The intensification of platinum (Pt), cobalt (Co) and nickel (Ni) mining and processing results in the release of salts and metals into the environment. This calls for the identification of halophytes with an ability to tolerate and desalinate metal-contaminated sites while simultaneously allocating metals (Pt, Ni, and Co) into harvestable biomass. Tamarix usneoides E. Mey ex Bunge is an indigenous exo-recretohalophyte that has been used for erosion control and for the desalination and allocation of metals from gold and uranium mine tailings and land contaminated by metallurgical effluent. The aim of this study was to investigate the uptake, translocation, and tolerance of Pt, Ni, and Co by T. usneoides from liquid medium (in vitro) and soil contaminated by base metal refinery effluent spillages and previous overspray from the enhanced evaporation spray system (in situ). More specifically, the in situ study investigated the utility of mature T. usneoides trees in the desalination of soil contaminated by previous metallurgical spillages and overspray emissions through the extraction of sulphur and metals Pt, Ni, and Co into harvestable biomass. Four T. usneoides trees were categorised into different size classes based on tree measurements and allometric equations. Seven soil pits (four “planted” and three “unplanted” – control) were excavated and opposite faces of the soil profile were sampled at 20 different intervals (0 – 340 cm). Soil samples were freeze-dried and analysed for total element concentrations. Root systems were harvested by excavating soil pits (maximum depth of 3.5 meters) using a mechanical excavator. Trees were harvested and immediately separated into above (leaves, twigs, wood, and flowers) and belowground (coarse and fine roots) plant organs. Tree biomass was further separated into different above (outer bark, inner bark, and sapwood and heartwood) and belowground (epidermis, cortex, and stele) tissue types. Plant material was rinsed three times in tap water to remove unbound residual metals and residual substrate from root and shoot surfaces. It must be noted that the determined metal concentrations are a combined measure of metals adsorbed on the root surface, assimilated in planta, and excreted on the plant surface from the foliar salt glands. Metals were allocated in trees (across plant organs and tissue types) in the order: Ni (59.46 ± 4.67 mg/kg) > Co (2.65 ± 0.34 mg/kg) > Pt (50 ± 6 µg/kg) whereas sulphur (S) was hyperaccumulated in tree leaves [39 900 ± 861 mg/kg (3.9% ± 0.7 %)]. Platinum was bioaccumulated [bioconcentration factor (BCF) > 1.5] and translocated [translocation factor (TF) > 1] in the leafy shoots of one individual tree, Ni in one (BCF = 1.03), and Co in another replicate (BCF = 1.02). Soil chemical properties (pH, electrical conductivity, and redox potential) differed between planted and unplanted pits whereby pH and EC were lower in planted pits [pH 6.0; EC = 3 499 µS/cm (34.99 mM NaCl)] compared with unplanted [pH 7.6; EC = 9 644 µS/cm (96.44 mM NaCl)] (ANOVA, p < 0.01). The lower EC, along with S hyperaccumulation (BCF > 20; TF > 1), supports the potential use of T. usneoides for phytoextraction of S and Ni in shoot tissues and Co and Pt in roots. At a spacing of 1333 trees / ha, T. usneoides trees could remove an estimated 2.23 ± 0.30 mg Pt/ha, 3.02 ± 0.83 kg Ni/ha, 1.28 ± 0.90 kg Co/ha, and 1.28 ± 0.09 tons S/ha, excluding excreted salts. Excreted salts were visible but could not be quantified without confounding surface dust contamination. The first in vitro study determined factors influencing the rhizogenesis of T. usneoides in order to develop a mass propagation protocol. Explant establishment in vitro was influenced by various donor plant factors, viz. growing conditions (contaminated < non-contaminated; Kruskal-Wallis (KW), p < 0.05), physiological age (younger > older donor plants; ANOVA, p < 0.05), genotype (KW, p < 0.001), season of culturing (higher establishment in winter; KW, p < 0.05), length of explant (40 mm > 25 mm; KW, p < 0.05), and volume of growth vial (50 mL > 15 mL; KW, p < 0.05) but not pH, chronological age, strength of plant growth medium, or auxin pulse treatments. This study indicates that propagation protocols can be developed by controlling factors influencing explant establishment. A standardised and rapid in vitro protocol was developed for the mass propagation of T. usneoides explants. This in vitro protocol was used for the metal uptake studies whereby established explants were exposed to 25 % Murashige and Skoog standard plant growth medium supplemented with Pt, Co, or Ni (as sulphate complexes) at 0, 25, 50, or 100 mg/L at pH 5.5 or 7.5 over a 14-day exposure period. On completion of the metal exposure period, plantlets were harvested, separated into roots and shoots, freeze-dried, and analysed for metal concentrations. Higher metal concentrations (Ni > Co >> Pt) were accumulated in roots (combined measure of metals adsorbed on the root surface and assimilated in planta) compared with shoots whereby BCF > 1 (excluding Pt) and TF < 1. Metal BCF (Ni > Co >> Pt; KW, p < 0.05) and TF (Co > Ni >> Pt; KW, p < 0.05) increased in a dose-dependent fashion and were not influenced by pH level. Cobalt and Ni (≤ 50 mg/L) uptake dynamics did not v differ suggesting similar uptake dynamics, when treated separately. Platinum (defined in this study as Pt > 1 – 4 mg/kg), Ni (> 1 000 mg/kg), and Co (> 300 mg/kg) were hyperaccumulated in roots (“rhizo-hyperaccumulation”) across treatments with possible Co-hyperaccumulation in shoots by two genotypes. Genotype influenced Co allocation in shoots but not Ni or Pt. Tolerance indices did not differ [Co (97 %) > Pt (82 %) > Ni (77 %)] between pH, metal, treatment concentration, or the interplay between these factors. Metal treatments did not impact measured morphological parameters (excluding Ni treatments which promoted shoot length increment) (KW, p < 0.05). Plantlet survival differed between pH and metals [Pt (90 %) > Ni (81 %) > Co (62 %)] (KW, p < 0.05). Variability in Co accumulation capacity between genotypes indicated that selective breeding, using the developed in vitro mass propagation protocol, for improved rhizofiltration and phytoextraction traits is feasible. Results demonstrate that T. usneoides has the potential for recovery of Ni and Co (and Pt to a lesser degree) from effluents, exhibiting a tolerance to Ni, Co, and Pt at 1, 10, and 10,000 times the average soil crustal abundance, respectively, under moderately acidic (pH 5.5) and alkaline (pH 7.5) conditions and across a wide metal concentration range. Results from the in situ study indicate that 9- year-old T. usneoides trees can be used for the decontamination of sulphate-contaminated soils under study site conditions which are more conducive to the survival of glycophytes. Tamarix usneoides is thus able to assimilate, translocate, and tolerate Ni, Co, and Pt (to a lesser degree) when exposed to metals across a wide pH and metal concentration range, under different (in situ and in vitro) experimental conditions. This opens the possibility for the species to be used in a range of phytotechnologies.

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