Foliar application of plant growth regulators changes the physiological quality of crambe seeds

The application of plant growth regulators can manipulate the development of cultivated plants, and their physiological effects may be reflected in the physiological quality of the seeds. This study evaluated the effects of plant growth regulators (PGR) on the physiological quality of crambe seeds produced in the 2014 and 2015 harvests. During crop development, we applied two foliar sprayings of the following treatments: 1) control treatment with distilled water; 2) indole-3-acetic acid 100 mg L-1 (IAA); 3) 3-gibberellic acid P.A. 100 mg L-1 (GA3); 4) commercial PGR Stimulate® 6 mL L-1. The seeds were harvested and evaluated for water content, germination percentage and germination speed index (GSI), electrical conductivity and concentrations of leached Ca, Mg and K ions. Means were compared by Tukey ́s and Dunnett tests (p<0.05). Applying Stimulate® increased the germination percentage and the germination speed index, while IAA application showed an opposite effect, increasing the electrical conductivity of crambe seeds. Calcium leaching decreased with IAA application, and all regulators reduced K leaching. We therefore infer that the application of plant growth regulators influences the physiological quality of crambe seeds.

Genetic stability of micropropagated plants of Crambe abyssinica Hochst using ISSR markers.

Crambe (Crambe abyssinica) is a non-edible annual herb, which was first cultivated to extract oil for industry, and now has great potential for biodiesel production. The objective of this investigation was to evaluate the genetic stability of micropropagated plants of the C. abyssinica Hochst cultivar 'FMS brilhante' using polymerase chain reaction techniques based on inter-simple sequence repeat (ISSR) molecular markers. The aim was to develop a protocol for the in vitro regeneration of these plants with low genetic variation as compared to the donor plant. For micropropagation, shoot tips from in vitro germinated seedlings were used as explants and were initially cultivated for 90 days on MS medium with 5.0 µM 6-benzylaminopurine (BAP), which at 90 days, led to the highest number of shoots per explant (NSE) (12.20 shoots) being detected. After 120 days, the interaction between BAP concentration and naphthalene acetic acid (NAA) was tested, and the highest NSE was observed following exposure to 0.0/0.5 µM BAP/NAA (11.40 shoots) and 1.0/0.0 µM BAP/NAA (11.00 shoots). The highest proportion of rooting phase were observed following exposure to 0.5 µM NAA (30%). The 13 ISSR primers used to analyze genetic stability produced 91 amplification products, of which only eight bands were polymorphic and 83 were monomorphic for all 10 regenerated crambe plants, compared to the donor plant explant. These results indicate that crambe shoot tips are a highly reliable explant that can be used to micropropagate genetically true-to-type plants or to maintain genetic stability, as verified using ISSR markers.

Reduced Silver Nanoparticle Phytotoxicity in Crambe abyssinica with Enhanced Glutathione Production by Overexpressing Bacterial γ-Glutamylcysteine Synthase.

Silver nanoparticles (Ag NPs) are widely used in consumer products, and their release has raised serious concerns about the risk of their exposure to the environment and to human health. However, biochemical mechanisms by which plants counteract NP toxicity are largely unknown. We have previously engineered Crambe abyssinica plants expressing the bacterial γ-glutamylecysteine synthase (γ-ECS) for enhancing glutathione (GSH) levels. In this study, we investigated if enhanced levels of GSH and its derivatives can protect plants from Ag NPs and AgNO3 (Ag(+) ions). Our results showed that transgenic lines, when exposed to Ag NPs and Ag(+) ions, were significantly more tolerant, attaining a 28%-46% higher biomass and 34-49% more chlorophyll content, as well as maintaining 35-46% higher transpiration rates as compared to those of wild type (WT) plants. Transgenic γ-ECS lines showed 2-6-fold Ag accumulation in shoot tissue and slightly lower or no difference in root tissue relative to levels in WT plants. The levels of malondialdehyde (MDA) in γ-ECS lines were also 27.3-32.5% lower than those in WT Crambe. These results indicate that GSH and related peptides protect plants from Ag nanotoxicity. To our knowledge, this is the first direct report of Ag NP detoxification by GSH in transgenic plants, and these results will be highly useful in developing strategies to counteract the phytotoxicty of metal-based nanoparticles in crop plants.

Plant-pollinator interactions in Crambe abyssinica Hochst. (Brassicaceae) associated with environmental variables.

Despite the economic importance of Crambe abyssinica, the present study is the first report on bees that occur with this species, and could aid in developing alternative methods for controlling insect pests without seriously impacting pollinators. The present study examined the following questions: (1) Which species are potential pollinators of C. abyssinica? (2) How do environmental conditions influence pollinator visitation fluctuations? Insects were sampled on a weekly basis between 08:00 and 16:00 during five weeks of flowering. When the results of analyses of variance were significant, the data was adjusted using regression equations at a 5% level of probability; the environmental variables were examined using Pearson's correlation coefficient. Apis mellifera, Geotrigona mombuca, Exomalopsis (Exomalopsis) fulvofasciata, Plebeia sp. and Dialictus sp. were observed visiting C. abyssinica flowers. A. mellifera and G. mombuca were observed to be potential pollinators, with the former demonstrating visitation peaks during flowering weeks II and IV at 12:00 and 08:00, respectively and the latter visiting during weeks III and IV at 12:00 and 10:00, respectively. Environmental factors such as temperature, relative humidity, and wind speed have different effect on the activity of bees.

Bottlenecks in erucic acid accumulation in genetically engineered ultrahigh erucic acid Crambe abyssinica.

Erucic acid is a valuable industrial fatty acid with many applications. The main producers of this acid are today high erucic rapeseed (Brassica napus) and mustard (Brassica juncea), which have 45%-50% of erucic acid in their seed oils. Crambe abyssinica is an alternative promising producer of this acid as it has 55%-60% of erucic acid in its oil. Through genetic modification (GM) of three genes, we have previously increased the level of erucic acid to 71% (68 mol%) in Crambe seed oil. In this study, we further investigated different aspects of oil biosynthesis in the developing GM Crambe seeds in comparison with wild-type (Wt) Crambe, rapeseed and safflower (Carthamus tinctorius). We show that Crambe seeds have very low phosphatidylcholine-diacylglycerol interconversion, suggesting it to be the main reason why erucic acid is limited in the membrane lipids during oil biosynthesis. We further show that GM Crambe seeds have slower seed development than Wt, accompanied by slower oil accumulation during the first 20 days after flowering (DAF). Despite low accumulation of erucic acid during early stages of GM seed development, nearly 86 mol% of all fatty acids accumulated between 27 and 50 DAF was erucic acid, when 40% of the total oil is laid down. Likely bottlenecks in the accumulation of erucic acid during early stages of GM Crambe seed development are discussed.