Arabidopsis arenosa is a good candidate for phytoremediation due to its high tolerance to Zn and Cd as well as its accumulation ability. However, its small size and low biomass are the largest obstacles to applying it on a broad scale. The aim was to obtain polyploid specimens, which tend to have higher biomass to increase the accumulation and translocation capacity of heavy metals in this metal-tolerant plant. Doubled polyploids (octaploids) were obtained via indirect organogenesis on a ½ MS medium supplemented with 1 mg L⁻¹ TDZ, followed by rooting on the same medium without growth regulators. Callus tissue of a high endopolyploidy level (the (Ʃ>2C)/2C ratio over 2.5) obtained on seedling fragments on ½ MS supplemented with 2 mg L⁻¹ 2,4-D + 2 mg L⁻¹ BAP served as a source material. Among the regenerants successfully obtained (without using antimitotic agents), over half, regardless of the stage of regenerant development, were octaploid (54–78%; 2C DNA = 1.642 pg). Octaploids were not affected by ploidy or in vitro culture conditions; they were fully fertile, produced normal pollen (~97% of viability), and set seeds capable of germinating (78%). Their cell and organ size was affected by genome doubling resulting in longer stomata, bigger pollen grains, and flowers with a larger area and width in comparison with tetraploid regenerants and initial plants. The promising results of measurements of morpho-anatomical, physiological, and reproductive parameters indicate that, in the future, after passing tolerance tests, the obtained polyploids could be used in phytoremediation of metal-contaminated areas.

Many species of the Viola L. genus (violets) colonize areas with high concentrations of trace elements in the soil, e.g., nickel, cadmium, zinc, and lead. Although tolerance to heavy metals is a common phenomenon in violets, it is not clear whether this is the result of gradual microevolutionary processes as a part of the adaptation to the specific conditions, or whether the tolerance was inherited from the ancestor(s). We developed cell suspension cultures of five plant species: two non-metallophytes—Arabidopsis thaliana (Col-0) and Viola · wittrockiana, and three metallophytes—V. philippica, V. tricolor, and Silene vulgaris subsp. humilis for tolerance tests. The aim of the study was to measure the level of tolerance of violets in comparison with species from the other genera to verify the hypothesis of the high, innate tolerance of the former. We measured cell viability, non-enzymatic antioxidant content, and the accumulation of heavy metals after cell treatment with Zn or Pb. The results indicate they are innate and independent on the ecological status (metallophyte vs. non-metallophyte) and high in comparison with other species tolerance to Zn and Pb in violets. Viability of the cells after Zn and Pb (1000 μM) exposure for 72 h was the highest in violets. Antioxidant content, after heavy metal treatment, increased significantly, particularly in metallophyte violets, indicating their high responsivity to metals. In all species, lead was detected in the protoplasm of the cells, not in the vacuole or cell wall. All violets were characterized by the accumulation capacity of lead. Here, we clearly show that the physiological and biochemical studies conducted with the use of heavy metals on plant cells translate into the heavy metal tolerance of the species.

Background: Autopolyploids generally exhibit larger phenotypes than their progenitors, suggesting higher biomass and enhanced phytoremediation potential. This study tested this hypothesis using a previously uncharacterized, novel artificial autooctaploid of a Zn- and Cd-tolerant ecotype of Arabidopsis arenosa , derived from a natural tetraploid cytotype. Vegetative development, metal tolerance, and accumulation capacity were compared between the parental tetraploids and the derived autooctaploids under controlled laboratory conditions. Results: During the intermediate growth period (days 56–72), octaploid plants had significantly lower biomass than tetraploids, but this difference was no longer significant by day 84. This transient difference coincided with the earlier entry of octaploids into the generative phase. Despite their autopolyploid origin, the octaploid specimens were fully fertile, enabling sexual reproduction via seeds. They could also be propagated using encapsulated vegetative tissues (artificial seeds). Under extreme heavy metal stress (8 mM Zn and/or 0.2 mM Cd for 9 days), octaploids showed a significantly higher Zn concentration in their roots than tetraploids, while both cytotypes exhibited similar overall resilience to metal toxicity. Conclusions: The autooctaploids demonstrate significant potential for land reclamation due to their ability to tolerate and sequester extreme concentrations of zinc and cadmium without visible signs of toxicity. Under the controlled conditions used here, octaploids showed a > 50% higher Zn concentration in roots than tetraploids, whereas shoot Zn concentrations did not differ, supporting potential for phytostabilisation rather than enhanced phytoextraction. The more rapid life cycle of octaploids may be relevant to future applications, but any advantage under field conditions remains to be tested.