Plasmid heterogeneity and identification of a Tn5281-like element in clinical isolates of high-level gentamicin-resistant Enterococcus faecium isolated in the UK

Ten clinical isolates of high-level gentamicin-resistant (HLGR) Enterococcu s faecium, collected from six hospitals throughout the UK, were studied to determine whether HLGR was attributed to widespread dissemination of a sing le plasmid or whether different plasmid types were implicated in the dissem ination of this phenotype. HLGR was attributed to the presence of the AAC6' -APH2 " bifunctional aminoglycoside modifying enzyme. The aac6'-aph2 " gene was present on a 70 kb plasmid in all ten isolates. Conjugation studies in dicated that the HLGR marker could transfer with varying frequency, with or without the associated 70 kb plasmid. Detailed molecular genetic analysis suggested that four of the isolates harboured a transposon similar to Tn528 1, originally identified in Enterococcus faecalis strain HH22 isolated in t he USA. The UK transposon, however, lacked the two symmetrically located Ha eIII sites found in Tn5281. The six remaining isolates appeared to have a T n5281-truncated structure in which the aac6'-aph2 " gene is flanked by an I S256 element at the 5' end. Further stud ies with nine restriction endonucl eases showed that the aac6'-aph2 " gene was associated with two different p lasmid types in E. faecium. Pulsed-field gel electrophoresis (PFGE) analysi s identified three different patterns. The four E. faecium isolates harbour ing the Tn5281-like structure were indistinguishable from each other, while the remaining six isolates exhibited two distinct PFGE patterns. This is t he first study to indicate that there is heterogeneity among the plasmids t hat confer the HLGR phenotype in E. faecium isolates in the UK and to repor t on the presence of a transposon, similar to Tn5281, in E. faecium harbour ing the aac6'-aph2 " gene.