Diversity of plasmids and Tn1546-type transposons among VanA Enterococcus faecium in Poland

E Wardal,W Hryniewicz,E Sadowy,I Gawryszewska,A Kuch,D Żabicka

doi:10.1007/s10096-016-2804-8

Abstract

The objective of this study was to investigate the antimicrobial resistance, Tn1546 transposon variability and plasmid diversity among Polish vancomycin-resistant Enterococcus faecium (VREfm) isolates of VanA phenotype in the context of their clonal structure. Two hundred sixteen clinical VREfm isolates collected between 1997 and 2010 were studied by antimicrobial susceptibility testing, MLST, MLVA and detection of IS16, espEfm, pilA, intA and plasmid-specific genes by PCR. Tn1546 structure was revealed by overlapping PCR and sequencing. Selected isolates were subjected to PFGE-S1 and Southern hybridization analyses. The vast majority of the isolates (95.8 %) belonged to lineages 17/18 (during the whole study period 1997–2010) and 78 (mostly in 2006–2010) of hospital-adapted meroclone of E. faecium. All isolates displayed a multi-drug resistance phenotype. Twenty-eight Tn1546 types (including 26 novel ones) were associated with eight different ISs (IS1216, IS1251, ISEfa4, ISEfa5, ISEfm2, ISEf1, IS3-like, ISEfm1-like). The vanA-determinant was typically located on plasmids, which most commonly carried rep2pRE25, rep17pRUM, rep18pEF418, rep1pIP501, ω-ε-ζ and axe-txe genes. VanA isolates from 1997–2005 to 2006–2010 differed in clonal composition, prevalence of gentamicin- and tetracycline-resistance and plasmidome. Our analysis revealed high complexity of Tn1546-type transposons and vanA-plasmids, and suggested that diverse genetic events, such as conjugation transfer, recombination, chromosomal integration and DNA mutations shaped the structure of these elements among Polish VREfm.

Highlights

In the past 20 years, vancomycin-resistant enterococci (VRE) have emerged as nosocomial pathogens worldwide
In the present study we aimed at the characterization of clonality of VanA-VRE outbreak due to Enterococcus faecium (VREfm) and genetic elements associated with this resistance determinant
All isolates belonging to hospital meroclone, as expected, were resistant to both ampicillin and ciprofloxacin, and enriched in putative virulence traits / markers such as IS16, espEfm, intAEfm and pili genes

Summary

Introduction

In the past 20 years, vancomycin-resistant enterococci (VRE) have emerged as nosocomial pathogens worldwide. In Poland, the first VRE outbreak due to Enterococcus faecium (VREfm) of VanA phenotype started in December 1996 in the Gdańsk Medical University [1]. The vast majority of VREfm observed worldwide belongs to a specific hospital meroclone, initially described as clonal complex (CC17), later divided into three distinct lineages 17, and 78 based on multilocus sequence typing (MLST) analyses [2, 3]. Strains belonging to the hospital meroclone are ciprofloxacin- and ampicillin-resistant, enriched in putative virulence traits, and show a distinct genetic repertoire, including cell surface protein genes (fms), regulatory genes, putative pathogenicity islands, plasmids, insertion sequences (IS) and integrated phages, which promote their adaptation [5,6,7]. The presence of IS16 and the E. faecium-specific esp gene (espEfm), carried on the integrative conjugative element ICEEfm, together with the intA integrase gene, are

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