Inter-species gene flow drives ongoing evolution of Streptococcus pyogenes and Streptococcus dysgalactiae subsp. equisimilis.

Streptococcus dysgalactiae subsp. equisimilis (SDSE) is an emerging cause of human infection with invasive disease incidence and clinical manifestations comparable to the closely related species, Streptococcus pyogenes. Through systematic genomic analyses of 501 disseminated SDSE strains, we demonstrate extensive overlap between the genomes of SDSE and S. pyogenes. More than 75% of core genes are shared between the two species with one third demonstrating evidence of cross-species recombination. Twenty-five percent of mobile genetic element (MGE) clusters and 16 of 55 SDSE MGE insertion regions were shared across species. Assessing potential cross-protection from leading S. pyogenes vaccine candidates on SDSE, 12/34 preclinical vaccine antigen genes were shown to be present in >99% of isolates of both species. Relevant to possible vaccine evasion, six vaccine candidate genes demonstrated evidence of inter-species recombination. These findings demonstrate previously unappreciated levels of genomic overlap between these closely related pathogens with implications for streptococcal pathobiology, disease surveillance and prevention.

Characterization of an experimental model to determine streptococcal M protein-induced autoimmune cardiac and neurobehavioral abnormalities.

Group A streptococcal (GAS) infection is associated with a spectrum of autoimmune diseases including acute rheumatic fever/rheumatic heart disease (ARF/RHD) and neurobehavioral abnormalities. Antibodies against GAS M proteins cross-react with host tissue proteins in the heart and brain leading to the symptomatology observed in ARF/RHD. As throat carriage of Streptococcus dysgalactiae subspecies equisimilis (SDSE) has been reported to be relatively high in some ARF/RHD endemic regions compared with GAS, and both SDSE and GAS express coiled-coil surface protein called M protein, we hypothesized that streptococci other than GAS can also associated with ARF/RHD and neurobehavioral abnormalities. Neurobehavioral assessments and electrocardiography were performed on Lewis rats before and after exposure to recombinant GAS and SDSE M proteins. Histological assessments were performed to confirm inflammatory changes in cardiac and neuronal tissues. ELISA and Western blot analysis were performed to determine the cross-reactivity of antibodies with host connective, cardiac and neuronal tissue proteins. Lewis rats injected with M proteins either from GAS or SDSE developed significant cardiac functional and neurobehavioral abnormalities in comparison to control rats injected with phosphate-buffered saline. Antibodies against GAS and SDSE M proteins cross-reacted with cardiac, connective and neuronal proteins. Serum from rats injected with streptococcal antigens showed higher immunoglobulin G binding to the striatum and cortex of the brain. Cardiac and neurobehavioral abnormalities observed in our experimental model were comparable to the cardinal symptoms observed in patients with ARF/RHD. Here for the first time, we demonstrate in an experimental model that M proteins from different streptococcal species could initiate and drive the autoimmune-mediated cardiac tissue damage and neurobehavioral abnormalities.

Group A streptococcal antigen exposed rat model to investigate neurobehavioral and cardiac complications associated with post-streptococcal autoimmune sequelae.

Background: The neuropsychiatric disorders due to post-streptococcal autoimmune complications such as Sydenham's chorea (SC) are associated with acute rheumatic fever and rheumatic heart disease (ARF/RHD). An animal model that exhibits characteristics of both cardiac and neurobehavioral defects in ARF/RHD would be an important adjunct for future studies. Since age, gender, strain differences, and genotypes impact on the development of autoimmunity, we investigated the behavior of male and female Wistar and Lewis rat strains in two age cohorts (<6 weeks and >12 weeks) under normal husbandry conditions and following exposure to group A streptococcus (GAS). Methods: Standard behavioral assessments were performed to determine the impairments in fine motor control (food manipulation test), gait and balance (beam walking test), and obsessive-compulsive behavior (grooming and marble burying tests). Furthermore, electrocardiography, histology, and behavioral assessments were performed on male and female Lewis rats injected with GAS antigens. Results: For control Lewis rats there were no significant age and gender dependent differences in marble burying, food manipulation, beam walking and grooming behaviors. In contrast significant age-dependent differences were observed in Wistar rats in all the behavioral tests except for food manipulation. Therefore, Lewis rats were selected for further experiments to determine the effect of GAS. After exposure to GAS, Lewis rats demonstrated neurobehavioral abnormalities and cardiac pathology akin to SC and ARF/RHD, respectively. Conclusion: We have characterised a new model that provides longitudinal stability of age-dependent behavior, to simultaneously investigate both neurobehavioral and cardiac abnormalities associated with post-streptococcal complications.

