Fitness cost of vancomycin-resistant Enterococcus faecium plasmids associated with hospital infection outbreaks

Abstract

ABSTRACTBackgroundVancomycin resistance is mostly associated with Enterococcus faecium due to Tn1546-vanA located on narrow- and broad-host plasmids of various families. The study’s aim was to analyse the effects of acquiring Tn1546-plasmids with proven epidemicity in different bacterial host backgrounds.MethodsWidespread Tn1546-plasmids of different families RepA_N (n=5), Inc18 (n=4) and/or pHTβ (n=1), and prototype plasmids RepA_N (pRUM) and Inc18 (pRE25, pIP501) were analysed. Plasmid transferability and fitness cost were assessed using E. faecium (GE1, 64/3) and Enterococcus faecalis (JH2-2/FA202/UV202) recipient strains. Growth curves (Bioscreen C) and Relative Growth Rates were obtained in presence/absence of vancomycin. Plasmid stability was analysed (300 generations). Whole genome sequencing (Illumina-MiSeq) of non-evolved and evolved strains (GE1/64/3 transconjugants, n=49) was performed. SNP calling (breseq software) of non-evolved strains was used for comparison.ResultsAll plasmids were successfully transferred to different E. faecium clonal backgrounds. Most Tn1546-plasmids and Inc18 and RepA_N prototypes reduced host fitness (−2%-18%) while the cost of Tn1546 expression varied according to the Tn1546-variant and the recipient strain (9-49%). Stability of Tn1546-plasmids was documented in all cases, often with loss of phenotypic resistance and/or partial plasmid deletions. Point mutations and/or indels associated with essential bacterial functions were observed on the chromosome of evolved strains, some of them linked to increased fitness.ConclusionsThe stability of E. faecium Tn1546-plasmids in the absence of selective pressure and the high intra-species conjugation rates might explain the persistence of vancomycin resistance in E. faecium populations despite the significant burden they might impose on bacterial host strains.