Solanum nigrum is an underutilised vegetable that accumulates high leaf nutrient levels. Although numerous micropropagation protocols are reported for its mass production, there have been no follow-on studies detailing the pollen and seed viabilities of clones or clonal genotypes. It is important to determine these via-bilities in cases where the inheritance of desired traits in the F1 progenies of micropropagated clones are required. In the present study, two high Fe-accumulating S. nigrum genotypes (G6 and G15) identified in a previous investigation were clonally propagated and acclimatised until sexual maturity using an established protocol. Following flower and fruit production, viability tests were performed on clonal pollen (Lugol's staining) and seeds (tetrazolium chloride test) in addition to determinations of berry and seed numbers, seed set, seed length, weight, germination and ultrastructure. A wild type (WT) population was used as the con-trol. Compared to the WT population (82%, 75%, 200, 11,400, 57 and 51%), the G6 and G15 clones, respec-tively, resulted in significantly lower pollen (51 and 39%) and seed viabilities (4 and 17%), berry (23 and 16) and seed (162 and 116) numbers, seed sets (7 and 7) and germination (7 and 8%). Microscopic analyses revealed normal, abnormal, viable and nonviable pollen and seeds for G6, G15 and the WT in addition to abnormal or damaged embryonic and endosperm tissues for both clonal genotypes. Micropropagation of S. nigrum genotypes decreases their clonal pollen and seed viabilities, berry and seed production, and germina-tion success of their F1 progenies.(c) 2022 SAAB. Published by Elsevier B.V. All rights reserved.

Solanum nigrum is an orphan crop known for its medicinal and nutritional value and phenotypic plasticity. Four genotypes, previously identified as accumulating high levels of Ca (G5 and G20) and Fe (G6 and G15), were clonally propagated. Following acclimatization, the clonal progenies were subjected to two watering regimes (50 and 100% water holding capacities) in a greenhouse and clones of each genotype were harvested at 15, 30 and 45 days of ex-vitro growth. Full water supply and aging led to the greatest biomass production by the clonal genotypes while both watering regimes resulted in overall increases in leaf water contents. Ca and Fe contents were not affected by age or changing water availability except for the G6 clones which accumulated less Fe under water stress. Genotypic variations occurred for all tested parameters except Fe content (full water supply), and Ca content, root fresh and dry weights (water stress). With age, plasticities for biomass increased, those for leaf water content decreased and remained unaffected for Ca and Fe contents. Genotypic effects were observed for shoot and root dry weight plasticities (G6 > G15). The clones of G5, G6 and G20 could be grown at 50% WHC and harvested after 15 days of ex-vitro growth as full plants for consumption but since shoot biomass was found to be a critical factor influencing Ca and Fe content, a greater number of clones will need to be propagated to gain the maximum nutritional benefit at day 15.

This study aimed to screen and clonally propagate high and low Ca-and Fe-producing Amaranthus dubius genotypes and to test the resulting clonal progeny ex vitro for their Fe and Ca content. Seven genotypes (G) with varying levels of Ca and Fe were selected from 50 A. dubius seedlings and then micropropagated using nodal explants. They were acclimatised and grown in the greenhouse for 90 days on either soil or soil and vermiculite (1:1). Ca and Fe levels of the clones were determined at days 15, 30, 60, 80, and 90 following 21 days of acclimatisation and compared to those of 60-day-old parent genotypes. In general, the Ca levels of the clones of the tested genotypes were similar to those of their parents at day 15 and then increased up to 90 days, regardless of the substrate. In contrast, on both substrates, at day 15, the Fe levels were the same or higher in the clones compared with their parents and then decreased with time. Amongst the clonal genotypes, the mean plasticity indices (MPIs) were low and ranged from 0.12 to 0.43, 0.13 to 0.42, and 0.11 to 0.21 for shoot growth, Ca content, and Fe content, respectively. The clones of G41 were found to be the most plastic (MPI = 0.28), while the clones of G15 were the least plastic (MPI = 0.15). Overall, there is an indication that the number of leaves, their biomass, and their Ca and Fe contents are not negatively affected by low nutrient soils, which may be useful to communities who cannot afford to apply fertilisers.

