Genotypic Characterization of a Nosocomial Outbreak of VanAEnterococcus faecalis

Abstract

Despite growing concern about vancomycin-resistant enterococci (VRE) as nosocomial pathogens, especially in the United States, in Italy VRE still represent an uncommon and occasional experience for most diagnostic laboratories. We report a genotypic characterization of the first reported nosocomial outbreak of VRE in Italy. Some experiments, including plasmid analysis and pulsed-field gel electrophoresis (PFGE) assays, aimed at investigating the genetic relatedness of the VRE isolates. Other experiments, based on hybridization and polymerase chain reaction (PCR) assays, aimed at characterizing the vancomycin resistance determinants. Over a 6-month period, 21 VRE, all identified as Enterococcus faecalis, were isolated from eight patients (all treated earlier with glycopeptide antibiotics) in a neurosurgical intensive care unit. All isolates had the same biochemical profile and antibiotic susceptibility pattern, including high-level resistance to aminoglycosides and vancomycin and teicoplanin MICs of 256 and 128 micrograms/ml, respectively. Three plasmids, one strongly hybridizing with a vanA probe, were detected in all but the last of the 21 VRE isolates. The last isolate of the cluster lacked the smallest of the three plasmids. Similar restriction profiles were obtained after plasmid DNA digestion with several endonucleases, with minor differences appreciated only in the first and last isolates. Analysis of genomic DNA restriction fragment patterns by PFGE confirmed that the reported cluster of VRE isolations was due to a single nosocomial strain of E. faecalis, despite some modifications in plasmid DNA at the beginning and at the end of the outbreak. Completely different PFGE patterns were yielded by vancomycin-susceptible E. faecalis strains isolated during the same period from inpatients in the same intensive care unit. Hybridization experiments with vanA and vanS-vanH probes and DNA amplification assays using 14 PCR primer pairs specific for vanA cluster genes (vanR, vanS, vanH, vanA, and vanY), orf1, orf2, vanB, and vanC showed identical organization of resistance determinants in all epidemic VRE isolates. This organization appeared to be the same as that described for Tn1546 in VanA prototype strain E. faecium BM4147.

Antibiotic resistance and genotypic characterization by PFGE of clinical and environmental isolates of enterococci

“…Interestingly, PFGE type 1 was exclusive of biliary stent group, and accounted for six of the 13 isolates (Table 3); as all six isolates were recovered from patients hospitalized in the same ward, undergoing the same invasive procedure, this is suggestive of a ward‐associated clonal spread [19]. E. faecalis isolates recovered from UTI were mainly PFGE types 3 and 14 (Table 3).…”

Vancomycin-ResistantEnterococcus faeciumIsolates Causing Hospital Outbreaks in Northern Italy Belong to the Multilocus Sequence Typing C1 Lineage

“…16,35,36 Isolation of VREF in Italy has been reported since 1993. 3,4,13,19 Over the period 1993-1999, a single vanA clone and various related subtypes persisted in a hospital in northeastern Italy. 26 VRE were also found to be widely distributed among farm animals in the same geographical area, 5 as well as in meat and dairy produce.…”

VanA-Type Enterococci from Humans, Animals, and Food: Species Distribution, Population Structure, Tn 1546 Typing and Location, and Virulence Determinants

“…The VanA phenotype, expressing inducible, high-level vancomycin and teicoplanin resistance, is the most common in Europe (3,5,25,29,37). The vanA cluster-detected primarily in E. faecium and E. faecalis and less frequently in other enterococcal species-is carried by Tn1546 and is transferable by conjugation (2,9).…”