Karin Kollárová

Interaction of silicon and cadmium in Brassica juncea and Brassica napus

The objective of this study was to determine the effect of silicon (Si) and cadmium (Cd) on root and shoot growth and Cd uptake in two hydroponically cultivated Brassica species (B. juncea (L.) Czern. cv. Vitasso and B. napus L. cv. Atlantic). Both species are potentially usable for phytoextraction. Inhibitory effects of Cd on root elongation were diminished by the impact of Si. Primary roots elongation in the presence of Cd + Si compared with Cd was stronger and the number of lateral roots was lower in B. juncea than in B. napus. Cd content per plant was higher in B. napus roots and shoots compared with B. juncea. Suberin lamellae were formed closer to the root apex in Cd + Si than in Cd treated plants and this effect was stronger in B. napus than in B. juncea. Accelerated maturation of endodermis was associated with reduced Cd uptake. Cd decreased the content of chlorophylls and carotenoids in both species, but Si addition positively influenced the content of photosynthetic pigments which was higher in B. napus than in B. juncea. Si enhanced more substantially translocation of Cd into the shoot of B. napus than of B. juncea. Based on our results B. napus seems to be more suitable for Cd phytoextraction than B. juncea because these plants produce more biomass and accumulate higher amount of Cd. The protective effect of Si on Cd treated Brassica plants could be attributed to more extensive development of suberin lamellae in endodermis.

Further biological characteristics of galactoglucomannan oligosaccharides

The biological activity of cell wall-derived galactoglucomannan oligosaccharides (GGMOs) was dependent on their chemical structure. Galactosyl side chains linked to the glucomanno-core influenced their inhibition of elongation growth of pea (Pisum sativum L. cv. Tyrkys) stem segments induced by 2,4-dichlorophenoxyacetic acid (2,4-D). Reduction of the number of galactosyl side chains in GGMOs caused stimulation of the endogenous growth. Modification on the glucomanno-reducing end did not affect significantly the activity of these oligosaccharides. GGMOs inhibited also the elongation induced by indole-3-acetic acid (IAA) and gibberellic acid (GA3). In the presence of IAA the elongation growth was inhibited to 20 – 35 % after 24 h of incubation depending on GGMOs concentrations (1 μM, 10 nM, 0.1 nM), similarly as in the presence of 2,4-D, which confirms the hypothesis of GGMOs antiauxin properties. The elongation induced by GA3 was inhibited to 25 – 60 %, however, the time course of inhibition was different compared with IAA and 2,4-D. The highest inhibition was determined already after 6 h of incubation with a significant decrease after this time. The results indicated a competition between GGMOs and growth regulators.

An easy method for cutting and fluorescent staining of thin roots

BACKGROUND AND AIMS Cutting plant material is essential for observing internal structures and may be difficult for various reasons. Most fixation agents such as aldehydes, as well as embedding resins, do not allow subsequent use of fluorescent staining and make material too soft to make good-quality hand-sections. Moreover, cutting thin roots can be very difficult and time consuming. A new, fast and effective method to provide good-quality sections and fluorescent staining of fresh or fixed root samples, including those of very thin roots (such as Arabidopsis or Noccaea), is described here. METHODS To overcome the above-mentioned difficulties the following procedure is proposed: fixation in methanol (when fresh material cannot be used) followed by en bloc staining with toluidine blue, embedding in 6 % agarose, preparation of free-hand sections of embedded material, staining with fluorescent dye, and observation in a microscope under UV light. KEY RESULTS Despite eventual slight deformation of primary cell walls (depending on the species and root developmental stage), this method allows effective observation of different structures such as ontogenetic changes of cells along the root axis, e.g. development of xylem elements, deposition of Casparian bands and suberin lamellae in endodermis or exodermis or peri-endodermal thickenings in Noccaea roots. CONCLUSIONS This method provides good-quality sections and allows relatively rapid detection of cell-wall modifications. Also important is the possibility of using this method for free-hand cutting of extremely thin roots such as those of Arabidopsis.

