Endosymbiotic bacteria nodulating a new endemic lupine Lupinus mariae-josephi from alkaline soils in Eastern Spain represent a new lineage within the Bradyrhizobium genus

Abstract

Lupinus mariae-josephi isa recently described endemic Lupinus species from a small area in Eastern Spain where it thrives in soils with active lime and high pH. The L. mariae-josephi root symbionts were shown to be very slow-growing bacteria with different phenotypic and symbiotic characteristics from those of Bradyrhizobium strains nodulating other Lupinus. Their phylogenetic status was examined by multilocus sequence analyses of four housekeeping genes (16S rRNA, glnl I, recA, and atpD) and showed the existence of a distinct evolutionary lineage for L. mariae-josephi thatalso included Bradyrhizobiumjicamae. Within this lineage, the tested isolates clustered in three different sub-groups that might correspond to novel sister Bradyrhizobium species. These core gene analyses consistently showed that all the endosymbiotic bacteria isolated from other Lupinus species of the Iberian Peninsula were related to strains of the B. canariense or B. japonicum lineages and were separate from the L. mariae-josephi isolates. Phylogenetic analysis based on nodC symbiotic gene sequences showed that L. mariae-josephi bacteria also constituted a new symbiotic lineage distant from those previously defined in the genus Bradyrhizobium. In contrast, the nodC genes of isolates from other Lupinus spp. from the Iberian Peninsula were again clearly related to the B. canariense and B. japonicum bv. genistearum lineages. Speciation of L. mariae-josephi bradyrhizobia may result from the colonization of a singular habitat by their unique legume host.

Diverse Bacteria Affiliated with the Genera Microvirga, Phyllobacterium, and Bradyrhizobium Nodulate Lupinus micranthus Growing in Soils of Northern Tunisia

“…The analysis showed a remarkable diversity of symbiotic bacteria, including isolates belonging to three genera, Bradyrhizobium, Microvirga, and Phyllobacterium, from three different families within the order Rhizobiales, namely Bradyrhizobiaceae, Methylobacteriaceae, and Phyllobacteriaceae, respectively. In previous studies, a clear predominance of Bradyrhizobium lineages was described for most Lupinus species (6,7,9,12,42). In lupine species from the Old World, B. canariense and B. japonicum are the two dominant rhizobial species in root nodules (8,9).…”

“…Thus, Microvirga strains appear to have a preference for alkaline soils, similar to that of the remaining Bradyrhizobium groups (II, III, IV and V) nodulating L. micranthus. In this respect, it is worth noting that groups III and IV are similar to B. valentinum and B. retamae, respectively, and these have been associated with alkaline soils (12,34,49). Overall, the available evidence suggests that symbiotic bacteria with certain genotypes are adapted to particular ecosystems where soil pH might play an important role.…”

“…(i) Symbiovar genistearum was described by Vinuesa et al (45) and includes many strains nodulating a large number of plants from the tribe Genisteae, including the genera Lupinus, Cytisus, Retama, Chamaecytisus, and Spartium, growing in Africa, Europe, and America (12,33,39,40,45). The majority of L. micranthus-associated isolates related to B. lupini (group I), and those similar to B. cytisi/B.…”

“…PCR amplicons of genes coding for DNA recombination and repair protein RecA (recA), DNA gyrase subunit B (gyrB), 16S rRNA (rrs), glutamine synthetase 2 (glnII), chaperone protein DnaK (dnaK), and N-acetylglucosaminyltransferase (nodC) were obtained with primers and conditions previously described for recA, rrs, glnII, and nodC (12,16) and for gyrB and dnaK (21). Amplicons were sequenced according to the procedures described by Sánchez-Cañizares et al (12) and were used for phylogenetic analysis. Unincorporated primers and deoxynucleoside triphosphates (dNTPs) were removed from PCR products with the NucleoSpin Extract II (MachereyNagel) or, when needed, by gel electrophoresis followed by band purification with the same kit.…”

Conservation of Endangered Lupinus mariae-josephae in Its Natural Habitat by Inoculation with Selected, Native Bradyrhizobium Strains

“…Sanchez-Cañizares et al [17] characterized Lmj-nodulating bacteria from soils of a specific site in the Llombai (Valencia) area, and concluded that they belong to a new Bradyrhizobiun lineage unrelated to B. canariense or B. japonicum . Later on, a wider study on the genetic diversity of Lmj-nodulating bacteria characterized up to nineteen groups of Bradyrhizobium strains belonging to up to six genospecies that were able to nodulate Lmj plant populations in the Valencia province [18].…”