Lytic Bacteriophages Facilitate Antibiotic Sensitization of Enterococcus faecium

Abstract

Enterococcus faecium, a commensal of the human intestine, has emerged as a hospital-adapted, multi-drug resistant (MDR) pathogen. Bacteriophages (phages), natural predators of bacteria, have regained attention as therapeutics to stem the rise of MDR bacteria. Despite their potential to curtail MDR E. faecium infections, the molecular events governing E. faecium-phage interactions remain largely unknown. Such interactions are important to delineate because phage selective pressure imposed on E. faecium will undoubtedly result in phage resistance phenotypes that could threaten the efficacy of phage therapy. In an effort to understand the emergence of phage resistance in E. faecium, three newly isolated lytic phages were used to demonstrate that E. faecium phage resistance is conferred through an array of cell wall-associated molecules, including secreted antigen A (SagA), enterococcal polysaccharide antigen (Epa), wall teichoic acids, capsule, and an arginine-aspartate-aspartate (RDD) protein of unknown function. We find that capsule and Epa are important for robust phage adsorption and that phage resistance mutations in sagA, epaR, and epaX enhance E. faecium susceptibility to ceftriaxone, an antibiotic normally ineffective due to its low affinity for enterococcal penicillin binding proteins. Consistent with these findings, we provide evidence that phages potently synergize with cell wall (ceftriaxone and ampicillin) and membrane-acting (daptomycin) antimicrobials to slow or completely inhibit the growth of E. faecium. Our work demonstrates that the evolution of phage resistance comes with fitness defects resulting in drug sensitization and that lytic phages could serve as effective antimicrobials for the treatment of E. faecium infections.

Genetically distant bacteriophages elicit unique genomic changes in Enterococcus faecalis

“…Enterococcal phage phylogeny was determined using OrthoMCL (39) as described previously (24). Enterococcal phage genomes were downloaded from the INPHARED phage genome database (18).…”

“…Both phages VPE25 and VFW bind to E. faecalis through cell surface polysaccharides, and infection proceeds following viral DNA entry which requires PIP EF (23). Studies in E. faecium have identified cell wall polysaccharides, secreted antigen A, and RNA polymerase to be involved in phage infection (12, 24). Other studies have identified the enterococcal polysaccharide antigen (Epa) as a co-receptor for E. faecalis phages (25, 26).…”

Sugarcoating it: Enterococcal polysaccharides as key modulators of host–pathogen interactions

“…The absence/variability of epa genes suggests that the EPA polysaccharides of the 2 species have different sugar compositions, and, in consequence, may confer diverse physiological functions. The epa loci were adapted from [ 10 , 20 , 32 , 54 , 104 , 105 , 107 ]. All genes are drawn to scale.…”

“…Changes in the composition of EPA caused by mutations in the epa loci have also been shown to decrease sensitivity to antibiotics that target cell envelope synthesis, such as ceftriaxone or carbapenems in agar plates with Etest strips [ 54 , 55 ]. This adaptation is thought to result in part from reduced rhamnose and/or increased mannose abundance in some of these strains’ cell wall polysaccharides [ 55 ].…”

“…faecium , although to a lesser extent than similar mutations in E . faecalis [ 54 ]. These adaptive traits act as gatekeepers to phage absorption, conferring additional means through which Enterococcus thrives.…”