RecombinantStaphylococcus aureusExfoliative Toxins Are Not Bacterial Superantigens

Abstract

Staphylococcal scalded-skin syndrome is an exfoliative dermatitis characterized by the separation of the epidermis at the stratum granulosum. This disruption is mediated by one of two Staphylococcus aureus exotoxins, exfoliative toxins A and B (ETA and ETB). Both ETA and ETB have been reported to be bacterial superantigens. A controversy exists, however, as other data indicate that these exotoxins are not superantigens. Here we demonstrate that recombinant exfoliative toxins produced in Escherichia coli do not act as T-cell mitogens and thus are not bacterial superantigens. These data fit the clinical profile of the disease, which is not associated with the classic symptoms of a superantigen-mediated syndrome.Staphylococcal scalded-skin syndrome (SSSS) is an exfoliative dermatitis of infants and children that results from infection with exfoliative-toxin-producing Staphylococcus aureus (1,15). SSSS is characterized by the formation of large bullae without inflammatory cell infiltrate and separation of extended areas of the epidermis at the stratum granulosum leaving the keratinocytes intact. Two biologically and serologically distinct S. aureus exotoxins are responsible for the skin manifestations of SSSS in humans, exfoliative toxin A (ETA) and exfoliative toxin B (ETB) (25). ETA (26.9 kDa) is encoded on the bacterial chromosome and shares 40% amino acid identity with the plasmid-borne ETB (27.3 kDa) (2, 13, 19). Despite extensive studies, the exact mechanism responsible for the skin disruption is not known.X-ray crystallographic structures of ETA and ETB (5,20,23,24) suggest that the toxins are members of the trypsin-like serine protease family. Protease activity has not been demonstrated for either toxin in vitro, but both ETA and ETB have intrinsic esterase activity, which is associated with serine proteases (3). Thus, it is likely that both toxins are proteases. In addition to having possible protease activity, both ETA and ETB are reported to be bacterial superantigens (14,16,17,24). Bacterial superantigens are a family of proteins able to bind simultaneously to the major histocompatibility complex and to the T-cell receptor (TCR), resulting in stimulation of a large number of T cells expressing specific V␤ subsets of the TCR repertoire (14).Previously, Fleischer and Bailey reported that recombinant ETA expressed in a superantigen-free S. aureus background does not have mitogenic activity (7). They conclude that the activity seen by others is due to contamination of the toxin preparations with other superantigens. However, since this report, additional literature has addressed the superantigenic activity of ETA and ETB. Here we demonstrate that recombinant exfoliative toxins produced in an Escherichia coli background do not act as T-cell mitogens and thus are not bacterial superantigens.Isolation, expression, and purification of recombinant exfoliative toxins. DNA fragments encoding ETA and ETB were obtained by utilizing PCR and DNA from exfoliative-toxinproducing bacterial strains. Toxins were expressed a...

Staphylococcal scalded skin syndrome related to an exfoliative toxin A- and B-producing strain in preterm infants

“…The presence of ETA was confirmed by Western blot analysis using anti-ETA polyclonal antiserum. Toxin activity was confirmed by injection of 10 ll of the partially purified culture supernantant into the subcutaneous tissue at the nape of the neck of newborn Balb/c mice as described [14,15]. The mice were placed back with lactating mothers and were observed at 4 h post-injection for signs of exfoliation.…”

Bullous Impetigo in Children Infected with Methicillin-Resistant Staphylococcus aureus Alone or in Combination with Methicillin-Susceptible S. aureus: Analysis of Genetic Characteristics, Including Assessment of Exfoliative Toxin Gene Carriage

“…The causative agent has traditionally been considered to be methicillin-susceptible S. aureus (MSSA) producing exfoliative toxin (ET) (9). ET from clinical isolates is serologically divided into three types, ETA, ETB, and ETD, which are serine proteases that specifically bind and cleave desmoglein 1, a cadherin-type intercellular adhesion molecule in desmosomes (1,9,10,13). The ETA gene (eta), ETB gene (etb), and ETD gene (etd) are carried by phage (18), plasmid (19), and a pathogenicity island (PI) (20), respectively.…”

Toxin Levels in Serum Correlate with the Development of Staphylococcal Scalded Skin Syndrome in a Murine Model

“…In the original work with the mouse model of SSSS, it was demonstrated that neonatal CD1 mice would exfoliate in response to injection with exfoliative toxin producing S. aureus, but that mice more than 7 days of age would not exfoliate. As a first step to determine the mode of resistance in adult mice, we wished to determine the time course of this apparent decrease in susceptibility to the toxin.BALB/c mice, bred and housed under barrier conditions in a pathogen-free environment at the Division of Veterinary Resources at the University of Miami School of Medicine, were inoculated with rETA previously demonstrated to be active in a neonatal mouse model (14). Mice at 1 to 10 days of life were given a single subcutaneous injection at the nape of the neck with a dose of 5 g of toxin per g of body weight and were examined at 16 h postinjection for signs of exfoliation.…”

“…BALB/c mice, bred and housed under barrier conditions in a pathogen-free environment at the Division of Veterinary Resources at the University of Miami School of Medicine, were inoculated with rETA previously demonstrated to be active in a neonatal mouse model (14). Mice at 1 to 10 days of life were given a single subcutaneous injection at the nape of the neck with a dose of 5 g of toxin per g of body weight and were examined at 16 h postinjection for signs of exfoliation.…”