Investigation of the skin microbiome: swabs vs. biopsies.

BACKGROUND: Human skin is populated by diverse bacteria and there is increasing evidence that resident bacteria play a key role initiating immune responses in cutaneous diseases such as atopic dermatitis, psoriasis and hidradenitis suppurativa. Bacteria are present at all layers of the skin but many studies have relied on swabs to profile the skin microbiota. OBJECTIVES: As the pathogenesis of many skin conditions is dermal, we wanted to compare the microbiota obtained in swabs (surface) and biopsies (dermis). METHODS: Using 16S rRNA gene sequencing we established the microbial profiles of skin swabs and skin biopsies in 16 patients. RESULTS: We found differences in both diversity and taxonomic composition of the microbiome obtained from swabs and biopsies of the same individual. Several taxa were found to be more abundant in the swabs, which displayed significantly higher community richness, but Clostridiales and Bacteroidetes were significantly enriched in the biopsies. Most published research on cutaneous microbiota has been based on skin swabs, which represent the surface of the skin. CONCLUSIONS: Our study demonstrated a clear difference between the microbiome observed from skin swabs and skin biopsies. These findings may be highly relevant in disorders such as psoriasis where pathogenesis arises in the dermis. What's already known about this topic? 16S RNA gene sequencing has facilitated study of the skin microbiome. Several studies have sequenced the microbiome sampled by skin swabs. What does this study add? The microbiome data obtained using swabs and biopsies were different. Diseases that are predominantly dermal should be studied using both swabs and biopsies.

Is chronic plaque psoriasis triggered by microbiota in the skin?

There is a known association between psoriasis and Crohn disease (CD). Patients with CD are five times more likely to develop psoriasis, and, conversely, patients with psoriasis are more likely to develop CD. Many gastroenterologists now accept that CD results from a breakdown of immune tolerance to the microbiota of the intestine in genetically susceptible individuals. The microbiota of the skin have recently been investigated in psoriasis. Firmicutes was the most common phylum, and Streptococcus the most common genus identified. Beta-haemolytic streptococci have been implicated in both guttate and chronic plaque psoriasis. Furthermore, the innate immune system has been shown to be activated in psoriasis, and many of the genes associated with the disease are concerned with the signalling pathways of the innate immune system, notably interleukin-23 and nuclear factor κB. Patients with psoriasis also have an increased incidence of periodontitis, a disease thought to be due to an abnormal response to normal oral commensals. Based on the similarities between CD and psoriasis, we propose that psoriasis is due to a breakdown of immune tolerance to the microbiota of the skin. In support of this hypothesis we provide evidence for microbiota in the skin, activation of the innate immune system, and genetic abnormalities involving the innate immune system.

Concepts in psoriasis: psoriasis and the extracellular matrix.

Post-streptococcal guttate psoriasis only sometimes progresses to the persistent plaque psoriasis. We have previously proposed that psoriasis is driven by multiple extradomain A+ fibronectin (EDA+ FN) 'feedback loops'. The psoriasis-specific loop, the 'keratinocyte loop', depends upon expression by keratinocytes of α5ß1 integrin. In normal skin, α5ß1 expression is minimal or absent in resting epithelial cells, where basal cells are anchored to the laminin component of the basement membrane (via integrins α6ß4 and α3ß1). However, when the laminin layer is disrupted, due to wounding for instance, α5ß1 is then strongly expressed by basal cells as a means for anchoring. At uninvolved skin sites in patients with psoriasis the laminin layer within the basement membrane is disrupted, with a consequential increase in the expression by basal keratinocytes of EDA+ FN and α5ß1. We are postulating that in guttate psoriasis streptococcal lytic enzymes, and in particular streptokinase, may - via plasminogen activation - initiate a disruption of the laminin layer in the basement membrane. It is also possible that the expression by proliferating keratinocytes of plasminogen-activating factor might result in additional laminin digestion. The suggestion is that progression of guttate psoriasis to full-blown plaque psoriasis, or to its spontaneous resolution, is governed largely by the triggering of humoral immune responses. Thus, if streptococcal lytic enzymes are neutralized by host antibodies in advance of irreversible and lasting damage to the basement membrane of the dermal papillae, then guttate psoriasis will resolve spontaneously. In contrast, if permanent damage is induced prior to effective neutralization of the relevant bacterial toxins, then keratinocytes will become chronically destabilized. The consequence will be to liberate keratinocytes to proliferate in response to various stimuli, and to continue to elaborate plasminogen activation enzymes; the combined effect being, in concert with immunological priming, to precipitate and sustain plaque psoriasis.