Requirements for a Robust Animal Model to Investigate the Disease Mechanism of Autoimmune Complications Associated With ARF/RHD.

The pathogenesis of Acute Rheumatic Fever/Rheumatic Heart Disease (ARF/RHD) and associated neurobehavioral complications including Sydenham's chorea (SC) is complex. Disease complications triggered by Group A streptococcal (GAS) infection are confined to human and determining the early events leading to pathology requires a robust animal model that reflects the hallmark features of the disease. However, modeling these conditions in a laboratory animal, of a uniquely human disease is challenging. Animal models including cattle, sheep, pig, dog, cat, guinea pigs rats and mice have been used extensively to dissect molecular mechanisms of the autoimmune inflammatory responses in ARF/RHD. Despite the characteristic limitations of some animal models, several rodent models have significantly contributed to better understanding of the fundamental mechanisms underpinning features of ARF/RHD. In the Lewis rat autoimmune valvulitis model the development of myocarditis and valvulitis with the infiltration of mononuclear cells along with generation of antibodies that cross-react with cardiac tissue proteins following exposure to GAS antigens were found to be similar to ARF/RHD. We have recently shown that Lewis rats injected with recombinant GAS antigens simultaneously developed cardiac and neurobehavioral changes. Since ARF/RHD is multifactorial in origin, an animal model which exhibit the characteristics of several of the cardinal diagnostic criteria observed in ARF/RHD, would be advantageous to determine the early immune responses to facilitate biomarker discovery as well as provide a suitable model to evaluate treatment options, safety and efficacy of vaccine candidates. This review focuses on some of the common small animals and their advantages and limitations.

In Search of the Holy Grail: A Specific Diagnostic Test for Rheumatic Fever.

Current diagnosis of Acute Rheumatic Fever and Rheumatic Heart Disease (ARF/RHD) relies on a battery of clinical observations aided by technologically advanced diagnostic tools and non-specific laboratory tests. The laboratory-based assays fall into two categories: those that (1) detect "evidence of preceding streptococcal infections" (ASOT, anti-DNAse B, isolation of the Group A Streptococcus from a throat swab) and (2) those that detect an ongoing inflammatory process (ESR and CRP). These laboratory tests are positive during any streptococcal infection and are non-specific for the diagnosis of ARF/RHD. Over the last few decades, we have accumulated considerable knowledge about streptococcal biology and the immunopathological mechanisms that contribute to the development, progression and exacerbation of ARF/RHD. Although our knowledge is incomplete and many more years will be devoted to understanding the exact molecular and cellular mechanisms involved in the spectrum of clinical manifestations of ARF/RHD, in this commentary we contend that there is sufficient understanding of the disease process that using currently available technologies it is possible to identify pathogen associated peptides and develop a specific test for ARF/RHD. It is our view that with collaboration and sharing of well-characterised serial blood samples from patients with ARF/RHD from different regions, antibody array technology and/or T-cell tetramers could be used to identify streptococcal peptides specific to ARF/RHD. The availability of an appropriate animal model for this uniquely human disease can further facilitate the determination as to whether these peptides are pathognomonic. Identification of such peptides will also facilitate testing of potential anti-streptococcal vaccines for safety and avoid potential candidates that may pre-dispose potential vaccine recipients to adverse outcomes. Such peptides can also be readily incorporated into a universally affordable point of care device for both primary and tertiary care.

A longitudinal study of antibody responses to selected host antigens in rheumatic fever and rheumatic heart disease.