Wild leafy vegetables are of increasing interest because many have higher concentrations of some beneficial minerals, vitamins and nutrients than commercial crops. Some reports also indicate that they exhibit high levels of tolerance to abiotic stressors associated with climate change. The aim of this study was to investigate whether selected biochemical, physiological (related to photosynthesis) and morphological traits could be used to screen for elevated temperature tolerance in two wild leafy vegetable species, viz. Amaranthus dubius (C-4) and Galinsoga parviflora (C-3). One-month-old seedlings were transferred from a greenhouse to growth chambers and subjected to either ambient [25 degrees C (day)/14 degrees C (night)] or elevated [30 degrees C (day)/19 degrees C (night)] temperatures for 15 days. On days 0, 5, 10 and 15, gas exchange, chlorophyll fluorescence, selected biochemical, physiological and morphological traits were determined. Data for the 24 traits measured were used in plant trait network analyses to establish the populations' phenotypic plasticity, including changes in the interactions/relationships amongst the traits when exposed to elevated temperatures. Sixteen traits showed significant differences between ambient and elevated temperatures on day 15, 11 in G. parviflora only, one in A. dubius only and four in both species. Those shared by the two species were intracellular hydrogen peroxide, electrolyte leakage, ascorbate peroxidase, and superoxide dismutase, while specific leaf area was significantly different in A. dubius only. Both species altered biochemical leaf traits under elevated temperatures, while morphological leaf traits were altered in A. dubius only and photosynthetic leaf traits in G. parviflora only. In G. parviflora, the photosynthetic traits were more sensitive to elevated temperatures than in A. dubius. This suggests that photosynthetic traits may be ineffective for screening C-4 species, due to them being highly evolved to survive in warmer climates. However, the results showed that photosynthetic traits could be useful as screening tools for C-3 photosynthetic pathway species. The plant trait network analyses showed that both species displayed high phenotypic plasticity with a 5 degrees C increase in temperature. We recommend when screening wild leafy vegetables for high temperature tolerance that photosynthetic traits be considered in the context of their phenotypic plasticity. This approach could fast-track the identification of more climate-resilient wild edible species. (c) 2022 SAAB. Published by Elsevier B.V. All rights reserved.

Mutagenic breeding is an approach being developed especially for those characteristics that are not inherited through conventional genetic hybridization. Soil acidity and aluminum (Al3+) toxicity limit sugarcane production, particularly in leached soils in the coastal cultivation regions in South Africa. An in vitro screening procedure was developed based on plantlet re-rooting in Al3+-containing medium to select putative Al3+ tolerant variants produced by induced mutagenesis. A preliminary test did not show any difference among three cultivars in their ability to re-root in the presence of Al3+. Subsequent work focused on Saccharum sp. ‘NCo376’ and two Al3+ concentrations (15 and 45 mM), representing LC50 and LC90 re-rooting inhibition. In an attempt to increase the stringency of the selection conditions, the possible effects of inorganic nitrogen (N) forms in the culture media on Al3+ toxicity were investigated. The form of N affected root regrowth and citrate exudation. Sugarcane plantlets showed an uptake preference for ammonium (NH4+) over nitrate (NO3−), and 10 and 20 mM NH4+ appeared to alleviate Al3+ toxicity compared with NO3−. Although in agricultural soils NO3− is the dominant form of N, it can readily leach into the sub-soil, in which deep roots encounter Al3+ toxicity in the presence of NO3−. Therefore, the ability to withstand the toxic effects of Al3+ in the presence of NO3− could be beneficial for N use efficiency and access to water at depths during drought.

