clinical, microbial, and biochemical aspects of the exfoliative … · staphylococcal scalded skin...

CLINICAL MICROBIOLOGY REVIEWS,0893-8512/99/$04.0010

Apr. 1999, p. 224–242 Vol. 12, No. 2

Copyright © 1999, American Society for Microbiology. All Rights Reserved.

Clinical, Microbial, and Biochemical Aspects of the ExfoliativeToxins Causing Staphylococcal Scalded-Skin Syndrome

SHAMEZ LADHANI,* CHRISTOPHER L. JOANNOU, DENISE P. LOCHRIE,ROBERT W. EVANS, AND SUSAN M. POSTON

Division of Biomolecular Sciences, King’s College London, London SE1 9RT, United Kingdom

INTRODUCTION .......................................................................................................................................................224HISTORICAL PERSPECTIVES...............................................................................................................................224CLINICAL FEATURES .............................................................................................................................................226STAPHYLOCOCCUS AUREUS ..................................................................................................................................227

Carriers of S. aureus ...............................................................................................................................................227Outbreaks of SSSS .................................................................................................................................................227Exfoliative Toxin-Producing S. aureus Strains ...................................................................................................228

EXFOLIATIVE TOXINS ...........................................................................................................................................228Serotypes ..................................................................................................................................................................228Site of Action ...........................................................................................................................................................229Putative Mechanism of Action ..............................................................................................................................230

SUPERANTIGENIC ACTIVITY ...............................................................................................................................232Exfoliative Toxins and the Skin............................................................................................................................233Exfoliative Toxins and Other Human Diseases..................................................................................................233

SUSCEPTIBILITY AND SEVERITY .......................................................................................................................234DIFFERENTIAL DIAGNOSIS..................................................................................................................................235DIAGNOSIS AND INVESTIGATIONS ...................................................................................................................235TREATMENT..............................................................................................................................................................236PREVENTION.............................................................................................................................................................237FUTURE DIRECTIVES AND CONCLUDING REMARKS.................................................................................237REFERENCES ............................................................................................................................................................237

INTRODUCTION

Staphylococcal scalded skin syndrome (SSSS) is the termused for a collection of blistering skin diseases induced by theexfoliative (epidermolytic) toxins (ETs) of Staphylococcus au-reus. It primarily affects neonates and young children (Fig. 1),although adults with underlying diseases are also susceptible(Fig. 2). The severity of SSSS varies from a few blisters inlocalized SSSS to severe exfoliation affecting almost the wholebody surface in generalized SSSS. Because of the relative rarityof the disease and ease of treatment, SSSS has not received asmuch attention as it deserves by either clinicians or research-ers. Staphylococcal infections are on the increase in all agegroups worldwide, and show an increasing resistance to con-ventional antibiotics; despite the availability of a wide range ofantibiotics, these infections still carry a significant morbidityand mortality, particularly among adults. Furthermore, al-though the condition was described over a century ago, ourunderstanding of it began only 25 years ago, when the toxinswere first discovered. Even now, their mechanism of action isstill not certain. However, recent exciting data from computermodelling and three-dimensional crystallography of the toxinshas provided us with a clearer and more defined approach tounderstanding the pathologic processes of the disease. Eluci-dating the mechanism of action of the toxins holds excitingprospects for understanding the normal physiology of the skin,targeting drugs to very specific regions of the skin, and devel-

oping antitoxins and toxoids that may soon play a vital role inthe treatment and prevention of SSSS. The aim of this reviewis to provide both the clinician and the researcher with ourcurrent understanding—from sources worldwide and spanningmore than a century—of the epidemiology, clinical features,diagnosis, differential diagnoses, investigations, treatment andprevention of the disease. The review also discusses the organ-isms responsible for SSSS, probable mechanisms of action ofthe ETs, and their potential role in diseases other than SSSS.It is hoped that the information provided will (i) help theclinician to make a correct diagnosis of SSSS by using theavailable routine investigations and to manage the disease ap-propriately, (ii) discourage repetitive research by bringing re-searchers up to date with the current knowledge of the disease,and (iii) encourage collaboration between various researchgroups working on SSSS around the world.

HISTORICAL PERSPECTIVES

The clinical features of SSSS were first described over acentury ago by a German physician, Baron Gotfried Ritter vonRittershain, who observed 297 such cases among young chil-dren in a 10-year period while working in a Czechoslovakianfoundling asylum; he called it dermatitis exfoliativa neonato-rum but was unable to determine its cause (275). The term“Ritter’s disease,” named after its describer, is still used whengeneralized SSSS occurs in neonates. “Pemphigus neonato-rum” describes a milder, self-limiting disease of infants thatproduces a few blisters (162). S. aureus was first isolated in1891 from a patient with pemphigus neonatorum by Almquist(7), who called the organism Micrococcus pemphigi neonato-rum, and then by Winternitz in 1898 (281).

* Corresponding author. Mailing address: c/o Dr. R. W. Evans,Division of Biomolecular Sciences, King’s College London, Guy’sCampus, London Bridge, London SE1 9RT, United Kingdom. Phone:(44) 171 955 4525. Fax: (44) 171 955 8881. E-mail: [email protected].

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FIG. 1. Generalized SSSS in a previously well neonate. The neonate (A and B) shows the characteristic well-demarcated erythematous superficial exfoliation withareas of skin sparing. Courtesy of J. Etienne, Faculte de Medecine RTH Laennec, Laboratoire de Bacteriologie, Centre National de Reference Des Toxemies aStaphylocoques, France.

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The disease received little attention until 1956, when Lyelldescribed a skin eruption that resembled scalded skin andcalled it toxic epidermal necrolysis (TEN) (159). He speculatedthat the condition might be due to a circulating toxin thatdamages the skin and causes necrosis, although he was notaware that he was describing two distinct, unrelated conditions.In 1967, however, Lyell reviewed 128 cases that fitted his di-agnostic criteria of TEN and found that almost one-quarter—all children under 10 years of age—presented predominantlywith a staphylococcal infection (160). A link between staphy-lococcal infection in children and exfoliation was also observedby Lowney et al. (158).

It soon became apparent that the exfoliation associated withS. aureus infection occurred specifically within the midepider-mis whereas exfoliation that was not associated with staphylo-coccal infection occurred at the dermoepidermal junction (158,161). As a result, the former condition was termed SSSS andthe latter was termed TEN. The etiology and pathogenesis ofSSSS are very different from those of TEN, and although theymay be clinically difficult to distinguish, correct diagnosis isessential because the treatment for the two conditions is dif-ferent (236).

By this time, S. aureus had also been implicated in bullousimpetigo (60, 200), staphylococcal scarlantiniform rash (74,253), and outbreaks of Ritter’s disease (30, 200, 221). While acirculating staphylococcal exotoxin was widely accepted tocause SSSS (158, 229), it was not until the elegant demonstra-tion by Melish and Glasgow that injection of newborn micewith group II staphylococci isolated from patients with local-ized SSSS such as bullous impetigo or from patients with gen-eralized SSSS resulted in exfoliation that SSSS was appropri-ately termed (170).

CLINICAL FEATURES

SSSS is primarily a disease of neonates and children (48, 96,174), and white children may be more prone to SSSS thanblack children, according to a study of 75 cases published more

than 2 decades ago (211). The clinical features of SSSS varyfrom localized blisters to severe exfoliation affecting over 90%of the entire body surface. In localized forms of SSSS, alsoknown as bullous impetigo, tropical bullous impetigo, measlespemphigoid (162), and bullous varicella (174), characteristicfragile, thin-roofed, flaccid bullae are formed, which ruptureeasily to release fluid that varies from a thin, cloudy, amberliquid to purulent, opaque, white or yellow pus (174). This isthe mildest form of the disease, in which the surrounding skinremains normal and there are no systemic symptoms or signs(100). In neonates, the lesions are found mostly on the peri-neum, periumbilical area, or both, while in older children, thelesions are found most often on the extremities (174).

In the generalized forms of SSSS, widespread involvementof the entire skin surface can occur but the mucous membranesare usually spared (172, 208). The disease usually follows alocalized infection of the upper respiratory tract, inner ear,conjunctiva, or umbilical stump (24), although rare cases ofSSSS caused by staphylococci isolated from patients with pneu-monia, septic arthritis (174), pyomyositis (284), and maternalbreast abscesses (213) have been reported. In adults, in whomisolated cases tend to occur, bacteria are reported to enter viacatheterization, abscesses, septic arthritis, infection of arterio-venous shunts, and parenteral infections, although in manycases, no primary localized staphylococcal infection is found(48). In neonates, the age of onset is usually between 3 and 16days, and only one case of SSSS has been reported to occurwithin 24 h of birth, where a term infant, delivered by emer-gency cesarian section to a 20-year-old woman who presentedwith clinical signs of septicemia, developed SSSS. S. aureus wasisolated from the baby’s blood and urine (157). Patients withgeneralized SSSS initially develop fever, malaise, and lethargy,with poor feeding and irritability, followed by a generalized,tender erythematous rash, which generally begins on the headand neck and spreads to the rest of the body within a few days(110, 114, 193). The rash is more marked in flexural creases. Soonthereafter, large, fragile, thinroofed blisters appear, which rapidly

FIG. 2. Generalized SSSS in a 77-year-old adult with renal impairment. The photograph shows extensive exfoliation of the trunk and arm. Although the conditionis rare in adults, the observed lesions are similar to those in neonates. Reprinted from reference 103 with permission of the publisher.

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rupture on the slightest pressure (with a positive Nikolskysign), resulting in large sheets of epidermis sloughing off toleave extensive areas of raw, denuded, varnish-like skin. Thesepatients have poor temperature control, can lose extensiveamounts of fluids, and may develop secondary infection; suchsecondary complications of epidermal loss significantly con-tribute to mortality in neonates and young children (110, 135).They may also develop primary staphylococcal or secondarysepsis and may present with hypotension, tachycardia, neutro-penia, and/or respiratory distress (78, 157).