Introduction. Group A streptococci can trigger autoimmune responses that lead to acute rheumatic fever (ARF) and rheumatic heart disease (RHD).Gap Statement. Some autoantibodies generated in ARF/RHD target antigens in the S2 subfragment region of cardiac myosin. However, little is known about the kinetics of these antibodies during the disease process.Aim. To determine the antibody responses over time in patients and healthy controls against host tissue proteins - cardiac myosin and peptides from its S2 subfragment, tropomyosin, laminin and keratin.Methodology. We used enzyme-linked immunosorbent assays (ELISA) to determine antibody responses in: (1) healthy controls; (2) patients with streptococcal pharyngitis; (3) patients with ARF with carditis and (4) patients with RHD on penicillin prophylaxis.Results. We observed significantly higher antibody responses against extracellular proteins - laminin and keratin in pharyngitis group, patients with ARF and patients with RHD when compared to healthy controls. The antibody responses against intracellular proteins - cardiac myosin and tropomyosin were elevated only in the group of patients with ARF with active carditis. While the reactivity to S2 peptides S2-1-3, 8-11, 14, 16-18, 21-22 and 32 was higher in patients with ARF, the reactivity in the RHD group was high only against S2-1, 9, 11, 12 when compared to healthy controls. The reactivity against S2 peptides reduced as the disease condition stabilized in the ARF group whereas the reactivity remained unaltered in the RHD group. By contrast antibodies against laminin and keratin persisted in patients with RHD.Conclusion. Our findings of antibody responses against host proteins support the multistep hypothesis in the development of rheumatic carditis. The differential kinetics of serum antibody responses against S2 peptides may have potential use as markers of ongoing cardiac damage that can be used to monitor patients with ARF/RHD.

First Description of the Composition and the Functional Capabilities of the Skin Microbial Community Accompanying Severe Scabies Infestation in Humans.

Epidemiological studies link Sarcoptes scabiei infection and impetigo. Scabies mites can promote Streptococcus pyogenes (Group A Streptococcus) and Staphylococcus aureus infections by breaching the skin barrier and excreting molecules that inhibit host innate immune responses. However, little is known about the composition and the function of the scabies-associated microbiota. Here, high-throughput whole-metagenome sequencing was used to explore the scabies-associated microbiome. Scabies mites including their immediate microenvironments were isolated from two patients with severe scabies in Northern Australia. Two ~45-50 million paired-end reads Illumina libraries were generated of which ~2 (5.1%) and 0.7 million (1.3%) microbial reads were filtered out by mapping to human (hg19) and mite draft genomes. Taxonomic profiling revealed a microbial community dominated by the phylum Firmicutes (A: 79% and B: 59%) and genera that comprise Streptococcus, Staphylococcus, Acinetobacter, and Corynebacterium. Assembly of the metagenome reads resulted in genome bins representing reference genomes of Acinetobacter baumannii, Streptococcus dysgalactiae (Group C/G), Proteus mirablis and Staphylococcus aureus. The contigs contained genes relevant to pathogenicity and antibiotics resistance. Confocal microscopy of a patient skin sample confirmed A. baumannii, Streptococci and S. aureus in scabies mite gut and faeces and the surrounding skin. The study provides fundamental evidence for the association of opportunistic pathogens with scabies infection.

High-throughput metagenome analysis of the Sarcoptes scabiei internal microbiota and in-situ identification of intestinal Streptomyces sp.

Multiple parasitic arthropods of medical importance depend on symbiotic bacteria. While the link between scabies and secondary bacterial infections causing post infective complications of Group A streptococcal and staphylococcal pyoderma is increasingly recognized, very little is known about the microbiota of Sarcoptes scabiei. Here we analyze adult female mite and egg metagenome datasets. The majority of adult mite bacterial reads matched with Enterobacteriaceae (phylum Proteobacteria), followed by Corynebacteriaceae (phylum Actinobacteria). Klebsiella was the most dominant genus (78%) and Corynebacterium constituted 9% of the assigned sequences. Scabies mite eggs had a more diverse microbial composition with sequences from Proteobacteria being the most dominant (75%), while Actinobacteria, Bacteroidetes and Firmicutes accounted for 23% of the egg microbiome sequences. DNA sequences of a potential endosymbiont, namely Streptomyces, were identified in the metagenome sequence data of both life stages. The presence of Streptomyces was confirmed by conventional PCR. Digital droplet PCR indicated higher Streptomyces numbers in adult mites compared to eggs. Streptomyces were localized histologically in the scabies mite gut and faecal pellets by Fluorescent In Situ Hybridization (FISH). Streptomyces may have essential symbiotic roles in the scabies parasite intestinal system requiring further investigation.

