ABSTRACT
Staphylococcus aureus is a human and animal pathogen as well as a commensal bacterium. It can be a causative agent of severe, life-threatening infections with high mortality, e.g., toxic shock syndrome, septic shock, and multi-organ failure. S. aureus strains secrete a number of toxins. Exotoxins/enterotoxins are considered important in the pathogenesis of the above-mentioned conditions. Exotoxins, e.g., superantigen toxins, cause uncontrolled and polyclonal T cell activation and unregulated activation of inflammatory cytokines. Here we show the importance of genomic analysis of infectious strains in order to identify disease-causing exotoxins. Further, we show through functional analysis of superantigenic properties of staphylococcal exotoxins that even very small amounts of a putative superantigenic contaminant can have a significant mitogenic effect. The results show expression and production of two distinct staphylococcal exotoxins, SEC and SEL, in several strains from clinical isolates. Antibodies against both toxins are required to neutralise the superantigenic activity of staphylococcal supernatants and purified staphylococcal toxins.
Subject(s)
Shock, Septic , Staphylococcal Infections , Animals , Cytokines/metabolism , Enterotoxins/genetics , Enterotoxins/toxicity , Exotoxins/genetics , Humans , Staphylococcal Infections/microbiology , Staphylococcus aureus/metabolism , Superantigens/genetics , Superantigens/toxicityABSTRACT
Thrombocytopenia or platelet dysfunction is a risk factor for severe infection. Staphylococcus aureus (S. aureus) releases a variety of virulence factors especially toxic shock syndrome toxin 1 (TSST-1), which may cause toxic shock syndrome. S. aureus, when carrying the tst gene, is more prone to cause toxic shock syndrome and is responsible for an especially high rate of mortality. However, the effect of TSST-1 protein on platelets is unknown. Patients with the tst gene positive S. aureus bacteremia showed more serious infection, higher mortality and lower platelet count. The tst gene positive S. aureus strains induce more platelet apoptosis and activation and corresponding up-regulation of Bak and down-regulation of Bcl-XL in addition to the activation of Caspase-3. C57BL/6 mice infected with the tst gene positive strains resulted in both a decrease in platelet count and an increase in platelet apoptosis and/or activation events and mortality. Moreover, TSST-1 protein, encoded by tst gene, caused the decrease of platelet count, the increase of platelet apoptosis and activation events and the level of inflammatory cytokines in vivo. However, TSST-1 protein was unable to induce traditional activation and apoptosis on human platelets in vitro. These results suggested that TSST-1 protein may exert indirect effects on platelet activation and apoptosis in vivo.
Subject(s)
Shock, Septic , Staphylococcal Infections , Animals , Apoptosis , Bacterial Toxins , Enterotoxins , Humans , Mice , Mice, Inbred C57BL , Platelet Activation , Staphylococcal Infections/metabolism , Staphylococcus aureus , Superantigens/genetics , Superantigens/metabolism , Superantigens/toxicityABSTRACT
In the 1980s, menstrual toxic shock syndrome (mTSS) became a household topic, particularly among mothers and their daughters. The research performed at the time, and for the first time, exposed the American public as well as the biomedical community, in a major way, to understanding disease progression and investigation. Those studies led to the identification of the cause, Staphylococcus aureus and the pyrogenic toxin superantigen TSS toxin 1 (TSST-1), and many of the risk factors, for example, tampon use. Those studies in turn led to TSS warning labels on the outside and inside of tampon boxes and, as important, uniform standards worldwide of tampon absorbency labeling. This review addresses our understanding of the development and conclusions related to mTSS and risk factors. We leave the final message that even though mTSS is not commonly in the news today, cases continue to occur. Additionally, S. aureus strains cycle in human populations in roughly 10-year intervals, possibly dependent on immune status. TSST-1-producing S. aureus bacteria appear to be reemerging, suggesting that physician awareness of this emergence and mTSS history should be heightened.