Staphylococcal scarlet fever, also called scarlatiniform eryth-roderma/rash, was first described in the 1920s and until re-cently was generally considered to be a milder or abortive formof SSSS (24, 172, 174, 266), although very little literature existsto confirm this association. Patients usually develop a gener-alized tender erythroderma with a roughened, sandpaper-liketexture, associated with fever. This is followed by thick flakesdeveloping within a few days and the entire skin desquamatingover the next week (174). Unlike generalized SSSS, the scar-latiniform eruption is not associated with the formation ofbullae or exfoliation and is very difficult to differentiate fromother infectious erythrodermal causes such as toxic shock syn-drome (TSS) and streptococcal scarlet fever. TSS is a multi-systemic disease, caused by staphylococcal toxic shock syn-drome toxin 1 (TSST-1) and other superantigens, that mayrapidly progress to death (242). It occurs mainly in menstru-ating women and is associated with tampon use, although it hasbeen found in nonmenstruating women, postmenopausalwomen, children, and men (79, 90). Recently, Lina et al. (154)analyzed 60 strains of S. aureus isolated from children withsuspected SSSS. They demonstrated that although the strainsisolated from patients with localized or generalized exfoliationproduced the ETs and caused a positive Nikolsky sign wheninjected into neonatal mice, only 1 of the 17 strains (6%)isolated from patients with staphylococcal scarlet fever pro-duced ET and the other 16 produced TSST-1, enterotoxins, orboth. Therefore, it is likely that most cases of staphylococcalscarlet fever are a clinical manifestation of mild TSS, with theenterotoxins (40) occasionally mimicking this condition. Thefew cases of staphylococcal scarlet fever caused by the ETs thatLina et al. (154) have reported may be a mild form of SSSSpreviously described by Melish et al. (172), where a transientpositive Nikolsky sign can be demonstrated in most patients. Itshould also be noted that TSS occasionally coexists with SSSS(42, 269).

STAPHYLOCOCCUS AUREUS

S. aureus remains one of the most common causes of com-munity- and hospital-acquired localized and systemic infec-tions; indeed, large epidemiological studies, including the Na-tional Nosocomial Infections Surveillance in the United States,consistently show that the incidence of S. aureus infections hasbeen increasing steadily over the past few decades in all agegroups (20, 206), including neonates (122). Despite the recentexplosion in antibiotic development, S. aureus infections stillcarry a significantly high morbidity and mortality in childrenand adults (122, 141, 169). Isolates of S. aureus are able toproduce a range of more than 30 different extracellular pro-teins, most of which play a direct role in pathogenicity orenhance virulence (112). S. aureus is probably the most fre-quently implicated organism in bacterial illness, although themechanisms by which infection is established is not alwaysknown (5).

Carriers of S. aureus

Nasal carriage of S. aureus occurs in 35% of the normalpopulation (54, 191) but varies according to age (19) and race(176). Carriage rates of S. aureus are higher in certain sub-populations such as patients suffering from atopic dermatitis(8, 152), contact dermatitis (190), psoriasis (164), and cutane-ous T-cell lymphoma (115). These carriers can be responsiblefor SSSS outbreaks (108). In neonates, organisms are alsocommon on the skin, eyes, perineum (51), wound sites (194),circumcision wounds (11), and umbilical stumps (282). Thehospital remains one of the common sources of S. aureus,mainly due to inadequate adherence to infection control prac-tices, such as barrier nursing care, hand-washing, and cleaningstethoscopes between patients (123, 155). About 30% of neo-nates are colonized by S. aureus strains within a week of birth(51, 183, 241), although some studies have shown carriage ratesof 60 to 90% of newborn babies discharged from hospitals,particularly in areas where the use of antiseptic umbilical cordcare is discouraged and during staphylococcal epidemics (105,239, 240). In hospitals, neonatal colonization is more likely tooriginate from nursery attendants than mothers (51, 183, 241),who, in turn, are more likely to be responsible for outbreaks.

A study published more than two decades ago showed thatventilation shafts and other inanimate objects such as stetho-scope bells, ophthalmoscope and otoscope handles, laundrycarts, and nursery magazines in hospitals may also harbourET-producing strains and were responsible for prolonged ep-idemics of staphylococcal infections in neonatal nurseries(123). More recently, a Nigerian study revealed that healthyanimals may also be carriers of ET-producing S. aureus strains,with 3.9% of animal S. aureus strains producing ETs (4). How-ever, their importance as reservoirs of SSSS is not known.

Outbreaks of SSSSOccasional isolated cases of SSSS can occur after organisms

colonize the infant from the maternal vaginal flora acquiredduring delivery (157). There has been one recent reported caseof a breast-fed infant developing SSSS from S. aureus thatpossibly originated from the mother’s breast abscess (213).However, more commonly, as a consequence of cross-infec-tion, SSSS infections tend to occur in outbreak clusters (29, 51,52). During the 1980s, several papers were published showingthat the use of antiseptics such as chlorhexidine on neonatalumbilical stumps delayed the time to cord separation (26, 142,227, 272). Because of the resulting increase in communitymidwives’ workload due to the delay in cord separation, as wellas rare isolated cases of hexachlorophane toxicity causingspongiform myelinopathy after percutaneous absorption (10,237), hospitals have gradually stopped using antiseptic umbil-ical cord care. This move has occurred despite previous studiesdemonstrating that the use of antiseptics for cord care is es-sential for preventing staphylococcal infection and cross-infec-tion in hospitals (47, 244).

As a consequence of stopping antiseptic cord care, manyclinicians have observed an increase in the rate of staphylo-coccal colonization and infections among neonates (6, 251,276). Furthermore, several studies have demonstrated that re-implementation of antiseptic cord care resulted in a decreasein neonatal staphylococcal colonization and infections (168,239, 240, 246). Therefore, many clinicians are very concernedthat if hospitals do not start reimplementing the antiseptic cordcare policy, the number of neonatal staphylococcal infections islikely to increase.

Over the decades, many outbreaks have been reported, of-ten with dozens of babies affected over several months and



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usually due to several asymptomatic carriers of ET-producingS. aureus strains (49, 51, 52, 55, 95). Nursery-associated out-breaks usually manifest themselves during the first month, af-ter the baby has been discharged from hospital (29, 174).Along with the recent pressure to increase day surgery andencourage early discharges from hospitals, outbreaks such asthose associated with SSSS are being dissipated into the com-munity, where infections are often treated by the primaryhealth care team—an observation made almost two decadesago (174). As a result, the hospital carrier of the offendingorganism, usually nursery attendants in close contact with theinfants, such as midwives and pediatricians (29, 51, 52, 55, 95),will continue to spread the infection over months until theoutbreak is acknowledged and the carrier is removed fromwork. Recently, for example, four babies aged 6 to 8 days werefound to have blistering rashes characteristic of SSSS during a6-week period. An inquiry and retrospective case finding re-vealed that 36 babies had been infected with an ET-producingstrain of S. aureus. Swabs from suspect health care personnelrevealed the source, a pediatrician who was suspended fromduty until shown to be free of staphylococcal colonization (29).

Exfoliative Toxin-Producing S. aureus Strains

Most S. aureus strains giving rise to SSSS in Europe and theUnited States belong to phage group II, particularly types 71and 55/71 (50, 170, 199), which account for 7 to 25% of humanS. aureus strains isolated in the United Kingdom (54). In Ja-pan, most strains causing SSSS belong to phage groups otherthan II (132, 186, 230). The only study published on the ETs inAfrica revealed that almost 60% of toxin-producing staphylo-coccal strains were nontypeable, with phage group II strainsaccounting for only 18% (4). It should be noted that only 31 to40% of phage group II S. aureus strains produce ETs (85, 128,279).

Unfortunately, due to the lack of a suitable and standardizedin vitro assay, there is very little literature available on theepidemiology of ET-producing S. aureus strains. One prospec-tive study of 944 S. aureus isolates from 577 dermatologicalpatients of all age groups revealed a 5.1% carriage rate ofET-producing strains, although only 32% of these patientspresented with any symptoms of SSSS, indicating that a largeproportion were simply asymptomatic carriers (71). Anotherepidemiological survey revealed that 164 (33%) of 500 preg-nant women attending an antenatal clinic carried S. aureus inthe nose, axilla, and/or perineum. Of the 184 S. aureus strainsisolated, 5 produced ET as determined by the neonatal-mouseassay (54). Other studies reported toxin production in 6% of2,632 S. aureus strains isolated from hospitalized patients of allages (203), 4.4% of 194 strains isolated from human diarrheaand wounds (4), 3.9% of 666 strains isolated from apparentlyhealthy animals (4), 19% of 100 S. aureus strains isolated fromthe mouths of children with dental disease (179), and 40% ofS. aureus isolated from superficial skin infections (17).

EXFOLIATIVE TOXINS

Although the presence of a soluble toxin in the pathogenesisof SSSS was the subject of speculation as far back as the 1950s(159), the first clue of its existence came only after Melish andGlasgow showed that sterile fluid obtained from intact bullaeand from phage group II S. aureus culture medium was able toproduce a positive Nikolsky sign in neonatal mice (170). Evenintraperitoneal inoculation resulted in exfoliation, indicatingthat the toxin had its specific target in the skin. Within a year,the active component in the supernatant was isolated, purified,

characterized, termed exfoliatin (125), and confirmed by oth-ers (13, 171). Soon thereafter, it was realized that at least twodifferent ET serotypes existed (131). The second serotype wasisolated and characterized as a heat-labile toxin that has asimilar molecular weight to the original toxin. This second ETserotype was able to elicit a positive Nikolsky sign in the neo-natal-mouse bioassay; the discoverers termed this toxin ETB(132).

