Diversity of IncP-9 plasmids of Pseudomonas

IncP-9 plasmids are important vehicles for degradation and resistance genes that contribute to the adaptability of Pseudomonas species in a variety of natural habitats. The three completely sequenced IncP-9 plasmids, pWW0, pDTG1 and NAH7, show extensive homology in replication, partitioning and transfer loci (an approximately 25 kb region) and to a lesser extent in the remaining backbone segments. We used PCR, DNA sequencing, hybridization and phylogenetic analyses to investigate the genetic diversity of 30 IncP-9 plasmids as well as the possibility of recombination between plasmids belonging to this family. Phylogenetic analysis of rep and oriV sequences revealed nine plasmid subgroups with 7-35 % divergence between them. Only one phenotypic character was normally associated with each subgroup, except for the IncP-9beta cluster, which included naphthalene- and toluene-degradation plasmids. The PCR and hybridization analysis using pWW0- and pDTG1-specific primers and probes targeting selected backbone loci showed that members of different IncP-9 subgroups have considerable similarity in their overall organization, supporting the existence of a conserved ancestral IncP-9 sequence. The results suggested that some IncP-9 plasmids are the product of recombination between plasmids of different IncP-9 subgroups but demonstrated clearly that insertion of degradative transposons has occurred on multiple occasions, indicating that association of this phenotype with these plasmids is not simply the result of divergent evolution from a single successful ancestral degradative plasmid.