Aggregation Substance Promotes Adherence, Phagocytosis, and Intracellular Survival ofEnterococcus faecaliswithin Human Macrophages and Suppresses Respiratory Burst

Abstract

The aggregation substance (AS) of Enterococcus faecalis, encoded on sex pheromone plasmids, is a surfacebound glycoprotein that mediates aggregation between bacteria thereby facilitating plasmid transfer. Sequencing of the pAD1-encoded Asa1 revealed that this surface protein contains two RGD motifs which are known to ligate integrins. Therefore, we investigated the influence of AS on the interaction of E. faecalis with human monocyte-derived macrophages which constitutively express ␤ 2 integrins (e.g., CD18). AS was found to cause a greater-than-fivefold increase in enterococcal adherence to macrophages and a greater-than-sevenfold increase in phagocytosis. Adherence was mediated by an interaction between the RGD motif and the integrin CD11b/CD18 (complement receptor type 3) as demonstrated by inhibition studies with monoclonal antibodies and RGD peptide. AS-bearing enterococci were significantly more resistant to macrophage killing during the first 3 h postinfection, probably due to inhibition of the respiratory burst as indicated by reduced concentrations of superoxide anion.Enterococci are gram-positive cocci which inhabit the gastrointestinal tract as well as the vagina and the oral cavity. Enterococcus faecalis accounts for 90% of human enterococcal infections, the most common being urinary tract infections, followed by abdominal infections, wound infections, bacteremia, and infective endocarditis (31, 39). Although infections due to E. faecalis have increased substantially during the last 10 years, the understanding of virulence mechanisms is still limited (24). One of the postulated virulence factors is the aggregation substance (AS), a sex pheromone plasmid-encoded surface protein which promotes the conjugative transfer of sex pheromone plasmids by formation of mating aggregates between donor and recipient cells (6,13,52). DNA sequencing of the structural gene for the pAD1-encoded AS revealed the presence of two Arg-Gly-Asp (RGD) sequences (16); RGD is a well-known motif recognized by a family of eukaryotic receptors, the integrins (38). Integrins consist of noncovalently linked ␣ and ␤ chains and are expressed on leukocytes, thrombocytes, endothelium, and various epithelial cells (21, 37, 42). Our group first suggested an interaction of AS with integrins, since we found that AS augmented adherence to porcine renal tubular cells which could be inhibited competitively by an RGD-Ser (RGDS) peptide (26). This hypothesis was corroborated by in vitro experiments with human polymorphonuclear leukocytes (PMN) which demonstrated that AS promotes opsonin-independent binding of E. faecalis via a ␤ 2 integrinmediated mechanism (46). It is assumed that many enterococcal infections are endogenous, originating from the intestinal tract (25, 51). Wells et al. speculated that macrophages may serve as a vehicle facilitating translocation from the intestinum into the lymph system and bloodstream (49, 50). However, this can occur only if enterococci are able to survive within macrophages. Indeed, Gentry-Weeks et al. demons...

Coxiella burnetii Avoids Macrophage Phagocytosis by Interfering with Spatial Distribution of Complement Receptor 3

“…The engagement of CR3 by Mycobacterium tuberculosis did not affect its intracellular survival (39) and CR3 is not involved in the outcome of M. tuberculosis infection (40). The aggregation substance of Enterococcus faecalis interacts with CR3, causes increased bacterial uptake, and prevents microbicidal activity of macrophages (41). In this report, whereas C. burnetii replicated slowly in control THP-1 monocytes, their treatment with RANTES prevented the replication phase but did not result in bacterial elimination.…”

Virulence Factor and Biofilm Formation in Clinical Enterococcal Isolates of the West of Iran

“…The asa1 is encoded by pheromoneresponsive plasmids, which often harbour antibiotic resistance genes (26,27). The protein causes clumping of E. faecalis cells and survival inside polymorphonuclear leucocytes, internalisation by intestinal cells, and increases in bacterial binding to cultured renal epithelial cells (28,29).…”

Cultivos primarios de células endoteliales aisladas de cordón umbilical humano: un modelo biológico para el estudio de los mecanismos de infección por enterococos.

“…Ya se ha demostrado que las cepas de Enterococcus faecalis resistentes a los antibióticos son capaces de trasladarse a través del epitelio intestinal de ratones (12,13) y los estudios sobre los factores bacterianos involucrados en las interacciones célula-patógeno han identificado sustancias que desempeñan un papel importante en el ingreso de la bacteria en enterocitos y macrófagos cultivados (14)(15)(16). Como se cree que muchas infecciones enterocócicas del torrente sanguíneo tienen origen entérico (17), los estudios in vitro se han enfocado a entender los mecanismos empleados por los enterococos para superar la barrera epitelial y sobrevivir en los macrófagos.…”