Group G Streptococcus Induces an Autoimmune Carditis Mediated by Interleukin 17A and Interferon γ in the Lewis Rat Model of Rheumatic Heart Disease.

Acute rheumatic fever and rheumatic heart disease (ARF/RHD) have long been described as autoimmune sequelae of Streptococcus pyogenes or group A streptococcal (GAS) infection. Both antibody and T-cell responses against immunodominant GAS virulence factors, including M protein, cross-react with host tissue proteins, triggering an inflammatory response leading to permanent heart damage. However, in some ARF/RHD-endemic regions, throat carriage of GAS is low. Because Streptococcus dysgalactiae subspecies equisimilis organisms, also known as ß-hemolytic group C streptococci and group G streptococci (GGS), also express M protein, we postulated that streptococci other than GAS may have the potential to initiate or exacerbate ARF/RHD. Using a model initially developed to investigate the uniquely human disease of ARF/RHD, we have discovered that GGS causes interleukin 17A/interferon γ-induced myocarditis and valvulitis, hallmarks of ARF/RHD. Remarkably the histological, immunological, and functional changes in the hearts of rats exposed to GGS are identical to those exposed to GAS. Furthermore, antibody cross-reactivity to cardiac myosin was comparable in both GGS- and GAS-exposed animals, providing additional evidence that GGS can induce and/or exacerbate ARF/RHD.

Complement inhibition by Sarcoptes scabiei protects Streptococcus pyogenes - An in vitro study to unravel the molecular mechanisms behind the poorly understood predilection of S. pyogenes to infect mite-induced skin lesions.

BACKGROUND: On a global scale scabies is one of the most common dermatological conditions, imposing a considerable economic burden on individuals, communities and health systems. There is substantial epidemiological evidence that in tropical regions scabies is often causing pyoderma and subsequently serious illness due to invasion by opportunistic bacteria. The health burden due to complicated scabies causing cellulitis, bacteraemia and sepsis, heart and kidney diseases in resource-poor communities is extreme. Co-infections of group A streptococcus (GAS) and scabies mites is a common phenomenon in the tropics. Both pathogens produce multiple complement inhibitors to overcome the host innate defence. We investigated the relative role of classical (CP), lectin (LP) and alternative pathways (AP) towards a pyodermic GAS isolate 88/30 in the presence of a scabies mite complement inhibitor, SMSB4. METHODOLOGY/PRINCIPAL FINDINGS: Opsonophagocytosis assays in fresh blood showed baseline immunity towards GAS. The role of innate immunity was investigated by deposition of the first complement components of each pathway, specifically C1q, FB and MBL from normal human serum on GAS. C1q deposition was the highest followed by FB deposition while MBL deposition was undetectable, suggesting that CP and AP may be mainly activated by GAS. We confirmed this result using sera depleted of either C1q or FB, and serum deficient in MBL. Recombinant SMSB4 was produced and purified from Pichia pastoris. SMSB4 reduced the baseline immunity against GAS by decreasing the formation of CP- and AP-C3 convertases, subsequently affecting opsonisation and the release of anaphylatoxin. CONCLUSIONS/SIGNIFICANCE: Our results indicate that the complement-inhibitory function of SMSB4 promotes the survival of GAS in vitro and inferably in the microenvironment of the mite-infested skin. Understanding the tripartite interactions between host, parasite and microbial pathogens at a molecular level may serve as a basis to develop improved intervention strategies targeting scabies and associated bacterial infections.

Predicted Coverage and Immuno-Safety of a Recombinant C-Repeat Region Based Streptococcus pyogenes Vaccine Candidate.