Myalgic encephalomyelitis: International Consensus Criteria.

The label 'chronic fatigue syndrome' (CFS) has persisted for many years because of the lack of knowledge of the aetiological agents and the disease process. In view of more recent research and clinical experience that strongly point to widespread inflammation and multisystemic neuropathology, it is more appropriate and correct to use the term 'myalgic encephalomyelitis' (ME) because it indicates an underlying pathophysiology. It is also consistent with the neurological classification of ME in the World Health Organization's International Classification of Diseases (ICD G93.3). Consequently, an International Consensus Panel consisting of clinicians, researchers, teaching faculty and an independent patient advocate was formed with the purpose of developing criteria based on current knowledge. Thirteen countries and a wide range of specialties were represented. Collectively, members have approximately 400 years of both clinical and teaching experience, authored hundreds of peer-reviewed publications, diagnosed or treated approximately 50 000 patients with ME, and several members coauthored previous criteria. The expertise and experience of the panel members as well as PubMed and other medical sources were utilized in a progression of suggestions/drafts/reviews/revisions. The authors, free of any sponsoring organization, achieved 100% consensus through a Delphi-type process. The scope of this paper is limited to criteria of ME and their application. Accordingly, the criteria reflect the complex symptomatology. Operational notes enhance clarity and specificity by providing guidance in the expression and interpretation of symptoms. Clinical and research application guidelines promote optimal recognition of ME by primary physicians and other healthcare providers, improve the consistency of diagnoses in adult and paediatric patients internationally and facilitate clearer identification of patients for research studies.

Psoriasis and extra domain A fibronectin loops.

We have previously postulated that as well as T-helper (Th) 1 and Th17 cells, the transforming growth factor (TGF)-beta/fibronectin (FN)/alpha5beta1 pathway is central to psoriasis pathogenesis. EDA+ FN refers to an alternatively spliced isoform of FN with an additional domain known as extra domain A. EDA+ FN has two important properties pertinent to psoriasis lesions: it stimulates keratinocyte hyperproliferation, and, through stimulation of Toll-like receptor (TLR) 4, stimulates production of proinflammatory cytokines. EDA+ FN production induced by TGF-beta stimulation can be maintained in psoriasis lesions via two main feedback loops. Firstly, EDA+ FN stimulates proliferation of keratinocytes, which, in an autocrine fashion, will release more EDA+ FN. Secondly, EDA+ FN stimulates TLR4 expressed by antigen-presenting cells resulting in the production of proinflammatory cytokines such as tumour necrosis factor-alpha, interleukin (IL)-1, IL-6 and IL-12. The resultant promotion of cutaneous inflammation results in the recruitment of Th1 cells, which also produce EDA+ FN. We propose that these 'FN loops' contribute to the maintenance and progression of psoriatic lesions. Finally, although the association between psoriasis and heart/thrombotic disease remains unclear one plausible link may be the promotion of atherosclerosis and thrombotic heart disease by EDA+ FN.

Psoriasis and streptococci: the natural selection of psoriasis revisited.

We have previously postulated that surviving invasive streptococcal infections may have been a factor in psoriasis becoming a common skin disease in some parts of the world. Many of the candidate genes linked to psoriasis are associated with the acquired or innate immune system, which are also important in host defence to invasive streptococcal infections. High rates of positive streptococcal throat swabs among patients with chronic plaque psoriasis suggest that they are efficient at internalizing/carrying beta-haemolytic streptococci. Internalization of streptococci in the throat is dependent upon the transforming growth factor (TGF)-beta/fibronectin/alpha 5 beta 1 integrin pathway. The immune cell Th17 and its related cytokine network are important in mucosal defence, being very effective against extracellular microbes but having little effect on intracellular organisms. The TGF-beta/fibronectin/alpha 5 beta 1 integrin pathway and the Th17 cell network also appear to be operative in psoriasis, animal models of both TGF-beta and alpha 5 beta 1 cutaneous overexpression being associated with characteristic psoriasis lesions. We postulate that some of the genotypic/phenotypic changes in different immunological pathways in psoriasis, including the acquired T-cell response, the innate immune response, the TGF-beta/fibronectin/alpha 5 beta 1 integrin pathway and the Th17 cell system, confer protection against mortality during epidemics of invasive streptococcal infections, heightened efficiency in internalizing and allowing carriage of streptococci as well as predisposition to the development of psoriasis.

