Oral Presentation BACPATH 2017

The genomic diversity of the type I restriction modification systems of Staphylococcus epidermidis and use of plasmid artificial modification to bypass the restriction barrier (#4)

Jean YH Lee 1 , Ian R Monk 1 , Sacha J Pidot 1 , Torsten Seemann 2 3 , Timothy P Stinear 1 2 , Benjamin P Howden 2 4 5
  1. Microbiology & Immunology, Doherty Institute for Infection & Immunity, University of Melbourne, Melbourne, Victoria, Australia
  2. Doherty Applied Microbial Genomics, Department of Microbiology & Immunology, Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, Victoria, Australia
  3. Victorian Life Sciences Computation Initiative, University of Melbourne, Melbourne, Victoria, Australia
  4. Microbiological Diagnostic Unit Public Health Laboratory, Department of Microbiology & Immunology, Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, Victoria, Australia
  5. Infectious Diseases Department, Austin Health, Melbourne, Victoria, Australia

S. epidermidis is a significant opportunistic pathogen of humans, with a single lineage, sequence type (ST) 2, accounting for most clinical disease. Molecular based studies in this species have been limited by the presence of restriction modification systems (RMS) that pose a barrier to the exchange of foreign DNA. Here we present the first systematic review of the genomic diversity of type I RMS for both S. epidermidis (n=247) and S. aureus (n=128). Using PacBio SMRT sequencing, we report the methylomes (and completed genomes) of six S. epidermidis isolates, expanding on the six already described. Functional assessment of genomic predictions were performed with transformation and phage transduction experiments on: wild-type and type I specificity gene deletion (ΔhsdS) mutants (n=5) sets; using plasmid isolated from Escherichia coli hosts (n=5) constructed to heterologously express the relevant S. epidermidis type I systems, compared to a non-specific DC10B host. Contrary to current assumptions, genomic analysis revealed marked differences in the subunit arrangement, chromosomal location and movement of type I systems within S. epidermidis compared to S. aureus. Unlike S. aureus, the type I RMS of S. epidermidis were not reliably preserved within ST lineages. Of particular interest was clinically significant ST2 isolate, BPH0736, determined to have naturally disrupted restriction subunit, rendering it highly amenable to both transformation and phage transduction. For isolates with intact systems, plasmid artificial modification (PAM) was demonstrated to be an efficient way to overcome the type I RM barrier, with transformation efficiencies equivalent to that of the corresponding ΔhsdS mutant. Our findings indicated that the genetic manipulation of S. epidermidis using PAM typically requires construction of an E. coli host tailored to the specific strain of interest. In this context, BPH0736, our newly characterised isolate with a naturally defective restriction barrier is an ideal candidate for future laboratory studies.