Detection of superantigens in Streptococcus pyogenes isolates based on whole genome sequencing data.

Streptococcus pyogenes causes a variety of human diseases ranging from uncomplicated respiratory tract and skin infections to severe invasive diseases possibly involving toxic shock syndrome. Besides the emm gene-encoded M protein, important virulence factors are pyrogenic exotoxins, referred to as superantigens. The National Reference Laboratory for Streptococcal Infections has newly introduced bioinformatics tools for processing S. pyogenes whole genome sequencing data. Using the SRST2 software and BV-BRC platform, WGS data of 10 S. pyogenes isolates from patients with invasive disease were analysed, and emm type, sequence type, and superantigen encoding gene profiles were determined. The Unicycler assembly pipeline with the SPAdes de novo assembler was used to assemble genome sequences from short reads.

Optimized surface plasmon resonance immunoassay for staphylococcal enterotoxin G detection using silica nanoparticles.

Staphylococcal enterotoxins are one of the most important causative agents of food poisoning. These molecules function as both gastrointestinal toxins and superantigens (SAgs) which can simultaneously bind MHC-II and T cell receptor leading to a non-specific polyclonal T cell activation and massive proinflammatory cytokine release. Common symptoms include vomiting and diarrhea; however, in more severe cases, systemic dissemination may result in toxic shock syndrome and can be lethal in a few hours. Only small amounts of these heat-stable toxins are needed to cause the disease. Therefore, it is highly important to detect quickly low concentrations of SAgs in biological samples. In this work, we report a surface plasmon resonance (SPR)-based capture immunoassay for the detection of the SAg SEG. We analyzed the use of different amplification strategies. The SPR-based double-antibody sandwich approach could detect picomolar levels of SEG. The use of antibody-coated silica nanoparticles (AbSiNPs) as an alternative enhancing reagent also detected SEG in the picomolar range. Although AbSiNPs did not improve the limit of detection, for the same amount of SAg tested, AbSiNPs gave a higher response level than free antibodies. This work highlights the suitability of silica nanoparticles for signal amplification in SPR-based biosensors. Overall, SPR biosensors offer the capability for continuous real-time monitoring and high sensitivity that can be befitting for the detection of enterotoxins in food industries, laboratories and regulatory agencies.

Highly Sensitive and Specific Detection of Staphylococcal Enterotoxins SEA, SEG, SEH, and SEI by Immunoassay.

Staphylococcal food poisoning (SFP) is one of the most common foodborne diseases worldwide, resulting from the ingestion of staphylococcal enterotoxins (SEs), primarily SE type A (SEA), which is produced in food by enterotoxigenic strains of staphylococci, mainly S. aureus. Since newly identified SEs have been shown to have emetic properties and the genes encoding them have been found in food involved in poisoning outbreaks, it is necessary to have reliable tools to prove the presence of the toxins themselves, to clarify the role played by these non-classical SEs, and to precisely document SFP outbreaks. We have produced and characterized monoclonal antibodies directed specifically against SE type G, H or I (SEG, SEH or SEI respectively) or SEA. With these antibodies, we have developed, for each of these four targets, highly sensitive, specific, and reliable 3-h sandwich enzyme immunoassays that we evaluated for their suitability for SE detection in different matrices (bacterial cultures of S. aureus, contaminated food, human samples) for different purposes (strain characterization, food safety, biological threat detection, diagnosis). We also initiated and described for the first time the development of monoplex and quintuplex (SEA, SE type B (SEB), SEG, SEH, and SEI) lateral flow immunoassays for these new staphylococcal enterotoxins. The detection limits in buffer were under 10 pg/mL (0.4 pM) by enzyme immunoassays and at least 300 pg/mL (11 pM) by immunochromatography for all target toxins with no cross-reactivity observed. Spiking studies and/or bacterial supernatant analysis demonstrated the applicability of the developed methods, which could become reliable detection tools for the routine investigation of SEG, SEH, and SEI.

Effect of non-absorbent intravaginal menstrual/contraceptive products on Staphylococcus aureus and production of the superantigen TSST-1.

