Endocarditis and biofilm-associated pili of Enterococcus faecalis

Abstract

Increasing multidrug resistance in Enterococcus faecalis, a nosocomial opportunist and common cause of bacterial endocarditis, emphasizes the need for alternative therapeutic approaches such as immunotherapy or immunoprophylaxis. In an earlier study, we demonstrated the presence of antibodies in E. faecalis endocarditis patient sera to recombinant forms of 9 E. faecalis cell wall-anchored proteins; of these, we have now characterized an in vivo-expressed locus of 3 genes and an associated sortase gene (encoding sortase C; SrtC). Here, using mutation analyses and complementation, we demonstrated that both the ebp (encoding endocarditis and biofilm-associated pili) operon and srtC are important for biofilm production of E. faecalis strain OG1RF. In addition, immunogold electron microscopy using antisera against EbpA-EbpC proteins as well as patient serum demonstrated that E. faecalis produces pleomorphic surface pili. Assembly of pili and their cell wall attachment appeared to occur via a mechanism of cross-linking of the Ebp proteins by the designated SrtC. Importantly, a nonpiliated, allelic replacement mutant was significantly attenuated in an endocarditis model. These biologically important surface pili, which are antigenic in humans during endocarditis and encoded by a ubiquitous E. faecalis operon, may be a useful immunotarget for studies aimed at prevention and/or treatment of this pathogen.

Vancomycin-Resistant Enterococcus faecalis Endocarditis: Linezolid Failure and Strain Characterization of Virulence Factors

“…These included the gelatinase gene (22,26,28,30); recently described pilus-encoding genes (16); genes encoding putative MSCRAMMs (microbial surface components recognizing adhesive matrix molecules) with predicted immunoglobulin (Ig)-like folds (17,18,27; J. Sillanpää, S. R. Nallapareddy, and B. E. Murray, unpublished data); genes, including esp (33), in a predicted pathogenicity island (PAI) (15); and an acquired gene that contributes to biofilm formation (32) ( Table 1). The strain was examined for phenotypic production of gelatinase (22), hemolytic activity on Bacto Tryptic Soy Agar (Becton Dickinson and Company, Sparks, MD) plus 5% human blood agar plates, and biofilm formation (14).…”

“…Subsequently, a family of seven genes encoding MSCRAMM-like proteins was found in 100% (nine out of nine) of the E. faecalis endocarditis strains tested, and elevated titers of IgG to these MSCRAMMlike proteins were found in the sera of nine patients with E. faecalis infections (27). Three of these genes, ebpA, ebpB, and ebpC (endocarditis-and biofilm-associated pili), control sur- face pilus formation and may be important in endocarditis pathogenesis (16). Biofilm formation, which is modulated by many genes, including esp and the fsr locus, likely serves as an important factor in E. faecalis infections (16,32).…”

“…Three of these genes, ebpA, ebpB, and ebpC (endocarditis-and biofilm-associated pili), control sur- face pilus formation and may be important in endocarditis pathogenesis (16). Biofilm formation, which is modulated by many genes, including esp and the fsr locus, likely serves as an important factor in E. faecalis infections (16,32). In one study, E. faecalis endocarditis isolates produced biofilm more often than did E. faecalis isolates from nonendocarditis sources and from hospital fecal specimens (14).…”

Enterococcus faecalis Ebp pili are important for cell-cell aggregation and intraspecies gene transfer

“…The endocarditis-and biofilm-associated pili (Ebp) are surface-associated filamentous structures considered to play a pivotal role in Enterococcus faecalis virulence (Nallapareddy et al, 2006(Nallapareddy et al, , 2011bKemp et al, 2007;Singh et al, 2007;Nielsen et al, 2012;Sillanpää et al, 2013). Ebp pili are encoded by the ubiquitous ebpABC-bps cluster, which codes for the cell wall anchor pilin EbpB, the major shaft pilin EbpC and the fibre tip pilin EbpA.…”

“…Ebp pili are encoded by the ubiquitous ebpABC-bps cluster, which codes for the cell wall anchor pilin EbpB, the major shaft pilin EbpC and the fibre tip pilin EbpA. A pilusspecific class C sortase (Bps, for biofilm-and pilusassociated sortase), catalyses the assembly of the structural subunits into pili, before the housekeeping sortase A (SrtA) covalently binds the elongated pili to the cell wall via EbpB (Nallapareddy et al, 2006;Sillanpää et al, 2013). Studies on ebp regulation identified the upstream ebpR gene (Bourgogne et al, 2007) and the rnjB gene (Gao et al, 2010) as activators at the mRNA level, while the Fsr quorum-sensing system was described as a weak repressor (Bourgogne et al, 2006).…”