The C-terminal region of the M-protein of Streptococcus pyogenes is a major target for vaccine development. The major feature is the C-repeat region, consisting of 35-42 amino acid repeat units that display high but not perfect identity. SV1 is a S. pyogenes vaccine candidate that incorporates five 14mer amino acid sequences (called J14i variants) from differing C-repeat units in a single recombinant construct. Here we show that the J14i variants chosen for inclusion in SV1 are the most common variants in a dataset of 176 unique M-proteins. Murine antibodies raised against SV1 were shown to bind to each of the J14i variants present in SV1, as well as variants not present in the vaccine. Antibodies raised to the individual J14i variants were also shown to bind to multiple but different combinations of J14i variants, supporting the underlying rationale for the design of SV1. A Lewis Rat Model of valvulitis was then used to assess the capacity of SV1 to induce deleterious immune response associated with rheumatic heart disease. In this model, both SV1 and the M5 positive control protein were immunogenic. Neither of these antibodies were cross-reactive with cardiac myosin or collagen. Splenic T cells from SV1/CFA and SV1/alum immunized rats did not proliferate in response to cardiac myosin or collagen. Subsequent histological examination of heart tissue showed that 4 of 5 mice from the M5/CFA group had valvulitis and inflammatory cell infiltration into valvular tissue, whereas mice immunised with SV1/CFA, SV1/alum showed no sign of valvulitis. These results suggest that SV1 is a safe vaccine candidate that will elicit antibodies that recognise the vast majority of circulating GAS M-types.

Seropositivity for Antibodies to DRS-G, a Virulence Factor from Streptococcus dysgalactiae subsp. equisimilis, Is an Independent Risk Factor for Poststreptococcus Glomerulonephritis and Chronic Kidney Disease in Mumbai, India.

The disease spectrum caused by Streptococcus dysgalactiae subsp. equisimilis resembles that of S. pyogenes (group A streptococcus [GAS]). These two bacterial species are closely related and possess many common virulence characteristics. While some GAS strains express virulence factors called streptococcal inhibitor of complement (SIC) and distantly related to SIC (DRS), some S. dysgalactiae subsp. equisimilis isolates express an orthologue of DRS, which is referred to as DRS-G. We reported previously that seropositivity for either anti-SIC or anti-DRS antibodies (Abs) is associated with poststreptococcal glomerulonephritis (PSGN). However, only seropositivity for anti-SIC Abs is associated with chronic kidney disease (CKD). We now extend the study to test whether seropositivity for anti-DRS-G Abs is also associated with these renal diseases. Stored serum samples collected for our previous study were tested by an enzyme-linked immunosorbent assay (ELISA) for Abs to DRS-G. The samples represented sera from 100 CKD adult patients, 70 adult end-stage renal disease (ESRD) patients, 25 PSGN pediatric patients, and corresponding age-matched control subjects. The proportion of PSGN, CKD, and ESRD patients who showed seroreaction to anti-DRS-G Abs was significantly higher than that of the corresponding age-matched controls, who in general exhibited seropositivity rates commensurate with the isolation rate of drsG-positive S. dysgalactiae subsp. equisimilis in the community during this study period. Since higher rates of seropositivity for anti-DRS-G Abs in the renal disease categories are resultant of previous infections with DRS-G-positive S. dysgalactiae subsp. equisimilis strains, we conclude the seropositivity is an additional risk factor for these renal diseases. In this regard, anti-DRS-G Abs have attributes similar to those of the anti-SIC Abs.

Antibodies to group A streptococcal virulence factors, SIC and DRS, increase predilection to GAS pyoderma.