Tampons are associated with toxic shock syndrome (mTSS). One reason for this association is oxygen introduction within tampons into the anaerobic vagina. Oxygen is required for Staphylococcus aureus to produce TSS toxin-1 (TSST-1). There have been changes in use of medical devices to control menstrual flow, including increased use of menstrual discs and cups. These devices composed of solid, flexible materials do not absorb menstrual fluid and thus do not trap oxygen. This study evaluates tampons and non-absorbent devices for effect on S. aureus and TSST-1 production. There are three in vitro tests to evaluate devices for effect on TSST-1 production: (1) stationary flask, (2) shake flask, and (3) tampon sac. In this study, 100% rayon and 100% cotton tampons with three absorbencies, contraceptive diaphragms, and menstrual discs and cups were tested for effect on S. aureus growth and TSST-1 production. Product composition did not affect bacterial growth or TSST-1 production. Tampons showed no effect on S. aureus growth compared with no-tampon controls, but tampons showed enhanced TSST-1 production as a function of trapped oxygen in stationary cultures and tampon sacs but not in shake flasks. The non-absorbent devices showed no enhanced S. aureus growth or TSST-1 production compared with no-device controls. These studies are consistent with the association of tampons with mTSS as a function of absorbency, but they suggest the occasional association of mTSS with non-absorbent devices may be coincidental as opposed to co-causative.

High Prevalence of Staphylococcus aureus Enterotoxin Gene Cluster Superantigens in Cystic Fibrosis Clinical Isolates.

BACKGROUND: Staphylococcus aureus is a highly prevalent respiratory pathogen in cystic fibrosis (CF). It is unclear how this organism establishes chronic infections in CF airways. We hypothesized that S. aureus isolates from patients with CF would share common virulence properties that enable chronic infection. METHODS: 77 S. aureus isolates were obtained from 45 de-identified patients with CF at the University of Iowa. We assessed isolates phenotypically and used genotyping assays to determine the presence or absence of 18 superantigens (SAgs). RESULTS: We observed phenotypic diversity among S. aureus isolates from patients with CF. Genotypic analysis for SAgs revealed 79.8% of CF clinical isolates carried all six members of the enterotoxin gene cluster (EGC). MRSA and MSSA isolates had similar prevalence of SAgs. We additionally observed that EGC SAgs were prevalent in S. aureus isolated from two geographically distinct CF centers. CONCLUSIONS: S. aureus SAgs belonging to the EGC are highly prevalent in CF clinical isolates. The greater prevalence in these SAgs in CF airway specimens compared to skin isolates suggests that these toxins confer selective advantage in the CF airway.

Magnetic nanocomposites: versatile tool for the combination of immunomagnetic separation with flow-based chemiluminescence immunochip for rapid biosensing of Staphylococcal enterotoxin B in milk.

Immunomagnetic separation (IMS) was combined with flow-based chemiluminescence sandwich immunoassays (CL-SIA) for the quantification of Staphylococcal enterotoxin B in milk. Therefore, iron oxide-shell silica-core magnetic nanocomposites were conjugated to biotinylated anti-SEB antibodies (MNC-IgGs). MNC-IgGs were applied successfully for (i) capturing SEB in milk samples by an affinity reaction, (ii) magnetophoretic collection on antibody spots in a channel of a flow-based immunochip, and (iii) sensitive enzymatic chemiluminescence detection of biotin labels by poly(horseradish peroxidase)-streptavidin. IMS was performed in 0.6 mL and 100 mL milk samples resulting in detection limits of 50 ng L-1 and 0.39 ng L-1, respectively, for the combined analytical method. It was shown that the assay sensitivity was dramatically improved by the combination of IMS with flow-based CL-SIA compared to CL-SIA directly applied with milk samples (detection limit 130 ng L-1). The IMS-CL-SIA has a time-to-result of 2-3 h. The reported combined analytical method can be used for a rapid control of SEB in complex food matrices such as milk. In future, even the monitoring of multiple contaminants in food or water may be performed by IMS-CL-SIA. Graphical abstract.

Intact Staphylococcus Enterotoxin SEB from Culture Supernatant Detected by MALDI-TOF Mass Spectrometry.

