Annie Claessens

Alleviation of Drought Stress and Metabolic Changes in Timothy (Phleum pratense L.) Colonized with Bacillus subtilis B26

Drought is a major limiting factor of crop productivity worldwide and its incidence is predicted to increase under climate change. Drought adaptation of cool-season grasses is thus a major challenge to secure the agricultural productivity under current and future climate conditions. Endophytes are non-pathogenic plant-associated bacteria that can play an important role in conferring resistance and improving plant tolerance to drought. In this study, the effect of inoculation of the bacterial endophyte Bacillus subtilis strain B26 on growth, water status, photosynthetic activity and metabolism of timothy (Phleum pratense L.) subjected to drought stress was investigated under controlled conditions. Under both drought-stress and non-stressed conditions, strain B26 successfully colonized the internal tissues of timothy and had a positive impact on plant growth. Exposure of inoculated plant to a 8-week drought-stress led to significant increase in shoot and root biomass by 26.6 and 63.8%, and in photosynthesis and stomatal conductance by 55.2 and 214.9% respectively, compared to non-inoculated plants grown under similar conditions. There was a significant effect of the endophyte on plant metabolism; higher levels of several sugars, notably sucrose and fructans and an increase of key amino acids such as, asparagine, glutamic acid and glutamine were recorded in shoots and roots of colonized plants compared to non-colonized ones. The accumulation of the non-protein amino acid GABA in shoots of stressed plants and in roots of stressed and unstressed plants was increased in the presence of the endophyte. Taken together, our results indicate that B. subtilis B26 improves timothy growth under drought stress through the modification of osmolyte accumulation in roots and shoots. These results will contribute to the development of a microbial agent to improve the yield of grass species including forage crops and cereals exposed to environmental stresses.

Genomic Prediction of Biomass Yield in Two Selection Cycles of a Tetraploid Alfalfa Breeding Population

Alfalfa (Medicago sativa L.) is a widely planted perennial forage legume grown throughout temperate and dry subtropical regions in the world. Long breeding cycles limit genetic improvement of alfalfa, particularly for complex traits such as biomass yield. Genomic selection (GS), based on predicted breeding values obtained using genome‐wide molecular markers, could enhance breeding efficiency in terms of gain per unit time and cost. In this study, we genotyped tetraploid alfalfa plants that had previously been evaluated for yield during two cycles of phenotypic selection using genotyping‐by‐sequencing (GBS). We then developed prediction equations using yield data from three locations. Approximately 10,000 single nucleotide polymorphism (SNP) markers were used for GS modeling. The genomic prediction accuracy of total biomass yield ranged from 0.34 to 0.51 for the Cycle 0 population and from 0.21 to 0.66 for the Cycle 1 population, depending on the location. The GS model developed using Cycle 0 as the training population in predicting total biomass yield in Cycle 1 resulted in accuracies up to 0.40. Both genotype × environment interaction and the number of harvests and years used to generate yield phenotypes had effects on prediction accuracy across generations and locations, Based on our results, the selection efficiency per unit time for GS is higher than phenotypic selection, although accuracies will likely decline across multiple selection cycles. This study provided evidence that GS can accelerate genetic gain in alfalfa for biomass yield.

Relationship between soil nitrate accumulation and in-season corn N nutrition indicators

Ziadi, N., Bélanger, G. and Claessens, A. 2012. Relationship between soil nitrate accumulation and in-season corn N nutrition indicators. Can. J. Plant Sci. 92: 331-339. Nitrogen management tools are required to optimize crop growth and yield while minimizing the likelihood of N losses to the environment. We previously determined that non-limiting N conditions for near maximum corn (Zea mays L.) grain yield are reached with the following threshold values for three in-season plant-based indicators of corn N nutrition determined at approximately the V12 stage of development: N nutrition index (NNI)=0.88, leaf N (NL) concentration =32.7 mg N g-1 leaf DM, and relative chlorophyll meter (RCM) values =0.95. Our objective was to study the relationship between these plant-based indicators and soil NO3-N content in an effort to develop tools to reduce the likelihood of soil NO3-N accumulation without affecting grain yield. This study at 5 site-years in Québec consisted of six N fertilizer rates (20-250 kg N ha-1). The NNI, NL concentrations, RCM values, and soil (0-0.15 m) NO3-N content were measured weekly from July to early August, while soil NO3-N content to a 0.90-m depth was measured in late August and October. During the growing season from July to early August, the proportion of data points above the average soil NO3-N content was greater under non-limiting N conditions (NNI ≤ 0.88, NL concentrations ≤ 32.7 mg N g-1 leaf DM, or RCM values =0.95) than under limiting N conditions. Furthermore, the mean soil NO3-N content of the data points above the general average was much higher under non limiting than limiting N conditions in late August (167 vs. 78 kg NO3-N ha-1 for NNI and RCM; 166 vs. 112 kg NO3-N ha-1 for NL concentration) and October (68 vs. 49 kg NO3-N ha-1). High soil NO3-N accumulation during the season and at harvest occurs only when in-season plant-based N indicators are greater than their threshold values.

