Novel maintenance determinant carried by staphylococcal multiresistance plasmids

  • Dr Slade Jensen, University of Sydney, Australia
  • Dr Rebecca LeBard, University of New South Wales, Australia
  • Ms Danielle Davies, University of Sydney, Australia
  • Mr William Figgett, University of Sydney, Australia
  • Ms Evelyn Lai, University of Sydney, Australia
  • Prof Ronald Skurray, University of Sydney, Australia
  • Neville Firth, University of Sydney, Australia

    • Clinical staphylococci commonly contain several plasmids, the largest usually carrying multiple resistance genes, often in the form of cointegrated copies of small resistance plasmids and/or associated with transposable elements. We are studying the plasmids pSK1 and pSK41 as prototypes of staphylococcal multiresistance and conjugative plasmids, respectively, to understand the mechanisms that contribute to their spread and persistence. Members of the pSK1 and pSK41 plasmid families variously confer resistance to beta-lactams, trimethoprim, aminoglycosides, bleomycin, mupirocin, antiseptics/disinfectants and notably, vancomycin. These plasmids utilise an evolutionarily common theta-mode replication system that is also found in plasmids from other Gram-positive genera, including Enterococcus, Streptococcus, Lactococcus, Bacillus and Lactobacillus.

    Both pSK1 and pSK41 possess two distinct plasmid maintenance determinants. pSK41 encodes a resolvase and a type II partitioning system which we have shown to extend plasmid segregational stability in S. aureus. pSK1 also encodes a resolvase, but does not possess a typical partitioning system. Instead, a single protein-encoding gene, now designated par, located immediately upstream of, and divergently transcribed from the plasmid’s replication initiation gene, rep, has been shown to enhance plasmid maintenance. Genes homologous to pSK1 par appear to be ubiquitous on staphylococcal multiresistance plasmids, with the pSK41-like conjugative plasmids being the exception.

    DNA binding studies have shown that pSK1 Par binds to a DNA site located immediately upstream of its coding sequence, and transcriptional reporter gene fusions have confirmed that the gene is transcriptionally autoregulated. Site-directed mutagenesis has been used to investigate the activities of functional domains of the Par protein, and the cellular localisation of the protein has been examined in S. aureus using fluorescence microscopy. The results provide an insight into a determinant that contributes to the persistence of staphylococcal multiresistance plasmids, even in the absence of selection for the resistance phenotypes that they confer.