Routine identification of pathogens by MALDI-TOF MS (matrix-assisted laser desorption ionisation time-of-flight mass spectrometry) is based on the fingerprint of intracellular proteins. This work evaluated the use of MALDI-TOF MS for the identification of extracellular pathogen factors. A Staphylococcus aureus isolate from a food contaminant was exponentially grown in liquid cultures. Secreted proteins were collected using methanol⁻ chloroform precipitation and analysed by MALDI-TOF MS. A main peak m/z 28,250 was demonstrated, which was identified as S.aureus enterotoxin type B (SEB) by using the pure authentic SEB reference of 28.2 kDa and by amino acid sequence analysis. SEB was also detected in this intact form following pasteurization and cooking treatments. Further application of the elaborated MALDI-TOF MS protocol resulted in the detection of SEA at m/z 27,032 and SEC at m/z 27,629. In conclusion, a simple sample preparation from S.aureus cultures and an easy-to-perform identification of pathogen factors SE in intact form represents a promising next-generation application of MALDI-TOF MS.

[Staphylococcal toxic shock syndrome should be considered in the event of diffuse erythema with fever and shock]. / Érythème généralisé fébrile et choc : choc toxinique staphylococcique.

BACKGROUND: Toxic shock syndrome (TSS) was first described by Todd in 1978. The relevant Lancet publication reported 7 cases of children with fever, exanthema, hypotension and diarrhoea associated with multiple organ failure. An association between TSS and use of hyper-absorbent tampons in menstruating women was discovered in the 1980s. Following the market withdrawal of such tampons, TSS virtually disappeared. Herein we report a new case of TSS in a 15-year-old girl. PATIENTS AND METHODS: A 15-year-old patient was admitted to intensive care for severe sepsis and impaired consciousness associated with diffuse abdominal pain. Dermatological examination revealed diffuse macular exanthema. Laboratory tests showed hepatic cytolysis (ASAT 101 U/L, ALAT 167 U/L, total bilirubin 68µmol/L) and an inflammatory syndrome. Lumbar puncture and blood cultures were sterile while thoraco-abdomino-pelvic and brain scans were normal. The patient was menstruating and had been using a tampon over the previous 24hours. Vaginal sampling and tampon culture revealed TSST-1 toxin-producing S. aureus. Management consisted of intensive care measures and treatment with amoxicillin-clavulanic acid and clindamycin for 10 days. CONCLUSION: In case of septic shock associated with diffuse macular exanthema a diagnosis of TSS must be envisaged, particularly in menstruating women.

Subinhibitory concentrations of tedizolid potently inhibit extracellular toxin production by methicillin-sensitive and methicillin-resistant Staphylococcus aureus.

PURPOSE: Potent extracellular toxins including alpha-haemolysin, Panton-Valentine leukocidin (PVL) and toxic-shock syndrome toxin 1 (TSST-1) significantly contribute to Staphylococcus aureus pathogenesis, thus, toxin suppression is a primary focus in treatment of staphylococcal disease. S. aureus maintains complex strategies to regulate toxin expression and previous data have demonstrated that subinhibitory concentrations of beta-lactam antibiotics can adversely increase S. aureus exotoxin production. The current study evaluates the effects of subinhibitory concentrations of tedizolid, a second-generation oxazolidinone derivative, on expression of staphylococcal exotoxins in both methicillin-resistant and methicillin-sensitive S. aureus. METHODOLOGY: S. aureus exotoxin expression levels were compared at 12 and 24 h following treatment with tedizolid, linezolid, nafcillin or vehicle control. RESULTS: Our findings show that the level of antibiotic required to alter toxin production was strain-dependent and corresponds with the quantity of toxin produced, but both tedizolid and linezolid could effectively reduce expression of alpha-haemolysin, PVL and TSST-1 toxin at subinhibitory concentrations. In contrast, nafcillin showed less attenuation and, in some S. aureus strains, led to an increase in toxin expression. Tedizolid consistently inhibited toxin production at a lower overall drug concentration than comparator agents. CONCLUSION: Together, our data support that tedizolid has the potential to improve outcomes of infection due to its superior ability to inhibit S. aureus growth and attenuate exotoxin production.

Analysis of the VIDAS® Staph Enterotoxin III (SET3) for Detection of Staphylococcal Enterotoxins G, H, and I in Foods.