Subject(s)
Bacterial Toxins/toxicity , Enterotoxins/toxicity , Menstrual Hygiene Products/adverse effects , Shock, Septic/epidemiology , Shock, Septic/microbiology , Staphylococcal Infections/microbiology , Superantigens/toxicity , Female , Humans , Menstruation , Risk Factors , Staphylococcus aureus/pathogenicityABSTRACT
Staphylococcal and streptococcal superantigens are virulence factors that cause toxic shock by hyperinducing inflammatory cytokines. Effective T-cell activation requires interaction between the principal costimulatory receptor CD28 and its two coligands, B7-1 (CD80) and B7-2 (CD86). To elicit an inflammatory cytokine storm, bacterial superantigens must bind directly into the homodimer interfaces of CD28 and B7-2. Recent evidence revealed that by engaging CD28 and B7-2 directly at their dimer interface, staphylococcal enterotoxin B (SEB) potently enhances intercellular synapse formation mediated by B7-2 and CD28, resulting in T-cell hyperactivation. Here, we addressed the question, whether diverse bacterial superantigens share the property of triggering B7-2/CD28 receptor engagement and if so, whether they are capable of enhancing also the interaction between B7-1 and CD28, which occurs with an order-of-magnitude higher affinity. To this end, we compared the ability of distinct staphylococcal and streptococcal superantigens to enhance intercellular B7-2/CD28 engagement. Each of these diverse superantigens promoted B7-2/CD28 engagement to a comparable extent. Moreover, they were capable of triggering the intercellular B7-1/CD28 interaction, analyzed by flow cytometry of co-cultured cell populations transfected separately to express human CD28 or B7-1. Streptococcal mitogenic exotoxin Z (SMEZ), the most potent superantigen known, was as sensitive as SEB, SEA and toxic shock syndrome toxin-1 (TSST-1) to inhibition of inflammatory cytokine induction by CD28 and B7-2 dimer interface mimetic peptides. Thus, superantigens act not only by mediating unconventional interaction between MHC-II molecule and T-cell receptor but especially, by strongly promoting engagement of CD28 by its B7-2 and B7-1 coligands, a critical immune checkpoint, forcing the principal costimulatory axis to signal excessively. Our results show that the diverse superantigens use a common mechanism to subvert the inflammatory response, strongly enhancing B7-1/CD28 and B7-2/CD28 costimulatory receptor engagement.
Subject(s)
B7-1 Antigen/immunology , B7-2 Antigen/immunology , Bacterial Toxins/toxicity , CD28 Antigens/immunology , Staphylococcus aureus/immunology , Streptococcus pyogenes/immunology , Superantigens/toxicity , T-Lymphocytes/immunology , Bacterial Toxins/immunology , HEK293 Cells , Humans , Inflammation/chemically induced , Inflammation/immunology , Inflammation/pathology , Lymphocyte Activation/drug effects , Superantigens/immunology , T-Lymphocytes/pathologyABSTRACT
Staphylococcal enterotoxin B (SEB) and related superantigenic toxins produced by Staphylococcus aureus are potent activators of the immune system. These protein toxins bind to major histocompatibility complex (MHC) class II molecules and specific Vß regions of T-cell receptors (TCRs), resulting in the activation of both monocytes/macrophages and T lymphocytes. The bridging of TCRs with MHC class II molecules by superantigens triggers an early "cytokine storm" and massive polyclonal T-cell proliferation. Proinflammatory cytokines, tumor necrosis factor α, interleukin 1 (IL-1), IL-2, interferon γ (IFNγ), and macrophage chemoattractant protein 1 elicit fever, inflammation, multiple organ injury, hypotension, and lethal shock. Upon MHC/TCR ligation, superantigens induce signaling pathways, including mitogen-activated protein kinase cascades and cytokine receptor signaling, which results in NFκB activation and the phosphoinositide 3-kinase/mammalian target of rapamycin pathways. In addition, gene profiling studies have revealed the essential roles of innate antimicrobial defense genes in the pathogenesis of SEB. The genes expressed in a murine model of SEB-induced shock include intracellular DNA/RNA sensors, apoptosis/DNA damage-related molecules, endoplasmic reticulum/mitochondrial stress responses, immunoproteasome components, and IFN-stimulated genes. This review focuses on the signaling pathways induced by superantigens that lead to the activation of inflammation and damage response genes. The induction of these damage response genes provides evidence that SEB induces danger signals in host cells, resulting in multiorgan injury and toxic shock. Therapeutics targeting both host inflammatory and cell death pathways can potentially mitigate the toxic effects of staphylococcal superantigens.
