Acta Biologica Cracoviensia s. Botanica

Abstract In recent years there has been growing interest in selenium (Se) as an important micronutrient not only for animals and humans but also for plants. In particular, its protective effect in plants exposed to stress conditions has been suggested. In spite of many studies, the mechanism of Se action is not fully understood. In this review, possible ways of interaction of Se with stress factors leading to optimal growth and development of plants are presented. As the majority of experiments have focused on the effects of Se application under stress conditions induced by heavy metals, special attention is paid to the results obtained in such studies. Changes of physiological and biochemical properties of plant cells, with particular regard to the influence of Se on the activation of enzymatic and non-enzymatic antioxidants under this stress, are summarized. Experiments in which Se was used in some other environmental stresses (drought, UV, cold and high temperature) are also cited. On the basis of the presented literature it is suggested that a positive effect of Se depends on both its doses and on chosen plant genotypes and is mainly connected with activation of antioxidative defense in plant cells.

Abstract In the last decade contradictory results have been published as to whether exogenous salicylic acid (SA) can increase salt stress tolerance in cultivated plants by inducing an antioxidant response. Salt stress injury in tomato was mitigated only in cases when the plant was hardened with a high concentration of SA (~10−4 M), low concentrations were ineffective. An efficient accumulation of Na+ in older leaves is a well-known response to salt stress in tomato plants (Solanum lycopersicum cv. Rio fuego) but it remains largely unexplored whether young and old leaves or root tissues have a distinct antioxidant status during salt stress after hardening with 10−7 M or 10−4 M SA. The determination of superoxide dismutase (SOD) and catalase (CAT) activity revealed that the SA-induced transient increases in these enzyme activities in young leaf and/or root tissues did not correlate with the salt tolerance of plants. Salt stress resulted in a tenfold increase in ascorbate peroxidase (APX) activities of young leaves and significant increases in APX and glutathione reductase (GR) activities of the roots hardened with 10−4 M SA. Both total ascorbate (AsA) and glutathione pools reached their highest levels in leaves after 10−7 M SA pre-treatment. However, in contrast to the leaves, the total pool of AsA decreased in the roots under salt stress and thus, due to low APX activity, active oxygen species were scavenged by ascorbate non-enzymatically in these tissues. The increased GR activities in the roots after treatment with 10−4 M SA enabled plants to enhance the reduced glutathione (GSH) pool and maintain the redox status of AsA under high salinity, which led to increased salt tolerance.

Abstract The study investigates the genetic differentiation among two subspecies of Allium ursinum L., namely A. ursinum subsp. ursinum and subsp. ucrainicum as well as their putative hybrid that is represented by individuals with intermediate morphology. Inter-Simple Sequence Repeats (ISSR) were applied to determine the status of intermediate morphotypes in terms of their genetic pattern and to assess the level of genetic variability within and between various populations of A. ursinum. The study comprises 144 specimens from nine populations along the east-west transect in Poland, which includes localities of both subspecies and their putative hybrid. Among the examined populations, 48 bands were amplified, of which 45 were found to be polymorphic. The principal coordinate analysis (PCoA), the neighbour-net analysis and Mantel test showed a strong correlation between genetic variability and geographic distance. Analysis of molecular variance (AMOVA) revealed that a greater proportion of total genetic variation resided within populations rather than among them. The Structure Bayesian clustering analysis revealed the presence of three distinct genetic groups within studied populations, where ‘eastern’ genotypes correspond to A. ursinum subsp. ucrainicum, and ‘western’ to subsp. ursinum; whereas the third genetic group has the largest share in the individuals occurring at the border of the distribution ranges of both subspecies. The emergence of the third genetic group is probably an effect of hybridization events occurring within the secondary contact zone. Typical morphologically intermediate populations occur only in a relatively narrow geographical zone, but the hybrid zone revealed by molecular markers is actually much wider than it is suggested by the morphological pattern of individuals. The current distribution pattern of both subspecies of A. ursinum and their hybrid zone is related to the two main directions of postglacial migration of Fagus sylvatica to the area of Poland. The hybrid zone arose as an effect of the secondary contact of two divergent lineages of A. ursinum.