BACKGROUND: Staphylococcal food poisoning (SFP) frequently causes illnesses worldwide. SFP occurs from the ingestion of staphylococcal enterotoxins (SEs) preformed in foods by enterotoxigenic strains of Staphylococcus species, primarily S. aureus. SEG, SEH, and SEI induce emesis and have been implicated in outbreaks. Immunological-based methods are deemed the most practical methods for the routine analysis of SEs in foods given their ease of use, sensitivity, specificity, and commercial availability. These kits are routinely used to test for SEA-SEE. However, only recently has a kit been developed to detect SEG, SEH, and SEI. OBJECTIVE: Our research examined the performance of the novel VIDAS® Staph Enterotoxin III (SET3) for the detection of staphylococcal enterotoxins SEG, SEH, and SEI in foods. METHODS: Here we assess the sensitivity and specificity of SET3 using duplicate test portions of six foods at varying concentrations of inclusivity and exclusivity inocula: pure SEG, SEH, SEI, S. aureus strain extracts positive for seg, seh, and sei, as well as SEA, SEB, SEC, SED, and SEE. RESULTS: The overall detection limit was less than 2.09 ng/mL for foods inoculated with SEG, SEH, and SEI, with no cross reactivity observed. HIGHLIGHTS: Integrating concurrent testing to detect the presence of SEA-SEE and SEG-SEI utilizing the SET3 along with the VIDAS SET2, Ridascreen® SET total, or other comparable kits will be instrumental for the future food assessments in our laboratory and may become the new standard for SE analysis of foods.

Impact of Currently Marketed Tampons and Menstrual Cups on Staphylococcus aureus Growth and Toxic Shock Syndrome Toxin 1 Production In Vitro.

Fifteen currently marketed intravaginal protection products (11 types of tampon and 4 types of menstrual cup) were tested by the modified tampon sac method to determine their effect on Staphylococcus aureus growth and toxic shock syndrome toxin 1 (TSST-1) production. Most tampons reduced S. aureus growth and TSST-1 production, with differences based on brand and composition, and the level of S. aureus growth was higher in destructured than in unaltered tampons. We observed higher levels of S. aureus growth and toxin production in menstrual cups than in tampons, potentially due to the additional air introduced into the bag by cups, with differences based on cup composition and size.IMPORTANCE Menstrual toxic shock syndrome is a rare but severe disease. It occurs in healthy women vaginally colonized by Staphylococcus aureus producing toxic shock syndrome toxin 1 using intravaginal protection, such as tampons or menstrual cups. Intravaginal protection induces TSS by the collection of catamenial products, which act as a growth medium for S. aureus Previous studies evaluated the impact of tampon composition on S. aureus producing toxic shock syndrome toxin 1, but they are not recent and did not include menstrual cups. This study demonstrates that highly reproducible results for S. aureus growth and TSST-1 production can be obtained by using a simple protocol that reproduces the physiological conditions of tampon and cup usage as closely as possible, providing recommendations for tampon or cup use to both manufacturers and consumers. Notably, our results do not show that menstrual cups are safer than tampons and suggest that they require similar precautions.

Detección y expresión de superantígenos y de resistencia antimicrobiana en aislamientos obtenidos de mujeres portadoras de Staphylococcus aureus que cuidan y alimentan niños.

Introducción. Staphylococcus aureus coloniza mucosas y piel, y causa graves infecciones en el hombre y los animales. Es importante establecer el estatus de portadoras de cepas enterotoxigénicas de este microorganismo en manipuladoras de alimentos, con el fin de prevenir intoxicaciones alimentarias.Objetivo. Establecer las correlaciones entre los genes de enterotoxinas clásicas, el gen tsst-1, la producción de toxinas en cultivo y la resistencia antimicrobiana en aislamientos de S. aureus provenientes de manipuladoras de alimentos que cuidan niños en sus comunidades.Materiales y métodos. Se cultivaron muestras de las fosas nasales y las yemas de los dedos de las manos, y se identificó S. aureus empleando las pruebas de rutina y métodos automatizados. La extracción de ADN se hizo mediante el método de bromuro de cetil-trimetil-amonio (Cetyl-Trimethyl-Ammonium Bromide, CTAB) modificado. Para la detección de superantígenos se emplearon pruebas de reacción en cadena de la polimerasa (PCR) simple y múltiple, y para la de toxinas, estuches comerciales.Resultados. Se encontró que el 22,0 % de los aislamientos correspondía a portadoras de S. aureus: 17,0 % en los aislamientos de fosas nasales; 5,0 % en los de las manos y 6,7 % simultáneamente en los dos sitios. La prevalencia de superantígenos fue de 73,7 %. El genotipo más frecuente fue el seatsst-1, con 10,0 %. La resistencia a un solo antibiótico fue de 74,7 % y, a cuatro antibióticos, de 3,2 %; de los aislamientos, el 93,7 % correspondía a cepas productoras de betalactamasas. La detección de genes clásicos y de tsst-1 mediante PCR fue de 48,4 % y la de toxinas en el sobrenadante, de 42,1 %,con una correlación de 95,7 %. Las mayores correlaciones se establecieron entre las toxinas TSST-1 (22/22) y SEA (17/18). La correlación del gen tsst-1 con la proteína y la resistencia fue de 100 %. Todos los aislamientos con el genotipo sea-tsst-1 t fueron resistentes y productores de las toxinas.Conclusión. La tasa de aislamientos de S. aureus toxigénicos y resistentes obtenidos de mujeres que cuidan y preparan alimentos para niños fue de más de 70 %, lo que demostró su gran virulencia y la consecuente necesidad de aplicar estrictamente las normas higiénicas y sanitarias vigentes para evitar el riesgo de intoxicación alimentaria.