Subject(s)
Bacterial Toxins/toxicity , Pyrogens/toxicity , Shock, Septic/etiology , Staphylococcus , Superantigens/toxicity , Animals , Cell Death , Cytokines/immunology , Humans , Oxidative Stress , Receptors, Antigen, T-Cell/immunology , Shock, Septic/prevention & control , Signal TransductionABSTRACT
Streptococcal pyrogenic exotoxin (Spe) A expression is epidemiologically linked to streptococcal tonsillo-pharyngitis and outbreaks of scarlet fever, although the mechanisms by which superantigens confer advantage to Streptococcus pyogenes are unclear. S. pyogenes is an exclusively human pathogen. As the leucocyte profile of tonsil is unique, the impact of SpeA production on human tonsil cell function was investigated. Human tonsil cells from routine tonsillectomy were co-incubated with purified streptococcal superantigens or culture supernatants from isogenic streptococcal isolates, differing only in superantigen production. Tonsil cell proliferation was quantified by tritiated thymidine incorporation, and cell surface characteristics assessed by flow cytometry. Soluble mediators including immunoglobulin were measured using enzyme-linked immunosorbent assay. Tonsil T cells proliferated in response to SpeA and demonstrated typical release of proinflammatory cytokines. When cultured in the absence of superantigen, tonsil preparations released large quantities of immunoglobulin over 7 days. In contrast, marked B cell apoptosis and abrogation of total immunoglobulin (Ig)A, IgM, and IgG production occurred in the presence of SpeA and other superantigens. In SpeA-stimulated cultures, T follicular helper (Tfh) cells showed a reduction in C-X-C chemokine receptor (CXCR)5 (CD185) expression, but up-regulation of OX40 (CD134) and inducible T cell co-stimulator (ICOS) (CD278) expression. The phenotypical change in the Tfh population was associated with impaired chemotactic response to CXCL13. SpeA and other superantigens cause dysregulated tonsil immune function, driving T cells from Tfh to a proliferating phenotype, with resultant loss of B cells and immunoglobulin production, providing superantigen-producing bacteria with a probable survival advantage.
Subject(s)
Bacterial Proteins/immunology , Exotoxins/immunology , Membrane Proteins/immunology , Palatine Tonsil/immunology , Streptococcus pyogenes/immunology , Adaptive Immunity , Antigens, Bacterial/immunology , Antigens, Bacterial/toxicity , B-Lymphocytes/immunology , B-Lymphocytes/pathology , Bacterial Proteins/toxicity , Cell Death/immunology , Cell Proliferation , Cytokines/metabolism , Exotoxins/toxicity , Humans , Immunoglobulins/biosynthesis , In Vitro Techniques , Lymphocyte Activation , Membrane Proteins/toxicity , Palatine Tonsil/pathology , Phenotype , Streptococcal Infections/immunology , Streptococcal Infections/pathology , Streptococcus pyogenes/pathogenicity , Superantigens/immunology , Superantigens/toxicity , T-Lymphocytes, Helper-Inducer/immunology , T-Lymphocytes, Helper-Inducer/pathologyABSTRACT
Superantigens (SAgs) play a major role in the pathogenesis of Staphylococcus aureus and are associated with several diseases, including food poisoning, bacterial arthritis, and toxic shock syndrome. Monoclonal antibodies to these SAgs, primarily TSST-1, SEB and SEA have been shown to provide protection in animal studies and to reduce clinical severity in bacteremic patients. Here we quantify the pre-existing antibodies against SAgs in many human plasma and IVIG samples and demonstrate that in a major portion of the population these antibody titers are suboptimal and IVIG therapy only incrementally elevates the anti-SAg titers. Our in vitro neutralization studies show that a combination of antibodies against SEA, SEB,and TSST-1 can provide broad neutralization of staphylococcal SAgs. We report a single fusion protein (TBA225) consisting of the toxoid versions of TSST-1, SEB and SEA and demonstrate its immunogenicity and protective efficacy in a mouse model of toxic shock. Antibodies raised against this fusion vaccine provide broad neutralization of purified SAgs and culture supernatants of multiple clinically relevant S. aureus strains. Our data strongly supports the use of this fusion protein as a component of an anti-virulence based multivalent toxoid vaccine against S. aureus disease.
Subject(s)
Enterotoxins/toxicity , Recombinant Fusion Proteins/pharmacology , Staphylococcal Toxoid/pharmacology , Staphylococcus aureus , Superantigens/toxicity , Animals , Enterotoxins/chemistry , Enterotoxins/genetics , Enterotoxins/immunology , Female , Humans , Mice , Mice, Inbred BALB C , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/immunology , Staphylococcal Toxoid/chemistry , Staphylococcal Toxoid/genetics , Staphylococcal Toxoid/immunology , Staphylococcus aureus/chemistry , Staphylococcus aureus/genetics , Staphylococcus aureus/immunology , Superantigens/chemistry , Superantigens/genetics , Superantigens/immunologyABSTRACT
The relationship between infection and Kawasaki disease (KD) remains unclear. Infection has long been considered a key predisposing factor for KD. Bacterial and viral agents may be related to the onset of KD because of superantigen and cytokine production. Various bacterial and viral infections have been reported to be associated with KD, but the actual mechanism remains unknown. The higher association between KD and enterovirus has been well documented by using Taiwan National Health Insurance Research Database. However, no evidence has been obtained that various bacterial and viral infections induce KD. Comprehensive research, including infectious agents, should be conducted to elucidate the pathogenesis of KD.