Abstract Several studies have used A. thaliana as a model to identify the physiological and molecular mechanisms underlying iron deficiency tolerance in plants. Here, Arabidopsis thaliana and Thellungiella salsuginea were used to investigate the differential responses to iron deficiency of these two species. Plants were cultivated in hydroponic medium containing 5 or 0 μM Fe, for 10 days. Results showed that rosette biomass was more reduced in T. salsuginea than in A. thaliana when grown on Fe-deficient medium. As a marker for iron deficiency tolerance, the induction of ferric chelate reductase (FCR) and phosphoenolpyruvate carboxylase (PEPC) activities was observed only in A. thaliana roots. In addition, we found that the accumulation of phenolic acids in roots of N1438 ecotype of A. thaliana was stimulated by Fe deficiency. Furthermore, an increase of flavonoids content in the root and exudates was observed under Fe-deficiency in this ecotype. Unlike other abiotic stresses, it appears that iron deficiency effects were more pronounced in Thellungiella than in Arabidopsis. The higher tolerance of the Arabidopsis plant to iron deficiency may be due to the metabolic changes occurring in the roots.

Abstract In this study, wheat (Triticum aestivum L.) roots were treated with hypoxic water. The staining of cell preparations with DAPI revealed morphological changes of the cells such as nuclear condensation, deformation and fragmentation. Under TEM, cellular membrane shrinkage and breakage, chromatin condensation and apoptotic-like bodies were displayed. The number of mitochondria increased dramatically; their cristae were damaged; the interior became a cavitation and only some flocculent materials were distributed. Indirect immunofluorescence staining indicated that cytochrome C diffused from mitochondria to nucleoplasm and cytoplasm. TUNEL positive nuclei indicated double strand breaks of DNA. DAB staining was used for the identification of hydrogen peroxide and examination showed that the longer the treating time, the darker the staining of the meristematic zones of the roots which suggested the increased accumulation of these Reactive Oxygen Species (ROS). The elevation of hydrogen peroxide production was paralleled with the increase of SOD and POD activities. A negative correlation between the exposure time under hypoxia and the contents of soluble proteins was found. No obvious effect of hypoxia on MDA was established. The obtained results demonstrate that hypoxia causes programmed cell death in the root-tip meristematic cells of Triticum aestivum L. which is most probably attributed to the accumulation of large amounts of ROS.

Abstract The aim of the study was to determine the effect of exogenous sucrose and cytokinin on ethylene production and responsiveness in relation to the shoot formation of Pelargonium × hortorum ‘Bergpalais’ in vitro. Increasing the concentration of sucrose from 15 to 40 g L−1 in medium containing meta-topolin (mT) resulted in a two-fold decrease in the number of shoots and leaves as well as a reduction in ethylene production. The addition of ethylene synthesis inhibitor (AVG) to mT-medium significantly reduced the ethylene production and the shoot growth, but it had no significant influence on the shoot formation. The mT-induced shoot formation was, however, significantly reduced in the presence of ethylene action inhibitor (AgNO3), in a manner dependent on sucrose levels. At the end of the subculture period, increased sucrose concentrations (15–40 g L−1) in the presence of mT and AgNO3 resulted in a 3.7-fold increase in ethylene emission. At the same time, the supply of sucrose caused a 2.8-fold increase in the level of endogenous abscisic acid (ABA). Our results may suggest that the inhibitory effect of high sucrose concentration (30 and 40 g L−1) may depend on its influence on ethylene sensitivity. It also suggests that sucrose-regulation of the shoot formation of Pelargonium in vitro is mediated by ABA.

Abstract Germination responses of Galium cracoviense Ehrend. (Rubiaceae), a narrow endemic species from southern Poland, were tested in light and dark conditions at three constant temperatures (5, 10, or 22°C), before and after cold-wet stratification. Additionally, seeds were germinated under different calcium carbonate (CaCO3) concentrations (1, 5, 10, 15, 20, or 25 mM/L CaCO3) at 22°C in light. The high germination capacity of seeds incubated at different temperatures, shortly after collection, already suggested the absence of dormancy in this species. Thus, the seeds are ready to germinate immediately in the field when water resources are available and the temperature is adequate. Light was a significant factor for G. cracoviense; more seeds germinated in light than in darkness at all temperatures tested. Cold stratification decreased germination especially at higher temperatures. The light requirement for G. cracoviense germination ensures their successful germination on or near the soil surface, and in cracks and crevices in limestone, when temperature and edaphic conditions are favourable. Seeds of this species show temperature enforced dormancy throughout the winter. Germination was significantly affected by calcium carbonate. Non-germinated seeds germinated well after being transferred from higher CaCO3 concentrations to distilled water. The results indicate that the seeds of this species can endure CaCO3 stress without losing their viability and start germination once CaCO3 concentration is reduced. It can be concluded that the seeds of this species require lower Ca2+ ion concentration, moderate temperatures and the presence of light to germinate.