Interaction of galactoglucomannan oligosaccharides with auxin in mung bean primary root

In the present paper timing of galactoglucomannan oligosaccharides (GGMOs) with exogenously added indole-3-butyric acid (IBA) action on early germination stage (24 h) and primary root elongation of mung bean (Vigna radiata (L.) Wilczek) has been studied. GGMOs inhibited primary root elongation induced by low concentration (10(-8) M) of IBA. This inhibition was considerably higher after preincubation with GGMOs compared with other timing experiments. The most intensive inhibition of elongation has been ascertained at the 10(-8) M concentration of GGMOs. On the other hand GGMOs stimulated this elongation inhibited by high IBA concentration (10(-4) M). This stimulation was the most intensive by simultaneous addition of IBA and GGMOs at the beginning of the experiment and subsequent seeds incubation in distilled water. Our results indicate competition between GGMOs and auxin. The root growth inhibition, induced by GGMOs and/or IBA, was accompanied by the increase of cell wall-associated peroxidase activity and by a higher number of peroxidase isoenzymes. The presence of different peroxidase isoenzymes in experiments with distinct treatment of GGMOs and IBA could indicate variations in the mechanism of interaction between GGMOs and IBA.

Effect of auxins on Karwinskia humboldtiana root cultures

Effects of auxins (IAA, IBA and NAA) on K. humboldtiana root culture cultivated in 16-h photoperiod or in dark have been observed. Light affected positively the production of biomass when cultivated on medium supplemented with NAA in 10 and 25 μ mol −1 concentrations. In the presence of IAA and IBA these values were significantly lower. The growth dynamics of root cultures depended on the auxin used. The best adventitious roots elongation and lateral roots induction on media supplemented with IBA has been ascertained. Morphological and anatomical differences in dependence on auxin used were observed. NAA supported the formation of huge callus-like mass besides mostly very short roots, especially under the light. Similarly IAA induced short roots, and IBA seems to be the most effective substance for the root elongation in this model system. NAA induced roots with larger diameter under the light compared with the other two auxins used. The reason is in the different anatomical structure of roots which was characterized by higher number of cell layers and large intercellulars in the cortex. The shape of cortical cells in the presence of IBA depended on the light conditions. Isodiametric cortical cells were present in roots cultivated in 16-h photoperiod, irregularly-shaped cells in the dark. The effect of light conditions was the smallest in the case of roots grown on IAA enriched media.

Oligosaccharides induce changes in protein patterns of regenerating spruce protoplasts

Galactoglucomannan oligosaccharides (GGMOs, d.p. 4–8 and fractions d.p. 3, 4, 5, 6–7), used in culture media for spruce protoplasts derived from callus cells showed a pleiotropic effect. They influenced both, quality and quantity of extracellular proteins in regenerating protoplasts. GGMOs d.p. 4, 5 at pH 6.0 and the mixture of d.p. 4–8 (pH 3.8 and 6.0) after 48 h of culture increased the amount of extra- and intracellular proteins and the viability of protoplasts. The most significant effect on protoplasts viability in the presence of GGMOs d.p. 4–8 without the growth hormone supplementation at pH 3.8, and with lower efficiency in the presence of NAA (1 mg/l) has been observed. The most significant differences were observed in the molecular mass intervals Mr∼17–21 kDa, 25–30 kDa, and 45–60 kDa. After 24 h of culture extracellular acid proteins with Mr∼30; 41; 68; and 90 in media supplemented with GGMOs d.p. 3, 4, 5, 6–7 (pH 6.0) with or without NAA were identified to belong to the group of β-1,3-glucanases. Extracellular proteins p27.5; 30; 41; 50; 52 and 90 exhibited chitinase activity after 24 h of protoplast cultivation. GGMOs probably fulfil a protective role in this process of spruce protoplast regeneration.

