biparental inbreeding resulting from the pollen and seed flow limitations. The seed germination capacity was very low (2.70%); however, the populations characterised by the lowest and highest values of clonal diversity parameters did not differ significantly in the number of germinated seeds, which indicates that clonality is not responsible for seed production failure.