Subject(s)
Bacterial Infections/complications , Mucocutaneous Lymph Node Syndrome/etiology , Virus Diseases/complications , Endogenous Retroviruses/pathogenicity , Humans , Superantigens/toxicityABSTRACT
Toxic shock syndrome toxin-1 (TSST-1), a superantigen produced by Staphylococcus aureus is a causative agent of toxic shock syndrome (TSS) that is frequently associated with tampon use. It has long been suggested that TSS is induced when TSST-1 circulates through the body. However, the systemic distribution of TSST-1 from vagina or uterus has never been demonstrated. In this study, a mouse cervicovaginal infection model was established. Transcervical inoculation with a virulence strain of S. aureus and its derivative TSST-1-deficient mutant demonstrated that TSST-1 distributed to the bloodstream and spleen, and promoted systemic inflammation without bacteremia. Transcervical administration with the wild-type toxin and a superantigen-deficient mutant of TSST-1 (mTSST-1) demonstrated that the superantigenic activity of TSST-1 was essential to stimulate the systemic inflammation. Furthermore, this activity was not promoted by co-transcervical inoculation with lipopolysaccharides. The circulating TSST-1 and systemic inflammation rapidly reduced at 48 h after administration, suggesting that persistence of S. aureus in the uterus may be involved in long-term complications of TSS. Transcervical inoculation with mTSST-1-producing S. aureus showed that this toxin promoted bacterial number, uterine tissue damage, and localization of bacterial cells around uterine cavity. The results suggest that TSST-1 enhances S. aureus burden in uterine cavity, the secreted TSST-1 distributes into circulation system, and then systemic inflammation is induced.
Subject(s)
Bacterial Toxins/toxicity , Endometritis/complications , Enterotoxins/toxicity , Shock, Septic/physiopathology , Staphylococcal Infections/pathology , Staphylococcus aureus/metabolism , Superantigens/toxicity , Animals , Bacterial Load , Disease Models, Animal , Endometritis/microbiology , Endometritis/pathology , Female , Mice, Inbred C57BL , Shock, Septic/pathology , Staphylococcus aureus/growth & development , Uterus/microbiologyABSTRACT
Staphylococcus aureus infective endocarditis (IE) is a fast-progressing and tissue-destructive infection of the cardiac endothelium. The superantigens (SAgs) toxic shock syndrome toxin 1 (TSST-1), staphylococcal enterotoxin C (SEC), and the toxins encoded by the enterotoxin gene cluster (egc) play a novel and essential role in the etiology of S. aureus IE. Recent studies indicate that SAgs act at the infection site to cause tissue pathology and promote vegetation growth. The underlying mechanism of SAg involvement has not been clearly defined. In SAg-mediated responses, immune cell priming is considered a primary triggering event leading to endothelial cell activation and altered function. Utilizing immortalized human aortic endothelial cells (iHAECs), we demonstrated that TSST-1 directly activates iHAECs, as documented by upregulation of vascular and intercellular adhesion molecules (VCAM-1 and ICAM-1). TSST-1-mediated activation results in increased monolayer permeability and defects in vascular reendothelialization. Yet stimulation of iHAECs with TSST-1 fails to induce interleukin-8 (IL-8) and IL-6 production. Furthermore, simultaneous stimulation of iHAECs with TSST-1 and lipopolysaccharide (LPS) inhibits LPS-mediated IL-8 and IL-6 secretion, even after pretreatment with either of the proinflammatory cytokines tumor necrosis factor alpha (TNF-α) and IL-1ß. IL-8 suppression is not mediated by TSST-1 binding to its canonical receptor major histocompatibility complex class II (MHC-II), supporting current evidence for a nonhematopoietic interacting site on SAgs. Together, the data suggest that TSST-1 differentially regulates cell-bound and secreted markers of endothelial cell activation that may result in dysregulated innate immune responses during S. aureus IE. Endothelial changes resulting from the action of SAgs can therefore directly contribute to the aggressive nature of S. aureus IE and development of life-threatening complications.
Subject(s)
Aorta/cytology , Bacterial Toxins/toxicity , Endothelial Cells/drug effects , Enterotoxins/toxicity , Superantigens/toxicity , Cell Movement/drug effects , Cell Proliferation/drug effects , Cells, Cultured , Gene Expression Regulation/drug effects , Humans , Interleukin-6/genetics , Interleukin-6/metabolism , Interleukin-8/genetics , Interleukin-8/metabolismABSTRACT
Superantigens (SAgs) are extremely potent bacterial toxins, which evoke a virulent immune response, inducing nonspecific T-cell proliferation, rapid cytokine release, and lethal toxic shock, for which there is no effective treatment. We previously developed a small molecule, S101, which potently inhibits proliferating T cells. In a severe mouse model of toxic shock, a single injection of S101 given together with superantigen challenge rescued 100% of the mice. Even when given 2 hours after challenge, S101 rescued 40% of the mice. S101 targets the T-cell receptor, inflammatory response, and actin cytoskeleton pathways. S101 inhibits the aryl hydrocarbon receptor, a ligand-activated transcription factor that is involved in the differentiation of T-helper cells, especially Th17, and regulatory T cells. Our results provide the rationale for developing S101 to treat superantigen-induced toxic shock and other pathologies characterized by T-cell activation and proliferation.
