Abstract
Background Enterococcus faecalis is one of the leading causes of nosocomial infections. Due to its innate and acquired resistance to most antibiotics, identification of new targets for antimicrobial treatment of E. faecalis is a high priority. The multiple peptide resistance factor MprF, which was first described in Staphylococcus aureus, modifies phosphatidylglycerol with lysin and reduces the negative charge of the membrane, thus increasing resistance to cationic antimicrobial peptides. We studied the effect of mprF in E. faecalis regarding influence on bacterial physiology and virulence.ResultsTwo putative mprF paralogs (mprF1 and mprF2) were identified in E. faecalis by BLAST search using the well-described S. aureus gene as a lead. Two deletion mutants in E. faecalis 12030 were created by homologous recombination. Analysis of both mutants by thin-layer chromatography showed that inactivation of mprF2 abolishes the synthesis of three distinct amino-phosphatidylglycerols (PGs). In contrast, deletion of mprF1 did not interfere with the biosynthesis of amino-PG. Inactivation of mprF2 increased susceptibility against several antimicrobial peptides and resulted in a 42% increased biofilm formation compared to wild-type mprF. However, resistance to opsonic killing was increased in the mutant, while virulence in a mouse bacteremia model was unchanged.ConclusionOur data suggest that only mprF2 is involved in the aminoacylation of PG in enterococci, and is probably responsible for synthesis of Lys-PG, Ala-PG, and Arg-PG, while mprF1 does not seem to have a role in aminoacylation. As in other Gram-positive pathogens, aminoacylation through MprF2 increases resistance against cationic antimicrobial peptides. Unlike mprF found in other bacteria, mprF2 does not seem to be a major virulence factor in enterococci.
Highlights
Enterococcus faecalis is part of the normal flora in the gastrointestinal tract of humans and animals
LysPG is synthesized by the integral membrane protein MprF, which transfers a lysyl group from lysyl-tRNA to PG and subsequently translocates Lys-PG from the inner to the outer leaflet of the cytoplasmic membrane [4]
MprF is considered a virulence factor, since it allows bacteria to evade neutrophil killing and enhances the virulence of S. aureus in mice [8]
Summary
Enterococcus faecalis is part of the normal flora in the gastrointestinal tract of humans and animals. The two major bacterial phospholipids are phosphatidylglycerol (PG) and diphosphatidylglycerol (DPG) Their head groups are negatively charged, thereby imparting anionic properties to the membrane surface. The reduced negative net charge of the cell membrane leads to the repulsion of cationic peptides, decreasing the sensitivity against these peptides [5]. This mechanism bestows S. aureus with resistance to cationic antibiotics such as daptomycin, vancomycin [6], and gentamicin [7]. The multiple peptide resistance factor MprF, which was first described in Staphylococcus aureus, modifies phosphatidylglycerol with lysin and reduces the negative charge of the membrane, increasing resistance to cationic antimicrobial peptides. We studied the effect of mprF in E. faecalis regarding influence on bacterial physiology and virulence
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