sp. BNC1 1 3 Chr Chr Methylobacterium this website extorquens AM1 1 4 Chr Chr M. radiotolerans JCM2831 1 8 Chr Chr M. nodulans ORS2060 1 7 Chr pMNOD2 Bradyrhizobium sp. BTAi1 1 1 Chr Chr Nitrobacter hamburgensis X14 1 3 Chr Chr Xantobacter autotrophicus Ry2 1 1 Chr Chr Abbreviations are as follows: Chr, chromosome of those Rhizobiales with one chromosome; Chr I and Chr II, chromosome I and chromosome II respectively

in those Rhizobiales harboring two chromosomes; p, plasmid. *Rhizobium species in which localization of panCB genes was done by Southern blot hybridization of plasmid profiles. †Plasmids with very similar electrophoretic mobility gave as result ambiguous plasmid PRIMA-1MET localization of panC and panB homologous sequences. Phylogenetic analysis of rhizobial panCB genes indicates a common origin of chromosomal and plasmid-borne sequences Two possible hypotheses

were considered to explain the presence of panCB genes in plasmids of R. etli and R. leguminosarum strains: (1) an intragenomic rearrangement of panCB genes from chromosome to plasmid, which must have occurred in the last common ancestor of both species; (2) by xenologous gene displacement, that is, a horizontal transfer event in which a gene is displaced by a horizontally transferred ortholog acquired from another lineage [16]. In the latter hypothesis we assume that the presence of these xenolog genes in plasmids conferred a selective advantage that may have eventually led to the loss of the chromosome-EX 527 cost located panCB genes. To test these hypotheses the phylogeny of 16 rhizobial species inferred from ten orthologous single copy housekeeping genes (fusA, guaA, ileS, infB, recA, rplB, rpoB, rpoC, secY and valS) located on primary out chromosomes, was

compared with the phylogeny of the same rhizobial species inferred from the panCB genes located on plasmids and chromosomes. The rationale for this comparison was that if the plasmid-borne panCB phylogeny agrees with the current phylogeny of the Rhizobiales, inferred from the housekeeping genes, it would support the hypothesis of intragenomic transfer of the panCB genes. On the other hand, if both phylogenies are incongruent, it would favor the hypothesis of horizontal transfer of the panCB genes. Concatenated nucleic acids multiple alignments were used to infer both phylogenies with the maximum likelihood method described in materials and methods. The resulting phylogenetic trees are shown in Figure 2. The housekeeping genes inferred tree (Figure 2a) was consistent with the recently reported phylogeny of 19 Rhizobiales performed on a data set of 507 homologous proteins from the primary chromosome [17]. Both trees are in close agreement with the phylogeny inferred from the panCB genes (Figure 2b). Thus the phylogeny of R. etli and R.