Subject(s)
Immunologic Factors/administration & dosage , Shock, Septic/prevention & control , Shock, Septic/therapy , Superantigens/toxicity , T-Lymphocytes/drug effects , Animals , Disease Models, Animal , Female , Injections, Intravenous , Mice , Mice, Inbred BALB C , Survival Analysis , Treatment OutcomeABSTRACT
Superantigens (SAgs) are potent exotoxins secreted by Staphylococcus aureus and Streptococcus pyogenes. They target a large fraction of T cell pools to set in motion a "cytokine storm" with severe and sometimes life-threatening consequences typically encountered in toxic shock syndrome (TSS). Given the rapidity with which TSS develops, designing timely and truly targeted therapies for this syndrome requires identification of key mediators of the cytokine storm's initial wave. Equally important, early host responses to SAgs can be accompanied or followed by a state of immunosuppression, which in turn jeopardizes the host's ability to combat and clear infections. Unlike in mouse models, the mechanisms underlying SAg-associated immunosuppression in humans are ill-defined. In this work, we have identified a population of innate-like T cells, called mucosa-associated invariant T (MAIT) cells, as the most powerful source of pro-inflammatory cytokines after exposure to SAgs. We have utilized primary human peripheral blood and hepatic mononuclear cells, mouse MAIT hybridoma lines, HLA-DR4-transgenic mice, MAIThighHLA-DR4+ bone marrow chimeras, and humanized NOD-scid IL-2Rγnull mice to demonstrate for the first time that: i) mouse and human MAIT cells are hyperresponsive to SAgs, typified by staphylococcal enterotoxin B (SEB); ii) the human MAIT cell response to SEB is rapid and far greater in magnitude than that launched by unfractionated conventional T, invariant natural killer T (iNKT) or γδ T cells, and is characterized by production of interferon (IFN)-γ, tumor necrosis factor (TNF)-α and interleukin (IL)-2, but not IL-17A; iii) high-affinity MHC class II interaction with SAgs, but not MHC-related protein 1 (MR1) participation, is required for MAIT cell activation; iv) MAIT cell responses to SEB can occur in a T cell receptor (TCR) Vß-specific manner but are largely contributed by IL-12 and IL-18; v) as MAIT cells are primed by SAgs, they also begin to develop a molecular signature consistent with exhaustion and failure to participate in antimicrobial defense. Accordingly, they upregulate lymphocyte-activation gene 3 (LAG-3), T cell immunoglobulin and mucin-3 (TIM-3), and/or programmed cell death-1 (PD-1), and acquire an anergic phenotype that interferes with their cognate function against Klebsiella pneumoniae and Escherichia coli; vi) MAIT cell hyperactivation and anergy co-utilize a signaling pathway that is governed by p38 and MEK1/2. Collectively, our findings demonstrate a pathogenic, rather than protective, role for MAIT cells during infection. Furthermore, we propose a novel mechanism of SAg-associated immunosuppression in humans. MAIT cells may therefore provide an attractive therapeutic target for the management of both early and late phases of severe SAg-mediated illnesses.