"Sleepiness" is serious in adolescence: two surveys of 3235 Canadian students.

BACKGROUND: Evidence is growing that sleep problems in adolescents are significant impediments to learning and negatively affect behaviour, attainment of social competence and quality of life. The objectives of the study were to determine the level of sleepiness among students in high school, to identify factors to explain it, and to determine the association between sleepiness and performance in both academic and extracurricular activities METHODS: A cross-sectional survey of 2201 high school students in the Hamilton Wentworth District School Board and the Near North District School Board in Ontario was conducted in 1998/9. A similar survey was done three years later involving 1034 students in the Grand Erie District School Board in the same Province. The Epworth Sleepiness Scale (ESS) was used to measure sleepiness and we also assessed the reliability of this tool for this population. Descriptive analysis of the cohort and information on various measures of performance and demographic data were included. Regression analysis, using the generalised estimating equation (GEE), was utilized to investigate factors associated with risk of sleepiness (ESS>10). RESULTS: Seventy per cent of the students had less than 8.5 hours weeknight sleep. Bedtime habits such as a consistent bedtime routine, staying up late or drinking caffeinated beverages before bed were statistically significantly associated with ESS, as were weeknight sleep quantity and gender. As ESS increased there was an increase in the proportion of students who felt their grades had dropped because of sleepiness, were late for school, were often extremely sleepy at school, and were involved in fewer extracurricular activities. These performance measures were statistically significantly associated with ESS. Twenty-three percent of the students felt their grades had dropped because of sleepiness. Most students (58-68%) reported that they were "really sleepy" between 8 and 10 A.M. CONCLUSION: Sleep deprivation and excessive daytime sleepiness were common in two samples of Ontario high school students and were associated with a decrease in academic achievement and extracurricular activity. There is a need to increase awareness of this problem in the education and health communities and to translate knowledge already available to strategies to address it.

HLA Cw*06 is not essential for streptococcal-induced psoriasis.

BACKGROUND: Streptococcal throat infections and HLA Cw6 (Cw*06) have been implicated in the pathogenesis of psoriasis, particularly in the guttate form. OBJECTIVES: To study 105 Irish patients with psoriasis to investigate the relationship between streptococcal infections and Cw*06. METHODS: The patients were divided into two groups: those with guttate psoriasis or guttate flare (guttate group, GG, n=64) and those with chronic plaque psoriasis (chronic plaque group, CPG, n=41). RESULTS: The incidence of Cw*06 was 86% in the GG and 73% in the CPG, which was not significantly different (P=0.1725) but the incidence in both groups was significantly higher than in an Irish control group (18%) (P<0.0001 vs. GG and P<0.0001 vs. CPG). Evidence for streptococcal infection was higher in the GG (56%) than in the CPG (32%) (P=0.0231). Of those patients with evidence of streptococcal infection, 30 of 36 GG (83%) and nine of 13 CPG (69%) patients possessed the Cw*06 genotype. CONCLUSIONS: Thus, not all patients with streptococcal-related psoriasis carry Cw*06. The role of Cw*06 in psoriasis, if any, has yet to be determined.

Peptidoglycan and peptidoglycan-specific Th1 cells in psoriatic skin lesions.

We have previously demonstrated, in psoriatic skin lesions, the presence of a subset of dermal CD4+ T cells that produce interferon-gamma (IFN-gamma) in response to a mixture of cell wall proteins extracted from group A streptococci. However, the identity of the antigen(s) involved is unknown. To investigate the hypothesis that peptidoglycan (PG), the major constituent of the streptococcal cell wall, acts as a T cell activator in psoriasis, we performed in situ analysis to detect antigen-presenting cells containing PG in lesional versus non-lesional skin, and determined proliferation and IFN-gamma responses of lesional skin T cells. Increased numbers of PG-containing cells were detected in the dermal papillae and cellular infiltrates of guttate and chronic plaque skin lesions compared with normal and non-lesional psoriatic skin. A varying proportion of these were CD68+ macrophages, but the remaining cells did not double stain for either Langerhans' or dendritic cell markers. Psoriatic dermal streptococcal-specific CD4+ T cell lines proliferated and produced IFN-gamma in a self HLA-DR allele-restricted manner in response to streptococcal PG, excluding mitogenic or superantigenic stimulation, but were unresponsive to staphylococcal PG. Similarly, psoriatic staphylococcus-specific T cell lines recognized staphylococcal, but not streptococcal, PG by IFN-gamma production. The presence of PG-containing macrophages in close association with PG-specific CD4+ T cells in lesional skin suggests that PG may be responsible, at least in part, for T cell activation in psoriasis.

