Host-specific competitiveness to form nodules in Rhizobium leguminosarum symbiovar viciae

Fabeae legumes such as pea and faba bean form symbiotic nodules with a large diversity of soil Rhizobium leguminosarum symbiovar viciae (Rlv) bacteria. However, bacteria competitive to form root nodules (CFN) are generally not the most efficient to fix dinitrogen, resulting in a decrease in legume crop yields. Here, we investigate differential selection by host plants on the diversity of Rlv. A large collection of Rlv was collected by nodule trapping with pea and faba bean from soils at five European sites. Representative genomes were sequenced. In parallel, diversity and abundance of Rlv were estimated directly in these soils using metabarcoding. The CFN of isolates was measured with both legume hosts. Pea/faba bean CFN were associated to Rlv genomic regions. Variations of bacterial pea and/or faba bean CFN explained the differential abundance of Rlv genotypes in pea and faba bean nodules. No evidence was found for genetic association between CFN and variations in the core genome, but variations in specific regions of the nod locus, as well as in other plasmid loci, were associated with differences in CFN. These findings shed light on the genetic control of CFN in Rlv and emphasise the importance of host plants in controlling Rhizobium diversity

Inoculation requirement of pea and faba bean and selection of Rlv strains

Soil is an heterogeneous and fluctuating environment. Water limitation in agronomic soils is often partial but lead locally to the suppression of symbiotic activity and therefore to reduction of N acquisition by the plant. A split-root system was developed in pea to evaluate the capacities of Pea-Rhizobium leguminosarum bv viciae (Rlv) symbiotic associations to compensate a local reduction of water availability. Root systems are split in two parts watered separately. Soil water content was monitored. Water limitation was exerted by stopping watering of a half root system. Arrest of nitrogen fixation is very rapidly observed in root directly exposed to water stress. This local and partial water limitation of the plant triggered a systemic signaling on well-watered roots of the same plant. The compensatory response occurs by increasing nodule biomass (and therefore nitrogen fixation capacities) of these roots not directly exposed to the water limitation. In preliminary experiments pea plants were inoculated with a reference Rlv strain. Response of and root and nodule development that compensate the local water limitation was confirmed. Interestingly, the compensatory response varies according to the Rlv strain as another Rlv strain, displaying similar nitrogen fixation efficiency in watered conditions, was not able to promote equivalent compensatory response. This experimental system has been used as a screen to select Rlv strains able to promote rapid compensatory responses to water stress

Genetic diversity for partner choice in a core collection of pea accessions inoculated by a mix of five <em>Rhizobium</em> <em>leguminosarum</em> bv. <em>viciae</em> genotypes

National audienc

Genetic bases of variation in plant architecture and rhizobial partner choice along the pea domestication gradient

International audienceLegumes play an increasingly important role in sustainable agriculture due to their ability to form a beneficial symbiotic interaction with nitrogen-fixing Rhizobium bacteria. Legumes are also a valuable source of protein for both feed and food, but are not grown as extensively as expected in Europe due to their high yield variability. Many genomic approaches are being developed to improve stress tolerance traits. However, to date, little attention has been paid to improving the interaction between symbiotic partners. The establishment of the symbiotic interaction is a complex evolutionary process in which the interests of both partners are not always aligned. No evidence was found in pea for co-selection of competitiveness for nodulation and nitrogen (N) fixation efficiency (Bourion et al., 2018). Furthermore, several data indicated that N fixation and plant growth could be suboptimal in fields where pea is exposed to populations of heterogenous rhizobial strains with contrasting effects on nodule, root and shoot development (Laguerre et al., 2007). We performed Genome-Wide Association Studies to decipher the genetic determinants and relationships between the complex trait of pea choice between rhizobial strains in mixture and plant architecture. A large panel of 340 pea accessions including very diverse cultivars, wild accessions and landraces, all inoculated with the same mixture of 28 diverse rhizobial strains, was grown in two successive experiments, on a high throughput non-destructive phenotyping platform. The proportion of each strain in the nodules of each pea at harvest was determined by DNA metabarcoding, and 20 variables of nodulated root architecture or plant growth traits were estimated by image analysis or measured. The results highlighted differential variation and largely uncoupled genetic bases between rhizobial partner choice and architectural or growth traits, along the pea domestication gradient

Anthyllis vulneraria/Mesorhizobium metallidurans, an efficient symbiotic nitrogen fixing association able to grow in mine tailings highly contaminated by Zn, Pb and Cd

The excessive concentrations of toxic heavy metals in mine tailings and their very low N content make soil reclamation strategies by phytostabilization difficult. Our objective was to test if the symbiotic association between the legume Anthyllis vulneraria subsp. carpatica and the bacteria Mesorhizobium metallidurans originating from highly polluted mine tailings is able to increase N concentration in soils with contrasting Zn, Pb and Cd contents. Plants of A. vulneraria subsp. carpatica from a mine site and of a non-metallicolous subsp. praeopera from non-polluted soil were inoculated with a metallicolous or a non-metallicolous compatible Mesorhizobium spp. and grown on low and high heavy metal-contaminated soils. In contaminated soil, many nodules were observed when the metallicolous A. vulneraria was inoculated with its rhizobium species M. metallidurans, whereas the non-metallicolous A. vulneraria died after a few weeks regardless of the rhizobium inoculant. Eighty percent of the total nitrogen was derived from biological nitrogen fixation through the association between metallicolous A. vulneraria and the rhizobium grown on metal-enriched soil. The ability of the metallicolous A. vulneraria to develop a high nitrogen fixing potential opens new possibilities for promoting a low-maintenance plant cover and for stabilizing the vegetation in high heavy metal-contaminated soils

