Variation in the plasmid backbone and dif module content of R3-T33 Acinetobacter plasmids

The predominant type of plasmids found in Acinetobacter species encode a Rep_3 initiation protein and many of these carry their accessory genes in dif modules. Here, available sequences of the 14 members of the group of Rep_3 plasmids typed as R3-T33, using a threshold of 95% identity in the repA gene, were compiled and compared. These plasmids were from various Acinetobacter species. The pdif sites were identified allowing the backbone and dif modules to be defined. As for other Rep_3 plasmids carrying dif modules, orfX encoding a protein of unknown function was found downstream of repA followed by a pdif site in the orientation XerC binding site-spacer-XerD binding site. Most backbones (n = 12) also included mobA and mobC genes but the two plasmids with the most diverged repA and orfX genes had different backbone contents. Although the gene content of the plasmid backbone was largely conserved, extensive recombinational exchange was detected and only two small groups carried identical or nearly identical backbones. Individual plasmids were associated with 1 to 13 dif modules. Many different dif modules were identified, including ones containing antibiotic or chromate resistance genes and several toxin/antitoxin gene pairs. In some cases, modules carrying the same genes were significantly diverged. Generally, the orientation of the pdif sites alternated such that C modules (XerC binding sites internal) alternated with D modules (XerD binding sites internal). However, fusions of two dif modules via mutational inactivation or loss of a pdif site were also detected.