Phenotypic and phylogenetic characterization of endosymbiotic bacteria from Lupinus mariae-josephi

Abstract

Lupinus mariae-josephi is a Lupinus species that thrives in a Southeastern area of Spain (Valencia) in soils of singularly high pH and active lime content. It is nodulated by extra-slow growing bacteria symbiotically and phylogenetically distant to endosymbiotic strains nodulating other Lupinus sp. native of the Iberian Peninsula and adapted to growth in acid soils. Cross-inoculation experiments revealed that the L. mariae-josephi endosymbiotic bacteria are unable to nodulate or efficiently fix nitrogen with well-known Lupinus spp. Their species affiliation was examined by a multilocus sequence analysis of four housekeeping genes (16S rDNA, glnII, recA, atpD) and the symbiotic nodC gene. Single and concatenated phylogenetic analyses of these genes consistently revealed that L. mariae-josephi endosymbiotic bacteria belong to a clade, within the Bradyrhizobium genus, highly differentiated from the Bradyrhizobium clade that includes currently named Bradyrhizobium species as well as the endosymbiotic bacteria from Lupinus species tested in this study. Within this new clade the L. mariae-josephi bacteria nested in several subgroup that may correspond to novel sister species. The phylogenetic analysis based on the nodC gene showed that L. mariae-josephi endosymbiotic bacteria define a novel branch of the nodC Bradyrhizobium tree and likely have a common unique ancestor for the symbiotic genes with nodule isolates from Retama spp.