Selective response of dermal Th-1 cells to 20-50 kDa streptococcal cell-wall proteins in chronic plaque psoriasis.

We have recently described a dermal Th-1 subset in skin lesions of psoriasis which recognizes cell-wall extract isolated from group A streptococci (GAS). As a first step in the identification of the streptococcal proteins involved, dermal T-cell lines (TCL) cultured from the lesional skin of 12 human leucocyte antigen (HLA)-typed psoriasis patients were stimulated with GAS cell-wall extract and 14 fractions (MWt approximately 20-100 kDa) separated from the cell-wall extract by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and electroelution, stained for intracellular interferon-gamma(IFN-gamma) expression and analysed by flow cytometry. All the TCL responded to GAS cell-wall extract to varying extents (3.5-27.6% IFN-gamma+). This response was consistently directed against 20-50 kDa cell-wall fractions and inhibited by anti-HLA-DR antibody. TCL with higher responses to GAS cell-wall extract recognized a larger number of fractions within this range than the lower responder TCL. No difference between the level and pattern of response to the fractions was observed for TCL from HLA-DR7+ (n = 6) and HLA-DR7- (n = 6) individuals. This preliminary study has shown a selective response to lower MWt proteins expressed on GAS cell wall by skin Th-1 cells in psoriasis. Further studies are required to identify the proteins involved.

Normal keratinocytes express Toll-like receptors (TLRs) 1, 2 and 5: modulation of TLR expression in chronic plaque psoriasis.

BACKGROUND: Toll-like receptors (TLRs) are part of the innate immune system involved in the response to microbial pathogens. TLR2 recognizes various ligands expressed by Gram-positive bacteria, while TLR3, TLR4 and TLR5 are specific for double-stranded RNA, Gram-negative lipopolysaccharides and bacterial flagellin, respectively. OBJECTIVES: To determine, firstly, whether epidermal keratinocytes of normal skin express TLRs and, secondly, whether modulation of TLR expression occurs in psoriasis, an inflammatory skin disease associated with certain microorganisms such as streptococci, staphylococci and yeasts. METHODS: Eight samples of normal, and 15 samples of lesional and nonlesional psoriatic skin were stained with polyclonal antibodies specific for TLR1-5 using an avidin-biotin-peroxidase technique. RESULTS: Epidermal keratinocytes in normal skin constitutively expressed TLR1, TLR2 and TLR5, while TLR3 and TLR4 were, in most cases, barely detectable. Cytoplasmic TLR1 and TLR2 were expressed throughout the epidermis, with higher staining of the latter on basal keratinocytes, while TLR5 expression was concentrated in the basal layer. In contrast, in lesional epidermis from patients with psoriasis, TLR2 was more highly expressed on the keratinocytes of the upper epidermis than on the basal layer, while TLR5 was downregulated in basal keratinocytes compared with corresponding nonlesional psoriatic epidermis. In addition, nuclear TLR1 staining was observed in the upper layers of both nonlesional and lesional psoriatic epidermis, but not in that of normal skin. CONCLUSIONS: These findings suggest that TLRs expressed by epidermal keratinocytes constitute part of the innate immune system of the skin. The relevance of altered keratinocyte TLR expression in psoriasis remains to be determined.

Epidermal CD8+ T cells reactive with group A streptococcal antigens in chronic plaque psoriasis.