Detección y expresión de superantígenos y de resistencia antimicrobiana en aislamientos obtenidos de mujeres portadoras de Staphylococcus aureus que cuidan y alimentan niños / Detection and expression of superantigens and resistance in isolates from women carriers of Staphylococcus aureus who take care of and feed children

Resumen Introducción. Staphylococcus aureus coloniza mucosas y piel, y causa graves infecciones en el hombre y los animales. Es importante establecer el estatus de portadoras de cepas enterotoxigénicas de este microorganismo en manipuladoras de alimentos, con el fin de prevenir intoxicaciones alimentarias. Objetivo. Establecer las correlaciones entre los genes de enterotoxinas clásicas, el gen tsst-1, la producción de toxinas en cultivo y la resistencia antimicrobiana en aislamientos de S. aureus provenientes de manipuladoras de alimentos que cuidan niños en sus comunidades. Materiales y métodos. Se cultivaron muestras de las fosas nasales y las yemas de los dedos de las manos, y se identificó S. aureus empleando las pruebas de rutina y métodos automatizados. La extracción de ADN se hizo mediante el método de bromuro de cetil-trimetil-amonio (Cetyl-Trimethyl- Ammonium Bromide, CTAB) modificado. Para la detección de superantígenos se emplearon pruebas de reacción en cadena de la polimerasa (PCR) simple y múltiple, y para la de toxinas, estuches comerciales. Resultados. Se encontró que el 22,0 % de los aislamientos correspondía a portadoras de S. aureus: 17,0 % en los aislamientos de fosas nasales; 5,0 % en los de las manos y 6,7 % simultáneamente en los dos sitios. La prevalencia de superantígenos fue de 73,7 %. El genotipo más frecuente fue el sea-tsst-1, con 10,0 %. La resistencia a un solo antibiótico fue de 74,7 % y, a cuatro antibióticos, de 3,2 %; de los aislamientos, el 93,7 % correspondía a cepas productoras de betalactamasas. La detección de genes clásicos y de tsst-1 mediante PCR fue de 48,4 % y la de toxinas en el sobrenadante, de 42,1 %, con una correlación de 95,7 %. Las mayores correlaciones se establecieron entre las toxinas TSST-1 (22/22) y SEA (17/18). La correlación del gen tsst-1 con la proteína y la resistencia fue de 100 %. Todos los aislamientos con el genotipo sea-tsst-1 t fueron resistentes y productores de las toxinas. Conclusión. La tasa de aislamientos de S. aureus toxigénicos y resistentes obtenidos de mujeres que cuidan y preparan alimentos para niños fue de más de 70 %, lo que demostró su gran virulencia y la consecuente necesidad de aplicar estrictamente las normas higiénicas y sanitarias vigentes para evitar el riesgo de intoxicación alimentaria.

