Global Spread of the hyl Efm Colonization-Virulence Gene in Megaplasmids of the Enterococcus faecium CC17 Polyclonal Subcluster

Abstract

Enterococcus faecium has increasingly been reported as a nosocomial pathogen since the early 1990s, presumptively associated with the expansion of a human-associated Enterococcus faecium polyclonal subcluster known as clonal complex 17 (CC17) that has progressively acquired different antibiotic resistance (ampicillin and vancomycin) and virulence (esp(Efm), hyl(Efm), and fms) traits. We analyzed the presence and the location of a putative glycoside hydrolase hyl(Efm) gene among E. faecium strains obtained from hospitalized patients (255 patients; outbreak, bacteremic, and/or disseminated isolates from 23 countries and five continents; 1986 to 2009) and from nonclinical origins (isolates obtained from healthy humans [25 isolates], poultry [30], swine [90], and the environment [55]; 1999 to 2007). Clonal relatedness was established by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Plasmid analysis included determination of content and size (S1-PFGE), transferability (filter mating), screening of Rep initiator proteins (PCR), and location of vanA, vanB, ermB, and hyl(Efm) genes (S1/I-CeuI hybridization). Most E. faecium isolates contained large plasmids (>150 kb) and showed variable contents of van, hyl(Efm), or esp(Efm). The hyl(Efm) gene was associated with megaplasmids (170 to 375 kb) of worldwide spread (ST16, ST17, and ST18) or locally predominant (ST192, ST203, ST280, and ST412) ampicillin-resistant CC17 clones collected in the five continents since the early 1990s. All but one hyl(Efm)-positive isolate belonged to the CC17 polyclonal subcluster. The presence of hyl(Efm) megaplasmids among CC17 from Europe, Australia, Asia, and Africa since at least the mid-1990s was documented. This study further demonstrates the pandemic expansion of particular CC17 clones before acquisition of vancomycin resistance and putative virulence traits and describes the presence of megaplasmids in most of the contemporary E. faecium isolates with different origins.