Genetic diversity of nodulated root structure in a very diverse pea collection

National audienceThe root system is responsible for nitrogen (N) acquisition, which in legumes, combines mineral acquisition and symbiotic fixation in nodules. Despite these two complementary pathways, N nutrition may be a limiting factor of legumes yield because nodules are very sensitive to their local environment and N fixing legume root system is poorly developed which may limit soil exploration [1]. Pea establishes in root nodules a symbiotic association with Rhizobium leguminosarum sv viciae bacteria (Rlv) [2]. This study assessed the potential of naturally occurring genetic variability of nodulated root structure and functioning traits to improve yield pea performance. Two successive glasshouse experiments were performed on a wide 336-pea panel consisted of wild, landraces and cultivars from diverse geographic origins [3]. Plants were inoculated by a mixture of strains representative of the Rlv diversity and grown in innovative RhizoTubes© on the 4PMI high throughput phenotyping platform allowing daily automatic imaging of shoots and nodulated root systems and their analysis [4]. Significant variations between pea accessions were observed for traits describing shoot and nodulated root system architecture. After genotyping of the pea panel by exome capture, genome wide association analyses were performed using 3.9 millions SNPs to identify the genetic determinants of these traits. They will be useful for breeding new pea cultivars with increased root system size, sustained nodule number, and improved N nutrition

Diversité du choix de partenaires symbiotiques parmi une collection de pois inoculée par un mélange de souches de<em> rhizobium</em>

National audienceLes légumineuses sont des cultures de grand intérêt pour répondre aux enjeux de sécurité alimentaire et de développement durable. Dans un contexte de forte croissance démographique, ce sont des sources de protéines pour les alimentations humaine et animale. Un autre atout des légumineuses est qu’elles peuvent s’affranchir d’un apport d’engrais azoté grâce à leur capacité à former des associations symbiotiques au sein de nodosités racinaires avec des bactéries du sol (les rhizobia) qui fixent l’azote de l’air. Cependant, la fixation symbiotique n’est pas toujours optimale ; elle est très sensible aux stress abiotiques et dépend directement de l’efficacité symbiotique du partenaire bactérien. Les populations naturelles de rhizobia sont quantitativement et qualitativement hétérogènes et peuvent aboutir à des symbioses inefficaces. Une approche pluridisciplinaire a été menée afin d’évaluer l’impact du génotype de pois et des rhizobia qui nodulent cette espèce sur l’établissement et l’efficacité de la symbiose fixatrice d’azote. Des essais ont été réalisés en serre sur une collection de 104 accessions de pois avec des historiques de sélection et des origines géographiques diverses. Ces 104 accessions ont été inoculées par un mélange de cinq souches de Rhizobium leguminosarum sv. viciae (Rlv) choisies pour leur diversité. Une forte variation du choix entre les partenaires symbiotiques a été observée. Ceci a permis de mettre en évidence des liens entre la diversité génétique des accessions de pois et l’établissement préférentiel de la symbiose avec certaines souches de Rlv. Une expérimentation complémentaire sur un sous-ensemble de 18 génotypes de pois, inoculés chacun séparément avec chacune des cinq souches de Rlv, a révélé que la plante ne s’associe pas toujours préférentiellement avec la souche de rhizobium qui permet la fixation symbiotique la plus efficace. Des effets d’interaction pois x rhizobium ont été mis en évidence. Cette expérimentation a montré la grande variabilité de l’interaction symbiotique entre pois et Rlv et son importance pour l’établissement d’un rendement optimal. Des expérimentations supplémentaires permettront de préciser les déterminants génétiques de la plante et de la bactérie qui pilotent les capacités d’association des partenaires symbiotiques et/ou leurs efficacités. Des stratégies d’inoculation des semences de pois par des bactéries rhizobiacées, qui sont actuellement très peu pratiquées en Europe, combinées à une meilleure prise en compte de l’interaction symbiotique dans les processus de création de variétés sont des leviers à travailler pour accroître les performances des légumineuses dans des systèmes de cultures à bas intrants

Co-inoculation of a Pea Core-Collection with Diverse Rhizobial Strains Shows Competitiveness for Nodulation and Efficiency of Nitrogen Fixation Are Distinct traits in the Interaction

Pea forms symbiotic nodules with Rhizobium leguminosarum sv. viciae (Rlv). In the field, pea roots can be exposed to multiple compatible Rlv strains. Little is known about the mechanisms underlying the competitiveness for nodulation of Rlv strains and the ability of pea to choose between diverse compatible Rlv strains. The variability of pea-Rlv partner choice was investigated by co-inoculation with a mixture of five diverse Rlv strains of a 104-pea collection representative of the variability encountered in the genus Pisum. The nitrogen fixation efficiency conferred by each strain was determined in additional mono-inoculation experiments on a subset of 18 pea lines displaying contrasted Rlv choice. Differences in Rlv choice were observed within the pea collection according to their genetic or geographical diversities. The competitiveness for nodulation of a given pea-Rlv association evaluated in the multi-inoculated experiment was poorly correlated with its nitrogen fixation efficiency determined in mono-inoculation. Both plant and bacterial genetic determinants contribute to pea-Rlv partner choice. No evidence was found for co-selection of competitiveness for nodulation and nitrogen fixation efficiency. Plant and inoculant for an improved symbiotic association in the field must be selected not only on nitrogen fixation efficiency but also for competitiveness for nodulation