Serotypes

At present, at least two serologically distinct toxins, ETAand ETB, can cause disease in humans and are produced by asmall but significant proportion of S. aureus strains (205). Al-though they differ in some physicochemical properties, bothtoxins produce the dermatological effects described above inneonatal mice. The toxins are species specific, affecting hu-mans, monkeys, mice, and hamsters but not rats, rabbits, dogs,hedgehogs, voles, guinea pigs, chickens, or frogs (24, 68, 125).Both toxins have been identified and fully sequenced; theyhave 40% sequence homology (21, 145, 196). ETA and ETBconsist of 242 and 246 amino acids, respectively, with molec-ular masses of 26,950 and 27,274 kDa (145). The gene for ETAis chromosomal, while that for ETB is plasmid located (91);both genes have been fully sequenced and cloned into plasmidvectors for protein expression in Escherichia coli (116, 224).ETA is heat stable and retains its exfoliative activity even afterbeing heated at 60°C for 30 min (112, 125). Initial work withEDTA, a metal ion chelator, suggested that ETA but not ETBrequired a metal ion for activity (113). However, this is nowuncertain. Comparison of the primary amino acid sequencesrevealed that amino acid residues 93 to 107 of ETA weresignificantly homologous to a region of rat intestinal and pla-cental calcium-binding protein and supported the evidencethat ETA but not ETB may require calcium for activity (53).On the other hand, calcium-binding motifs have not been iden-tified in the two recently determined crystallographic struc-tures of ETA (44, 270) (see below).

The prevalence of different ET serotypes shows a significantgeographical distribution. In the United Kingdom and Ireland,32% of 116 S. aureus isolates from patients with suspectedSSSS produced ETA, 27% produced both ETA and ETB, and12% produced ETB only, with neither toxin being detected in34 strains (56). A large epidemiological study of dermatologi-cal patients in Germany reported that 98% of the ET-produc-ing strains produced ETA (71). A French study found thatmost cases of childhood SSSS were associated with ETA alone(89%) and that only a few were due to both ETA and ETB(8%) or ETB alone (4%) (203, 279). Similarly, a Nigerianstudy reported that ETA accounted for 91.1% of the 34 toxin-producing strains isolated from a range of human and animalsources (4). In the United States, 40% of S. aureus isolatesfrom SSSS patients produced ETA, 36% produced both, and16% produced ETB only (174). On the other hand, ETB seemsto be the predominant toxin in Japan, with two studies report-ing ETB, ETA and ETB, and ETA production rates of 52, 23,and 25% of 61 strains (186) and 35, 37, and 21% of 43 strainsisolated from patients with SSSS (133), respectively.

Two other serologically different staphylococcal ETs thataffect animals have also been reported (232, 233, 263). Staph-ylococcus hyicus is known to cause exudative epidermitis inpiglets younger than 1 month; the infection is characterized byexudation, exfoliation, and vesicle formation with skin erosion(121, 144). The responsible toxin, termed S. hyicus exfoliativetoxin (ShET), has been isolated and purified from culturesupernatants of S. hyicus recently (232). This toxin, a monomer



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with a molecular mass of 27 kDa, does not cross-react withETA or ETB in immunodiffusion tests and causes exfoliationin piglets less than 1 month old and in 1-day-old chickens butnot in older pigs, 15-day-old chickens, mice, or guinea pigs(263). While still in its infancy, work on ShET has already shedlight on possible receptors for the ETs (see below) and mayprovide a more acceptable model for studying the mechanismof action of the toxins. A serologically different ET isolatedand purified from a horse strain of S. aureus was termed ETCby the authors on the basis of its similarities to the human ETs(233). ETC is a heat-labile, 27,000-kDa toxin that can produceintraepidermal splitting in both neonatal mice and chickens.However, the role of these toxins in human SSSS is still to bedetermined.

Site of Action

Most of the work on ETs has been done with ETA and theneonatal-mouse model. The toxins are made during the post-exponential phase of bacterial growth (24) and excreted fromcolonizing staphylococci before being absorbed into the sys-temic circulation (173). The toxins reach the zona granulosa ofthe epidermis by diffusing through dermal capillaries. Workwith 125I-labelled ET injected into neonatal mice has shownthat apart from the skin, there is no significant binding of thetoxins to any other body organ (94).

Histological studies have shown that addition of ETs toconfluent keratinocyte cultures isolated from pieces of humanskin obtained from plastic surgery or mouse organotypic skincultures results in the disappearance of small vesicles that areusually present between the cells (153, 166); this is followed byformation of intercellular fluid-filled gaps in the granulosa-spinosum interface, which eventually leads to the characteristicmidepidermal splitting seen in SSSS (Fig. 3). Furthermore, thebiological actions of ETA and ETB are histologically indistin-guishable (97). Cytolysis or necrolysis does not occur; an in-flammatory response or cell degeneration has not been ob-

served in the neighboring region (58, 68, 109, 285). Although S.aureus is sometimes seen in localized forms of SSSS, it is onlyrarely seen in generalized SSSS (27).

Recently, immunoperoxidase staining on histological speci-mens by Gentilhomme et al. (97) revealed that cells that bor-dered both the upper and lower edges of the cleavage sitecaused by the ET were positive for keratohyalin granules. Elec-tron microscopy of ferritin-labelled toxin and light microscopyof fluorescein thiocarbamyl toxin were previously used to showthat the toxin bound intensely to keratohyalin granules in thestratum granulosum of the epidermis and also that ETB had aparticularly high affinity toward profilaggrin (360 kDa) andfilaggrin (30 kDa) in epidermal extracts (247, 248). However,there is no evidence suggesting that the toxins have an intra-cellular mechanism of action. Profilaggrin is synthesized duringthe pathway of keratinocyte development and incorporatedinto keratohyalin granules, where it is hydrolyzed to interme-diate filaggrins and then to amino acids in the stratum corneum(24). The authors speculated that ET binding to these intra-cellular granular proteins may lead to disruption of intercellu-lar cohesion because the keratohyalin granules are linked tothe keratin intermediate filaments, which extend to desmo-somes at the cell surface. These observations fitted well withprevious electron micrographic observations suggesting thatdesmosomes, which link adjacent cells, were disrupted (172).However, several histological studies have demonstrated thateven though intercellular edema can be demonstrated, thedesmosomes are initially intact and desmosomal disruption is asecondary event, occurring only as the edema progresses (70,109, 153). Along with the above observation, some authors (62,285) also noted that adding the toxin to mouse epidermisresulted in the development of intracytoplasmic clearing alongthe cell membrane at the granulosa-spinosum interface, fol-lowed by disappearance of the cell membranes. It was specu-lated that the toxins may act by weakening cell membranes andmaking them more fragile and that the desmosome disruption

FIG. 3. Histological characteristics of skin splitting of patients with SSSS. A photomicrograph of a skin biopsy specimen from the adult depicted in Fig. 2 showsepidermal splitting at the granular layer of the epidermis. Hematoxylin and eosin stain; magnification, 3200. Reprinted from reference 103 with permission of thepublisher.



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was a secondary event (62). Therefore, the role of the desmo-some in the epidermolysis seen in SSSS is still unclear, and itis not likely to be the primary site of action of the toxins.

A more promising primary site of toxin action may be theputative receptor first identified in mice (223). Water-insolublesodium dodecyl sulfate fractions of neonatal and adult mouseepidermal extracts containing various gangliosides were incu-bated with ETA for 3 h and then subcutaneously injected intoneonatal mice. ETA incubated with neonatal (but not adult)mouse epidermis fractions was unable to induce exfoliation,indicating that an ET-inhibitory substance may be present inneonatal epidermis (223). This ability was not lost after trypsinor pronase digestion, heating at 100°C for 30 min, or neur-aminidase treatment, thereby suggesting that the receptor wasnot a protein but possibly an amino sugar. The presence of thereceptor in neonatal but not adult epidermis may explain theincreased susceptibility of neonates to SSSS, and the authorsspeculated that the identified receptor may be masked orchanged as the neonate evolves into an adult (223).

More recently, ETA and ETB were shown to bind GM4-likeglycolipid extracted from the skin of suckling mice (264). Fur-thermore, ShET from S. hyicus bound to skin extracted from1-day-old chickens but not to skin extracted from adult chick-ens. Exfoliation in the respective neonatal animal models wasabolished when the ETs and ShET were preincubated withGM4-like, but not GM1-, GM2-, or GM3-like, glycolipids fromthe skin of 1-day-old chickens and suckling mice, respectively.Their results strongly suggest that the GM4-like glycolipidsthat are present in susceptible neonatal but not adult or non-susceptible animals are possible receptors for the toxins andwould account for the age and species specificity of the toxins.Furthermore, the majority of adults who develop SSSS have anunderlying disease which may alter the normal physiology ofadult skin, making it susceptible to exfoliative toxins (48). In-flammation, for example, will induce the expression of majorhistocompatibility complex (MHC) class II molecules on thesurface of keratinocytes (245). However, despite these recentsuccesses, three decades of intensive research have been un-able to elucidate the exact mechanisms by which the toxinsinduce exfoliation.

Putative Mechanism of Action

Initial attempts to determine the mechanisms involved inexfoliation were made by studying the enzyme pattern of S.aureus isolated from clinical cases of localized and extensiveSSSS. It was initially suggested that the potent dermonecroticeffect of delta-hemolysin may be responsible, but this theorywas soon rejected after the discovery of the exfoliative toxins(96). Histological studies on the epidermal effects of ETs led tospeculations that the initial appearance followed by the disap-pearance of intercellular vesicles may be due to the release ofproteolytic lysosomal enzymes by nearby cells (153). Lyso-some-like lamellar bodies inside adjacent cells are known torelease lipids and enzymes into the surrounding extracellularspace (92, 102); proteases will produce a similar nonspecificdisruption of the extracellular matrix to some extent (24, 252).Subsequent histochemical studies did not identify any differ-ence between enzyme reactions in ET-injected, saline-injected,and uninjected specimens of mouse epidermis (62). Native ETwas also shown to have no intrinsic arylsulfatase, b-galactosi-dase, or cathepsin activity (these enzymes are normally presentin lysosomes) (278). The toxins did not have any enzymaticactivity toward known nonspecific substrates, including casein(134, 218), and using sensitivity plates (15, 21), and their ac-tivity was unaffected by any of a wide range of metabolic

inhibitors tested (24, 249). Even attempts at crystallizing eitherETA (289) or ETB (180) to determine their three-dimensionalstructure, which might shed light on their mechanism of action,met with little success because of difficulty in obtaining stablecrystals.