The production of regenerable embryogenic callus is essential to sugarcane genetic improvement. However, some sugarcane cultivars display poor calli yields using established in vitro protocols. In this study, we tested the impact of methylglyoxal (MG) on embryogenic callus and plantlet development in cultivars NCo376 and N41. Calli were exposed to 0–10 mM MG at embryo maturation and germination stages. For both cultivars, the 2 and 4 mM MG treatments increased callus dry mass by up to 48%, but 70–80% decreases were observed when calli were exposed to 7 and 10 mM. The 2 and 4 mM MG treatments also produced more compact white embryogenic callus than the control. Incorporation of MG at the same levels during embryo germination promoted faster shoot morphogenesis in both cultivars and increase plantlet yield in NCo376 by 130% when treated with 4 mM MG. In both cultivars, MG levels higher than 7 mM had a negative or no effect plantlet production. Metabolic profiling revealed higher levels of sugars in MG-treated than in control calli, which may have contributed to development of more white compact calli. Separate clustering of NCo376 and N41 MG-treated calli in principal component and hierarchical clustering analyses of the metabolic profiles, suggested variations in MG metabolism among the genotypes that may account for variations in the MG-induced effect on somatic embryogenesis between the two cultivars. Although the effect may be genotype-dependant, low MG concentrations can induce improved embryogenic callus and plantlet development in sugarcane.

Simple in vitro protocols were developed or improved for in vitro manipulations of sugarcane (Saccharum species hybrids) to free up resources, deal with back-logs and increase throughput in research and commercial laboratories. These include: (1) in vitro germination of field-derived axillary buds for meristem isolation; (2) a 'common' medium for rapid shoot multiplication resulting from single genetic modification events; (3) shoot storage for 12 months; (4) 'holding-back' in vitro plantlets before acclimation. The novel strategies have proven reliable and effective in managing changing demands on resources and priorities.

The aims and objectives of the current study were to investigate the level of genotypic variation in the leaf calcium (Ca) and iron (Fe) contents within a relatively small population of Solanum nigrum, to select high and low Ca- and Fe-producing genotypes and clonally propagate them in vitro and to test the resulting clonal progeny ex vitro for their Ca and Fe content and growth, based on their physiological age and on the use of different substrates. Firstly, chilled and fresh S. nigrum seeds were acid-scarified and sown onto potting soil in a greenhouse. Optimal germination (95%) was achieved in seeds chilled for 3 weeks and scarified with 0.01 M HCl for 10 min. Thereafter, fifty 6-week old seedlings were screened using Inductively Coupled Plasma-Optical Emission Spectrometry resulting in ranges of leaf calcium (Ca) and iron (Fe) of 331.0-916.3 mg 100 g(-1) dry mass (DM) and 0.64-14.95 mg 100 g(-1) DM, respectively. When a micropropagation protocol using seedling nodal and leaf explants was investigated, direct organogenesis using leaves on shoot multiplication medium containing 3 mg l(-1) benzylaminopurine (BAP) and rooting on hormone-free MS medium was most successful resulting in 80% explants producing shoots, 50 shoots/explant and 80% rooted shoots. After acclimatisation in nutrient-rich potting soil, the yield was 25 plants/explant. Subsequently, genotypes for high Ca (G20), high Fe (G15), low Ca (G43) and low Fe (G35) were micropropagated using the established protocol. At certain physiological ages, G43 was the only genotype whose clones had better shoot and root growth on the nutrient-poor substrate than on the nutrient-rich one while G35 was the only genotype whose clones had better root growth on the nutrient-rich substrate than on the nutrient-poor one. The clones of G20 produced high numbers of leaves and shoot and root fresh and dry masses on both substrate types but the overall growth was better on soil. For the G15 clones, shoot and root growth increased with age with no overall difference in growth between the substrates. The levels of phenotypic plasticity for the selected genotypes were found to be G43 > G35 > G20 > G15 and in cases where phenotypic plasticity was observed, more clones with higher overall biomass were found on the nutrient-poor substrate. When compared to their respective parents, the G20 clones produced the same amount of Ca on both tested substrates while G43 exhibited higher Ca production on the nutrient-rich substrate. The Fe levels in the clones of G15 decreased with physiological age, and were lower than that of the parent plant, on both the nutrient-rich and poor substrates while the Fe levels of the G35 clones were higher than that of the parent plant on nutrient-rich soil at all ages. (c) 2018 SAAB. Published by Elsevier B.V. All rights reserved.

