Acta Biologica Cracoviensia s. Botanica

Abstract Biscutella laevigata L. is known as a Tl hyperaccumulator. In Poland Biscutella laevigata occurs in the Tatra Mts (Western Carpathians) and on the calamine waste heap in Bolesław near Olkusz (Silesian Upland). The purpose of this work was to evaluate whether plants of both populations were able to accumulate an elevated amount of thallium in their tissues. The plants were cultivated in calamine soil in a glasshouse for a season and studied at different ages – from 2-week-old seedlings to 10-month-old adults. Additionally, the plants were grown for ten weeks in calamine soil with EDTA to enhance Tl bioavailability. The total content of Tl in plant tissues after digestion was determined by ICP-MS, whereas its distribution in leaves was studied by LA-ICP-MS. Of the total content of Tl in the soil in the range of (15.2–66.7) mg∙kg−1d.m., only (1.1–2.1) mg∙kg−1d.m. was present in a bioavailable form. The mean content in all the plants grown on the soil without EDTA was 98.5 mg∙kg−1d.m. The largest content was found in leaves – 164.9 mg∙kg−1d.m. (max. 588.2 mg∙kg−1d.m.). In the case of plants grown on the soil enriched with EDTA, the mean content in plants increased to 108.9 mg∙kg−1d.m., max. in leaves – 138.4 mg∙kg−1d.m. (max. 1100 mg∙kg−1d.m.). The translocation factor was 6.1 in the soil and 2.2 in the soil with EDTA; the bioconcentration factor amounted to 10.9 and 5.8, respectively. The plants from both populations did not contain a Tl amount clearly indicating hyperaccumulation (100–500 mg∙kg−1d.m.), however, high (>1) translocation and bioconcentration factors suggest such an ability. It is a characteristic species-wide trait; B. laevigata L. is a facultative Tl hyperaccumulator. The largest Tl amount was located at the leaf base, the smallest at its top. Thallium also occurred in trichomes, which was presented for the first time; in this way plants detoxify Tl in the above-ground parts. Leaves were much more hairy in the Bolesław plants. This is an adaptation for growth in the extreme conditions of the zinc-lead waste heap with elevated Tl quantity.

Abstract The male-specific DNA markers are very useful in molecular sexing of non-flowering plants and seeds of dioecious species. In this paper we identified ten Y chromosome-specific RAPD primers suitable for identification of male plants in three Cannabaceae species with sex chromosomes (Humulus lupulus, XX/XY; H. japonicus, XX/XY1Y2; Cannabis sativa, XX/XY). Basing on the nucleotide sequence of the OPJ-09 RAPD product we developed the HJY09 SCAR marker, which is very efficient in sexing of Japanese hop.

Abstract Rhododendron tomentosum Harmaja (formerly Ledum palustre L.) is a medicinal peat bog plant native to northern Europe, Asia and North America. This plant has a distinctive aroma thanks to the presence of essential oil, to which it also owes its anti-inflammatory, analgesic, antimicrobial and insecticidal properties. However, in Europe R. tomentosum is classified as an endangered species, mainly due to degradation of peatlands. In the present work, the micropropagation protocol for R. tomentosum was established for the first time, providing both an ex situ conservation tool and a means of continuous production of in vivo and in vitro plant material for further studies. R. tomentosum microshoots were initiated from leaf explants and further multiplied using Schenk-Hildebrandt (SH) medium supplemented with 9.84 μM 2iP and 1.00 μM TDZ. The shoots were elongated on the SH medium supplemented with 24.6 μM 2iP and subsequently rooted using the perlite substrate saturated with half-strength Woody Plant medium supplemented with 1.0% sucrose and 4.92 μM IBA. The regenerated plants were hardened on the phytohormone-free SH medium and acclimatized using 3:1:1 deacidified peat:perlite:gravel substrate. The identity of the mother plant was confirmed at morphological and molecular levels and Random Amplified Polymorphic DNA (RAPD) method was implemented to assess the genetic fidelity of the regenerants. The essential oil content of the maternal plant, in vitro shoots and the regenerants was determined by steam-distillation, and the obtained volatile fractions were analyzed by GC/MS.