Takiuchi et al. (262) were the first group to provide evidencethat ETs may act as proteases. Epidermis from 1- to 3-day-oldmice (jla-ddY) was incubated with 200 mg of partially purifiedET, ET boiled for 40 min at 100°C, epidermis alone, or ETalone as negative controls. The mixtures were incubated at37°C for 12 h, and the supernatant was then tested for case-inolytic activity after removing the epidermis by centrifuging at1,000 3 g for 15 min. Incubation of ET with epidermis induceda fourfold increase in caseinolytic activity compared to thethree other negative controls; this activity was inhibited bya2-macroglobulin, indicating the activation of proteases or in-hibition of protease inhibitors in the epidermis. Contaminationwith other proteases is unlikely, because ETA or epidermisalone did not demonstrate any caseinolytic activity. However,the authors did not identify the protease in the supernatant orprovide any evidence supporting that ETA was the proteaseinduced.

A major step forward followed after both ETs were shown tohave significant primary amino acid sequence homology (25%)to the staphylococcal V8 protease, particularly in the regioncontaining the protease active site (53). V8 protease is a mem-ber of the trypsin-like serine protease family; it has an unusualspecificity for cleaving the carboxyl-terminal side of acidicamino acid residues and does not possess any disulfide bridges(64). The active site in question consists of a catalytic triad ofserine, histidine, and aspartic acid and forms the active site ofall known eukaryotic trypsin-like serine proteases. The highdegree of sequence conservation around these regions wasspeculated to preserve the active site of possible protease ac-tivity of the toxins. It was also shown that phenylmethylsulfonylfluoride (PMSF), a specific serine protease inhibitor, was ableto delay exfoliation by the ETA in neonatal mice; however,inhibition was not achieved because higher doses of PMSFwere lethal to the animals (53).

At the same time, Bailey and Smith (22) observed that ra-diolabelled diisopropyl phosphorofluoridate (i2Pr2P-F), whichhas the ability to covalently modify the active site of knownserine proteases such as V8 protease and chymotrypsin, wasalso able to label the Ser-195 of the putative active site of ETA,supporting the theory proposed by Dancer et al. that the ETsmay act as serine proteases. However, it should be noted thati2Pr2P-F could label only 6% of the total ETA (22), suggestingthat the active site of the toxin was not easily accessible toi2Pr2P-F.

Soon thereafter, several papers appeared which further sup-ported this hypothesis. Two groups independently reportedthat site-directed replacement of the Ser-195 active site of thecatalytic triad of ETA resulted in a complete loss of biologicalactivity when excess amounts were injected into newborn mice(210, 214).

Redpath et al. used oligonucleotide site-directed mutagen-esis to replace the active-site Ser-195 of ETA with glycine andshowed that injection of 1 and 10 mg of wild-type ETA into3-day-old Sha-Sha mice gave a positive Nikolsky sign within2.5 h whereas the mutated ETA had no effect at doses up to100 mg, indicating that Ser-195 is essential for the biologicalactivity of ETA (214). Prevost et al. (210) also performed asimilar experiment, replacing the Ser-195 with cysteine, andshowed that a positive Nikolsky sign was obtained in neonatalmice with as little as 0.1 mg of wild-type ETA but not with upto 300 mg of mutated ETA.



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Research on the ETs was hampered by the lack of a suitablemodel. Neonatal and hairless mice still remain the “gold stan-dard” for demonstrating the toxins as causes of exfoliation.However, this model, as well as isolated epidermis from vari-ous sources, does not allow for accurate, quantitative, andreproducible studies of toxin activity (69). A search for an invitro model for the toxins soon began in an attempt to developa rapid method to routinely identify the toxins and possibly aidin elucidating their mechanism of action. Bailey and Redpath(23) tried to determine whether the ETs might have an estero-lytic activity on the basis that proteases are always esterasesand serine proteases tend to have a higher catalytic efficiencytoward esters (76). They demonstrated that both ETA andETB have very low but significant N-t-butoxycarbonyl L-glu-tamic acid a-phenyl (boc-L-Glu-O-phenyl) esterase activity,which was not present in a mutant form of ETA where theSer-195 of the putative catalytic site was replaced by glycine(23). Furthermore, the pH dependence of both ETA and ETBin this reaction was typical of the histidine-associated pKafound with all serine proteases. On the other hand, Jaulhac etal. (118) modified the original neonatal-mouse model (262)and demonstrated an induction in proteolytic activity by usinga supernatant of the epidermis. An epidermal extract was pre-pared by grinding neonatal-mouse epidermis in liquid nitro-gen, suspending it in buffer, and subjecting it to high-speedcentrifugation. The supernatant was then incubated with ETAor ETB and different protein substrates, and the appearance ofsoluble peptides was estimated by the Lowry titration assay.Significant proteolytic activity in the supernatant with ETAand ETB (but not with biologically inactive toxin mutants) wasinduced after 20 h (118).

Many authors have argued that the proteolytic activity ob-served by both Takiuchi et al. (262) and Jaulhac et al. (118) wasdue to initial protease contamination of the toxin preparation(24, 44). However, this is very unlikely, because a threefoldinduction in supernatant proteolytic activity was obtained com-pared to the native toxin preparation or epidermis alone, andthis activity was no different from the negative controls.Jaulhac et al. (118) also showed that mutants of ETA had noproteolytic activity. This provides strong evidence that ETA isthe serine protease in the supernatant, although it is still pos-sible that altering the active site of the toxins also resulted indestruction of their ability to release or activate serine pro-tease(s) from epidermal cells. Furthermore, the authors dem-onstrated proteolytic activity against whole bovine casein, b-ca-sein, and a-casein but not k-casein or bovine serum albumin(118). Preliminary purification of one of the digested peptidesby high-performance liquid chromatography also revealed thatETA might cleave a peptide bond between glutamic acid andvaline, although the authors did not search for other cleavedpeptides (118). Their results suggested that the toxin mightcleave the peptide bond between a glutamic acid and a hydro-phobic amino acid; this has been supported by recent structuralfindings (see below).

Probably the strongest evidence supporting the toxins asserine proteases comes from some recent spectacular studieson computer models and crystallographic data. By using thestructures of known glutamate-specific trypsin-like serine pro-teases to model the three-dimensional structure of ETA andother serine proteases, a 15 to 20% sequence homology toother known glutamate-specific serine proteases (including V8protease), particularly in the region of the putative catalytictriad, was demonstrated (25). The computer model predictedthat ETA consisted of two domains, S1 and S2, each made upof six antiparallel b-strands that form a b-barrel common to allmembers of the trypsin family, although the toxin had a larger

N-terminal portion than the other members. The model alsodemonstrated that Thr-190 and His-213 (chymotrypsin num-bering), which form the core elements of the S1 pocket, wereunconditionally conserved in all serine proteases that are glu-tamate specific as well as in the ETs. However, because bothtoxins differed from the others in that they had a Lys-216protruding into the S1 pocket, it was thought that its positivecharge would help stabilize binding to a negatively chargedsubstrate, such as glutamic acid.

Crystallographic data from two independent studies con-firmed all these speculations and provided even more insightinto the structure and function of the toxins. Vath et al. (270)reported that both toxins were unique in that they possessed ahighly charged N-terminal a-helix (aN helix) containing fourpositive and four negative charges (Fig. 4). This aN helixinteracts with loop 2, which forms part of the S1 pocket, in sucha way as to partially block the pocket entrance. Furthermore,the Pro-192–Gly-193 peptide bond is flipped 180° compared tothat in the other glutamate-specific proteases. This allows theglycine to form a hydrogen bond with Asp-164 of loop D of theS2 pocket and the proline to form a hydrogen bond with theSer-195 of the active site, thereby preventing any activity bythe toxin. The authors proposed that binding of the highlycharged aN helix to a specific epidermal receptor results in aconformational change which opens the S1 pocket and flips theproline-glycine peptide bond by 180° to allow the proline toform a hydrogen bond with Asp-164 instead of the glycine. Thisconformational change then allows the active site to cleave thecarboxyl-terminal side of glutamic acid.

Cavarelli et al. (44) confirmed most aspects of the abovemodel with their own crystallographic data (44). Their free-energy simulations showed that the proline-glycine flip re-quires very little energy, less than 3 kCal per mol, and thatthere were no steric hindrances to prevent the proline-glycineflip from occurring. The possibility of binding to a specificmolecule that induces a conformational change leading totoxin activation had been proposed before (112); this event isnot novel, since thrombin (98, 99) and the hepatitis C virusNS3 protease also demonstrate such a phenomenon (130).

FIG. 4. Ribbon diagram of the structural element of ETA. Helices are in red;b strands are in blue; loops 1, 2, and D are in magenta, cyan, and yellow,respectively. Reprinted from reference 270 with permission of the publisher.



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However, the highly charged N-terminal region of the toxinshave significant sequence homology to that of trypsinogen. It istherefore possible that cleavage (by an as yet unknown mech-anism) of, for example, the first few N-terminal amino acids ofETA and ETB is sufficient (as in trypsinogen) to induce theconformational change that activates the toxins (53). However,it should be noted that larger deletions of 10 or 20 amino acidsin this region reduce toxin activity (44).

