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We investigated the ability of two overlapping fragments of staphylococcal enterotoxin B (SEB), which encompass the whole toxin, to induce protection and also examined if passive transfer of chicken anti-SEB antibodies raised against the holotoxin could protect rhesus monkeys against aerosolized SEB. Although both fragments of SEB were highly immunogenic, the fragments failed to protect mice whether they were injected separately or injected together. Passive transfer of antibody generated in chickens (immunoglobulin Y [IgY]) against the whole toxin suppressed cytokine responses and was protective in mice. All rhesus monkeys treated with the IgY specific for SEB up to 4 h after challenge survived lethal SEB aerosol exposure. These findings suggest that large fragments of SEB may not be ideal for productive vaccination, but passive transfer of SEB-specific antibodies protects nonhuman primates against lethal aerosol challenge. Thus, antibodies raised in chickens against the holotoxin may have potential therapeutic value within a therapeutic window of opportunity after SEB encounter.The staphylococcal enterotoxins (SEs) are a family of bacterial superantigens (BSAgs) produced by Staphylococcus aureus. These protein toxins bind to major histocompatibility complex (MHC) class II molecules and, with less affinity, to the T-cell antigen receptors without MHC molecules, resulting in intense stimulation of the immune system that triggers acute pathological effects (8,11,20). BSAgs are associated with several serious diseases, including food poisoning, bacterial arthritis, and lethal toxic shock syndrome (7, 12). In addition, viral infections may predispose patients to toxic shock syndrome caused by BSAg-associated secondary streptococcal or staphylococcal infection (4, 16). The main component of the intoxication process depends on the ability of BSAgs to activate a large number of T cells, causing a massive release of inflammatory cytokines (7,20).Because SEs can cause severe pathologies and are considered potential biowarfare agents, there is considerable need to develop vaccines and therapeutic approaches capable of eliminating their toxicity. Previously, we showed that genetically altered staphylococcal enterotoxin A (SEA) and SEB inactivated by a site-directed mutagenesis strategy and lacking superantigenic effects were highly immunogenic in mice and rhesus monkeys (2,20,21). These recombinant vaccines elicited neutralizing antibodies that were detected in in vitro surrogate assays and protected the vaccinees against wild-type (WT) SEA and SEB. The experiments reported here were initiated to find fragments of SEB that could be used for vaccine purposes and to examine the suitability of passive immunotherapy with anti-SEB antibody developed in chickens (immunoglobulin Y [IgY]) against lethal effects of SEB in mice and rhesus monkeys. The data presented here highlight a useful therapeutic maneuver that could be employed to reduce or eliminate BSAg-mediated toxic shock syndrome and possibly other associated disorders. M...

Bacterial superantigens (BSAgs) cause massive stimulation of the immune system and are associated with various pathologies and diseases. To address the role of antibodies in protection against BSAgs, we screened the sera of 29 human volunteers for antibodies to the SAgs staphylococcal enterotoxin A (SEA), SEB, SEC1, and toxic shock syndrome toxin 1 (TSST-1). Although all volunteers had detectable levels of antibodies against SEB and SEC1, many (9 out of 29 volunteers) lacked detectable antibody to SEA or had minimal titers. Antibody titers to TSST-1 were well below those to SEB and SEC1, and three volunteers lacked detectable antibody to this BSAg. In addition, pooled immunoglobulin preparations obtained from different companies had antibody titers against SEs and TSST-1. There was a good correlation between antibody titers and inhibition of superantigenic effects of these toxins. Transfer of SEB-specific antibodies, obtained from pooled sera, suppressed in vitro T-cell proliferation and totally protected mice against SEB. These data suggest that the inhibitory activity of human sera was specific to antibodies directed against the toxins. Thus, it may be possible to counteract with specific antibodies BSAg-associated pathologies caused by stimulation of the immune system.Bacterial superantigens (BSAgs), such as staphylococcal enterotoxins (SEs) and toxic shock syndrome toxin 1 (TSST-1), are pyrogenic virulence factors produced by Staphylococcus aureus (9,11,13,26). These microbial SAgs bind to both human major histocompatibility antigen class II molecules on the surface of antigen-presenting cells and germ line-encoded variable domain sequences of the specific T-cell receptor variable ␤ chain on T lymphocytes (9, 11). Thus, BSAgs bypass the normal antigen-specific restrictions by creating a wedge between T-cell receptor and class II molecules and hence activate significantly greater numbers of T lymphocytes. The majority of stimulated T cells are programmed to acquire susceptibility to cell death by Fas-and Fas ligand-mediated apoptosis, or alternatively they enter into a state of specific nonresponsiveness (anergy), which may last for several months after the initial encounter with the BSAg. The activation of antigenpresenting cells and T cells results in production of pathological levels of proinflammatory cytokines that contribute to several serious pathologies and lethal toxic shock syndrome (11,17,22,26).Low serum antibody titers to BSAgs have been associated with the recurrence of toxic shock syndrome (10,23,28). Vaccination with nonsuperantigenic forms of BSAgs mitigates many of the symptoms of SE exposure (4,14,27). Vaccinated animals had high protective antibody titers against SEs and were fully protected against lethal challenge (4, 27). Thus, antibody responses may play a major role in protection against BSAgs. Here, we studied the prevalence of anti-SE and anti-TSST-1 antibodies in normal human volunteers and several pooled intravenous immunoglobulin (IVIG) products and examined if there is a correla...

