Effect of the environment during seed development on brassica seed quality

High quality seed is essential for the establishment of a good crop. New Zealand grown brassica seeds usually have high germination but often have variable seed vigour. The latter can result in poor crop establishment and storability. High temperature stress during seed development is known to reduce seed vigour in some species, but whether temperature stress is responsible for seed vigour loss in brassica species was not known. The effects of high temperature during seed development on forage rape (Brassica napus) seed quality were determined by assessing seed mass, germination and vigour using a sowing date trial and field and controlled environment experiments. A time of sowing trial was conducted in the 2011-12 season. A late flowering forage rape cultivar “Greenland” was sown on 25 March and 13 April, 2011 with sowings replicated four times in a randomized complete block design. Seed quality was assessed at three seed development stages (determined by seed moisture content (SMC)): at physiological maturity (PM) (≈50% SMC), pre-desiccation final stage (≈25% SMC) and harvest maturity (≈14% SMC). Seed had attained PM at between 47-52% SMC which was similar to other brassica species. The seed quality testing results demonstrated that sowing time had no effect on seed germination in the prevailing environmental conditions in that season, and at PM there were no differences in seed vigour. However, seed vigour was significantly reduced in seeds harvested at the pre-desiccation (≈25% SMC) and harvest maturity (HM) (≈14% SMC) stages for the early sowing. This was explained by a longer time of exposure to conditions which caused weathering during maturation for the March sowing. In a controlled growth room, set at 30/25 ˚C (day/night, 12 hours each, R.H 70%), plants received heat stress for four days (240 ˚Ch) at (i) seed filling ii) PM and iii) seed filling plus PM before being returned to the field until seed harvest for two consecutive seasons, 2011-12 and 2012-13. Heat stress decreased seed quality in all three treatments. In both years seed vigour was adversely affected by the heat stress, but seed germination was not. High temperature stress during seed filling produced smaller seeds but this did not occur with heat stress at PM. Seed developed at the top of the raceme was smaller and had lower germination compared with seed developed at the middle and basal raceme positions. This difference in seed quality between raceme positions became greater after heat stress. A field trial was conducted in the same two seasons with artificially created high field temperature conditions (using plastic sheet cages) during forage rape seed development. The heat stress was imposed during phase-I (seed filling to PM) and phase-II (PM to HM) and at both Phase-I+II. Heat stress during phase-I significantly reduced seed germination, vigour and seed mass, confirming the results of the controlled environment experiment. Imposition of heat stress during phase-II (after PM), however, significantly reduced seed germination and vigour but did not affect seed mass. Hourly thermal time (HTT) at a base temperature (Tb) of 25 ˚C and the number of hours that temperature remained above 25 ˚C during phase-II (from PM to HM) were significantly correlated with germination and vigour, but not seed mass. The data suggested that for a Tb of 25 ˚C, at least 100 ˚Ch before PM and 300 ˚Ch after PM were required before vigour loss occurred. The effects of high temperature during seed development were further studied at a physiological and ultrastructural level using heat stressed and non-stressed seeds from the controlled environment experiment. Both reactive oxygen species (ROS) (H2O2) and lipid peroxidation were measured. H2O2 and malondialdehyde (MDA) were both significantly higher in heat stressed seeds than in non- stressed seeds. Loss of seed vigour was associated with an accumulation of H2O2 and lipid peroxidation. H2O2 in heat stressed seeds was strongly correlated with seed vigour loss, suggesting that lipid peroxidation was not the only cause of seed deterioration. Seed vigour loss was also characterized by a marked decrease in the ROS scavenging antioxidant enzymes, superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR) activities following heat stress. A significant negative effect of heat stress on the adenine nucleotides pool and adenylate energy charge (AEC) was recorded which indicated the altered metabolic system. This was mainly due to a decrease in cellular adenosine triphosphate (ATP), resulting in a decrease of AEC. Electron microscopy revealed significant cellular damage in heat stressed seeds, particularly in the cell membranes and mitochondria. The decreased level of nucleotides and energy levels, and higher electrolyte leakage recorded in heat stressed seeds was associated with this structural damage. Mitochondrial ATP synthesis provides an important source of energy to complete the germination process. The mitochondrial damage in this study as a result of heat stress suggests that the mitochondria were unable to synthesize sufficient energy for the active oxidative phosphorylation required to complete successful germination