Ralstonia solanacearum is a widely distributed phytopathogenic bacterium that is known to invade more than 200 host species, mainly in tropical areas. Reference strain GMI1000 is naturally transformable at in vitro and also in planta conditions and thus has the ability to acquire free exogenous DNA. We tested the ubiquity and variability of natural transformation in the four phylotypes of this species complex using 55 strains isolated from different hosts and geographical regions. Eighty per cent of strains distributed in all the phylotypes were naturally transformable by plasmids and/or genomic DNA. Transformability can be considered as a ubiquitous physiological trait in the R. solanacearum species complex. Transformation performed with two independent DNA donors showed that multiple integration events occurred simultaneously in two distant genomic regions. We also engineered a fourfold-resistant R. solanacearum GMI1000 mutant RS28 to evaluate the size of DNA exchanged during natural transformation. The results demonstrated that this bacterium was able to exchange large DNA fragments ranging from 30 to 90 kb by DNA replacement. The combination of these findings indicated that the natural transformation mechanism could be the main driving force of genetic diversification of the R. solanacearum species complex.

Horizontal gene transfer (HGT) is recognized as the major force for bacterial genome evolution. Yet, numerous questions remain about the transferred genes, their function, quantity and frequency. The extent to which genetic transformation by exogenous DNA has occurred over evolutionary time was initially addressed by an in silico approach using the complete genome sequence of the Ralstonia solanacearum GMI1000 strain. Methods based on phylogenetic reconstruction of prokaryote homologous genes families detected 151 genes (13.3%) of foreign origin in the R. solanacearum genome and tentatively identified their bacterial origin. These putative transfers were analyzed in comparison to experimental transformation tests involving 18 different genomic DNA positions in the genome as sites for homologous or homeologous recombination. Significant transformation frequency differences were observed among these positions tested regardless of the overall genomic divergence of the R. solanacearum strains tested as recipients. The genomic positions containing the putative exogenous DNA were not systematically transformed at the highest frequencies. The two genomic “hot spots”, which contain recA and mutS genes, exhibited transformation frequencies from 2 to more than 4 orders of magnitude higher than positions associated with other genes depending on the recipient strain. These results support the notion that the bacterial cell is equipped with active mechanisms to modulate acquisition of new DNA in different genomic positions. Bio-informatics study correlated recombination “hot-spots” to the presence of Chi-like signature sequences with which recombination might be preferentially initiated. The fundamental role of HGT is certainly not limited to the critical impact that the very rare foreign genes acquired mainly by chance can have on the bacterial adaptation potential. The frequency to which HGT with homologous and homeologous DNA happens in the environment might have led the bacteria to hijack DNA repair mechanisms in order to generate genetic diversity without losing too much genomic stability.