Aggregation and Binding Substances Enhance Pathogenicity in Rabbit Models ofEnterococcus faecalisEndocarditis

Abstract

We investigated the importance of enterococcal aggregation substance (AS) and enterococcal binding substance (EBS) in rabbit models of Enterococcus faecalis cardiac infections. First, American Dutch belted rabbits were injected intraventricularly with 108 CFU and observed for 2 days. No clinical signs of illness developed in animals given AS− EBS−organisms, and all survived. All rabbits given AS−EBS+ organisms developed signs of illness, including significant pericardial inflammation, but only one of six died. All animals given AS+ EBS− organisms developed signs of illness, including pericardial inflammation, and survived. All rabbits given AS+ EBS+ organisms developed signs of illness and died. None of the rabbits receiving AS+ EBS+ organisms showed gross pericardial inflammation. The lethality and lack of inflammation are consistent with the presence of a superantigen. Rabbit and human lymphocytes were highly stimulated in vitro by cell extracts, but not cell-free culture fluids, of AS+ EBS+ organisms. In contrast, cell extracts from AS− EBS− organisms weakly stimulated lymphocyte proliferation. Culture fluids from human lymphocytes stimulated with AS+/EBS+enterococci contained high levels of gamma interferon and tumor necrosis factor alpha (TNF-α) and TNF-β, which is consistent with functional stimulation of T-lymphocyte proliferation and macrophage activation. Subsequent experiments examined the abilities of the same strains to cause endocarditis in a catheterization model. New Zealand White rabbits underwent transaortic catheterization for 2 h, at which time catheters were removed and animals were injected with 2 × 109 CFU of test organisms. None of the animals given AS− EBS− organisms developed vegetations or showed autopsy evidence of tissue damage. Rabbits given AS− EBS+ or AS+ EBS−organisms developed small vegetations and had splenomegaly at autopsy. All rabbits given AS+ EBS+ organisms developed large vegetations and had splenomegaly and lung congestion at autopsy. Similar experiments that left catheters in place for 3 days revealed that all rabbits given AS− EBS− or AS+ EBS+ organisms developed vegetations, but animals given AS+ EBS+ organisms had larger vegetations and autopsy evidence of lung congestion. These experiments provide direct evidence that these two cell wall components play an important role in the pathogenesis of endocarditis as well as in conjugative plasmid transfer.

Transcriptional analysis of virulence-related genes in enterococci from distinct origins

“…To assess if enterococcal virulence determinants are differentially expressed in environments related to colonization and infection, isolates from food products (milk and cheese), clinical strains from both human and veterinarian origin, as well as reference strains from international culture collections were selected. The eight virulence genes selected for this study were previously identified (Jett et al 1992;Kreft et al 1992;Schlievert et al 1998;Singh et al 1998;Shankar et al 2001Shankar et al , 2004 and included both cytolysin, an important cellular toxin, and proteins involved in the adhesion to host tissues, which is a crucial step in the infection process.…”

Enterococcus faecalis aggregation substance promotes opsonin-independent binding to human neutrophils via a complement receptor type 3-mediated mechanism

“…However, preliminary results indicate that INY1801 does act in a similar manner to E. faecalis containing wild-type pCF10 in vivo (Hirt, H., Schlievert, P. and Dunny, G.M., in preparation). In addition, the importance of AS as a virulence factor has been demonstrated in animal models of endocarditis [45,46]. AS-containing E. faecalis have been frequently found amongst clinical isolates [47] and have been less frequently seen in isolates from stools of healthy volunteers [48], again suggesting its importance as a virulence determinant of E. faecalis.…”

“…First, AS may facilitate the attachment of enterococci to renal [7] and intestinal [8] epithelial cells and colonization of these surfaces. Second, it may act as a superantigen and stimulate tumor necrosis factor production by macrophages and lymphocytes [45], which could subsequently increase the amount of CR3 on the surface of PMNs [49]. This could provide a route for greater AS-mediated bacterial binding to PMNs.…”

The Enterococcus faecalis cytolysin: a novel toxin active against eukaryotic and prokaryotic cells

“…Among known virulence traits encoded within the pathogenicity island, in addition to the cytolysin, is a gene for a surface adhesin, designated esp , found enriched among clinical isolates (Shankar et al ., 1999), and recently shown to play a role in urinary tract colonization (Shankar et al ., 2001). The surface protein, aggregation substance, previously noted for its contribution to endocarditis (Chow et al ., 1993;Schlievert et al ., 1998), is also encoded within the pathogenicity island. PAI genes that may contribute to the ability of clinical isolates to colonize the patient gastrointestinal tract include a bile acid hydrolase, new carbohydrate utilization pathways and other virulence, stress response and metabolic pathways (Shankar et al ., 2002).…”