Galactoglucomannan oligosaccharides inhibition of elongation growth is in pea epicotyls coupled with peroxidase activity

The effect of galactoglucomannan oligosaccharides — GGMOs, GGMOs-r (GGMOs with reduced reducing ends), and GGMOs-g (GGMOs with reduced number of d-galactose units) on peroxidase activity was determined in pea epicotyls. GGMOs didn’t significantly modify the activity of soluble peroxidases. However, cell wall-associated peroxidases activity increased after GGMOs and GGMOs-r treatment, while in the presence of GGMOs-g this activity was significantly lower. These results are inversely related to the GGMOs, GGMOs-r, and GGMOs-g effect on elongation growth induced by 2,4-D (2,4-dichlorophenoxyacetic acid) in pea epicotyls. It can be concluded that GGMOs evoked inhibition of the elongation growth induced by auxin is probably associated with cell wall modifications catalysed by peroxidase.

Growth and anatomical parameters of adventitious roots formed on mung bean hypocotyls are correlated with galactoglucomannan oligosaccharides structure.

The effect of galactoglucomannan oligosaccharides (GGMOs) compared with chemically modified oligosaccharides, GGMOs-g (with reduced number of D-galactose side chains) and GGMOs-r (with reduced reducing ends) on mung bean (Vigna radiata (L.) Wilczek) adventitious roots formation, elongation, and anatomical structure have been studied. All types of oligosaccharides influenced adventitious root formation in the same way: stimulation in the absence of exogenous auxin and inhibition in the presence of exogenous auxin. Both reactions are probably related with the presence/content of endogenous auxin in plant cuttings. However, the adventitious root length was inhibited by GGMOs both in the absence as well as in the presence of auxin (IBA or NAA), while GGMOs-g inhibition was significantly weaker compared with GGMOs. GGMOs-r were without significant difference on both processes, compared with GGMOs. GGMOs affected not only the adventitious root length but also their anatomy in dependence on the combination with certain type of auxin. The oligosaccharides influenced cortical cells division, which was reflected in the cortex area and in the root diameter. All processes followed were dependent on oligosaccharides chemical structure. The results suggest also that GGM-derived oligosaccharides may play an important role in adventitious roots elongation but not in their formation.

Cadmium impact, accumulation and detection in poplar callus cells

Effect of cadmium cations and their interaction with silicon cations was determined in poplar calli and expressed as changes in callus growth, cell viability and cadmium cation accumulation. Cell viability throughout culture versus cadmium cation accumulation in cells is discussed. At the same time, the study sought appropriate methods for cadmium cation detection in callus cells and also in experiments with low plant material (e.g. protoplasts). Cadmium cations were determined by atomic absorption spectroscopy and using fluorescence microscopy with a specific cadmium cation fluorescent dye. The detection of cadmium cations in callus cells by the latter method appears suitable because the callus cells are surrounded by primary cell walls without auto-fluorescence and these values fit well with atomic absorption spectroscopy quantification. However, the visualisation method has some limits discussed below.

Impact of galactoglucomannan oligosaccharides and Cd stress on maize root growth parameters, morphology, and structure

Biologically active oligosaccharides, including galactoglucomannan oligosaccharides (GGMOs), affect plant growth and development. The impact of GGMOs is dependent on their concentration, and the plant species and plant parts affected. The aim of this article is to ascertain the effects of GGMOs, GGMOs + Cd2+, on growth parameters, morphology, and the structure of maize (Zea mays L.) roots. We undertook this research because, in monocots, the effect of these oligosaccharides is so far unknown. In our study, GGMOs stimulated primary root elongation, induction and elongation of lateral roots, and biomass production. Their effect was dependent on the concentration used. Simultaneously, GGMOs moderated the negative effect of Cd2+ on root elongation growth. Besides, GGMOs affected the primary root structure, proven in the earlier development of xylem and Casparian bands, but not of suberin lamellae (compared to the control). The presence of Cd2+ shifted the apoplasmic barriers closer to the root apex in comparison to samples treated with GGMOs + Cd2+. GGMOs do not inhibit Cd uptake into the root directly, but they moderate its effect, and therefore their influence at the structural and metabolic level seems possible. Their positive impact on plant vitality, even in contaminated conditions, strongly indicates their potential application in remediation technologies.