BACKGROUND: Streptococcus pyogenes (group A streptococcus; GAS) is an etiological agent for pharyngitis, pyoderma, and invasive infections in humans. Pharyngitis and pyoderma may lead to serious immune sequelae such as rheumatic heart disease and post-streptococcal glomerulonephritis (PSGN). Streptococcal Inhibitor of Complement (SIC) and its orthologue, distantly related to SIC (DRS), are virulence factors expressed by only four of more than 100 M types of GAS. These four types (M1, M57, M12 and M55) are among the M types, which are associated with PSGN. In several populations PSGN has been shown to be a risk factor for chronic kidney disease (CKD) and end-stage renal disease (ESRD). Previous studies showed SIC or DRS antibody-prevalence was associated with PSGN, and seroprevalence of SIC antibodies is significantly high among CKD and ESRD patients in Mumbai. METHODS: Streptococcal isolates recovered from GAS pyoderma cases were typed. Seropositivity for SIC and DRS antibodies in subjects with pyoderma, PSGN pediatric cases, age matched healthy controls and non-GAS pyoderma cases were determined. RESULTS: We confirm in this study an association between seroprevalence to SIC and DRS antibodies, and PSGN in Mumbai population despite low point prevalence of M1, M12, M55 and M57. In addition we extended the study to GAS-pyoderma and non-GAS pyoderma cases. To our surprise, we found a positive association between the seroprevalence to SIC and DRS antibodies, and GAS-pyoderma owing to infection with diverse M types. The mechanism of increased predisposition to pyoderma owing to infection by diverse GAS among SIC or DRS antibody-positive population is not clear. Nonetheless, our findings could be explained by a phenomenon akin to antibody-dependent enhancement (ADE). CONCLUSIONS: This is the first report showing a small number of GAS M types conferring predisposition to pyoderma by diverse types. Implications of this ADE-like phenomenon are discussed in the light of evolutionary advantage to GAS, vaccine design and control of renal diseases.

Conjugative transfer of ICESde3396 between three ß-hemolytic streptococcal species.

BACKGROUND: Integrative conjugative elements (ICEs) are mobile genetic elements (MGEs) that possess all genes necessary for excision, transfer and integration into recipient genome. They also carry accessory genes that impart new phenotypic features to recipient strains. ICEs therefore play an important role in genomic plasticity and population structure. We previously characterised ICESde3396, the first ICE identified in the ß-hemolytic Streptococcus dysgalactiae subsp equisimilis (SDSE) and demonstrated its transfer to single isolates of Streptococcus pyogenes (group A streptococcus, GAS) and Streptococcus agalactiae (group B streptococcus, GBS). While molecular studies found the ICE in multiple SDSE and GBS isolates, it was absent in all GAS isolates examined. RESULTS: Here we demonstrate that ICESde3396:km is transferable from SDSE to multiple SDSE, GAS and GBS isolates. However not all strains of these species were successful recipients under the same growth conditions. To address the role that host factors may have in conjugation we also undertook conjugation experiments in the presence of A549 epithelial cells and DMEM. While Horizontal Gene Transfer (HGT) occurred, conjugation efficiencies were no greater than when similar experiments were conducted in DMEM. Additionally transfer to GAS NS235 was successful in the presence of DMEM but not in Todd Hewitt Broth suggesting that nutritional factors may also influence HGT. The GAS and GBS transconjugants produced in this study are also able to act as donors of the ICE. CONCLUSION: We conclude that ICEs are major sources of interspecies HGT between ß-hemolytic streptococci, and by introducing accessory genes imparting novel phenotypic characteristics, have the potential to alter the population structure of these species.

A systematic and functional classification of Streptococcus pyogenes that serves as a new tool for molecular typing and vaccine development.

Streptococcus pyogenes ranks among the main causes of mortality from bacterial infections worldwide. Currently there is no vaccine to prevent diseases such as rheumatic heart disease and invasive streptococcal infection. The streptococcal M protein that is used as the substrate for epidemiological typing is both a virulence factor and a vaccine antigen. Over 220 variants of this protein have been described, making comparisons between proteins difficult, and hindering M protein-based vaccine development. A functional classification based on 48 emm-clusters containing closely related M proteins that share binding and structural properties is proposed. The need for a paradigm shift from type-specific immunity against S. pyogenes to emm-cluster based immunity for this bacterium should be further investigated. Implementation of this emm-cluster-based system as a standard typing scheme for S. pyogenes will facilitate the design of future studies of M protein function, streptococcal virulence, epidemiological surveillance, and vaccine development.

DrsG from Streptococcus dysgalactiae subsp. equisimilis inhibits the antimicrobial peptide LL-37.

