Mobile genetic elements (MGEs) such as plasmids and insertion sequences (IS) are important drivers of bacterial evolution. We report here a new IS element from Pseudomonas stutzeri, and we show that a synergistic interaction occurs between this IS  and pUC-based plasmids, resulting in these plasmids becoming capable of replication in P.stutzeri strain Q (PstQ), a soil isolate.

The plasmid pUS23 is a pUC19 derivative that contains silent marker genes (gfp, aadB). The pU23 plasmid does not transform P.stutzeri effectively, but upon prolonged incubation, Ap-R and Gm-R transformants can be obtained. The plasmid content of five such transformants was studied. From 35 plasmids screened, 12 had 1.3 kb insertions, and these were of three different restriction types. One of each type (13A, 23P1, 10C) was sequenced.

All the sequenced plasmids contained an identical IS, which was designated ISPSt4 – this was 1312 bp in length, and part of the IS3 family. ISPst4 has 27 bp terminal inverted repeats (IRs), and created 3 bp direct repeats at the insertion site. Plasmid types 13A and 23P1 had ISPst4 inserted between bla and ori in the pUC plasmid backbone while type 10C had ISPst4 inserted upstream of the aadB gene.

Plasmid types 13A and 23P1 were capable of robust replication in P.stutzeri, while plasmid type 10C and the original pUS23 were not.  It was hypothesised that ISPst4 was providing a promoter to drive expression of the pUC origin of replication in P.stutzeri. Sequential deletions localised this promoter activity to a 74 bp NcoI—SacI restriction fragment.

We believe this is the first report of IS transposition directly leading to an expansion of plasmid host range. This work reveals new emergent properties arising from the interaction of two different MGEs, and adds a new dimension to our understanding of the relationship between plasmids and IS elements.