In vitro enzymatic studies by the same authors (44) withsynthetic substrates confirmed previous work by Bailey andRedpath (23) showing that the toxins could hydrolyze the car-boxyl terminal of glutamic acid of the ester boc-L-Glu-O-phe-nyl. These authors also showed that the toxins were unable tohydrolyze boc-L-Asp-O-phenyl, indicating the glutamic acidspecificity of the S1 pocket. This activity, as well as the abilityto produce exfoliation in neonatal mice, was lost when any ofthe three amino acids constituting the putative catalytic triad ofETA was replaced by another amino acid, confirming theirimportance in the observed activity.

However, there is some evidence, speculated as far back as1969, that ETs might have intrinsic lipase activity, since anal-ysis of clinical isolates of S. aureus from patients with localizedor extensive SSSS revealed that a significant proportion of thestrains had lipolytic activity toward a range of lipids (12, 261).Soon thereafter, Wiley and Rogolsky (278) demonstrated thatincubation of purified ET with either egg yolk emulsion orbovine brain sphingomyelin resulted in a dose-dependent re-lease of phosphate, suggesting that ETs have intrinsic phos-pholipase activity (278). However, it should be noted that otherinvestigators were unable to detect any lipase activity towardtriacylglycerol or phospholipids (210).

More recently, a triacylglycerol acylhydrolase of Mucor mie-hei (37) has been speculated to have structural homology toserine proteases, with His-257, Asp-203, and Ser-144 formingthe catalytic triad of the lipase. As with serine proteases, lipaseactivity could be reduced with the serine protease inhibitorPMSF. A chemically analogous but structurally different aspar-tate-histidine-serine catalytic triad has also been found in hu-man pancreatic lipases which hydrolyze triglycerides into di-glycerides and subsequently into monoglycerides and free fattyacids (280). Regarding the ETs, ETA also has the sequenceGly-Asn-Ser195-Gly-Ser-Gly, which resembles the Gly-X1-Ser-X2-Gly consensus sequence of all known mammalian and mi-crobial lipases (201). More recently, on the basis of the Lys-216residue unique to both the ETs, computer models for thetoxins were used to demonstrate that the Ser-195 active site ofthe toxins might act on lipids by cleaving phosphodiester bondsin a similar manner to alkaline phosphatase (25). The modelshowed that a phosphonyl group of a phosphodiester, insteadof a peptide, would fit into the active site of the toxin and thatLys-216, along with His-213, would serve to stabilize the neg-ative charge on the phosphonyl group. Similarly, lipases with aserine protease catalytic triad may also have an active site thatis covered by a protein region displaced during the activationprocess (37, 41, 280).

Although the ETs might have no inherent lipase activity, it ispossible that enzyme activity is induced only after activation inthe epidermis. For example, the human pancreatic lipase in itsnative form has little enzyme activity, and data from its three-dimensional structure suggest that the serine residue repre-senting the catalytic site is covered by a short one-turn a-helixsurface loop which undergoes a conformational change to ex-pose the active site at a lipid-aqueous interface (280).

Recently, preliminary work was presented in which an oc-tapeptide derived from ETB prevented the formation of aprecipitation line between ETB and anti-ETB serum. Further-

more, a subcutaneous injection of 2 mg of the peptide resultedin a positive Nikolsky sign in neonatal mice 16 h later (225).This work has not been published in a peer-reviewed journal;it is unlikely that such a small peptide could simulate the wholetoxin molecule, and the positive Nikolsky sign may be due to atoxic rather than an exfoliative effect.

In summary, current evidence suggests that ETs induce ex-foliation by possessing an atypical glutamic acid-specific tryp-sin-like serine protease activity which may be activated onlylocally by an as yet unknown mechanism. The GM4 ganglio-sides, toward which the ETs have a strong affinity, are unlikelyto be the activating factors, since such binding abolishes exfo-liation in the neonatal-animal models. It is, however, possiblethat binding to cell surface GM4 gangliosides results in inter-nalization of the toxin by keratinocytes to reach keratohyalingranules; during this journey, the toxin is activated either bypartial cleavage by an intracellular protease or by attachmentto coreceptor to induce the exfoliation seen. On the otherhand, the possibility of ETs acting as lipases, as well as theirpotential ability to activate or release other serine proteasesfrom surrounding cells, still exists. In the endless search toidentify the complex mechanism of action of the toxins, simplerexplanations should not be forgotten. A very specific substratein the epidermis, the requirement of specific microenviron-ment conditions for toxin activation, or physical disruption ofthe intercellular matrix by excess fluid-filled vesicles are only afew such possibilities.

SUPERANTIGENIC ACTIVITY

Whether staphylococcal ETs possess superantigenic activityis currently under considerable debate. Superantigens are agroup of bacterial and viral proteins that can activate a largenumber of T-cell clones simultaneously. They have been re-viewed recently (137). Conventional antigens require digestionby antigen-presenting cells (APCs) followed by surface presen-tation of a small peptide fragment complexed to an MHCprotein in order to activate a small proportion (less than 0.1%)of T cells bearing specific T-cell receptors (245). In contrast,superantigens can bypass this process of intracellular process-ing (80, 81) and can bind directly to the MHC molecule on thesurface of the APC outside the antigen-binding groove (59).This complex can then cross-link with the variable Vb region ofthe b chain of the T-cell receptor (77, 124), resulting in poly-clonal CD41- and CD81-T-cell activation (up to 20% of all Tcells) that is characteristic of superantigens (245).

S. aureus produces several exotoxins that also possess super-antigenic activity, including TSST-1 (2, 209) and the staphylo-coccal enterotoxins (SE), including SEA, SEB, SEC1-3, SED,and SEE (77, 109, 137, 198, 234, 260); new toxins continue toemerge (259). While the ETs are globular proteins with mo-lecular masses of 24 to 30 kDa, similar to the masses of othersuperantigens, the degree of primary sequence and structuralhomology such as that seen between other superantigens ispoor (165). Initial reports indicated that the ETs had mito-genic activity toward murine T lymphocytes (181), and ETAhas been shown to preferentially stimulate Vb2 human (46)and Vb3 murine T cells (106).

The main argument against the ETs being superantigensarises from work by Fleischer and Bailey (82). Cloning ETAinto a non-toxin-producing strain of S. aureus by using a shuttlevector, these investigators showed that purified recombinantETA (confirmed by ETA-specific rabbit antiserum on Westernblots), and even crude concentrated extract of the recombinanttoxin, was unable to stimulate human or murine T cells even atconcentrations of 10 mg/ml (83). On the other hand, 10 ng of



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commercially available ETA, as well as TSST-1 and SEB, perml could stimulate activity that was consistent with superanti-gens (46, 124). The authors speculated that because superan-tigens exert their effects at concentrations that are not detect-able by conventional biochemical methods, previousdemonstration of superantigenic activity with ET was probablydue to contamination by other staphylococcal enterotoxins (82,83), as was demonstrated with protein A (235). They alsomentioned that another group of researchers (107) could notprecipitate MHC class II molecules with commercial ETA,possibly because the toxin did not bind to the molecule. Sim-ilarly, ETA and ETB purified from S. aureus strains couldstimulate spleen cells of transgenic mutant mice deficient inthe expression of MHC class II antigens (45), as well as MHCclass II deficient macrophages, but did not bind to class IImolecules (28). Further arguments against superantigenic ac-tivity presented by Cavarelli et al. (44) are as follows: (i) thecharacteristic symptoms of cytokines (including rash, fever,and hypotension) induced by the action on lymphocytes ofother superantigens such as TSST-1 and the enterotoxins is notseen with the ETs; (ii) histological examination of skin frompatients with SSSS does not show any recruitment of immunesystem cells; and (iii) none of the other known superantigenspossess the speculated hydrolytic activity of the ETs.

On the other hand, Vath et al. (270) cloned ETA into asuperantigen-free strain of S. aureus and showed that the pu-rified toxin had mitogenic activity toward T cells. The authorsargued that since superantigenic activity could be demon-strated only after cloning ETA into the superantigen-freestrain of S. aureus, ETA must be the source of the superanti-genic activity and contamination by other superantigens wasunlikely. Also, recombinant ETA with a mutated active site,Ser195Cys-ETA, was shown to possess a similar mitogenicactivity, indicating that the mitogenic activity was separatefrom the exfoliative activity of the ETs. This dual action is alsoseen with the separate emetic activity and mitogenicity of theenterotoxins (33) and with the lethality and mitogenicity ofTSST-1 (188). The ETs also activate lipopolysaccharide-re-sponsive murine macrophages to produce tumor necrosis fac-tor alpha and high levels of interleukin-6 (IL-6) (compared toTSST-1 and SE) and nitric oxide (which was not produced byeither TSST-1 or the SE) resulting in contact-dependent cyto-toxicity of transformed embryo fibroblast cells, although thelevel of killing was lower than that for TSST-1 and SE (84).

Furthermore, SSSS does share many clinical features withother superantigen-mediated syndromes, including the tendererythematous rash, tachycardia, hypotension, fever, and shock(29, 114, 157, 174, 245). Since S. aureus in SSSS produces ETsat a site distant from the exfoliation (174), it is very likely thatthese systemic symptoms, particularly the characteristic rash,are a direct result of the toxins.