SIC and DRS are related proteins present in only 4 of the >200 Streptococcus pyogenes emm types. These proteins inhibit complement-mediated lysis and/or the activity of certain antimicrobial peptides (AMPs). A gene encoding a homologue of these proteins, herein called DrsG, has been identified in the related bacterium Streptococcus dysgalactiae subsp. equisimilis. Here we show that geographically dispersed isolates representing 14 of 50 emm types examined possess variants of drsG. However, not all isolates within the drsG-positive emm types possess the gene. Sequence comparisons also revealed a high degree of conservation in different S. dysgalactiae subsp. equisimilis emm types. To examine the biological activity of DrsG, recombinant versions of two major DrsG variants, DrsGS and DrsGL, were expressed and purified. Western blot analysis using antisera raised to these proteins demonstrated both variants to be expressed and secreted into culture supernatants. Unlike SIC, but similar to DRS, DrsG does not inhibit complement-mediated lysis. However, like both SIC and DRS, DrsG is a ligand of the cathelicidin LL-37 and is inhibitory to its bactericidal activity in in vitro assays. Conservation of prolines in the C-terminal region also suggests that these residues are important in the biology of this family of proteins. This is the first report demonstrating the activity of an AMP-inhibitory protein in S. dysgalactiae subsp. equisimilis and suggests that inhibition of AMP activity is the primary function of this family of proteins. The acquisition of the complement-inhibitory activity of SIC may reflect its continuing evolution.

Molecular markers for the study of streptococcal epidemiology.

Diseases caused by Streptococcus pyogenes (Group A streptococcus, GAS) range from superficial infections such as pharyngitis and impetigo to potentially fatal rheumatic heart disease and invasive disease. Studies spanning emm-typing surveillance to population genomics are providing new insights into the epidemiology, pathogenesis, and biology of this organism. Such studies have demonstrated the differences that exist in the epidemiology of streptococcal disease between developing and developed nations. In developing nations, where streptococcal disease is endemic, the diversity of GAS emm-types circulating is much greater than that found in developed nations. An association between emm-type and disease, as observed in developed countries is also lacking. Intriguingly, comparative genetic studies suggest that emm-type is not always a good predictor of the evolutionary relatedness of geographically distant isolates. A view of GAS as a highly dynamic organism, in possession of a core set of virulence genes that contribute to host niche specialization and common pathogenic processes, augmented by accessory genes that change the relative virulence of specific lineages is emerging. Our inability to definitively identify genetic factors that contribute to specific disease outcome underscores the complex nature of streptococcal diseases.

Complement inhibitors from scabies mites promote streptococcal growth--a novel mechanism in infected epidermis?

BACKGROUND: Scabies is highly prevalent in socially disadvantaged communities such as indigenous populations and in developing countries. Generalized itching causes discomfort to the patient; however, serious complications can occur as a result of secondary bacterial pyoderma, commonly caused by Streptococcus pyogenes (GAS) or Staphylococcus aureus. In the tropics, skin damage due to scabies mite infestations has been postulated to be an important link in the pathogenesis of disease associated with acute rheumatic fever and heart disease, poststreptococcal glomerulonephritis and systemic sepsis. Treatment of scabies decreases the prevalence of infections by bacteria. This study aims to identify the molecular mechanisms underlying the link between scabies and GAS infections. METHODOLOGY/PRINCIPAL FINDINGS: GAS bacteria were pre-incubated with blood containing active complement, phagocytes and antibodies against the bacteria, and subsequently tested for viability by plate counts. Initial experiments were done with serum from an individual previously exposed to GAS with naturally acquired anti-GAS antibodies. The protocol was optimized for large-scale testing of low-opsonic whole blood from non-exposed human donors by supplementing with a standard dose of heat inactivated human sera previously exposed to GAS. This allowed an extension of the dataset to two additional donors and four proteins tested at a range of concentrations. Shown first is the effect of scabies mite complement inhibitors on human complement using ELISA-based complement activation assays. Six purified recombinant mite proteins tested at a concentration of 50 µg/ml blocked all three complement activation pathways. Further we demonstrate in human whole blood assays that each of four scabies mite complement inhibitors tested increased GAS survival rates by 2-15 fold. CONCLUSIONS/SIGNIFICANCE: We propose that local complement inhibition plays an important role in the development of pyoderma in scabies infested skin. This molecular link between scabies and bacterial infections may provide new avenues to develop alternative treatment options against this neglected disease.