Molecular profiling of microbial communities associated with seeds of Beta vulgaris subsp. Vulgaris (sugar beet)

The composition of microbial communities on and within seeds may effect their storage and field performance, whether they are indigenous or applied as biocontrol agents. In this study, we have explored the usefulness of profiling small subunit ribosomal (SSR) gene fragments for studying the microflora associated with seeds. DNA was amplified by the polymerase chain reaction (PCR) and the amplicons separated using denaturing gradient gel electrophoresis (DGGE). Primers targeting eukaryotic SSRs were used to investigate fungal communities, and primers targeting bacterial SSRs were employed to study the eubacterial microflora. As a case study, we attempted to profile the fungi and bacteria associated with seeds of Beta vulgaris (sugar beet) to permit an insight into the varying field performance of several well-characterised commercial seed lots. Serious interference with the microbial signals was observed from the plant's own nuclear 18S rRNA genes and chloroplast 16S rRNA genes using standard PCR conditions and DNA extracted from whole seeds as template. Hot-start and touchdown PCR made no appreciable improvement to these signals. Seed imbibition and dissection into operculum and fruit wall and true seed prior to DNA extraction improved signal recovery in the fruit fraction. With primer modification, bacteria and fungi were detected in an excess of plant DNA of 100:1 and 10:1, respectively. With this method, microbial communities on seeds could be profiled, however, it is likely that targeted depletion of plant rDNA targets will be a necessary extra step before this approach can be used to screen seeds routinely.     

The impact of mechanical impedance on the emergence of carrot and onion seedlings

The development of a method using wax layers to simulate the effect of strong soil crusts on seedling emergence is described. Wax layers of different strengths were prepared by melting together white soft paraffin and paraffin wax in different proportions. The wax discs were placed above seeds planted in wet, but well-aerated sand in controlled environments. The effect of adding charcoal to the wax discs to prevent transmission of light was also tested. When wax layer penetrometer pressure was 0.2–0.25 MPa, light transmission greatly decreased the final emergence of carrot, but not onion seedlings. Penetrometer pressures above 0.25 MPa greatly decreased emergence through black wax layers in both carrot and onion. The presence of 2–4 mm stones immediately below wax layers (to simulate aggregates) decreased the emergence of onion shoots through wax layers with penetrometer pressures of 0.25 MPa and above. The emergence of carrot and onion seedlings from wax layers was compared to emergence in the field from different soil types and aggregate sizes. In both laboratory and field experiments, carrot gave better emergence than onion when emergence was relatively poor. Results were consistent with the hypothesis that mechanical impedance is a major factor in poor crop emergence in temperate conditions.     

The effect of temperature on reproduction in the summer and winter annual Arabidopsis thaliana ecotypes Bur and Cvi

Background and Aims

Seed yield and dormancy status are key components of species fitness that are influenced by the maternal environment, in particular temperature. Responses to environmental conditions can differ between ecotypes of the same species. Therefore, to investigate the effect of maternal environment on seed production, this study compared two contrasting Arabidopsis thaliana ecotypes, Cape Verdi Isle (Cvi) and Burren (Bur). Cvi is adapted to a hot dry climate and Bur to a cool damp climate, and they exhibit winter and summer annual phenotypes, respectively.

Methods

Bur and Cvi plants were grown in reciprocal controlled environments that simulated their native environments. Reproductive development, seed production and subsequent germination behaviour were investigated. Measurements included: pollen viability, the development of floral structure, and germination at 10 and 25 °C in the light to determine dormancy status. Floral development was further investigated by applying gibberellins (GAs) to alter the pistil:stamen ratio.

Key Results

Temperature during seed development determined seed dormancy status. In addition, seed yield was greatly reduced by higher temperature, especially in Bur (>90 %) compared with Cvi (approx. 50 %). The reproductive organs (i.e. stamens) of Bur plants were very sensitive to high temperature during early flowering. Viability of pollen was unaffected, but limited filament extension relative to that of the pistils resulted in failure to pollinate. Thus GA applied to flowers to enhance filament extension largely overcame the effect of high temperature on yield.