Abstract Plants belonging to the family Oleaceae have been traditionally used in treatment of various inflammatory skin disorders. One of the most well-known species is Olea europaea L. (olive), cultivated in the Mediterranean countries. Another species is Ligustrum vulgare L. (common privet), occurring particularly in Northern Europe and Asia. The aim of the study was a comparison of the composition of aqueous and ethanolic extracts from leaves of O. europaea and L. vulgare (HPLC-DAD-MS), and determination of the total content of phenolics and flavonoids, as well as the content of the major compound, oleuropein. Secondly, we aimed to study the protective effect of extracts on reactive oxygen species (ROS) production by human fibroblast cells (NHDF), cell viability (MTT assay), and apoptosis rate (Annexin V/propidium iodide staining) after UVA-irradiation. The phytochemical analysis allowed us to identify compounds belonging to the groups of flavonoids, phenylpropanoids and secoiridoids in the extracts. The compounds from the group of lignans (olivil) were identified as being unique to O. europaea extracts. Echinacoside, ligustroflavone and ligustaloside A were identified in L. vulgare extracts in contrast to olive preparations. It was established that the aqueous and ethanolic extracts from leaves of both species, except the privet aqueous extract at a concentration of 5 μg/ml, did not show any significant inhibition of ROS production after UVA-irradiation in the model of NHDF cell line. The aqueous extracts of both species at concentrations of 5 and 25 μg/ml had a protective effect on the viability of UVA-treated cells in contrast to the ethanolic extract. In conclusion, no significant difference in the activity of olive and privet leaf extracts has been observed, which suggests that both plant materials’ extracts, particularly aqueous ones, are effective herbal medicines and photoprotectors, which – to some extent – confirms the use of their preparations in skin disorders.

Abstract Floral epidermal cells of most species of Bulbophyllum Thouars studied to date produce both lipid-rich food-rewards and fragrance. Since fragrances largely consist of terpenoids and have an affinity for lipophilic stains, the simultaneous presence of lipid-rich food-rewards frustrates identification of fragrance-secreting cells by conventional histochemistry. Furthermore, since both lipid-rich food-rewards and fragrances are probably synthesized by a similar complement of organelles, interpretation of TEM images can prove difficult. All members of section Racemosae Benth. & Hook. f. investigated to date, however, are unusual in their secretion of a predominantly proteinaceous food-reward, and lipids are seemingly absent. This might enable their use as models for the identification and characterization of fragrance-secreting tissues and organelles. Three members of sect. Racemosae were chosen, namely Bulbophyllum dissitiflorum Seidenf., B. lilacinum Ridl. and B. tricorne Seidenf. & Smitinand. All produced food-rewards. Of these, one (B. dissitiflorum) lacked fragrance and was used as a control, whereas the remaining two species produced fragrance. Having established that the food-reward was mainly proteinaceous in each case, and did not test positively for lipid, we undertook further histochemical investigations, as well as light microscopy, SEM and TEM. Specialized palisade-like epidermal cells of all species contained protein bodies and rough endoplasmic reticulum consistent with the production and secretion of a protein-rich food-reward. Cuticular pores were also present. In fragrant species, these cells also contained abundant smooth endoplasmic reticulum, oil droplets and many, well-developed, spherical plastids with numerous plastoglobuli, similar to those found in the osmophores (fragrance-producing structures) of other orchids. Indeterminate, osmiophilic cytoplasmic inclusions were also present. By contrast, the non-fragrant species lacked oil droplets and other osmiophilic inclusions and the plastids were scant, poorly developed, often elongate or irregular in shape and contained few plastoglobuli. Smooth endoplasmic reticulum was also less frequent. Since food-rewards tested negatively for lipid, it is probable that any oil droplets present were involved in fragrance production, especially since they were absent from the non-fragrant species. Thus, the unusual absence of lipids from the food-rewards of sect. Racemosae provided a rare opportunity, permitting, for the first time, the unraveling of these two secretory processes (food-reward and fragrance) in Bulbophyllum and clearly demonstrating the plasticity of these cells and their dual role in secretion.