Abstract Introduction: Staphylococcus aureus colonizes mucous membranes and skin causing severe infections in humans and animals. It is important to determine carrier status of enterotoxigenic strains of this microorganism in food handlers to prevent food poisoning. Objective: To establish the correlations among classic enterotoxigenic genes, tsst-1 gene, the production of toxins in cultures and antimicrobial resistance in S. aureus isolates from women who handle the food, feed and take care of children in their communities. Materials and methods: Nasal swab and finger samples were cultured and S. aureus was identified using routine methods and automated systems. DNA extraction was done by the CTAB modified method, and superantigen detection by simple and multiplex PCR, while toxins were detected using commercial kits. Results: We found that 22.0% of subjects were S. aureus carriers: 17.0% corresponded to nose samples, 5.0% to hands and 6.7% to both nose and hands. The prevalence of superantigens was 73.7%. The most frequent genotype was sea-tsst-1 with 10%. Resistance to one antibiotic was 74.7%, and to four antibiotics, 3.2%; 93.7% of the isolates were betalactamase-positive. Classical genes and tsst-1 gene were detected by PCR in 48.4% of samples and toxins in supernatant were detected in 42.1% of them with 95.7% of correlation.The highest correlations were established for TSST-1 and SEA with 100% and 94.4%, respectively. The correlation of tsst-1 gene with toxin production and resistance was 100%. All isolates with genotype sea-tsst-1 were toxin-positive and resistant. Conclusion: The rate of toxigenic and resistant S. aureus isolates from women in charge of feeding and taking care of children was higher than 70%, which demonstrates its high virulence. This requires the strict application of hygienic and sanitary regulations in order to avoid the risk of food poisoning.

Epidemiology and distribution of 10 superantigens among invasive Streptococcus pyogenes disease in Germany from 2009 to 2014.

A nationwide laboratory-based surveillance study of invasive S. pyogenes infections was conducted in Germany. Invasive isolates (n = 719) were obtained between 2009 and 2014. Most isolates were obtained from blood (92.1%). The proportions of isolates from cerebrospinal fluid, pleural fluid, synovial fluid and peritoneal fluid were 3.9%, 1.8%, 1.7% and 0.6%, respectively. The most common emm types were emm 1 (31.8%), emm 28 (15.4%) and emm 89 (14.5%). The most common superantigen genes (speA, speC, speG, speH, speI, speJ, speK, speL, speM, ssa) identified from S. pyogenes were speG (92.1%), speJ (50.9%), and speC (42.0%). Significant associations of superantigen genes with underlying conditions or risks were observed in speG, speH, speJ, and speK. Significant associations between emm types or superantigen genes with clinical complications were observed in emm type 3 and in superantigen gene speA 1-3. Most frequent clinical manifestations included sepsis 59.4%, STSS 6.3%, meningitis 5.4%, and necrotizing fasciitis 5.0% (significantly associated with emm1).

Do Staphylococcus aureus superantigens play a role in the pathogenesis of otitis media with effusion in children?

OBJECTIVES: Staphylococcal enterotoxins (SEs), acting as superantigens, have been reported to be involved in the pathogenesis of chronic inflammatory diseases of the upper and lower airway. There has been no previous study investigating the role of SEs in otitis media with effusion (OME). Therefore, this study was designed to analyze middle ear aspirates from children with and without OME for the presence of SEs. METHODS: Middle ear aspirates were obtained from 24 patients and 24 controls. All samples were processed for bacterial culture and detection of five staphylococcal SEs (SEA, SEB, SEC and SED) and toxic shock syndrome toxin-1 using the Rapid Latex Agglutination Test. RESULTS: In bacterial culture assays, six samples (25%) of the study group and five samples (20.8%) of the control group showed bacterial growth. At least one SE was demonstrated in 6 of 24 patients and in 3 of 24 controls. There was no statistically significant difference between the two groups with respect to the presence of SEs. CONCLUSION: Although there is evidence that SEs have a potential role in the pathogenesis of chronic inflammatory diseases, there is no evidence that the inflammation process is initiated by SEs in patients with OME.

Determining the Presence of Superantigens in Coagulase Negative Staphylococci from Humans.

Superantigens (SAgs) are important virulence factors in S. aureus. Recent studies identified their presence in animal coagulase-negative staphylococci (CNS). The emergence of human-associated SAg+ CNS would mark a prodigious shift in virulence capabilities. We examined CNS isolates from healthy human nares and diseased individuals, and determined that no known SAgs were present.

Streptococcus pyogenes strains in Sao Paulo, Brazil: molecular characterization as a basis for StreptInCor coverage capacity analysis.

