Vancomycin-Variable Enterococci Can Give Rise to Constitutive Resistance during Antibiotic Therapy

Abstract

e Vancomycin-resistant enterococci (VRE) are notorious clinical pathogens restricting the use of glycopeptide antibiotics in the clinic setting. Routine surveillance to detect VRE isolated from patients relies on PCR bioassays and chromogenic agar-based test methods. In recent years, we and others have reported the emergence of enterococcal strains harboring a "silent" copy of vancomycin resistance genes that confer a vancomycin-susceptible phenotype (vancomycin-susceptible enterococci [VSE]) and thus escape detection using drug sensitivity screening tests. Alarmingly, these strains are able to convert to a resistance phenotype (VSE¡VRE) during antibiotic treatment, severely compromising the success of therapy. Such strains have been termed vancomycin-variable enterococci (VVE). We have investigated the molecular mechanisms leading to the restoration of resistance in VVE isolates through the whole-genome sequencing of resistant isolates, measurement of resistance gene expression, and quantification of the accumulation of drug-resistant peptidoglycan precursors. The results demonstrate that VVE strains can revert to a VRE phenotype through the constitutive expression of the vancomycin resistance cassette. This is accomplished through a variety of changes in the DNA region upstream of the resistance genes that includes both a deletion of a likely transcription inhibitory secondary structure and the introduction of a new unregulated promoter. The VSE¡VRE transition of VVE can occur in patients during the course of antibiotic therapy, resulting in treatment failure. These VVE strains therefore pose a new challenge to the current regimen of diagnostic tests used for VRE detection in the clinic setting. Fig. 1) (5). VanX is a dipeptidase that removes the D-Ala-D-Ala that continues to be generated by the constitutively expressed chromosomal cell wall biosynthesis machinery. VanH supplies D-Lac by the biochemical reduction of pyruvate, while VanA is an ATP-dependent ligase that synthesizes the ester D-Ala-D-Lac. This depsipeptide is then incorporated into intracellular cell wall precursor biosynthesis, which supplies the reagents required for extracellular peptidoglycan synthesis, which is necessary for cell division and growth.The expression of the vanHAX three-gene cassette is controlled by the two-component regulatory system VanRS, in which VanR is a response regulator and VanS is an integral membrane histidine kinase that recognizes the presence of vancomycin (by direct binding in the case of VanS B [3,6]). The autophosphorylation of VanS and transfer of the phosphate to a VanR aspartyl side chain generates the induction-competent version of VanR that binds to elements upstream of vanH, inducing the expression of vanHAX, which results in resistance (Fig. 1). The vanRS and vanHAX genes have been found on a number of mobile genetic elements, including plasmids and transposons, such as Tn1546. As a result, they have spread globally, and VRE remains an important drug-resistant pathogen in many clinical settings (1). Co...

Substrate Inhibition of VanA by d -Alanine Reduces Vancomycin Resistance in a VanX-Dependent Manner

“…This spontaneous integration event in vanS had been observed before in both Streptomyces and Enterococcus strains and results in constitutive upregulation of the vancomycin resistance cluster (37,66,67). The ddl suppressor mutant LAG2 had a level of vancomycin resistance similar to that of the parental strain, with an MIC of 128 g/ml (Table 1).…”

Molecular Genetic Profiling of Clinical and Foodborne Strains of Enterococci with High Level Resistance to Gentamicin and Vancomycin

“…In recent years, worldwide, colonization with vancomycin-resistant E. faecium was considered endemic in many hospitals. Antimicrobial use in patients seems to be a critical factor related to vancomycin-resistant enterococci infections [20][21][22].…”

Vancomycin-Variable Enterococcal Bacteremia

“…A patient colonized with this organism was recently described, and it was shown in that case that the isolate became resistant to vancomycin in vivo over time during vancomycin therapy (1). A recent study reported that exposure of VVE strains to vancomycin in vitro leads to the development of vancomycin resistance by the constitutive expression of vanHAXYZ gene clusters (10). The use of vancomycin to treat VVE infections could provide selective pressure that may lead to the development of resistance on therapy and resultant treatment failure.…”