Exfoliative Toxins and the Skin

Recently, ETs were shown to stimulate the skin associatedlymphoid tissue (SALT) by demonstrating that the toxins ac-tivated murine splenic T cells in the presence of Langerhans’cells and MHC class II-bearing keratinocytes to produce arange of cytokines, including IL-1 and tumor necrosis factoralpha, capable of stimulating other accessory and immune sys-tem cells (268). Furthermore, binding of ETs to Langerhans’cells resulted in dose-dependent depletion of these cells (asmeasured by the number of ATPase-positive cells per squaremillimeter) in cultured mouse skin, presumably due to migra-tion of the cells to draining lymph nodes, where they may actas APCs for T lymphocytes (202). SALT is part of the immune

system that deals with antigenic challenges common to the skinand includes antigen-presenting Langerhans’ cells, keratino-cytes, and T-cell subsets with skin-homing receptors (258).Despite the presence of MHC class II molecules, which areinduced on the cell surface during inflammatory processes(273), keratinocytes are unable to act as conventional APC andseem to function only in the presence of superantigens (268).Activation of SALT by ETs may therefore result in the ery-thematous rash associated with SSSS. For example, there issome evidence that the ability of the superantigen TSST-1 toactivate cutaneous lymphocyte-associated antigens may be re-sponsible for the rash and desquamation seen in TSS (149),just as the superantigens of group A streptococci might play arole in the development of the characteristic cutaneous swell-ing, erythema, and desquamation seen in streptococcal infec-tions (254). However, the lack of immune cells on histologicalspecimens from patients with SSSS cannot be easily explainedand requires more research. It may be that the very specific(possibly intracellular) site of action of the toxins does notallow them to exert superantigenic activity in terms of durationor location or that activation of the toxins to proteases, whichprobably requires a structural change (25, 44, 270), results inloss of superantigenic activity at the site of action.

The ability of the ETs to activate the SALT subpopulation ofthe immune system has important consequences for severaldermatological conditions, including cutaneous T-cell lym-phoma and atopic dermatitis. ETA is capable of inducing theproliferation of Vb2.1-bearing cutaneous T-cell lymphomacells in vitro, and this proliferative response is enhanced byadding IL-1. This fits in with observations showing that activa-tion of SALT by ETA results in IL-1 release (268), and IL-1,along with IL-6, plays an important role in enhancing theactivity of other superantigens (136).

Patients with atopic dermatitis have a higher staphylococcalcolonization rate than does the general population (8, 152).Antibiotic treatment of patients with atopic dermatitis hasresulted in significant clinical improvement of the condition(150), suggesting that staphylococci play a role in the patho-genesis of the disease. Recently, it has been shown that a largeproportion of S. aureus strains isolated from patients withatopic dermatitis release superantigens, including enterotox-ins, TSST-1, and ETs (147, 167). It has been speculated thatsuperantigens bind to surface MHC class II molecules foundon keratinocytes during inflammation and thereby activatepolyclonal T cells locally (257).

Exfoliative Toxins and Other Human Diseases

The Vb-expansion of T cells characteristic of superantigenshas also been demonstrated in other conditions, where the roleof staphylococcal toxins is still unclear. Kawasaki disease, alsoknown as mucocutaneous lymph node syndrome, is a multisys-tem disorder in children that is characterized by fever, rash,conjunctivitis, mucosal inflammation, erythematous peeling ofthe hands and feet, and, rarely, coronary artery aneurysms(147, 175). A selective expansion of Vb2- and Vb8-expressingT cells has been detected in the blood of patients with Ka-wasaki disease (1), and, while TSST-1 has been detected in aproportion of these patients (146, 147, 148), the potential roleof the ETs has not been studied. Other conditions where ex-pansion of Vb-expressing T cells, often by staphylococcal su-perantigens, has been demonstrated include hematogenouslyacquired staphylococcal nephritis (138, 271), staphylococcalseptic arthritis (38, 290), various autoimmune diseases (137,184), rheumatoid arthritis (197), multiple sclerosis (39, 256),contact sensitivity (228), and guttate and chronic plaque pso-



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riasis (151, 288); another possible role for the expansion ofVb-expressing T cells is in the gradual decline in the number ofCD41 T cells in human immunodeficiency virus infection(143). Unfortunately, many of the studies on the superanti-genic effects of ETs in various diseases have used commerciallyprepared toxins, which, as many authors have pointed out, maybe contaminated by other superantigens. Future studies shouldtake this factor into consideration and use appropriate precau-tions to eliminate the problem.

Several other conditions have demonstrated a significantassociation with staphylococcal toxins; sudden infant deathsyndrome (SIDS) is probably one of the most important (31).S. aureus strains producing multiple toxins have been isolatedfrom infants with SIDS (265). In addition, S. aureus is the mostcommon organism isolated from 2- to 4-month-old infants,being found in almost 50% of the infants (222). This age groupis also associated with the highest incidence of SIDS (31). Leeet al. (145) have shown that bacterial isolates, particularly S.aureus and Escherichia coli, from infants with SIDS were lethalto gnotobiotic rats. Therefore, some authors have proposedthat common bacterial toxins absorbed through the respiratorytract might be involved (182). Furthermore, staphylococcalinfection in combination with influenza virus infection wasshown to greatly increase the production of SE, resulting in anexaggerated inflammatory response in ferrets (117). This ac-tivity might explain the higher prevalence of SIDS in winter(43). Although TSST-1 was demonstrated in renal tubular cellsof some infants with SIDS but not control infants (189), therole of the ETs in SIDS has, unfortunately, never been studied.It is possible that an abnormal inflammatory response due tothe superantigenic properties of the ETs, which may be pro-duced locally in the nasopharynx and then systemically ab-sorbed (189), coupled with enhancement by concurrent viralinfection will result in sudden death in infants.

SUSCEPTIBILITY AND SEVERITY

A whole range of host and organism factors govern thedevelopment and severity of SSSS, and many of these are notfully understood. Furthermore, it is still not clear why neonatesare more susceptible to SSSS than adults. Initially, anatomicaldifferences in the skin were believed to be the reason, basedmainly on the fact that the toxins induced exfoliation in neo-natal but not adult mice and that, in humans, the condition wasseen primarily in neonates. However, this premise has nowbeen refuted for several reasons: (i) healthy adults of thehairless mice strain will develop SSSS when injected with ET(126); (ii) adult mice can also develop localized SSSS at the siteof toxin injection (93) and can even develop generalized SSSSif they are immunosuppressed with drugs such as prednisoloneand azathioprine (277); (iii) SSSS cases have been reported inhealthy human adults and in adults with underlying disease(48, 194); and (iv) intradermal injection of purified ET into theforearm of a healthy human adult will result in blister forma-tion characteristic of SSSS (127, 277). Nevertheless, anatomi-cal differences are the most likely candidates in determiningspecies susceptibility (69).

Poor renal clearance of the toxins by neonates and by adultswith impaired renal function is a major risk factor for devel-oping SSSS and may partly explain why neonates may be moresusceptible to developing SSSS. Adult mice excrete one-thirdof a test dose of intravenous ETA within 3 h compared to 1/15of a test dose in newborn mice (94). As a consequence, toxinlevels reach a higher peak in the newborn and decline moreslowly. Furthermore, nephrectomized (but not hepatecto-mized) adult mice develop generalized SSSS when challenged

with ET. However, renal function alone cannot explain whyinjection of very high doses of ET into adult mice, which wouldbe expected to more than compensate for the rapid renalclearance, does not result in exfoliation (94).

Antibody status may also have a protective function, and thishas been shown in mice, in which inoculation of small doses oftoxigenic S. aureus, for example, resulted in exfoliation only inneonatal mice that had not previously been immunized withpurified extracts of ET (173). A large epidemiological studylooking at the antibody status of patients over the whole rangeof SSSS spectrum was also performed (173). Anti-toxin anti-bodies were found in 88% of infant cord blood samples. Thesefindings reflect passive maternal antibodies, which, in turn,may protect newborn infants. This finding is further supportedby the observation that infants involved in neonatal-nurseryoutbreaks tend to have the localized rather than the general-ized form of the disease. In addition, work with mice showedthat maternal antibodies play an important role in protectingoffspring from subcutaneous challenges of ETA (163).

The large epidemiological study also showed that antibodylevels fell to 30% in patients between 3 and 24 months of ageand then rose steadily to 50% in those older than 10 years andto 91% in those older than 40 years. Furthermore, the sameauthors noted that 13 (62%) of 21 and 15 (100%) of 15 patientswith localized SSSS had anti-toxin antibodies in acute-phase (5days) and convalescent-phase (14 days) sera, respectively, incontrast to 0 of 10 and 5 of 5 patients with generalized SSSS(173). The authors concluded that while antibodies may notalways protect against exfoliation, they may serve to localizethe infection around the port of entry of the responsible S.aureus strain. In support of this, the blisters of localized SSSS,such as bullous impetigo, contain S. aureus while the blisters ofgeneralized SSSS, where ETs are produced by S. aureus at adistant site, tend to be sterile (70). Although the study involvedETA only, the overall conclusions are unlikely to be affected byexcluding ETB, and at least two other studies have confirmedthe above findings (173).

Protective antibodies may also explain why only a proportionof nasal carriers of toxin-producing S. aureus are infected.However, it is possible that the strains harbored by these car-riers are not active producers of the toxins. The antibody the-ory also provides an explanation for observations that breast-feeding is protective in neonates.

While adult SSSS is very uncommon, more than 30 caseshave been reported (this is likely to be a gross underestimation,since the disease was linked to the ETs only in 1970) and havebeen reviewed recently (48). Of the 32 reported cases, 21(66%) were in males, and the ages of the patients ranged from19 to 91 years; more than half of the patients were older than60 years. Most of the adults with SSSS had an underlyingdisorder such as poor renal function (63); immunosuppressionwith immunosuppressive drugs; infections such as HIV andAIDS (63); malignancy including Hodgkin’s and non-Hodgkin’slymphomas, metastatic breast carcinoma, and acute andchronic myeloid leukaemias; and other less common causessuch as chronic alcohol abuse, heroin misuse, diabetes mellitus,cachexia, and rheumatic fever (96, 173). Another review ofadult SSSS found that 78% of patients had factors suggestingpossible impaired immunity such as those described above(32). Furthermore, renal impairment was found in 73% ofthose who had their renal function tested, and many had frankrenal failure.