Abstract The co-occurrence of hybrids and parental species in similar ecological niches poses a question on the role of traits additivity and overdispersion (emergence of new traits) in microevolutionary processes. We analysed genetic polymorphism of Bromus benekenii, B. ramosus and the spontaneous hybrid B. benekenii × B. ramosus in sympatric and allopatric parts of the species distribution in Europe, based on non-coding regions of the taxon genomes (ISSR genetic fingerprinting). We tested 68 individuals in 7 populations, including a hybrid population in N France. Altogether 233 polymorphic ISSR bands (loci) were obtained. We found that the parent species were genetically distinct and the hybrids had an additive pattern of ISSR bands found in the putative parental species (NMDS, STRUCTURE); however, there was evidence of introgression towards B. ramosus (NEWHYBRIDS, UPGMA classifications, Nei's D genetic distance). Bromus benekenii had 72, B. ramosus 21 and the hybrids 9 private bands (genetic overdispersion), probably resulting from the rearranged genomes. Based on its low genetic divergence index DW, the hybrid population seems to be at a young age. We argue that in the face of anthropogenic landscape transformations favouring secondary contacts, the hybrids may competitively replace the parental species in sympatric areas.

Abstract The electrical penetration graph (EPG) method was used to quantify the effect of surface waxes on probing behaviour of the grain aphid Sitobion avenae F. (Hemiptera: Aphididae). The experiments showed that wax removal significantly affected probing behaviour of S. avenae. Generally, the aphids feeding on the plants without wax had a shortened non-probing (EPG-pattern np) and prolonged penetration of peripheral tissues – epidermis and mesophyll (EPG-pattern C). The EPG tests also showed that the three tested extracts of surface waxes from waxy plants RAH 122 were active as aphicides against the grain aphid.

Abstract Since M. sinensis Anderss., M. sacchariflorus (Maxim.) Hack. and M. ×giganteus J.M.Greef & Deuter ex Hodk. and Renvoize have considerably the highest potential for biomass production among Miscanthus Anderss. species, there is an urgent need to broaden the knowledge about cytological characteristics required for their improvement. In this study our objectives were to assess the genome size variation among eighteen Miscanthus accessions, as well as estimation of the monoploid genome size (2C and Cx) of the M. sinensis cultivars, which have not been analyzed yet. The characterization of three Miscanthus species was performed with the use of flow cytometry and analysis of the stomatal length. The triploid (2n = 3x = 57) M. sinensis ‘Goliath’ and M. ×giganteus clones possessed the highest 2C DNA content (8.34 pg and 7.43 pg, respectively). The intermediate 2C-values were found in the nuclei of the diploid (2n = 2x = 38) M. sinensis accessions (5.52–5.72 pg), whereas they were the lowest in the diploid (2n = 2x = 38) M. sacchariflorus ecotypes (4.58–4.59 pg). The presented study revealed interspecific variation of nuclear DNA content (P<0.01) and therefore allowed for recognition of particular taxa, inter- and intraspecific hybrids and prediction of potential parental components. Moreover, intraspecific genome size variation (P<0.01) was observed in M. sinensis cultivars at 3.62%. The values of the stomatal size obtained for the triploid M. ×giganteus ‘Great Britain’ (mean 30.70 μm) or ‘Canada’ (mean 29.67 μm) and diploid M. sinensis ‘Graziella’ (mean 29.96 μm) did not differ significantly, therefore this parameter is not recommended for ploidy estimation.

