ABSTRACT
Enterococcus faecalis are a major cause of nosocomial infection worldwide, and the spread of vancomycin resistant strains (VRE) limits treatment options. Tigecycline-resistant VRE began to be isolated from inpatients at a Brazilian hospital within months following the addition of tigecycline to the hospital formulary. This was found to be the result of a spread of an ST103 E. faecalis clone. Our objective was to identify the basis for tigecycline resistance in this lineage. The genomes of two closely related tigecycline-susceptible (MICâ¯=â¯0.06â¯mg/L), and three representative tigecycline-resistant (MICâ¯=â¯1â¯mg/L) ST103 isolates were sequenced and compared. Further, efforts were undertaken to recapitulate the emergence of resistant strains in vitro. The specific mutations identified in clinical isolates in several cases were within the same genes identified in laboratory-evolved strains. The contribution of various polymorphisms to the resistance phenotype was assessed by trans-complementation of the wild type or mutant alleles, by testing for differences in mRNA abundance, and/or by examining the phenotype of transposon insertion mutants. Among tigecycline-resistant clinical isolates, five genes contained non-synonymous mutations, including two genes known to be related to enterococcal tigecycline resistance (tetM and rpsJ). Finally, within the in vitro-selected resistant variants, mutation in the gene for a MarR-family response regulator was associated with tigecycline resistance. This study shows that E. faecalis mutates to attain tigecycline resistance through the complex interplay of multiple mechanisms, along multiple evolutionary trajectories.
Subject(s)
Anti-Bacterial Agents/pharmacology , Cross Infection/microbiology , Drug Resistance, Bacterial , Enterococcus faecalis/drug effects , Enterococcus faecalis/genetics , Gram-Positive Bacterial Infections/microbiology , Tigecycline/pharmacology , Cross Infection/drug therapy , Cross Infection/epidemiology , DNA Transposable Elements , Enterococcus faecalis/classification , Gene Expression Regulation, Bacterial , Genes, Bacterial , Genome, Bacterial , Genomics/methods , Gram-Positive Bacterial Infections/drug therapy , Gram-Positive Bacterial Infections/epidemiology , Humans , Microbial Sensitivity Tests , Mutagenesis, Insertional , PhylogenyABSTRACT
In 2009 during surveillance in a Brazilian hospital, many patients were confirmed to be colonized by vancomycin-resistant enterococci (VRE) and a few infection cases occurred. Among 14 isolates of Enterococcus faecalis, most had the same pulsotype, virulence profile (agg(+)elrA(+)gelE(+)), and were sequence type (ST)103, indicating dissemination of a clone. The 47 Enterococcus faecium were separated into four pulsotypes, the predominant virulence profile being esp(+)acm(+). All of them harbored the hospital marker IS16, and three randomly chosen isolates were ST412, belonging to the Clonal Complex 17. E. faecalis were all susceptible to penicillin and ampicillin, while all E. faecium were resistant to them. All isolates were susceptible to daptomycin and tigecycline. There were no rep-family genes common to all VRE. The VanA element of all E. faecium lost its left-side inverted repeat (IRL) region and had a specific IS insertion. On the other hand, all E. faecalis presented intact Tn1546. The size of plasmids containing the vanA gene as well as its rep-families varied between and within species. The lack of a vanA plasmid common to all VRE, together with the differences among VanA elements, despite the fact that some patients were colonized by both species during their hospitalization, leads us to suggest at least two different Tn1546 origins.