There have been only two reports of apparently healthyimmunocompetent young adults developing SSSS, and bothcases were due to ETB (185); it is not known whether this ismerely a coincidence or whether the ETB subtype is more



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pathogenic. Another epidemiological study (185) did note thatalthough both toxins were equally responsible for localizedSSSS such as bullous impetigo, ETB was more frequently iso-lated from children with generalized SSSS than was ETA alone(20 of 24 and 2 of 24, respectively). Recently, a case of SSSSwas reported in an immunocompetent 73-year-old man whohad very mild renal impairment and who was taking ibuprofen(103). Nonsteroidal anti-inflammatory drugs have been previ-ously reported to predispose to SSSS (129), possibly by a com-bined effect of inhibiting prostaglandin-mediated renal vasodi-lation and inhibiting neutrophil function. However, as theauthors correctly point out, because such drugs are so widelyused, this association may be purely coincidental.

DIFFERENTIAL DIAGNOSIS

While the mortality rate from SSSS in neonates and childrenis low, it is important to make a correct diagnosis in order toinitiate early and appropriate treatment, as well as to preventmedicolegal implications, since the disease may result in alle-gations of burns by neglect (160) or child abuse (231). Differ-ential diagnoses of generalized SSSS in neonates are few andinclude drug- or virus-mediated TEN, burns, epidermolysisbullosa, bullous erythema multiforme, listeriosis, syphilis, dif-fuse cutaneous mastocytosis, and graft-versus-host rejection(65). Chemical burns from petrol, paraffin, boric acid, ethylene,acrylonitrile fumigant spray, tribromofluorene, and monosul-firam, which act at the dermoepidermal junction, must also beexcluded (162). TEN, also known as Lyell’s syndrome after theauthor who first described it (159), is probably the most im-portant differential diagnosis of SSSS, since their treatmentsare very different (236). TEN is an uncommon but life-threat-ening cutaneous disorder that occurs as frequently in childrenas in adults (215). More than 100 different drugs, includingantibiotics and anti-inflammatory drugs, and occasionally im-munizations and infections such as with members of the familyHerpesviridiae, have been implicated in TEN (236). TEN oc-curs 1 to 3 weeks after drug ingestion, although reexposure tothe offending drug may result in a more rapid and very severepresentation, with involvement of multiorgan systems includ-ing the liver, kidneys, gut, and lungs (212, 236). The dermato-logical features are characterized by an initial prodrome ofpyrexia, malaise, anorexia, pharyngitis, and a tender, morbill-iform rash followed by epidermal exfoliation in sheets (236).As in SSSS, fragile blisters are also seen in TEN and theNikolsky sign is positive (18, 236). However, unlike SSSS, erod-ing mucosal lesions of the mouth, conjunctiva, trachea, bron-chi, esophagus, anus, and vagina (18) are almost always presentin TEN, as demonstrated in one study of 87 patients (215), andcan be differentiated histologically from SSSS lesions (see be-low).

Localized SSSS such as bullous impetigo is usually charac-terized by blisters that dry up without producing any markedcrusting (162, 178). On the other hand, in streptococcal impe-tigo there is copious exudation from raw surfaces that driesinto a characteristic thick, dirty yellow crust, which soon re-forms if removed. Furthermore, glomerulonephritis is a knowncomplication of the latter but not the former condition (162,174). However, staphylococcal impetigo and streptococcal im-petigo are often clinically indistinguishable, and antibiotictreatment should cover both groups of organisms.

The initial generalized erythematous rash seen in SSSS maybe confused with streptococcal scarlet fever, but the very ten-der skin, exfoliation, lack of a strawberry tongue, and possibleperioral and periorbital crusting seen in SSSS should confirmthe diagnosis (174). Congenital epidermolysis bullosa can

present with exfoliation of the skin upon slightest mechanicalpressure, but, unlike SSSS, it presents from birth. Other con-genital bullous diseases include acrodermatitis enteropathicaand bullous ichthyosis. Occasionally, bacterial (staphylococci,streptococci, E. coli, or, rarely, Haemophilus spp.) (207), fungal(Candida spp.), and viral (herpesviruses, vaccinia virus, vari-cella-zoster virus) infections resemble SSSS. Isolated cases ofdiffuse cutaneous mastocytosis (195), Kawasaki disease (79),psoriasis (217), and TSS (66) have been reported to mimicSSSS.

DIAGNOSIS AND INVESTIGATIONS

Most cases of SSSS are diagnosed on clinical grounds andare easily treated with antibiotics, which rapidly eliminate thestaphylococci producing the toxin. Laboratory investigationsare required only if the clinical findings are equivocal or whenoutbreaks occur. Because the condition is the result of exotox-ins which may be produced by staphylococci at a distant site,the blister fluid in generalized SSSS tends to be sterile whereasthe fluid in localized bullous impetigo will contain S. aureus(70, 158, 208). Staphylococci producing ET can usually becultured from the nares, conjunctiva, or nasopharynx (50, 158,229). Blood cultures are usually negative because the organ-isms are frequently noninvasive, particularly in children (49,114, 157, 208, 211). In one study, only 3% of children had apositive blood culture (101), in contrast to 20 (62.5%) of 32adults (48). Of note, children with renal disease tend to havebacteremia with generalized SSSS rather than the localizedform (34). Bacteriological studies of the skin are not usuallyuseful in differentiating SSSS from TEN, since the skin ofpatients with generalized SSSS is usually sterile and that ofpatients with TEN is often colonized by S. aureus (67, 236).Indeed, up to half of the deaths in patients with TEN are dueto secondary bacterial infection of the denuded skin (18).

Phage typing is not routinely available in hospital laborato-ries. In the United Kingdom, bacterial swabs of S. aureus canbe sent to the Public Health Laboratory at Colindale, London,for phage typing, despite this test having poor reproducibilityand requiring special reagents and a large number of tests (95).Many authors assume that the presence of characteristic skinlesions, including a positive Nikolsky sign, along with the iso-lation of phage group II S. aureus from a distal site is sufficientfor the diagnosis of SSSS (for a review, see reference 72).However, it is now generally agreed that the production of ETis not associated with any particular phage type, since phagetypes other than group II will also produce the ETs and only aproportion of phage II strains produce ETs (56, 85, 174). Forexample, Falk and King (72) reported the case of a 43-year-oldschizophrenic who presented with fever, erythema, and exfoli-ation with a positive Nikolsky sign but no mucosal involvementand whose routine skin swabs contained S. aureus phage type71. He was treated for SSSS until a blister biopsy revealed adiagnosis of TEN. Phage typing is currently not acceptable forconfirming the diagnosis of SSSS since it is neither sensitivenor specific (56, 174, 185).

A biopsy of the blister is one of the most definitive diagnostictests in SSSS. One study revealed a positive blister biopsyresult with intraepidermal cleavage in all 30 adults with SSSS(48). Blister biopsy is also essential in ruling out other bullousdermatoses (42, 75, 269). Histological testing will clearly ex-clude conditions involving subepidermal separation, includingerythema multiforme and TEN, bullous lichen planus, poly-morphic light eruptions, graft-versus-host rejection, and non-inflammatory processes such as bullous amyloidosis, Wilson’sdisease, drug ingestion, epidermolysis bullosa acquisita, and



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porphyria (75). To rapidly differentiate TEN from SSSS, aTzanck preparation from a freshly denuded area will showlarge polygonal epithelial cells with large nuclei and no inflam-matory cells in SSSS and only a few rounded or cuboidalepithelial cells with many inflammatory cells in TEN (9, 73).

Histologically, SSSS needs to be distinguished from otherintraepidermal conditions, including TSS and nutritional dis-orders. TSS is usually diagnosed clinically (90), with or withoutlaboratory tests to detect TSST-1 and other superantigens(178). Histological tests for the diagnosis of TSS are onlyoccasionally required and show spongiosis, scattered necrotickeratinocytes, and scattered neutrophils, with a perivascularand interstitial mixed-cell infiltrate and dilatation of the super-ficial plexus and edema in the dermis (111). Nutritional defi-ciency syndromes, including pellagra, zinc deficiency, biotindeficiency, essential fatty acid deficiency, and the glucagonomasyndromes (necrolytic migratory erythema), may present witherythematous, scaly, and vesicular eruptions (3). All have ahistologically similar picture, with intercellular edema in theupper half of the stratum spinosum that may be severe enoughto cause intraepidermal vesiculation (3). Along with SSSS,subcorneal pustular dermatosis and pemphigus foliaceus aretwo other causes of subcorneal blistering; they can be distin-guished from SSSS by the presence of a large number ofneutrophils in the blister and by distinct immunohistochemicalfindings respectively (75). Lesions from bullous impetigo arecharacterized by stratum granulosum epidermal cleavage witha local infiltrate of staphylococci and polymorphonuclear leu-cocytes (174).

If further confirmation is required, the Public Health Lab-oratory in the United Kingdom may attempt to detect the ETsby using the Ouchterlony method. This method is known to becrude, unreliable, and time-consuming with poor sensitivityand is unsuitable for batch testing of large samples (139).Furthermore, our work (139) and work with diagnostic kits forTSST-1 (219, 238) have shown that protein A in culture super-natants is produced by more than 90% of S. aureus strains (88,89), resulting in high rates of false-positive results due to non-specific binding to the detecting antibodies (36, 255).

