Anthony G. Kachenko

Heavy metal tolerance in common fern species

The effects of cadmium (Cd), chromium (Cr), copper (Cu), nickel (Ni), lead (Pb) and zinc (Zn) on the growth and uptake of 10 fern species was investigated under a controlled environment in order to evaluate their suitability for phytoremediation. Fern species included Adiantum aethiopicum, Blechnum cartilagineum, Blechnum nudum, Calochlaena dubia, Dennstaedtia davallioides, Doodia aspera, Hypolepis muelleri, Nephrolepis cordifolia, Pellaea falcata and the arsenic (As) hyperaccumulating Pteris vittata. Ferns were exposed to four levels of metals at concentrations of 0, 50, 100 and 500 mg kg–1 for a period of 20 weeks. The response of ferns significantly varied among species and metals. In general, heavy-metal translocation was limited, with metals being absorbed and held in roots, suggesting an exclusion mechanism as part of the ferns’ tolerance to the applied metals. Similar metal-accumulation patterns were observed for all species in that accumulation generally increased with increasing metal treatments; in most cases a sharp increase in metal accumulation was observed between 100 and 500 mg kg–1 treatments, suggesting a breakdown in tolerance mechanisms and unrestricted metal transport. This was corroborated by enhanced visual toxicity symptoms and a reduction in survival rates among ferns when exposed to 500 mg kg–1 metal treatments; and to a lesser extent 100 mg kg–1 metal treatments. Of the species investigated, N. cordifolia and H. muelleri were identified as possible candidates in phytostabilisation of Cu, Pb, Ni or Zn contaminated soils. Similarly, D. davallioides appeared favourable for use in phytostabilisation of Cu and Zn contaminated soils. These species had high survival rates and accumulated high levels of the aforementioned metals relative to other ferns investigated. Ferns belonging to the family Blechnaceae (B. nudum, B. cartilagineum and D. aspera) and C. dubia (Family Dicksoniaceae) were least tolerant to most metals, had a low survival rate and were classified as being unsuitable for phytoremediation purposes. Metal tolerance was also observed in P. vittata when exposed to Cd, Cr and Cu; however, no hyperaccumulation was observed.

Arsenic speciation in tissues of the hyperaccumulator P. calomelanos var. austroamericana using X-ray absorption spectroscopy.

The fate and chemical speciation of arsenic (As) during uptake, translocation, and storage by the As hyperaccumulating fern Pityrogramma calomelanos var. austroamericana (Pteridaceae) were examined using inductively coupled plasma-atomic emission spectrometry (ICP-AES) and synchrotron-based micro-X-ray absorption near edge structure (micro-XANES) and micro-X-ray fluorescence (micro-XRF) spectroscopies. Chemical analysis revealed total As concentration was ca. 6.5 times greater in young fronds (5845 mg kg(-1) dry weight (DW)) than in old fronds (903 mg kg(-1) DW). In pinnae, As concentration decreased from the base (6822 mg kg(-1) DW) to the apex (4301 mg kg(-1) DW) of the fronds. The results from micro-XANES and micro-XRF of living tissues suggested that more than 60% of arsenate (As(V)) absorbed was reduced to arsenite (As(III)) in roots, prior to transport through vascular tissues as As(V) and As(III). In pinnules, As(III) was the predominant redox species (72-90%), presumably as solvated, oxygen coordinated compounds. The presence of putative As(III)-sulphide (S(2-)) coordination throughout the fern tissues (4-25%) suggests that S(2-) functional groups may contribute in the biochemical reduction of As(V) to As(III) during uptake and transport at a whole-plant level. Organic arsenicals and thiol-rich compounds were not detected in the species and are unlikely to play a role in As hyperaccumulation in this fern. The study provides important insights into homeostatic regulation of As following As uptake in P. calomelanos var. austroamericana.

Effect of soil properties on arsenic hyperaccumulation in Pteris vittata and Pityrogramma calomelanos var. austroamericana.