Effect of photoperiod prior to cold acclimation on freezing tolerance and carbohydrate metabolism in alfalfa (Medicago sativa L.)

Cold acclimation proceeds sequentially in response to decreases in photoperiod and temperature. This study aimed at assessing the impact of photoperiod prior to cold acclimation on freezing tolerance and related biochemical and molecular responses in two alfalfa cultivars. The fall dormant cultivar Evolution and semi-dormant cultivar 6010 were grown in growth chambers under different photoperiods (8, 10, 12, 14 or 16h) prior to cold acclimation. Freezing tolerance was evaluated as well as carbohydrate concentrations, levels of transcripts encoding enzymes of carbohydrate metabolism as well as a K-3dehydrin, before and after cold acclimation. The fall dormant cultivar Evolution had a better freezing tolerance than the semi-dormant cultivar 6010. The effect of photoperiod prior to cold acclimation on the level of freezing tolerance differed between the two cultivars: an 8h-photoperiod induced the highest level of freezing tolerance in Evolution and the lowest in 6010. In Evolution, the 8h-induced superior freezing tolerance was associated with higher concentration of raffinose-family oligosaccharides (RFO). The transcript levels of sucrose synthase (SuSy) decreased whereas those of sucrose phosphatase synthase (SPS) and galactinol synthase (GaS) increased in response to cold acclimation in both cultivars. Our results indicate that RFO metabolism could be involved in short photoperiod-induced freezing tolerance in dormant alfalfa cultivars.

Breeding for Improved Nonstructural Carbohydrates in Alfalfa

Extensive degradation of proteins combined with insufficient readily fermentable energy in alfalfa (Medicago sativa L.) results in poor forage N utilization by ruminants and substantial N losses into the environment. Selection for increased nonstructural carbohydrate (NSC) concentration in forage crops is considered to be an effective approach to improve the feeding value of forage. Our objective was to assess the extent of genetic variability for NSC concentration and to identify DNA polymorphisms associated with that trait in alfalfa. A divergent selection was applied for forage NSC concentration among 560 alfalfa genotypes. Ten plants with high (NSC+), 10 plants with low (NSC-) and 10 plants randomly selected within the base population (NSC0) were selected to make three populations. Genetic variability for NSC concentration in alfalfa was observed (ranging from 73 to 230 mg/g DM under field conditions). Genotypes that consistently maintained high or low NSC concentrations were singled out under repeated assessments and were subsequently confirmed with clonal propagules. Although it is effective, phenotypic selection for NSC is resource intensive. Our identification of DNA polymorphism associated with this trait suggests the possibility of developing marker-assisted approaches to facilitate the identification of superior genotypes.

An Indoor Screening Method for Reduced Fall Dormancy in Alfalfa

Fall dormancy (FD) is a determinant component of alfalfa yield in northern climate, but selection for that trait is often done at the expense of freezing tolerance. Field selection for low dormancy is a tedious process that depends on unpredictable fall conditions to identify less dormant genotypes. Here we describe two screening methods for low dormancy in alfalfa which can be entirely performed indoor. Both methods rely on plant exposure to a critical photoperiod during four weeks to discriminate plants according to their FD level. One method also includes successive freezing stress to simultaneously improve freezing tolerance (TF) and FD. These approaches can be integrated in a recurrent selection program to produce populations combining reduced FD and high TF.