Evidence for a Role for Coagulase-Negative Staphylococci and their Biofilms in the Historic and Future Evolution of Staphylococcus Aureus Including MRSA
Creators: name: Budri, Paulo Eduardo
Publisher: name: Royal College of Surgeons in Ireland
Year: 2019
Subjects: Uncategorized

description: Rates of methicillin-resistant <em>Staphylococcus aureus</em> (MRSA) infections in Ireland are decreasing since 2006. However, MRSA and methicillin-susceptible <em>S. aureus</em> (MSSA) remain a major cause of healthcare-associated infections in Europe. Recent evolutionary trends indicate that MRSA lineages associated with infections in the community are causing infections in the healthcare setting with increasing frequency. It is therefore important to investigate potential reservoirs in the community that may contribute to MRSA and <em>S. aureus</em> evolution. The methicillin resistance gene, <em>mecA</em>, is located within a transferable genetic element, staphylococcal Cassette Chromosome (SCC), referred to as SCC<em>mec</em>, which is also frequently found in coagulase-negative staphylococci (CoNS) including <em>Staphylococcus epidermidis.</em> <em>S. epidermidis</em> is the predominant human skin colonizer and clinically, its most important virulence trait is biofilm production. Indirect evidence suggests that MRSA clones arose from genetic transfer of SCC<em>mec</em> from CoNS to MSSA. Adjacent to SCC<em>mec,</em> other antibiotic resistance genes, most notably <em>fusC</em>, encoding fusidic acid resistance, have also been found associated with SCC. It is thought that CoNS may be a reservoir of these antibiotic-resistances and the close proximity of these staphylococcal species in the human nasopharynx provides opportunity for such gene acquisitions. In this thesis, we describe the investigation of community-associated colonizing staphylococcal species as a reservoir that may underpin the evolution of <em>S. aureus</em>. In Chapter 3, DNA microarray analysis of nasal colonising staphylococcal isolates revealed a greater frequency of the antimicrobial resistance genes <em>mecA, fusC, fusB, ileS2, qacA/qacC </em>and the arginine catabolite mobile element (ACME) among colonising CoNS compared to <em>S. aureus</em> among 137 <em>S. aureus</em>-carriers with no previous healthcare exposure, identified from 444 medical students in their pre-clinical years. Among <em>S. aureus</em> carriers, MRSA and methicillin-resistant CoNS (MRCoNS) rates were 6.5% (9/137) and 13.1 % (18/137), respectively. Simultaneous carriage of MRSA and MRCoNS was found in 1/137 students studied. The clonal lineages of colonizing <em>S. aureus</em> included several clonal complexes (CC)s associated with the acquisition of SCC<em>mec</em>. In Chapter 4 an <em>in-vitro</em> study is described in which the efficiency of three mechanisms of horizontal gene transfer (transformation, transduction and conjugation) were investigated when applied in a staphylococcal biofilm model, for the transfer of clinically relevant antibiotic resistance genes identified in Chapter 3. Within a staphylococcal biofilm of the <em>S. aureus</em> strain RN4220, transduction was the most efficient genetic transfer mechanism, followed by conjugation and transformation. Transfer of the SCC<em>mec-fus </em>element, identified among <em>S. aureus </em>carriers, was successful in this model when exposed to sub-inhibitory concentrations of fusidic acid. In Chapter 5, the further detailed analysis of a subset of isolates from <em>S. aureus</em>-carriers (MRSA, MSSA and methicillin-susceptible <em>S. epidermidis</em> (MRSE)) using a combination of DNA microarray and whole genome sequence (WGS) analyses, is described. Using this approach, a high homology of SCC elements and <em>fusC</em> among <em>S. aureus</em> and <em>S. epidermidis</em> isolates from healthy people was revealed including 100% sequence similarity among the SCC<em>fus</em> elements. Overall, investigation of the staphylococcal species among 30.8% of medical students positive for <em>S. aureus</em>, prior to their exposure to the clinical environment, indicates a low rate of MRSA but the presence of several lineages into which SCC<em>mec</em> is capable of inserting. Furthermore, the finding of the composite element SCC<em>fus </em>carried in association with SCC<em>mec </em>V in the CC1 background, is concerning as it suggests the recent expansion of this clone in the community possibly against the selective pressure of un-regulated fusidic acid use. The stable transfer potential of this element with its associated phenotypic antibiotic resistance traits, determined in an <em>in-vitro</em> biofilm model, particularly by transduction further supports this evolutionary trend. Data presented support the hypothesis that the close proximity and co-evolution of CoNS and <em>S. aureus</em> in the human host, even in the absence of confirmed antibiotic pressure, underpinned by horizontal gene transfer events within staphylococcal biofilms plays an important and often underestimated role in antimicrobial resistance evolution in <em>S. aureus</em>. Further epidemiological investigation and follow up of the co-evolution of these species in the transition from community to clinical setting may provide further insights to explain the recent but sustained success of community strains in the healthcare setting.