While several techniques have been proposed in the past tostudy the effects of ETs, few have been developed. Subcutane-ous injection of small amounts of the partially purified toxininto neonatal (,5-day-old) Sha-Sha, BALB/c, CD-1, TA, ICR,Swiss-Webster, or Park mice (56, 57, 170) or adult hairlessmice (14, 126) results in a positive Nikolsky sign 2 to 120 minafter inoculation, although the adult hairless-mouse model isless sensitive. While such mouse models remain the gold stan-dard for confirming the diagnosis of SSSS, they are clearly notsuitable for routine rapid identification of ETs in hospital lab-oratories. Gel immunoprecipitation (Mancini test) is expensiveand has limited sensitivity (16). Radioimmunological assays(RIA) are more recent tests developed for detecting the toxins(288), since they can detect ETA at 100 pg/ml by using 125I(173); these tests are very sensitive and quantitative but are tooexpensive and time-consuming for routine use, and they re-quire the handling of radioactive materials. Simple enzyme-linked immunosorbent assays (204) are not as sensitive as RIA,but a double-antibody procedure, in which a secondary ratpolyclonal antibody is used, has attained the sensitivity of RIA(156). Other methods developed include slide latex agglutina-tion (187), electrosyneresis and DNA hybridization with oli-godeoxynucleotide probes (204, 216), and genomic DNA fin-gerprinting after SmaI digestion of DNA extracted from S.aureus strains (95). Unfortunately, most of these tests havebeen developed for research purposes and are not suitable as

routine tests in hospital laboratories because they tend to betime-consuming, laborious, and expensive.

More recently, PCR with specific ETA and ETB primers hasbeen used to identify the toxin genes in suspected strains of S.aureus (120, 226). For example, Sakurai et al. correctly iden-tified all 30 ETA-, 20 ETB-, and 6 ETA- and ETB-producingstrains within 3 h (226). Thus, this technique is rapid andsensitive and has been used to confirm the diagnosis of SSSS insuspected clinical cases (288). Random amplified polymorphicDNA analysis, which is simple and rapid to perform, has alsobeen used to demonstrate that the S. aureus strain isolatedfrom a mother’s breast abscess was identical to the straincausing SSSS in her breast-feeding infant (213).

TREATMENT

Although SSSS can be very severe, it is easily managed withantibiotics to eradicate the offending toxin-producing S. aureusstrain and appropriate nursing care to prevent and/or treat anysecondary symptoms. Neonates must be isolated from otherbabies and strict barrier precautions must be applied to pre-vent cross-infection (79). A focal source of infection, whichmay be absent in more than half the patients (48), must besought and treated to prevent further spread of the disease.This includes blood, wound, eye exudates, and throat swabsinitially, followed by osteomyelitis, septic arthritis, and occultfoci if the source is not obvious (174). Potential carriers oftoxin-producing strains must also be identified and treated toprevent cross-transmission, which may result in outbreaks. Thisinvolves culturing nasal swabs for S. aureus from all personnelwho have handled the infected baby (79).

Mild forms of SSSS such as bullous impetigo should bemanaged with cleaning and drying of the lesions with com-presses and administration of oral antibiotics that are effectiveagainst penicillin-resistant staphylococci as well as strepto-cocci. Methicillin, nafcillin, dicloxacillin, cloxacillin, and flu-cloxacillin have all been used to treat S. aureus, and otherproposed regimens include cloxacillin plus tobramycin, anderythromycin (48). Multidrug-resistant strains are rarely en-countered in phage group II S. aureus (48, 279). One recentstudy of 61 ET-producing S. aureus strains revealed that 7%were resistant to tetracycline, 5% were resistant to gentamicin,and 2% were resistant to chloramphenicol. None of the strainswere resistant to methicillin, cephalosporins, or vancomycin(186). Another epidemiological study of 82 methicillin-resis-tant S. aureus (MRSA) strains isolated from outbreaks in neo-natal intensive-care units in Japan revealed that only 1 strainproduced ETA and none produced ETB (287). However, thesame authors also reported a case involving a 6-month-oldinfant with SSSS due to a methicillin-resistant ETB-producingphage II S. aureus strain in Japan, who was successfully treatedwith minocycline (287).

Topical antibiotics are often not effective and should beavoided due to unpredictable absorption through denudedskin; even localized forms of SSSS such as bullous impetigoshould be treated with oral antibiotics, which have been shownto be more effective than topical antibiotics (61). Steroids arecontraindicated (157, 208) on the basis of both experimental(171) and clinical (220) evidence.

Severe forms require more aggressive treatment with intra-venous antistaphylococcal antibiotics and extra care of de-nuded skin to prevent secondary infection and fluid losses andto maintain body temperature, especially in neonates. Addi-tional broad-spectrum antibiotics should be considered if sec-ondary skin infection is suspected, and they should coverPseudomonas species (110). Intravenous crystalloids should be



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used to replace fluids, and an incubator should be used tomaintain body temperature. Sterile wrapping and minimalhandling may be used to decrease conductive heat, fluid losses,and the risk of secondary infections. Semipermeable dressings,such as those used to cover graft donor sites, have been used toprovide a barrier against evaporative fluid losses (177). Topicalapplications such as silver sulfadiazine, which are sometimesused for burns and TEN, are not recommended for SSSSbecause of enhanced systemic absorption through denudedskin (86). Because the complications of severe SSSS resemblethose of severe burns, many clinicians prefer treating suchpatients in specialized burns units (243). A few severe cases ofSSSS have benefited from extracorporeal detoxification involv-ing sorption detoxification and separation plasmapheresis(250).

Within 2 to 3 days, fever usually subsides, the erythemabegins to resolve, and no new bullae form (157). Desquama-tion of the skin occurs 3 to 5 days after onset, and completeresolution occurs in most cases, usually within 2 to 3 weeks,with no permanent sequalae (100, 114, 157). Serious compli-cations, including lung abscesses, staphylococcal pneumonia,and osteomyelitis, have been reported (87, 104). Mild emol-lients may be used if the skin becomes very dry during thehealing process (114).

The prognosis of SSSS in children who are appropriatelytreated is good, with a mortality of less than 5% (48, 96, 174).Nevertheless, some children will die despite antibiotic therapy(211). In contrast, mortality rates in adults are very high, usu-ally due to underlying diseases, despite aggressive treatment.One review of adult SSSS reported that 19 (59%) of 32 patientsreported in the literature died despite antibiotic therapy (48);all but 2 of those who died had severe underlying disease, andthe prognosis was not necessarily better in those with localizedforms of SSSS.

PREVENTION

Hospitals remain the most important source of S. aureus,and simple barrier nursing techniques are still considered to bethe most effective method of preventing transmission and out-breaks of infection. Using MRSA as a parallel example, rein-forcement of hygiene measures has consistently led to a de-crease in MRSA colonization and infection rates (35, 119, 155,185). Simple measures such as hand-washing, minimal han-dling, cleaning objects such as stethoscopes and thermometersbefore use, and avoiding unnecessary invasive catheterizationall contribute to prevention of cross-infection (119). Such asep-tic technique should be encouraged among all health careprofessionals and reenforced at all levels.

Any patient developing SSSS should be immediately iso-lated. Anyone else at risk of developing SSSS who may havebeen exposed to ET-producing S. aureus strains will benefitfrom prompt administration of prophylactic anti-staphylococ-cal antibiotics, which, in mice, have shown to abort exfoliationif given early enough (172). Furthermore, carriers of toxin-producing strains should be rapidly identified and treated toprevent outbreaks. It is important to emphasize that any oneoutbreak may have more than one carrier (49, 52, 55). Unfor-tunately, a rapid diagnostic kit for SSSS is not yet available. Asa consequence, there is often a delay in diagnosis as well asidentification of carriers. The organism has to be isolated andsent to the Public Health Laboratory for phage typing and thenpossibly for culturing to produce the ETs, which are thentested for by the Ouchterlony method (140).

FUTURE DIRECTIVES AND CONCLUDING REMARKS

Research into SSSS is now reaching an exciting era. Eluci-dation of the three-dimensional structure of the ETs has givenus an insight into the mode of action of the toxins. The precisemechanisms by which they split the epidermis will be deter-mined in the very near future, and their ability to target to avery specific region of the epidermis can be exploited in manyways (140). In particular, the targeting domain of the toxinsmay be used to deliver drugs to precise intraepidermal loca-tions for the treatment of dermatological conditions such asneoplasms, as has already been demonstrated with Corynebac-terium diphtheriae and Pseudomonas toxins (283). Further-more, if the ETs are confirmed to be superantigens, they mayplay a wider role in a whole range of dermatological and otherdiseases.

Similarly, there have been studies showing that antitoxinantibodies will protect neonatal mice from lethal doses of ET,even if the antitoxin is administered late during the course ofthe disease (127). Because antibiotics are targeted to the of-fending organism, exfoliation will continue for 24 to 36 h afterantibiotic administration. Development of an effective anti-toxin based on the structure and function of the toxin may beinvaluable in halting the progression of exfoliation in severecases of generalized SSSS, particularly in adults, in whom mor-tality rates are high (140). Although the rarity of the diseasewould not support large-scale immunization, the use of toxoidsin immunization against SSSS may also play a role for at-riskgroups such as neonates with immune deficiency or adults withcancer or renal failure.

The need for a rapid diagnostic kit for SSSS can no longer beignored. Current methods of diagnosis are neither sensitivenor specific, and delay in diagnosing the disease will result inincreased morbidity and mortality, particularly among adults.Most hospital laboratories have facilities to detect a wide rangeof microbial toxins, and the ETs should be no exception. Rapiddetection of the ETs is important not only in diagnosing SSSSbut also in detecting carriers and screening at-risk patients forET-producing S. aureus carriage to prevent the development ofSSSS.

Clinicians should also be aware of the need for increasedsurveillance of staphylococcal infections in general. The pres-sure to discharge babies early from hospitals, along with therecent trend to stop using antiseptic neonatal umbilical cordcare, is likely to increase the prevalence of staphylococcal in-fections in the community. Therefore, it might be useful todevelop a central reporting system, whereby primary-care phy-sicians report to the local hospital any neonatal infection in thecommunity so that possible outbreaks dissipated into the com-munity would be rapidly recognized and contained and emerg-ing trends such as antibiotic resistance would be quickly de-tected and dealt with.

It is hoped that the relative rarity of the disease will not lullresearchers and clinicians into a false sense of security and thatthe recent exciting developments in the field will strengthenefforts to conquer the disease once and for all.

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