Two arsenic (As) hyperaccumulators, Pteris vittata L. and Pityrogramma calomelanos var. austroamericana, were grown in As-contaminated soils of contrasting properties. Ferns were exposed to three levels of As in soil at concentrations of 0, 600 and 2400 μmol kg−1 for a period of 22 weeks. Plant biomass and As concentration in fronds and roots varied significantly between the two species. At 600 μmol kg−1 As level, As hyperaccumulation was not observed in both the fern species. However at the 2400 μmol kg−1 As level, both the species accumulated very high levels (>1000 mg kg−1) of As in fronds. Arsenic concentration and uptake in fronds of both species followed the order Kurosol (Box Hill) > Vertosol (Narrabri) > Ferrosol (Robertson). In the studied soils, P. vittata possessed higher frond biomass and As accumulation, and thus was more efficient in removing soil As than P. calomelanos var. austroamericana. Soil properties such as free Fe, clay and organic matter contents appear to have affected the bioavailability of As in the studied soils. These results show that soil properties influence the As extraction efficiency of hyperaccumulating plants and must be considered in context of the phytoextraction technology of As contaminated soils.

Influence of Drought Stress on the Nickel-Hyperaccumulating Shrub Hybanthus floribundus (Lindl.) F.Muell. subsp. floribundus

Hyperaccumulation of nickel (Ni) in certain plants may play a role in drought resistance under water stress. This article tests the influence of water stress on the Ni-hyperaccumulating shrub Hybanthus floribundus Lindl. F.Muell. subsp. floribundus. Plants grown in 1000 mg kg−1 Ni-amended Clastic Rudosol were exposed to five levels of soil water potentials (−33 [field capacity], −60, −400, −600, and −1000 kPa) for 12 wk. Water stress did not induce significant changes in growth rate, relative water content, rates of gas exchange, or carbon isotope discrimination. Water use efficiency (WUE) values were approximately threefold lower in plants at water potentials <−400 kPa than they were in those at water potentials of −33 kPa. Low WUE values suggest that this species possesses an efficient water conservation mechanism that enables its survival in competitive water-limited environments. A 38% decline in water potential and a 68% decline in osmotic potential occurred between −1000- and −33-kPa water potentials (), indicating that osmotic adjustment (OA) may have provided turgor maintenance in response to increasing water stress. However, Ni concentration in plants did not significantly increase in response to decreasing water potentials and is therefore unlikely to play a role in OA.

The role of low molecular weight ligands in nickel hyperaccumulation in Hybanthus floribundus subspecies floribundus

The mechanisms responsible for nickel (Ni) hyperaccumulation in Hybanthus floribundus (Lindl.) F.Muell. subspecies floribundus are obscure. In this study, organic acids and free amino acids (AAs) were quantified in 0.025 M HCl H. floribundus subsp. floribundus shoot extracts using HPLC and ultra performance liquid chromatography (UPLC). In a 20 week pot experiment, plants exposed to five levels of Ni (0–3000 mg kg–1 Ni) accumulated up to 3200 mg Ni kg–1 dry weight in shoots, and the shoot : root Ni concentration ratios were >1.4. Concentration of organic acids followed the order malic acid > citric acid > oxalic acid. Citric acid concentration significantly increased upon Ni exposure, with concentrations between 2.3- and 5.9-fold higher in Ni treated plants that in control plants. Molar ratios of Ni to citric acid ranged from 1.3 : 1 to 1.7 : 1 equivalent to >60% of the accumulated Ni. Malic acid concentration also increased upon exposure to applied Ni. However, concentrations were statistically at par across 0–3000 mg kg–1 Ni treatments, suggesting that the production of malic acid is a constitutive property of the subspecies. Total AA concentrations were stimulated upon exposure to external Ni treatment, with glutamine, alanine and aspartic acids being the predominant acids. These AAs accounted for up to 64% of the total free AA concentration in control plants and up to 75% for the 2000 mg kg–1 Ni treatment plants. These results suggest that citric acid in addition to the aforementioned AAs are synthesised in H. floribundus subsp. floribundus plants following exposure to elevated concentrations of Ni and may act as potential ligands for detoxification and possibly storage of accumulated Ni.