Subject(s)
Antigens, Bacterial/toxicity , Clonal Anergy , Models, Immunological , Mucosal-Associated Invariant T Cells/immunology , Staphylococcus aureus/immunology , Streptococcus pyogenes/immunology , Superantigens/toxicity , Animals , Antigens, Bacterial/metabolism , Bone Marrow Cells/cytology , Bone Marrow Cells/drug effects , Bone Marrow Cells/immunology , Bone Marrow Cells/metabolism , Cell Line , Cells, Cultured , Clonal Anergy/drug effects , Crosses, Genetic , Enterotoxins/metabolism , Enterotoxins/toxicity , Female , Humans , Hybridomas , Immunity, Innate , Leukocytes, Mononuclear/cytology , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/immunology , Leukocytes, Mononuclear/metabolism , Lymphocyte Activation/drug effects , Mice , Mice, Inbred NOD , Mice, Knockout , Mice, SCID , Mice, Transgenic , Mucosal-Associated Invariant T Cells/cytology , Mucosal-Associated Invariant T Cells/drug effects , Mucosal-Associated Invariant T Cells/metabolism , Specific Pathogen-Free Organisms , Staphylococcus aureus/metabolism , Streptococcus pyogenes/metabolism , Superantigens/metabolism , Transplantation Chimera/blood , Transplantation Chimera/immunology , Transplantation Chimera/metabolismABSTRACT
The present report describes three patients with toxic shock syndrome toxin (TSST)-1-associated exanthematous disease. In all patients, fever and systemic erythema without hemodynamic disturbance occurred following cellulitis of the lower limbs. At the site of infection, TSST-1 producing Methicillin-susceptible Staphylococcus aureus was detected. They defervesced and erythema resolved in response to administration of an antimicrobial drug, thereby avoiding severe illness. These patients did not meet the criteria for a clinical diagnosis of toxic shock syndrome. Measurement of T-cell receptor Vß2-positive T cells in the peripheral blood early after onset of symptoms was useful for diagnosis.
Subject(s)
Bacterial Toxins/toxicity , Enterotoxins/toxicity , Exanthema/etiology , Exanthema/microbiology , Methicillin-Resistant Staphylococcus aureus/pathogenicity , Shock, Septic/etiology , Shock, Septic/microbiology , Staphylococcal Infections/complications , Superantigens/toxicity , Anti-Infective Agents/therapeutic use , Child , Child, Preschool , Exanthema/drug therapy , Female , Humans , Male , Methicillin Resistance/drug effects , Methicillin-Resistant Staphylococcus aureus/drug effects , Shock, Septic/drug therapy , Staphylococcal Infections/drug therapyABSTRACT
The 1928 Bundaberg disaster is one of the greatest vaccine tragedies in history. Of 21 children immunized with a diphtheria toxin-antitoxin preparation contaminated with Staphylococcus aureus, 18 developed life-threatening disease and 12 died within 48â h. Historically, the deaths have been attributed to α-toxin, a secreted cytotoxin produced by most S. aureus strains, yet the ability of the Bundaberg contaminant microbe to produce the toxin has never been verified. For the first time, the ability of the original strain to produce α-toxin and other virulence factors is investigated. The study investigates the genetic and regulatory loci mediating α-toxin expression by PCR and assesses production of the cytotoxin in vitro using an erythrocyte haemolysis assay. This analysis is extended to other secreted virulence factors produced by the strain, and their sufficiency to cause lethality in New Zealand white rabbits is determined. Although the strain possesses a wild-type allele for α-toxin, it must have a defective regulatory system, which is responsible for the strain's minimal α-toxin production. The strain encodes and produces staphylococcal superantigens, including toxic shock syndrome toxin-1 (TSST-1), which is sufficient to cause lethality in patients. The findings cast doubt on the belief that α-toxin is the major virulence factor responsible for the Bundaberg fatalities and point to the superantigen TSST-1 as the cause of the disaster.
Subject(s)
Bacterial Toxins/toxicity , Enterotoxins/toxicity , Staphylococcal Infections/microbiology , Staphylococcus aureus/metabolism , Superantigens/toxicity , Animals , Australia , Humans , Rabbits , Staphylococcal Infections/mortality , Staphylococcus aureus/geneticsABSTRACT
Treatment of Staphylococcus aureus infections has become complicated owing to growing antibiotic resistance mechanisms and due to the multitude of virulence factors secreted by this organism. Failures with traditional monovalent vaccines or toxoids have brought a shift towards the use of multivalent formulas and neutralizing antibodies to combat and prevent range of staphylococcal infections. In this study, we evaluated the efficacy of a fusion protein (r-ET) comprising truncated regions of staphylococcal enterotoxin A (SEA) and toxic shock syndrome toxin (TSST-1) in generating neutralizing antibodies against superantigen induced toxicity in murine model. Serum antibodies showed specific reactivity to both SEA and TSST-1 native toxins. Hyperimmune serum from immunized animals protected cultured splenocytes from non-specific superantigen induced proliferation completely. Passive antibody administration prevented tissue damage from acute inflammation associated with superantigen challenge from S. aureus cell free culture supernatants. Approximately 80% and 50% of actively and passively immunized mice respectively were protected from lethal dose against S. aureus toxin challenge. This study revealed that r-ET protein is non-toxic and a strong immunogen which generated neutralizing antibodies and memory immune response against superantigen induced toxic effects in mice model.
