Sinorhizobium meliloti and Sinorhizobium medicae are two closely related species of the genus Sinorhizobium showing a similar host range, nodulating leguminous species of the genera Medicago, Melilotus and Trigonella, but their phylogenic relationship has not been elucidated yet. In this paper we report the application of three different molecular markers, (i) RFLP of nodD genes, (ii) 16S-23S rDNA intergenic gene spacer fingerprinting and (iii) amplification fragment length polymorphism to S. meliloti and S. medicae strains isolated from the Caucasian area, which is the region of origin of the host plant Medicago. The analysis of data could suggest the origin of S. medicae strains from an ancestral S. meliloti population.

Twenty-two rhizobia strains isolated from three distinct populations (North Ossetia, Dagestan, and Armenia) of a relict legume Vavilovia formosa were analysed to determine their position within Rhizobium leguminosarum biovar viciae (Rlv). These bacteria are described as symbionts of four plant genera Pisum, Vicia, Lathyrus, and Lens from the Fabeae tribe, of which Vavilovia is considered to be closest to its last common ancestor (LCA). In contrast to biovar viciae, bacteria from Rhizobium leguminosarum biovar trifolii (Rlt) inoculate plants from the Trifolieae tribe. Comparison of house-keeping (hkg: 16S rRNA, glnII, gltA, and dnaK) and symbiotic (sym: nodA, nodC, nodD, and nif H) genes of the symbionts of V. formosa with those of other Rlv and Rlt strains reveals a significant group separation, which was most pronounced for sym genes. A remarkable feature of the strains isolated from V. formosa was the presence of the nodX gene, which was commonly found in Rlv strains isolated from Afghanistan pea genotypes. Tube testing of different strains on nine plant species, including all genera from the Fabeae tribe, demonstrated that the strains from V. formosa nodulated the same cross inoculation group as the other Rlv strains. Comparison of nucleotide similarity in sym genes suggested that their diversification within sym-biotypes of Rlv was elicited by host plants. Contrariwise, that of hkg genes could be caused by either local adaptation to soil niches or by genetic drift. Long-term ecological isolation, genetic separation, and the ancestral position of V. formosa suggested that symbionts of V. formosa could be responsible for preserving ancestral genotypes of the Rlv biovar. may be traced using specialised symbiotic (sym) genes representing the accessory parts of bacterial genomes, which differ in their natural histories from housekeeping genes (hkg) representing the core parts of genomes [4]. As a result of co-evolutionary processes, symbiosis is formed between tightly co-adapted cross-inoculation groups of rhizobia and legumes, and their coevolution is directed by a set of symbiosis-specific genes from each partner [5][6][7]. In some rhizobia, sym genes are more susceptible to autonomous horizontal gene transfer than hkg genes, because they are located on plasmids-mobile elements of the genome [3]. This results in an intensive recombination of host specific and chromosomal markers [8]. For example, Rhizobium leguminosarum is composed of two biovars, which have diverged based on their plasmid-encoded host ranges [9]. Biovar viciae (Rlv) nodulates legumes from the Fabeae tribe, while biovar trifolii (Rlt) nodulates clovers from the Trifolieae tribe; however, they show a conservative chromosomal arrangement of hkg markers (Figure 1).Even so, divergent evolution of rhizobia is not restricted to sym genes. Application of the average nucleotide identity (ANI) method has demonstrated that a local R. leguminosarum population could be separated into five genomic species, differing in their hkg genes, representing their cor...