Mutagenesis had no effect on number of stalks/plot, stalk height, fibre and sucrose content of mutants. Imazapyr tolerance is likely due to a S622N mutation in the acetolactate synthase gene.

To-date, assessments of nitrogen use efficiency (NUE) of sugarcane have not included the contribution of its components, nitrogen uptake efficiency (NUpE) and nitrogen utilization efficiency (NUtE). This study determined these values, based on biomass and plant nitrogen (N) content, in two four-month-old pot-grown genotypes. The treatments included six N regimes, with nitrate (NO3-N) or ammonium (NH4+-N) supplied alone, or as NO3--N for the first 6 weeks and then NH4+-N until harvest, each as 4 or 20 mM. Regardless of the N form, NUE was higher at four than at 20 mM due to significantly higher NUpE at low N supply. The results indicated that there was luxury N uptake and preference for NH4+-N nutrition, which resulted in the highest determined NUE. There were significant differences between genotypes in biomass, morphological growth parameters, N uptake, total plant N and NUE, the latter matching previously established sucrose yield-based NUE field rankings.

Successful adventitious root induction from macro-, mini- and micro-cuttings is critical for Eucalyptus forestry establishments. As for all the aspects of plant development, adventitious rooting is controlled by plant growth regulators (PGRs), primarily the auxins and their interaction with the cytokinins. Nevertheless, the approach to eucalyptus root induction from vegetative tissues remains empirical, with protocols that are often only clone-specific. Recent advancements have allowed for a more guided approach with respect to exogenous PGR supply for root induction. This review explores the genetic and phytohormone control of root development as it relates to woody plants and commercially-important Eucalyptus clones, and offers some suggestions with respect to the selection of PGRs for enhanced root induction and subsequent development.

Seven imazapyr-tolerant mutant sugarcane plants, previously generated by in vitro mutagenesis, were studied. The imazapyr concentrations required to inhibit their acetolactate synthase (ALS basal activity) (IC50 as μmoles acetoin h-1 mg-1 protein) were 0.77 - 5.36 times greater than that of the N12 ‘parent’. The basal ALS activities of Mut1 and Mut6 were 1.4-fold higher than that of N12. When the mutants were sprayed with Arsenal® GEN 2 (312 and 624 g a.i. imazapyr ha-1), 2 months after field planting, and evaluated 9 months later, live stalk height and number were significantly lowest in Mut2, Mut3 and the control N12. No differences in sucrose, fibre and estimated yield were observed amongst lines in untreated plots. Mutant plants germinated and grew in soil treated with the herbicide (at the lethal dose of 1248 g a.i. ha-1). The Mut lines tested in this study offer improved options for weed control.

Six cultivars with a range of known field responses to drought were tested in vitro. Firstly, the two on each end of the drought tolerance scale (NCo376, most susceptible and N41, most tolerant) were screened on 204–1493 mM mannitol for  % re-rooting after 10 days, following plantlet root trimming. From the results, the LD50 and LD90 of re-rooting were calculated as 332 and 606 mM mannitol for NCo376 and 851 and 1493 mM mannitol for N41, respectively. These mannitol concentrations were then used as benchmarks for testing four other cultivars. At 606 mM mannitol, % re-rooting in vitro was: N41 (80 ± 10.7), N26 (80 ± 10.7), N36 (60 ± 13.1), N12 (60 ± 13.1), N19 (20 ± 10.7) and NCo376 (13.3 ± 9.1) (p < 0.05). The consequent ‘ranking’ in terms of mannitol tolerance in vitro matched well with those of field observations for drought tolerance, with N41 being the most drought tolerant and NCo376 the most susceptible. Other parameters assessed as possible drought indices such as leaf chlorophyll content (by soil plant analysis development measurements) and histochemical staining were difficult to perform on the small in vitro leaves, and relative electrolyte leakage did not distinguish drought tolerant from susceptible cultivars. Therefore, the re-rooting assessment provides a rapid (3 weeks) ranking measure and has potential application when screening mutagenic and genetically modified lines.