The exotoxins produced by Staphylococcus aureus, staphylococcal enterotoxins (SE) A-E and toxic shock syndrome toxin (TSST)-1, which are associated with serious diseases, including food poisoning and toxic shock syndrome, are termed superantigens (SAgs). To examine whether common antigenic epitopes were present and whether vaccination with 1 bacterial SAg could protect against challenge with a different SE or TSST-1, mice were vaccinated with SEA, SEB, SEC1, or TSST-1 individually or in combination. Mice injected with a single toxin developed high antibody titers against other SAgs. Marked improvement in survival was observed when immunized mice were challenged with a heterologous toxin. Mice vaccinated with a mixture of toxins were fully protected against 1 or multiple toxin challenges, indicating no interference effects of multivalent vaccinations. More importantly, higher titers were found against each SAg with the multivalent vaccination than with injection with a single SAg. Thus, immunizations with 1 SAg can induce cross-protective antibodies to heterologous SAgs, and multicomponent vaccination can enhance antibody responses against each bacterial SAg.

BackgroundThere are currently no licensed vaccines available for prevention of botulism in humans. The vaccination is not desirable due to expanding therapeutic indications of botulinum toxins. The only available specific treatment for botulism is antitoxin to remove circulating toxin, thus, preventing further neuronal damage. BAT® (Botulism Antitoxin Heptavalent (A, B, C, D, E, F, G)—(Equine)) has been developed and its therapeutic efficacy evaluated against botulinum neurotoxin serotype A (BoNT/A) in Rhesus macaques.Methods and findingsIn a post-exposure prophylaxis (PEP) study, animals were exposed to 4x LD50/kg of BoNT/A and administered intravenously with either BAT (1x or 0.1x scaled human dose), or placebo at 4 hours post-exposure. The animals were monitored for 14 days. For the therapeutic intervention studies, animals were exposed to a 1.7x LD50/kg of BoNT/A and treated intravenously with either placebo or BAT at a 1x scaled human dose at the onset of clinical signs. Animals were monitored on an hourly basis for 14 or 21 days. In the PEP study, all animals tolerated equine based antitoxin without any adverse clinical signs. A 100% survival was observed in groups treated with the BAT compared to 0% survival in those treated with the placebo (p<0.001, Fisher’s exact test). BAT antitoxin prevented the development of signs of neurotoxicity of botulinum toxin. In a therapeutic study, treatment with the BAT at scaled 1x human dose after the onset of clinical signs significantly enhanced survival compared to the placebo (46.6% vs. 0%, p<0.0001, Fisher’s exact test). Additionally, treatment with the BAT delayed the progression of signs (muscular weakness, respiratory distress, oral/nasal discharge) of toxin intoxication and reduced the severity of the disease.ConclusionsA single dose of BAT, when administered to symptomatic monkeys, resulted in a statistically significant survival benefit compared to the placebo. Additionally, BAT completely protected monkeys from the clinical signs of intoxication and subsequent death when administered as PEP treatment. These data in part supported the licensure of BAT under the Animal Rule in the United States by the Food and Drug Administration.