Effects of boron and calcium nutrition on the establishment of the Rhizobium leguminosarum-pea (Pisum sativum) symbiosis and nodule development under salt stress

The effects of modifying boron (B) and calcium (Ca2+) concentrations on the establishment and development of rhizobial symbiosis in Pisum sativum plants grown under salt stress were investigated. Salinity almost completely inhibited the nodulation of pea plants by Rhizobium leguminosarum bv. viciae 3841. This effect was prevented by addition of Ca2+ during plant growth. The capacity of root exudates derived from salt-treated plants to induce Rhizobium nod genes was not significantly decreased. However, bacterial adsorption to roots was highly inhibited in plants grown with 75 mM NaCl. Moreover, R. leguminosarum 3841 did not grow in minimal media containing such salt concentration. High Ca2+ levels enhanced both rhizobial growth and adsorption to roots, and increased nodule number in the presence of high salt. Nevertheless, the nodules developed were not functional unless the B concentration was also increased. Because B has a strong effect on infection and cell invasion, these processes were investigated by fluorescence microscopy in pea nodules harbouring a R. leguminosarum strain that expresses green fluorescent protein. Salt-stressed plants had empty nodules and only those treated with high B and high Ca2+ developed infection threads and exhibited enhanced cell and tissue invasion by Rhizobium. Overall, the results indicate that Ca2+ promotes nodulation and B nodule development leading to an increase of salt tolerance of nodulated legumes.