First nosocomial outbreak of vancomycin-resistant Enterococcus faecium expressing a VanD-like phenotype associated with a vanA genotype

Abstract

Although enterococci expressing acquired vancomycin resistance phenotype have been reported increasingly worldwide, they have been rarely reported in France. From August to December 2004 we faced an outbreak of vancomycin-resistant Enterococcus faecium (VRE) isolates in the nephrology department at Bicêtre Hospital (K.-Bicêtre, France). The expression of the glycopeptide resistance varied among the 26 VRE isolates, with vancomycin MICs ranging from 12 to >256 microg/ml, whereas teicoplanin MICs ranged from 4 to 48 microg/ml. However, several strains appeared to be susceptible to glycopeptides according to disk diffusion testing and expressed resistance only after subculture with glycopeptides. In addition, a heterogeneous expression of glycopeptide resistance was also observed. This so-called VanD-like phenotype of resistance (low-level resistance to vancomycin and mostly susceptibility to teicoplanin) was surprisingly associated with a vanA gene. Plasmid extraction and mating-out experiments indicated that the vanA gene was located on a 200-kb self-transferable plasmid. Pulsed-field gel electrophoresis identified mostly dissemination of a single clone, whereas diffusion of the VanA-positive plasmid in different genomic backgrounds had also occurred. The vanA gene was part of a vanA-type operon for expression of resistance located on a Tn1546-like transposon. Sequencing of this transposon identified insertion of insertion sequence IS16 in the vanY gene that encodes a d,d-carboxypeptidase that might explain in part the peculiar VanD-type phenotype of resistance. This report is the first description of a VRE outbreak in France and underlines the difficulty in detecting this organism due to variability on the expression of the glycopeptide resistance trait, if any.

FIG. 1.

Teicoplanin (TP) and vancomycin (VA) MICs determined by E-test for E. faecium isolate 11 (A), E. faecium isolate 13 (B), a colony of E. faecium isolate 13 grown in the inhibition zone next to the E-test strip (C), and E. faecium isolate 12 that had a homogeneous and high level of resistance to vancomycin and teicoplanin (D).

FIG. 2.

Analysis of SmaI-digested genomic DNA by PFGE. Lanes 1 to 29 represent the 26 VRE strains studied and three control strains (lanes 6, 7, and 26) as indicated in Table 1. Lane M, bacteriophage lambda concatemers were used as molecular size markers (Bio-Rad).

FIG. 3.

Schematic representation of Tn1546-like transposons. The orientation of the genes is indicated by an arrow. (A) Structure of the prototype Tn1546 transposon (37); (B) structure of the Tn1546-like transposon characterized from E. faecium isolate 11. Primers used for IS16 mapping are indicated by an arrow. TSD, target site duplication generated by IS16 upon insertion into vanY gene.