Chronic plaque psoriasis is a T cell mediated disease associated with group A streptococci (GAS). We have previously shown the presence of a psoriasis-specific dermal Th1 subset that recognizes GAS antigens. To assess whether GAS-reactive T cells are also present in lesional epidermis, fresh cell suspensions or T cell lines isolated from lesional epidermis of 33 psoriasis patients were stained for intracellular interferon-gamma after stimulation with GAS antigens. The patients were typed by PCR for HLA-DR7 and HLA-Cw6 expression. A subset of GAS-reactive CD8+ T cells (2.4% +/- 2.4) was found in 14/21 (67%) fresh cell suspensions. A smaller subset of GAS-reactive CD4+ T cells (0.9% +/- 0.9) was found in 13/21 (62%) fresh cell suspensions, which was expanded in the T cell lines. There was a significant inverse correlation between the proportions of GAS-reactive CD4+ and CD8+ T cells in the fresh suspensions (r = -0.48, P = 0.0277). The presence of GAS-reactive CD4+ or CD8+ T cells did not correlate with HLA-DR7 or HLA-Cw6 expression, respectively. This study has demonstrated GAS-reactive CD8+, and to a lesser extent CD4+, T cell subsets in psoriatic epidermis and provides further evidence that GAS antigens may play a role in the pathogenesis of chronic plaque psoriasis.

Rescue therapy with inhaled nitric oxide in critically ill patients with severe hypoxemic respiratory failure (Brief report).

PURPOSE: To evaluate the efficacy of inhaled nitric oxide (iNO) on oxygenation, shunt, and pulmonary vascular resistance index (PVRI) in severely hypoxemic, ventilated patients. METHODS: In a two-period double-blind crossover design, 14 critically ill, hypoxemic, ventilated patients were randomized to receive iNO 10 ppm in 100% oxygen or no iNO in 100% oxygen for 30 min followed by a 30-min washout period and then crossed over to the other intervention. Responders to iNO then received iNO, which was increased from 5 ppm to 25 ppm in 5 ppm increments. Severity of illness scores and cardiorespiratory variables were measured. RESULTS: Nitric oxide decreased shunt (P=0.002) and PVRI (P=0.033) and increased oxygenation (P=0.011) although the latter two were not statistically significant after adjustment for multiple comparisons. Treatment by period interactions were observed. CONCLUSION: Our findings suggest that iNO improves oxygenation to a clinically significant extent in critically ill patients who are severely hypoxemic.

Skin T cell proliferative response to M protein and other cell wall and membrane proteins of group A streptococci in chronic plaque psoriasis.

To determine and compare the T cell response to M protein and other group A streptococcal (GAS) antigens, T cell lines (TCL) were cultured from the lesional skin of 33 psoriatic patients and 17 disease controls. GAS-reactive skin TCL were tested in proliferation assays with recombinant M6 protein, and extracts of cell wall and membrane from type M6 GAS and its corresponding M gene deletion mutant. Initially, GAS-reactive skin TCL were obtained from 16 of 25 (64%) psoriasis, and from seven of 17 (41%) control patients. Eleven psoriatic and four control GAS-reactive TCL proliferated to M6 cell wall extract, whereas all the TCL from both groups responded to the extract of M6 membrane proteins. This difference in response to the two extracts was significant for both groups of patients (psoriasis, P = 0.0335, controls, P = 0.0156). GAS-reactive TCL from a further eight psoriasis patients showed no difference in response to cell wall extract from M6 GAS (containing the M protein minus its C-terminus) compared to that of its corresponding M gene deletion mutant. Furthermore, GAS-reactive TCL did not proliferate to recombinant M6 protein. However, a small, but significant reduction in proliferation by the eight psoriatic GAS-reactive TCL to the M-negative (lacking the M protein C-terminus) compared to M6-positive membrane extract was observed (P = 0.01). These findings suggest that GAS-reactive T cells in skin lesions of chronic plaque psoriasis proliferate to streptococcal membrane and, to a lesser extent, cell wall proteins. However, psoriatic skin T cells do not recognize cell wall M protein.

Non-M protein(s) on the cell wall and membrane of group A streptococci induce(s) IFN-gamma production by dermal CD4+ T cells in psoriasis.