BACKGROUND: Several human diseases are caused by Streptococcus pyogenes, ranging from common infections to autoimmunity. Characterization of the most prevalent strains worldwide is a useful tool for evaluating the coverage capacity of vaccines under development. In this study, a collection of S. pyogenes strains from Sao Paulo, Brazil, was analyzed to describe the diversity of strains and assess the vaccine coverage capacity of StreptInCor. METHODS: Molecular epidemiology of S. pyogenes strains was performed by emm-genotyping the 229 isolates from different clinical sites, and PCR was used for superantigen profile analysis. The emm-pattern and tissue tropism for these M types were also predicted and compared based on the emm-cluster classification. RESULTS: The strains were fit into 12 different emm-clusters, revealing a diverse phylogenetic origin and, consequently, different mechanisms of infection and escape of the host immune system. Forty-eight emm-types were distinguished in 229 samples, and the 10 most frequently observed types accounted for 69 % of all isolates, indicating a diverse profile of circulating strains comparable to other countries under development. A similar proportion of E and A-C emm-patterns were observed, whereas pattern D was less frequent, indicating that the strains of this collection primarily had a tissue tropism for the throat. In silico analysis of the coverage capacity of StreptInCor, an M protein-conserved regionally based vaccine candidate developed by our group, had a range of 94.5 % to 59.7 %, with a mean of 71.0 % identity between the vaccine antigen and the predicted amino acid sequence of the emm-types included here. CONCLUSIONS: This is the first report of S. pyogenes strain characterization in Sao Paulo, one of the largest cities in the world; thus, the strain panel described here is a representative sample for vaccine coverage capacity analysis. Our results enabled evaluation of StreptInCor candidate vaccine coverage capacity against diverse M-types, indicating that the vaccine candidate likely would induce protection against the diverse strains worldwide.

Superantigens in Staphylococcus aureus isolated from prosthetic joint infection.

Staphylococcus aureus is a common cause of prosthetic joint infection (PJI). The prevalence of superantigens (SAgs) among PJI-associated S. aureus is unknown. Eighty-four S. aureus isolates associated with PJI isolated between 1999 and 2006 were studied. SAg genes, sea, seb, sec, sed, see, seg, seh, sei, and tst, were assayed by PCR. Seventy-eight (92.9%) isolates carried at least 1 SAg gene studied, with 61 (72.6%) harboring more than 1. seg was most commonly (70.2%), and seh was least frequently (4.8%) detected. tst-positive isolates were associated with early infection and increased erythrocyte sedimentation rate at diagnosis (P=0.006 and P=0.021, respectively). seg and sei were associated with methicillin resistance (P=0.008 and P=0.002, respectively). A majority of PJI-associated isolates studied produced biologically active SAgs in both planktonic and biofilm growth modes. SAg genes are prevalent in S. aureus causing PJI.

Superantigens of Staphylococcus aureus from patients with diabetic foot ulcers.

BACKGROUND: Diabetic foot ulcer (DFU) infections are challenging. Staphylococcus aureus is the most commonly isolated pathogen in DFUs. Superantigens (SAgs) are causative in many S. aureus infections. We hypothesized both that DFU S. aureus will produce large SAg numbers, consistent with skin infections, and that certain SAgs will be overrepresented. We assessed the SAg and α-toxin profile of isolates from patients with DFU, compared with profiles of isolates from other sources. MATERIALS: Twenty-five S. aureus isolates from patients with DFU were characterized. Polymerase chain reaction was used to detect genes for methicillin-resistance and SAgs. Some SAgs and the α-toxin were quantified. We compared the SAg profile of DFU isolates with SAg profiles of S. aureus isolates from skin lesions of patients with atopic dermatitis and from vaginal mucosa of healthy individuals. RESULTS: Most DFU isolates were methicillin susceptible (64%), with USA100 the most common clonal group. The SAg gene profile of DFU isolates most closely resembled that of isolates from patients with atopic dermatitis, with the highest number of different SAg genes per isolate and a high prevalence of staphylococcal enterotoxin D and the enterotoxin gene cluster. DFU isolates also had a high prevalence of staphylococcal enterotoxin-like X. CONCLUSIONS: Comparison of the SAg profile of DFU isolates to SAg profiles of skin lesion isolates and vaginal mucosa isolates revealed that the SAg profile of DFU isolates was more similar to that of skin lesion isolates. SAgs offer selective advantages in facilitating DFU infections and suggest that therapies to neutralize or reduce SAg production by S. aureus may be beneficial in management of patients with DFU.