Identification of Factors Contributing to T-Cell Toxicity of Staphylococcus aureus Clinical Isolates

Abstract

We examined the ability of 206 clinical isolates of Staphylococcus aureus to lyse T cells and found differences between Agr groups. We found that the beta and delta hemolysins are involved and that methicillin-resistant S. aureus strains are less toxic than methicillin-susceptible S. aureus strains.Staphylococcus aureus is a major human pathogen, and its ability to secrete toxins significantly contributes to host damage (3). Studies linking disease characteristics with specific genes in S. aureus are often hampered by the difficulty with the genetic manipulation of clinical isolates (18), the fact that laboratory strains are no longer a faithful representation of those causing disease today (5), and the fact that S. aureus often has multiple genes whose activities can be substituted for by other genes (4). In this study, we adopted an alternative approach, in that we took a large S. aureus strain collection and assayed a specific characteristic of each isolate. Because we had a large amount of genetic information (e.g., clonality by multilocus sequence type [MLST] [2] and the presence of 33 adhesins and toxins [11,13]) relating to this strain collection, we sought to make associations between these factors and toxicity.T cells are an important component of vertebrate immunity, although their role in protection against S. aureus infections is unclear. Using an immortalized T-cell line (T2 cells [15]), we developed an assay in which washed bacterial cells (grown in brain heart infusion medium overnight at 37°C shaking) were incubated with T cells (grown in RPMI [Gibco] supplemented with 10% fetal bovine serum [Gibco], 1 mM L-glutamine, 200 units/ml penicillin, and 0.1 mg/ml streptomycin [Sigma] at 37°C with 10% CO 2 ) for 30 min. The number of T cells lysed by the de novo production of proteins by S. aureus was measured by using trypan blue (Sigma) in a Fast-Read counting chamber (Immune Systems, United Kingdom). Table 1 shows the factors found to be associated with T-cell toxicity. The data could not be normalized, and so nonparametric tests were performed to identify associations between T-cell toxicity and a specific genetic factor(s). The first factor that was found to be associated with T-cell toxicity was Agr (accessory gene regulator) function. The Agr proteins are responsible for the down-regulation of surface proteins and the up-regulation of exoproteins when they reach a quorum (1) and are measured by determination of the level of expression of the delta toxin (11). The mean T-cell survival achieved when the T cells were incubated with strains expressing delta toxin was 59.2%, whereas 72.4% survival was achieved when the T cells were incubated with strains not expressing the delta toxin (P Ͻ 0.001, Mann-Whitney U test). As the delta toxin is itself a pore-forming protein, it is possible that our observation could be explained, at least in part, by the loss of this protein's expression alone and not as a result of the loss of other Agrregulated proteins.We next examined whether the presence of 16 other ...

Offsetting virulence and antibiotic resistance costs by MRSA

“…This has been partially attributed to diminished growth rates associated with antibiotic resistance, but our findings suggest that lower levels of toxicity may also contribute to reduced virulence in healthy individuals (Collins et al, 2008). MRSA strains containing the smaller type IV SCCmec element are successful both in and outside health-care settings, suggesting that these MRSA strains may be more toxic than strains carrying the type II element.…”

“…Work by us and others have reported diverse virulence phenotypes among MRSA and MSSA (methicillin-sensitive Staphylococcus aureus) strains (Collins et al, 2008;Fowler et al, 2004;Vaudaux et al, 1998;O'Neill et al, 2008). Here we examine the ability of both clinical and isogenic MSSAs and MRSAs with the large type II (hospital-associated) and smaller type IV (hospital-and communityassociated) SCCmec elements to lyse cells.…”

Predicting the virulence of MRSA from its genome sequence

“…We next measured the gross lytic activity of these isolates using an immortalized T-cell line (Collins et al 2008;Rudkin et al 2012) (sensitive to beta toxin, gamma toxin, delta toxin, LukED and PSMalpha1, alpha2 and alpha3) and lipid vesicles ) (sensitive to delta toxin and PSMalpha1, alpha2 and alpha3). No differences in lytic abilities were observed across these two assays (Supplemental Fig.…”

“…2). The ability of the isolates to lyse T cells, which measured beta toxin, gamma toxin, delta toxin, PSMalpha1, alpha2, and alpha3 activity was performed as described previously (Collins et al 2008;Rudkin et al 2012). Lipid vesicles, which are susceptible to delta toxin, PSMalpha1, alpha2, and alpha3, were prepared as described previously .…”

Meticillin-resistantStaphylococcus aureusand meticillin-susceptibleS. aureuspneumonia: comparison of clinical and thin-section CT findings

“…In the present study, the frequency of bronchial wall thickening and centrilobular nodules were significantly higher in cases of S. aureus pneumonia than in patients with K. pneumoniae pneumonia (MRSA, p,0.001 and p,0.001; MSSA, p,0.001 and p,0.001, respectively) or acute C. pneumoniae pneumonia (MRSA, p,0.011 and p,0.001; MSSA, p,0.001 and p,0.001, respectively). Moreover, Collins et al [34] examined the toxicity of a genetically diverse population of clinical S. aureus and reported that the MSSA strains were significantly more toxic than MRSA strains. These findings suggest that the rapid exudation of numerous polymorphonuclear leukocytes showed the higher prevalence of bronchial wall thickening and centrilobular nodular with tree-in-bud pattern in the MSSA group than in the MRSA group.…”