Abstract Viola uliginosa (bog violet) is a declining species throughout its range due to – mostly anthropogenic – drying out of the wet habitats it occupies. Using AFLP markers, we aimed to estimate the genetic diversity in Polish populations, that may give an insight into the situation of plant populations facing rapid loss of natural habitats. Bog violet from several dispersed Polish populations is generally characterized by very low genetic diversity (HT = 0.048), even lower than several other endangered violets; therefore, we suggest that it should preserve at least EN rank in the red lists/red data books. The mean gene diversity within all populations (HS) was much lower than gene diversity (GST) between populations (0.020 versus 0.583, respectively) which supports the prevalence of clonal propagation of the species (mainly by stolons) but may also point to some significance of autogamy in cleisto- and chasmogamous flowers. A high FST value and the Mantel test for all populations revealed significant isolation by distance. Geographically neighboring pairs of populations formed genetic clusters supported by all (in the case of two closest populations) or most statistical analyses applied. Special attention should be paid to the locus classicus of the species in Rząska, consisting of a small number of individuals, forming a genetically distinct group, revealing very low gene diversity (Hj = 0.009) and the longest genetic distance to the remaining populations. Our results can contribute to planning future protection measures for the species at this and other locations. Genetic structure of the studied populations suggests local affinities of populations but does not generally support hypothesized recent continuity of V. uliginosa range along the river valleys of southern Poland; this view may, however, be altered with widening of the scope of studied populations and chosen molecular markers.

Abstract This study deals with populations of the European-South-Siberian geoelement Adenophora liliifolia (L.) A. DC. in the Czech Republic, Slovakia, Hungary, Romania, and Poland, where this species has its European periphery distribution. We studied the population size, genetic variability, site conditions, and vegetation units in which A. liliifolia grows. Recent and historical localities of A. liliifolia were ranked into six vegetation units of both forest and non-forest character. A phytosociological survey showed differences in the species composition among localities. Only a weak pattern of population structure was observed (only 22% of total genetic variation present at the interpopulation level, AMOVA analysis), with moderate values for gene diversity (Hj = 0.141) and polymorphism (P = 27.6%). Neighbor-joining and Bayesian clusterings suggest a similar genetic background for most of the populations from Slovakia, the Czech Republic, and Poland, contrary to the populations from Hungary, Romania, as well as two populations from Central and South Slovakia. This might be explained by a relatively recent fragmentation of the A. liliifolia populations in Central Europe. Nevertheless, it seems that several populations in Romania, South Hungary, and Slovakia were isolated for a longer period of time and their genetic differentiation is more evident.

Abstract This is the first study to report an efficient anther culture (AC) method for spelt wheat, which has an increasing importance not only in applied research but also in organic farming and changing nutritional standards. In this study, an efficient AC protocol has been described for ‘GK Fehér’ spelt wheat. The number of AC-derived embryo-like structures (ELS) was 62.2/100 anthers, from which we were able to regenerate 30.6 green plantlets per 100 anthers. The percentage of green plantlets production was 89.0% among the regenerated plantlets, while the phenomenon of albinism was restricted (3.8/100 anthers). Altogether, from AC of ‘GK Fehér’ 306 green plantlets were produced in vitro and 241 plants were acclimatized to the greenhouse conditions. Based on ploidy level analyses, 83 spontaneous doubled haploid (DH) plants were produced (8.3 DH plants/100 anthers), so the percentage of spontaneous rediploidization was 34.4%. The spontaneous DH plants produced fertile spikes, while a few seeds were harvested from seven partially fertile plants.