This paper explores the relationship between the genetic diversity of rhizobia and the morphological diversity of their plant hosts. Rhizobium galegae strains were isolated from nodules of wild Galega orientalis and Galega officinalis in the Caucasus, the center of origin for G. orientalis. All 101 isolates were characterized by genomic amplified fragment length polymorphism fingerprinting and by PCR-restriction fragment length polymorphism (RFLP) of the rRNA intergenic spacer and of five parts of the symbiotic region adjacent to nod box sequences. By all criteria, the R. galegae bv. officinalis and R. galegae bv. orientalis strains form distinct clusters. The nod box regions are highly conserved among strains belonging to each of the two biovars but differ structurally to various degrees between the biovars. The findings suggest varying evolutionary pressures in different parts of the symbiotic genome of closely related R. galegae biovars. Sixteen R. galegae bv. orientalis strains harbored copies of the same insertion sequence element; all were isolated from a particular site and belonged to a limited range of chromosomal genotypes. In all analyses, the Caucasian R. galegae bv. orientalis strains were more diverse than R. galegae bv. officinalis strains, in accordance with the gene center theory.Rhizobium galegae (14) is a species that forms an effective symbiosis with plants of Galega orientalis and Galega officinalis, the only species in the genus Galega (Fabaceae) that have been studied for symbiosis. This symbiotic system represents a sharply defined cross-inoculation group. However, there are some differences in symbiotic performance. R. galegae strains are able to infect both Galega species, but strains isolated from G. officinalis form effective nodules on that plant and ineffective nodules on G. orientalis, while the converse is true for strains from G. orientalis (16). This finding, along with a wide range of phenotypic and genotypic approaches using numerical taxonomy (15), phage typing, DNA homology (13, 40), lipopolysaccharide and protein patterns (16), plasmid profiling (29), phylogeny of ribosomal genes (25, 35), randomly amplified polymorphic DNA, and repetitive PCR (21, 28), led to the proposal of two biovars for strains forming an effective symbiosis with G. officinalis (R. galegae bv. officinalis) and G. orientalis (R. galegae bv. orientalis) (24). Symbiosis-related genetic traits were found to be the main factor in genetic divergence between the biovars (24). G. orientalis is a good fodder plant with some widely used cultivars, whereas G. officinalis is rather poisonous because of its high alkaloid content and has no agricultural importance.The interesting symbiotic properties, together with the extensive taxonomic knowledge about R. galegae, prompted us to use the species to answer important questions related to the diversity and evolution of rhizobia and their symbiosis with legumes. A recent study indicated that R.galegae bv. orientalis strains have lower genetic diversity than R. galegae ...

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Gram-stain-negative strains V5/3MT, V5/5K, V5/5M and V5/13 were isolated from root nodules of Vicia alpestris plants growing in the North Ossetia region (Caucasus). Sequencing of the partial 16S rRNA gene (rrs) and four housekeeping genes (dnaK, gyrB, recA and rpoB) showed that the isolates from V. alpestris were most closely related to the species Microvirga zambiensis (order Rhizobiales, family Methylobacteriaceae) which was described for the single isolate from root nodule of Listia angolensis growing in Zambia. Sequence similarities between the Microvirga-related isolates and M. zambiensis WSM3693T ranged from 98.5 to 98.7 % for rrs and from 79.7 to 95.8 % for housekeeping genes. Cellular fatty acids of the isolates V5/3MT, V5/5K, V5/5M and V5/13 included important amounts of C18 : 1ω7c (54.0-67.2 %), C16 : 0 (6.0-7.8 %), C19 : 0 cyclo ω8c (3.1-10.2 %), summed feature 2 (comprising one or more of iso-C16 : 1 I, C14 : 0 3-OH and unknown ECL 10.938, 5.8-22.5 %) and summed feature 3 (comprising C16 : 1ω7c and/or iso-C15 : 02-OH, 2.9-4.0 %). DNA-DNA hybridization between the isolate V5/3MT and M. zambiensis WSM3693T revealed DNA-DNA relatedness of 35.3 %. Analysis of morphological and physiological features of the novel isolates demonstrated their unique phenotypic profile in comparison with reference strains from closely related species of the genus Microvirga. On the basis of genotypic and phenotypic analysis, a novel species named Microvirga ossetica sp. nov. is proposed. The type strain is V5/3MT (=LMG 29787T=RCAM 02728T). Three additional strains of the species are V5/5K, V5/5M and V5/13.