Study of alternative pathways to the LiaFSR and YycFGHIJ systems that lead to the decrease of Daptomycin susceptibility in Enterococcus faecium

Abstract

Daptomycin is one of the last alternatives for treatment of infections caused by vancomycin-resistant enterococci (VRE), especially in clonal complex 17 E. faecium, so-called of "high risk". However, resistance and therapeutic failure have already been observed and genes with mutations, such as liaFSR and yycFGHIJ have been suggested as responsible. However, the mechanism of resistance is not complete because sensitive isolates with mutations in such genes have already been found and, on the other hand, these genes are not always mutated in resistant isolates. In addition, the fact that the survival of E. faecalis in daptomycin after changing the composition of its membrane caused by the incorporation of fatty acids from the medium where they were incubated is a novelty that is not well explained and nothing is known about this in E. faecium. Recently, our group identified 3 daptomycin-nonsusceptible E. faecium isolates and, among several susceptible isolates, we identified a "supersusceptible" with MIC below the CIM90 described in the literature. It is intended to study the mechanisms involved in the resistant isolates based on what is already in the literature and to select resistant isolates in vitro from the "supersusceptible" to determine new pathways of resistance by sequencing the genomes and subsequent overexpression of mutated genes. In addition, the in vitro and susceptible parental isolates will be characterized in terms of membrane depolarization and fluidity , cell wall thickness, biofilm formation, time kill assay with daptomycin and ampicillin. (AU)