ace , Which Encodes an Adhesin in Enterococcus faecalis , Is Regulated by Ers and Is Involved in Virulence

Abstract

Enterococcus faecalis is an opportunistic pathogen that causes numerous infectious diseases in humans and is a major agent of nosocomial infections. In this work, we showed that the recently identified transcriptional regulator Ers (PrfA like), known to be involved in the cellular metabolism and the virulence of E. faecalis, acts as a repressor of ace, which encodes a collagen-binding protein. We characterized the promoter region of ace, and transcriptional analysis by reverse transcription-quantitative PCR and mobility shift protein-DNA binding assays revealed that Ers directly regulates the expression of ace. Transcription of ace appeared to be induced by the presence of bile salts, probably via the deregulation of ers. Moreover, with an ace deletion mutant and the complemented strain and by using an insect (Galleria mellonella) virulence model, as well as in vivo-in vitro murine macrophage models, we demonstrated for the first time that Ace can be considered a virulence factor for E. faecalis. Furthermore, animal experiments revealed that Ace is also involved in urinary tract infection by E. faecalis.Enterococcus faecalis is a natural member of the intestinal microflora of warm-blooded animals and humans. However, E. faecalis remains an important opportunistic pathogen and represents one of the principal causes of nosocomial infections in the United States and Europe (33; for a review, see reference 20). Especially for immunocompromised patients, these infections include endocarditis, meningitis, pneumonia, peritonitis, visceral abscesses, urinary infections, and septicemia (13). About a dozen putative virulence factors have been identified in E. faecalis, but mechanisms of virulence remain not fully understood. These factors are involved in different steps of the infection process, such as attachment to host cells or extracellular matrix (ECM), macrophage resistance, tissue damage, and immune system evasion (20).For extracellular pathogens such as E. faecalis or Staphylococcus aureus, components of ECM or serum (i.e., collagen, fibronectin, and fibrinogen) are preferred targets for adhesion. During important steps of the infectious process, they interact with bacterial surface-exposed molecules, which include microbial surface components recognizing adhesive matrix molecules (MSCRAMMs). Ace, an adhesin that binds collagen (types I and IV) and laminin and belongs to the MSCRAMM family, was identified in E. faecalis by sequence homology with the virulence factor Cna, a well-characterized MSCRAMM in S. aureus (16,26). Synthesis of the A domain of Ace by S. aureus increased its arthritogenic potential to a level similar to that of S. aureus expressing Cna (34). Study of the sequence diversity of ace among several strains of E. faecalis has revealed an important variation in the number of repeated sequences (17). However, the role played by these regions has not yet been elucidated. The expression of Ace is significantly induced by high temperature (culture at 46°C) and in vivo by the presence of serum or ECM com...

Cold-shock RNA-binding protein CspR is also exposed to the surface of Enterococcus faecalis

“…It is possible that CspR may be one of the 'moonlighting' protein group, meaning that it can have two unrelated functions according to its intra-or extracellular location. To date, for the prevention of E. faecalis infections, in addition to lipoteichoic acid and capsular polysaccharides, three proteins known as virulence factors have been proposed as possible immunotherapy targets: the general stress protein Gls24, the collagen adhesion Ace, and the endocarditis-and biofilm-associated pilus Ebp (Giard et al, 2000;Huebner et al, 2000;Lebreton et al, 2009;Murray, 1990;Singh & Murray, 2012;Singh et al, 2007;Teng et al, 2005). Our results showed that a cold-shock RNA-binding protein can present a double localization, both intercellularly and at the surface of the bacteria, and that anti-CspR antibody protects insect larvae against infection by E. faecalis.…”

“…mellonella infection experiments. Infection of G. mellonella larvae with E. faecalis was accomplished as previously described by Lebreton et al (2009). Briefly, using a syringe pump (KD Scientific), larvae (about 0.3 g and 3 cm long) were infected subcutaneously with washed E. faecalis WT, DcspR and corresponding complemented strains from an overnight culture in GM17, with 6610 6 ± 0.6610 6 c.f.u.…”

“…Therefore, the virulence of E. faecalis has been intensively studied over the past 20 years and~12 putative virulence genes have been reported, including several transcriptional and post-transcriptional regulators. The recently identified cold-shock RNA-binding protein CspR is part of this last category of virulence-associated factors (Fox et al, 2009;Hancock & Gilmore, 2002;Lebreton et al, 2009;Michaux et al, 2011 Michaux et al, , 2012Qin et al, 2001; Shankar et al, 2001). In some Gram-positive bacteria such as Staphylococcus aureus and Bacillus subtilis, cold-shock polypeptides have been shown to be involved in several aspects of bacterial life, i.e.…”

Genes de virulencia y bacteriocinas en cepas de Enterococcus faecalis aisladas desde diferentes muestras clínicas en la Región del Maule, Chile

“…Microbiología proteína de superfi cie extracelular (Esp), la cual promueve la adhesión, colonización y evasión del sistema inmune, también participa en la generación de biopelícula y aparentemente cumple un rol en la resistencia antimicrobiana; citolisina (Cyl), también denominada hemolisina, que posee propiedades β-hemolíticas en humanos y es bactericida contra bacterias grampositivas; gelatinasa (Gel) que provee de nutrientes a la bacteria degradando el tejido del hospedero y además presenta algún tipo de función en la formación de biopelícula; hialuronidasa que actúa despolimerizando la molécula de mucopolisacárido del tejido conectivo lo que facilita la diseminación de la bacteria; determinantes de feromonas (Eep) son capaces de modular de modo importante la respuesta infl amatoria in vivo; antígeno A (EfaA) que se asocia a la adherencia de la bacteria tanto a células bióticas y superfi cies abióticas, además de participar en ciertas etapas de la formación de biopelícula; proteína de superfi cie celular (Ace) que exhibe una fuerte similitud con la proteína del mismo nombre producida por Staphylococcus aureus, la cual posee la capacidad de unirse al colágeno y presentaría un rol en la patogénesis de la endocarditis 12 ; sustancia de agregación (Agg) que actúa mediando la unión específi ca de la bacteria al epitelio intestinal, células del epitelio renal, neutrófi los humanos y macrófagos además de aumentar la internalización y vida intracelular del microorganismo 13 . La presencia de un pilus de E. faecalis es importante en la producción de la biopelícula, la cual se sustenta en la proteína Bps.…”

The prolipoprotein diacylglyceryl transferase (Lgt) of Enterococcus faecalis contributes to virulence

“…Infection of Galleria mellonella larvae with E. faecalis was accomplished as previously described by Lebreton et al (2009). Briefly, using a syringe pump (KD Scientific), larvae (about 0.3 g and 3 cm in length) were infected subcutaneously with washed E. faecalis cells from an overnight culture in GM17, with 1.5610 6 ±0.15610 6 or 3610 6 ±0.3610 6 c.f.u.…”