Investigating the effect of cadmium and aluminium on growth and stress-induced responses in the micropropagated medicinal plant Hypoxis hemerocallidea.

Hypoxis hemerocallidea is a highly utilized medicinal plant in South Africa. Its cultivation has received considerable attention in order to meet the high demand. High levels of cadmium (Cd) and aluminum (Al) in H. hemerocallidea plants sold in traditional medicinal markets was previously reported. The present study used an in vitro propagation model to investigate the uptake of Cd and Al by H. hemerocallidea and their effect on plant growth, elemental uptake and some stress-induced responses such as pigment, malondialdehyde (MDA), proline content and ultrastructural changes. Shoot and root growth of plantlets exposed to Cd, Cd:Al and high concentrations of Al was significantly reduced. Highest concentrations of Cd accumulated in the corms of Cd-treated plantlets while highest Al concentrations occurred in the leaves and roots. There was higher accumulation of Cd and Al when applied singularly compared to the Cd:Al combination treatments. Cd and Al also reduced accumulation of trace elements in micropropagted H. hemerocallidea with lowest concentrations in the Cd:Al combination treatments. Exposure to Cd, Al and Cd:Al significantly reduced the level of chlorophyll but increased the levels of carotenoids, MDA and proline. Ultrastructural changes were also observed in H. hemerocallidea exposed to Cd and Al. All these factors contributed to the inhibition of plant growth and could potentially affect the ability of this important medicinal plant to synthesize bioactive compounds. It is thus necessary to understand heavy metal stress-induced responses in this highly valued medicinal plant to ensure a high quality product for the consumer.

Effects of Cd and Al stress on secondary metabolites, antioxidant and antibacterial activity of Hypoxis hemerocallidea Fisch. & C.A. Mey.

This study investigated the effects of cadmium (Cd) and aluminium (Al) on the accumulation of phenolics, flavonoids and the bioactive compound hypoxoside in Hypoxis hemerocallidea. In addition, antioxidant scavenging and antibacterial activity were assessed to evaluate if Cd and Al stress affect the accumulation of bioactive compounds in H. hemerocallidea. In vitro grown plantlets of H. hemerocallidea were acclimatized for seven months in a greenhouse. Thereafter plants were exposed to various concentrations of Cd and Al both singularly and in combination in the form of Cd(NO3)2 (2, 5, 10 mg Cd/L); Al3(NO3)3 (500, 1000, 1500 mg Al/L) and combinations of Cd and Al (Cd 2:Al 500, Cd 5:Al 1000 and Cd 10:Al 1500 mg/L) for a further six weeks. The highest amounts of Cd and Al translocated to the shoot were 34 and 1608 mg/L respectively. Phytochemical screening showed significantly high amounts of total phenolics and flavonoids at the moderate Cd treatment (5 mg/L) compared to the controls. Exposure to Cd and Al significantly decreased the accumulation of hypoxoside. There was a significant increase in diphenylpicrylhydrazyl (DPPH) antioxidant scavenging activity in most of the metal-treated plants compared to the positive control ascorbic acid. Extracts from Cd 2 mg/L treatment exhibited moderate antibacterial activity against Staphylococcus aureus compared to the control. The results of the present study revealed that cultivating H. hemerocallidea on metal contaminated soils affects the accumulation of the bioactive compound, hypoxoside.

GA3-mediated dormancy alleviation in the reputed African potato, Hypoxis hemerocallidea.

BACKGROUND: Hypoxis hemerocallidea, arguably the most well-known medicinal species in South Africa, has been the subject of intensive harvesting from the wild leading to recent conservation concerns. The seeds of this species do not propagate easily and can lie dormant for up to twelve months. MATERIALS AND METHODS: In the in vitro germination experiments water, acid and chemical pre-sowing treatments were performed to determine the germination response of this species in both light and dark conditions. In the ex vitro experiment, intact seeds were sown and left to germinate in a potting soil mix under greenhouse conditions. RESULTS: Highest levels of germination (36.7-60.0% in the light and 36.7-46.7% in the dark) were achieved by treating mechanically scarified seeds with GA3 at various concentrations for 24 h. This was followed by scarified seeds soaked in water (26.7% in the light and 23.3% in the dark). Similar results (23.3 and 26.7%) were obtained in the 1% and 4% KNO3 treatments, respectively, under light conditions only. The fastest time to germinate was two days and was obtained in the 1200 ppm GA3 treatment in both light and dark conditions. Ex vitro germination of H. hemerocallidea seeds under greenhouse conditions was unsuccessful. CONCLUSION: H. hemerocallidea displays physical and non-deep physiological dormancy where pre-sowing treatments are required before the seeds will germinate.

Efficient in vitro regeneration of fertile plants from corm explants of Hypoxis hemerocallidea landrace Gaza -- the "African Potato".

We present efficient protocols for the regeneration of fertile plants from corm explants of Hypoxis hemerocallidea Fisch. and C. A. Mey. landrace Gaza, either by direct multiple shoot formation or via shoot organogenesis from corm-derived calluses. The regeneration efficiency depended on plant growth regulator concentrations and combinations. Multiple direct shoot formation with high frequency (100% with 5-8 shoots/explant) was obtained on a basal medium (BM) supplemented with 3 mg/l kinetin (BM1). However, efficient indirect regeneration occurred when corm explants were first plated on callus induction medium (BM2) with high kinetin (3 mg/l) and naphthalene acetic acid (NAA 1 mg/l), and then transferred to shoot inducing medium (BM3) containing BA (1.5 mg/l) and NAA (0.5 mg/l). Shoot regeneration frequency was 100% and 30-35 shoots per explant were obtained. The regenerated shoots were rooted on a root inducing medium (BM4) containing NAA (0.1 mg/l). Rooted plantlets were transferred to the greenhouse. The regenerants were morphologically normal and fertile. Flow cytometric analyses and chloroplast counts of guard cells suggested that the regenerants were diploid. Efficient cloning protocols described here, have the potential not only to substantially reduce the pressure on natural populations but also for wider biotechnological applications of Hypoxis hemerocallidea-an endangered medicinal plant.