Conclusions

High temperature in the maternal environment reduced dormancy and negatively affected the final seed yield of both ecotypes; however, the extent of these responses differed, demonstrating natural variation. Reduced seed yield in Bur resulted from altered floral development not reduced pollen viability. Future higher temperatures will impact on seed performance, but the consequences may differ significantly between ecotypes of the same species.     

Development of combined imbibition and hydrothermal threshold models to simulate maize (Zea mays) and chickpea (Cicer arietinum) seed germination in variable environments

* The ability of hydrothermal time (HTT) and virtual osmotic potential (VOP) models to describe the kinetics of maize (Zea mays) and chickpea (Cicer arietinum) seed germination under variable conditions of water potential was investigated with a view to gaining an improved understanding of the impact of on-farm seed priming on seedling establishment through simulation. * Germination and/or imbibition time courses were recorded over a wide range of constant temperatures and water potentials and simple stepwise changes in water potential. * Both models adequately described germination under constant environmental conditions, but not conditions of water potential that varied. To test the hypothesis that this inaccuracy resulted from the use of ambient water potential, a parsimonious model of seed imbibition was developed to calibrate the HTT and VOP models (IHTT and IVOP) and drive them with estimates of seed water potential. * The IHTT and IVOP models described germination during stepwise changes in water potential more accurately than the conventional models, and should contribute to improved predictions of germination time in the field.     

Development of a hydrothermal time model that accurately characterises how thermoinhibition regulates seed germination

Thermoinhibition is the decline in germinability within a seed population as soil temperatures increase above the optimum for germination. Hydrothermal time (HTT) models have been developed that describe the thermoinhibition response as a function of increases in the threshold water potential for seed germination [seed base water potential, Ψ(b) (G)]. Although these models assume a normal distribution of Ψ(b) (G) and a linear upward shift in Ψ(b) (G) with increasing temperature, little research has tested these assumptions. Using germination data obtained from four unrelated plant species, we fitted HTT models that use the Weibull and normal distribution to describe Ψ(b) (G) and compared the accuracy and bias of these two HTT models. For all four species, Ψ(b) (G) and germination were more accurately described by the Weibull than the normal distribution HTT model. At supra-optimal temperatures, Ψ(b) (G) of the earliest germinating seeds showed little thermoinhibition effect so that the seeds germinated very rapidly under moist conditions. However, for the rest of the population, Ψ(b) (G) increased progressively in response to supra-optimal temperatures so that the slower germinating seeds were thermoinhibited. The fitted HTT models reveal aspects of seed thermoinhibition that appear to have adaptational value under variable conditions of soil temperature and moisture.     

Seed reserve-dependent growth responses to temperature and water potential in carrot (Daucus carota L.)

Both temperature and soil moisture vary greatly in the surface layers of the soil through which seedlings grow following germination. The work presented studied the impact of these environmental variables on post-germination carrot growth to nominal seedling emergence. The rapid pre-crook downward growth of both the hypocotyl and root was consistent with their requirement for establishment in soil drying from the surface. At all temperatures, both hypocotyl and root growth rates decreased as water stress increased and there was a very distinct temperature optimum that tended to occur at lower temperatures as water stress increased. A model based on the thermodynamics of reversible protein denaturation was adapted to include the effects of water potential in order to describe these growth rate responses. In general, the percentage of seedlings that reached the crook stage (start of upward hypocotyl growth) decreased at the extremes of the temperature range used and was progressively reduced by increasing water stress. A model was developed to describe this response based on the idea that each seedling within a population has lower and upper temperature thresholds and a water potential threshold which define the conditions within which it is able to grow. This threshold modelling approach which applies growth rates within a distribution of temperature and water potential thresholds could be used to simulate seedling growth by dividing time into suitable units.