Abstract Flow cytometry estimation of 2C nuclear DNA content of the examined Nasturtium species resulted in taxonomic identification of N. × sterile in eight new localities, N. microphyllum in four new localities and N. officinale in one new locality in western Poland. Scanning electron microscopy proved a few of the micromorphological traits of seeds and fruits (size and shape of cells on the fruit septum surface, their anticlinal walls; secondary sculpture on the outer periclinal walls of cells on the siliqua valve internal surface) to be of taxonomic importance.

Abstract The total antioxidant activity of three apple cultivars conventionally farmed (Jonagold, Golden Delicious and Idared) and two cultivars organically farmed (Jonagold and Golden Delicious) were investigated by electron paramagnetic resonance (EPR) spectroscopy. The spin label used for experiments was TEMPO radical (2,2,6,6-tetramethylpiperidine-1-oxyl) with a well-defined EPR spectrum consisting of three equidistant peaks. The results obtained indicated difference in the total antioxidant activity of apples grown in two different ways, conventional and organic, as well as between different cultivars grown in the same way. The Golden Delicious cultivar is richer in antioxidants than the Jonagold and Idared, regardless of the method of farming. The difference between them is higher in conventionally grown apples than in organically farmed apples. Experimental data was fitted with non-linear curve fit for exponential decay of the first order based on the Levenberg-Marquardt method. The high values of the R2 parameters for all data indicated correctness in the proposition of exponential decay of the first order as a model describing the dynamic properties of spin labels which diminish in the presence of fresh apple juice.

Abstract Studies of plant breeding systems are particularly important when they involve facultative processes, which can be overlooked in natural conditions. This especially applies to species that are considered allogamous, due to their pollination syndrome and floral architecture. We examined the potential level and factors enabling or limiting spontaneous autogamy in three populations of the lepidopteran orchid Gymnadenia conopsea. Using a bagging experiment, we noted the stages of the anther thecae and the positions of the pollinaria at the five phenological stages of the flower, as well as the quantity of autogamously set fruits and the number of properly formed seeds. In the studied populations, autogamy represents an accidental character, with a maximum of 3.3% of fruits set spontaneously per analysed sample in a given population, and with seed numbers ranging from 29 to 354. This process is an environmentally dependent co-product of the mechanisms that enable a position appropriate for touching the stigma (bending of the caudicle) and increase male fitness (disintegration of the massulae), preceded by the gradual opening of the anther chambers. Autopollination of G. conopsea may occur in the flowers at various flowering stages (excluding the beginning of anthesis) at each position on the inflorescence.

Abstract The influence of methyl jasmonate on anthocyanin accumulation in roots of Kalanchoe blossfediana plants was studied. Methyl jasmonate (JA-Me), at a concentration of 5.0 to 40.0 mg.l−1, substantially increased anthocyanin accumulation in roots of intact plants, when it was applied as a solution under natural light conditions. The production of anthocyanin depended on the concentration of methyl jasmonate and the age of the plant. The stimulatory effect was higher in older plants of K. blossfeldiana than in younger ones. When leaves were removed methyl jasmonate slightly stimulated anthocyanin accumulation compared with intact plants. The obtained results indicate that leaves are necessary for the anthocyanin accumulation in the roots. In isolated roots methyl jasmonate did not affect the accumulation of anthocyanins in light conditions. Seven anthocyanins were documented in the roots of control plants and 8 anthocyanins in the roots of JA-Me treated ones. JA-Me increased the level of anthocyanins in roots of old K. blossfeldiana plants 6.8, 6.0 and 3.6-folds, after 4, 8 and 14-days of treatment, respectively.

Abstract Pollen grains of Pinus mugo, P. sylvestris and P. ×rhaetica (= P. mugo × P. sylvestris) were examined by light and scanning electron microscopy. The pollen grains were bisaccate and monosulcate. The corpus-saccus attachment was distinct. The pollen corpus exine sculpture was verrucate-rugulate and deeply sculptured. The surface of the tectum was covered with small grana and it was perforate. The saccus sexine ornamentation was reticulate and irregularly perforate. The tectum surface characters in the proximal and distal view of the corpus and saccus were less variable and they did not provide good criteria to identify the species under study. However, some differences were observed in the size of elevation on the corpus between pollen grains of the same species and between the parental species. This study of the pollen grain morphology of the corpus and saccus provided some important new data.