Subject(s)
Bacterial Toxins/toxicity , Enterotoxins/toxicity , Recombinant Fusion Proteins/pharmacology , Staphylococcus aureus/immunology , Superantigens/toxicity , Toxoids/pharmacology , Animals , Antibodies, Bacterial/blood , Antibodies, Bacterial/immunology , Antibodies, Neutralizing/blood , Antigens, Bacterial/blood , Antigens, Bacterial/toxicity , Cell Proliferation/drug effects , Disease Models, Animal , Drug Resistance, Multiple, Bacterial , Female , Lethal Dose 50 , Mice , Mice, Inbred BALB C , Protein Conformation , Sequence AlignmentABSTRACT
Toxic shock syndrome (TSS) results from the host's overwhelming inflammatory response and cytokine storm mainly due to superantigens (SAgs). There is no effective specific therapy. Application of immunoglobulins has been shown to improve the outcome of the disease and to neutralize SAgs both in vivo and in vitro. However, in most experiments that have been performed, antiserum was either pre-incubated with SAg, or both were applied simultaneously. To mirror more closely the clinical situation, we applied a multiple dose (over five days) lethal challenge in a rabbit model. Treatment with toxic shock syndrome toxin 1 (TSST-1) neutralizing antibody was fully protective, even when administered late in the course of the challenge. Kinetic studies on the effect of superantigen toxins are scarce. We performed in vitro kinetic studies by neutralizing the toxin with antibodies at well-defined time points. T-cell activation was determined by assessing T-cell proliferation (3H-thymidine incorporation), determination of IL-2 release in the cell supernatant (ELISA), and IL-2 gene activation (real-time PCR (RT-PCR)). Here we show that T-cell activation occurs continuously. The application of TSST-1 neutralizing antiserum reduced IL-2 and TNFα release into the cell supernatant, even if added at later time points. Interference with the prolonged stimulation of proinflammatory cytokines is likely to be in vivo relevant, as postexposure treatment protected rabbits against the multiple dose lethal SAg challenge. Our results shed new light on the treatment of TSS by specific antibodies even at late stages of exposure.
Subject(s)
Antibodies, Neutralizing/therapeutic use , Antitoxins/therapeutic use , Bacterial Toxins/antagonists & inhibitors , Disease Models, Animal , Enterotoxins/antagonists & inhibitors , Shock, Septic/drug therapy , Animals , Antibodies, Neutralizing/pharmacology , Antitoxins/pharmacology , Bacterial Toxins/genetics , Bacterial Toxins/metabolism , Bacterial Toxins/toxicity , Cell Proliferation/drug effects , Cells, Cultured , Enterotoxins/genetics , Enterotoxins/metabolism , Enterotoxins/toxicity , Female , Gene Expression Regulation/drug effects , Humans , Interleukin-2/genetics , Interleukin-2/metabolism , Leukocytes, Mononuclear/cytology , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/immunology , Leukocytes, Mononuclear/metabolism , Lymphocyte Activation/drug effects , Male , Mutant Proteins/antagonists & inhibitors , Mutant Proteins/metabolism , Mutant Proteins/toxicity , Rabbits , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Recombinant Proteins/toxicity , Shock, Septic/etiology , Shock, Septic/immunology , Shock, Septic/metabolism , Superantigens/genetics , Superantigens/metabolism , Superantigens/toxicity , Survival Analysis , T-Lymphocytes/drug effects , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Toxicokinetics , Tumor Necrosis Factor-alpha/metabolismABSTRACT
Bacterial sepsis is a major cause of fatality worldwide. Sepsis is a multi-step process that involves an uncontrolled inflammatory response by the host cells that may result in multi organ failure and death. Both gram-negative and gram-positive bacteria play a major role in causing sepsis. These bacteria produce a range of virulence factors that enable them to escape the immune defenses and disseminate to remote organs, and toxins that interact with host cells via specific receptors on the cell surface and trigger a dysregulated immune response. Over the past decade, our understanding of toxins has markedly improved, allowing for new therapeutic strategies to be developed. This review summarizes some of these toxins and their role in sepsis.