Recently we have demonstrated that a disease-specific subpopulation of CD4+ T cells isolated from skin lesions of chronic plaque psoriasis produces interferon-gamma in response to group A streptococcal (GAS) antigens. To determine if these T cells recognize M or non-M protein, extracts from cell wall of type M6 GAS (M6W) and its isogenic M gene deletion mutant (M-W), M6 membrane extract (M6M) and recombinant M6 protein (rM6) were used to stimulate GAS-reactive T-cell lines from nine patients with chronic plaque psoriasis. T-cell lines were incubated with or without streptococcal extracts for 18 h in the presence of a transport inhibitor, stained for surface CD4 and intracellular cytokine expression, and analysed by flow cytometry. Variable numbers (0.2-34%) of CD4+ T cells produced interferon-gamma, in all but one of the T-cell lines tested, in response to M6W, M-W and M6M extracts. No significant difference between the response to M6W and M-W extracts was detected. In addition, rM6 protein failed to increase CD4+/interferon-gamma+ T-cell numbers in seven of nine T-cell lines compared to medium alone. For the group, there was a highly significant correlation between the responses to the three extracts (M6W vs M-W, P = 0.0005; M6W vs M6M, P = 0.0003; M-W vs M6M, P = 0.0001). Low or minimal numbers of interleukin-4- and interleukin-10-producing CD4+ T cells were occasionally induced. These findings suggest that a subpopulation of CD4+ T cells isolated from skin lesions of chronic plaque psoriasis patients produces interferon-gamma in response to non-M protein(s) present on the cell wall and membrane of GAS.

Epidermal CD8+ T cells in chronic plaque psoriasis are Tc1 cells producing heterogeneous levels of interferon-gamma.

The majority of epidermal CD8+ T cells in chronic plaque psoriasis are activated Tc1 cells producing interferon-gamma and no interleukin-4, a small proportion of which express NK-T receptors. To quantitate their level of cytokine production and characterize them further, CD8+ T cells were isolated from epidermal cell suspensions of lesional biopsies from 24 patients with chronic plaque psoriasis. T-cell lines (TCL) were established by culture of CD8+ T cells with feeders and IL-2 for 11 days and expansion with PHA. Ten TCL were stained for surface markers; 6 were cloned with PHA by limiting dilution. Interferon-gamma, interleukin-4 and interleukin-10 production was measured by ELISA after PMA/anti-CD3 activation of 15 TCL and 39 CD8+ T-cell clones. The 10 TCL stained were CD8alphabeta+ (93.3%), T-cell receptor-alphabeta+ (99.5%), costimulatory molecule CD28+ (90.1%), with a small CD8alphaalpha+ population (2.3%). No NK-T-cell receptor CD158a or CD158b expression was detected, whilst CD94 was expressed on 6.2% of cells in 6/9 TCL. All the TCL and 37/39 CD8+ T-cell clones produced interferon-gamma but no or minimal interleukin-4 or interleukin-10. The TCL produced a wide range of interferon-gamma levels (138 to 15,020 pg/ml). Clones from 3 patients showed low levels (60 to 1,410 pg/ml), from 2 patients high levels (6,105 to 43,040 pg/ml) and from 1 patient a wide range (405 to 36,010 pg/ml) of interferon-gamma production. Thus epidermal CD8+ Tc1 cells in chronic plaque psoriasis produce highly heterogeneous levels of interferon-gamma, which may reflect clinical diversity.

Stronger proliferative response to membrane versus cell-wall Streptococcal proteins by peripheral blood T cells in chronic plaque psoriasis.

Proliferative responses of peripheral blood mononuclear cells (PBMC) to group A streptococcal (GAS) antigens have been studied in 24 patients with psoriasis and 15 disease controls. Extracts of cell wall (including M protein) from types M4 and M12 GAS, recombinant M6 protein, and both cell-wall and cell-membrane extracts from type M6 (M6+) GAS and its corresponding M gene deletion mutant (M6-) were tested. PBMC from psoriatic patients proliferated more strongly to cell-wall extracts from M12 versus M4 (P = 0.0348), and to M6+ versus M6- (P = 0.0019) GAS with, in most cases, moderate proliferation to recombinant M6 protein. The psoriatic response to M12 cell wall was significantly increased compared to the controls (P = 0.0032). In psoriatics, M6+ membrane extracts induced a markedly greater proliferation than those of cell wall (P = 0.0002); responses to M6+ (P = 0.0039) and M6- (P = 0.0114) membrane extracts were higher than those of the control PBMC. Both groups showed a decreased response to the M6- versus M6+ membrane extracts (P = 0.0030; P = 0.0181, respectively). This study has demonstrated that patients with psoriasis have a heightened circulating T-cell response to cell wall M protein and to non-M proteins present on the cell wall and membrane of GAS.