Subject(s)
Bacterial Toxins/toxicity , Gram-Negative Bacteria/pathogenicity , Gram-Positive Bacteria/pathogenicity , Sepsis/microbiology , Systemic Inflammatory Response Syndrome/microbiology , ADP Ribose Transferases/toxicity , Antigens, Bacterial/toxicity , Exotoxins/toxicity , Gram-Negative Bacterial Infections/pathology , Gram-Positive Bacterial Infections/pathology , Humans , Lipopolysaccharides/toxicity , Superantigens/toxicity , Virulence Factors/toxicity , Pseudomonas aeruginosa Exotoxin AABSTRACT
Menstrual toxic shock syndrome (mTSS) is a rare, recognizable, and treatable disease that has been associated with tampon use epidemiologically. It involves a confluence of microbial risk factors (Staphylococcus aureus strains that produce the superantigen-TSST-1), as well as environmental characteristics of the vaginal ecosystem during menstruation and host susceptibility factors. This paper describes a series of experiments using the well-characterized model of porcine vaginal mucosa ex-vivo to assess the effect of these factors associated with tampon use on the permeability of the mucosa. The flux of radiolabeled TSST-1 and tritiated water ((3)H2O) through porcine vaginal mucosa was determined at various temperatures, after mechanical disruption of the epithelial surface by tape stripping, after treatment with surfactants or other compounds, and in the presence of microbial virulence factors. Elevated temperatures (42, 47 and 52°C) did not significantly increase flux of (3)H2O. Stripping of the epithelial layers significantly increased the flux of labeled toxin in a dose-dependent manner. Addition of benzalkonium chloride (0.1 and 0.5%) and glycerol (4%) significantly increased the flux of (3)H2O but sodium lauryl sulfate at any concentration tested did not. The flux of the labeled toxin was significantly increased in the presence of benzalkonium chloride but not Pluronic® L92 and Tween 20 and significantly increased with addition of α-hemolysin but not endotoxin. These results show that the permeability of porcine vagina ex-vivo to labeled toxin or water can be used to evaluate changes to the vaginal environment and modifications in tampon materials, and thus aid in risk assessment.
Subject(s)
Bacterial Toxins/toxicity , Enterotoxins/toxicity , Mucous Membrane/drug effects , Superantigens/toxicity , Vagina/drug effects , Animals , Cell Membrane Permeability/drug effects , Epithelial Cells/drug effects , Epithelial Cells/metabolism , Female , Hemolysin Proteins/toxicity , In Vitro Techniques , Lipopolysaccharides/isolation & purification , Lipopolysaccharides/metabolism , Mucous Membrane/pathology , Risk Factors , Salmonella typhimurium/metabolism , Shock, Septic/microbiology , Shock, Septic/pathology , Staphylococcus aureus , Surface-Active Agents/pharmacology , Swine , Temperature , Vagina/pathology , Virulence Factors/toxicityABSTRACT
Staphylococcal enterotoxin B (SEB) and related bacterial toxins cause diseases in humans and laboratory animals ranging from food poisoning, acute lung injury to toxic shock. These superantigens bind directly to the major histocompatibility complex class II molecules on antigen-presenting cells and specific Vß regions of T-cell receptors (TCR), resulting in rapid hyper-activation of the host immune system. In addition to TCR and co-stimulatory signals, proinflammatory mediators activate signaling pathways culminating in cell-stress response, activation of NFκB and mammalian target of rapamycin (mTOR). This article presents a concise review of superantigen-activated signaling pathways and focuses on the therapeutic challenges against bacterial superantigens.
Subject(s)
Enterotoxins/toxicity , Shock, Septic/therapy , Superantigens/toxicity , Animals , Antibodies/therapeutic use , Humans , Protective Agents/therapeutic use , Shock, Septic/immunology , Signal Transduction , Staphylococcal Vaccines , Staphylococcus/immunology , T-Lymphocytes/immunologyABSTRACT
Noninvasive vaginal infections by Staphylococcus aureus strains producing the superantigen TSST-1 can cause menstrual toxic shock syndrome (mTSS). With the objective of exploring the basis for differential susceptibility to mTSS, the relative responsiveness to TSST-1 of healthy women has been investigated. Peripheral blood mononuclear cells from healthy donors were incubated with purified TSST-1 or with the T-cell mitogen phytohemmaglutinin (PHA), and proliferation was measured. The concentrations of TSST-1 and PHA required to elicit a response equivalent to 15% of the maximal achievable response (EC15) were determined. Although with PHA, EC15 values were comparable between donors, subjects could be classified as being of high, medium, or low sensitivity based on responsiveness to TSST-1. Sensitivity to TSST-1-induced proliferation was associated with increased production of the cytokines interleukin-2 and interferon-γ. When the entire T lymphocyte population was considered, there were no differences between sensitivity groups with respect to the frequency of cells known to be responsive to TSST-1 (those bearing CD3(+) Vß2(+)). However, there was an association between sensitivity to TSST-1 and certain HLA-class II haplotypes. Thus, the frequencies of DR7DQ2, DR14DQ5, DR4DQ8, and DR8DQ4 haplotypes were greater among those with high sensitivity, a finding confirmed by analysis of responses to immortalized homozygous B cell lines. Collectively, the results reveal that factors other than neutralizing antibody and the frequency of Vß2(+) T lymphocytes determine immunological responsiveness to TSST-1. Differential responsiveness of lymphocytes to TSST-1 may form the basis of interindividual variations in susceptibility to mTSS.